REVIEW NOTES IN CLINICAL CHEMISTRY Flashcards by Arriane Cyrel (MTI)

: the act of obtaining a blood sample from a vein using a needle attached to a syringe or a stoppered evacuated tube; it is the most common way to collect blood specimens

VENIPUNCTURE

How well did you know this?

Not at all

Perfectly

THE MAJOR VEINS FOR VENIPUNCTURE are in the antecubital fossa, the area of the arm in front of the elbow. The H pattern is displayed by approximately 70% of the population and includes the following veins:

Median
Cephalic
Basilic

How well did you know this?

Not at all

Perfectly

Located near the center of the antecubital fossa

Median cubital vein

How well did you know this?

Not at all

Perfectly

• Preferred vein because it is typically large, closer to the surface and the most stationary

Median cubital vein

How well did you know this?

Not at all

Perfectly

Easiest and least painful to puncture

Median cubital vein

How well did you know this?

Not at all

Perfectly

• Least likely to bruise

Median cubital vein

How well did you know this?

Not at all

Perfectly

Second-choice vein

Cephalic vein

How well did you know this?

Not at all

Perfectly

• Often harder to palpate than medial cubital vein

Cephalic vein

How well did you know this?

Not at all

Perfectly

• Fairly well-anchored

Cephalic vein

How well did you know this?

Not at all

Perfectly

• Often the only vein felt in obese patients

Cephalic vein

How well did you know this?

Not at all

Perfectly

Last choice

Basilic vein

How well did you know this?

Not at all

Perfectly

Not well anchored and rolls easily

Basilic vein

How well did you know this?

Not at all

Perfectly

• Increased risk of puncturing a median cutaneous nerve branch or the brachial artery

Basilic vein

How well did you know this?

Not at all

Perfectly

• Not recommended unless no other vein in either arm is more prominent

Basilic vein

How well did you know this?

Not at all

Perfectly

OTHER VEINS
Although antecubital veins are used most frequently, veins on the [?] may also be used for venipuncture. Veins on the [?], however, should never be used for venipuncture. [?] are sometimes used but not without permission of the patient’s physician, due to a potential for significant medical complications.

back of the hand and wrist

underside of the wrist

Leg, ankle, and foot veins

How well did you know this?

Not at all

Perfectly

• normally a clear, pale yellow fluid

Serum

How well did you know this?

Not at all

Perfectly

• non-fasting serum can be cloudy due to lipids

Serum

How well did you know this?

Not at all

Perfectly

• separated from clotted blood by centrifugation (approx. 10 minutes at an RCF of 1,000 to 2,000g)

Serum

How well did you know this?

Not at all

Perfectly

• many chemistry tests are performed on serum

Serum

How well did you know this?

Not at all

Perfectly

• normally a clear to slightly hazy, pale yellow fluid

Plasma

How well did you know this?

Not at all

Perfectly

• separates from the cells when blood in an anticoagulant tube is centrifuged

Plasma

How well did you know this?

Not at all

Perfectly

• contains fibrinogen (serum does not because it was used in clot formation)

Plasma

How well did you know this?

Not at all

Perfectly

• Stat and other tests requiring a fast turnaround time (TAT) are often collected in tubes containing heparin anticoagulant because they can be centrifuged immediately to obtain plasma

Plasma

How well did you know this?

Not at all

Perfectly

• contains both cells and plasma

Whole blood

How well did you know this?

Not at all

Perfectly

• must be collected in an anticoagulant tube to keep it from clotting

Whole blood

• used for most hematology tests and many point-of-care tests (POCTs), especially in acute care and stat situations.

Whole blood

preferred method because blood is collected directly from the vein into a tube, minimizing the risk of specimen contamination and exposure to the blood.

Evacuated tube system (ETS)

• discouraged by CLSI due to safety and specimen quality issues

Needle and syringe

• sometimes used on small, fragile, or damaged veins

Needle and syringe

• can be used with the ETS or a syringe

Butterfly set

• often used to draw blood from:

Butterfly set

infants and children; hand veins; in other difficult-draw situations

Butterfly set

Fibrin degradation products

Light blue

Inhibits thrombin formation

Light green/black Green Tan (glass) Royal blue (heparin)

Inhibits glycolysis

Gray

WBC preservative

Yellow

Clot activator

Yellow/gray and orange

Silica clot

Red/gray and gold

CAPILLARY PUNCTURE Length of lancet should be

less than 2.0 mm to avoid penetrating bone

CAPILLARY PUNCTURE Sites:

Palmar surface of 3rd and 4th fingers Lateral plantar heel surface (newborns) Earlobes

Syringes are used instead of evacuated tubes because of the pressure in an arterial blood vessel.

ARTERIAL PUNCTURE

ARTERIAL PUNCTURE Preferred anticoagulant:

Heparin

Collect without a tourniquet

ARTERIAL PUNCTURE

ARTERIAL PUNCTURE Primary arterial sites (in order of preference):

radial, brachial and femoral arteries

ARTERIAL PUNCTURE Major complications of arterial puncture:

thrombosis, hemorrhage, and possible infection

done before the collecting arterial blood from radial artery

Modified Allen Test

to determine whether the ulnar artery can provide collateral circulation to the hand after the radial arter puncture

Modified Allen Test

Albumin, ALP (↑older), phosphorus, cholesterol

Age

(↑older)

ALP

: Albumin, ALP, creatine, calcium, uric acid, CK, AST, PO4, BUN, magnesium, bilirubin, cholesterol

↑ males

: Fe, cholesterol, gamma-globulins, a-lipoproteins

↑ females

Peaks 4-6 AM; lowest 8 PM-12 AM; 50% lower at 8 PM than at 8 AM

Cortisol

Lower at night

ACTH, Plasma renin activity, Aldosterone, Insulin

Lower at night; higher standing than supine

Plasma renin activity

Higher in afternoon and evening

Acid phosphatase, Growth hormone

Higher levels at 4 and 8 AM and at 8 and 10 PM

Prolactin

Peaks early to late morning; decreases up to 30% during the day

Iron

≥20% for ALT, bilirubin, Fe, TSH, triglycerides

Day-to-day variation

↑ Glucose, insulin, triglycerides, gastrin, ionized calcium

Recent Food Ingestion

↓ chloride, phosphorus, potassium, amylase, ALP

Recent Food Ingestion

: albumin, cholesterol, aldosterone, calcium

↑ when standing

: CK

↑ in ambulatory patients

: lactic acid, creatine, protein, CK, AST, LD, thyroxine

↑ with exercise

: cholesterol and triglycerides

↓ with exercise

↑ACTH, cortisol, catecholamines, prolactin

Stress

: ↑TP, ↓ albumin

Black

: ↑CK/LD

Black males

: IgG ↑40% and IgA ↑20%

Black male vs white male

: ↑cholesterol and triglycerides

White & >40 years old

FBS, GTT, Triglycerides, lipid panel, gastrin, insulin, aldosterone/ renin

Require fasting

Lactic acid, ammonia, blood gas (if not cooled = ↓ pH and pO2)

Require Ice (Immediate Cooling)

↑ potassium, PO4, Fe, magnesium, ALT, AST, LD, ALP, catecholamines, CK

Hemolysis

(marked hemolysis)

: method of determining the concentration of substance in solution by measuring the amount of light absorbed by that solution after appropriate treatment.

SPECTROPHOTOMETRY

- photons travelling in waves

Electromagnetic radiation

- distance between two peaks

Wavelength

- distance between peak and trough

Amplitude

Visible light:

400-700nm

states that the concentration of a substance is directly proportional to the amount of light absorbed or inversely proportional to the logarithm of the transmitted light

BEER-LAMBERT LAW (BEER'S LAW)

Light Source = most common source of light for work in the visible and near-infrared regions

a. Incandescent tungsten or tungsten-iodide lamp

= most commonly used for ultraviolet (UV) work

Deuterium - discharge lamp & Mercury - arc lamp

= most commonly used Monochromator

Diffraction gratings

= advantage over round cuvets in that there is less error from the lens effect, orientation in the spectrophotometer, and refraction.

Square

= used for applications in the visible range

Glass

= for applications requiring UV radiation.

Quartz

= least expensive

Barrier - layer cell or photo cell

= used in instruments designed to be extremely sensitive to very low light levels and light flashes of very short duration

Photomultiplier (PM) tube

PRINCIPLE: measures the quantity of light reflected by a liquid sample that has been dispensed onto a grainy or fibrous solid support

REFLECTOMETRY

COMPONENTS are very similar to those of a photometer

REFLECTOMETRY

APPLICATION: urine dipstick analysis dry slide chemical analysis

REFLECTOMETRY

PRINCIPLE: measurement of concentration is done by detecting the absorption of electromagnetic by atoms rather than molecules. When a ground-state atom absorbs light energy, an excited atom is produced. The excited atom ther returns to the ground state, emitting light of the same energy as it absorbed.

ATOMIC ABSORPTION SPECTROPHOTOMETRY

COMPONENTS Hollow-cathode lamp Flame Monochromator

ATOMIC ABSORPTION SPECTROPHOTOMETRY

- usual light source

• Hollow-cathode lamp

- breaks chemical bonds and form free, unexcited atoms; serves as sample cells (instead of a cuvet)

Flame

- used to isolate the desired wavelength; also protects photodetector from excessive light emanating from flame emissions.

Monochromator

APPLICATION: measurement of unexcited trace metals e.g. calcium and magnesium

ATOMIC ABSORPTION SPECTROPHOTOMETRY

APPLICATION: measurement of unexcited trace metals e.g. calcium and magnesium

ATOMIC ABSORPTION SPECTROPHOTOMETRY

PRINCIPLE: measurement of light emitted by excited atoms

FLAME PHOTOMETRY

APPLICATION: Widely used before to determine the concentration of Na*, K+ or Lit

FLAME PHOTOMETRY

PRINCIPLE: measurement of the concentration of solutions that contain fluorescing molecules

FLUOROMETRY

COMPONENTS Xenon lamp - most common light source

FLUOROMETRY

APPLICATION: is used to measure small particles, such as drugs.

FLUOROMETRY

PRINCIPLE: Chemical energy generated in a chemiluminiscent reaction produces excited intermediates that decay a ground state with the emission of photons; no excitation is required unlike in fluorometry

CHEMILUMINESCENCE

PRINCIPLE: measurements are made with a spectrophotometer to determine concentration of particulate matter in sample. The amount of light blocked by a suspension of particles depends not only on concentration but also on size.

TURBIDIMETRY

APPLICATIONS 1. Detection of bacterial growth and bacterial culture 2. Antibiotic sensitivity 3. Coagulation studies 4. Protein concentration in CSF and urine

TURBIDIMETRY

PRINCIPLE: light scattered by small particles is measured at an angle to the beam incident to the cuvet

NEPHELOMETRY

measure particles which are too large for spectrophotometry, such as antibody-antigen complexes formed in enzyme immunoassays.

NEPHELOMETRY

PRINCIPLE: involves measurement of the current or voltage generated by the activity of specific ions. techniques include potentiometry, coulometry, voltammetry, and amperometry.

ELECTROCHEMISTRY

Measurement of potential (voltage) between two electrodes in a solution to measure analyte concentration

Potentiometry

pH, pCO2, Nat, Ca?*, K*, NH4*

Potentiometry

measurement of the current flow produced by an oxidation-reduction reaction

Amperometry

pOz (Clark electrode), glucose, peroxidase

Amperometry

Electrochemical titration in which the titrant is electrochemically generated

Coulometry

Potential is applied to an electrochemical cell and the resulting current is measured

Voltammetry

Anodic stripping voltametry (for lead and iron)

Voltammetry

PRINCIPLE: separation of charged compounds based on their electrical charge

ELECTROPHORESIS

COMPONENTS 1. A driving force (electrical power) 2. Support more (electrical power) a. Filter paper b. Agarose c. Cellulose acetate d. Polyacrylamide 3. Buffer 4. Sample 5. Detecting system

ELECTROPHORESIS

: movement of buffer ions and solvent relative to the fixed support

ELECTROENDOSMOSIS

- most common and reliable way for quantitation of separated protein fractions

DENSITOMETRY

PRINCIPLE: separation of complex mixtures on the basis of different physical attractions between the individual compounds and the stationary phase of the system

CHROMATOGRAPHY

COMPONENTS 1. Mobile phase (gas or liquid) 2. Stationary phase (solid or liquid) 3. Column 4. Eluate

CHROMATOGRAPHY

: carries the complex mixture

1. Mobile phase (gas or liquid)

: substance through which the mobile phase flows

2. Stationary phase (solid or liquid)

: holds the stationary phase

3. Column

: separated components

4. Eluate

CHROMATOGRAPHIC PROCEDURES

1. Thin-Layer Chromatography 2. High-Performance Liquid Chromatography (HPLC) 3. Gas Chromatography (GC)

- uses pressure for faster separations

2. High-Performance Liquid Chromatography (HPLC)

- separate mixtures of compounds that are volatile or can be made volatile

3. Gas Chromatography (GC)

PRINCIPLE: Sample in a MS is first volatilized and then ionized to form charged molecular ions and fragments that are separated according to their mass-to-charge (m/Z) ratio

MASS SPECTROMETRY

Allows definitive identification when used on samples eluting from GC or HPLC

MASS SPECTROMETRY

Gold standard for drug testing when coupled with GC

MASS SPECTROMETRY

PRINCIPLE Two-step procedure: (1) MALDI, then (2) Time-of-Flight (TOF) Mass

MALDI-TOF MS (Matrix-Assisted Laser Desorption lonization Time-of-Flight) analysis

• A laser pulse irradiates the sample, causing desorption and ionization of both the matrix and the sample.

MALDI-TOF MS (Matrix-Assisted Laser Desorption lonization Time-of-Flight) analysis

• lons from the sample are focused into the mass spectrometer.

MALDI-TOF MS (Matrix-Assisted Laser Desorption lonization Time-of-Flight) analysis

The molecular weight of the proteins acquired by mass spectrometry is used to determine the identity of the sample and is helpful in determining posttranslational modifications that may have occurred.

MALDI-TOF MS (Matrix-Assisted Laser Desorption lonization Time-of-Flight) analysis

APPLICATION: used for the analysis of biomolecules, such as peptides and proteins

MALDI-TOF MS (Matrix-Assisted Laser Desorption lonization Time-of-Flight) analysis

- process by which lab ensures quality results by closely monitoring preanalytical, analytical, & postanalytical stages of testing.

QUALITY ASSURANCE

- everything that precedes test performance,

• Preanalytical

e.g., test ordering, patient preparation, patient ID, specimen collection, specimen transport, specimen processing.

• Preanalytical

- everything related to assay,

• Analytical

e.g., test analysis, quality control (QC), reagents, calibration, preventive maintenance.

• Analytical

- everything that comes after test analysis, e.g., verification of calculations & reference ranges, review of results, notification of critical values, result reporting, test interpretation by physician, follow-up patient care.

• Postanalytical

- part of analytical phase of quality assurance

QUALITY CONTROL (QC)

process of monitoring results from control samples to verify accuracy of patient results.

QUALITY CONTROL (QC)

- most frequently used measure of variation

b. Standard deviation (SD)

- an index of precision used to compare the dispersion of two or more groups of data with different units / concentrations

c. Coefficient of variation (CV)

- used to determine if there is a significant difference between the MEANS of two groups of data

T-test

- used to determine if there is a significant difference between the SD of two groups of data

F-test

describes many continuous laboratory variables and deviation shares several unique characteristics

The Gaussian Distribution (Normal Distribution)

the mean, median, and mode are [?]; the distribution is [?] — meaning half the values fall to the left of the mean, and the other half fall to the right (the symmetrical shape is often referred to as a "bell curve.") The total area under the gaussian curve is [?].

identical symmetric 1.0, or 100%

summarizes the above relationships between the area under a Gaussian distribution and the SD.

"68-95-99 Rule"

"68-95-99 Rule" In other words, given any Gaussian distributed data of the data fall between ‡1 SD from the mean of the data fall between +2 SDs from the mean fall between ‡3 SDs from the mean

68% 295% 99%

- nearness or closeness of assayed values to the true value

1. Accuracy

- nearness or closeness of assayed values to each other

2. Precision (Reproducibility)

- ability of an analytical method to maintain accuracy and precision over an extended period of time

3. Reliability

- degree by which a method can easily be repeated

4. Practicability

- ability to measure the smallest concentration of the analyte of interest

5. Analytical sensitivity

- ability to measure only the analyte of interest

6. Analytical specificity

- also known as linearity; range of values over which lab can verify accuracy of test system

7. Reportable range

Formerly called normal value.

8. Reference interval

Can vary for different patient populations (age, gender, race).

8. Reference interval

Established by testing minimum of 120 healthy subjects & determining range in which 95% fall.

8. Reference interval

Verifying a reference interval (transference) can required as few as 20 study individuals

8. Reference interval

Reporting a positive result in a patient who has the disease

True positive (TP)

Reporting a positive result in a patient who doesn't have the disease

False positive (FP)

Reporting a negative result in a patient who doesn't have the disease

True negative (TN)

Reporting a negative result in a patient who has the disease

False negative (FN)

% of population with the disease that test positive

Diagnostic sensitivity

ability of the analytical method to detect the proportion of individuals with the disease

Diagnostic sensitivity

% of population without the disease that test negative

Diagnostic specificity

ability of the analytical method to detect the proportion of individuals without the disease

Diagnostic specificity

Number individuals without the disease with a negative test × 100%

Diagnostic Specificity (%)

% of time that a positive result is correct

Positive predictive value (PPV)

% of time that a negative result is correct

Negative predictive value (NPV)

• Assayed on a regular schedule to verify that a laboratory procedure is performing correctly

QC SAMPLES

• Generally, two different concentrations are necessary for adequate statistical QC

QC SAMPLES

• Chemically & physically similar to unknown specimen & is tested in exactly the same manner

QC SAMPLES

• New instrument or new lot of reagents: analyze QC materials for 20 days

QC SAMPLES

CHARACTERISTICS OF IDEAL QC MATERIALS

1. Must resemble human samples 2. Inexpensive and stable for long periods 3. No communicable disease 4. No known matrix effects 5. With known analyte concentrations (for assayed controls) 6. Convenient packaging for easy dispensing and storage

Also called a Shewart plot

LEVEY-JENNINGS CONTROL CHART

• Most common presentation for evaluating QC results

LEVEY-JENNINGS CONTROL CHART

shows each QC result sequentially over time

LEVEY-JENNINGS CONTROL CHART

- control values increasing or decreasing for six consecutive runs

Trend

Trend Main cause:

DETERIORATION OF REAGENTS

- six consecutive control values on the same side of the mean

Shift

Shift Main cause:

IMPROPER CALIBRATION OF INSTRUMENT

- highly deviating values

Outliers

control result outside established limits

Outliers

1 control >‡ 2s from mean.

1(2S)

1 control >‡ 2s from mean.

1(2S)

Warning flag of possible change in accuracy or precision.

1(2S)

Initiates testing of other rules (warning rule). If no violation of other rules, run is considered in control.

1(2S)

1 control >‡ 3s from mean

1(3S)

2 consecutive controls >2s from mean on same side

2(2S)

Random; Rejection rule

Systematic; Rejection rule

2 consecutive controls differ by >4s

R(4S)

4 consecutive controls > 1s from mean on same side

4(1S)

10 consecutive controls on same side of mean

10x

Present in all measurements; due to chance; no means of predicting it

1. Random error

Error that doesn't recur in regular pattern

1. Random error

Associated with violations of the 12s, 13s and R4s Westgard rules

1. Random error

• Error that influences ALL observations consistently in one direction

2. Systematic error

Recurring error inherent in test procedure

2. Systematic error

• Associated with violations of the 22s and 41s Westgard rules

2. Systematic error

• Also known as external quality assessment

PROFICIENCY TESTING

• consists of evaluation of method performance by comparison of results versus those of other

PROFICIENCY TESTING

• laboratories for the same set of samples

PROFICIENCY TESTING

PROFICIENCY TESTING • Basic procedure:

PT providers circulate a set of samples among a group of laboratories. Each laboratory includes the PT samples along with patient samples in the usual assay process. Results for the PT samples are reported to the PT provider for evaluation.

Error due to dirty glassware

RANDOM ERRORS

Dirty photometer

SYSTEMATIC ERRORS

Use of wrong pipet

RANDOM ERRORS

Faulty ISE

SYSTEMATIC ERRORS

Voltage fluctuation

RANDOM ERRORS

Evaporation or contamination of standards or reagents

SYSTEMATIC ERRORS

Sampling error

RANDOM ERRORS

Anticoagulant or drug interference

RANDOM ERRORS

• Comparison of patient data with previous results.

Delta checks

• Detects specimen mix-up & other errors.

Delta checks

• When limit is exceeded, must determine if due to medical change in patient or lab error.

Delta checks

Test results that indicate a potentially life-threatening situation.

Critical values

List typically includes glucose, Na+, K+, total CO2, Ca2+, Mg2+, phosphorus, total billrubin (neonates), blood gases

Critical values

• Patient care personnel must be notified immediately.

Critical values

Critical values• Example: Serum glucose

<40 mg/dL >500 mg/dL

Schedule of maintenance to keep equipment in peak operating condition; must be documented & must follow manufacturer' s specifications & frequencies.

Preventive maintenance

CARBOHYDRATES 1. Contain C, H and 0; Empiric formula: (CH20)n 3. Can be reducing or non-reducing sugars ; Can be classified according to the number of sugar units

CARBOHYDRATES

CARBOHYDRATES Functions: Major energy source (?) Storage form of energy e.g. [?] Components of cell membranes e.g. [?] Structural component in plants, bacteria, insects (e.g. ?)

glucose glycogen glycoproteins chitin, cellulose

- one sugar unit e.g. glucose, fructose, galactose - 2 sugar units linked together by a glycosidic bond e.g. sucrose, lactose, maltose - 3 to 10 sugar units - more than 10 sugar units (e.g. starch, glycogen, cellulose)

a. Monosaccharides b. Disaccharides Oligosaccharides d. Polysaccharides

Glucose + fructose

Sucrose-Sucrase

Glucose + galactose

Lactose-Lactase

Glucose + glucose

Maltose-Maltase

Metabolism of glucose molecule to pyruvate or lactate for production of energy

Glycolysis

Formation of glucose-6-phosphate from non-carbohydrate sources

Gluconeogenesis

Breakdown of glycogen to glucose for use as energy

Glycogenolysis

Conversion of glucose to glycogen for storage

Glycogenesis

Conversion of carbohydrates to fatty acids

Lipogenesis

Decomposition of fat

Lipolysis

Carbohydrates in the diet constitute about 50% of the calories in the average diet: - 60% - 30% C. - 5% - 5% (part of dietary fiber)

a. Starch and dextrins b. Sucrose C. Lactose d. Other sugars e. Cellulose

breakdown polymers to dextrins and disaccharides.

Salivary amylase (ptyalin) and pancreatic amylase (amylopsin)

are further hydrolyzed into monosaccharides by specific enzymes (disaccharidases)

Disaccharides

are absorbed by the gut via active transport (glucose and galactose) or facilitated diffusion (fructose).

Monosaccharides

They are then transported into the liver through the portal circulation.

Monosaccharides

is the only carbohydrate to be used directly for energy.

Glucose

After glucose enters the cell, it undergoes phosphorylation into glucose-6-phosphate through the action of

hexokinase or glucokinase.

Glucose-6-phosphate is then shunted into the following metabolic pathways:

a. Glycolysis (Embden-Meyerhof pathway) b. Glycogenesis c. Hexose-Monophosphate shunt

• Produced by the beta cells of the islets of Langerhans (pancreas)

Insulin

→ insulin

• Preproinsulin → proinsulin

• Target: most cells of the body

Insulin

Increases utilization of glucose by the cells by increasing cellular uptake and hepatic glycolysis

Insulin

Increases glycogenesis and inhibits glycogenolysis; Inhibits gluconeogenesis

Insulin

Stimulates lipogenesis while inhibiting lipolysis

Insulin

Stimulates protein synthesis and stimulates uptake of amino acids into muscles

Insulin

• Produced by the alpha cells of the islets of Langerhans

GLUCAGON

• Target: liver

GLUCAGON

• Target: liver

GLUCAGON

Promotes liver glycogenolysis

GLUCAGON

Increases gluconeogenesis

GLUCAGON

Inhibits glycolysis

GLUCAGON

Increases gluconeogenesis

Cortisol (Glucocorticoids)

Decreases glucose uptake and utilization by extrahepatic tissues

Cortisol (Glucocorticoids)

Stimulates glycogenolysis

Catecholamines

Increases glucose absorption in the small intestines

Thyroid hormone

Inhibit glucagon and insulin secretion

Somatostatin

Increases liver gluconeogenesis

Growth hormone

Inhibits glucose transport

Growth hormone

- heterogeneous group of multifactorial, polygenic syndromes characterized by an elevated fasting blood glucose caused by a relative or absolute deficiency in insulin

DIABETES MELLITUS

- characterized by an absolute deficiency of insulin caused by an autoimmune attack on the beta cells of the pancreas

1. Type 1 DM

- characterized by a combination of insulin resistance and dysfunctional beta cells

2. Type 2 DM

Juvenile Onset DM; Insulin Dependent DM; Most common in children and young adults

TYPE 1 DIABETES

Adult Onset DM; Non-insulin Dependent DM; Most common with advancing age

TYPE 2 DIABETES

5-10%

TYPE 1 DIABETES

90-95%

TYPE 2 DIABETES

Genetic, autoimmune, environmental (e.g. viral infection)

TYPE 1 DIABETES

HLA DR3/4 ; Autoantibodies -Anti-islet cell cytoplasmic antibody -Insulin autoantibodies -Anti-GAD (glutamic acid decarboxylase)

TYPE 1 DIABETES

Genetic, obesity, sedentary lifestyle, race/ethnicity

TYPE 2 DIABETES

Destruction of pancreatic beta cells, usually autoimmune

TYPE 1 DIABETES

No autoimmunity; Insulin resistance and progressive insulin deficiency

TYPE 2 DIABETES

Very low or undetectable c-peptide

TYPE 1 DIABETES

Detectable c-peptide

TYPE 2 DIABETES

Low to absent plasma insulin

TYPE 1 DIABETES

High in early disease; low to absent in disease of long duration plasma insulin

TYPE 2 DIABETES

Prone to ketoacidosis and diabetic complications

TYPE 1 DIABETES

Not prone to ketoacidosis

TYPE 2 DIABETES

Insulin absolutely necessary; muliple

TYPE 1 DIABETES

Oral agents (insulin sometimes daily injections or insulin pump indicated)

TYPE 2 DIABETES

None known therapy

TYPE 1 DIABETES

Lifestyle, oral medicines

TYPE 2 DIABETES

- associated with secondary conditions E.g. genetic defects of beta cell function; pancreatic disease; endocrine disease; drug or chemical induced; insulin receptor abnormalities; other genetic syndromes

Other specific types of DM

Glucose intolerance with onset or first recognition during pregnancy

Gestational Diabetes Mellitus (GDM)

• Due to metabolic and hormonal changes

Gestational Diabetes Mellitus (GDM)

• Large % of patients develop DM Within 5 to 10 years

Gestational Diabetes Mellitus (GDM)

• Infants born to mothers with diabetes are at increased risk for RDS, hypocalcemia, hyperbilirubinemia and other complications

Gestational Diabetes Mellitus (GDM)

• Screening: 2-hour OGTT using a 75 g glucose load

Gestational Diabetes Mellitus (GDM)

Random plasma glucose

≥200 mg/dL (211.1 mmol/L), +symptoms of DM

Fasting plasma glucose

≥126 mg/dL (27.0 mmol/L)

Two-h plasma glucose

≥200 mg/dL (≥11.1 mmol/L)

N.B. In absence of unequivocal hyperglycemia, these criteria should be confirmed by repeat testing on a different day. The third measure (OGTT) is not recommended for routine clinical use.

Normal Fasting plasma glucose

<100 mg/dL <5.6 mmol/L

Normal 2-hour plasma glucose level (after 75 g load)

<140 mg/dL <7.8 mmol/L

Pre-diabetes HbA1c

5.7-6.4 %

Impaired fasting glucose Fasting plasma glucose

100-125 mg/dL 5.6-6.9 mmol/L

Impaired glucose tolerance 2-hour plasma glucose level (after 75 g load)

140-199 mg/dL 7.8-11.0 mmol/L

Diabetes mellitus Fasting plasma glucose

≥126 mg/dL ≥7.0 mmol/L

Diabetes mellitus 2-hour plasma glucose level (after 75 g load)

≥200 mg/dL ≥11.1 mmol/L

Diabetes mellitus HbA1c

≥6.5%

Use: detection of GDM

ORAL GLUCOSE TOLERANCE TEST (OGTT)

Before an OGTT is performed, individuals should ingest [?] preceding the test.

At least 150g/day of carbohydrates for the 3 days (no restriction of diet)

No limitation in physical activity

ORAL GLUCOSE TOLERANCE TEST (OGTT)

Test should be performed after an overnight 8- to 14-hour fast.

ORAL GLUCOSE TOLERANCE TEST (OGTT)

The individual should not eat food, drink tea, coffee, or alcohol, or smoke cigarettes during the test, and should be seated.

ORAL GLUCOSE TOLERANCE TEST (OGTT)

Venous glucose samples are preferably collected in gray-top tubes containing fluoride and an anticoagulant (Henry's, 23rd ed)

ORAL GLUCOSE TOLERANCE TEST (OGTT)

FBG is measured right before the administration of the glucose load. A FBG of greater than 140 mg/dL necessitates that the test be stopped immediately. Proceed with the glucose load if FBG is less than 140 mg/dL.

ORAL GLUCOSE TOLERANCE TEST (OGTT)

ORAL GLUCOSE TOLERANCE TEST (OGTT) • Glucose load for adults • Glucose load for children • Pregnant women

75 g 1.75g/kg bw (max: 75 g) 75 g or 100 g

Patient should finish glucose load within 5 - 15 minutes.

ORAL GLUCOSE TOLERANCE TEST (OGTT)

ORAL GLUCOSE TOLERANCE TEST (OGTT) Patient should NOT vomit. If patient vomits, [?]

Discontinue the test

DIAGNOSIS OF GESTATIONAL DIABETES MELLITUS Gestational diabetes mellitus is diagnosed if [?] plasma glucose levels are exceeded (American Diabetes Association, 2004)

≥2

• Rate of formation of Hbac is proportional to the average blood glucose concentration over the previous 3 months

GLYCOSYLATED HEMOGLOBIN (HbA1c)

• For every 1% increase in Hba1c, there is a corresponding 35mg/dL change in plasma glucose

GLYCOSYLATED HEMOGLOBIN (HbA1c)

• The ADA also recommends that it be tested at least twice a year to monitor long-term glycemic control.

GLYCOSYLATED HEMOGLOBIN (HbA1c)

• Spn: EDTA-WB → hemolysate

GLYCOSYLATED HEMOGLOBIN (HbA1c)

• False decrease: decreased RBC lifespan

GLYCOSYLATED HEMOGLOBIN (HbA1c)

• Monitoring glucose control over past 2-3 weeks

FRUCTOSAMINE (Glycated Albumin)

• Albumin has a life span of 20 days in circulation

FRUCTOSAMINE (Glycated Albumin)

• Affected by albumin levels; false decrease in patients with hypoalbuminemia

FRUCTOSAMINE (Glycated Albumin)

100 g OGTT plasma glucose/75 g OGTT plasma glucose Fasting

≥95 mg/dL ≥5.3 mmol/L

100 g OGTT plasma glucose/75 g OGTT plasma glucose 1 hour

≥180 mg/dL ≥10.0 mmol/L

100 g OGTT plasma glucose/75 g OGTT plasma glucose 2 hour

≥ 155 mg/dL ≥8.6 mmol/L

100 g OGTT plasma glucose 3 hour

≥140 mg/dL ≥7.8 mmol/L

HYPOGLYCEMIA The plasma glucose concentration at which glucagon and other glycemic factors are released is between [?]. At about [?], observable symptoms of hypoglycemia appear. Warning signs and symptoms are all related to the central nervous system

65 and 70 mg/dL 50 to 55 mg/dL

HYPOGLYCEMIA Most causes are secondary to other illnesses and resolve themselves when the primary disorder is treated. Examples:

Insulinoma Various liver disorders Gastrointestinal disorders and surgery

HYPOGLYCEMIA Possible specimens =

WB, serum, plasma, urine, CSF, serous fluid, synovial fluid

HYPOGLYCEMIA Standard clinical specimen

Fasting venous plasma

HYPOGLYCEMIA Fasting blood sugar should be obtained after

8 - 10 hours of fasting

HYPOGLYCEMIA Whole blood glucose levels

10-15% lower vs plasma levels

HYPOGLYCEMIA Glucose is metabolized at room temperature at a rate of

7 mg/dl/hour

HYPOGLYCEMIA At 4°C, glucose decreases by approximately

2 mg/dl/hour

HYPOGLYCEMIA Evacuated tube

Gray top (NaF)

HYPOGLYCEMIA CSF glucose levels

60-70% of plasma levels (decreased in bacterial meningitis)

HYPOGLYCEMIA As little as 10% contamination with 5% dextrose (D5W) will elevate glucose in a sample by

500 mg/dL or more

Principle: Glucose and other carbohydrates are capable of converting cupric ions in alkaline solution to cuprous ions.

Chemical Methods

GLUCOSE: Chemical Methods

a. Oxidation-reduction method i. Alkaline Copper Reduction Method Folin-Wu Nelson-Somogyi Neocuproine method Benedict's method ii. Alkaline Ferric Reduction Method (Hagedorn-Jensen) a. Condensation method (Dubowski)

GLUCOSE: Enzymatic Methods

a. Glucose oxidase method i. Colorimetric method ii. Polarographic method b. Hexokinase method

is the most specific enzyme reacting with only B-D-glucose

Glucose oxidase

- uses a side reaction that consumes H202

i. Colorimetric method

- measure the rate of disappearance of oxygen using an oxygen electrode

ii. Polarographic method

• More accurate than glucose oxidase methods because the coupling reaction using G6PDH is highly specific; therefore it has less interference than the coupled glucose oxidase procedure

b. Hexokinase method

• NADPH has a strong absorbance at 340 nm

b. Hexokinase method

Generally accepted as the reference method

b. Hexokinase method

• Not affected by ascorbic acid or uric acid

b. Hexokinase method

Most common cause of lactose intolerance

Lactase deficiency

Lactose is not digested at a normal rate and accumulates in the gut, where it is metabolized by bacteria. Bloating, abdominal cramps, and watery diarrhea result

Lactase deficiency

result of the deficiency of a specific enzyme that causes an alternation of glycogen metabolism

Glycogen storage diseases

Glycogen storage diseases Most common congenital form:

Von Gierke Disease

i. Autosomal recessive disease

Von Gierke Disease

ii. Characterized by hepatomegaly, severe hypoglycemia, metabolic acidosis, ketonemia, and elevated lactate and alanine

Von Gierke Disease

• Building blocks of lipids

FATTY ACIDS

• Hydrocarbon chains with a terminal COO- group

FATTY ACIDS

• 3 fatty acid molecules attached to one molecule of glycerol by ester bonds

TRIGLYCERIDES

• Serves as main storage form of energy, insulator, shock absorber and integral part of cell membrane

TRIGLYCERIDES

• Similar to triglycerides except that the third position on the glycerol backbone contains a phospholipid head group

PHOSPHOLIPIDS

• Contains polar and non-polar end

PHOSPHOLIPIDS

• Constituent of cell membranes

PHOSPHOLIPIDS

• Serves as part of cell membranes and as parent chain for cholesterol-based hormones, e.g. aldosterone, cortisol and the sex hormones

CHOLESTEROL

CHOLESTEROL• Exists in two forms - approximately 70% of total cholesterol - approximately 30% of total cholesterol

• Cholesterol esters • Free cholesterol

Typically spherical in shape with sizes ranging from 10 to 1200 nm

LIPOPROTEIN

Composed of lipids and proteins, called apolipoproteins

LIPOPROTEIN

Size particle correlates with its lipid content

LIPOPROTEIN

originally separated through ultracentrifugation

LIPOPROTEIN

• located on the surface of lipoprotein particles maintain structural integrity of lipoproteins

Apolipoproteins

• serve as ligands for cell receptors

Apolipoproteins

Apolipoproteins• Important types:

- largest -least dense - highest TG content - postprandial turbidity - fxn: transports exogenous / dietary triglycerides

CHYLOMICRONS

- 2nd largest -2nd least dense - 2nd highest TG content - fasting hyperlipidemic turbidity - fxn: transports endogenous/hepatic triglycerides

VERY LOW DENSITY LIPOPROTEIN (VLDL)

- small --> can cross BV walls --> deposition of lipid - highest cholesterol content - fxn: transports cholesterol to peripheral tissues →inc LDL --> in atherosclerosis - target for cholesterol lowering therapy

LOW DENSITY LIPOPROTEIN (LDL)

- smallest but densest - highest protein content - fxns: reverse transport cholesterol (peripheral tissues --> liver) - inc HDL --> dec atherosclerosis

HIGH DENSITY LIPOPROTEIN (HDL)

• Chylomicrons accumulate as a floating "cream" layer and can be detected visually. The presence of chylomicrons in fasting plasma is considered to be abnormal.

Standing Plasma Test

• A plasma sample that remains turbid after standing overnight contains excessive amounts of VLDL; if a floating "cream" layer also forms, chylomicrons are present as well. (Henry, 23rd ed)

Standing Plasma Test

Floating Beta-lipoprotein

1. Beta-VLDL

Increased in familial dysbetalipoproteinema

1. Beta-VLDL

• Sinking pre-beta lipoprotein

2. Lp(a)

• LDL - like particle

2. Lp(a)

Increased risk of premature coronary heart disease and stroke

2. Lp(a)

Seen in patient with biliary cirrhosis or cholestasis and in patients with mutations in the enzyme lecithin: cholesterol acyltransferase (LCAT)

3. Lpx

happens in the intestines

Absorption Pathway

CM transports exogenous TG

Exogenous Pathway

VLDL and hDL transports endogenous TG

Endogenous Pathway

LDL transports Cholesterol

Reverse Cholesterol Transport Pathway

was used to characterize lipid disorders; used electrophoresis and a standing plasma test for CM to correlate clinical disease syndromes with laboratory phenotypes. Note that each phenotype is not a specific disease but rather a variety of disorders that affect the same lipoproteins and therefore express the same lipid pattern.

FREDRICKSON CLASSIFICATION OF LIPID DISORDERS

-Hyperchylomicronemia -Familial LPL deficiency

Type 1

-Familial Hypercholesterolemia

Type 2a

-Familial Combined Hyperlipidemia

Type 2b

Familial Dysbetalipoproteinemia

Type 3

Familial Hypertriglyceridemia

Type 4

Low cardiac risk; eruptive xanthoma; recurrent pancreatitis

Type 1

High cardiac risk; xanthelasma; tendon xanthoma; corneal arcus; hypothyroidism and nephrotic syndrome

Type 2a

High cardiac risk

Type 2b

Eruptive and palmar xanthomas

Type 3

Low cardiac risk

Type 4

Low cardiac risk; eruptive xanthoma, may be associated with pancreatitis

Type 5

Fasting = 12 hours before venipuncture

LIPIDS AND LIPOPROTEINS

• Can be measured non-fasting = TC and HDL-C

LIPIDS AND LIPOPROTEINS

• Prolonged tourniquet application = causes hemoconcentration

LIPIDS AND LIPOPROTEINS

Reclined patients = decreased values

LIPIDS AND LIPOPROTEINS

Preferred sample = Serum or plasma but plasma preferred in electrophoresis and ultracentrifugation

LIPIDS AND LIPOPROTEINS

Capillary blood samples = generally lower values

LIPIDS AND LIPOPROTEINS

Lipemic samples = seen when triglyceride levels exceed 4.6 mmol/L (400 mg/dL).

LIPIDS AND LIPOPROTEINS

Initial extraction with zeolite to remove sterols

1. Abell-Kendall method

Redissolving of cholesterol

1. Abell-Kendall method

Hydrolysis of cholesterol esters to cholesteriol

1. Abell-Kendall method

Liebermann-Burchard reagent

glacial acetic acid sulfuric acid acetic anhydride

(+) formation of product which strongly absorbs at 410 nm

1. Abell-Kendall method

Recently, the reference method has changed to a [?] that now specifically measures cholesterol and does not detect related sterols. (Bishop 7h ed.)

GC-MS method

- definitive method

3. Isotope Dilution Mass Spectrometry (IDMS)

Hydrolysis of glycerol is accomplished using alcoholic KOH

2. Chemical methods (triglyceride)

Oxidation of glycerol by periodic acid, forming formaldehyde and formic acid

2. Chemical methods (triglyceride)

Formaldehyde combines with a variety of reagents:

2. Chemical methods (triglyceride)

Reagent: Chromotropic acid → blue colored compound

a. Van Handel & Zilversmit (Colorimetric method)

Reagent: acetylacetone (aka diacetyl acetone; reactant of choice)

Hantzsch (Fluorometric method)

Product has a strong absorption maximum at 412 nm and also has good fluorescence

Hantzsch (Fluorometric method)

- new reference method for triglyceride measurements; involve the hydrolysis of fatty acids on triglycerides and the measurement of glycerol.

GC-MS method

: range in density observed among lipoprotein classes is a function of lipid and protein content and enables fractionation by density using ultracentrifugation

1. Ultracentrifugation

: takes advantage of differences in size and charge

2. Electrophoresis

: depends on particle size, charge and differences in the apolipoprotein content; primarily used in research labs only

3. Chemical precipitation

Uses polyanions (heparin sulfate, dextran sulfate and phosphotungstate) and divalent cations such as magnesium, calcium andmanganese e.g. HDL - dextran sulfate + magnesium

3. Chemical precipitation

: uses antibodies specific to apolipoproteins to bind and separate lipoprotein classes

4. Immunoassays

: takes advantage of size differences in molecular sieving methods or composition in affinity methods e.g. gel chromatography or affinity chromatography

5. Chromatographic methods

• The term [Plasma TG]/5 is used when concentrations are expressed in mg/dL.

K. FRIEDEWALD CALCULATION

It has been reported that the factor [Plasma TG]/2.825 gives a more accurate estimate of VLDL-C (DeLong, 1986). This is equivalent to Plasma TG/6.5, when concentrations are expressed in mg/dL.

K. FRIEDEWALD CALCULATION

a. Involves large- and medium-sized arteries (e.g. abdominal aorta, coronary artery, popliteal artery, internal carotid artery)

1. Atherosclerosis

b. LDL - increased risk ; HDL - decreased risk

1. Atherosclerosis

c. Complications Narrowing of blood vessels result in impaired blood flow and ischemia leading to: (i) Peripheral vascular disease (ii) Angina (iii) Ischemic bowel disease

1. Atherosclerosis

Plaque rupture → thrombosis → myocardial infarction and stroke Plaque rupture → embolization atherosclerotic embolism Weakening of blood vessel wall results in aneurysm

1. Atherosclerosis

An extreme form of hypoalphalipoproteinemia (isolated decrease in circulating HDL)

2. Tangier Disease

Associated with HDL cholesterol concentrations as low as 1-2 mg/dL (0.03-0.05 mmol/L) in homozygotes, accompanied by total cholesterol concentrations of 50 to 80 mg/dL (1.3-2.1 mmol/L).

2. Tangier Disease

Associated with increased risk of premature coronary heart disease (CHD).

2. Tangier Disease

Linear polymers of amino acids; Perform diverse functions

PROTEIN

Regulate metabolism

PROTEIN

• Facilitate contraction in the muscle

PROTEIN

Provide structural framework

PROTEIN

Shuttle molecules in the bloodstream

PROTEIN

• Component of the immune system

PROTEIN

: determined by amino acid sequence : folding of short segments of polypeptide into geometrically ordered units (e.g. alpha-helix, beta-sheet) : overall 3-dimensional shape of the protein (globular vs fibrous) : number and types of polypeptide units of oligomeric proteins and their spatial arrangement

Primary Secondary Tertiary Quaternary

Indicator of malnutrition; binds thyroid hormones and retinol-binding protein

Prealbumin (Transthyretin)

Binds bilirubin, steroids, fatty acids; major contributor to oncotic pressure

Albumin

Protease inhibitor

Alpha-1-antitrypsin

Principal fetal protein

Alpha-1-fetoprotein

May be related to immune response

Alpha-1- acid glycoprotein

Binds hemoglobin Transports copper; peroxidase activity

Haptoglobin Ceruloplasmin

Inhibits thrombin, trypsin and pepsin

Alpha-2-macroglobulin

Transports iron Binds heme Immune response Precursor of fibrin Opsonin

Transferrin Hemopexin Complement Fibrinogen C-reactive protein

1. Most plasma proteins are synthesized in the [?] and secreted by the hepatocyte into the circulation. The immunoglobulins are exceptions because they are synthesized in plasma cells.

Liver (hepatocytes)

2. The nitrogen content of serum proteins is, on average,

16%

: site of protein synthesis within the cell

3. Ribosomes

• Aka Transthyretin • Migrates before albumin in the serum protein electrophoresis

1. PREALBUMIN

• Function: Transport protein for thyroid hormones; transports vitamin A by forming a complex with retinol-binding protein

1. PREALBUMIN

Decreased in hepatic damage, acute-phase inflammatory response, and tissue necrosis

1. PREALBUMIN

- A low prealbumin level is a sensitive marker of poor nutritional status

1. PREALBUMIN

Increased in patients receiving steroids, in alcoholism, and in chronic renal failure

1. PREALBUMIN

• Protein present in the highest concentration in serum

2. ALBUMIN

Provide nearly 80% of colloid osmotic pressure (COP) of intravascular fluid

2. ALBUMIN

Buffers pH

2. ALBUMIN

Binds to various substances in blood (e.g. some hormones, drugs, electrolytes, unconjugated bilirubin)

2. ALBUMIN

Negative acute-phase reactant

2. ALBUMIN

Decreased: liver disease, malnutrition, malabsorption, kidney loss, hemodilution

2. ALBUMIN

Increased: dehydration

2. ALBUMIN

• Most important function: inhibition of the protease neutrophil elastase

3. ALPHA-1-ANTITRYPSIN

• Abnormal form of AAT can also accumulate in the liver and cause cirrhosis

3. ALPHA-1-ANTITRYPSIN

• Major component of a1-globulin band → deficiency of AAT seen as lack of an a1-globulin band on SPE

3. ALPHA-1-ANTITRYPSIN

One of the COPs (chronic obstructive pulmonary diseases)

Emphysema

Most common cause: smoking

Emphysema

• Pathophysiology: excessive inflammation or lack of AAT leads to destruction of alveolar air sacs → loss of elastic recoil and collapse of airways during exhalation → obstruction and air trapping

Emphysema

• Dyspnea, cough with minimal sputum

Emphysema

"Pink puffers", "barrel-chest", hypoxemia

Emphysema

Synthesized by the developing embryo and fetus; thought to protect the fetus from immunologic attack by the mother

ALPHA-1-FETOPROTEIN

• No known function in normal adults

ALPHA-1-FETOPROTEIN

: neural tube defects (e.g. spina bifida), presence of twins : increased risk for Down syndrome (trisomy 21)

Elevated AFP Low AFP

- Tumor marker for hepatocellular carcinoma, some testicular carcinomas

AFP

• Copper-containing (contains >90% of total serum copper)

CERULOPLASMIN

• Used in the diagnosis of Wilson's disease

CERULOPLASMIN

• Autosomal recessive • Decreased levels of ceruloplasmin

Wilson's disease

Wilson's disease • Excess storage of copper in various organs Liver → - Brain → - Cornea →

hepatic cirrhosis neurologic damage Kayser-Fleischer rings

• Large protein that inhibits proteases such as trypsin, thrombin, kallikrein, and plasmin

ALPHA-2-MACROGLOBULIN

• Increased in nephrotic syndrome (large size aids in its retention)

ALPHA-2-MACROGLOBULIN

• Glomerular disorder characterized by proteinuria (>3.5 g/day)

Nephrotic syndrome

• Pathophysiology: Disruption of the electrical charges that produce the tightly fitting podocyte barrier resulting in massive loss of protein and lipids

Nephrotic syndrome

Nephrotic syndrome • Manifestations - pitting edema - increased risk of infection - due to the loss of anti-thrombin III - may result in fatty casts in the urine

- Hypoalbuminemia - Hypogammaglobulinemia - Hypercoagulable state - Hyperlipidemia and hypercholesterolemia

Function: bind free hemoglobin to prevent loss of hemoglobin and its constituent, iron, into the urine

HAPTOGLOBIN

Used primarl to holp detect and evaluate hemolylic anemia

HAPTOGLOBIN

Transports two molecules of ferric iron

TRANSFERRIN

• Negative acute-phase reactant

TRANSFERRIN

• Major component of the beta-globulin fraction

TRANSFERRIN

• Tested to determine cause of anemia (e.g. increased levels in IDA)

TRANSFERRIN

• Function: scavenge heme released or lost by the turnover of heme proteins such as hemoglobin → protect body from oxidative damage that free heme can cause

HEMOPEXIN

• Low levels are diagnostic of hemolytic anemia

HEMOPEXIN

• Precipitates with C substance, a polysaccharide of pneumococci

C-REACTIVE PROTEIN

• Functions in opsonization

C-REACTIVE PROTEIN

One of the first acute-phase proteins to rise in inflammatory disease

C-REACTIVE PROTEIN

• High or increasing amount of CRP suggests an acute infection or inflammation

C-REACTIVE PROTEIN

• Glycoprotein produced by fetal membranes responsible for the cellular adhesiveness of placenta and membranes to the decidua.

FIBRONECTIN

• Fetal fibronectin is produced at the boundary between the amniotic sac and the decidua (the lining of the uterus) and functions to maintain the adherence of the placenta to the uterus.

FIBRONECTIN

• Test for assessment of the risk for [PRE-TERM DELIVERY] in women between 24 to 35 weeks gestational age.

FIBRONECTIN

• Proteolytic fragments of collagen I formed during bone resorption

CROSS-LINKED C-TELOPEPTIDES

• CTX is a biochemical marker of bone resorption that can be detected in serum and urine.

CROSS-LINKED C-TELOPEPTIDES

Govern excitation-contraction coupling in muscle TROPONIN• Three subunits: Troponin T (cTnT), Troponin C (TnC), Troponin I (cTnl)

TROPONIN

is used as an AMI indicator because of specificity and early rise in serum concentration following AMI rises within 3-4 hours, peaks in 10-24 hours, returns to normal in 10-24 days. rises within 3-6 hours, peaks in 14-20 hours, and returns to normal in 5-10 days.

cTnT or cTnI cTnT cTnl

• Now known as the "gold standard" for diagnosis of MI

TROPONIN

• Heme-containing protein that binds oxygen with cardiac and skeletal muscle

MYOGLOBIN

• Levels are related to muscle mass and activity (reasonable sensitivity but poor specificity)

MYOGLOBIN

Increased in skeletal injuries, muscular dystrophy, and AMI

MYOGLOBIN

is released early in cases of AMI, rising in 1-3 hours and peaks in 5-12 hours, and returns to normal in 18-30 hours. However, it is not tissue specific. It is better used as a negative predictor in the first 2-4 hours following chest pain.

MYOGLOBIN

are neurohormones that affect body fluid homeostasis (through natriuresis and diuresis) and blood pressure

BRAIN NATRIURETIC PEPTIDE (BNP) AND N-TERMINAL-BRAIN NATRIURETIC PEPTIDE (NT-BNP)

BNP has become a popular marker for

CONGESTIVE EART FAILURE

are found in largest concentration in the left ventricular myocardium but are also detectable in atrial tissue as well as in the myocardium of the right ventricle.

NT-proBNP and BP

ELECTROPHORETIC PATTERNS OF SERUM PROTEINS

share the property of showing elevations in concentrations in response to stressful or inflammatory states that occur with infection, injury, surgery, trauma, or other tissue necrosis.

acute phase reactant proteins

They include AAT, orosomucoid, haptoglobin, ceruloplasmin, fibrinogen, serum amyloid A protein, and CRP. Others are Factor VIII, ferritin, lipoproteins, complement proteins, and immunoglobulins.

acute phase reactant proteins

Opsonization, complement activation

CRP

Removal of cholesterol

Serum amyloid A

Protease inhibitor

Alpha1-antitrypsin

Binds hemoglobin

Haptoglobin

Clot formation

Fibrinogen

Binds copper, oxidizes iron

Ceruloplasmin

Opsonization, lysis

С3

Complement activation

Mannose-binding protein

• Total protein level less than the reference interval

Hypoproteinemia

• occurs in any condition where a negative nitrogen balance exists (excessive loss, decreased intake, decreased synthesis, accelerated catabolism)

Hypoproteinemia

• Increase in total plasma proteins

Hyperproteinemia

• Not an actual disease state but is the result of dehydration

Hyperproteinemia

When excess water is lost from the vascular system, the proteins, because of their size, remain within the blood vessels

Hyperproteinemia

May also be cause by excessive production, primarily of gamma-globulins e.g. multiple myeloma

Hyperproteinemia

Digestion of protein; measurement of nitrogen content

Kjeldahl

Reference method; assume average nitrogen content of 16%

Kjeldahl

Measurement of refractive index due to solutes in serum

Refractometry

Rapid and simple; assume nonprotein solids are present in same concentration as in the calibrating serum

Refractometry

Formation of violet-colored chelate between Cu?+ ions and peptide bonds

Biuret

Routine method; requires at least two peptide bonds and an alkaline medium

Biuret

Protein binds to dye and causes a spectral shift in the absorbance maximum of the dye

Dye binding

Research use

Dye binding

Globulins are precipitated in high salt concentrations; albumin in supernatant is quantitated by biuret reaction

Salt precipitation

Labor intensive

Salt precipitation

Albumin binds to dye; causes shift in absorption maximum

Methyl orange HABA [2,4'-hydroxyazobenzene)-benzoic acid] BCG (bromcresol green) BCP (bromcresol purple)

Nonspecific for albumin

Methyl orange

Many interferences (salicylates, bilirubin)

HABA [2,4'-hydroxyazobenzene)-benzoic acid]

Sensitive; overestimates low albumin levels; most commonly used dye

BCG (bromcresol green)

Specific, sensitive, precise

BCP (bromcresol purple)

Proteins separated based on electric charge

Electrophoresis

Accurate; gives overview of relative changes in different protein fractions

Electrophoresis

Performed when an abnormality in the total protein or albumin is found

Protein Electrophoresis

Principle: separation of proteins based on their charge density

Protein Electrophoresis

Regions are stained using: Coomassie Blue, Amido Black, Ponceau S

Protein Electrophoresis

Electrophoretic patterns:

Protein Electrophoresis

Probably the most significant finding from an electrophoretic pattern is (Bishop, 6th ed)

monoclonal immunoglobulin disease

Beta-gamma bridging: Monoclonal spike: ↑a2, ↑B, ↓albumin: ↓a1-antitrypsin: ↑B:

Cirrhosis Multiple myeloma Nephrotic Emphysema Inflammation

NPN present in the highest corientation in blood

UREA

Major excretory product of protein metabolism

UREA

• Following synthesis in the liver, urea is carried in the blood to the kidney, where it is readily filtered from the plasma by the glomerulus. Most of the urea in the glomerular filtrate is excreted in the urine.

UREA

• The concentration of urea in the plasma is determined by: i. renal function and perfusion ii. the protein content of the diet iii. rate of protein catabolism

UREA

• evaluate renal function

UREA

• assess hydration status aid in the diagnosis of renal disease

UREA

• verify frequency of dialysis

UREA

• Urea nitrogen concentration is converted to urea concentration by multiplying [?]

• Enzymatic methods are used most frequently in clinical laboratories.

UREA

UREA: Used on many automated instruments; best as kinetic measurement

GLDH coupled enzymatic

UREA: Used in automated systems, multilayer film reagents, and dry reagent strips

Indicator dye

UREA: Specific and rapid

Conductimetric

UREA: Proposed reference method

Isotope dilution mass spectrometry

An elevated concentration in the blood is called azotemia. Very high plasma concentration accompanied by renal failure is called uremia, or the uremic syndrome.

urea

This condition is eventually fatal if not treated by dialysis or transplantation.

uremia, or the uremic syndrome

Low protein intake Severe vomiting and diarrhea Liver disease Pregnancy

DECREASED UREA CONCENTRATION

Congestive heart failure Shock, hemorrhage Dehydration Increased protein catabolism High-protein diet

INCREASED UREA CONCENTRATION: Prerenal

Acute and chronic renal failure Renal disease, including glomerular nephritis, tubular necrosis

INCREASED UREA CONCENTRATION: Renal

Urinary tract obstruction

INCREASED UREA CONCENTRATION: Postrenal

1. Product of the catabolism of purines (guanine and adenosine)

URIC ACID

2. Filtered by the glomerulus and secreted by the distal tubules into the urine, but mostly reabsorbed in the proxima tubules and reused

URIC ACID

3. Relatively insoluble in plasma, and at high concentrations, can be deposited in the joints and tissue, causing painful inflammation; mostly present as monosodium urate in plasma, wherein it is insoluble at around pH 7

URIC ACID

• Assess inherited disorders of purine metabolism

URIC ACID

Confirm diagnosis and monitor the treatment of gout

URIC ACID

Assist in the diagnosis of renal calculi

URIC ACID

• Prevent uric acid nephropathy during chemotherapeutic treatment

URIC ACID

• Detect kidney dysfunction

URIC ACID

Liver disease

DECREASED URIC ACID CONCENTRATION

Defective tubular reabsorption (Fanconi syndrome)

DECREASED URIC ACID CONCENTRATION

Chemotherapy with azathioprine or 6-mercaptopurine

DECREASED URIC ACID CONCENTRATION

Overtreatment with allopurinol

DECREASED URIC ACID CONCENTRATION

Enzyme deficiencies Lesch - Nyhan Syndrome (hypoxanthine guanine phosphoribosyltransferase deficiency) Phosphoribosy|pyrophosphate synthetase deficiency Glycogen storage disease type I (glucose-6-phosphatase deficiency) Fructose intolerance (fructose-1-phosphate aldolase deficiency)

INCREASED URIC ACID CONCENTRATION

Gout Treatment of myeloproliferative disease with cytotoxic drugs Hemolytic and proliferative processes Chronic renal disease Toxemia of pregnancy Lactic acidosis Drugs and poisons Purine-rich diet Increased tissue catabolism or starvation

INCREASED URIC ACID CONCENTRATION

is formed from creatine (synthesized primarily in the liver from arginine, glycine and methionine) and creatine phosphate in muscle

Creatinine

Excreted in plasma at a constant rate related to muscle mass

CREATININE

Daily excretion is fairly stable thus it is commonly used to assess renal filtration function

CREATININE

Determine sufficiency of kidney function and severity of disease

CREATININE

Monitor the progression of kidney disease

CREATININE

Measure of completeness of 24 hour collections

CREATININE

Abnormal renal function

INCREASED CREATININE CONCENTRATION

Muscle disease: Muscular dystrophy Poliomyelitis Hyperthyroidism Trauma

INCREASED CREATININE CONCENTRATION

The normal BUN/CREATININE RATIO is [?] In prerenal disease, it rises to well [?] In true renal disease, both BUN and creatinine rise together, maintaining BUN/Creatinine ratio at [?] (Bishop, 6th ed.)

10:1 to 20:1. 10:1 to 20:1. 10-20:1.

Formed through the deamination of amino acids during protein metabolism

AMMONIA

Removed from the circulation and converted to urea in the liver

AMMONIA

Free ammonia is toxic; however ammonia is present in the plasma in low concentrations

AMMONIA

• Provide useful information on clinical conditions such as hepatic failure, Reye's syndrome and inherited deficiencies of the urea cycle enzymes

AMMONIA

Measurement of urine ammonia can be used to confirm the ability of the kidneys to produce ammonia

AMMONIA

Severe liver disease Inherited deficiencies of enzymes of the urea cycle

INCREASED AMMONIA CONCENTRATION

- protein catalysts; increase velocity of a chemical reaction without being consumed during the reaction they catalyze. This is achieved by decreasing the energy of activation (Ea) of a chemical reaction.

Enzymes

Majority are proteins so they can be denatured by certain agents (acids, strong bases, detergents, etc.)

Enzymes

Majority are proteins so they can be denatured by certain agents (acids, strong bases, detergents, etc.) Active site - site where substrate interacts with enzymes Allosteric site- site other than the active site Isoenzyme - multiple forms of an enzyme with different genetic origin Isoform - results when an enzyme is subject to post-translational modifications Cofactors - nonprotein molecule that must bind to particular enzymes for enzyme reactions to occur a. Activators - inorganic cofactors b. Coenzymes - organic cofactor (e.g. vitamins) Holoenzyme = apoenzyme + prosthetic group (a coenzyme tightly bound to its enzyme) Proenzyme / zymogen - inactive form IZYME CLASSIFICATION AND NOMENCLATURE The International Union of Biochemistry (IUB) Enzyme Commission categorized all enzymes into six (6) classes based on the catalytic activity of an enzyme:

Enzymes

are highly specific for their substrates and products. Many recognize only a single compound as a substrate.

Enzymes

- site where substrate interacts with enzymes - site other than the active site

Active site Allosteric site

- multiple forms of an enzyme with different genetic origin

Isoenzyme

- results when an enzyme is subject to post-translational modifications

Isoform

- nonprotein molecule that must bind to particular enzymes for enzyme reactions to occur

Cofactors

- inorganic cofactors - organic cofactor (e.g. vitamins)

a. Activators b. Coenzymes

= apoenzyme + prosthetic group (a coenzyme tightly bound to its enzyme)

Holoenzyme

- inactive form

Proenzyme / zymogen

categorized all enzymes into six (6) classes based on the catalytic activity of an enzyme

The International Union of Biochemistry (IUB) Enzyme Commission

- catalyze an oxidation-reduction reaction between two substrates

1. Oxidoreductases

- catalyze the transfer of a group other than hydrogen from one substrate to another

2. Transferases

- catalyze hydrolysis of various bonds

3. Hydrolases

- catalyze removal of groups from substrates without hydrolysis; the product contains double bonds

4. Lyases

- catalyze the interconversion of geometric, optical or positional isomers

5. Isomerases

- catalyze the joining of two substrate molecules, coupled with the breaking of the pyrophosphate bond in ATP or a similar compound

Ligases

- rate of reaction is almost directly proportional to substrate concentration at low level

FIRST-ORDER KINETICS

- rate of reaction is almost directly proportional to substrate concentration at low level

FIRST-ORDER KINETICS

• substrate is high enough to saturate all available enzymes reaction rate is unaffected by increase in substrate concentration

ZERO-ORDER KINETICS

• dependent on enzyme concentration only

ZERO-ORDER KINETICS

• when maximum velocity is reached, the rate of increase in velocity is "zero"

ZERO-ORDER KINETICS

pH a. most enzymes react at pH b. controlled through

7.0 - 8.0 buffer solutions

Temperature a. Increased temperatures increase b. For each 10°C, rate of reaction is c. Temperatures which are too high can

rate of reaction doubled denature proteins

- required for enzymatic activity but are not consumed in the process (unlike substrates)

Cofactors

- the presence of inhibitors can affect the reaction velocity

Inhibitors

- compete for the substrate at the active site of the enzyme and form an enzyme - inhibitor complex; competitive inhibition can be reversed by increasing substrate [S]

a. Competitive inhibitors

- bind to the enzyme or enzyme-substrate at a site distinct from the active site, decreasing the activity of the enzyme. Inhibition cannot be overcome by increasing substrate.

b. Non-competitive inhibitors

- inhibitor binds to the enzyme-substrate complex, preventing the formation of a product. Increasing substrate concentration further increases inhibition.

Uncompetitive inhibitors

: reaction is stopped then measurement is done : multiple measurements are done at different intervals

Fixed time Continuous monitoring or kinetic

1 International Unit: amount of enzyme that will catalyze the reaction of [?] 1 Katal: amount of enzyme that will catalyze the reaction of [?] 1 IU = [?]

1 umol of substrate in 1 minute 1 mole of substrate in 1 second 17nkat

HIGH SPECIFICITY - RBC & Prostate - Liver - Pancreas & Salivary glands - Pancreas MODERATE SPECIFICITY - Liver, Heart, Skeletal muscle - Heart, Skeletal muscle, brain - Liver, Bone, Kidney, Placenta LOW SPECIFICITY - all tissues

ACP ALT AMS LIPASE AST CK ALP LDH

HEART

CK-MB, AST, LD1>LD2

BONE

ALP

LIVER Hepatocellular disorders: Biliary tract obstruction:

AST, ALT, LD5 ALP, GGT

PANCREAS

Amylase Lipase

SKELETAL MUSCLE

CK-MM, AST, LD, Aldolase

BRAIN

СК-BB

PROSTATE

ACP

Widely distributed in skeletal muscle, brain and cardiac muscle

CREATINE KINASE

May be detected in nerve tissue, testicular tissue, amniotic fluid and in certain malignant tissues

CREATINE KINASE

exists as a dimer (subunits: M or B)

CREATINE KINASE

• Major form in sera of healthy people and in striated muscle

СК-ММ

• Increased in hypothyroidism, IM injections, mild to strenuous activity

СК-ММ

• Cardiac tissue contains significant amounts

СК-МВ

• Value in detection of AMI

СК-МВ

Rise within 4-8 hours, peak at 12-24 hours, return to normal within 48 to 72 hours

СК-МВ

Highest concentration in CNS

СК-ВВ

• Increased with extensive damage to the brain and carcinoma of various organs

СК-ВВ

Frequently elevated in disorders of cardiac and skeletal muscles

CREATINE KINASE

Sensitive indicator of AMI and Duchenne-type muscular dystrophy

CREATINE KINASE

Separation of total CK into its various isoenzyme fractions is considered a more specific indicator of various disorders than total levels:

CREATINE KINASE

• Optimum pH: 9.0 • Coupled with pyruvate kinase and lactate dehydrogenase (NADH → NAD+) • Absorbance at 340 nm is determined

CK: Forward Reaction (Tanzer-Gilvarg)

• More commonly performed method in the laboratory • Reverse reaction is two to six times faster and has less interferences • Optimum pH: 6.8

CK: Reverse Reaction (Oliver-Rosalki)

Sources of Error Anticoagulants inhibit enzyme activity Avoid hemolysis Serum should be stored in the dark Muscular activity and muscle mass have effects on levels

CREATINE KINASE

Widely distributed in the tissues of the body

LACTATE DEHYDROGENASE

High concentrations in the heart and liver

LACTATE DEHYDROGENASE

Five major isoenzyme fractions ; Each isoenzyme is composed of four subunits. The subunits are of two different structures: (heart) and M (muscle) polypeptide

LACTATE DEHYDROGENASE

LACTATE DEHYDROGENASE Serum concentration: LD flipped pattern:

LD-2 > LD-1 > LD-3 > LD-4 > LD-5

Heart, RBCs

LD-1 LD-2

Myocardial infarction Hemolytic anemia

LD-1

Megaloblastic anemia Acute renal infarct Hemolyzed specimen

LD-2

Lung, lymphocytes, spleen and pancreas

LD-3

Pulmonary embolism Extensive pulmonary pneumonia Lymphocytosis Acute pancreatitis Carcinoma

LD-3

Liver Hepatic injury or inflammation

LD-4

Skeletal muscles Skeletal muscle injury

LD-5

Because of its widespread activity in numerous body tissue, [?] is elevated in a variety of disorders.

LDH

Highest levels are seen in pernicious anemia and hemolytic disorders

LDH

In AMI: rises 12-24 hours after onset, peaks 48-72 hours, remains elevated for 10 days An elevated total value is a nonspecific finding. Assays therefore assume more clinical significance when separated into isoenzyme fractions.

LDH

Optimum pH: 8.3 to 8.9

LDH: Forward reaction (Wacker et al)

Optimum pH: 7.1 to 7.4 Rate of reaction is three times faster → smaller sample volumes and shorter reaction times More susceptible to substrate exhaustion and loss of linearity

LDH: Reverse reaction (Wrobleuski & La Due)

Sources of Error Avoid hemolysis Do not freeze specimen Serum is the sample of choice since some anticoagulants (e.g. oxalate) inhibit the enzyme

LDH

increases within 4-8 hours of MI, peaks at 12-24 hours, elevated 3-4 days increases within 12-24 hours, peaks at 72 hours, elevated 10 days

CK LD

[?] are heat stable; [?] labile [?]↑with liver and skeletal muscle disease. [?] with hemolysis

LD1 and LD2; LD5 LD4 and LD5 LD1 > LD2

Widely distributed in human tissue

Aspartate aminotransferase (SGOT)

Highest concentration: cardiac tissue, liver & skeletal muscle

Aspartate aminotransferase (SGOT)

Smaller amounts: kidney, pancreas & RBCs

Aspartate aminotransferase (SGOT)

Bilocular enzyme: cytoplasmic & mitochondrial isoenzyme

Aspartate aminotransferase (SGOT)

Distributed in many tissues

Alanine aminotransferase (SGPT)

Comparatively high concentrations in the liver (more liver-specific enzyme of the aminotransferases)

Alanine aminotransferase (SGPT)

catalyze the interconversion of amino acids & alpha-ketoacids by transfer of amino groups

AMINOTRANSFERASES

acts as the coenzyme for both AST and ALT

Pyridoxal phosphate

AMI: rises within 6-8 hours, peaks 24 hours, returns to normal in 5 days

AST

measurements are mainly confined to evaluation of hepatic disorders

• ALT

• With most forms of acute hepatocellular injury (e.g. hepatitis) [?] will be higher than ALT initially, because of the higher activity of [?] in hepatocytes. Within 24 - 48 hours, particularly if ongoing damage occurs, ALT will become higher than AST, based on its longer half-life

AST AST

In alcoholic hepatitis, the elevations in ALT are comparatively lower than AST, resulting in an AST/ALT ratio (De Ritis ratio) [?]. (Bishop)

greater than 2

Oxaloacetate from AST activity reacts with Malate dehydrogenase as the indicator enzyme Monitors the change in absorbance at 340 nm continuously as NADH is oxidized to NAD Optimal pH: 7.3 - 7.8

Karmen Method (for AST)

Pyruvate from ALT activity reacts with lactate dehydrogenase as the indicator enzyme

Walker et al Method (for ALT)

Sources of Error [?] can be dramatically increased in hemolyzed specimens while [?] is relatively unaffected.

AST ALT

Involved in the cleavage of phosphate-containing compounds in alkaline pH

ALKALINE PHOSPHATASE

facilitates the transfer of metabolites across cell membranes associated with lipid transport and the calcification process in osseous tissues

ALKALINE PHOSPHATASE

Located in a wide variety of tissues None in erythrocytes

ALKALINE PHOSPHATASE

ALKALINE PHOSPHATASE: Separation is done through the following methods [?]- reduces activity of intestinal and placental isoenzymes [?]- Reduces activity of bone and liver enzyme Heat fractionation Electrophoretic fractionation

Inhibition with phenylalanine Inhibition with levamisole

Often used in evaluation of hepatobiliary (obstructive conditions) and bone disorders (osteoblast involvement)

ALKALINE PHOSPHATASE

Highest elevation of ALP is seen in [?] Physiologic elevation of ALP can be seen in [?]

Sources of Error Use serum or heparinized plasma only Increased activity in specimens left standing on the clot for a long time due to a gradual development of a more basic pH in the system as CO2 is lost

ALKALINE PHOSPHATASE

ACID PHOSPHATASE: Tissue distribution and isoenzymes - band 1; inhibited by tartrate - bands 2 & 4 - band 3; major form in plasma - band 5; osteoclasts

Prostatic ACP Granulocytic ACP Platelets, RBs & Monocytes Bone isoenzyme

Separation is done through the following methods Electrophoretic separation Chemical inhibition - pACP inhibited by L-tartrate Immunoassays

ACID PHOSPHATASE

Prostatic cancer: ACP is inferior to PSA

↑ Prostatic ACP

Prostatic hyperplasia and prostatic infarction

↑ Prostatic ACP

Urinary tract obstruction, carcinoid tumors of rectum and prostatic massage

↑ Prostatic ACP

assays have proven useful in forensic clinical chemistry, particularly in the investigation of rape. Vaginal washings are examined for seminal fluid (presumptive evidence of rape in such cases).

ACID PHOSPHATASE

Sources of Error Store at appropriate pH and temperature Sample of choice: plasma (to minimize contamination by platelets) Preferred anticoagulant citrate buffered to a pH of 6.2 to 6.6 Avoid hemolysis

ACID PHOSPHATASE

Catalyze the hydrolytic cleavage of peptides to form amino acids or smaller peptides

GAMMA-GLUTAMYLTRANSFERASE

Plasma membrane-bound on cells

GAMMA-GLUTAMYLTRANSFERASE

Widely distributed in a number of tissues

GAMMA-GLUTAMYLTRANSFERASE

Highest amounts of GGT in the kidneys / Significant amounts in pancreas and liver

GAMMA-GLUTAMYLTRANSFERASE

Useful marker for liver damage

GAMMA-GLUTAMYLTRANSFERASE

Increased in obstructive liver disease, inflammation of the liver, obstruction of the biliary tract

GAMMA-GLUTAMYLTRANSFERASE

Increase over time in patients on long-term medications

GAMMA-GLUTAMYLTRANSFERASE

Most useful application for GGT measurements:

GAMMA-GLUTAMYLTRANSFERASE Reference method: Product of the reaction: 5-amino-2-nitrobenzoate (410 nm)

Szasz Assay

Sources of Error Preferred specimen: serum Some anticoagulants inhibit activity

GAMMA-GLUTAMYLTRANSFERASE

Primarily responsible for starch digestion Splits complex CHO made up of a-D glucose units

AMYLASE

Two major sources:

AMYLASE

Major clinical reason for increased serum amylase:

cancers kidney impairment (e.g. renal failure) conditions affecting salivary glands

↑ AMYLASE

: number of milligrams of glucose released in 30 minutes at 37 deg C under specific assay conditions

Somogyi unit

: inhibits salivary amylase

Wheat germ lectin

Electrophoresis: salivary amylase is more

anodal

Measures the rate of disappearance of the starch substrate

Amyloclastic (lodometric)

Measures the amount of reducing sugars produced by the hydrolysis of starch

Saccharogenic (Nelson Somogyi mod. By Henry & Chiamon)

Measures the increasing color from production of product coupled with a chromogenic dye

Chromogenic (Klein, Foreman, Searcy)

This measures the change in turbidity of starch solution over a short reaction period. This is used for stat analysis to rule out acute upper abdominal pain

Turbidimetry and Nephelometry (Peralta & Reinhart)

AMYLASE Classic reference method:

Enzyme that hydrolyzes glycerol esters of long chain fatty acids (e.g. triglycerides)

LIPASE

Full activity in the presence of bile salts and colipase

LIPASE

Highest concentrations found in pancreatic tissue and secretions Assessment of acute pancreatitis

LIPASE

• Rise: 2 - 12 hours • Peak value within 48 - 72 hours • Remains elevated for 10 - 14 days

LIPASE

LPS: Cherry-Crandall method Substrate:

Liberated fatty acids were measured after a 24-h incubation

LPS: Cherry-Crandall method

Modifications - used triolein as substrate

LPS: Cherry-Crandall method

Simpler and more rapid

LPS: Turbidimetric method

Based on coupled reactions with enzymes such as peroxidase or glycerol kinase

LPS: Colorimetric method

Sources of Error Bacterial contamination - false elevation Avoid hemolysis - hemoglobin inhibits lipase thus causing falsely lower values

LPS

Sources include the adrenal cortex, spleen, thymus, lymph nodes, lactating mammary gland and RBCs

GLUCOSE-6-PHOSPHATE DEHYDROGENASE

Little activity found in normal serum

GLUCOSE-6-PHOSPHATE DEHYDROGENASE

Specimen RBC hemolysate: to detect enzyme deficiencies Serum: to detect enzyme evaluations

GLUCOSE-6-PHOSPHATE DEHYDROGENASE

Inherited sex-linked trait Drug-induced hemolytic anemia Increased levels in MI and megaloblastic anemias (but not routinely tested in these conditions)

G6PD deficiency

Reaction catalyzed: hydrolysis of the neurotransmitter acetylcholine into choline and acetic acid, a reaction necessary to allow a cholinergic neuron to return to its resting state after activation

CHOLINESTERASE

Uses only acetylcholine as a substrate (high substrate specificity)

"TRUE" CHOLINESTERASE

Hydrolyzes a variety of choline esters

"PSEUDO" CHOLINESTERASE

Also known as acetylcholinesterase

"TRUE" CHOLINESTERASE

Referred to as "cholinesterase"

"PSEUDO" CHOLINESTERASE

Primary location are synapses of nerve cells. Also seen in RBCs, lung, brain, spleen

"TRUE" CHOLINESTERASE

Found in serum and in the white matter of the central and peripheral nervous system Also in the heart, liver and pancrease

"PSEUDO" CHOLINESTERASE

Important part of the process for the transmission of nerve impulses

"TRUE" CHOLINESTERASE

Protective function in the body; hydrolyzes choline esters (other than acetylcholine) which can inhibit acetylcholinesterase

"PSEUDO" CHOLINESTERASE

CHOLINESTERASE Pathological values are Important roles in the diagnosis and management of

Richest source in prostate. Also in bone, liver, spleen, kidneys, RBCs and platelets.

Acid phosphatase (ACP)

↑in prostatic carcinoma, bone disease

Acid phosphatase (ACP)

Catalyze hydrolysis of phosphomonoesters = alcohol + phosphate ion

Acid phosphatase (ACP) Alkaline phosphatase (ALP)

Reacts optimally at pH 5.0

Acid phosphatase (ACP)

Differentiate prostatic portion using tartrate as inhibitor (inhibited by tartrate).

Acid phosphatase (ACP)

Also used in rape investigation.

Acid phosphatase (ACP)

Activity associated with osteoclasts.

Acid phosphatase (ACP)

Separate serum from RBCs ASAP

Acid phosphatase (ACP)

Freeze or acidify to

Acid phosphatase (ACP)

Avoid hemolysis.

Acid phosphatase (ACP) Alkaline phosphatase (ALP) AST

Most human tissues. Highest concentrations in intestines, liver, bone, spleen, placenta and kidney.

Alkaline phosphatase (ALP)

↑in hepatobiliary disease and bone disorders; highest ↑with Paget's disease

Alkaline phosphatase (ALP)

Reacts optimally at pH 9.0 to 10.0

Alkaline phosphatase (ALP)

Requires Mg2+

Alkaline phosphatase (ALP)

Associated with osteoblast activity

Alkaline phosphatase (ALP)

Diet may induce elevations in ALP activity of blood group B and O individuals who are secretors.

Alkaline phosphatase (ALP)

Primarily in kidney, brain, prostate, pancreas and liver.

Gamma-glutamyltransferase (GGT)

↑ with biliary tract obstruction, chronic alcoholism

Gamma-glutamyltransferase (GGT)

Glutathione serves as gamma-glutamyl donor in most biologic systems.

Gamma-glutamyltransferase (GGT)

Lower values in females.

Gamma-glutamyltransferase (GGT)

Hemolysis does not interefere.

Gamma-glutamyltransferase (GGT)

Many tissues; Considered the liver-specific enzyme of the transferases

Alanine aminotransferase (ALT)

↑ with liver disease

Alanine aminotransferase (ALT)

Formerly SGPT. Pyridoxal phosphate as coenzyme.

Alanine aminotransferase (ALT)

Relatively unaffected by hemolysis.

Alanine aminotransferase (ALT)

Many. Highest in liver, heart, skeletal muscle.

Alanine aminotransferase (ALT)

↑ with liver disease, myocardial infarction (MI, muscular dystrophy

Alanine aminotransferase (ALT)

Formerly SGOT. Pyridoxal phosphate as coenzyme.

Alanine aminotransferase (ALT)

Marked elevation with viral hepatitis.

Alanine aminotransferase (ALT)

Don't use ammonium heparin

Alanine aminotransferase (ALT)

All. Highest in liver, heart, skeletal muscle, RBCs.

Lactate dehydrogenase (LD)

↑ with MI, liver disease, pernicious anemia

Lactate dehydrogenase (LD)

Catalyzes lactic acid › pyruvic acid

Lactate dehydrogenase (LD)

Serum should be separated in clot immediately to prevent ↑in LD1 and LD2.

Lactate dehydrogenase (LD)

Unstable; Store at 25 °C

Lactate dehydrogenase (LD)

Highest levels with pernicious anemia.

Lactate dehydrogenase (LD)

Enzyme that stays elevated longest with MI.

Lactate dehydrogenase (LD)

Some anticoagulants interfere.

Lactate dehydrogenase (LD)

Cardiac muscle, skeletal muscle, brain.

Creatine kinase (CK)

↑ with MI and muscular dystrophy

Creatine kinase (CK)

Catalyzes phosphocreatinine + ADP → creatine +ATP.

Creatine kinase (CK)

Most sensitive enzyme for skeletal muscle disease.

Creatine kinase (CK)

Highest levels with muscular dystrophy.

Creatine kinase (CK)

First enzyme to increase with MI.

Creatine kinase (CK)

Inhibited by all anticoagulants except heparin.

Creatine kinase (CK)

Salivary glands, pancreas

Amylase (AMS)

↑ with acute pancreatitis

Amylase (AMS)

Breaks down starch to simple sugars.

Amylase (AMS)

Saccharogenic method measures sugar produced. lodometric or amyloclastic measures starch remaining. Chromogenic method measures dye released from the breakdown of polysaccharide. Kinetic method measures change of NAD to NADH at 340 nm.

Amylase (AMS)

Ca2+ and Cl required.

Amylase (AMS)

Don't use EDTA, citrate or oxalate.

Amylase (AMS)

Urine levels stay elevated longer than serum.

Amylase (AMS)

Pancreas;↑ with acute pancreatitis

Lipase (LPS)

Breaks down triglycerides into fatty acids and glycerol.

Lipase (LPS)

Olive oil substrate.

Lipase (LPS)

Levels usually parallel amylase, but may peak a little later and stay elevated longer.

Lipase (LPS)

RBCs;↓ with hereditary predisposition to hemolytic crises after ingestion of oxidant drugs such as primaquine, infection or diabetic ketoacidosis

Glucose-6-phosphate dehydrogenase (G6PD)

Involved in first step of glucose metabolism.

Glucose-6-phosphate dehydrogenase (G6PD)

Measure in hemolysate of whole blood.

Glucose-6-phosphate dehydrogenase (G6PD)

Liver, brain

Pseudocholinesterase (PChE)

↑ after exposure to organophosphorus compounds found in insecticides and nerve gases, with hypersensitivity to succinylcholine and liver disease

Pseudocholinesterase (PChE)

1.2 to 1.5 kg Extremely vascular - blood supply comes from the hepatic artery and portal vein 1500 mL blood/min Functional unit: LOBULE

LIVER

-Six-sided with one portal triad (comprised of a hepatic artery, a portal vein and a bile duct) on each side -Two major cell types: • Kupffer cells • Hepatocytes (80%)

LOBULE

major waste product of heme catabolism Approximately 200-300 mg produced per day

BILIRUBIN

Synthesis: Proteins - albumin, cholinesterase, coagulation proteins, cholesterol, bile salts, glycogen Cholesterol Bile salts Glycogen

BILIRUBIN

Metabolism: Glucose to acetyl-CoA, gluconeogenesis, amino acid conversions, fatty acids

BILIRUBIN

Detoxification: Bilirubin, drugs, ammonia

BILIRUBIN

Excretion: Bile acids

BILIRUBIN

Jaundice becomes noticeable to the naked eye once bilirubin levels reach

>3.0 mg/dL.

: increased amounts of bilirubin are being presented to the liver

a. Prehepatic Jaundice

: primary problem is within the liver

b. Hepatic Jaundice

: lower production of UDPGT due to a genetic lesion and overall lower enzymatic activity; an additional defect related to a transport deficit in the sinusoidal membrane of the hepatocyte may be present

Gilbert Disease

: more serious disorder; multiple mutations in the gene coding for UDPGT results in the production of mildly dysfunctional to completely nonfunctional UDPGT.

Crigler- Najjar Syndrome

: more serious form; homozygously nonfunctioning proteins

• Crigler- Najjar Syndrome Type I

: severe deficiency

• Crigler- Najjar Syndrome Туре Il

Infants who are affected with Crigler-Najjar syndrome, especially the more severe form, develop severe unconjugated hyperbilirubinemia, which typically leads to kernicterus, the deposition of bilirubin in the brain. Motor dysfunction and retardation can result. The danger of kernicterus is a certainty at levels exceeding

20 mg/dL.

: removal of conjugated bilirubin from the liver cell & the excretion into the bile are defective

Dubin-Johnson

• Caused by deficiency of the canalicular transporter protein (MDR2/cMOAT)

Dubin-Johnson

• Appearance of dark-stained granules on a liver biopsy

Dubin-Johnson

: hypothesized to be due to a reduction in the concentration or activity of intracellular binding proteins; Liver biopsy does NOT show dark pigmented granules

iv. Rotor syndrome

v. Physiologic jaundice:

: biliary obstructive disease; stool loses color (clay-colored)

c. Posthepatic jaundice

: biliary obstructive disease; stool loses color (clay-colored)

c. Posthepatic jaundice

• Clinical condition in which tissue scar replaces normal, healthy liver tissue

Cirrhosis

• Most common causes include chronic alcoholism and chronic hepatitis C infection

Cirrhosis

are more common than primary liver cancers

a. Metastatic liver cancers

is the most common malignant tumor of the liver

b. Hepatocellular carcinoma

Reye Syndrome • Almost exclusively seen in • Often preceded by a • Strong association with intake of

• Acute illness characterized by non-inflammatory encephalopathy and fatty degeneration of the liver

Reye Syndrome

• Mild hyperbilirubinemia; threefold changes in ammonia and aminotransferases (AST and ALT)

Reye Syndrome

Drug and Alcohol-related Disorders Drugs cause injury most commonly via immune-mediated injury to the [?] [?]: most important drug which can cause liver damage; can lead to alcoholic cirrhosis [?] can cause fatal hepatic necrosis

hepatocytes Acetaminophen

• Bilirubin + diazotized sulfanilic acid → azobilirubin (pink-purple)

Classic Diazo Reaction

Reaction Initially done before on urine samples

Classic Diazo

• Can be done on serum samples, but only in the presence of accelerators (solubilizer)

Classic Diazo Reaction

Classic Diazo Reaction • Evelyn-Malloy reaction: • Jendrassik-Grof :

Total Bilirubin =

unconjugated bilirubin + conjugated bilirubin + delta bilirubin

Attached to one or two glucuronic acid molecules

CONJUGATED BILIRUBIN

Reacts DIRECTLY with the color reagent

CONJUGATED BILIRUBIN

Noncovalently attached to albumin

UNCONJUGATED BILIRUBIN

Does not react with the color reagent until the bilirubin is first dissociated from albumin using an accelerator (INDIRECT)

UNCONJUGATED BILIRUBIN

Non-polar; water insoluble

UNCONJUGATED BILIRUBIN

AKA hemobilirubin / slow-reacting bilirubin

UNCONJUGATED BILIRUBIN

Polar; water soluble

CONJUGATED BILIRUBIN

AKA cholebilirubin / one-minute / prompt bilirubin

CONJUGATED BILIRUBIN

• Third fraction of bilirubin

Delta bilirubin

• Conjugated bilirubin that is covalently bound to albumin.

Delta bilirubin

Seen only when there is significant hepatic obstruction.

Delta bilirubin

• When present, will react in most laboratory methods as conjugated bilirubin.

Delta bilirubin

Bilirubin is very sensitive to and is destroyed by light; therefore, specimens should be protected from light. If left unprotected from light, bilirubin values may reduce by

30% to 50% per hour.

WATER BALANCE • Average water content of the human body ranges from [?] • Located within the [?] compartments. • Concentrations of ions within cells are maintained both by [?]

40% to 75% of total body weight. intracellular (2/3) and extracellular (1/3) active and passive transport.

Physical property based on the concentration of solutes (colligative property)

OSMOLALITY

The sensation of thirst and arginine vasopressin hormone (AVP formerly ADH; posterior pituitary gland) secretion are stimulated by the hypothalamus in response to an increased osmolality of blood

OSMOLALITY

Normal plasma osmolality =

275 - 295 mOsm/kg of plasma H20

Indirectly indicates the presence of osmotically active substances other than Na+, urea or glucose, such as: Ethanol Lactate Methanol Betanydroxybutyrate Ethylene glycol

OSMOLAL GAP

Volume and osmotic regulation

Sodium, potassium, chloride

Myocardial rhythm and contractility

Potassium, magnesium, calcium

Cofactors in enzyme activation

Magnesium, calcium, zinc

Regulation of ATPase ion pumps

Magnesium

Acid-base balance

Bicarbonate, potassium, chloride

Blood coagulation

Calcium, magnesium

Neuromuscular excitability

Potassium, calcium, magnesium

Production and use of ATP from glucose

Magnesium, phosphate

Greatly depends on intake and excretion of water and on renal regulation of Na*; AVP, aldosterone, angiotensin Il and ANP

Sodium

Kidneys are important in regulation; aldosterone stimulates secretion into urine

Potassium

Aldosterone secretion conserves Cl; changes usually parallel Na*

Chloride

Most reabsorbed in kidneys as CO2

Bicarbonate

Regulation controlled largely by kidneys; regulation can be related to Ca?+ and Na*; PTH increases renal reabsorption and intestinal absorption; aldosterone and thyroxine increase renal excretion

Magnesium

↑: PTH, vitamin D ↓: calcitonin

Calcium

Kidneys play major role in regulation ↓: PTH ↑: Vitamin D and Growth hormone

Phosphate

Not specifically regulated; liver is the major organ for removing lactate

Lactate

1. Difference between unmeasured anions and unmeasured cations

ANION GAP

2. Useful in indicating an increase in one or more of the unmeasured anions in the serum and also as a form of QC for the analyzer used to measure these electrolytes

ANION GAP

Hypoalbuminemia

LOW ANION GAP

Severe hypercalcemia

LOW ANION GAP

uremia/ renal failure ketoacidosis methanol, ethanol, ethylene glycol, salicylate poisoning lactic acidosis hypernatremia instrument error

ELEVATED ANION GAP

is amperometric (Clarke electrode).

• p02 measurement

are potentiometric, using the Severinghaus and glass membrane electrodes respectively.

pCO2 and pH measurements

• Several acid-base parameters can be calculated from measured pH and pCO2 values: 1. calculation is based on the Henderson Hasselbach equation. 2. can be calculated using the solubility coefficient of CO2 in plasma at 37°C 3. is the bicarbonate plus the dissolved CO2 (carbonic acid) plus the associated CO2 with proteins (carbamates).

HCO3 Carbonic acid concentration Total CO2 content

Mixture of a weak acid and its salt with the capability of combining with protons and releasing protons in response to external shifts in pH

BUFFERS

Purpose: maintain a defined pH

BUFFERS

BUFFER SYSTEMS IN THE BODY

A. Plasma bicarbonate buffer B. RBC hemoglobin/oxyhemoglobin buffer C. Organic and inorganic phosphate buffer D. Plasma proteins Carbon dioxide = driving force in the bicarbonate carbonic acid system

TRH, CRF, GnRH, others

Hypothalamus

TSH, ACTH, FSH, LH, prolactin, GH

Anterior Pituitary

Vasopressin, oxytocin

Posterior Pituitary

Epinephrine, norepinephrine

Adrenal Medulla

Cortisol, 11-deoxycortisol, aldosterone

Adrenal Cortex

T3, T4, calcitonin

Thyroid

Parathyroid hormone (PTH)

Parathyroid

Insulin, glucagon

Pancreas

Estrogens

Ovaries

Testosterone, other androgens

Testes

Released directly from the tissue into the bloodstream and carried to the specific site of action.

HORMONES

Acts at a specific site or sites (target cells) to induce certain characteristic biochemical changes.

HORMONES

Adrenal glands, gonads, and placenta

STEROID HORMONES

Anterior pituitary, placenta, and parathyroid glands

PROTEIN HORMONES

Thyroid and adrenal glands

AMINE HORMONES

Cholesterol

STEROID HORMONES

Protein

PROTEIN HORMONES

Amino acids

AMINE HORMONES

Synthesized as needed, not stored

STEROID HORMONES

Synthesized, then stored in cell as secretory granules until needed

PROTEIN HORMONES

Synthesized, then stored in the cell as secretory granules until needed

AMINE HORMONES

Lipid Water

STEROID HORMONES PROTEIN HORMONES ; AMINE HORMONES

Lipid Water

STEROID HORMONES PROTEIN HORMONES ; AMINE HORMONES

Protein Do not need protein Require a carrier protein and others do not

STEROID HORMONES PROTEIN HORMONES AMINE HORMONES

Cortisol Aldosterone Testosterone Estrogen, and Progesterone

STEROID HORMONES

FSH* LH* TSH* hCG* Glucagon Parathyroid hormone Growth hormone Prolactin

PROTEIN HORMONES

Epinephrine Norepinephrine Thyroxine Triiodothyronine

AMINE HORMONES

- Composed of two polypeptide chains containing carbohydrate. Alpha chains are the same in all. Beta chains determine specificity

*Glycoproteins

Gonad (tropic)

Luteinizing Hormone (LH) Follicle-stimulating hormone (FSH)

Maturation of follicles, ovulation, production of estrogen progesterone, testosterone

Luteinizing Hormone (LH)

• Regulated by GnRH from hypothalamus.

Luteinizing Hormone (LH)

• Sharp ↑ just before ovulation.

Luteinizing Hormone (LH)

↑ FSH & LH in

ovarian failure, menopause

Sperm and egg production

Follicle-stimulating hormone (FSH)

• Regulated by gonadotropin-releasing hormone (GnRH) from hypothalamus

Follicle-stimulating hormone (FSH)

Thyroid (tropic)

Thyroid-stimulating hormone (TSH)

Production of T3 and T4 by thyroid

Thyroid-stimulating hormone (TSH)

• Thyrotropin-releasing hormone (TRH) from hypothalamus.

Thyroid-stimulating hormone (TSH)

Adrenal (tropic)

Adrenocorticotropic hormone (ACTH)

Production of adrenocortical hormones by adrenal cortex

Adrenocorticotropic hormone (ACTH)

• Regulated by corticotropin-releasing hormone (CRH) from hypothalamus.

Adrenocorticotropic hormone (ACTH)

• Diurnal variation: highest levels in early am, lowest in late afternoon.

Adrenocorticotropic hormone (ACTH)

↑ ACTH in

Cushing's disease.

(GH; aka somatotropin)

Growth hormone

Multiple (direct effector)

Growth hormone

Allows an individual to transition from a fed state to a fasting state

Growth hormone

↑ protein synthesis in skeletal muscle and other tissues

Growth hormone

Antagonizes the effects of insulin

Growth hormone

• Regulated by growth-hormone releasing hormone (GHRH) & somatostatin from hypothalamus.

Growth hormone

↑ GH in ↓ GH in

Gigantism & Acromegaly Dwarfism.

Breasts (direct effector)

Prolactin

Lactation

Prolactin

Regulated by prolactin-releasing factor (PRF) & prolactin inhibiting factor (PIF) from hypothalamus.

Prolactin

Breasts and uterus (direct effector)

Oxytocin

Critical role in lactation

Oxytocin

Major role in labor and parturition

Oxytocin

Produced in hypothalamus. Stored in posterior pituitary

Oxytocin

Kidneys (direct effector)

Vasopressin (Antidiuretic hormone)

Regulation of renal free water excretion

Vasopressin (Antidiuretic hormone)

Produced in hypothalamus. Stored in posterior pituitary.

Vasopressin (Antidiuretic hormone)

Release stimulated by ↑ osmolality, ↓ blood volume or blood pressure.

Vasopressin (Antidiuretic hormone)

↓ in diabetes insipidus.

Vasopressin (Antidiuretic hormone)

• Located near larynx; two lobes connected by a thin piece of tissue

THYROID GLAND

Exerts significant control over the rate of metabolism in humans

THYROID GLAND

THYROID GLAND Hormones produced:

T3, T4, calcitonin

Stimulated by TSH (from the anterior pituitary to produce and secrete T3, T4

THYROID GLAND

THYROID GLAND Functional unit:

thyroid follicle

Comprised of follicular cells surrounding a central colloid

thyroid follicle

Colloid contains thyroglobulin, which is rich in tyrosine (the amino acid that forms the backbone for the thyroid hormone molecules)

thyroid follicle

- contains tyrosine which forms the backbone for the thyroid hormone molecules

•Thyroglobulin

- contains tyrosine which forms the backbone for the thyroid hormone molecules

•Thyroglobulin

= T3 (3, 5, 3'-triiodothyronine)

• Monoiodotyrosine + dodotyrosine

= T4 (3, 5, 3', 5'-tetraiodothyronine)

• Diiodotyrosine + diodotyrosine

is more potent (T4 is converted to T3 suggesting that T3 is more important)

• T3

Major transport protein for T3 and T4

Thyroxine-binding globulin (TBG)

: 0.03-0.05% is unbound (almost completely bound to proteins) : 0.5% free (weaker attachment to proteins)

T4 T3

RIA, fluorometric enzyme immunoassay, fluorescence polarization immunoassay (FPIA)

Thyroxine

RIA, microparticle enzyme immunoassay, fluorometric enzyme immunoassay

Triiodothyronine

Resin uptake, FPIA

Thyroid hormone binding ratio, T3 uptake, T uptake

Equilibrium dialysis, immunometric assay (chemiluminescence)

Free T4

RIA

Free T3

Calculation from T4 and THBR

Free T4 index, Free thyroxine index (FTI), T7

Calculation from T3 and THBR

Free T3 index

RIA, Immunometric assay (IMA)

Thyroid-stimulating hormone

Cretinism Hashimoto's thyroiditis

Hypothyroidism

• Hypothyroidism in neonates and infants

Cretinism

• Characterized by mental retardation, short stature with skeletal abnormalities, coarse facial features, enlarged tongue and umbilical hernia

Cretinism

• Causes: Maternal hypothyroidism during early pregnancy Thyroid agenesis lodine deficiency Dyshormonogenetic goiter (congenital defect in thyroid hormone production; most commonly involves thyroid peroxidase)

Cretinism

• Most common cause of hypothyroidism in regions where iodine levels are adequate

Hashimoto's thyroiditis

• Autoimmune destruction of thyroid gland

Hashimoto's thyroiditis

• Anti-thyroglobulin and anti-thyroid peroxidase antibodies are often present

Hashimoto's thyroiditis

• Associated with HLA-DR5

Hashimoto's thyroiditis

Subclinical hypothyroidism = Subclinical hyperthyroidism =

↑ TSH, normal FT4 ↓ TSH, normal FT4

Grave's disease

Hyperthyroidism

• Most common cause of hyperthyroidism

Grave's disease

• Autoantibody (IgG) stimulates TSH receptor → increased synthesis and release of thyroid hormones

Grave's disease

• Clinical features: - Hyperthyroidism - Diffuse goiter - Exophthalmos - Pretibial myxedema

Grave's disease

• Composed of three layers; each secretes predominantly one class of hormones.

ADRENAL CORTEX

produces mineralocorticoids (e.g. aldosterone)

Zona glomerulosa

produces glucocorticoids (e.g. cortisol)

Zona fasciculata

produces androgens (e.g. DHEA, dehydroepiandrosterone)

Zona reticularis

are derived from cholesterol.

Cortical hormones

= Fluid and electrolyte balance

a. Mineralocorticoids

= Fluid and electrolyte balance

a. Mineralocorticoids

= Glucose production and protein metabolism

b. Glucocorticoids

= Regulate sexual development and control many aspects of pregnancy

c. Sex steroids

All adrenal steroids are derived by sequential enzymatic conversion of a common substrate, cholesterol.

STEROIDOGENESIS

Non-specific; carries many steroids

Albumin

Cortisol and derivatives; progesterone

Cortisol-binding globulin

Testosterone and estradiol

Sex hormone- binding globulin

• critical for sodium retention, (volume), potassium, and acid-base homeostasis

MINERALOCORTICOIDS (ALDOSTERONE)

Most commonly due to bilateral adrenal hyperplasia (60%) or adrenal adenoma (40%, Conn syndrome)

Primary Hyperaldosteronism

Arises with activation of renin-angiotensin system (e.g. renovascular hypertension)

Secondary Hyperaldosteronism

Muscle weakness with thin extremities - cortisol increases muscle break down to produce amino acids for gluconeogenesis