Mechanisms & Genetics of Diabetes - Scott

Question 1

What are the 3 pathways that function in normal cells to maintain glucose homeostasis?

Answer 1

Glucose stimulates release of insulin from pancreatic β cells.

Insulin stimulates uptake of glucose by skeletal muscle and adipose tissue
via Glut4.

Insulin stimulates glycogen synthesis and conversion of fatty acids to
triglycerides in liver, inhibits gluconeogenesis

Question 2

What are the various categories (types) of diabetes?

Answer 2

DM Type 1

DM Type 2

Gestational Diabetes

Maturity Onset Diabetes of the Young (MODY)

Question 3

What causes Maturity Onset Diabetes of the Young (MODY)?

Answer 3

Rare, autosomal dominant mutations in single genes that disrupt pancreatic β cell function.

Give clues to important functions in β cells.
MODY1 hepatocyte nuclear transcription factor (HNF)-4α
2 glucokinase
3 HNF-1α
4 PDX1 (Insulin Promoter Factor-1
5 HNF-1β
6 NeuroD1

***All are transcription factors that effect pancreatic beta cells.

Question 4

How is insulin secretion regulated?

Answer 4

Regulated mainly by glucose:

Glucose taken up by GLUT1 or GLUT2

phosphorylated by glucokinase

Glucose-6-phosphate&raquo_space;> ATP

ATP inhibits K+ channel > depolarization

Depolarization > opening of voltage

dependent Ca2+ channels

Influx of Ca2+ > secretion of insulin

Question 5

MODY 1

Answer 5

HNF-4α- is a β cell transcription factor. Targets include GLUT2 and pyruvate kinase.

Question 6

MODY 2

Answer 6

Glucokinase catalyzes rate limiting step in regulation of insulin secretion

Question 7

MODY 4

Answer 7

PDX1 is a β cell transcription factor necessary for early development, and for transcription of insulin, glucokinase, GLUT2.

Question 8

What is the etiology of Type 1 Diabetes?

Answer 8

Insulin is not made because pancreatic beta cells are
destroyed by an autoimmune process.

Genetics-
Identical twin concordance 40-60%.
50% of genetic risk comes from polymorphisms in MHCII alleles.

Immunological characteristics-
	Autoantibodies to islet proteins 	
	Insulitis- infiltration of T cells. 
	T cells recognize and proliferate in 
	response to β cell antigens.
	Autoantibodies before development of T1D.

Question 9

What are the major genetic variants associated with Type 1 Diabetes?

Answer 9

polymorphisms in MHCII genes => alter peptide binding sequences

Specific MHCII alleles are strongly associated with T1D:
DQA10301
DQB10302
DQB1*0201

Question 10

Why/how are MHC II proteins important in immune system function?

Answer 10

MHCII proteins and the immune response:

MHCII present antigen peptides to T cell with cognate TCR.
TCR interacts with complex of particular MHCII + peptide activates T cell => Initiates adaptive immune response.

MHCII restriction:

Different MHCII alleles encode MHCII proteins with different peptide binding capabilities.
Repertoire of MHCII proteins determines which peptides will be presented to TCR.
Repertoire of MHCII proteins influences immune response and tolerance.

MHCII proteins and tolerance to “self” antigens:

During T cell development MHCII proteins present “self” peptides on cell surface.
T cells expressing TCR that strongly bind to specific MHCII + “self” peptide complex are targeted for apoptosis.
Tolerance requires that individual’s MHCII proteins be able to bind self antigens.

Question 11

What is the model of Autoimmune etiology of Type 1 Diabetes?

Answer 11

1. High risk variants of MHCII can not bind pancreatic beta cell “self” peptides strongly.
2. T cell receptors (TCR) recognizing self antigens of pancreatic beta cells are not weeded out during development
3. These T cells have the potential to mount an autoimmune attack when the right environmental trigger occurs.
4. Specific environmental triggers lead to inflammatory response (possibly specific viral infections or microbiome)
5. Autoimmune response:
   T cells expressing TCR which recognize beta cell peptides are clonally expanded => mount immune response against beta cells.
6. Individuals become symptomatic when ~80% of beta cells are destroyed. Only recourse is to supply artificial insulin.

Question 12

What are four of the immunomodulatory therapies being tested for Type 1 Diabetes treatment?

Answer 12

Teplizumab = Anti-CD3- alters T cell immune response at several levels

Oral insulin as a decoy for anti-insulin antibodies

CTLA4-Ig (Abatacept) for Prevention of Abnormal Glucose Tolerance = Activates CTLA4 leading to T cell inhibition

T1DM Immunotherapy Using CD4+CD127lo/-CD25+ Polyclonal Tregs (Treg) = adoptive transfer of autologous transfer of Tregs to treat recently diagnosed T1D

Question 13

What is the etiology begin Type II Diabetes?

Answer 13

Excess nutrients at cellular level => Hyperglycemia, circulating free fatty acids =>

Inflammation, dysregulated lipid metabolism =>

Insulin resistance =>

Pancreatic Compensation (no DM) or

Relative insulin insufficiency =>

Beta cell destruction, absolute insufficiency => TYPE II DM

***Each step amplifies the previous step!

Question 14

What is the genetic risk in Type II Diabetes?

Answer 14

Specific genes not known:
Many genes associated with beta cell function with polymorphisms that confer a small risk have been identified mainly through genome wide association studies (GWAS) = 
TF7L2
Ppar gamma
K+channel
zinc transporter
IRS
calpain 10

Question 15

What are the processes that contribute to insulin resistance? Involved tissues?

Answer 15

Processes: inflammation, dysregulation of lipid metabolism

Tissues: adipose, skeletal muscle and liver

Question 16

How does adipose tissue become the site of initiation for inflammation in Type II Diabetes?

Answer 16

Chronic inflammation of adipose tissue develops over several stages:

1. Non-pathogenic- initial increase in size due to increased uptake of glucose, storage of triglycerides.
2. Pro-inflammatory- Secretion of Monocyte Chemotractant Protein 1 (MCP-1) by adipocytes
3. Inflammatory- recruitment of macrophages which secrete TNFα

Question 17

How does lipid dysregulation begin in Adipose Tissue?

Answer 17

Excess nutrition and inflammation: Favors free fatty acid release not TG formation.

TNFα inhibits TG synthesis and promotes release of FFA by inhibiting PPAR-gamma

Insulin resistance inhibits TG formation

Decreased glucose uptake + Activation HSL

Question 18

How does local insulin resistance develop in Adipose Tissue?

Answer 18

FA’s bind to TLR2 or TLR4, initiate signaling through inflammatory NFKB pathway (JNK)

TNF alpha activates NFKB and JNK pathways.

JNK = Promotes nuclear translocation of AP-1 pro-inflammatory transcription factor. Phosphorylates and inactivates IRS = > inhibits insulin receptor signaling pathway.

1. Translocation of the insulin dependent glucose transporter GLUT4 to the cell surface is inhibited.
2. HSL is activated so that TG formation is inhibited.

Question 19

How do Adipose tissues promote systemic insulin resistance?

Answer 19

Adipocytes attract macrophages

Macrophages secrete TNF-alpha

Increased free fatty acids in circulation => activates more TLRs on skeletal muscle

Question 20

How does insulin resistance develop in skeletal muscles?

Answer 20

1. Toll-like receptor (TLR) pathway:
   Saturated fatty acids bind TLR => Activate Jun kinase (JNK) => JNK inhibits IRS adapters via serine phosphorylation => Inhibition of downstream signaling results in insulin resistance.
2. Diacylglycerol pathway:
   Increased fatty acid flux > increased DAG => DAG activates protein kinase C (PKC) => PKC inhibits IRS adapters via serine phosphorylation => Inhibition of downstream signaling results in insulin resistance

Question 21

What does insulin resistance mean in adipose/skeletal tissues in the development of Type II Diabetes?

Answer 21

Insulin binding to insulin receptor is intact but downstream signaling is impaired.

Insulin is ineffective; tissue is “resistant” to the effects of insulin

Question 22

How does insulin resistance develop in hepatocytes?

Answer 22

Hepatocytes take up glucose via Glut2 which is not regulated by insulin signaling.

1.Inhibition of glycogen synthesis:
Decreased phosphorylation of glycogen synthase 3 (GSK3) => Decreased activity of glycogen synthase (GS)

2. Increased gluconeogenesis:
   Translocation of FOXO1 to nucleus => Transcription of gluconeogenesis enzymes

Insulin resistance > decreased storage and greatly increased production of glucose

Question 23

What are the interactions of insulin resistance between tissues in Type II Diabetes?

Answer 23

Adipose tissue- Increased glucose > chronic inflammation > release of free fatty acids (FFA)

Skeletal muscle- insulin resistance > inability to take up glucose > further increase in blood glucose

Hepatocyte- insulin resistance > increased
gluconeogenesis > further increase in blood glucose

Pancreatic beta cell response to initial hyperglycemia
and amplified insulin resistance :
1. Increase in insulin output.
2. Relative insufficiency due to insulin resistance.
3. Destruction of beta cells due to inflammatory process.

Question 24

How does pancreatic beta cell dysfunction occur via oxidative stress?

Answer 24

Excess nutrients cause overload of electron transport chain in mitochondria, excess proton gradient

Transfer of electrons to Complex III is inhibited => Electrons are transferred to O2 instead, thus generating reactive oxygen species (ROS) =>

Increased levels of ROS cause oxidative stress =>

Pancreatic β cells have low levels of anti-oxidant enzymes
and so are highly susceptible to damage by ROS

Question 25

How does pancreatic beta cell dysfunction occur via Endoplasmic reticulum (ER) stress?

Answer 25

Overload of unfolded proteins creates ER stress. ER stress activates a set of protective signaling pathways called the unfolded protein response (UPR) Under extreme conditions the unfolded protein response initiates apoptosis Normal pancreatic beta cells operate on the edge of ER stress. Insulin resistance increases the need for insulin and can push the UPR to apoptosis

Question 26

How does pancreatic beta cell dysfunction occur via the Stress Apoptosis Pathway?

Answer 26

Oxidative stress (ROS) and ER stress (unfolded protein response) can activate an apoptotic pathway. Activation of caspase-1 in the inflammasome. Caspase-1 cleaves and activates the cytokine IL1β. Secreted IL1β binds to pancreatic β cell receptors => APOPTOSIS! ***Excess nutrients + insulin resistance lead to cellular stresses that activate IL-1β mediated apoptosis of pancreatic β cells.