And Flashcards
Which gene in males inhibits Wnt4 expression in the undifferentiated gonad and promotes male sexual development?
Sry
When during gestation do cells migrate from the yolk sac to the gonadal ridge?
4-6 weeks gestation
In the female, which which duct regresses to allow for differentiation into the genital tract ( oviduct, uterus, cervix and upper one third of the vagina)?
Wolffian duct regresses ( mullerian duct remains and differentiates)
Organs in the female genital tract are composed of which 3 layers?
A protective outer layer, a middle wall of smooth muscle, and an inner mucosal layer
The pH of the cervix is higher, lower or same as the vagina?
higher (pH 7-8)
The cervix secretes buffers that..?
Neutralize the acidic environment of the vagina, increasing the pH from 4-5 to 6-7 at the opening of the cervix
MII oocytes have elevated levels of what substance that regulates the reduction of disulfide bonds between sperm chromatin-associated protamines allowing the sperm nucleus to decondense to support normal processing of the male pronucleus?
Glutathione
Plot used to see performance over time
Levey-Jennings Plot
expected-observed=difference
Differenced is summed together over tiem and should remain constant
Cusum plot (cumulative sum plot)
acceptable range for Sperm morph QC (levey Jennings)
+/-1 or 2 SD
CV%
SD/mean observed observations (usually 10-15% range)
PSA
Prostate-specific antigen
(enzyme secreted by the prostate gland to degrade structural proteins semenogelin I and II (secreted by seminal vesicles) during liquefaction 30-60 min post ejaculation
Which hormone aids in mucus production in the cervix to help sperm transport?
Estrogen
Uterine transport of sperm
uterine myometrium contractions
Sperm entry into the isthmus of the oviduct via the utero-tubal junction requires?
Correct sperm morphology, motility, and surface proteins (oviduct is free of immunological substrates and can maintain sperm viability for hrs or days)
Increased paternal age
reduced sperm motility and volume, increased DNA damage (DNA fragmentation); no observed reduction in concentration. Increased mutational load but no observed increase in aneuploidy
functional maturation involving process that does not alter the cell structure but changes its potential for fertilization
Capacitation. fert competence gained within the female reproductive tract
During capacitation, which changes in the plasma membrane prepare the sperm to undergo the acrosome reaction?
Shedding of proteins and cholesterol, post-translational and structural modification of proteins, change in tail movement (increase in the bend amplitude but a decrease in progressive movement)
Capacitation at the cellular level requires
the removal of inhibitory factors from sperm + alterations in membrane proteins and lipids
Capacitation at the molecular level
changes in intracellular pH, [Ca2+], and cAMP-dependent tyrosine phosphorylation/ dephosphorylation
sperm hyperactivated motility, ability to bind to the zone pellucida, and undergo the acrosome reaction are characteristics of what?
Capacitation.
Hyperactivation of sperm
occurs in the female tract and allows the sperm to move to the oocyte and penetrate viscous substances like mucus in the oviduct and through cumulous cells.
Capacitation in vitro
removal of sperm from seminal fluid for 2-4 hrs held in buffered media
long range guidance mechanism for sperm to find the oocyte
Thermotaxis- temperature gradient (lower in the isthmus and higher in the ampulla- gets warmer as closer to ovary and egg) guide capacitated sperm to fert site
short range guidance mechanism
Chemotactic method- may happen once sperm are closer to the egg (along with oviductal contractions) using chemoreceptors on the spermincluding olfactory receptors (chemoattractants may be secreted by follicle or mature egg)
Acrosome reaction is exocytotic meaning
things are released from sperm to the environment
induced following sperm binding to the ZP
acrosome reaction (must happen prior to sperm fusing with the oocyte (oolema essentially) happens at the ZP
Acrosomal contents are released when
the outer acrosomal membrane fuses with the the overlying sperm plasma membrane
Membrane bound organelle on sperm that originates from the golgi apparatus
acrosome
Hyaluranidase is present
sperm head- either plasma membrane or released from acrosome
What could zona binding tell you about the sperm or oocyte?
could tell you if the zona of the ova were abnormal or sperm that failed to bing were abnormal
guanine nucleotide-binding regulatory protein + PLCbeta1 and tyrosine kinase (TK) receptor coupled to PLCgamma
receptor-mediated signaling pathways in sperm plasma membrane involved in acrosome reaction caused by fusion with ZP
acrosome reaction by ZP caused by
progesterone and ZP proteins. Both lead to an increase in intra cellular Ca2+ within the sperm
What event occurs to induce depolarization of sperm membrane potential, increased intracellularpH, increased intracellular Ca2+ in the capacitated sperm?
ZP-sperm binding
ZP-sperm binding»_space;fusion of sperm plasma membrane with outer acrosomal membrane»
release of acrosomal contents
ZP1, ZP2, ZP3, ZP4 can work as a complex
all 4 ZP proteins play a role in sperm egg binding and more than one ZP proteins induces the acrosome reaction
ZP1, 3, 4 bind to which part of the sperm?
capacitated head (pre-acrosome reaction)…that comes next
ZP2 binds to which part of the sperm?
the acrosome-reacted sperm
sperm penetrate the ZP through two mechanisms
- mechanical forces (motility)
- acrosomal proteases that hydrolyze the ZP glycoproteins
Role of ZP x4
spermbinding
induction of Acrosome rxn
acrosomal exocytosis
block polyspermy
2 blocks to polyspermy
- fast block: oocyte-membrane block»_space;depolarization of the oocyte membrane following binding of first sperm
- Slow block: zona rxn/cortical rxn»_space; hardening of ZP proteins by enzymes (hydrolyses, proteinases, peroxides) released in cortical granules after sperm-oocyte binding
can a sperm that has already started the AR still bind with the zona p?
yes- sperm binding is a redundant process-many ways for it to happen in case something goes wrong. other sperm surface proteins that can interact with the ZP
ROS can impact capacitation, hyper activation and AR induction? T/F?
true
Sperm in the PV space fuse where with the oolema?
fusion occurs btwn the post-acrosomal region of the sperm plasma membrane and the oocytes microvillous surfaces- except can’t bind where 2nd metephase spindle and 1st polar body are located
2 plasma membrane proteins essential for sperm-oolema binding
- Izumo-1(sperm) - must move from outer acrosomal membrane to mid plasma membrane region during AR (AR incomplete sperm cannot bind oocyte)
- Juno (oocyte)- shed from the oolema rapidly following fert and may be another mechanisms for polyspermy block
SPA
sperm penetration assay (zona-free hamster eggs can be fertilized by sperm from many mammalian species ) test for functionality of human sperm
Oocyte activation mechanisms induced by sperm
Sperm introduces a soluble cytosolic factor PLC(zeta), into oocyte cytoplasm acting as molecular trigger of activation causing cytoplasmic calcium oscillations
The cortical reaction
Receptor modification and Zona hardening and loss of sperm binding capacity
Protease Ovastacin and fetuin-B
cortical granule protease that hardens zona; inhibitor of ovastacin to sustain sperm binding
In spermiogenesis, somatic (non-gametic) histones in DNA are replaced with
Protamines, for chromatin condensation
what type of bond hold compacted DNA in place in elongated spermatids?
Disulfide bonds (formed by oxidation of the sulphhydral groups)
Syngamy
two adjacent pronuclear formation
First step in sperm nucleus decondensation?
what happens after sperm-oocyte fusion and demembrenated sperm are exposed to oocyte factors (in nuclear and cytoplasmicalaly mature oocytes)?
reduction of disulfide bonds btwn protamines by oocyte-derived disulfide bond reducer glutathione
Removed sperm protamines are replaced by what after sperm nucleus decondensation?
replaced by oocyte histones, organizing them into nucleosomes
reduction of sperm protamines
glutathione and heparin sulfate (protein acceptor)
sperm decondensation is dependent on which environmental factors?
time, pH, temperature
Sperm centriole becomes zygotic centrosome, needed for syngamy
SRY and SOX9 actions are required for testis determination
SRY on Y chromosome up regulates SOX9 (master testis determining factor)
Fetal leydig cells at 8 weeks (promoted by hCG) produce
testosterone, needed to for maintenance of wolffian ducts and masculinization of genitalia
Germ cells arrive at gonadal ridge at
5 weeks gestation
Exocrine function of testis
production of sprm, involving sertoli cells, serve a nurse cells and maintain the blood-testis barrier
endocrine function of testis
production and secretion of androgens from the Leydig cells (exocrine function is dependent on this endocrine function)
Paracrine function of testis
local controls of spermatogenesis through paracrine factors like cytokines, growth factors.
Leydig cells at 8 weeks gestation produce which hormone, neede for maintenance of wolffian ducts?
testosterone
From which embryonic tissue layer are wolffian ducts derived from?
Intermediate mesoderm - give rise to kidney and male genital tract (in the female, wolffian ducts regress at 2 months- same time as male leydig cells start producing T)
From which embryonic tissue layer are mullarian ducts derived from?
Coelomic epithelium - give rise to falopian tubes, ducts in pelvic region fure to form the uterovaginal canal
blood-testis barrier
Maintained by sertoli cells. tight junctions in sertoli cells serve to prevent proteins and antibodies from reaching spermatogenic cells and eliciting an immune response
FSH fxn in sertoli cell
Pituitary FSH stimulates receptors in sertoli cells to produce sertoli-derived inhibin that makes negative feedback loop to inhibit GnRH>FSH and activin release
Testosterone fxn in sertoli cell
androgens are ABSOLUTELY required for spermatogenesis. androgen receptors on sertoli cells facilitate spermatogenesis (no androgen receptors on germ cells- all actions of anderogens are on the sertoli cells for sperm production)
Spermiation
release of mature sperm into the lumen of the seminiferous tubule
T/F - # of sertoli cells dictates the spermatogenic potential of a man?
True. > sertoli cells = > # of sperm that can be produced
Blood-testis barrier
Tight junctions (not gap junctions) between neighboring sertoli cells at the basal laminae of the lumen.
Basal compartment of seminiferous tubule
between sertoli junctions and basal laminae containing the early spermatogonia
adluminal compartment of seminiferous tubule
above the sertoli cell tight junctions containing more advanced germ cells
Sertoli cells produce which form of inhibin, to provide negative feedback for FSH production?
Inhibin B
(if damage to germinal epithelium (sertoli cells), results in increased FSH seretion.
Sertoli cells produce which hormone during fetal development to regress the mullerian ducts?
AMH
Leydig cells primary function
steroid hormone makers- synthesize steroid hormones from cholestrol
Leydig cell androgen production is stimulated by
pituitary LH
rate limiting step of steroidogenesis in leydig cells
mobilization of cholesterol from intracellular stores to leydig mitochondrial membrance where it is converted into pregnenolone
Which protein is responsible for transporting cholesterol into mitochondria?
acute steroidogenic regulatory protein (StAR)
testosterone is reduced in peripheral tissue to dihydrotestosterone (DHT) by which enzyme?
5alpha reductase
testis also produce estrodiol from conversion of testosterone to estrogen by which hormone
aromatase (CYP19)
(estrogen receptor found in leydig cells-estrogens may regulate testosterone production)
Where are Leydig cells located?
between seminiferous tubules inthe interstitial space close to blood vessels and lymphatics
When are Leydig steroidogenically cells active?
8-18 weeks fetal develpoment and puberty onward
Leydig cell function is regulated by which two hormones
LH: stimulates T biosynthesis
Prolactin: induces expression of the LH receptor
Which germ cell population are undergoing meiosis?
spermatocytes
spermatogonia are
slowly dividing stem cell population
which germ cell population are already haploid and undergoing morphological changes?
spermatids
order of spermato -
goina»cytes»tids
Which cells in the body have the highest rate of mitosis?
Spermatogonia in the testes
spermatogonial stem cells
in basal compartment of blood-testes barrier next to sertoli cells and can can regenerate post toxic insult
Type A spermatogonia
least mature sperm cells located directly along basal lamina. require retinoic acid to convert type A to type B. Type A spermatogonia are connected my small cytoplasmic bridges
2 types of Type A spermatogonia
A pale: spermatogonia
A dark: considered to be stem cells
Type B spermatogonia are more differentiated than Type A?
True
What is needed to begin meiosis in sperm?
retinoic acid acts directly on spermatogonia to stimulate entry into meiosis which creates Type B from Type A
What is the ploidy of Spermatogonia
Diploid. Located along basal lamina of seminiferous epithelium. (outside of blood testis barrier- ie wont cause issue because still diploid cells, on the blood side of the tight junctions between sertoli cells)
Meiosis in sperm cells:
Type B spermatogonia> enter meiotic pathway
DNA replication> 4N (tetraploid gamete)
introduction of genetic variation of gametes from recombination, double-stranded breaks and crossing over, DNA repair
> 2 successive reductive devisions» haploid spermatids
4N/ 2N/ 2= 1N (haploid -23 chromosome spermatids)
Primary spermatocytes (4C) characterized by
large, 4C chromosome value, now have moved into the adluminal compartment inside the blood-testis barrier as non-diploid cells. Will then divide into 2 secondary spermatocytes (2C DNA content)
Secondary spermatocytes(2N)
2C; formed by completion of first meiotic division, will undergo one more division to form 2 sister spermatids with haploid (1N- 23 chromosomes) chromosome content
Which stage do germs cells end in at the completion of spermatogenesis (genetic component- end of meiotic divisions)
Spermatids
Spermiogensis is the process of
maturing of sperm cells
spermatids» mature spermatozoon
3 phases of spermiogenesis:
- Development of the acrosome: development of acrosomal cap where storage of hydrolytic enzymes (golgi stage, cap phase, acrosomal phase, maturation phase)
- Maturation phase: nucleus elongates, cytoplasm is phagocytosed by sertoli cell, spermatozia released into lumen
- Condensation of the nucleus: somatic chromatin histones are replaced by highly basic protamines, chromatin fibers condense and compact nulcear material into a tightly packed structure, transcriptional activity stops (packed up into the suitcase until get to final destination)
What part of the sperm contains hydrolytic enzymes?
The acrosome- the acrosome contains hyaluronidase, neuraminidase, acid phosphatase, and trypsin like protease needed by the mature sperm to transverse the cumulus complex and zona pellucida
sperm flagellum is formed by what
?
one of the sperm centrioles differentiates to form flagellum (mid piece of the tail is thickened region of flagellum where movement is generated. mitochondria migrate along developing axoneme)
What is the flagellum made of?
axoneme (9+2 microtubule doublets in a concentric arrangement) surrounded by fibrous sheath and keratin containing dense outer fibers. Mitochondria in mid-piece provide energy
Sperm Type A can be pale (Ap) or dark (Ad). Which type goes on tp make Type B?
Pale. Type Ap go on to make 2 Type B through mitotic division, which will then progress to enter meiosis.
Type Ad, dark, can either produce type Ap(pale) or more Type Ad (dark-stem cell)
Spermiogenesis
differentiation of the round spermatid to undergo morphological changes to develop an acrosome and flagella and undergo nuclear condensation and extrusion of cytoplasm
tetratozoospermia
low % normal morphology
Example:
round headed sperm/ globospermia due to the absence of the acrosome or acrosome atrophy
Length of the cycle of spermatogenesis (spermatogonia to spermatozoa)?
64 days
on diagram be able to define
radial spoke, central sheath, dynein arms, subfiber A and B
Factors that can decrease motility
tobacco, sexual lubricants, saliva, alcohol and drugs
What powers the dynein arms in sperm axoneme to move
ATP hydrolysis
Which stages of spermatogenesis are testosterone dependent?
Transition from primordial germ cell to type A spermatogonia during fetal development (meiosis), reductive devision of primary to secondary spermatocytes (4N»2N»1N) (meiosis), spermiation (sperm maturation)
(non-mitotic division)
What is the role of Thyroxine in fetal gonad development?
hormone/factor that regulates Sertoli cell numbers during development
Abnormalities in the pampiniform plexus, network of interconnected veins to cool arterial blood entering the testis, such as varicocele, causes what?
adverse effects on spermatogenesis by interfering with cooling mechanisms of the plexus
what is a varicocele
vericose veins in the spermatic cord of the scrotum
Where in the male tract do sperm acquire motility?
Epididymis (head-caput, body-corpus, cauda-tail).
How long does it take for sperm epidydimal maturation(to the head of the epididymis)?
10 days
How long until sperm reach the end of the epididymis and are ready for ejaculation?
16 days (so 90 days from start of spermatogenesis until ready for ejaculation) so give 3 months from insult to see if sperm quality reappears)
sperm go from
seminiferous tubule»
lumen»
caput (head) epididymis»
corpus(body) epididymis»
cauda(tail) epididymis»
vas deferens»urethra
While maturing in the epididymis sperm change by
gain motility, acquire additional surface glycoproteins, change net surface charge, increased capacity for glycolysis, increased ability to adhere to ZP, changes in immunoreactivity) these changes are regulated by androgens
release of sperm from cauda epididymis by
short adrenergic contractions by cauda epi. and vas def during ejactulation
emission vs ejaculation
The ejaculation process consists of two phases: emission (when semen is created by fluids from the prostate, seminal vesicles, and vas deferens and released into the penis) and expulsion (which follows the emission phase as the ejaculation climaxes).
Summary of epidydimal fxn
- sperm maturation
- stabilization of condensed chromatin
- changes in surface charge of the plasma membrane
- new sperm surface proteins
- sperm storage
- sperm transport by peristalsis
spermatic cords contains
vas deferens, cremaster muscle, spermatic artery and veins (pampiniform plexus)
Prostate
largest accessory gland (contributes 15-30% or 0.5mL to semen)clear, slightly acidic fluid (high citric acid content to maintain osmotic equilibrium in semen). Prostatic fluid contains enzymes required for liquefaction of the ejaculate coagulum.
Seminal vesicles
Fluid from seminal vesicles contributes 45-80% (2-2.5mL) of the volume of the semen. Rich in fructose (major source of glycolytic energy for the sperm) and prostaglandins. contain enzymes required for formation of coagulum, androgen dependent organ.
What can elevated or absent levels of fructose in semen mean?
Function of seminal vesicles or ejaculatory duct disfunction/obstruction in azoospermic men. no fructose may mean congenital bilateral absence of vas deferens and seminal vesicles
T/F vas deferens and seminal vesicles are derived from sam embryonic origin
true. if vas deferens is congenitally absent, seminal vesicles and fructose in semen will also be absent
seminal prostaglandins do what?
urethral muscle contractions during ejaculation and may aid sperm transport through female repro tract.
Cowper (bulbourethral) gland and Littre glands secrete?
first fluid portion of ejaculate. 0.1-0.2mL, low volume to coat/lubricate urethra, clear fluid rich in mucoproteins
Seminal plasma
combination of fluids from all accesory sex glands