Fertilization and Embryo Development 

The stage of the embryo that is finally eligible for implantation on the uterine surface 

THE  OOCYTE AT OVULATION 

ovulation in the human embryo when the oocyte has reached the meta phase of the second meiotic division. it carries a haploid set of 23 chromosome which lies in the previtline space.the human oocyte is approximately 100 microns in diameter and surrounded by two layers of distinctly recognize layer of cells. the immidiate surrounded layer is the corona radiates . which has cell process interdigitating with the micro-villi of oocyte,transfering nutrients, and proteins.

THE SPERMATOZOON AFTER SPERMATOGENESIS 

The human spermatozoon once relesed in the ejaculate consist of a head, mid piece and a tail. The entire spermatozoon like other cell is enclosed within a plasma membrane. The head has a flattened pyriform shape, measuring 4-5 micrometer in length and 2.5 - 3.5 micrometer width. The head has a large nucleus made up 23 compacted chromosome and an apical ridge 

CHANGES IN GAMETES  PRIOR TO FERTILIZATION 

The oocyte unlike the sperm undergoes less changes prior to fertilization while getting in the ampula of the fallopian tube. perheps the main changes undergone by the oocyte are maturation   accomplish by expansion and loosening of the cumulus  and corona cell and migration of the cortical granules before fertilization. since oocyes are aspirated at various stage of maturation in the in vitro fertilizzation procedure, a brief incubation 4-6 hours at 37`c, in a 6% CO2 incubator is necessary to complete maturation in many oocyte. The sperm before it can engage itself in fertilization undergoes change as follows 

capacitation 

Acrosome rea ion 

Hypermotility 

FERTILIZATION 

Fertilization involves the union of two germ cells, sperm and oocyte and this occurs in the ampula  of the Fallopian tube. fertilization sperm usually binds to the surface of the zona pellucida before penetrating it. There is a strong correlation between the degree of zonal binding and fertilization rates. Zonal receptor and sperm receptor have been recently shown to exist and these complement each other before fertilization once the spern enters the previtline space, the inner membrene of the sperm fuse with the oolemma of the oocyte. The plasma membrane above the equatorial segment fuses first with the oolemma. upon fusion of the two membrane the oocyte undergo the activation phenomenon. This involves the following steps 

Resszumption of meiosis 

Exocitosis of cortical granules 

EARLY EMBRYONIC DEVELOPMENT 

After formation of the zygote, the early human preimplantation embnryo undergoes clevage through simple mitotic division while within the ampulaa of the isthmus of the fallopian tube up to the late morulla or early cavitation blastocyst stage. Thereafter it descends to the uterus is still surrounded by the zona poellucida. clevage take place reletively slowly in the human with the first division 

ANALYSIS OF EMBRYO MORPHOLOGY 

The pronuclear Embryo 

the early clevage 

the clevage stage embryo 

THE BLASTOCYSTS 

s

Oogenesis

                                                               INTRODUCTION  

Oogenisis is creation of an ovum.It is the female process of gametogenesis. The mitotic phase of human germ cells in females terminates at birth and by 5th month of fetal life. All oogenia have entered their first mitotic division to become primery oocyte. At puberty about 200,000 germ cells are available for  reproductive life 

Ooocyte deliver half of chromosomal complement to an embryo. However maternal and paternal genomic contribution is not the same. Ooocyte transmit not only genomic maternal but also mitochondrial DNA to the embryo which is majorly affected in aged oocytes 

Female Reproduction

                                                                INTRODUCTION  

Female Reproduction is a complex phenomenon involving the ovary tubes and uterus. The ovary is different in the 7th week and starts forming primordial germ cells which in sdult life from oocytes. After minarche the development of the follicle containing the oocyte mature under the control of hormonecente in th released from pituitary gland hypothalamus. Hence reproduction is control from higher  centers in the brain. The ovulation occurs with release of oocyte which passes through the fallopian tube and is fertilized there.The subsequent embryo implants in the uterus by day 6. The overy release hormone to support pregnancy. 

THE FETAL OVARY

The Beginning of Ovarian Development 

Sexual differentiation of the undifferentiated starts at around 7 th week of intrauterine life. The migration of the germ cells into the ovary formation of the follicles and the distinction into cortex and medulla are the key events in formation of the fetal ovary. Formation of the ovary first starts appearing as a thickening of the coelomic epithelium like cells ( GRELs) cells extendas deeply into the mesoderm thus forming distinct projection known as gonadal ridge.

Migration of the primordial Germ cells in the ovary 

The primordial germ cells are endodermal  in origin. They arise from the yolk sac and migrate from yolk sac to the genital ridge along the hindgut. Kit ligand and its receptor kit play a critical role in the survival of migrating germ cells. On reaching the gonadal ridge these stem cells which are diploid are termed as oogenia. oogenia along with stromal cells and vasculature enter into the devloping ovary by break down of the basal lamina underlying the mesonephric surface epithelium 

;b b naDevelopment of the Fetal Ovary a

The GREL cells enclose the oogenia and form the ovigerous cords. The ovigerious cords lie at the periphery of the developing ovarian cortex. The medulla contain the stroma with the capillaries and remnants of mesonpherious known as rate ovarii. The ovigerous cords break up into smaller cords by basal lamina forms the primordial follicle. aaThe first primordial cells lie at the corticomedullary junction of the ovary. The ovary is first identified histologically by 10 th week of gestation 

The Timeline of Human Ovarian Development 

In 28 th days mesonpherious develops. Medial of the mesonepheric duct gonadal ridge appears in 35 days. The undifferentiated gonads are visible by 42 days and further development of the ovary and differentiation into cortex and medulla is completed by 10 th week of gestation. Between 10 and 12 weeks  the diploid primordial germ cells cease mitosis and enter stage 2 meiosis 

Factors Affecting the ovarian Development in the Fetus

Development of the fetal ovary requires a complex interplay between genetic and epigenetic factors. It is no more a default process as was thought earlier. The initiation of ovarian development in xx karyotype requires activation of genes Wnt -4 and DAX-1

Development of the ovarian Follicles 

After the migration of the primordial germ cells (PGCs) to the ovary at 5 week of gestation, these cells undergo mitosis and form germ cells cysts in the fatal ovary. At 10-12 week of gestation thse seeds breakdown and form a primordial follicle pool. Each ovarian follicle then undergoes development in three stage 

* Gonadotropin independent 

* Gonadotropin responsive 

* Gonadotropin dependent 

Stages of folliculogenesis 

primordial follicles 

primary follicles 

secondary  follicles 

Tertiary Follicle or the Preantral Follicle (400 micro meter) 

This stage of the follicle is characterized by the appearance of cavity due to the accumulation of fluid in between the granulosa cells.The antral fluid is formed from transudation of plasma from thecal cappilaries and is rich in proteins,steroids and anticoagulants, This process is controlled by autocrine and paracrine factors and is not dependent on gonadotropins.

Grafian follicles or the Antral follicle 

The largest and most mature of the tertiary follicles are called Graffian follicle. These will be selected for maturation each menstrual cycle. characteristic structural unit of graffian follicle is an antrum containing fluid called follicular fluid .

ANATOMY AND PHYSIOLOGY OF FEMALE REPRODUCTIVE SYSTEM 

The hypothalamus, anterior pituitary gland and ovaries are responsible for regulating the female reproduction. The hypothalamus secretes a gonadotropin releasing hormone (GnRH)

To understand the function of the female reproductive system it can be divided into three compartments 

* Hypothalamic pituitary level

* ovarian level 

* uterus, Fallopian tube cervix and vagina 

Spermatogenesis

                                        Spermatogenesis

The term spermatogenesis describe and include all the process involve in the production of gametes. Spermatogenesis is the process by which precursor germ cell turned spermatogonea undergo a complex series  of division to give rise to spermatozoa . Spermatogenesis take place in the tubular compartment, consisting of the seminiferious tubules. For quantitatively and qualitatively normal  production of sperms, the integrity of tubular compartments, is necessary. The function of testis and of its other compartments are governed by the by the hypothalamus and the pituitary gland. These endocrine effects are mediated and attuned at the testicular level by local control mechanism. The interstitial compartment between the seminiferious tubules plays and inevitable role in regulating the over all process of spermatogenesis till ejaculation.

The Hypothalamo-Pituitary System ; Functional organization 

The successful initiation of testicular function is dependent on the hypothalamic secretion of  GnRH   which in turn stimulates FSH  and LH to act on the testis from anterior pituitary. These action initiate spermatogenesis  and testosterone production. Due to pulsatile nature of GnRH secretion, the release of gonadotropins also occurs in discreet peaks ,more obvious in case of LH , because of its shorter half life (20 minutes) as compared to FSH in circulation . Where as the activation of GPR54 receptor,located on the surface of the GnRH neurons are responsible for the secration of GnRH. The function of pituitary gland is also under control of gonadal steroids and peptides eithe directly or via hypothalamus. 

LH and FSH   

LH targets the leydig cells to stimulate androgen biosynthesis, and the resulting androgen ( testosterone and its androgen metabolites )  act on receptor within the seminiferious epithelium to stimulate and  report spermatogenesis.FSH target receptor in the sertoli cells directly to support spermatogenesis . however the role of either endocrine factors, such as vitamin A and its metabolic retionoic acid,are emerging while both androgen and FSH are required for optimal spermatogenesis, spermatogenesis relies on the local production of growth factors, signaling molecules and other intransic mechanism.

  Teststerone 

 Leydig cells are the site of production of androgen in the Testis,The synthesis and secretion both are under control of pituitary LH.In the absence of androgen, spermatogenesis would not produce beyond meiosis. Testosterone is the most abundent androgen in the blood.In normal healthy man about 66-7 grams of testosterone is synthesized daily by leydig cells in the intestinal sapace of the testis. The transformation of testosterone into DHT take place in prostrate. A small amount of testosterone is store in the testis because very high level does not support spermatogenesis. Testosterone bind eith albumin nor sex harmone binding globulin for the transportation (SHBG) for the transportation in the plasma produced by hypatocytes

                                          Feedback Mechanism

At the onset of puberty, the hypothalamus cause the release of FSH and LH into the male system for the first time. FSH enters the testes and stimulate the sertoli cells to begin facilitating spermatogenesis using negative feedback, as illustrated. Testosterone the hormone responsible for the secondry sexual characteristic that devlop in the male during adolescence, stimulate spermatogenesis 

In response to FSH release, which stimulate sertoli cell sertoli cell produce two factor- one inhabition ( which regulate the production of FSH from pituatiary ) second androgen binding- globulin (ABG)

Leydig Cells 

Leydig cells are present in the interstitial compartment of testes. These cells are approx. A class of hormone called androgen and peptide hormones INSL-3 are secreted by leydig cell under the influence of Leueinizing hormones. A class of hormone called androgen (19 c steroid ) and peptide hormones INSL-3 are secreted by the Leydig cell under the influence of Leteinizing hormones (LH) stimulated by anterior pituitary. 

Sertoli Cells 

Sertoli Cells are found within the germinal epithelium of basal membrane. These cells represent approximately 35-40%of total volume of germinal epithelium. 

Stage Of Spermatogenesis 

There are following stage of spermatogenesis 

* Spermatogoniogenesis (mitosis)

* Spermatidogenesis (meiosis1 and meiosis 2)

* spermiogenesis 

* spermiation 

Mitotic Division 

Spermatocytogenesis term is refered to as the male gametocytogenesis and result in the formation of spermatocytes possessing half the normal complement of the genetic material . spermatocytogenesis starts with the mitotic division of diploid spermatogonium ( from the basal compartment of the seminiferious tubules ) and produces two primery spermatocytes ( diploid intermediate cells). 

Spermatidogenesis 

The formation of spermatids from secondary spermatocytes is called spermatidogenesis 

Spermiogenesis and Spermiation 

During spermiogenesis the spermatids begin to form a tail by growing microtubules on one of the centrioles , which turns into basal body. These microtubules form an axonema. Later the centriole is modified in the process of centrosome reduction 

Semen Analysis 

Semen primery atributes two parameters 

* The total number of spermatozoa present in the ejaculate,which reflects the proper functioning  of testes and post testicular duct system 

* The total fluid volume assures the secretory activity of various accessory gland 

The first cohert of semen ejaculate hjas prosthatic fraction and basically sperm rich. There is noticeable difference in the quality of sperm ejaculated in a laboratory setup 

Semen Analysis 

It can divided in two steps ;

* Macroscopic Analysis 

* Microscopic Analysis 

* Macroscopic Analysis 

Collected semensample should be kept at 370C for 5 minutes, volume, PH apperence pf the semen sample, liquification should be mesured between 30 to 60 minutes 

Volume To mesure the ideal volume of collectd semen saampe 

Liquefaction The time at which the heterogenous mass of fluid starts to liquify at room tempreture is known as liquification time 

PH  PH should me measured after 30 minutes and consensus value of 7.2 is the lower threshold 

Microscopic Analysis 

 Microscopic analysis can be done by making a wet preparation of sample under phase contrast microscope 

* Aggregation 

* Agglutination 

* Sperm mortility 

* Progressive mortality ( PR) 

* Non progressive mortality (NP)

* Immotility 

* Sperm concentration and number 

* Staining 

Male Reproduction

 Introduction

In embryonic stage determination of sex can be done at the time of fertilization although the gonads do not acquire differential characteristic until the 7th week of devlopment .Gonads appear initially as longitudinal ridges,also known as genital or gonadal ridges 

These ridges are result of proliferation of the epithelium and condensation of mesenchyme  lying deep of epithelium lining. Germ cell usually appear after 6th week of development in the genital ridges 

PRIMORDIAL GERM CELLS 

During development primordial gem cells appear in the wall of yolk sac close to the allantois and migrate along the dorsal mesentary of the hind gut by an ameboid movement . These germ cells reach the primitive gonads by 5th week and stars invading in genital ridge  by 6th week 

If they fail to reach the genital ridges,honads do not form so the primordial germ cells have an inductive influence on devlopment of the gonads.Primordial germ cells have an inductive influence on devlopment of the gonads 

At rhe stage gonads are known as the indifferent gonads  

DEVELOPMENT OF TESTIS 

In the male embryo, the primordial germ cells have XY chromosome.The y chromosome has SRY gene ,which encodes an important factor known as the testis - determining factor, this factor promotes the proliferation of primitive sex cords and help them to penetrate deep into the medulla to form the testis or medullary cords. At the hilium of the gland, these cords break up into a channel of cell strands, which forms the tubulus of the testis. During the development, a fibrus connective tissue layer, the tunica albuginea, forms between the medullary cords of testis and the surface epithelium. By the fourth mounth of these testis cords become u shaped, and their terminal ends are the connected with the rate testis 

The testicular cords near composed of 

* Migrated primitive germ cells 

* Sertoli cells derived from the surface epithelium 

INTERSTITIAL CELLS OF LEYDIG 

The mesenchyme in the gonadal ridge, which is present between the developing testis cords, forms the Leydig cells. They develop in the mesenchyme after the onset of proliferation and differentiation of testis cords. By the 8th week of development.  Leydig cells start secretion of testosterone. This testosterone influence the sexual differentiation of the genital ducts and external genitalia Testis cords remainsolid until puberty, when they acquire a lumen, they known as the seminiferous tubules.

Thus the structural components of Testis are derived from three source 

*intermediate mesoderm ( mesenchyme) that forms the urogenital rtidges on the pisterior abdominal waal, giving rise to Leydig cells ( interistial cells) and myoid cells (peritubular contractile cells)  

* Mesodermal epithelium (surface epithelium of coelom) that lines the urogenital ridges and gives rise to sertoli cells 

*Primordial germ cells that migrate from the yolk sac into developing gonads, where they divide and differentiate into spermatogonia 

* FACTORS REGULATING DEVELOPMENT 

 The  components of developing genital system are 

* Gonads and primative sex gland 

* Genital 

* External genitalia 

All these components are initially in an indifferent stage in which they may devlop inti either a male or female genital system. The SRY gene on the Y chromosome encodes the testis determining factor which regulate ,male sexual development. SRY gene also influence the genes encoding steroidogenesis factor (SFI)and SOX9 that stimulate differentiation of sertoli and Leydig cells in the testes 

The SRY gene induce the 

* Devlopment of the testis ( medullary) cord 

* Formation of the tunica albuginea 

The SRY gene inhabits  the 

* Decvloopment of cortical (ovarian) cords

if the expression of the SRY gene is absent ,another gene DAXI, down regulate the steroidogenesis factor (SFI) , and along with that the  expression of gene WNT4 in the gonadal ridge, induce the formationn of ovaries with formation of epithelial cord in cortical part of gonads, instead of medullary cord 

ANATOMY AND FUNCTION OF MALE REPRODUCTIVE SYSTEM 

( INCLUDING ACCESSORY ORGAN SYSTEM AND FUNCTION)

The male reproductive system consists of the 

* Spermatic cord 

* Scrotum

* Testes

* Genital-excurrent duct 

* Accessory sex glands and penis 

Spermatic Cord 

During development, the testis traverses and abdominal wall and reaches into the scrotum ; and carries its vessels, nerve and ductile part with it. All these form the spermatic cord, which suspend the testis in the scrotum. The spermatic cord has coverings.Which are form by the layers of the abdominal wall. The spermatic cord contains ;

* Vas deferens 

* Artery to the vas deferens

* Testicular artery and veins 

* Cremasteric artery 

* Cremasteric nerve and the sympathetic components of the testicular plexus 

* Genital branch of the genitofemoral nerve 

* Lymph vessels draining the testis 

All of these structures are conjoined by loose connective tissue.

Scrotum

The scrotum is a fibromuscular sac covered by skin, containing the testes and part of the spermatic cords. It consist of 

* Skin 

* Dartos muscle 

* External spermatic 

* Cremasteric and internal spermatic fasciae  

The scrotum is devided into right and left mhalves by a cutaneous raphe, which continues ventrally to the inferior penile surface and dorsally along the midline of the perineum to the anus.

Testis 

Testis is a male gonad and importent component of reproductive system. The adult testis are paired ovoid organ that lie within the scrotum. Testis are connected by the spermatic cord to the abdominal wall and tethered to the scrotam by scrotal ligaments, the remnants of the gubernaculum. The testis devlop on the posterior wall  of the abdomen and latter on they dexcend into the scrotum 

Structure of testes 

Each testis is covered by a      thick connective tissue capsule, the tunica albuginea. The inner part of this capsulwe is a loose connective tissue that  contains blood vessels and known as the tunica vasculosa. This tunica vasculosa form deeper separation in testis, which devide the each testis into approximately 250 lobules. Along the posterior surface of testis , the tunica albuginea thickens and projects inward as the mediastinum testis.Blood vessels, lymphatic vessels and the genital ex current duct pass through the mediastinum  

Epididymis 

It is the first part of genital excurrent duct system. The epididymis lays posterio-lateral to the testis, and the vas deferens lies along its medial side. It has an expended head known as globulus major, a body and a tail

Paradidymis 

The paradidymis is a smalln collection nof convoluted tubules. It is present anteriorly in the spermatic cord above the epididymal head.

Vas Deferens 

The vas deferens is the distal continuation of the epididymis, starting as a muscular tube at the epididymis, starting as a muscular tube at he epididymal tail. It is 45 cm long and convey sperm to the ejaculatory duct 

Ejaculatory Ducts 

The ejaculatory ducts are formed by the union of ducts of the seminal vessicle and the terminbal part of the vas deferens.

Seminal vessicles 

The two seminal vesicles are sacculated tube located between the bladder and rectum 

Bulbourethral Glands 

The two bulbouretharal glands are small rounded, somewhat lobulated mass of approximately 1 cm diameter. 

Penis 

The penis consist of 

Root 

Free and pendulous Body 

The Root of the penis 

The root of penis consist of erectile tissue in the urogenital triangle, namely the two crura and the bulb, which are firmly attached to the pubic arch and perineal membrane respectively.

The Main Body of the Penis 

Body of the penis consists of the three masses of erectile tissue, the right and left corpora cavernosa, and the median corpus spongiosum, which are continuations of the crura and the bulb of the penis of root respectively. They become engoredn with blood during penile erection.

  

Cell Biology

          

                                                      INTRODUCTION 

A cell is the basic unit of any form of life and the study of the cell and its function is known as cell biology.The discipline  is very old and goes back to the times When Anton van Leeuwenhok was first discovered cell under the microscope . There are some basic organelles common to all cell types  and then some vary unique ones as the cell specialized . Improvement in instrument ,especially in micros cope and staining techniques made it possible to relate cell structure and function from the simple magnifying glass to electron microscope  and imaging techniques. There are many specialized sub fields in cell biology which have become independent subject themselves some of them being cell genetics ,epigenetics , cell cycle and cell metabolism .

THERE  ARE FOLLOWING TERMINOLOGY USED IN CELL BIOLOGY 

Deoxyribonucleic ( DNA)

Ribonucleic Acid ( RNA)

Nucleotide 

Gene 

Exons and Introns 

Chromosome 

Chromatid 

Centromere 

Chromatin 

Genome 

Codon 

Replication 

Gene expression 

Translation 

Transcription 

Epigenetics 

Gene Regulation 

Nucleus 

Mitochondria 

Ribosomes 

Endoplasmic Reticulam 

Golgi Appartus 

Cytoskeletal Protein 

Spindle Apparatus

Kinetochore 

Cell Cycle 

G1 Phase 

S Phase 

G2 Phase 

Mitosis 

Karyokinesis 

Meta phase

Anaphase 

Telophase 

Cytokinesis 

Regulation of cell cycle 

Meiosis 

Meiosis 1

Spermatozoa 

Oocytes  

Epigenitics 

 

   

EMBRYOLOGIST

 What is the role of embryologist :-

An embryologist is the fertility spacelist that helps to create viable embryos to eighter be used in ivf or frozen for latter use. Embryologist are highly trained medical professional usually holding a master degree or PHD due to nature of work. They are responsible for the careful maintenance and management   of genetic material use in creating embryos the collected sperm and egg a well as maintaining these embryos they devlop