B2 Spermatogonia Research Articles

Clonal organization of proliferating spermatogonial stem cells in adult males of two species of non-human primates, Macaca mulatta and Callithrix jacchus.

The present study examines the existence of clonogenic patterns in the proliferation and differentiation of spermatogonial stem cells in two species of non-human primates, the marmoset and the rhesus monkey. We developed a novel approach to detect proliferating spermatogonial clones in whole mounts of seminiferous tubules. Dual fluorescence labeling of bromodeoxyuridine and acrosin in conjunction with confocal microscopy allows the description of the clonogenic and spatial arrangement of proliferating spermatogonia at specific stages of the seminiferous epithelial cycle. Cross-sections of paraffin-embedded tissue were labeled by the same approach. For both monkey species we demonstrate the presence of proliferating spermatogonial clones of variable size at specific stages of the cycle of the seminiferous epithelium. Detailed analysis of the rhesus monkey reveals proliferating Apale spermatogonia at stages VII and IX of the cycle of the seminiferous epithelium, and of proliferating B spermatogonia at stages II, IV, VI, and XII. Proliferating Apale spermatogonia at stages VII and IX of the cycle are organized in pairs or quadruplets. B1 spermatogonia appear as quadruplets or eight-cell clones, and B2 spermatogonia as 8- or 16-cell clones. We conclude that spermatogenesis in the rhesus monkey is initiated by two divisions of duplets or quadruplets of Apale spermatogonia: a first division at stage VII, after which the clones of Apale spermatogonia separate, and a second division at stage IX, which leads to clones of B1 spermatogonia as well as pairs and quadruplets of Apale spermatogonia replenishing the seminiferous epithelium to maintain the original size of the A spermatogonial population.

Germ cell apoptosis in the testes of normal stallions

Apoptosis in testicular germ cells has been demonstrated in many mammalian species. However, little is known about the stallion (Equus caballus) and rates of apoptosis during spermatogenesis. Morphological and biochemical features of apoptosis reported in other species were used to confirm that the TdT-mediated dUTP Nick end labeling (TUNEL) assay is an acceptable method for identification and quantification of apoptotic germ cells in histological tissue sections from stallion testis. Seminiferous tubules from eight stallions with normal testis size and semen quality were evaluated according to stage of seminiferous epithelium to determine the germ cell types and stages where apoptosis most commonly occurs. Spermatogonia and spermatocytes were the most common germ cell types labeled by the TUNEL assay. A low rate of round and elongated spermatids were labeled by the TUNEL assay. Mean numbers of TUNEL-positive germ cells per 100 Sertoli cell nuclei were highest in stages IV (15.5±1.0) and V (13.5±1.1) of the seminiferous epithelial cycle (P<0.001). An intermediate level of apoptosis was detected in stage VI (P<0.02). These stages (IV–VI) correspond to meiotic divisions of primary spermatocytes and mitotic proliferation of B1 and B2 spermatogonia. Establishing basal levels of germ cell apoptosis is a critical step towards understanding fertility and the role of apoptosis in regulating germ cell numbers during spermatogenesis.

Germ cell apoptosis in the testes of normal stallions

Apoptosis in testicular germ cells has been demonstrated in many mammalian species. However, little is known about the stallion ( Equus caballus) and rates of apoptosis during spermatogenesis. Morphological and biochemical features of apoptosis reported in other species were used to confirm that the TdT-mediated dUTP Nick end labeling (TUNEL) assay is an acceptable method for identification and quantification of apoptotic germ cells in histological tissue sections from stallion testis. Seminiferous tubules from eight stallions with normal testis size and semen quality were evaluated according to stage of seminiferous epithelium to determine the germ cell types and stages where apoptosis most commonly occurs. Spermatogonia and spermatocytes were the most common germ cell types labeled by the TUNEL assay. A low rate of round and elongated spermatids were labeled by the TUNEL assay. Mean numbers of TUNEL-positive germ cells per 100 Sertoli cell nuclei were highest in stages IV (15.5±1.0) and V (13.5±1.1) of the seminiferous epithelial cycle ( P<0.001). An intermediate level of apoptosis was detected in stage VI ( P<0.02). These stages (IV–VI) correspond to meiotic divisions of primary spermatocytes and mitotic proliferation of B1 and B2 spermatogonia. Establishing basal levels of germ cell apoptosis is a critical step towards understanding fertility and the role of apoptosis in regulating germ cell numbers during spermatogenesis.

Seasonal Differences in Equine Spermatocytogenesis1

Spermatocytogenesis plays a pivotal role in regulation of spermatogenesis; however, its details remain relatively obscure in nonrodent species. The equine testis contains approximately 100% more spermatogonia in summer than in winter and appears to be a good model to identify the flexible components of spermatocytogenesis that cause seasonal changes in daily sperm production. Testes were taken from horses in the winter (n = 47) and in summer (n = 43). Tissues were fixed by glutaraldehyde-perfusion and submission in osmium, embedded in Epon or methacrylate, sectioned at 0.5 micron or 5 microns, stained with toluidine blue, and observed using bright-field microscopy. The combined total number of A1, A2, A3, and B1 (A plus B1) spermatogonia/testis and the numbers of B2 spermatogonia or early primary spermatocytes were determined by stereology of Epon sections involving testicular volume density and volume of spermatogonial nuclei. In a subset of horses, different spermatogonial subtypes (A1, A2, A3, and B1) were counted per 100 Sertoli cells in each of the 8 spermatogenic stages and expressed as percentage of all A plus B1 spermatogonia. The number of each spermatogonial subtype/testis for the large series of horses was calculated by multiplying the number of A plus B1 spermatogonia/testis (determined for each horse) by the percentage of that given spermatogonial subtype. Season did not significantly affect the number of any given subtype per 100 Sertoli cells in any stage or percentages of different subtypes of spermatogonia. Numbers of A1 (p less than 0.05), A2, A3, B1, or B2 spermatogonia (p less than 0.01) were greater in the breeding season.(ABSTRACT TRUNCATED AT 250 WORDS)

The seminiferous epithelial cycle and spermatogenesis in goats (Capra hircus)

SUMMARYThe development of the acrosomic system and the spermatid nucleus were used to define 14 stages of the seminiferous epithelial cycle in goats; these stages provided a basis for the examination of the behaviour of different spermatogenic cells which gave an idea of the efficiency of spermatogenesis. Eighteen steps of acrosome development (spermiogenesis) were observed in testicular material stained with periodic acid-Schiff. The first 14 steps were used to classify SEC into 14 (I–XIV) stages which in turn were employed to study the pattern of differentiation of spermatogenic cells by counting them in each stage of the cycle. Three generations of type A (A1, A2, A3), one generation of type intermediate (In) and two generations of type B (B1, B2) spermatogonia could be distinguished. A1 spermatogonia divided primarily in stages IX–X to produce A2 spermatogonia which in turn divided in stages XII–XIII to produce A3 spermatogonia and A, spermatogonia. A3 spermatogonia divided in stage XIV to produce In spermatogonia whereas A1 spermatogonia did not divide till the next cycle but underwent 26·3 % degeneration. In spermatogonia divided to form B1 spermatogonia in stages III–V which further divided to produce B2 spermatogonia in stage VI. Types A3, In and B2 spermatogonia showed 15·0, 25·0 and 25·8% degeneration respectively. B2 spermatogonia divided in stages VII–VIII to produce double the number of primary spermatocytes which persisted without any degeneration till stage XIII of the following cycle and divided at the beginning of stage XIV to form double the number of secondary spermatocytes. These cells divided at the end of stage XIV to form less than double the number of young round spermatids, showing 10·4% degeneration. It is concluded that the development of the acrosomic system as well as the spermatid nucleus could be conveniently used to study the behaviour of spermatogenic cells and that the process of spermatogenesis was less efficient than thought previously.

A study on the different types of spermatogonia in buffalo (Bubalus bubalis).

As a first step to understanding spermatogenesis in the buffalo bull the cytological details of different types of spermatogonia were determined in adult buffalo bulls. Morphological changes in the nuclear details were used as a basis for classifying the different types of spermatogonia. The type A spermatogonia had a spherical to ovoid nucleus with finely granulated chromatin, homogeneously dispersed in the nucleoplasm and having one to two nucleoli adhering to the nuclear membrane. The type A0 spermatogonia were characterized by nuclei containing moderately stained, finely granulated chromatin and a nucleolus attached to the nuclear envelope. The A1 type spermatogonia, on the other hand, have pale stained, finely granulated chromatin with the nucleolus adhering to the nuclear membrane. The nuclei of A2 type spermatogonia resembled those of type A1, but contained coarse granular chromatin dispersed in the pale nucleoplasm. The intermediate type of spermatogonia acquired a central position of the nucleolus, but the chromatin remained coarsely granulated and non-clumped. Three classes of type B (B1-B3) spermatogonia were determined on the degree of clumping of the chromatin and the central position of the nucleolus. The type B1 cells were characterized by nuclei containing a few flakes of lightly stained chromatin and a centrally located nucleolus. The type B2 cells showed comparatively more clumping of chromatin than type B1 spermatogonia, which was dispersed at random in the pale nucleoplasm and along the nuclear envelope. The type B3 spermatogonia demonstrated chromophilic chromatin dispersed in the slightly grey nucleoplasm and adhering along the nuclear membrane. Since there seems to be a succession of events following differentiation of type A1 spermatogonia till the last type B cell differentiates into resting primary spermatocytes, may intermediate stages between the presently described classes of type A (A0-A2) and type B (B1-B3) could also be located in sections of the seminiferous tubules.