Comparison of the spermatogenic process in three different mice strains: Swiss, Balb/C and C57BL/6

Many studies have been trying to establish standard protocols for animal experimentation, especially for animal species or strains, to master research variables with high precision. The main mouse strains used in the field of the biology of reproduction are Swiss, Balb/c, and C57BL/6. Since some of the strains show reproduction limitations, such as the size of the litter, the present study aimed to compare their spermatogenic processes to verify differences regarding the testicular parenchyma and germ cell populations, which could explain low offspring production. In addition, the present study provides additional information concerning the testicular parenchyma of such strains, which consequently would help researchers to choose the most suitable strain for reproductive studies. Six adult male mice were used for each of the strains. After euthanasia, the testes were weighed, fixated with Karnovsky fixative, embedded in methacrylate, sectioned, and stained with toluidine blue/sodium borate 1%. Morphometrical analyses from the testicular parenchyma (seminiferous tubules and interstitium) were made using the software ImageJ. Germ and Sertoli cells populations were counted in seminiferous tubules cross-sections at stage I of the seminiferous epithelium cycle. The lowest body and testicular weights were observed in C57BL/6 animals, followed by Balb/c and Swiss, however, the relative testes, parenchyma, and albuginea weights were significantly lower only in C57BL/6. Despite the seminiferous tubules and seminiferous epithelium proportions were lower in Swiss animals, their relative amount related to the body weight was the same among strains. The total number of germ cells was higher in Swiss animals, reflecting higher spermatogenic yield and daily sperm production. Due to the lower relative number of Sertoli cells, the Swiss animals showed the highest Sertoli cell index and support capacity. On the other hand, the lowest pathological indexes regarding the germ cells were observed in Balb/c animals, followed by Swiss and C57BL/6. In the interstitium, the proportion of blood vessels was lower in Swiss mice, while the lymphatic cell proportion was lower in C57BL/6 animals. Moreover, the highest proportions of Leydig cells and macrophages were noticed in Swiss mice, which may indicate increased testosterone levels. Altogether, such observations must be taken into account when choosing any of the studied strains for reproduction studies.

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Effect of Cryptorchidism on the Histomorphometry, Proliferation, Apoptosis, and Autophagy in Boar Testes

Animals ◽

10.3390/ani11051379 ◽

2021 ◽

Vol 11 (5) ◽

pp. 1379

Author(s):

Xiaorui Fan ◽

Yihui Liu ◽

Meishan Yue ◽

Weidong Yue ◽

Gaoya Ren ◽

...

Keyword(s):

Germ Cells ◽

Sertoli Cells ◽

Proliferating Cell Nuclear Antigen ◽

Molecular Mechanisms ◽

Inguinal Canal ◽

The Body ◽

Nuclear Antigen ◽

Seminiferous Tubules ◽

Mrna Levels ◽

Proliferating Cell

Spontaneous unilateral cryptorchid boars have one testis in the abdomen or inguinal canal, causing its temperature to be at or near the body temperature, which impairs spermatogenesis, although the histomorphometry and molecular mechanisms underlying this process remain unclear. The aim of the present study was to determine the histomorphometry, proliferation, apoptosis, and autophagy alterations in spermatogonia and Sertoli cells in unilateral cryptorchid, scrotal (contrascrotal), and preweaning piglet (preweaning) testes. Histomorphometrical analysis of cryptorchid testes showed that the seminiferous tubules contained only Sertoli cells and a few spermatogonia, but did not contain post-meiotic germ cells. The number of spermatogonia markedly decreased, and the number of Sertoli cells did not change remarkably in cryptorchid testes. TUNEL assay results showed that apoptosis signals were predominantly observed in spermatogonia. In cryptorchid and contrascrotal testes, proliferating cell nuclear antigen (PCNA) and LC3 were located in spermatogonia. The number of PCNA-positive, TUNEL-positive, and LC3-positive germ cells was low, and the protein and mRNA levels of PCNA and LC3 were significantly decreased in cryptorchid testes. Taken together, the number of Sertoli cells did not change remarkably, whereas the number of germ cells decreased in the cryptorchid testes, compared with that in the contrascrotal testes. Insufficient proliferation, excessive apoptosis, and autophagy were involved in the regulation of the decrease in spermatogonia in cryptorchid boar testes.

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BRG1 Is Dispensable for Sertoli Cell Development and Functions in Mice

International Journal of Molecular Sciences ◽

10.3390/ijms21124358 ◽

2020 ◽

Vol 21 (12) ◽

pp. 4358

Author(s):

Shuai Wang ◽

Pengxiang Wang ◽

Dongli Liang ◽

Yuan Wang

Keyword(s):

Sertoli Cell ◽

Germ Cells ◽

Sertoli Cells ◽

Cell Development ◽

Conditional Knockout ◽

Mouse Strains ◽

Seminiferous Tubules ◽

Supporting Cells ◽

Knockout Mouse Model ◽

Conditional Knockout Mouse

Sertoli cells are somatic supporting cells in spermatogenic niche and play critical roles in germ cell development, but it is yet to be understood how epigenetic modifiers regulate Sertoli cell development and contribution to spermatogenesis. BRG1 (Brahma related gene 1) is a catalytic subunit of the mammalian SWI/SNF chromatin remodeling complex and participates in transcriptional regulation. The present study aimed to define the functions of BRG1 in mouse Sertoli cells during mouse spermatogenesis. We found that BRG1 protein was localized in the nuclei of both Sertoli cells and germ cells in seminiferous tubules. We further examined the requirement of BRG1 in Sertoli cell development using a Brg1 conditional knockout mouse model and two Amh-Cre mouse strains to specifically delete Brg1 gene from Sertoli cells. We found that the Amh-Cre mice from Jackson Laboratory had inefficient recombinase activities in Sertoli cells, while the other Amh-Cre strain from the European Mouse Mutant Archive achieved complete Brg1 deletion in Sertoli cells. Nevertheless, the conditional knockout of Brg1 from Sertoli cells by neither of Amh-Cre strains led to any detectable abnormalities in the development of either Sertoli cells or germ cells, suggesting that BRG1-SWI/SNF complex is dispensable to the functions of Sertoli cells in spermatogenesis.

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Cycle of the seminiferous epithelium in a marsupial species, the brushtail possum (Trichosurus vulpecula), and estimation of its duration

Reproduction Fertility and Development ◽

10.1071/rd02059 ◽

2004 ◽

Vol 16 (3) ◽

pp. 307 ◽

Cited By ~ 6

Author(s):

Minjie Lin ◽

Amanda Harman ◽

Terry P. Fletcher

Keyword(s):

Germ Cells ◽

Cross Sections ◽

Seminiferous Epithelium ◽

Seminiferous Tubules ◽

Brushtail Possum ◽

Trichosurus Vulpecula ◽

Type B ◽

Semithin Sections ◽

Marsupial Species ◽

Kinetics Of

We investigated the cycle of the seminiferous epithelium in a marsupial, namely the brushtail possum (Trichosurus vulpecula), using semithin sections of seminiferous tubules embedded in Spurr’s resin. Using 14 steps of spermatid development as markers, we were able to class tubular cross-sections into 10 well-defined stages of the seminiferous epithelial cycle. The duration of one cycle was 13.5 days, as determined by injections of [3H]-thymidine and autoradiographic examination of the most advanced sperm cells at 2 h and 17 days after injection. The durations of stages I–X were 21.4, 66.4, 54.1, 47.0, 29.8, 28.5, 25.3, 25.0, 12.0 and 15.9 h, respectively, estimated by the relative percentage of occurrence of each stage. It was estimated that the life spans of the main germ cells were as follows: type B spermatogonia, 5.4 days; primary spermatocytes, 16.7 days; secondary spermatocytes, 0.7 days; and spermatids, 21.4 days. The results suggest that the kinetics of spermatogenesis in marsupials show a similar pattern to that in eutherians.

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Minireview: Transcriptional Regulation of Gonadal Development and Differentiation

Endocrinology ◽

10.1210/en.2004-1454 ◽

2005 ◽

Vol 146 (3) ◽

pp. 1035-1042 ◽

Cited By ~ 60

Author(s):

Susan Y. Park ◽

J. Larry Jameson

Keyword(s):

Germ Cells ◽

Sertoli Cells ◽

Leydig Cells ◽

Cell Types ◽

Gonadal Development ◽

Targeted Mutagenesis ◽

Seminiferous Tubules ◽

Molecular Pathways ◽

Theca Cells ◽

Main Cell

The embryonic gonad is undifferentiated in males and females until a critical stage when the sex chromosomes dictate its development as a testis or ovary. This binary developmental process provides a unique opportunity to delineate the molecular pathways that lead to distinctly different tissues. The testis comprises three main cell types: Sertoli cells, Leydig cells, and germ cells. The Sertoli cells and germ cells reside in seminiferous tubules where spermatogenesis occurs. The Leydig cells populate the interstitial compartment and produce testosterone. The ovary also comprises three main cell types: granulosa cells, theca cells, and oocytes. The oocytes are surrounded by granulosa and theca cells in follicles that grow and differentiate during characteristic reproductive cycles. In this review, we summarize the molecular pathways that regulate the distinct differentiation of these cell types in the developing testis and ovary. In particular, we focus on the transcription factors that initiate these cascades. Although most of the early insights into the sex determination pathway were based on human mutations, targeted mutagenesis in mouse models has revealed key roles for genes not anticipated to regulate gonadal development. Defining these molecular pathways provides the foundation for understanding this critical developmental event and provides new insight into the causes of gonadal dysgenesis.

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Studies of Sl/Sld in equilibrium with +/+ mouse aggregation chimaeras. II. Effect of the steel locus on spermatogenesis

Development ◽

10.1242/dev.102.1.117 ◽

1988 ◽

Vol 102 (1) ◽

pp. 117-126 ◽

Cited By ~ 2

Author(s):

H. Nakayama ◽

H. Kuroda ◽

H. Onoue ◽

J. Fujita ◽

Y. Nishimune ◽

...

Keyword(s):

Mast Cells ◽

Germ Cell ◽

Germ Cells ◽

Cross Sections ◽

Sertoli Cells ◽

Type A ◽

Precursor Cells ◽

Intrinsic Defect ◽

Serial Sections ◽

Type A Spermatogonia

Mutant mice of Sl/Sld genotype are deficient in melanocytes, erythrocytes, mast cells and germ cells. Deficiency of melanocytes, erythrocytes and mast cells is not attributable to an intrinsic defect in their precursor cells but to a defect in the tissue environment that is necessary for migration, proliferation and/or differentiation. We investigated the mechanism of germ cell deficiency in male Sl/Sld mice by producing aggregation chimaeras from Sl/Sld and +/+ embryos. Chimaeric mice with apparent white stripes were obtained. Two of four such chimaeras were fertile and the phenotypes of resulting progenies showed that some Sl/Sld germ cells had differentiated into functioning sperms in the testis of the chimaeras. In cross sections of the testes of chimaeras, both differentiated and nondifferentiated tubules were observed. However, the proportions of type A spermatogonia to Sertoli cells in both types of tubules were comparable to the values observed in differentiated tubules of normal +/+ mice. We reconstructed the whole length of four tubules from serial sections. Differentiated and nondifferentiated segments alternated in a single tubule. The shortest differentiated segment contained about 180 Sertoli cells and the shortest nondifferentiated segment about 150 Sertoli cells. These results suggest that Sertoli cells of either Sl/Sld or +/+ genotype make discrete patches and that differentiation of type A spermatogonia does not occur in patches of Sl/Sld Sertoli cells.

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Developmental stage- and spermatogenic cycle-specific expression of transcription factor GATA-1 in mouse Sertoli cells

Development ◽

10.1242/dev.120.7.1759 ◽

1994 ◽

Vol 120 (7) ◽

pp. 1759-1766 ◽

Cited By ~ 6

Author(s):

K. Yomogida ◽

H. Ohtani ◽

H. Harigae ◽

E. Ito ◽

Y. Nishimune ◽

...

Keyword(s):

Transcription Factor ◽

Developmental Stage ◽

Sertoli Cell ◽

Germ Cells ◽

Sertoli Cells ◽

Transcriptional Activation ◽

Germ Line ◽

Mouse Testis ◽

Seminiferous Tubules ◽

Specific Expression

GATA-1 is an essential factor for the transcriptional activation of erythroid-specific genes, and is also abundantly expressed in a discrete subset of cells bordering the seminiferous epithelium in tubules of the murine testis. In examining normal and germ-line defective mutant mice, we show here that GATA-1 is expressed only in the Sertoli cell lineage in mouse testis. GATA-1 expression in Sertoli cells is induced concomitantly with the first wave of spermatogenesis, and GATA-1-positive cells are uniformly distributed among all tubules during prepubertal testis development. However, the number of GATA-1-positive cells declines thereafter and were found only in the peripheral zone of seminiferous tubules in stages VII, VIII and IX of spermatogenesis in the adult mouse testis. In contrast, virtually every Sertoli cell in mutant W/Wv, jsd/jsd or cryptorchid mice (all of which lack significant numbers of germ cells) expresses GATA-1, thus showing that the expression of this transcription factor is negatively controlled by the maturing germ cells. These observations suggest that transcription factor GATA-1 is a developmental stage- and spermatogenic cycle-specific regulator of gene expression in Sertoli cells.

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Histochemical localization of enzymes of various metabolic pathways in the testes of buffaloes, goats and rams

The Journal of Agricultural Science ◽

10.1017/s002185960004257x ◽

1984 ◽

Vol 102 (2) ◽

pp. 269-274 ◽

Cited By ~ 1

Author(s):

G. S. Bilaspuri ◽

S. S. Guraya

Keyword(s):

Germ Cells ◽

Sertoli Cells ◽

Metabolic Pathways ◽

Maximum Activity ◽

The Other ◽

Seminiferous Tubules ◽

Interstitial Tissue ◽

Cell Type ◽

Glucose 6 Phosphate Dehydrogenase ◽

Enzyme Species

SummaryIsocitrate dehydrogenase (ICDH), succinate dehydrogenase (SDH), malate dehydrogenase (MDH), glutamate dehydrogenase (GDH), β-hydroxybutyrate dehydrogenase (β-OH-BDH) and glucose-6-phosphate dehydrogenase (G-6-PDH) were histochemically located in the testes of buffaloes, goats and rams. The enzyme activities varied with the enzyme, species and cell type. The activities in the seminiferous tubules were correlated with the stages of seminiferous epithelial cycle (SEC). During this cycle, the activities in the Sertoli cells, spermatogonia and spermatocytes remained unaltered in contrast to those in the spermatids. The activities of SDH, ICDH and MDH were relatively greater in buffalo, while goat and ram resembled each other quite closely. ICDH and MDH preferred NADP to NAD. In the three species, the activities of ICDH, SDH and MDH generally followed an increasing order. G-6-PDH was greater in the interstitial tissue of buffalo than in goat and ram; the maximum activity of this enzyme in each species was found in the spermatogonia. In comparison with G-6-PDH, GDH was less evident in the interstitial tissue of buffalo and goat; Sertoli cells and spermatogonia also showed relatively less MDH activity whereas the other germ cells may have relatively less, similar or more, GDH activity depending on the species. β-OHBDH activity was similar in the interstitial tissue of the three species, but in the seminiferous tubule, the activity was less in goat. But for GDH and β-OH-BDH which could show different results, the activities of other enzymes generally decreased from spermatogonia through spermatocytes to spermatids but increased during spermiogenesis. In spermatozoa, the enzymes were observed only in the mid-piece. The possible physiological significance of the results is discussed in relation to different metabolic pathways.

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148. HORMONALLY REGULATED miRNAS TARGET THE TUBULOBULBAR COMPLEX IN THE TESTIS

Reproduction Fertility and Development ◽

10.1071/srb10abs148 ◽

2010 ◽

Vol 22 (9) ◽

pp. 66

Author(s):

P. K. Nicholls ◽

P. G. Stanton ◽

K. L. Walton ◽

R. I. McLachlan ◽

L. O'Donnell ◽

...

Keyword(s):

Sertoli Cell ◽

Sertoli Cells ◽

Hormonal Regulation ◽

Focal Adhesions ◽

Intercellular Junctions ◽

Protein Translation ◽

Seminiferous Epithelium ◽

Seminiferous Tubules ◽

Micro Rnas

Spermatogenesis is absolutely dependent on follicle stimulating hormone (FSH) and androgens; acute suppression of these hormones inhibits germ cell development and thus sperm production. The removal of intercellular junctions and release of spermatids by the Sertoli cell, a process known as spermiation, is particularly sensitive to acute hormone suppression(1). To define the molecular mechanisms that mediate FSH and androgen effects in the testis, we investigated the expression and hormonal regulation of micro-RNAs (miRNA), small non-coding RNAs that regulate protein translation and modify cellular responses. By array analysis, we identified 23 miRNAs that were upregulated >2-fold in stage VIII seminiferous tubules following hormone suppression, and in vitro in primary Sertoli cells. We subsequently validated the expression and hormonal regulation of several miRNAs, including miR-23b, -30d and -690 by quantitative PCR in primary Sertoli cells. Bioinformatic analysis of potential targets of hormonally-suppressed miRNAs identified genes associated with Focal adhesions (54 genes, P = –ln(17.97)) and the Regulation of the actin cytoskeleton (52 genes, P = –ln(10.16)), processes known to be intimately associated with adhesion of spermatids to Sertoli cells(2, 3). Furthermore, this analysis identified numerous components of the testicular tubulobulbar complex (TBC) as being targets of hormonally sensitive miRNAs. The TBC is a podosome-like structure between Sertoli and adjacent spermatids in the testis, which internalises intact inter-cellular junctions by endocytotic mechanisms prior to spermiation(4). We then demonstrate the hormonal regulation of predicted miRNA target proteins, and validate novel inhibitory miRNA interactions with Pten, nWASP, Eps15 and Picalm by luciferase knockdown in vitro. We hypothesise that hormonally suppressed miRNAs inhibit TBC function, and subsequently, endocytosis of intercellular junctions. In conclusion, we have demonstrated that hormonal suppression in the testis stimulates the expression of a subset of Sertoli cell miRNAs that are likely regulators of cell adhesion protein networks involved in spermiation. (1) Saito K, O’Donnell L, McLachlan RI, Robertson DM 2000 Spermiation failure is a major contributor to early spermatogenic suppression caused by hormone withdrawal in adult rats. Endocrinology 141: 2779–2.(2) O’Donnell L, Stanton PG, Bartles JR, Robertson DM 2000 Sertoli cell ectoplasmic specializations in the seminiferous epithelium of the testosterone-suppressed adult rat. Biol Reprod 63: 99–108.(3) Beardsley A, Robertson DM, O’Donnell L 2006 A complex containing alpha6beta1-integrin and phosphorylated focal adhesion kinase between Sertoli cells and elongated spermatids during spermatid release from the seminiferous epithelium. J Endocrinol 190(3): 759–70.(4) Young JS, Guttman JA, Vaid KS, Vogl AW 2009 Tubulobulbar complexes are intercellular podosome-like structures that internalize intact intercellular junctions during epithelial remodeling events in the rat testis. Biol Reprod 80: 162–74.

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A novel Amh-Treck transgenic mouse line allows toxin-dependent loss of supporting cells in gonads

Reproduction ◽

10.1530/rep-14-0171 ◽

2014 ◽

Vol 148 (6) ◽

pp. H1-H9 ◽

Cited By ~ 8

Author(s):

Mai Shinomura ◽

Kasane Kishi ◽

Ayako Tomita ◽

Miyuri Kawasumi ◽

Hiromi Kanezashi ◽

...

Keyword(s):

Germ Cells ◽

Granulosa Cells ◽

Sertoli Cells ◽

Seminiferous Tubules ◽

Specific Cell ◽

Partial Recovery ◽

Dependent Manner ◽

Mouse Line ◽

Cell Degeneration

Cell ablation technology is useful for studying specific cell lineages in a developing organ in vivo. Herein, we established a novel anti-Müllerian hormone (AMH)-toxin receptor-mediated cell knockout (Treck) mouse line, in which the diphtheria toxin (DT) receptor was specifically activated in Sertoli and granulosa cells in postnatal testes and ovaries respectively. In the postnatal testes of Amh-Treck transgenic (Tg) male mice, DT injection induced a specific loss of the Sertoli cells in a dose-dependent manner, as well as the specific degeneration of granulosa cells in the primary and secondary follicles caused by DT injection in Tg females. In the testes with depletion of Sertoli cell, germ cells appeared to survive for only several days after DT treatment and rapidly underwent cell degeneration, which led to the accumulation of a large amount of cell debris within the seminiferous tubules by day 10 after DT treatment. Transplantation of exogenous healthy Sertoli cells following DT treatment rescued the germ cell loss in the transplantation sites of the seminiferous epithelia, leading to a partial recovery of the spermatogenesis. These results provide not only in vivo evidence of the crucial role of Sertoli cells in the maintenance of germ cells, but also show that the Amh-Treck Tg line is a useful in vivo model of the function of the supporting cell lineage in developing mammalian gonads.

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Effect of thyroid hormone on the pre- and post-natal development of the rat testis

Journal of Endocrinology ◽

10.1677/joe.0.1290035 ◽

1991 ◽

Vol 129 (1) ◽

pp. 35-NP ◽

Cited By ~ 70

Author(s):

S. Francavilla ◽

G. Cordeschi ◽

G. Properzi ◽

L. Di Cicco ◽

E. A. Jannini ◽

...

Keyword(s):

Germ Cells ◽

Sertoli Cells ◽

Serum Levels ◽

Seminiferous Tubules ◽

Testicular Development ◽

Postnatal Life ◽

Fetal Life ◽

Newborn Rats ◽

Total Thyroxine ◽

Delayed Maturation

ABSTRACT The relationship between thyroid function and testicular development in the rat was investigated. Hypothyroidism was induced during fetal or postnatal life by adding methimazole (MMI) to the drinking water of pregnant or lactating mothers. A group of newborn rats was treated with MMI and i.p. injections of l-tri-iodothyronine (l-T3). Hypothyroidism was shown by the reduced serum levels of total T3 and of total thyroxine (T4) in pregnant mothers and in pubertal rats. Testes were studied using light microscopy at 18 and 21 days post coitum or during puberty (21, 35 and 50 days after birth); serum levels of gonadotrophins were also evaluated in pubertal rats. Hypothyroidism had no effect on testicular development during fetal life and when induced in newborn rats it was associated at puberty with reduced serum levels of FSH and LH and with delayed maturation of the testis compared with control rats. The delay in maturation consisted of a reduction in the diameter of seminiferous tubules, and a reduction in the number of germ cells per tubule; this was associated with increased degeneration and arrested maturation of germ cells. In addition, Sertoli cells demonstrated retarded development, as indicated by a delay in the appearance of cytoplasmic lipids and in the development of a tubule lumen. Hormonal and morphological abnormalities were absent in rats treated with MMI plus l-T3. In conclusion, hypothyroidism occurring soon after birth caused reduced levels of gonadotrophins in the serum and a delay in pubertal spermatogenesis, possibly due to retarded differentiation of the Sertoli cells. Journal of Endocrinology (1991) 129, 35–42

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