scholarly journals Reduction of Spermatogenesis but Not Fertility in Creb3l4-Deficient Mice

2005 ◽  
Vol 25 (17) ◽  
pp. 7657-7664 ◽  
Author(s):  
Ibrahim M. Adham ◽  
Thomas J. Eck ◽  
Kerstin Mierau ◽  
Nicole Müller ◽  
Mahmoud A. Sallam ◽  
...  
Keyword(s):  
Germ Cells ◽  
Intracellular Signaling ◽  
Fluorescent Protein ◽  
Survival Rates ◽  
Seminiferous Tubules ◽  
Wild Type ◽  
Specific Expression ◽  
Term Survival ◽  
Mouse Prostate ◽  
Deficient Mice

ABSTRACT Creb3l4 belongs to the CREB/ATF family of transcription factors that are involved in mediating transcription in response to intracellular signaling. This study shows that Creb3l4 is expressed at low levels in all organs and in different stages of embryogenesis but is present at very high levels in the testis, particularly in postmeiotic male germ cells. In contrast to CREB3L4 in the human prostate, of which specific expression was detected, Creb3l4 transcripts in the mouse prostate could be detected only by RT-PCR. To identify the physiological function of Creb3l4, the murine gene was inactivated by replacement with the gene encoding green fluorescent protein. Surprisingly, Creb3l4-deficient mice were born at expected ratios, were healthy, and displayed normal long-term survival rates. Despite a significant reduction in the number of spermatozoa in the epididymis of Creb3l4 − / − mice, the breeding of mutant males with wild-type females was productive and the average litter size was not significantly altered in comparison to wild-type littermates. Further analyses revealed that the seminiferous tubules of Creb3l4 − / − mice contained all of the developmental stages, though there was evidence for increased apoptosis of meiotic/postmeiotic germ cells. These results suggest that Creb3l4 plays a role in male germ cell development, but its loss is insufficient to completely compromise the production of spermatozoa.

scholarly journals MFN1 and MFN2 Are Dispensable for Sperm Development and Functions in Mice

2021 ◽  
Vol 22 (24) ◽  
pp. 13507
Author(s):  
Junru Miao ◽  
Wei Chen ◽  
Pengxiang Wang ◽  
Xin Zhang ◽  
Lei Wang ◽  
...  
Keyword(s):  
Germ Cells ◽  
Knockout Mice ◽  
Seminiferous Tubules ◽  
Wild Type ◽  
Double Knockout ◽  
Wild Type Littermate ◽  
Mitofusin 2 ◽  
Sperm Development ◽  
Mitofusin 1 ◽  
Deficient Mice

MFN1 (Mitofusin 1) and MFN2 (Mitofusin 2) are GTPases essential for mitochondrial fusion. Published studies revealed crucial roles of both Mitofusins during embryonic development. Despite the unique mitochondrial organization in sperm flagella, the biological requirement in sperm development and functions remain undefined. Here, using sperm-specific Cre drivers, we show that either Mfn1 or Mfn2 knockout in haploid germ cells does not affect male fertility. The Mfn1 and Mfn2 double knockout mice were further analyzed. We found no differences in testis morphology and weight between Mfn-deficient mice and their wild-type littermate controls. Spermatogenesis was normal in Mfn double knockout mice, in which properly developed TRA98+ germ cells, SYCP3+ spermatocytes, and TNP1+ spermatids/spermatozoa were detected in seminiferous tubules, indicating that sperm formation was not disrupted upon MFN deficiency. Collectively, our findings reveal that both MFN1 and MFN2 are dispensable for sperm development and functions in mice.

scholarly journals Involvement of a membrane skeletal protein, 4.1G, for Sertoli/germ cell interaction

Reproduction ◽  
2010 ◽  
Vol 139 (5) ◽  
pp. 883-892 ◽  
Author(s):  
Nobuo Terada ◽  
Nobuhiko Ohno ◽  
Sei Saitoh ◽  
Yurika Saitoh ◽  
Masayuki Komada ◽  
...  
Keyword(s):  
Adhesion Molecule ◽  
Germ Cells ◽  
Cell Membranes ◽  
Cell Interaction ◽  
Seminiferous Tubules ◽  
Wild Type ◽  
Skeletal Protein ◽  
Deficient Mice

We previously reported that a membrane skeletal protein, 4.1G (also known as EPB41L2), is immunolocalized in mouse seminiferous tubules. In this study, the 4.1G immunolocalizaiton was precisely evaluated at various stages of the mouse seminiferous epithelial cycle with ‘in vivocryotechnique’ and also with pre-embedding immunoelectron microscopy in testicular tissues whose ultrastructures were well preserved with glycerol treatment before cryosectioning. In addition, 4.1G-deficient mice were produced, and the morphology of their seminiferous tubules was also evaluated. The 4.1G immunolocalization was different among stages, indicating that it was not only along cell membranes of Sertoli cells, but also those of spermatogonia and early spermatocytes. To confirm the 4.1G immunolocalization in germ cells,in vitroculture of spermatogonial stem cells (SSCs) was used for immunocytochemistry and immunoblotting analysis. In the cultured SSCs, 4.1G was clearly expressed and immunolocalized along cell membranes, especially at mutual attaching regions. In testicular tissues, cell adhesion molecule-1 (CADM1), an intramembranous adhesion molecule, was colocalized on basal parts of the seminiferous tubules and immunoprecipitated with 4.1G in the tissue lysate. Interestingly, in the 4.1G-deficient mice, histological manifestation of the seminiferous tubules was not different from that in wild-type mice, and the CADM1 was also immunolocalized in the same pattern as that in the wild-type. Moreover, the 4.1G-deficient male mice were fertile. These results were probably due to functional redundancy of unknown membrane skeletal molecules in germ cells. Thus, a novel membrane skeletal protein, 4.1G, was found in germ cells, and considering its interaction with CADM family, it probably has roles in attachment of both Sertoli–germ and germ–germ cells.

scholarly journals Tissue-specific expression of B7x protects from CD4 T cell–mediated autoimmunity

2011 ◽  
Vol 208 (8) ◽  
pp. 1683-1694 ◽  
Author(s):  
Joyce Wei ◽  
P’ng Loke ◽  
Xingxing Zang ◽  
James P. Allison
Keyword(s):  
T Cells ◽  
Pancreatic Islets ◽  
Peripheral Tolerance ◽  
Wild Type ◽  
Autoimmune Encephalomyelitis ◽  
Specific Expression ◽  
Tissue Specific ◽  
Disease Induction ◽  
Deficient Mice ◽  
Experimental Autoimmune

B7x, an inhibitory member of the B7/CD28 superfamily, is highly expressed in a broad range of nonhematopoietic organs, suggesting a role in maintaining peripheral tolerance. As endogenous B7x protein is expressed in pancreatic islets, we investigated whether the molecule inhibits diabetogenic responses. Transfer of disease-inducing BDC2.5 T cells into B7x-deficient mice resulted in a more aggressive form of diabetes than in wild-type animals. This exacerbation of disease correlated with higher frequencies of islet-infiltrating Th1 and Th17 cells. Conversely, local B7x overexpression inhibited the development of autoimmunity, as crossing diabetes-susceptible BDC2.5/B6g7 mice to animals overexpressing B7x in pancreatic islets abrogated disease induction. This protection was caused by the inhibition of IFN-γ production by CD4 T cells and not to a skewing or expansion of Th2 or regulatory T cells. The suppressive function of B7x was also supported by observations from another autoimmune model, experimental autoimmune encephalomyelitis, in which B7x-deficient mice developed exacerbated disease in comparison with wild-type animals. Analysis of central nervous system–infiltrating immune cells revealed that the loss of endogenous B7x resulted in expanded Th1 and Th17 responses. Data from these two autoimmune models provide evidence that B7x expression in the periphery acts as an immune checkpoint to prevent tissue-specific autoimmunity.

A γGT-AT1A receptor transgene protects renal cortical structure in AT1 receptor-deficient mice

2004 ◽  
Vol 18 (3) ◽  
pp. 290-298 ◽  
Author(s):  
Thu H. Le ◽  
Michael I. Oliverio ◽  
Hyung-Suk Kim ◽  
Harmony Salzler ◽  
Rajesh C. Dash ◽  
...  
Keyword(s):  
Blood Pressure ◽  
Transgenic Mice ◽  
Proximal Tubule ◽  
Physiological Role ◽  
Renal Proximal Tubule ◽  
Hypoxanthine Phosphoribosyl Transferase ◽  
Wild Type ◽  
Specific Expression ◽  
Low Levels ◽  
Deficient Mice

To understand the physiological role of angiotensin type 1 (AT1) receptors in the proximal tubule of the kidney, we generated a transgenic mouse line in which the major murine AT1 receptor isoform, AT1A, was expressed under the control of the P1 portion of the γ-glutamyl transpeptidase (γGT) promoter. In transgenic mice, this promoter has been shown to confer cell-specific expression in epithelial cells of the renal proximal tubule. To avoid random integration of multiple copies of the transgene, we used gene targeting to produce mice with a single-copy transgene insertion at the hypoxanthine phosphoribosyl transferase ( Hprt) locus on the X chromosome. The physiological effects of the γGT-AT1A transgene were examined on a wild-type background and in mice with targeted disruption of one or both of the murine AT1 receptor genes ( Agtr1a and Agtr1b). On all three backgrounds, γGT-AT1A transgenic mice were healthy and viable. On the wild-type background, the presence of the transgene did not affect development, blood pressure, or kidney structure. Despite relatively low levels of expression in the proximal tubule, the transgene blunted the increase in renin expression typically seen in AT1-deficient mice and partially rescued the kidney phenotype associated with Agtr1a−/− Agtr1b−/− mice, significantly reducing cortical cyst formation by more than threefold. However, these low levels of cell-specific expression of AT1 receptors in the renal proximal tubule did not increase the low blood pressures or abolish sodium sensitivity, which are characteristic of AT1 receptor-deficient mice. Although our studies do not clearly identify a role for AT1 receptors in the proximal tubules of the kidney in blood pressure homeostasis, they support a major role for these receptors in modulating renin expression and in maintaining structural integrity of the renal cortex.

Lower skeletal muscle mass in male transgenic mice with muscle-specific overexpression of myostatin

2003 ◽  
Vol 285 (4) ◽  
pp. E876-E888 ◽  
Author(s):  
Suzanne Reisz-Porszasz ◽  
Shalender Bhasin ◽  
Jorge N. Artaza ◽  
Ruoqing Shen ◽  
Indrani Sinha-Hikim ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Transgenic Mice ◽  
Muscle Mass ◽  
Skeletal Muscle Mass ◽  
Fluorescent Protein ◽  
Male Mice ◽  
Wild Type ◽  
Specific Expression ◽  
Muscle Weight ◽  
Myostatin Gene

Mutations in the myostatin gene are associated with hypermuscularity, suggesting that myostatin inhibits skeletal muscle growth. We postulated that increased tissue-specific expression of myostatin protein in skeletal muscle would induce muscle loss. To investigate this hypothesis, we generated transgenic mice that overexpress myostatin protein selectively in the skeletal muscle, with or without ancillary expression in the heart, utilizing cDNA constructs in which a wild-type (MCK/Mst) or mutated muscle creatine kinase (MCK-3E/Mst) promoter was placed upstream of mouse myostatin cDNA. Transgenic mice harboring these MCK promoters linked to enhanced green fluorescent protein (EGFP) expressed the reporter protein only in skeletal and cardiac muscles (MCK) or in skeletal muscle alone (MCK-3E). Seven-week-old animals were genotyped by PCR of tail DNA or by Southern blot analysis of liver DNA. Myostatin mRNA and protein, measured by RT-PCR and Western blot, respectively, were significantly higher in gastrocnemius, quadriceps, and tibialis anterior of MCK/Mst-transgenic mice compared with wild-type mice. Male MCK/Mst-transgenic mice had 18–24% lower hind- and forelimb muscle weight and 18% reduction in quadriceps and gastrocnemius fiber cross-sectional area and myonuclear number (immunohistochemistry) than wild-type male mice. Male transgenic mice with mutated MCK-3E promoter showed similar effects on muscle mass. However, female transgenic mice with either type of MCK promoter did not differ from wild-type controls in either body weight or skeletal muscle mass. In conclusion, increased expression of myostatin in skeletal muscle is associated with lower muscle mass and decreased fiber size and myonuclear number, decreased cardiac muscle mass, and increased fat mass in male mice, consistent with its role as an inhibitor of skeletal muscle mass. The mechanism of gender specificity remains to be clarified.

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

Development ◽  
1994 ◽  
Vol 120 (7) ◽  
pp. 1759-1766 ◽  
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.

scholarly journals Epiplakin Is Dispensable for Skin Barrier Function and for Integrity of Keratin Network Cytoarchitecture in Simple and Stratified Epithelia

2006 ◽  
Vol 26 (2) ◽  
pp. 559-568 ◽  
Author(s):  
Daniel Spazierer ◽  
Peter Fuchs ◽  
Siegfried Reipert ◽  
Irmgard Fischer ◽  
Matthias Schmuth ◽  
...  
Keyword(s):  
Actin Filaments ◽  
Primary Cultures ◽  
Embryonic Stem ◽  
Skin Barrier ◽  
Skin Barrier Function ◽  
Wild Type ◽  
Primary Hepatocytes ◽  
Specific Expression ◽  
Keratin Filament ◽  
Deficient Mice

ABSTRACT Epiplakin, a giant epithelial protein of >700 kDa, belongs to the plakin family of cytolinker proteins. It represents an atypical family member, however, as it consists entirely of plakin repeat domains but lacks any of the other domains commonly shared by plakins. Hence, its putative function as a cytolinker protein remains to be shown. To investigate epiplakin's biological role, we generated epiplakin-deficient mice by gene targeting in embryonic stem cells. Epiplakin-deficient mice were viable and fertile, without developing any discernible phenotype. Ultrastructurally, their epidermis revealed no differences compared to wild-type littermates, and cornified envelopes isolated from skin showed no alterations in shape or stability. Furthermore, neither embryonal formation nor later function of the epithelial barrier was affected. In primary cultures of epiplakin-deficient keratinocytes, the organization of actin filaments, microtubules, and keratin networks was found to be normal. Similarly, no alterations in keratin network organization were observed in simple epithelia of small intestine and liver or in primary hepatocytes. We conclude that, despite epiplakin's abundant and highly specific expression in stratified and simple epithelia, its absence in mice does not lead to severe skin dysfunctions, nor has it detectable consequences for keratin filament organization and cytoarchitecture of cells.

250 ALL REGIONS OF THE MOUSE EPIDIDYMIS ARE ABLE TO PHAGOCYTIZE IMMATURE SPERMATOGENIC CELLS

2013 ◽  
Vol 25 (1) ◽  
pp. 272
Author(s):  
P. Ramos-Ibeas ◽  
E. Pericuesta ◽  
R. Fernandez-Gonzalez ◽  
M. A. Ramirez ◽  
A. Gutierrez-Adan
Keyword(s):  
Quality Control ◽  
Green Fluorescent Protein ◽  
Germ Cells ◽  
Enhanced Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Sperm Quality ◽  
Seminiferous Tubules ◽  
Spermatogenic Cells ◽  
Green Fluorescent ◽  
Immature Cells

Successful mammalian fertilization requires gametes with an intact structure and functionality. Although it is well known that epididymal functions are sperm maturation, sustenance, transport, and storage, there is controversial information about its role in sperm quality control, and it has been suggested that some regions of the rat epididymis are able to phagocytize germ cells. Our objective was to analyse whether different segments of the mouse epididymal epithelium act as a selection barrier for abnormal spermatogenic cells by removing immature cells from the lumen by phagocytosis. To detect the presence of immature germ cells along the epididymis, transgenic mice expressing enhanced green fluorescent protein under a Deleted in Azoospermia-Like (mDazl) promoter were generated. The transgenic animals express specifically enhanced green fluorescent protein in spermatogonias, spermatocytes, and spermatids; thus, immature spermatogenic cells can be easily identified by fluorescence microscopy. Colchicine, a microtubule disruptor that leads to severe alterations in the architecture of the seminiferous tubules, was administered in the rete testis to induce the release of immature germ cells into the epididymis. Mice were killed daily, from Day 1 to 8 post-administration, and epididymides were collected and observed under a fluorescence stereoscope to determine the transit of immature germ cells along the epididymis. Epididymides from control mice without colchicine administration were also collected. Fluorescent immature germ cells were present in the caput epididymis 24 h after colchicine administration, and they progressed through the corpus and cauda, leaving the epididymis 7 days after colchicine administration. After fluorescence observation, epididymides were fixed, sectioned, and stained with hematoxylin solution. Immature germ cells and phagosomes were not observed in control epididymides. By contrast, the presence of phagosomes in the principal cells of the epididymal epithelium containing immature germ cells in different degrees of degradation was observed by light microscopy in mice injected with colchicine. Phagocytosis was observed along the epididymis following the main wave of fluorescent immature cells. Thus, when immature cells had reached the corpus epididymis, phagocytosis was detected in several segments of the caput epididymis. Later, once the immature cells had arrived to the cauda epididymis or had abandoned the epididymis, phagocytosis was observed in the corpus and cauda epididymis. The presence of phagosomes was observed in all epididymal tubules within a phagocytosis area. In conclusion, we demonstrated that the epididymal epithelium is engaged in sperm quality control by clearing immature germ cells after a massive shedding into the epididymal lumen, and that this phenomenon is not restricted to a specific segment of the epididymis.

scholarly journals Impaired Pulmonary NF-κB Activation in Response to Lipopolysaccharide in NADPH Oxidase-Deficient Mice

2001 ◽  
Vol 69 (10) ◽  
pp. 5991-5996 ◽  
Author(s):  
M. Audrey Koay ◽  
John W. Christman ◽  
Brahm H. Segal ◽  
Annapurna Venkatakrishnan ◽  
Thomas R. Blackwell ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Intracellular Signaling ◽  
Lung Inflammation ◽  
Nuclear Translocation ◽  
Binding Activity ◽  
Wild Type ◽  
Inflammatory Protein ◽  
Macrophage Inflammatory Protein ◽  
Deficient Mice

ABSTRACT Reactive oxygen species (ROS) are thought to be involved in intracellular signaling, including activation of the transcription factor NF-κB. We investigated the role of NADPH oxidase in the NF-κB activation pathway by utilizing knockout mice (p47phox−/−) lacking the p47phox component of NADPH oxidase. Wild-type (WT) controls and p47phox−/−mice were treated with intraperitoneal (i.p.) Escherichia coli lipopolysaccharide (LPS) (5 or 20 μg/g of body weight). LPS-induced NF-κB binding activity and accumulation of RelA in nuclear protein extracts of lung tissue were markedly increased in WT compared to p47phox−/− mice 90 min after treatment with 20 but not 5 μg of i.p. LPS per g. In another model of lung inflammation, RelA nuclear translocation was reduced in p47phox−/− mice compared to WT mice following treatment with aerosolized LPS. In contrast to NF-κB activation in p47phox−/− mice, LPS-induced production of macrophage inflammatory protein 2 in the lungs and neutrophilic lung inflammation were not diminished in these mice compared to WT mice. We conclude that LPS-induced NF-κB activation is deficient in the lungs of p47phox−/− mice compared to WT mice, but this abnormality does not result in overt alteration in the acute inflammatory response.

Testicular injection of busulfan for recipient preparation in transplantation of spermatogonial stem cells in mice

2016 ◽  
Vol 28 (12) ◽  
pp. 1916 ◽  
Author(s):  
Yusheng Qin ◽  
Ling Liu ◽  
Yanan He ◽  
Wenzhi Ma ◽  
Huabin Zhu ◽  
...  
Keyword(s):  
Germ Cells ◽  
Fluorescent Protein ◽  
Positive Control ◽  
Negative Control ◽  
Seminiferous Tubules ◽  
Red Fluorescent Protein ◽  
Efferent Duct ◽  
Testicular Injection ◽  
Transplantation Recipient

Intraperitoneal busulfan injections are used to prepare recipients for spermatogonial stem cell (SSC) transplantation but they are associated with haematopoietic toxicity. Testicular injections of busulfan have been proposed to overcome this limitation. To date, testicular injections have not been studied in the mouse model. Therefore, in the present study we used ICR mice as recipients for SSC transplantation and prepared these mice by testicular injection of busulfan on both sides (2, 3, 4 or 6 mg kg–1 per side). Following this, donor germ cells expressing red fluorescent protein (RFP) from transgenic C57BL/6J male mice were transplanted into recipients via the efferent duct on Days 16–17 after busulfan treatment. Positive control mice were prepared by intraperitoneal injection of 40 mg kg–1 busulfan and negative control mice were treated with bilateral testicular injection of 50% dimethyl sulfoxide. On Day 49 after transplantation, recipient mice that were RFP-positive by in vivo imaging were mated with ICR female mice. Donor-derived germ cell colonies with red fluorescence were observed on Day 60 after transplantation, and donor-derived offspring were obtained. The results demonstrated that endogenous germ cells were successfully eliminated in the seminiferous tubules via testicular busulfan administration, and that exogenous SSCs successfully undergo spermatogenesis in the testes of recipient mice prepared by testicular injections of busulfan. In addition to its effects on recipient preparation, this method was safe in rodents and could possibly be adapted for use in other species.

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