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 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 Vaccine-Induced Protection against Helicobacter pylori in Mice Lacking Both Antibodies and Interleukin-4

2003 ◽  
Vol 71 (6) ◽  
pp. 3628-3633 ◽  
Author(s):  
Christine A. Garhart ◽  
John G. Nedrud ◽  
Frederick P. Heinzel ◽  
Norma E. Sigmund ◽  
Steven J. Czinn
Keyword(s):  
Helicobacter Pylori ◽  
B Cell ◽  
Knockout Mice ◽  
Cellular Responses ◽  
Interleukin 4 ◽  
Wild Type ◽  
Double Knockout ◽  
Th2 Response ◽  
H Pylori ◽  
Deficient Mice

ABSTRACT To test the hypothesis that a Th2 response to Helicobacter pylori is necessary for protection and to address the possibility that humoral and Th2 cellular responses may compensate for each other, we generated mice deficient in both interleukin-4 (IL-4) and antibodies. The immunized double-knockout mice were protected from H. pylori challenge, as were the parental strains and wild-type C57BL/6 mice. Neutralization of IL-4 in B-cell-deficient mice did not prevent protection. Immunized IL-5-deficient mice were also protected. Thus, IL-4 and IL-5 are not essential for protection.

scholarly journals Dual Ablation of Grb10 and Grb14 in Mice Reveals Their Combined Role in Regulation of Insulin Signaling and Glucose Homeostasis

2009 ◽  
Vol 30 (5) ◽  
pp. 540-540
Author(s):  
Lowenna J. Holt ◽  
Ruth J. Lyons ◽  
Ashleigh S. Ryan ◽  
Susan M. Beale ◽  
Andrew Ward ◽  
...  
Keyword(s):  
Body Composition ◽  
Glucose Homeostasis ◽  
Insulin Signaling ◽  
Knockout Mice ◽  
Gene Knockout ◽  
Physiological Role ◽  
Whole Body ◽  
Regulatory Function ◽  
Double Knockout ◽  
Deficient Mice

ABSTRACT Growth factor receptor bound (Grb)10 and Grb14 are closely related adaptor proteins that bind directly to the insulin receptor (IR) and regulate insulin-induced IR tyrosine phosphorylation and signaling to IRS-1 and Akt. Grb10- and Grb14-deficient mice both exhibit improved whole-body glucose homeostasis as a consequence of enhanced insulin signaling and, in the case of the former, altered body composition. However, the combined physiological role of these adaptors has remained undefined. In this study we utilize compound gene knockout mice to demonstrate that although deficiency in one adaptor can enhance insulin-induced IRS-1 phosphorylation and Akt activation, insulin signaling is not increased further upon dual ablation of Grb10 and Grb14. Context-dependent limiting mechanisms appear to include IR hypophosphorylation and decreased IRS-1 expression. In addition, the compound knockouts exhibit an increase in lean mass comparable to Grb10-deficient mice, indicating that this reflects a regulatory function specific to Grb10. However, despite the absence of additive effects on insulin signaling and body composition, the double-knockout mice are protected from the impaired glucose tolerance that results from high-fat feeding, whereas protection is not observed with animals deficient for individual adaptors. These results indicate that, in addition to their described effects on IRS-1/Akt, Grb10 and Grb14 may regulate whole-body glucose homeostasis by additional mechanisms and highlight these adaptors as potential therapeutic targets for amelioration of the insulin resistance associated with type 2 diabetes.

Nanomechanics of Knockout Mouse Bones

2006 ◽  
Vol 975 ◽  
Author(s):  
N Beril Kavukcuoglu ◽  
Adrian B. Mann
Keyword(s):  
Mechanical Properties ◽  
Knockout Mice ◽  
Bone Mineralization ◽  
Bone Matrix ◽  
Wild Type ◽  
Knock Out ◽  
Bone Matrix Proteins ◽  
Fibrillin 1 ◽  
Deficient Mice ◽  
Radial Axis

ABSTRACTOsteocalcin (OC) and osteopontin (OPN) are among the most abundant non-collagenous bone matrix proteins. Both have drawn interest from investigators studying their function in osteoporosis and it is known that mutations of these proteins can also have dramatic effects on the properties of bone. Other proteins including fibrillin 1 and 2 (FBN2) have been less widely studied, but can be mutated in some individuals resulting in connective tissue disorders. It has been reported that abnormal fibrillin may play a role in decreased bone mass. In this study bones from osteopontin (OPN), osteocalcin (OC) and fibrillin-2 (FBN2) knockout mice have been investigated. The study has identified how these proteins affect the bone's nanomechanical properties (hardness and elastic modulus). Nanoindentation tests were performed on the radial axis of cortical femora bones from the knockout mice and their wildtype controls. The results showed that young (age< 12 weeks) OPN knock-out bones have significantly lower mechanical properties than wild-type bones indicate a crucial role for OPN in early bone mineralization. After 12 weeks of age, the OPN knockout and wild-type control bones did not show any statistical difference. In OC deficient mice the mechanical properties were found to increase in the cortical mid-shaft of femora from 1 year old mice, suggesting an increase in bone mineralization, but 3 month old FBN2 deficient mice bones showed a decrease in mechanical properties across the cortical radial axis of the mid- femora.

scholarly journals Rhythmic changes in colonic motility are regulated by period genes

2010 ◽  
Vol 298 (2) ◽  
pp. G143-G150 ◽  
Author(s):  
Willemijntje A. Hoogerwerf ◽  
Vahakn B. Shahinian ◽  
Germaine Cornélissen ◽  
Franz Halberg ◽  
Jonathon Bostwick ◽  
...  
Keyword(s):  
Knockout Mice ◽  
Clock Genes ◽  
Ex Vivo ◽  
Colonic Motility ◽  
Biological Clock ◽  
Circular Muscle ◽  
Organ Bath ◽  
Wild Type ◽  
Double Knockout ◽  
Muscle Contractility

Human bowel movements usually occur during the day and seldom during the night, suggesting a role for a biological clock in the regulation of colonic motility. Research has unveiled molecular and physiological mechanisms for biological clock function in the brain; less is known about peripheral rhythmicity. This study aimed to determine whether clock genes such as period 1 ( per1) and period2 ( per2) modulate rhythmic changes in colonic motility. Organ bath studies, intracolonic pressure measurements, and stool studies were used to examine measures of colonic motility in wild-type and per1per2 double-knockout mice. To further examine the mechanism underlying rhythmic changes in circular muscle contractility, additional studies were completed in neuronal nitric oxide synthase (nNOS) knockout mice. Intracolonic pressure changes and stool output in vivo, and colonic circular muscle contractility ex vivo, are rhythmic with greatest activity at the start of night in nocturnal wild-type mice. In contrast, rhythmicity in these measures was absent in per1per2 double-knockout mice. Rhythmicity was also abolished in colonic circular muscle contractility of wild-type mice in the presence of Nω-nitro-l-arginine methyl ester and in nNOS knockout mice. These findings suggest that rhythms in colonic motility are regulated by both clock genes and a nNOS-mediated inhibitory process and suggest a connection between these two mechanisms.

A1 adenosine receptor knockout mice exhibit increased mortality, renal dysfunction, and hepatic injury in murine septic peritonitis

2005 ◽  
Vol 289 (2) ◽  
pp. F369-F376 ◽  
Author(s):  
George Gallos ◽  
Thomas D. Ruyle ◽  
Charles W. Emala ◽  
H. Thomas Lee
Keyword(s):  
Organ Dysfunction ◽  
Knockout Mice ◽  
Adenosine Receptor ◽  
Ischemia Reperfusion ◽  
Cecal Ligation ◽  
Intercellular Adhesion Molecule ◽  
Myeloperoxidase Activity ◽  
Wild Type ◽  
Wild Type Littermate ◽  
Septic Peritonitis

Sepsis is a leading cause of multiorgan dysfunction and death in hospitalized patients. Dysregulated inflammatory processes and apoptosis contribute to the pathogenesis of sepsis-induced organ dysfunction and death. A1 adenosine receptor (A1AR) activation reduces inflammation and apoptosis after ischemia-reperfusion injury. Therefore, we questioned whether A1AR-mediated reduction of inflammation and apoptosis could improve mortality and organ dysfunction in a murine model of sepsis. A1AR knockout mice (A1 knockout) and their wild-type (A1 wild-type) littermate controls were subjected to cecal ligation and double puncture (CLP) with a 20-gauge needle. A1 knockout mice or A1 wild-type mice treated with 1,3-dipropyl-8-cyclopentylxanthine (a selective A1AR antagonist) had a significantly higher mortality rate compared with A1 wild-type mice following CLP. Mice lacking endogenous A1ARs demonstrated significant elevations in plasma creatinine, alanine aminotransferase, aspartate aminotransferase, keratinocyte-derived chemokine, and tumor necrosis factor-α 24 h after induction of sepsis compared with wild-type mice. The renal corticomedullary junction from A1 knockout mice also exhibited increased myeloperoxidase activity, intercellular adhesion molecule-1 protein, and mRNA encoding proinflammatory cytokines compared with renal samples from A1 wild-type littermate controls. No difference in renal tubular apoptosis was detected between A1 knockout and A1 wild-type mice. We conclude that endogenous A1AR activation confers a protective effect in mice from septic peritonitis primarily by attenuating the hyperacute inflammatory response in sepsis.

The Cbl E3 Ubiquitin Ligase Negatively Regulates Erythropoiesis through Expression of the Foxo3a Transcription Factor.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1374-1374
Author(s):  
Terri D Richmond ◽  
Monica L Bailey ◽  
Wallace Y Langdon ◽  
Dwayne Barber
Keyword(s):  
Signal Transduction ◽  
Cell Signaling ◽  
Ubiquitin Ligase ◽  
Knockout Mice ◽  
E3 Ubiquitin Ligase ◽  
Parp Cleavage ◽  
Signal Transduction Pathways ◽  
Wild Type ◽  
Erythroid Progenitors ◽  
Deficient Mice

Abstract Erythropoietin (EPO) is the primary cytokine regulator of red blood cell (RBC) progenitor growth, survival and differentiation. EPO stimulation is regulated by EPO binding to its cognate ligand, the EPO receptor (EPO-R), and activating the primary associated tyrosine kinase, JAK2. The critical importance of EPO, EPO-R and JAK2 to erythropoiesis is demonstrated by the fatal embryonic anemia that develops upon EPO, EPO-R or JAK2 deletion. Intracellular signal transduction pathways regulating growth, survival and differentiation downstream of the EPO-R and JAK2 are well documented. However, relatively little is known about down-regulation of EPO-R signal transduction pathways at this time. Our laboratory has previously demonstrated that EPO stimulation leads to Cbltyrosine phosphorylation and subsequent recruitment of Crk-C3G, leading to Rap1activation. In addition, Cbl serves as an adaptor protein linking to PI 3 kinase and Rasand targets receptor tyrosine kinases for ubiquitination and proteasomal degradation. Cbl knockout mice have been generated and have defects in stem and T cell signaling pathways. Elevated platelet numbers and splenomegaly was observed, suggesting that Cbl −/− mice may have defects in megakaryocyte/erythroid progenitors or more committed cells in each lineage. The objective of this studyis to determine whether Cbl affects erythropoiesis and EPO-dependent signaling. Resting Cbl −/− mice (in the C57Bl/6 background) have increased numbers of Burst Forming Unit-Erythroid and Colony Forming Unit-Erythroid (CFU-E) cells. Furthermore, there is a 3-fold elevation of splenic CFU-E numbers. Erythroid differentiation was monitored via expression of the Transferrin Receptor (CD71) and Ter119. Cbl-deficient mice have delayed differentiation in the bone marrow with diminished CD71-Ter119+ cells. Increased apoptosis is observed in Ter119+ erythroid cells isolated from Cbl −/− mice as determined by Annexin V staining and confirmed by increased PARP cleavage. Interestingly, reactive oxygen species in wild type and Cbl-deficient mice remain unchanged. Despite normal resting hematologic parameters, serum EPO concentrations are elevated in Cbl knockout mice. Serum VEGF levels are comparable between wild type and Cbl −/− mice, suggesting that the EPO effect is specific to the erythroid lineage and not an effect of hypoxia. Notable differences in wild type and Cbl −/− mice were observed when stress erythropoiesiswas induced by phenylhydrazine-mediated anemia. Cbl-deficient mice respond with enhanced hematocrit recovery and increased reticulocyte production. EPO-dependent Aktphosphorylation is hypersensitive in Cbl −/− splenic erythroblasts. Interestingly, expression ofFoxo3a was stabilized in Cbl −/− splenic erythroblasts, suggesting that Cbl degrades Foxo3a in a direct or indirect manner. Given the importance of Foxo3a in regulating erythropoiesis, we are currently determining whether Cbl targets Foxo3a for ubiquitin-mediated degradation. These data demonstrate the remarkable homeostatic ability of the mouse to retain normal RBC concentrations in the peripheral blood despite elevated erythroid progenitors and cell signaling. Importantly, these studies are the first to phenotypically explore the effects of genetic ablation of an EPO-responsive E3 ubiquitin ligase in erythropoiesis.

Defective water and glycerol transport in the proximal tubules of AQP7 knockout mice

2005 ◽  
Vol 289 (6) ◽  
pp. F1195-F1200 ◽  
Author(s):  
Eisei Sohara ◽  
Tatemitsu Rai ◽  
Jun-ichi Miyazaki ◽  
A. S. Verkman ◽  
Sei Sasaki ◽  
...  
Keyword(s):  
Knockout Mice ◽  
Water Channel ◽  
Physiological Role ◽  
Wild Type ◽  
Double Knockout ◽  
Straight Tubule ◽  
Glycerol Transport ◽  
Straight Tubules ◽  
Proximal Straight Tubule ◽  
Urinary Concentrating Ability

The aquaporin-7 (AQP7) water channel is known as a member of the aquaglyceroporins, which facilitate the transport of glycerol as well as water. Although AQP7 is abundantly expressed on the apical membrane of the proximal straight tubules in the kidney, the physiological role of AQP7 is still unknown. To investigate this, we generated AQP7 knockout mice. The water permeability of the proximal tubule brush-border membrane measured by the stopped-flow method was slightly but significantly reduced in the AQP7 knockout mice compared with that of wild-type mice (AQP7, 18.0 ± 0.4 × 10−3 cm/s vs. wild-type, 20.0 ± 0.3 × 10−3 cm/s). Although AQP7 solo-knockout mice did not exhibit a urinary concentrating defect, AQP1/AQP7 double-knockout mice had a reduction in urinary concentrating ability compared with AQP1 solo-knockout mice, suggesting that the amount of water reabsorbed through AQP7 in the proximal straight tubules is physiologically substantial. On the other hand, AQP7 knockout mice showed marked glyceroluria (AQP7, 1.7 ± 0.34 mg/ml vs. wild-type, 0.005 ± 0.002 mg/ml). This identified a novel glycerol reabsorption pathway in the proximal straight tubules. In two mouse models of proximal straight tubule injury, the cisplatin-induced acute renal failure (ARF) model and the ischemic ARF model, an increase in urine glycerol was observed (pretreatment, 0.007 ± 0.005 mg/ml; cisplatin, 0.063 ± 0.043 mg/ml; ischemia, 0.076 ± 0.02 mg/ml), suggesting that urine glycerol could be used as a new biomarker for detecting proximal straight tubule injury.

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