Type-A spermatogonia first appear at between 3-7 d postnatally in mice and are the only immortalized diploid cells that reproduce in adulthood in these animals. In our current study, we explored the feasibility of pro...Type-A spermatogonia first appear at between 3-7 d postnatally in mice and are the only immortalized diploid cells that reproduce in adulthood in these animals. In our current study, we explored the feasibility of producing stable transgenic mice using these cells. Enhanced pEGFP-N1 plasmids were suspended in ExGen500 transfection reagent and injected at different angles into the testes of 7-d-old male ICR mice. The resulting type-A spermatogonia-mediated gene transfer （TASMGT） mice were then mated with normal females at different stages of sexual maturity （6, 12, and 24 wk）. The integration and expression of the introduced EGFP gene was evaluated in the F1 transgenic offspring by PCR and Southern blotting analysis. The foreign gene integration rates for a low-dose group （15 μL gene suspension injected into each testis） and a high-dose group （30 μL suspensions injected） at the three stages of female sexual maturity tested were 11.76% （2/17）, 14.29% （3/21）, and 11.11% （2/18）, and 5% （1/20）, 5.56% （1/18）, and 0 （0/17）, respectively. The average integration rates for these two dose groups were 12.5% （7/56） and 3.64% （2/55）, respectively, which was a significant difference （P0.05）. Semi-quantitative RT-PCR analysis further showed that the introduced GFP gene was expressed in 3/9 integration mice. In addition, GFP expression was observed in the sperm cells from the TASMGT mice, and also in the embryos and F2 pups from the F1 generation transgenic mice. Hence, although the foreign gene integration rate for TASMGT is not high and the transgenic offspring show as yet unexplained defects, our results indicate that this method is a potentially feasible and reproducible new approach to creating transgenic mice.展开更多
Col la I （one of the subunit of collagen type I） is a collagen, which belongs to a family of extracellular matrix （ECM） proteins that play an important role in cellular proliferation and differentiation. However, ...Col la I （one of the subunit of collagen type I） is a collagen, which belongs to a family of extracellular matrix （ECM） proteins that play an important role in cellular proliferation and differentiation. However, the role of Col lal in spermatogenesis, especially in the control of proliferation and differentiation of spermatogonial stem cells （SSCs）, remains unknown. In this study, we explored effects of downregulation of Collal on differentiation and proliferation of mouse spermatogonia. Loss-of-function study revealed that Oct4 and Plzf, markers of SSC self-renewal, were significantly decreased, whereas the expression of c-kit and haprin, hallmarks of SSC differentiation, was enhanced after Col la I knockdown. Cell cycle analyses indicated that two-thirds of spermatogonia were arrested in S phase after Collal knockdown. In vivo experiments, DNA injection and electroporation of the testes showed that spermatogonia self-renewal ability was impaired remarkably with the loss-of-function of Collal. Our data suggest that silencing of Collal can suppress spermatogonia self-renewal and promote spermatogonia differentiation.展开更多
Aim： To investigate whether estrogen stimulates the proliferation of spermatogonia or induces spermatogenesis in cryptorchid mice. Methods： Mice were surgically rendered cryptorchid, then treated with different dose...Aim： To investigate whether estrogen stimulates the proliferation of spermatogonia or induces spermatogenesis in cryptorchid mice. Methods： Mice were surgically rendered cryptorchid, then treated with different doses of 17β- estradiol （E2） s.c. once a day. Mice were killed at sexual maturity （45 days of age）, and histological analysis and inununofluorescence were performed. Serum follicle stimulating hormone （FSH）, estradiol, testosterone and luteinizing hormone （LH） were measured. Results： Low doses of E2 had no notable effect on spermatogonia, but at higher doses, E2 stimulated the proliferation of spermatogonia. Conclusion： E2 has a dose-related mitogenic effect on spermatogonia.展开更多
Background:Spermatogenesis is a cellular differentiation process that includes three major events:mitosis of spermatogonia,meiosis of spermatocytes and spermiogenesis.Steady-state spermatogenesis relies on functions o...Background:Spermatogenesis is a cellular differentiation process that includes three major events:mitosis of spermatogonia,meiosis of spermatocytes and spermiogenesis.Steady-state spermatogenesis relies on functions of spermatogonial stem cells(SSCs).Establishing and maintaining a foundational SSC pool is essential for continued spermatogenesis in mammals.Currently,our knowledge about SSC and spermatogenesis is severely limited in domestic animals.Results:In the present study,we examined transcriptomes of testes from domestic yaks at four different stages(3,5,8 and 24 months of age)and attempted to identify genes that are associated with key developmental events of spermatogenesis.Histological analyses showed that the most advanced germ cells within seminiferous tubules of testes from 3,5,8 and 24 months old yaks were gonocytes,spermatogonia,spermatocytes and elongated spermatids,respectively.RNA-sequencing(RNA-seq)analyses revealed that 11904,4381 and 2459 genes were differentially expressed during the gonocyte to spermatogonia transition,the mitosis to meiosis transition and the meiosis to post-meiosis transition.Further analyses identified a list of candidate genes than may regulate these important cellular processes.CXCR4,a previously identified SSC niche factor in mouse,was one of the up-regulated genes in the 5 months old yak testis.Results of immunohistochemical staining confirmed that CXCR4 was exclusively expressed in gonocytes and a subpopulation of spermatogonia in the yak testis.Conclusions:Together,these findings demonstrated histological changes of postnatal testis development in the domestic yak.During development of spermatogonial lineage,meiotic and haploid germ cells are supported by dynamic transcriptional regulation of gene expression.Our transcriptomic analyses provided a list of candidate genes that potentially play crucial roles in directing the establishment of SSC and spermatogenesis in yak.展开更多
BACKGROUND:Regulatory peptide receptors have attracted the interest of oncologists as a new promising approach for cancer pathology,imaging and therapy.Although cholecystokinin (CCK) is a potent modulator of gallbladd...BACKGROUND:Regulatory peptide receptors have attracted the interest of oncologists as a new promising approach for cancer pathology,imaging and therapy.Although cholecystokinin (CCK) is a potent modulator of gallbladder contractility and plays a potential role in pancreatic carcinogenesis through CCK type-A receptor (CCKAR),its role in gallbladder cancer (GBC) is still unknown and immunohistochemical detection of CCKAR in the gallbladder has not yet been reported.This novel case-control study aimed to investigate the expression profile of CCKAR in GBC and gallstone disease (GSD).METHODS:This study included 162 samples of gallbladder:94 from GBC and 68 from GSD.Expression of CCKAR was analyzed by immunohistochemistry and immunoblotting.The results were statistically correlated with disease history including age,sex,presence of gallstone,stage and differentiation.RESULTS:CCKAR was positive in 30/68 (44.1%) of GSD and 72/94 (76.6%) of GBC samples.Fifty-one of the 72 (70.8%) CCKAR-positive GBC samples showed over-expression.Interestingly,consistent results also appeared in the immunoblotting study.CONCLUSIONS:CCKAR expression was significantly increased in GBC compared to GSD.Moreover,CCKAR expression was associated with the degree of tumor differentiation,i.e.,less expression in poorly-differentiated tumors.Thus,it has future prognostic and therapeutic implications in the management of GBC.展开更多
Background:Spermatogenesis is an intricate developmental process during which undifferentiated spermatogonia,containing spermatogonial stem cells(SSCs),undergo self-renewal and differentiation to generate eventually m...Background:Spermatogenesis is an intricate developmental process during which undifferentiated spermatogonia,containing spermatogonial stem cells(SSCs),undergo self-renewal and differentiation to generate eventually mature spermatozoa.Spermatogenesis occurs in seminiferous tubules within the testis,and the seminiferous tubules harbor Sertoli and germ cells.Sertoli cells are an essential somatic cell type within the microenvironment that support and steer male germ cell development,whereas spermatogonia are the primitive male germ cells at the onset of spermatogenesis.While the developmental progression of Sertoli cells and spermatogonia has been well established in mice,much less is known in other mammalian species including pigs.Results:To acquire knowledge of Sertoli cell and spermatogonial development in pigs,here we collected as many as nine ages of Duroc porcine testes from the neonate to sexual maturity,i.e.,testes from 7-,30-,50-,70-,90-,110-,130-,150-and 210-day-old boars,and performed histological and immunohistochemical analyses on testis sections.We first examined the development of spermatogenic cells and seminiferous tubules in porcine testes.Then,by immunofluorescence staining for marker proteins(AMH,SOX9,DBA,UCHL1,VASA,KIT,Ki67 and/or PCNA),we delved into the proliferative activity and development of Sertoli cells and of spermatogonial subtypes(pro-,undifferentiated and differentiating spermatogonia).Besides,by immunostaining forβ-catenin and ZO-1,we studied the establishment of the blood-testis barrier in porcine testes.Conclusions:In this longitudinal study,we have systematically investigated the elaborate Sertoli cell and spermatogonial developmental patterns in pigs from the neonate to sexual maturity that have so far remained largely unknown.The findings not only extend the knowledge about spermatogenesis and testicular development in pigs,but also lay the theoretical groundwork for porcine breeding and rearing.展开更多
Numerical simulations based on a high-resolution three-dimensional MIT general circulation model(MITgcm)using realistic topography and tidal forcing are conducted to investigate the generation and propagation of the s...Numerical simulations based on a high-resolution three-dimensional MIT general circulation model(MITgcm)using realistic topography and tidal forcing are conducted to investigate the generation and propagation of the so-called type-a waves(large-amplitude rank-ordered wave packets)and type-b waves(isolated wave packets)in the northern South China Sea.At first,we summarized and analyzed the generation and propagation characteristics of these waves.Then,energy budget at the Luzon Strait is calculated.Energy generation has three local maxima every day,of which the largest one corresponds to the emergence of the type-a wave.Energy flux at the west boundary of the Luzon Strait shows two local maxima each day.The larger one is consistent with the generation of the type-a wave and the smaller one is in correspondence with the generation of the type-b wave.Sensitivity experiments are designed to explore the role of the east and west ridge of the Luzon Strait on the generation and propagation of the type-a and type-b waves.It is found that the east ridge is indispensable on the generation of the type-a wave while the west ridge has little contribution.The west ridge diminishes the type-a waves'amplitude but hardly changes their propagation speed.The type-b waves also come from perturbation signals which originate from the east ridge and are enhanced in amplitude and reduced in propagation speed by the west ridge.展开更多
Objective The expression patterns of ribosomal large subunit protein 23 a(RPL23 a)in mouse testes and GC-1 cells were analyzed to investigate the potential relationship between RPL23 a expression and spermatogonia apo...Objective The expression patterns of ribosomal large subunit protein 23 a(RPL23 a)in mouse testes and GC-1 cells were analyzed to investigate the potential relationship between RPL23 a expression and spermatogonia apoptosis upon exposure to X-ray.Methods Male mice and GC-1 cells were irradiated with X-ray,terminal dUTP nick end-labelling(TUNEL)was performed to detect apoptotic spermatogonia in vivo.Apoptotic rate and cell cycle phase of GC-1 cells were analyzed with flow cytometry.Protein interactions were detected by Immunoprecipitation and protein localization as studied by immunofluorescence.Immunoblotting and real-time PCR were applied to analyze to protein and gene expression.Results Ionizing radiation(IR)increased spermatogonia apoptosis,the expression of RPL11,MDM2 and p53,and decreased RPL23 a expression in mice spermatogonia in vivo and in vitro.RPL23 a knockdown weakened the interaction between RPL23 a and RPL11,leading to p53 accumulation.Moreover,knockdown and IR decreased RPL23 a that induces spermatogonia apoptosis via RPL23 a-RPL11-MDM2-p53 pathway in GC-1 cells.Conclusion These results suggested that IR reduced RPL23 a expression,leading to weakened the RPL23 a-RPL11 interactions,which may have activated p53,resulting in spermatogonia apoptosis.These results provide insights into environmental and clinical risks of radiotherapy following exposure to IR in male fertility.The graphical abstract was available in the web of www.besjournal.com.展开更多
In vitro production of functional gametes can revolutionize reproduction by reducing generation intervals and accelerating genetic breeding in aquaculture,especially in fish with relatively long generations.Neverthele...In vitro production of functional gametes can revolutionize reproduction by reducing generation intervals and accelerating genetic breeding in aquaculture,especially in fish with relatively long generations.Nevertheless,functional sperm production from in vitro-cultured spermatogonia remains a challenge in most aquaculture fish.In this study,we isolated and characterized premeiotic spermatogonia from marine four-eyed sleepers(Bostrychus sinensis),which are prone to ovotesticular or sterile testicular development,and induced the differentiation of the spermatogonia into flagellated sperm in a three-dimensional(3D)culture system.Artificial insemination indicated that the in vitro-derived sperm were capable of fertilizing mature oocytes to develop into normal larvae.Furthermore,melatonin significantly promoted spermatogonia proliferation and differentiation through the ERK1/2 signaling pathway,and thus increased the efficiency in functional sperm production.The 3D culture system and resulting functional sperm hold great promise for improving the genetic breeding of aquaculture fish.展开更多
The present study was aimed at finding an effective method to isolate and purify the subtype of type A spermatogonial stem cells （SSCs） in juvenile rats. Testes from 9-days-old rats were used to isolate germ cells b...The present study was aimed at finding an effective method to isolate and purify the subtype of type A spermatogonial stem cells （SSCs） in juvenile rats. Testes from 9-days-old rats were used to isolate germ cells by using two-step enzymatic digestion. The expression of c-kit in the testes of the rats was immunohistochemically detected. After isolation, cell suspension was enriched further by discontinuous density gradient centrifugation. Then type A1-A4 spermatogonia was isolated from the purified spermatogonia with c-kit as the marker by using fluorescence-activated cell sorting （FACS）. Electron microscopy was used to observe their ultrastructure. Finally, highly purified and viable subtype of SSCs was obtained. Cells separation with discontinuous density gradient centrifugation significantly increased the concentration of c-kit positive cells [（18.65±1.69）% after the centrifugation versus （3.16±0.84）% before the centrifugation, P〈0.01]. Furthermore, the recovery and viability were also high [（65.9±1.24）% and （85.6±1.14）%]. It is concluded that FACS with c-kit as the marker in combination with discontinuous density gradient centrifugation can well enrich type A1-A4 spermatogonia from the testes of 9-days-old rats.展开更多
Purpose: The purpose of the study was to evaluate the efficacy of botulinum toxin type A (BTX-A) injection in patients with acquired VI nerve palsy. Methods: Twenty-eight patients (16 F and 12 M), mean age 36.4 +/-- 1...Purpose: The purpose of the study was to evaluate the efficacy of botulinum toxin type A (BTX-A) injection in patients with acquired VI nerve palsy. Methods: Twenty-eight patients (16 F and 12 M), mean age 36.4 +/-- 17.8 DS;range 10 - 69 years with acquired VI nerve palsy, have been treated with BTX-A injection into the ipsilateral medial rectus, at least 6 months after onset. At 6 months follow up, a paresis and a paralysis, were respectively diagnosed if the affected eye could be actively abducted or not, across the mid-line;a simple horizontal rectus muscle procedure was performed for the paresis whilst for the paralysis, both a horizontal and vertical muscle transportation procedure were required. Results: A gradual physiological recovery of the lateral rectus muscle was observed in 9 (32%) patients (GROUP I) and surgical treatment was therefore unnecessary;the remaining 19 cases (68%), of which 5 paresis (GROUP II) and 14 paralysis (GROUP III) underwent surgery and within 1 year were all restored to orthotropia. Conclusion: Botulinum toxin type-A (BTX-A) injection is an invaluable tool in the differential diagnosis between paresis versus paralysis of the VIth nerve, allowing the correct choice of surgical procedure.展开更多
A previous study has proposed a mathematical model of type-A medial vestibular nucleus neurons (mVNn). This model is described by a system of nonlinear ordinary differential equations, which is based on the Hodgkin-Hu...A previous study has proposed a mathematical model of type-A medial vestibular nucleus neurons (mVNn). This model is described by a system of nonlinear ordinary differential equations, which is based on the Hodgkin-Huxley formalism. The type-A mVNn model contains several ionic conductances, such as the sodium conductance, calcium conductance, delayed-rectifier potassium conductance, transient potassium conductance, and calcium-dependent potassium conductance. The previous study revealed that spontaneous repetitive spiking in the type-A mVNn model can be suppressed by hyperpolarizing stimulation. However, how this suppression is affected by the ionic conductances has not been clarified in the previous study. The present study performed numerical simulation analysis of the type-A mVNn model to clarify how variations in the different ionic conductance values affect the suppression of repetitive spiking. The present study revealed that the threshold for the transition from a repetitive spiking state to a quiescent state is differentially sensitive to variations in the ionic conductances among the different types of ionic conductance.展开更多
基金supported by the National Transgenic Breeding Project (2008ZX08010-004)the National Natural Science Foundation of China (30830080)+1 种基金the National 973 Program of China (G2006CB102105,2009CB941604)the National 863 Program of China (20060110Z1039, 2008AA10Z143)
文摘Type-A spermatogonia first appear at between 3-7 d postnatally in mice and are the only immortalized diploid cells that reproduce in adulthood in these animals. In our current study, we explored the feasibility of producing stable transgenic mice using these cells. Enhanced pEGFP-N1 plasmids were suspended in ExGen500 transfection reagent and injected at different angles into the testes of 7-d-old male ICR mice. The resulting type-A spermatogonia-mediated gene transfer （TASMGT） mice were then mated with normal females at different stages of sexual maturity （6, 12, and 24 wk）. The integration and expression of the introduced EGFP gene was evaluated in the F1 transgenic offspring by PCR and Southern blotting analysis. The foreign gene integration rates for a low-dose group （15 μL gene suspension injected into each testis） and a high-dose group （30 μL suspensions injected） at the three stages of female sexual maturity tested were 11.76% （2/17）, 14.29% （3/21）, and 11.11% （2/18）, and 5% （1/20）, 5.56% （1/18）, and 0 （0/17）, respectively. The average integration rates for these two dose groups were 12.5% （7/56） and 3.64% （2/55）, respectively, which was a significant difference （P0.05）. Semi-quantitative RT-PCR analysis further showed that the introduced GFP gene was expressed in 3/9 integration mice. In addition, GFP expression was observed in the sperm cells from the TASMGT mice, and also in the embryos and F2 pups from the F1 generation transgenic mice. Hence, although the foreign gene integration rate for TASMGT is not high and the transgenic offspring show as yet unexplained defects, our results indicate that this method is a potentially feasible and reproducible new approach to creating transgenic mice.
文摘Col la I （one of the subunit of collagen type I） is a collagen, which belongs to a family of extracellular matrix （ECM） proteins that play an important role in cellular proliferation and differentiation. However, the role of Col lal in spermatogenesis, especially in the control of proliferation and differentiation of spermatogonial stem cells （SSCs）, remains unknown. In this study, we explored effects of downregulation of Collal on differentiation and proliferation of mouse spermatogonia. Loss-of-function study revealed that Oct4 and Plzf, markers of SSC self-renewal, were significantly decreased, whereas the expression of c-kit and haprin, hallmarks of SSC differentiation, was enhanced after Col la I knockdown. Cell cycle analyses indicated that two-thirds of spermatogonia were arrested in S phase after Collal knockdown. In vivo experiments, DNA injection and electroporation of the testes showed that spermatogonia self-renewal ability was impaired remarkably with the loss-of-function of Collal. Our data suggest that silencing of Collal can suppress spermatogonia self-renewal and promote spermatogonia differentiation.
基金Acknowledgment This work was supported by the National Natural Science Foundation of China （No. 30200195）. We thank Dr Hai-Bin Wang for taking photographs and Dr Su-Hui Wu （Henan Normal University, China） for statistical analysis. We thank the faculty of Huanghuai University for supporting Dr En-Zhong Li.
文摘Aim： To investigate whether estrogen stimulates the proliferation of spermatogonia or induces spermatogenesis in cryptorchid mice. Methods： Mice were surgically rendered cryptorchid, then treated with different doses of 17β- estradiol （E2） s.c. once a day. Mice were killed at sexual maturity （45 days of age）, and histological analysis and inununofluorescence were performed. Serum follicle stimulating hormone （FSH）, estradiol, testosterone and luteinizing hormone （LH） were measured. Results： Low doses of E2 had no notable effect on spermatogonia, but at higher doses, E2 stimulated the proliferation of spermatogonia. Conclusion： E2 has a dose-related mitogenic effect on spermatogonia.
基金supported by National Key Research&Development Project grant(2016YFC0501805)Qinghai Department of Science and Technology grants(2017-NK-154 and 2016-ZJ-917Q)+2 种基金a STS grant from Chinese Academy of Sciences(KFJ-STS-QYZD-113)supported by the CAS“100 Talents” and Qinghai “1000 Talents” programsfunded by CAS “Light of West China Foundation”
文摘Background:Spermatogenesis is a cellular differentiation process that includes three major events:mitosis of spermatogonia,meiosis of spermatocytes and spermiogenesis.Steady-state spermatogenesis relies on functions of spermatogonial stem cells(SSCs).Establishing and maintaining a foundational SSC pool is essential for continued spermatogenesis in mammals.Currently,our knowledge about SSC and spermatogenesis is severely limited in domestic animals.Results:In the present study,we examined transcriptomes of testes from domestic yaks at four different stages(3,5,8 and 24 months of age)and attempted to identify genes that are associated with key developmental events of spermatogenesis.Histological analyses showed that the most advanced germ cells within seminiferous tubules of testes from 3,5,8 and 24 months old yaks were gonocytes,spermatogonia,spermatocytes and elongated spermatids,respectively.RNA-sequencing(RNA-seq)analyses revealed that 11904,4381 and 2459 genes were differentially expressed during the gonocyte to spermatogonia transition,the mitosis to meiosis transition and the meiosis to post-meiosis transition.Further analyses identified a list of candidate genes than may regulate these important cellular processes.CXCR4,a previously identified SSC niche factor in mouse,was one of the up-regulated genes in the 5 months old yak testis.Results of immunohistochemical staining confirmed that CXCR4 was exclusively expressed in gonocytes and a subpopulation of spermatogonia in the yak testis.Conclusions:Together,these findings demonstrated histological changes of postnatal testis development in the domestic yak.During development of spermatogonial lineage,meiotic and haploid germ cells are supported by dynamic transcriptional regulation of gene expression.Our transcriptomic analyses provided a list of candidate genes that potentially play crucial roles in directing the establishment of SSC and spermatogenesis in yak.
基金supported by a grant from the Indian Council of Medical Research(ICMR Project Ref No.3/2/2/187/2009/NCD-Ⅲ)
文摘BACKGROUND:Regulatory peptide receptors have attracted the interest of oncologists as a new promising approach for cancer pathology,imaging and therapy.Although cholecystokinin (CCK) is a potent modulator of gallbladder contractility and plays a potential role in pancreatic carcinogenesis through CCK type-A receptor (CCKAR),its role in gallbladder cancer (GBC) is still unknown and immunohistochemical detection of CCKAR in the gallbladder has not yet been reported.This novel case-control study aimed to investigate the expression profile of CCKAR in GBC and gallstone disease (GSD).METHODS:This study included 162 samples of gallbladder:94 from GBC and 68 from GSD.Expression of CCKAR was analyzed by immunohistochemistry and immunoblotting.The results were statistically correlated with disease history including age,sex,presence of gallstone,stage and differentiation.RESULTS:CCKAR was positive in 30/68 (44.1%) of GSD and 72/94 (76.6%) of GBC samples.Fifty-one of the 72 (70.8%) CCKAR-positive GBC samples showed over-expression.Interestingly,consistent results also appeared in the immunoblotting study.CONCLUSIONS:CCKAR expression was significantly increased in GBC compared to GSD.Moreover,CCKAR expression was associated with the degree of tumor differentiation,i.e.,less expression in poorly-differentiated tumors.Thus,it has future prognostic and therapeutic implications in the management of GBC.
基金The authors would like to acknowledge Dr.K.R.Mahadik,Principal,Poona College of Pharmacy,Bharati Vidyapeeth Deemed University,Pune,India for providing necessary facilities to carry out the study.Research support was provided by Indus Biotech Private Limited but played no role in collection,analysis and interpretation of data.
基金supported by the National Natural Science Foundation of China(Grant No.32002178 and 31772605)the Undergraduate Training Program for Innovation and Entrepreneurship(X202110712185).
文摘Background:Spermatogenesis is an intricate developmental process during which undifferentiated spermatogonia,containing spermatogonial stem cells(SSCs),undergo self-renewal and differentiation to generate eventually mature spermatozoa.Spermatogenesis occurs in seminiferous tubules within the testis,and the seminiferous tubules harbor Sertoli and germ cells.Sertoli cells are an essential somatic cell type within the microenvironment that support and steer male germ cell development,whereas spermatogonia are the primitive male germ cells at the onset of spermatogenesis.While the developmental progression of Sertoli cells and spermatogonia has been well established in mice,much less is known in other mammalian species including pigs.Results:To acquire knowledge of Sertoli cell and spermatogonial development in pigs,here we collected as many as nine ages of Duroc porcine testes from the neonate to sexual maturity,i.e.,testes from 7-,30-,50-,70-,90-,110-,130-,150-and 210-day-old boars,and performed histological and immunohistochemical analyses on testis sections.We first examined the development of spermatogenic cells and seminiferous tubules in porcine testes.Then,by immunofluorescence staining for marker proteins(AMH,SOX9,DBA,UCHL1,VASA,KIT,Ki67 and/or PCNA),we delved into the proliferative activity and development of Sertoli cells and of spermatogonial subtypes(pro-,undifferentiated and differentiating spermatogonia).Besides,by immunostaining forβ-catenin and ZO-1,we studied the establishment of the blood-testis barrier in porcine testes.Conclusions:In this longitudinal study,we have systematically investigated the elaborate Sertoli cell and spermatogonial developmental patterns in pigs from the neonate to sexual maturity that have so far remained largely unknown.The findings not only extend the knowledge about spermatogenesis and testicular development in pigs,but also lay the theoretical groundwork for porcine breeding and rearing.
基金The National Key Research and Development Plan of China under contract No.2016YFC1401300the National Natural Science Foundation of China under contract No.41276008the Taishan Scholars Program
文摘Numerical simulations based on a high-resolution three-dimensional MIT general circulation model(MITgcm)using realistic topography and tidal forcing are conducted to investigate the generation and propagation of the so-called type-a waves(large-amplitude rank-ordered wave packets)and type-b waves(isolated wave packets)in the northern South China Sea.At first,we summarized and analyzed the generation and propagation characteristics of these waves.Then,energy budget at the Luzon Strait is calculated.Energy generation has three local maxima every day,of which the largest one corresponds to the emergence of the type-a wave.Energy flux at the west boundary of the Luzon Strait shows two local maxima each day.The larger one is consistent with the generation of the type-a wave and the smaller one is in correspondence with the generation of the type-b wave.Sensitivity experiments are designed to explore the role of the east and west ridge of the Luzon Strait on the generation and propagation of the type-a and type-b waves.It is found that the east ridge is indispensable on the generation of the type-a wave while the west ridge has little contribution.The west ridge diminishes the type-a waves'amplitude but hardly changes their propagation speed.The type-b waves also come from perturbation signals which originate from the east ridge and are enhanced in amplitude and reduced in propagation speed by the west ridge.
基金the National Natural Science Foundation of China[31902339]the Foundation for Youth Doctor of Gansu Province[2021QB-026]+2 种基金the Youth Talent Program of“Fuxi”[Gaufx-03Y02]the Scientific Research Start-up Funds for Openly-Recruited Doctors of Gansu Agricultural University[2017RCZX-13]the Special Funds for Discipline Construction of Gansu Agricultural University[GAU-XKJS-2018-067]。
文摘Objective The expression patterns of ribosomal large subunit protein 23 a(RPL23 a)in mouse testes and GC-1 cells were analyzed to investigate the potential relationship between RPL23 a expression and spermatogonia apoptosis upon exposure to X-ray.Methods Male mice and GC-1 cells were irradiated with X-ray,terminal dUTP nick end-labelling(TUNEL)was performed to detect apoptotic spermatogonia in vivo.Apoptotic rate and cell cycle phase of GC-1 cells were analyzed with flow cytometry.Protein interactions were detected by Immunoprecipitation and protein localization as studied by immunofluorescence.Immunoblotting and real-time PCR were applied to analyze to protein and gene expression.Results Ionizing radiation(IR)increased spermatogonia apoptosis,the expression of RPL11,MDM2 and p53,and decreased RPL23 a expression in mice spermatogonia in vivo and in vitro.RPL23 a knockdown weakened the interaction between RPL23 a and RPL11,leading to p53 accumulation.Moreover,knockdown and IR decreased RPL23 a that induces spermatogonia apoptosis via RPL23 a-RPL11-MDM2-p53 pathway in GC-1 cells.Conclusion These results suggested that IR reduced RPL23 a expression,leading to weakened the RPL23 a-RPL11 interactions,which may have activated p53,resulting in spermatogonia apoptosis.These results provide insights into environmental and clinical risks of radiotherapy following exposure to IR in male fertility.The graphical abstract was available in the web of www.besjournal.com.
基金supported by the National Key R&D Program of China(2018YFD0901205)National Natural Science Foundation of China(31771587,31970535)Guangdong Basic and Applied Basic Research Foundation(2020A1515010358)。
文摘In vitro production of functional gametes can revolutionize reproduction by reducing generation intervals and accelerating genetic breeding in aquaculture,especially in fish with relatively long generations.Nevertheless,functional sperm production from in vitro-cultured spermatogonia remains a challenge in most aquaculture fish.In this study,we isolated and characterized premeiotic spermatogonia from marine four-eyed sleepers(Bostrychus sinensis),which are prone to ovotesticular or sterile testicular development,and induced the differentiation of the spermatogonia into flagellated sperm in a three-dimensional(3D)culture system.Artificial insemination indicated that the in vitro-derived sperm were capable of fertilizing mature oocytes to develop into normal larvae.Furthermore,melatonin significantly promoted spermatogonia proliferation and differentiation through the ERK1/2 signaling pathway,and thus increased the efficiency in functional sperm production.The 3D culture system and resulting functional sperm hold great promise for improving the genetic breeding of aquaculture fish.
基金the National Natural Sciences Foundation of China (No. 30371424)
文摘The present study was aimed at finding an effective method to isolate and purify the subtype of type A spermatogonial stem cells （SSCs） in juvenile rats. Testes from 9-days-old rats were used to isolate germ cells by using two-step enzymatic digestion. The expression of c-kit in the testes of the rats was immunohistochemically detected. After isolation, cell suspension was enriched further by discontinuous density gradient centrifugation. Then type A1-A4 spermatogonia was isolated from the purified spermatogonia with c-kit as the marker by using fluorescence-activated cell sorting （FACS）. Electron microscopy was used to observe their ultrastructure. Finally, highly purified and viable subtype of SSCs was obtained. Cells separation with discontinuous density gradient centrifugation significantly increased the concentration of c-kit positive cells [（18.65±1.69）% after the centrifugation versus （3.16±0.84）% before the centrifugation, P〈0.01]. Furthermore, the recovery and viability were also high [（65.9±1.24）% and （85.6±1.14）%]. It is concluded that FACS with c-kit as the marker in combination with discontinuous density gradient centrifugation can well enrich type A1-A4 spermatogonia from the testes of 9-days-old rats.
文摘Purpose: The purpose of the study was to evaluate the efficacy of botulinum toxin type A (BTX-A) injection in patients with acquired VI nerve palsy. Methods: Twenty-eight patients (16 F and 12 M), mean age 36.4 +/-- 17.8 DS;range 10 - 69 years with acquired VI nerve palsy, have been treated with BTX-A injection into the ipsilateral medial rectus, at least 6 months after onset. At 6 months follow up, a paresis and a paralysis, were respectively diagnosed if the affected eye could be actively abducted or not, across the mid-line;a simple horizontal rectus muscle procedure was performed for the paresis whilst for the paralysis, both a horizontal and vertical muscle transportation procedure were required. Results: A gradual physiological recovery of the lateral rectus muscle was observed in 9 (32%) patients (GROUP I) and surgical treatment was therefore unnecessary;the remaining 19 cases (68%), of which 5 paresis (GROUP II) and 14 paralysis (GROUP III) underwent surgery and within 1 year were all restored to orthotropia. Conclusion: Botulinum toxin type-A (BTX-A) injection is an invaluable tool in the differential diagnosis between paresis versus paralysis of the VIth nerve, allowing the correct choice of surgical procedure.
文摘A previous study has proposed a mathematical model of type-A medial vestibular nucleus neurons (mVNn). This model is described by a system of nonlinear ordinary differential equations, which is based on the Hodgkin-Huxley formalism. The type-A mVNn model contains several ionic conductances, such as the sodium conductance, calcium conductance, delayed-rectifier potassium conductance, transient potassium conductance, and calcium-dependent potassium conductance. The previous study revealed that spontaneous repetitive spiking in the type-A mVNn model can be suppressed by hyperpolarizing stimulation. However, how this suppression is affected by the ionic conductances has not been clarified in the previous study. The present study performed numerical simulation analysis of the type-A mVNn model to clarify how variations in the different ionic conductance values affect the suppression of repetitive spiking. The present study revealed that the threshold for the transition from a repetitive spiking state to a quiescent state is differentially sensitive to variations in the ionic conductances among the different types of ionic conductance.
