• Title/Summary/Keyword: Bovine Testicular

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Optimization of Bovine Testicular PH-20 hyaluronidase Production in Pichia pastoris (소의 히아론산 분해효소(PH-20)의 Pichia pastoris에서의 생산 최적화)

  • Shin, Hwa Shook;Kim, Eunki
    • Korean Chemical Engineering Research
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    • v.46 no.4
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    • pp.764-768
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    • 2008
  • Bovine testicular hyaluronidase PH-20 was cloned into pPIC9 vector and expressed in Pichia pastoris. Recombinant PH-20 was 75 kDa MW and 7460 units/L activity. Extracellular hyaluronidase activity was two times higher than that of intracellular activity. Non-buffered medium and $30^{\circ}C$ cultivation was favorable for PH-20 production. 1M sorbitol as an osmotic pressure and 0.3% methanol inducer increased cell growth and enzyme activity. 0.4 M arginine augmentation decreased the proteolytic degradation of recombinant hyaluronidase.

A Study on Growth of Human Testicular Tissue in 3-Dimensional Collagen Gel Tissue Culture (Collagen Gel을 이용한 사람의 고환 조직배양에 관한연구)

  • Lee, Choong-Hyun;Lee, Sang-Cheol;Lee, Sun-Joo;Sohn, Joon-Woong;Chang, Sung-Goo;Kim, Jin-Il;Chai, Soo-Eung
    • Clinical and Experimental Reproductive Medicine
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    • v.20 no.1
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    • pp.53-56
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    • 1993
  • A recently developed collagen gel culture technique has been applied to study on growth in tissue of human testicular tissue. Minimum Eagle's medium supplemented with amino acid, 10% Fetal Bovine Serum and 0.1mM non-essential amino acid are emploid. Tissue fragments on collagan gel are fixed at time intervals for the histologic findings of testis. The mature spermatids are maintained for 2 weeks and can be observed until four weeks. But the rate of glucose consumption is increased contrary to histologic findings.

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Optimal Milieu for Culturing Porcine Sertoli Cell

  • Jabed Md. Anower;Kamal Tania;Kim, Byung-Ki
    • Reproductive and Developmental Biology
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    • v.30 no.3
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    • pp.163-167
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    • 2006
  • The purpose of the present study was to establish culture conditions for the in vitro study of the neonatal piglet Sertoli cell. Isolation for the culture of Sertoli cell was established using collagenase and pancreatin digestion of testicular tissues. The effects of various culture media, fetal bovine serum(FBS), follicular stimulating hormone(FSH), epidermal growth factor(EGF) and insulin-transferrin-sodium selenite(ITS) on growth of neonatal piglet Sertoli cells were investigated. The mitogenic effects of Dulbecco's modified Eagle's medium+Ham's F-12 medium was higher than other media used in this experiment. The addition of 1% FBS in cultures was necessary for attachment of Sertoli cell clusters. However, except FBS and EGF, FSH and ITS did not stimulate Sertoli cell proliferation. When Sertoli cells isolated from neonatal piglets were cultured in Dulbecco's modified Eagle's medium+Ham's F-12 medium supplemented with 1% FBS, FSH EGF and ITS, the yield and plating efficiency of Sertoli cells were largely increased. Confluency of Sertoli cells was reached as early as 4 days of culture. The method described here reduces or eliminates many of the drawbacks of the conventional procedures used to isolate and culture of Sertoli cells, thus providing a useful tool in studies of growth kinetics and regulation of cell proliferation in vitro.

Antiviral effects of Bovine antimicrobial peptide against TGEV in vivo and in vitro

  • Liang, Xiuli;Zhang, Xiaojun;Lian, Kaiqi;Tian, Xiuhua;Zhang, Mingliang;Wang, Shiqiong;Chen, Cheng;Nie, Cunxi;Pan, Yun;Han, Fangfang;Wei, Zhanyong;Zhang, Wenju
    • Journal of Veterinary Science
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    • v.21 no.5
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    • pp.80.1-80.13
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    • 2020
  • Background: In suckling piglets, transmissible gastroenteritis virus (TGEV) causes lethal diarrhea accompanied by high infection and mortality rates, leading to considerable economic losses. This study explored methods of preventing or inhibiting their production. Bovine antimicrobial peptide-13 (APB-13) has antibacterial, antiviral, and immune functions. Objectives: This study analyzed the efficacy of APB-13 against TGEV through in vivo and in vitro experiments. Methods: The effects of APB-13 toxicity and virus inhibition rate on swine testicular (ST) cells were detected using 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT). The impact of APB-13 on virus replication was examined through the 50% tissue culture infective dose (TCID50). The mRNA and protein levels were investigated by real-time quantitative polymerase chain reaction and western blot (WB). Tissue sections were used to detect intestinal morphological development. Results: The safe and effective concentration range of APB-13 on ST cells ranged from 0 to 62.5 ㎍/mL, and the highest viral inhibitory rate of APB-13 was 74.1%. The log10TCID50 of 62.5 ㎍/mL APB-13 was 3.63 lower than that of the virus control. The mRNA and protein expression at 62.5 ㎍/mL APB-13 was significantly lower than that of the virus control at 24 hpi. Piglets in the APB-13 group showed significantly lower viral shedding than that in the virus control group, and the pathological tissue sections of the jejunum morphology revealed significant differences between the groups. Conclusions: APB-13 exhibited good antiviral effects on TGEV in vivo and in vitro.

Ferritin Overload Suppresses Male Fertility Via altered Acrosome Reaction

  • Kwon, Woo-Sung;Rahman, Md Saidur;Kim, Ye-Ji;Ryu, Do-Yeol;Kahtun, Amena;Pang, Myung-Geol
    • Reproductive and Developmental Biology
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    • v.39 no.4
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    • pp.117-125
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    • 2015
  • Iron is required for cell viability but is toxic in excess. While the iron-mediated malfunction of testicular cells is well appreciated, the underlying mechanism(s) of this effect and its relationship with fertility are poorly understood. Ferritin is a ubiquitous intracellular protein that controls iron storage, ferroxidase activity, immune response, and stress response in cells. Ferritin light chain protein (FTL) is the light subunit of the Ferritin. Previously, we had identified the FTL in bovine spermatozoa following capacitation. In present study, to investigate the role of Ferritin in sperm function, mice spermatozoa were incubated with multiple doses (1, 10 and $100{\mu}M$) of sodium nitroprusside (SNP), an iron donor. SNP was increased Ferritin levels in a dose-dependent manner. The Ferritin was detected on the acrosome in spermatozoa by immunocytochemistry. Short-term exposure of spermatozoa to SNP increased tyrosine phosphorylation and the acrosome reaction (AR). Finally, SNP affected a significant decrease in the rate of fertilization as well as blastocyst formation during early embryonic development. On the basis of these results, we propose that the effects of Ferritin on the AR may reduce overall sperm function leads to poor fertility in males and compromised embryonic development.

Characterization and In Vitro Differentiation of Korean Ring-Necked Pheasant (Phasianus colchicus) Male Germ Cells

  • Jeong, Dong Kee;Sharma, Neelesh;Nguyen, Thanh Luan;Kim, Jong Hyun;Oh, Sung Jong
    • Journal of Embryo Transfer
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    • v.29 no.4
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    • pp.351-359
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    • 2014
  • Phasianus colchicus is not only a beautiful bird but also a great value in science and under the threat of endanger. Hence, the aim of this study was to isolate the pheasant male germ cells (mGCs) and then induce them into elongated sperm-like cells in vitro. The mGCs were purified and enriched by a two-step plating method based on the different adherence velocities of mGCs and somatic cells. The percentage of the c-kit positive cells and c-kit negative cells examined by flow cytometry analysis (FCA) was 92.87% and 2.57%, respectively. Subsequently, the mGCs were induced for 48h in DMEM/F12 medium supplemented factors such as retinol acid, testosterone and bovine FSH, followed by 5 weeks in culture. We found that some elongated sperm-like cells appeared initially in vitro under inducement of stimulated factors. The elongated sperm-like cells showed in the expression of changed morphology and post-transcriptional marker such as spermatid associated (SPERT), spermatid perinuclear RNA binding protein (STRBP), round spermatid basic protein 1 (RSBN1) and SPER1L. Moreover, in DNA content identified assay, induced cells showed that the 1C DNA population markedly increased in differentiated group but it was not change in undifferentiated group. Successful in vitro differentiation of pheasant testicular germline cells into spermatids appears to offer extremely attractive potential for the conservation of endangered birds and treatment of male infertility.

Evaluating the effect of conditioned medium from mesenchymal stem cells on differentiation of rat spermatogonial stem cells

  • Hoda Fazaeli;Mohsen Sheykhhasan;Naser Kalhor;Faezeh Davoodi Asl;Mojdeh Hosseinpoor Kashani;Azar Sheikholeslami
    • Anatomy and Cell Biology
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    • v.56 no.4
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    • pp.508-517
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    • 2023
  • In cancer patients, chemo/radio therapy may cause infertility by damaging the spermatogenesis affecting the self-renewal and differentiation of spermatogonial stem cells (SSCs). In vitro differentiation of stem cells especially mesenchymal stem cells (MSCs) into germ cells has recently been proposed as a new strategy for infertility treatment. The aim of this study was to evaluate the proliferation and differentiation of SSCs using their co-culture with Sertoli cells and conditioned medium (CM) from adipose tissue-derived MSCs (AD-MSCs). Testicular tissues were separated from 2-7 days old neonate Wistar Rats and after mechanical and enzymatic digestion, the SSCs and Sertoli cells were isolated and cultured in Dulbecco's modified eagle medium with 10% fetal bovine serum, 1X antibiotic, basic fibroblast growth factor, and glial cell line-derived neurotrophic factor. The cells were treated with the CM from AD-MSCs for 12 days and then the expression level of differentiation-related genes were measured. Also, the expression level of two major spermatogenic markers of DAZL and DDX4 was calculated. Scp3, Dazl, and Prm1 were significantly increased after treatment compared to the control group, whereas no significant difference was observed in Stra8 expression. The immunocytochemistry images showed that DAZL and DDX4 were positive in experimental group comparing with control. Also, western blotting revealed that both DAZL and DDX4 had higher expression in the treated group than the control group, however, no significant difference was observed. In this study, we concluded that the CM obtained from AD-MSCs can be considered as a suitable biological material to induce the differentiation in SSCs.