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Not AvailableThe postthaw motility and fertility of buffalo and cattle semen is reduced when they are cryopreserved for artificial insemination. In the present study, an attempt was made to characterize the cryogenic changes in proteases and antiprotease activities (APA) of buffalo and cattle semen because these proteolysis regulators have been reported to be associated with sperm motility and fertility. Buffalo sperm demonstrated at least two major proteases of 45 and 42 kDa and three minor proteases of 95, 52, and 33 kDa. Similarly, cattle sperm demonstrated three major proteases of 62, 45, and 42 kDa and two minor proteases of 85 and 78 kDa. Buffalo seminal plasma demonstrated at least three major proteases of 78, 68, and 62 kDa and one minor protease of 98 kDa and cattle seminal plasma demonstrated one major protease of 68 kDa and two minor proteases of 78 and 75 kDa. Except for the 45 kDa protease, most of the previously mentioned proteases were found to be metalloproteinases. Compared with fresh sperm, cryopreserved buffalo and cattle sperm demonstrated a major protease band of 52/49 kDa and the activity of this protease reduced progressively with the duration of cryopreservation. On the contrary, compared with the fresh seminal plasma, cryopreserved buffalo and cattle semen extenders displayed the presence of a new protease band of 45 kDa and demonstrated that this protease activity was leaked from buffalo and cattle cryopreserved spermatozoa. Buffalo and cattle seminal plasmas displayed at least two major APA of 86 and 26 kDa. Compared with buffalo, cattle seminal plasma demonstrated significantly greater APA. Thus, the present study demonstrated the presence of an array of proteases and APA in buffalo and cattle semen and the activities of which changed during cryopreservation. The leakage of the specific protease activity and changes in the proteases and APA might be attributed to reduced motility and fertility of cryopreserved semen in these species.Not Availabl

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Not AvailableThe motility and fertility of mammalian spermatozoa are compromised when they are cryopreserved. Sperm mitochondrial proteins play a vital role in conferring motility. However, the effects of cryopreservation on mitochondria-specific proteins remain primarily unexplored in domestic animals, including buffaloes, so the present study aimed to evaluate this issue. Mitochondria were isolated from both non-cryopreserved and cryopreserved buffalo spermatozoa by sonication followed by sucrose density gradient ultracentrifugation. The purity of the mitochondrial preparation was assessed by cytochrome oxidase assay and electron microscopy. Mitochondria separated from cryopreserved buffalo spermatozoa were associated with significantly lower (P ≤ 0.05) cytochrome oxidase activity as compared with non-cryopreserved spermatozoa. The intensities of two low-molecular-mass mitochondrial proteins (30.1 kDa and 26.1 kDa) were significantly reduced as compared with the non-cryopreserved group. In addition, in cryopreserved buffalo sperm mitochondria, the intensities of three tyrosine phosphorylated proteins (126.6, 106.7 and 26 kDa) increased significantly compared with the non-cryopreserved group. Of these, tyrosine phosphorylation of the 26-kDa mitochondrial protein of cryopreserved sperm was very intense and unique because it could not be detected in the mitochondria of non-cryopreserved sperm. Thus, the study confirmed that both cytochrome oxidase activity and the proteins of buffalo sperm mitochondria undergo significant cryogenic changes in terms of quantity and quality after a cycle of freezing and thawing and this may be one of the important causes of reduced post-thaw motility and fertility of cryopreserved buffalo spermatozoa.ICAR-NIAN

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Not AvailableThe ejaculate composition is extremely complicated and variable among livestock species. The seminal proteins and enzymes are vital for ejaculate metabolism, spermatozoa performance, survival, and transport within the feminine reproductive tract. Proteases and proteinase inhibitors are secreted by the accessory sex glands of the male reproductive tract and find mixed with the spermatozoa throughout ejaculation. But an entire understanding of those enzymes and their performance in numerous class species is not obtainable. The performance of the assorted proteinases and protease inhibitors of ejaculate stay a mystery, and only a few studies are conducted concerning the characterization of those enzymes. Throughout this review, we tend to target the current understanding of the key proteases and their inhibitors of ejaculate from various mammalian species.Not Availabl

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Not AvailableLarge number of proteases have been identified in different parts of the male reproductive tract which play a crucial role during testicular development, epididymal sperm maturation, and sperm-oocyte fusion. The ADAMTS10 is a member of metalloproteinase family, the expression of which is important in sperm-oocyte interaction at the time of fertilization in mouse. In this species, the expression of ADAMTS10 has been demonstrated in testis, and on surface of spermatozoa from caput, corpus and cauda epididymis. Whether ADAMTS10 plays any role in male fertility of any of the domestic ruminant species including buffaloes is not known so far. Thus, a study was conducted to detect the expression of ADAMTS10 in testis, different parts of the epididymis and ejaculated spermatozoa of water buffalo (Bubalus bubalis) using real-time PCR, Western blot, and indirect immunofluorescence techniques. The highest expression of ADAMTS10 transcript was observed in corpus epididymis, followed by cauda, caput and testis. The Western blot detection of ADAMTS10 using the heterologous antibody has demonstrated 31-, 44-, 50-, 65 kDa immune-reactive protein bands from testis and 82 kDa protein from both testes and corpus epididymis and a major 36 kDa protein from ejaculated spermatozoa. The indirect immunofluorescence study has demonstrated high fluorescence signal in post-acrosome and the middle piece region of the ejaculated spermatozoa. Thus the differential expression pattern of ADAMTS10 in different parts of male reproductive tract of buffalo indicates their role in sperm maturation process. However, specific role of ADAMTS10 in buffalo male reproduction can only be ascertained after further studies.ICAR-NIAN