• Asian-Australasian Journal of Animal Sciences
• /
• 제29권9호
• /
• pp.1247-1255
• /
• 2016

Heparin binding proteins (HBPs) are produced by accessory glands. These are secreted into the seminal fluid, bind to the spermatozoa at the time of ejaculation, favour capacitation, acrosome reaction, and alter the immune system response toward the sperm. The present study was conducted with an objective to assess the effect of purified seminal plasma-HBPs (SP-HBPs) on cross bred cattle bull sperm attributes during two phases of cryopreservation: Pre freezing and freezing-thawing. SP-HBPs were purified from pooled seminal plasma by heparin affinity chromatography. Three doses of SP-HBPs i.e. 10, 20, $40{\mu}g/mLs$ semen were standardized to find out the optimum dose and $20{\mu}g/mLs$ was found to be an optimum dose. Semen as such and treated with SP-HBPs was diluted with sodium citrate-egg yolk diluter and cryopreserved as per the standard protocol. Sperm parameters i.e. motility, viability, Hypo-osmotic swelling test (HOST), acrosome damage, in vitro capacitation and lipid peroxidation were evaluated in SP-HBP treated and untreated (control) semen at both phases of cryopreservation. A considerable variation in percent sperm motility, viability, membrane integrity (HOST), acrosome damage, acrosome reaction and lipid peroxidation was observed at both phases among the bulls irrespective of the treatment. Incubation of neat semen with $20{\mu}g/mL$ SP-HBP before processing for cryopreservation enhanced the average motility, viability, membrane integrity by 7.2%, 1.5%, 7.9%, and 5.6%, 6.6%, 7.4% in pre-frozen and frozen-thawed semen in comparison to control. There was also an average increase of 4.1%/3.9% in in vitro capacitation and acrosome reaction in SP-HBPs-treated frozen-thawed semen as compared to control. However, binding of SP-HBPs to the sperm declined acrosome damage and lipid peroxidation by 1.3%/4.1% and 22.1/$32.7{\mu}M$/$10^9$ spermatozoa in SP-HBP treated pre-frozen/frozen-thawed semen as compared to control, respectively. Significant (p<0.05) effects were observed only in motility, HOST and in vitro acrosome reaction. It can be concluded that treatment of neat semen with SP-HBPs before cryopreservation minimized the cryoinjury by decreasing the generation of reactive oxygen species.

• Journal of Animal Science and Technology
• /
• 제49권4호
• /
• pp.443-450
• /
• 2007

본 연구에서는 엘크 사슴의 계절별 정액성상 변화, 동결정액의 융해 후 활력 및 발정동기화 후 인공수정 시간이 수태율에 미치는 영향을 조사하여 효율적인 인공수정 방법을 제시코자 시도하였다. 수사슴에서 정액의 연중 변화는 정액량, 정자 농도, 총 정자 수 및 정자 활력은 7월부터 급격히 증가 되어 정액량과 총 정자 수 및 정자 운동성은 10월에 최고치에 달했고, 정자 농도는 9월에 가장 많은 것으로 조사 되었다. 그 후 정액량, 정자농도, 총 정자 수 및 정자 운동성은 점진적으로 감소하여 정액 양, 정자 농도는 4~6월, 총 정자 수는 4~5월 및 정자 운동성은 6월에 최저 수준으로 감소하였다.번식계절(9~2월)과 비번식계절(3~8월)의 정자 농도, 총 정자 수 및 정자 운동성은 번식계절이 유의적으로 높게 조사되었다(P<0.05). 번식계절과 비번식계절에 채취한 정액의 동결과정 중에 정자 운동성의 결과는 채취 직후, 5℃냉각 후 및 동결 융해 후 활력이 번식계절이 높은 것으로 조사 되었다(P<0.05).인공수정 시간에 따른 수태율은 CIDR 제거 후 60시간에 인공수정 한 처리구가 수태율이 다른 시간들 보다 다소 높았으나 유의적인 차이는 나타나지 않았다.이러한 연구 결과는 엘크 사슴에서 정액 생산과 인공수정을 수행함에 있어 유용한 자료로 이용 될 수 있으며, 발정동기화 후 호르몬 변화, 배란 및 수정 적기 등에 관한 연구가 추가적으로 수행되어야 할 것으로 사료된다.

• Reproductive and Developmental Biology
• /
• 제36권3호
• /
• pp.163-166
• /
• 2012

This study was conducted to determine the relationship between elapsed time after semen preservation on the changes of bacteria and semen quality. Semen was diluted with BTS(Beltsville Thawing Solution) extender without antibiotic for 7 days and sperm parameter and fertility were measured. Sperm motility was measured by CASA and total bacteria number was counted after 22~24 hr incubation from counting agar plate in which sperm dilute to $10{\sim}10^6$ in 0.9% saline solution and inoculate to agar. Acrosomal integrity was measured by Chlortetracycline (CTC) staining. CTC patterns were uniform fluorescence over the whole head (pattern F), characteristic of incapacitated acrosome-intact spermatozoa; fluorescence-free band in the post-acrosomal region (pattern B), characteristic of capacitated acrosome-intact spermatozoa; and almost no fluorescence over the whole head except for a thin band in the equatorial segment (pattern AR), characteristic of acrosome reacted spermatozoa. Total number of bacteria was significantly increased (p<0.0001) 3 days after preservation. Sperm motility, viability, and morphological abnormality on elapsed time after preservation were lower from 5 ($77.24{\pm}6.47$, p<0.001) and 7 days ($77.24{\pm}6.47$, p<0.001) after preservation compared to 1 ($15.71{\pm}7.18$) and 3 days($18.39{\pm}7.22$) after preservation, respectively. Sperm viability was significantly lower ($53.25{\pm}35.03$, p<0.0001) at 7 days after preservation. Morphological abnormality of sperm was lower (p<0.001) at 1 ($15.71{\pm}7.18$) and 3 ($18.39{\pm}7.22$) days compared to 5 ($21.84{\pm}7.91$) and 7 ($22.59{\pm}9.93$) days after preservation. Acrosomal integrity and capacitation rate (pattern F) were significantly lower (p<0.001) from 5 days after preservation. Based on the data we obtained from this study suggested that semen preserved more than 5 days without antibiotic would not recommend use for artificial insemination.

• Animal Bioscience
• /
• 제34권2호
• /
• pp.192-197
• /
• 2021

Objective: The present study evaluated the preservation of ram semen at 0℃ using soybean lecithin with a Tris-fructose extender. Methods: Semen was collected by artificial vagina ejaculation from six rams with proven fertility. High quality ejaculates were diluted by soybean lecithin (0.25%, 0.5%, 0.75%, 1.0%, 1.25%) using Tris-fructose extender and control (Tris-fructose egg yolk extender), respectively. The ejaculates were diluted to a concentration of 5×108 sperm/mL, followed by cooling to 0℃ in 90 min and maintaining the temperature for 12 days. The diluted semen samples were examined and recorded for sperm progressive motility, acrosome integrity at 0, 24, 72, 144, 216, 288 h, respectively. Two hundred and twenty-three ewes were inseminated for 216 h with optimal soybean lecithin concentrated semen or control via trans-cervical insemination. Results: The results showed that there were no differences in sperm progressive motility at 0, 24, 72, and 144 h (p>0.05). After 216 h, the sperm progressive motility in the control group and 0.5% concentration groups was significantly higher when compared to 0.25% concentration (p<0.05). The 0.5% concentration group demonstrated the highest survival rate and had no difference with the control group (p>0.05). At 216 h, the sperm progressive motility of all groups was still above 50%. The acrosome integrity of all groups was decreased with prolongation of storage time, but there was no difference at each time point (p>0.05). There was no significant difference in the lambing rate and pregnancy rate between the 0.5% concentration group and the control group (p>0.05). Conclusion: These results suggest that ram sperm is capable of fertilization after preservation at 0℃ with 0.5% of soybean lecithin in Tris-based extender substituted for egg yolk and produce normal offspring after insemination.

• 한국동물생명공학회지
• /
• 제36권2호
• /
• pp.69-75
• /
• 2021

Cryopreservation is a widely-used efficient means of long-term sperm preservation. However, unlike other types of semen, cryopreserved boar semen has reduced fertility and the efforts continue to optimize post-thawing sperm recovery. In this study, we evaluated the effects of various washing solutions (Hulsen solution, lab-made DPBS and commercial DPBS) on post-thawing porcine sperm kinematics (CASA system), viability (SYBR-14/PI) and acrosome integrity (PSA/FITC). We also examined the effect of washing-centrifugation on frozen-thawed semen kinematics. The results indicate that type of washing solution and post-thawing centrifugation alters parameters linked to sperm quality (total motility, progressive motility, viability and acrosome integrity). Significantly higher (p < 0.05) motility and progressive motility were obtained when cryopreserved semen was processed with Hulsen solution. The post-thaw percentage of live and intact acrosomal sperm was significantly higher in group 1 (Hulsen solution) as compared to other groups. Following thawing-centrifugation, the results showed significantly higher motility and progressive motility in group 1 than other groups. However, the latter two DPBS groups did not differ statistically. Taken together, Frozen-thawed spermatozoa motility, acrosome integrity and viability can be affected by the type of washing solution used. Moreover, centrifugation of frozen-thawed semen has an unfavorable effect on total motility and progressive motility.

• Animal Bioscience
• /
• 제35권11호
• /
• pp.1666-1674
• /
• 2022

Objective: Letrozole, a potent aromatase inhibitor, is known to have the potential to modify male reproductive function by altering sex hormone levels. This study aimed to evaluate the semen and testicular characteristics and hormonal profile of aged Mrakhoz bucks (Capra hircus) treated with letrozole. Methods: Twelve Markhoz male goats, aged between 4.5 to 5.5 years with an average body weight (BW) of 61.05±4.97 kg were used for the study. Animals were randomly divided into two equal groups and subcutaneously received either 0.25 mg/kg BW of letrozole or a control every week for 2 months. The semen collections were performed every 10 days, and blood samples and testicular biometric records were collected at 20 days intervals. Results: Letrozole causes increased testosterone and follicle-stimulating hormone levels, testosterone to estradiol ratio, semen index and reaction time during the period from 20th to 60th days (p<0.05). Furthermore, letrozole-treated bucks had higher semen volume, sperm concentration, and total sperm per ejaculate from 30th to 60th days (p<0.05). However, no differences occurred between the groups in scrotal circumference, relative testicular volume, semen pH, abnormality, acrosome integrity, and membrane integrity of sperm during the study (p>0.05). The serum luteinizing hormone levels, sperm viability, motility, and progressive motility increased, and estradiol levels decreased after 40th to 60th days of letrozole treatment (p<0.05). Conclusion: Letrozole application to aged Markhoz bucks provokes reproductive hormonal axis which, in turn, induces enhancement of semen production and quality.

• Clinical and Experimental Reproductive Medicine
• /
• 제50권4호
• /
• pp.262-269
• /
• 2023

Objective: The aim of this study was to compare semen parameters and sperm DNA fragmentation (SDF) and explore the relationship between semen parameters and SDF between 2 and 7 days of abstinence and a short abstinence period (within 4 hours) in oligozoospermic infertile patients. Methods: Two semen samples were collected from infertile oligozoospermic men (n=34) after an abstinence period of 2 to 7 days and within 4 hours, respectively. Sperm parameters were compared between the two abstinence duration groups, including semen volume, sperm concentration, total sperm count, sperm motility, total motile sperm count (TMSC), morphology, and SDF. Results: The semen volume, concentration, and total sperm count were significantly decreased after 4 hours of abstinence than after 2 to 7 days of abstinence, with median differences of 1.2 mL (p<0.001), 2×10⁶/mL (p=0.011), and 9.6×10⁶/ejaculation (p<0.001), respectively. TMSC was significantly lower after a short abstinence, with a median difference of 4.24×10⁶/ejaculate (p<0.001). However, there were no significance differences in the percentage of motility, the SDF, and the percentage of sperm with normal morphology. Interestingly, volume, concentration, total sperm count, sperm motility, and SDF, but not TMSC, exhibited significant linear correlations between the two abstinence groups in univariate regression analysis, except for TMSC. Conclusion: In oligozoospermic men, the volume, concentration, and total sperm count were significantly lower after a short abstinence period, but without adverse effects on sperm motility and SDF.

• Animal Bioscience
• /
• 제34권8호
• /
• pp.1253-1270
• /
• 2021

Assessment of male fertility is based on the evaluation of sperm. Semen evaluation measures various sperm quality parameters as fertility indicators. However, semen evaluation has limitations, and it requires the advancement and application of strict quality control methods to interpret the results. This article reviews the recent advances in evaluating various sperm-specific quality characteristics and methodologies, with the help of different assays to assess sperm-fertility status. Sperm evaluation methods that include conventional microscopic methods, computer-assisted sperm analyzers (CASA), and flow cytometric analysis, provide precise information related to sperm morphology and function. Moreover, profiling fertility-related biomarkers in sperm or seminal plasma can be helpful in predicting fertility. Identification of different sperm proteins and diagnosis of DNA damage has positively contributed to the existing pool of knowledge about sperm physiology and molecular anomalies associated with different infertility issues in males. Advances in methods and sperm-specific evaluation has subsequently resulted in a better understanding of sperm biology that has improved the diagnosis and clinical management of male factor infertility. Accurate sperm evaluation is of paramount importance in the application of artificial insemination and assisted reproductive technology. However, no single test can precisely determine fertility; the selection of an appropriate test or a set of tests and parameters is required to accurately determine the fertility of specific animal species. Therefore, a need to further calibrate the CASA and advance the gene expression tests is recommended for faster and field-level applications.

• Asian-Australasian Journal of Animal Sciences
• /
• 제5권3호
• /
• pp.439-447
• /
• 1992

Eight healthy Holstein bulls, 4-6 years old were used to study the effect of season of the year on the biophysical characteristics of semen. Semen was collected twice a week by AV (artificial vagina) over one-year period. The analyses revealed that all the basic seminal traits studied were differed significantly due to season, except the ejaculate volume and consistency and the percentage of swollen spermatozoa in a hypo-osmotic fructose-citrate solution. Ejaculates collected during hot summer season had significantly lower sperm motility, concentration and total counts, and higher percentage of dead spermatozoa than those collected during winter time. Warm spring had moderate semen quality. The temperature-humidity index was calculated and it was associated (p < 0.01) negatively with the ejaculate pH, sperm concentration and total counts, and positively with the % of dead sperms. Ejaculate volume, percentage of swollen spermatozoa, individual motilities did not correlate significantly with the change in temperature-humidity index values. The total live, motile spermatozoa per ejaculate during both the winter and spring seasons showed significant increase of about 37% and 32% respectively over the summer season. Also, rectal temperatures of the bulls were elevated during the hot summer season, while the values of blood hemoglobin and packed-cell volume were decreased.

• 생약학회지
• /
• 제40권1호
• /
• pp.18-24
• /
• 2009

GCSB-5 preparation is a purified extract from a mixture six herbal medicines (Acanthopanacis Cortex, Achyranthis Radix, Saposhnikoviae Radix, Cibotii Rhizoma, Glycine Semen Nigra, Eucommiae Cortex) that have been widely used in traditional medicine to treat various bone disorders. This study was carried out to obtain the HPLC analysis method that can be used to establish quantitative analysis of Glycine Semen Nigra and Eucommiae Cortex for standardization of GCSB-5 preparation. HPLC analysis methods for the simultaneous determination of genistin (Glycine Semen Nigra) and geniposide (Eucommiae Cortex) were established for the quality control of herbal medicinal raw material and preparation. And validation of HPLC analysis methods were conformed for verification of HPLC methods by check to specificity, linearity, intra-day precision, inter-day precision and accuracy following ICH guideline. As the result of quantitative analysis, the contents of genistin and geniposide in the raw material of GCSB-5 preparation were 0.0426-0.0427 mg/g and 0.431-0.432 mg/g. And GCSB-5 preparation contained genistin of 0.0202-0.0203 mg/capsule and geniposide of 0.211-0.212 mg/capsule, respectively.