Asian Pacific Journal of Reproduction

279-286

Authors: Norihiro Tada, Fumio Kanai, Eri Nakamura, Hongmei Lu, Masahiro Sato

To examine whether functional spermatozoa can be obtained when a whole male gonadal tissue (testes, epididymides, and fat) isolated from neonatal mice is grafted underneath adult mouse testes. Methods Neonatal (1-day-old) male gonadal tissue, isolated from enhanced green fluorescent protein (EGFP)-transgenic (Tg) mice (C57BL/6-Tg(ACTB-EGFP)1Osb/J), was inserted deep in the testis of a non-Tg recipient mouse through a tunica albuginea incision. Two months after transplantation, the fluorescent grafted tissues were retrieved from recipient mice. Results Histological analysis demonstrated that epididymal architecture was well developed and that spermatogenesis in the testis occurred in 30–60% of each seminiferous tubule of all the grafted tissues examined. Interestingly, motile spermatozoa could be successfully retrieved from the portion corresponding to the cauda epididymis in 1 of the 7 transplants obtained. These obtained spermatozoa had transgenes and could support embryonic development when intracytoplasmic sperm injection was performed using frozen-thawed spermatozoa. Conclusion This present technique will be useful for study in various biological fields including the rescue of Tg lines with lethal postnatal phenotypes and cloned animals that die immediately after birth.

295-300

Authors: Mark Anthony C. Mamon, Cheska May M. Menodiado, Glenn L. Siasu, Gliceria B. Ramos

Abstract Objective To assess the histoarchitecture of the maternal liver and kidney as influenced by selenium supplementation within the periconception period and to determine which of the periconception stages of selenium intake is most favorable of the liver and kidney. Methods Thirty-six 7-week-old female mice were divided into four groups where all groups were given 6.0 g of food pellets per day. Un-supplemented group (U) received no selenium in the food pellet, Pre-gestation supplemented group (P) was given 3.0 μg selenium/d for 21 d, Pre-gestation to Gestation supplemented group (PG) was given 3.0 μg selenium/d for 37 d, and Gestation supplemented group (G) was given 3.0 μg selenium/d for 16 d only. Results The occurrences of vacuolated hepatocytes and congested central veins in the liver were observed. The P and the G groups incurred the lowest and highest percent occurrence, respectively, of murine mothers that exhibited presence of vacuolated hepatocytes. The mean percent occurrence of congested central veins did not significantly vary among groups. The occurrences of shrunken and swollen glomeruli were observed in the kidney. The mean percent occurrence of shrunken glomeruli showed that P and G groups were significantly lower than those of U and PG groups. The mean percent occurrences of swollen glomeruli did not vary significantly among groups. Conclusion The histological analyses of the liver and the kidney obtained from different stages of periconception indicate that selenium supplementation would be best during pregestation stage.

311-316

Authors: V.U. Nna, E.O. Ofutet, O.E. Ofem, D.C. Okpokam, E.E. Osim

Objective To determine the prevalence of erectile dysfunction (ED), the use of sex stimulants, the motivation behind the use of sex stimulants and sexual satisfaction with or without the use of sex stimulants among male subjects in Calabar, Nigeria. Methods Male subjects aged 15–74 years were recruited for this study. Exclusion criteria was employed to eliminate responses of subjects who have had no prior engagement in any sexual activity, and were currently having sex less than 4 times a month. After applying the exclusion criteria, 2 010 respondents were obtained. Results Out of the 2 010 subjects, 11 were illiterates (0.5%), 605 (30.1%) had primary education, 1 317 (65.5%) had secondary education, 50 (2.5%) were undergraduates and 27 (1.3%) had post graduate education. Out of the 2 010 respondents, 45 (2.2%) had knowledge of the causes of ED while 1 965 (97.8%) were ignorant of the causes of ED. Fifty one respondents (2.5%) were currently suffering ED, while 1 959 (97.5%) had no history of ED. A significant (P < 0.001) positive correlation (0.531) between age and incidence of ED was observed. Forty one (2%) respondents had never and were currently not using sex stimulants, while 1 969 (98%) were currently using sex stimulants. Out of the 2 010 respondents, 1 745 (86.8%) used sex stimulants to increase their genital size, of which 1 622 (92.9%) were aged 15–34 years. Conclusion The incidence of ED in Calabar, Nigeria, increases with increasing age. Although the respondents between the ages of 15–34 years had the least incidence of ED, this age group used sex stimulants the most following their perceived beneficial effect in increasing genital size.

321-325

Authors: Ali Olfati, Gholamali Moghaddam, Nasroallah Moradi Kor, Behzad Baradaran

Objective This study was conducted to investigate the trace mineral concentrations (Mg, Se, Zn, Cu and Fe) in amniotic fluid (AF) in 40 pregnancy toxemia Ghezel ewes at the time of parturition phase and its association with placental traits. Methods Animals were treated with controlled internal drug release for 14 d and injected 400 IU pregnant mare serum gonadotropin at the time of controlled internal drug release removal. After the detection of estrus by use of teaser rams, ewes were hand-mated. Ewes were classified as having subclinical pregnancy toxemia on the basis of beta-hydroxy butyrate (BHBA) results (BHBA > 0.86 mmol/L). Results The overall mean AF traces of mineral concentrations were 3.13 ng/mL, 22.1 μg/dL, 134.7 μg/dL, 122.5 μg/dL and 166.6 μg/dL, respectively. There was a significant positive correlation between placental efficiency and Zn concentration in AF ewes (r = 0.633, P < 0.01), while the relationship was significantly negative between total volume of amniotic fluid and Fe concentration in AF ewes (r = −0.717, P < 0.01). In this research, no relationship between Se, Mg and Cu trace minerals was observed in AF ewes with placental traits ewes. Results of laboratory analyses demonstrated no relationship between BHBA concentrations and placental traits (P > 0.01), except for placental weight (r = 0.808, P < 0.01). Also, no significant correlation was detected between BHBA with the above trace minerals. Conclusions Overall, determinations of these trace minerals in the AF ewes could have been used to obtain information on nutritional and reproductive status for the diagnosis of pregnancy toxemia in Ghezel ewes.

331-334

Authors: Reda I. El-Sheshtawy, Diya A. El-Badry, Gamal A. El-Sisy, Walid S. El-Nattat, Amal M. Abo Almaaty

Objective To investigate the effect of extender supplementation with different concentrations of a honey bee on post-thawed sperm motility, viability index, membrane and acrosome integrities in Arab stallion. Methods Five ejaculates from each of four Arabian stallions were subjected to cryopreservation with a modified INRA-82, without any supplementation (control) or supplemented with 1%, 2%, 3%, 4% and 5% honey bee. After thawing, all samples were maintained at 37 °C, while analyses were performed at 0, 1, and 2 and 3 h. Sperm motility percentage, viability index percentage, membrane integrity percentage and acrosome integrity percentage of each sample were determined by conventional laboratory methods. Results Relative to the control group, supplementation with honey (2%, 3% and 4% significantly improved (P < 0.01 at least) post-thaw sperm motility, viability index (P < 0.001 at least) and had a positive effect on membrane integrity and intact acrosome percentage (P < 0.001 at least) at 0, 1, 2 and 3 h post-thawing. For all semen parameters, the lower concentration of honey (1%) and higher concentration (5%) did not show significant differences (P > 0.05) compared with the control. Conclusion Honey bee supplementation in Arab stallion semen extenders provided a better protection of sperm parameters against cryopreservation injury, in comparison to the control groups.

340-344

Authors: Mohammad Rahim Ahmadi, Ali Kadivar, Mahmood Vatankhah

Objective This study evaluated the cervical sampling as an easy and safe method for the diagnosis of subclinical endometritis in dairy cattle. Methods One hundred ninety seven lactating Holstein cows were examined at 26–32 d in milk (DIM) for diagnosis of endometritis. Differential cellular counts were also made from stained smears of the cervical mucosa. Using the Receiver/Response Operating Characteristics (ROC) curve, presence of >17.5% polymorphonuclear (PMN) cells was calculated for detection of subclinical endometritis with sensitivity and specificity of 56.5% and 83.3% respectively. Results Cows with subclinical endometritis had significantly more open days and all service conception rate than normal cows. The results of survival analysis showed that normal cows became pregnant at a significantly faster rate than cows with subclinical endometritis. Conclusions The results of the present study introduced the cervical smear sampling as an easy and suitable method for diagnosis of subclinical endometritis in dairy cattle.

351-353

Authors: Ferjaoui Mohamed Aimen, Youssef Atef, Gharrad Majed, Achour Radhouane, Mbarki Manel, Malek Monia, Néji Khaled, Reziga Hedi

The vaginal aplasia is a rare Mullerian duct malformation characterized by the absence of the vagina. The management of this malformation is based on surgery to restore a normal sexual life and eventually to hope having a normal fertility. In this case, we presented a spontaneous pregnancy in a 15 year-old girl who underwent a vaginoplasty with re-implantation of the uterus corpus in the neovagina.

356

Authors: Beuy Joob, Viroj Wiwanitkit

no

179-181

Authors: Nirav R. Soni

Zika virus (ZIKV) is a member of the virus family Flaviviridae and the genus Flavivirus, transmitted by daytime-active Aedes mosquitoes, such as A. aegypti. ZIKV will continue to spread and it will be difficult to determine how the virus will spread over time. Sign and symptoms of ZIKAVD (Zika virus disease) were conjunctivitis (red eyes), back pain, birth defect-abnormal brain development known as microcephaly and it is diagnosed through PCR (polymerase chain reaction) and virus isolation from blood samples.

188-192

Authors: T. Auta, A.T. Hassan

Objective The study was designed to investigate the alterations in oestrus cycle and implantation in Mus musculus administered aqueous wood ash extract of Azadirachta indica. Methods Mice were randomly grouped into four groups (5/group) where 0, 5, 50 and 100 mg/kg doses of the extract were orally administered for 20 d and microscopy of vaginal smear carried out daily to determine anti-ovulatory activity. Oestrus cycle, including metestrus, diestrus and oestrus phases and histopathology of uterus were examined. To assess abortifacient activity, pregnant mice were also administered extracts for 7 d and laparotomised on the 10th day of pregnancy to observe implantation in the horns of the uterus. Results Duration of metestrus and diestrus phases varied significantly, while the frequency at which the oestrus phase occurred was lowest at 100 mg/kg, (2.8 ± 0.2) and (1.2 ± 0.4) d, with degeneration of epithelial cells of glands in the uterus. Failure of implantation (0% implantation) was recorded in mice exposed to 50 and 100 mg/kg of the extract. Conclusions Aqueous wood ash extract of A. indica altered the duration of metestrus, diestrus and oestrus phases of the oestrus cycle and compromised implantation in female mice.

198-203

Authors: R. Heru Prasetyo, Erma Safitri

Objective Auto-regeneration of the intestinal and ovarian tissue are experiencing degenerative due to protein energy malnutrition (PEM) through of auto-mobilization, increase of immune response and differentiation of endogenous stem cells. Methods Female rat model of PEM obtained through fasting meals for 5 d and only to drink, causing malnutrition and damage of the intestinal and ovarian tissue. Furthermore, the therapy of honey with a dose of 30% water (T1) and 50% water (T2), respectively for 5 d and compared with the positive control, were fasted without being given honey (T0+) and negative control, not fasted and without being given honey (T0−). Observations on: auto-mobilization, increased immune response, differentiation of stem cells, and regeneration of the intestinal and ovarian tissue with HE staining. Results Auto-mobilization of stem cells based on the expression of CD34+ and CD45+, which is a marker of endogenous stem cells (hematopoietic stem cells/HSCs). Increased immune response is based on Hsp70 expression and PGE2 in intestinal tissue. Differentiation of stem cells into progenitor cells that expected based expression of growth differentiation factor-9 (GDF-9) by immunohistochemistry in ovarian tissue. Conclusions Expression of CD34+ and CD45+, which significantly different in treatment 2 (2). Furthermore, increase of immune response (decrease Hsp70 expression and increased PGE2) in intestinal tissue. Increased immune response causes expression of GDF-9 in ovarian tissue. Decreased of Hsp70 expression, increased PGE2 and increased GDF-9 followed the process of regeneration of the intestinal and ovarian tissue.

210-213

Authors: Raichel Jacob, Chandirasekar Ramachandran, Calistus Jude, Uthayakumar Venkatachalam, Sasikala Keshav Rao

Objective To investigate the frequency of Pro12Ala polymorphism in PPAR gamma gene among PCOS of South Indian Population. Understanding this polymorphism may help us in better diagnosis prevention and therapeutic approaches towards management of PCOS. Method A total of 75 women with the diagnosis of PCOS and 75 healthy controls were included in this study. The Pro12Ala variant in the PPAR gamma gene was analysed by PCR-RFLP. Results Genotypic frequencies for PPAR gamma gene Pro12Ala polymorphism showed the frequency of Pro/Pro genotypes was 77% in PCOS and 81% in controls. The odds ratio was 0.71 (95% CI 0.31–1.63) and 2.66 (95% CI 0.21–32.9) and alleles expressed 0.87 (95% CI 0.42–1.79). Regarding hormone levels, there were significant differences between PCOS and non PCOS. Conclusion PPARγ2 gene Pro12Ala polymorphism was supposed to be susceptible genes in PCOS. The present study demonstrated that there is a statistical difference between the distributions of PPAR gamma Pro12Ala polymorphism in South Indian Population.

221-226

Authors: Amir Hossein Asgari Safdar, Ali Asghar Sadeghi

Objective This study was conducted in order to the administration of 50 and 100 mg of bovine somatotropin (bST) at the beginning of the estrus synchronization and natural mating of the sheep to evaluate the improvement of the ovulation rate, embryonic development and pregnancy rate of the transferred embryos. Methods Forty eight donors were treated with three different types of treatment; Group A: treated with bST-100 (n = 15), received 100 mg of bST at the beginning of the synchronization and natural mating, Group B: treated with 50 mg of bST (n = 15) same as the previous group and control (n = 18) did not receive any type of bST. Each recipient received two embryos, (n = 108): 30 recipients received the embryos from bST-100s, 45 recipients received the embryos from bST-50 and 33 recipients received embryos from the control group. Using SAS related GENMOD method, rate of superovulatory, recovered structure percentage, cleavage rate, transferable embryo percentage, quality of embryos, rates of pregnancy and embryonic development were analyzed. Using GLM procedure, numbers of corpus luteum and blastocyst cells were analyzed. Results The bST administration had no significant effect on rate of superovulatory, number of CL and recovered structures (P ≥ 0.05). Number of transferable embryos and embryos that had access to the blastocyst in bST-50 (P ≤ 0.01) was more than bST-100 and control group. Conclusions The treatment 50 mg bovine somatotropins enhance the ratio and growth of the transferable embryos. Embryos of bST-50 treatment indicated an improved embryonic development but bST did not affect the pregnancy rates of transferred embryos.

232-235

Authors: Syed Ali Abid Bukhari, Shujait Ali, Muhammad Zubair, Ijaz Ahmad, Ubaid Ur Rehman

Objective To evaluate the ovulation inducing effect of clomiphene citrate along with human chorionic gonadotropin (hCG) in prepubertal Sahiwal heifers. Clomiphene citrate (an anti-estrogen) causes gonadotropin secretion from pituitary gland by removing the negative feedback effect of estrogen. This causes more FSH secretion which ultimately stimulates follicular growth and upon hCG administration (luteinizing hormone activity) ovulation occurs. Methods Twelve prepubertal Sahiwal heifers were divided into experimental and control groups, with six animals in each group. The grouping of animals was made on the basis of plane ovary. The status of ovarian puberty was checked through rectal palpation and history of previous estrus signs. Experimental heifers were given clomiphene citrate at 300 mg/heifer for 9 d orally, while heifers in control group were given placebo tablets. The dose of clomiphene citrate was selected on the basis of previous literature. On day 10, hCG (IVF-C 5000 iu (hCG) LG Life Sciences, Korea) was given to all heifers of both groups at 2 500 IU/heifer through intravenous route. Two base line blood samples were collected before start of the treatment at each rectal palpation examination for selection of animals. Subsequently, during and after treatment three blood samples per week (on alternate days) for 4 weeks were collected for measuring the progesterone (P4) in serum. The concentration of progesterone was measured though enzyme linked immunosorbent assay. For the comparison of variables, student t-test was applied. Results The P4 concentrations in five heifers (83.3%) in the experimental group with (P4 > 1 ng/mL) and one heifer (16.7%) in the control group along with corpus luteum on ovary showed ovulation. One heifer in control group ovulated that might be due to hCG. Mean progesterone concentration was significantly higher (P < 0.05) in treated heifers than in control heifers. Conclusion Administration of clomiphene citrate and hCG can induce ovulation induction in prepubertal Sahiwal heifers.

240-246

Authors: Mark Anthony C. Mamon, Julie Andrea P. Añano, Louise C. Abanador, Genesis Julyus T. Agcaoili, Caira B. Sagum, Ria Lorraine H. Pagliawan, Jean Mherck B. Tapere, Jessie Bob M. Agravante, John Harold G. Arevalo, Abdul Jabbar A. Minalang

Objective To determine the effects of the water quality of Manila Bay on allometric parameters and histological biomarkers of selected organs of P. viridis. Methods Green mussels were collected from two coastal sites of Manila Bay, Las Piñas – Parañaque (LPP) and Bacoor, Cavite (BC). Twenty-four green mussels from each site were used for the assessment of allometric parameters, and six green mussels from LPP and eight from BC were used for the assessment of histological structures of gonads, gut, and digestive glands. Gonad development was categorized into five stages, whereas gut and digestive glands were scored into four categories. Results Allometric parameters that include shell height, weight, and total wet and dry soft tissue weight were significantly different between LPP and BC. It was also observed that exposure to the pollutants in Manila Bay resulted to delays in gonadal development, and detrimental changes and lesions in the histostructure of digestive gland and gut. Conclusions Pollutants in Manila Bay have detrimental effects to the growth, reproductive development, and histological structure of digestive organs of P. viridis.

252-255

Authors: Reda I. El-Sheshtawy, Walid S. El-Nattat, Samy I.A. Shalaby, Mohamed I. Shahba, Islam E. Al-Se'dawy

Objective The benefits of dates palm and their pollen grains in the human nourishment system have their consideration and importance in medical and nutritional point of view. The present study aimed to investigate a new perspective for the use of date palm pollen grains (DPPG) aqueous extract or infusion in preservation of chilled and frozen buffalo semen. Methods Pooled buffalo semen were diluted by Tris-Citrate-Fructose egg yolk (TCFY) diluent (considered as control) and diverse concentrations of clarified soaked pollen grains in Tris-Citrate-Fructose (TCF) diluent in concentrations of 50, 100, 150, 200 and 250 mg DPPG/5 mL TCF. Five concentrations were kept without egg yolk (TPG) while other five concentrations contained 20% egg yolk (TPGY). Each of the two dilutions was enumerated according to the concentration of DPPG. Extended semen is then chilled (for 7 d) and cryopreserved. Motility, alive sperm and intact sperm membrane (HOST) % were recorded as characteristic sperm parameters. Results After 7 d of chilling, significant (P < 0.000 1) high percent of motile sperms was recorded for TPG 50, 100, 150, 200 and 250 (66.67, 65.00, 61.67, 61.67 and 63.33, respectively). After freeze thawing, TPGY 150 and 250 showed the significant (P < 0.000 1) high percent of motile sperms (43.75 and 45.00, respectively) while TPGY 100, 200, 250 and TPG 50, 150 revealed the high results concerning HOST (80.61, 83.99, 80.61, 88.33 and 83.33, respectively) whereas the highest alive sperm percentage was recorded with TPGY 150 (96.30) compared to the control. Conclusions The aqueous cold infusion or extract of the DPPG, added to the TCF extender with or without the addition of egg yolk, proved its good preserving and maintaining capacity of chilled and thawed buffalo bull sperms which was expressed mainly by the sperm motility.

263-265

Authors: Achour Radhouane, Neji Khaled

Corpus callosum agenesis (CCA) was evaluated by ultrasound examination and magnetic resonance imaging (MRI) with many studies. Ultrasonography was able to suspect CCA by indirect signs but a definitive diagnosis of CCA was achieved in rare cases. MRI was able to diagnose complete CCA in majority of cases. Additional neurological abnormalities including heterotopia, gyration anomaly, asymmetry of the cerebral hemispheres, and Dandy-Walker variant were documented, as well as an ocular anomaly which was described, by MRI examination. Prenatal counseling for fetal agenesis of the corpus callosum is difficult as the prognosis is uncertain. The association with other cerebral abnormalities increases the likelihood of a poor outcome and ultrasonographic assessment of the fetal brain is limited. We found MRI to be a safe and useful additional procedure to complement ultrasonographic diagnosis or suspicion of CCA.

269

Authors: Beuy Joob, Viroj Wiwanitkit

no

96-101

Authors: B.O. Omontese, P.I. Rekwot, I.U. Ate, J.O. Ayo, M.U. Kawu, J.S. Rwuaan, A.I. Nwannenna, R.A. Mustapha, A.A. Bello

This review is aimed at providing information on current status of oestrus synchronization as a management tool in goats in Nigeria. Oestrus synchronisation is a reproductive tool that enables goat farmers breed their animals within a short pre-determined period. The principle of oestrus synchronisation is chiefly the control of the luteal phase of the oestrous cycle and two basic mechanisms are employed. These include the use of prostaglandins or its analogues to shorten luteal life/induce premature luteolysis or the use of exogenous progesterone to prolong luteal life, thereby simulating the activity of natural progesterone produced by the corpus luteum. The latter is advantageous where the reproductive status of the flock is unknown. However, the former is easy to apply and only effective in cycling animals. Pharmaceutical products that have been employed in ES protocols in Nigeria include Lutalyse®, Estrumate®, EstroPLAN®, Fluorogestone acetate®, Sil-Oestrus®, Medroxyl-progesterone acetate®, Synchromate-B®, PMSG®, and more recently, the Controlled Internal Drug Release® (CIDR) devices. Over the last three decades, many attempts have been made at assessing the response of goats to various oestrus synchronisation agents and protocols in Nigeria. However, the low availability of these pharmaceutical agents and cost preclude the widespread use of oestrus synchronisation technique in goat production. The inclusion of gonadotrophins in oestrus synchronisation protocols have been reported to improve oestrus responses in goats, while season has been reported to influence the efficiency of oestrus synchronisation programmes. Increasing the dose levels of exogenous hormones in oestrus synchronisation protocols has caused variable and sometimes inconsistent effects. The effects of combining the male stimulus with exogenous hormones, seasonal variation, extra-label use of products and short-term nutritional manipulation on oestrus responses in goats require further evaluation. It is concluded adoption of oestrus synchronisation practice portends enhanced goat production in Nigeria.

105-110

Authors: I. Onyeso, K.W. Nkpaa, S. Omenihu

Objective To investigate the effect of Citrillus lanatus (C. lanatus) seeds and caffeine on blood glucose levels and testicular functions of alloxan-induced diabetic male Wistar rats. Methods Alloxan was administered at a single dose of 150 mg/kg BW to induce diabetes. A dose of either 200 mg/kg C. lanatus or 100 mg/kg caffeine or both was administered daily to alloxan-induced diabetic rats for three weeks, after which results were compared with a normal control group and a positive control group that received both alloxan and glybenclamide. Results C. lanatus seeds extract significantly decreases (P < 0.05) blood glucose level and significantly (P < 0.05) increased sperm motility, sperm count, normal sperm morphology, sperm viable cells and testosterone in plasma level of alloxan-induced diabetic rats treated with C. lanatus seed extract and caffeine. Conclusions The present study showed that co-administration of caffeine and hydromethanolic fraction of C. lanatus seed extract have hypoglycemic effect and may consequently ameliorate the impaired testicular general architecture and inhibits sperm death or testicular damage caused by alloxan-induced diabetes.

116-122

Authors: P. Sushma Reddy, Nazia Begum, Sumith Mutha, Vasudha Bakshi

Objective To investigated beneficial effect of Curcumin (a phenolic curcuminoid derivative from Curcuma longa) in Letrozole induced PCOS in female Wistar rats. Methods Letrozole (1 mg/kg) was administered per orally (p.o) for a period of 21 days for the induction of PCOS, followed by dose of Curcumin (100 mg/kg and 200 mg/kg, p.o) for 15 days using 0.5% w/v CMC as vehicle. Results The administration of Letrozole led to abnormalcy in serum sex steroid profile, lipid profile, glucose and glycosylated hemoglobin levels and depletion in antioxidant activity. Curcumin was able to successfully exert its protective effect by restoring all the parameters to normal and disappearance of cysts in ovaries. Conclusion Curcumin showed beneficial effects in Letrozole induced PCOS in female Wistar rats. Its effect was comparable to that of Clomiphene citrate, most widely used treatment for ovulation induction in PCOS condition.

128-131

Authors: Sneha Panchal, Ramtej J. Verma

Objective To determine the toxic effect of diethanolamine on testicular steroidogenesis and its possible amelioration by curcumin in mice. Methods Adult Swiss strain male albino mice were orally administered with diethanolamine (110, 165 and 330 mg/kg body weight) for 45 days. In another set of experiment, male albino mice were orally administered with curcumin (10, 25 and 50 mg/kg body weight) along with diethanolamine (330 mg/kg body weight) for 45 days. On completion of treatment, animals were humanely sacrificed, testis dissected out, blotted free of blood and used for biochemical analysis. The obtained serum samples were analyzed for serum testosterone level. Results Diethanolamine caused significant (P < 0.05), dose-dependent reduction in total lipid and cholesterol contents, activities of 3β- and 17β-hydroxysteroid dehydrogenases in testis as well as significant reduction in serum testosterone level. However, co-treatment of curcumin along with diethanolamine caused significant (P < 0.05) dose-dependent amelioration in diethanolamine-induced changes in total lipid and cholesterol contents, activities of 3β- and 17β-hydroxysteroid dehydrogenases and serum testosterone levels. Conclusions The results of the present study showed that curcumin significantly ameliorates diethanolamine reduced testicular steroidogenesis.

140-143

Authors: Afifi Inayah, Sri Rahayu, Nashi Widodo, Widya Ayu Prasdini

Objective To determine the polymorphism of GDF-9 gene in FH cattle that may affect its interaction with the BMP-15 protein. Method Blood was taken from the jugular vein using 10 mL sterile syringes from ten Friesian Holstein cattle. The DNA were isolated from the whole blood and used as a template to amplify GDF-9 gene. The Amplicon (PCR product) was sequenced to identify the new SNP. The three-dimensional structure of GDF-9 protein was modeled by SWISS-MODEL. The characteristic of the protein structure was analyzed by using projectHOPE. The binding affinity GDF-9 into MBP was examined by PatchDock and FireDock. Results The result indicates a new variant G1358A changed amino acid residue at position 435 from Arginine into Histidine (R453H). In silico analysis using projectHOPE predicted that the variant altered side chain of GDF-9 and changed its interaction with BMP-15. Further study suggested that the polymorphism R453H also reduce the binding affinity of the GDF-9 into BMP-15 from −85.58 kcal/mol (R453) into −80.79 kcal/mol (H453). Conclusion The new variant G1358A at an estrous FH a cattle has potency alter GDF-9 profile that may affect to reproduction performance.

148-155

Authors: K.H. El-Shahat, M.I. Taysser, M.R. Badr, K.A. Zaki

Objective To determine the effect of different concentrations of heparin, caffeine or calcium ionophore and incubation time on motility, hyperactivity (HA) and acrosome reaction (AR) of ram sperm in vitro. Methods Semen samples were collected by artificial vagina from three mature ram. Split fractions (0.1 mL) of the pooled semen were layered under 1 mL of S-TALP medium supplemented with either heparin (10, 25, 50, 75 and 100 μg/mL), caffeine (0.5, 0.75, 1, 2 mg/mL), or calcium ionophore A23187 (0.5, 0.75, 1, 1.55 mM/mL). Individual motility, hyperactivity percentage and acrosomal status were recorded at 0, 1, 2, 3 and 4 h post-incubation for all treatments and control. Moreover, they were examined for ability to fertilize sheep oocyte in vitro. Results Heparin, caffeine and calcium ionophore A23187 at a concentration of 75 μg/mL, 1 mg/mL, and 1.55 mM/mL respectively can be used as a protocol to provide the best results for in vitro cap citation and acrosome reaction in ram. The penetration rates of rate of oocytes inseminated with spermatozoa treated with calcium ionophore A23187, and heparin were higher as compared with caffeine. Moreover, heparin achieved higher fertilization rates but without significant difference with others. Conclusion The best concentration of heparin, caffeine and ionophore A23187 are 75 μg/mL, 1 mg/mL, 1.55 mM/mL for 3, 1, 4 h incubation respectively and can be used for in vitro fertilization in sheep.

161-170

Authors: Loganathan Kousalya, V. Narmatha Bai

Objective An efficient in vitro plant regeneration protocol for Canscora decussata Schult. (Gentianaceae) (C. decussate), a threatened medicinal herb used in Ayurvedic system of medicine was developed. Seed germination was achieved on MS growth regulator free medium. Methods The nodal explants were excised from the in vitro raised seedlings and inoculated on MS medium supplemented with various plant growth regulators such as BAP, KIN, TDZ and Zeatin individually and in combinations with or without GA3. BAP (2 mg/L) was proved to be effective for multiple shoot induction (30.20 ± 6.53) among the cytokinin tested individually. Addition of NAA (1 mg/L) to cytokinin containing medium resulted in callus, KIN (3 mg/L) with NAA (1 mg/L) produced highest percentage of callus (82%) per explant. Results Among the various combination of cytokinin tested, BAP (0.5 mg/L) in combination with KIN (2 mg/L) induced highest number of multiple shoots (72.10 ± 1.05 shoot per explant). Addition of 1 mg/L of GA3 to the above medium induced highest number of shoots (100.80 ± 3.20) with an average shoot length of 6.98 ± 0.66 cm. Rooting was optimized in half-strength MS medium supplemented with IBA at 1.0 mg/L. The plantlets were successfully transferred to hardening medium containing vermiculite with 83% survival rate. Among the antioxidant activity of methanol extract of wild-grown plants and in vitro regenerants tested, half-MS medium supplemented with NAA (0.5 mg/L) derived callus has promising activity for total phenolics, DPPH, ABTS, FRAP and phosphomolymbdenum assays. Total flavonoid content was found to be high in callus derived from MS medium supplemented with KIN (2 mg/L) in combination with NAA (1 mg/L). Conclusion Our present study suggest that in vitro derived callus of C. decussata represent a promising alternative source to meet the pharmaceutical demands for commercial formulations and the protocol could effectively be applied for the conservation of C. decussata Schult.

10-13

Authors: Jafar Nouri Nojadeh, Hossein Daghghagh

In recent years, it has become consumedly clear that changing of epigenetic modification is essential during both early and late oogenesis and spermatogenesis. Also epigenetic modifications are involved in some cases such as embryo development and growth, diseases and responsible for X-chromosome inactivation and genomic imprinting. Epigenetic reprogramming can be explained as any mitotic or meiotic changing which does not result any alteration in DNA sequence but will have important effect on the normal embryonic development. Germline epigenetic reprogramming in addition to requiring epigenetic modification to compose the germline, the primordial germ cells uniquely undergo striking wave of epigenetic reprogramming that most other lineage do not undergo. Epigenetic modification is affected by both internal factors and environmental factors during pre- and post-natal development. Because all of the epigenetic modification steps are not clear, by means of understanding epigenetic modification, misreprogramming of these steps can be modified with the aid of drugs and nutrients. Moreover, epigenetic regulation is essential to obtain the biological intricacy of multicellular organisms, cloning and producing of offspring by assisted reproductive technology (ART). The objective of this review is to provide comprehensive summary of the current knowledge in the field of epigenetic modification in relation to male and female germline development and reproduction.

24-30

Authors: Ketaki R. Desai, Nilofer Moid, Pragnesh B. Patel, Hyacinth N. Highland

Objective To assess the adverse effect of Deltamethrin (DM) (technical grade) on reproductive organs and fertility indices of male Swiss albino mice, Mus musculus. Methods Forty male mice were divided into four experimental groups: control, vehicle control (peanut oil), high dose DM treated and low dose DM treated group. An oral dose of 3 mg/kg b.wt (low dose) and 6 mg/kg b.wt (high dose) of Deltamethrin was administered for a period of 45 days to male Swiss albino mice. Results DM caused a significant reduction in body and organ weights, sperm count, sperm motility percent, sperm viability, serum testosterone level, sialic acid content of cauda epididymis and fructose level of seminal vesicle. DM-treated groups also showed a significant decline in testicular 3β and 17β Hydroxysteroid Dehydrogenase (HSD) activities. Histological examinations revealed significant alterations in the testes of dosed groups. Conclusion Deltamethrin is a toxic pyrethroid pesticide that produced significant reproductive toxicity in treated male mice as revealed by the severely affected parameters and the altered gravimetric indices.

36-41

Authors: A. Lalithamma, A. Govardhan Naik, C. Changamma

Objective To study the histomorphological changes with reference to estradiol valerate administration in aged female albino rats. Methods The observations showed in young, aged and aged administered with estradiol valerate (Progynova tablets) at the dose of 2 mg/day through oral gavages for one week. The histomorphological changes were observed in ovary, uterus, vagina and liver. Results Estradiol velerate administration improves the ovarian stroma and germinal epithelium. The vascular hyaline degeneration reduces. Vascular protective effects of estrogen are well established. The administration may be associated with endometrial hyperplasia and neoplasia. The stroma was comparatively less edematous with less active endometrial glands. The uteri exhibited endometrial epithelial cell pseudo stratification and hypertrophy. The vaginal histoarchitecture showed increased epithelial proliferation of the vaginal wall and improving thickening. Estradiol velerate does not showed any effect on liver architecture. Conclusions The administration of estradiol valerate showed its vascular protective effects. It does not showed any effect on liver architecture.

46-50

Authors: Ehsanollah Sakhaee, Ladan Emadi, Hamidreza Siahkouhi

Objective To evaluate the supportive effects of Rosa damascene (R. damascene) essential oil on epididymal sperm quality and histopathology of testes following long term administration of copper sulfate in mice. Materials and methods The study comprised of four different groups of six mice as follows: group Cu, which received 0.1 mL copper sulfate solution at dose of 100 mg/kg, group R which received 0.1 mL R. damascene essential oil at dose of 1 mg/kg, treatment group (T) which received copper sulfate solution (100 mg/kg) and treated by R. damascene essential oil (1 mg/kg), and control group (C) which received the same volume of normal saline. The supplements were gavaged in all animals every other day, during experimental period. All animals of each experimental group were sacrificed 42 days after the beginning of experiment. Results Results showed that sperm concentration, motility and viability in group Cu were significantly decreased after 6 weeks, and severe degenerative changes were observed in testicular tissues in comparison with the control group. In treatment group, significant improve in the sperm count, motility and viability, and normal architecture in most seminiferous tubules with organized epithelium was observed compared to the group Cu. Conclusions Administration of the essential oil of R. damascene owning to its antioxidant properties is able to protect the testis and epididymal sperm from the adverse effects of copper poisoning in mice.

58-62

Authors: Angelica Garcia-Olivares, Cesar Garzon-Perez, Oscar Gutierrez-Perez, Alfredo Medrano

Objective To evaluate the effect of skim milk and/or coconut milk in a glycerol-free trehalose extender to improve cryosurvival of boar spermatozoa. Methods Sperm samples were diluted in an egg-yolk-based freezing extender containing 100 mM trehalose and 0.25% Equex STM supplemented with coconut milk or/and skim milk at 2% or 5% (w/v). Spermatozoa were cryopreserved in 0.5 mL straws and thawed by a rapid transient method (at 70 °C for 8 s) followed with a stabilizing procedure at 39 °C. Thawed samples were analyzed for motility, viability, high mitochondrial membrane potential (HMMP), and acrosome damage. Results Even on the presence of egg yolk, motility, HMMP and viability were significantly higher in extender supplemented with 2% skim milk than controls without skim milk (P < 0.05). Post-thaw viability significantly improved with the addition of 2% skim milk plus 2% coconut milk as well (P < 0.05). Acrosome damage was considerably lower when the extender was supplemented with 2% coconut milk (P < 0.05), whereas the benefit was masked in the presence of 2% skim milk. Conclusion 2% skim milk can be used as supplements for a glycerol-free trehalose and egg yolk-based extender to improve post-thaw survival of boar spermatozoa, whereas 2% coconut milk has an effect to protect boar spermatozoa from acrosome damage.

67-70

Authors: F.A. Lone, A.A. Malik, A. Khatun, M. Shabir, R. Islam

Objective To evaluate the return of ovarian cyclicity with four hormone based protocols during non-breeding season in infertile Corriedale sheep. Methods The return to ovarian cyclicity was considered in terms of percent estrus response rate (ERR). The time of exhibition of estrus after either sponge removal or last PGF2α injection, was considered as time to onset of estrus. Infertile Corriedale ewes were randomly selected and distributed into four groups corresponding to four hormonal protocols such as PsE (intravaginal progesterone sponges for 12 days and eCG (equine chorionic gonadotropin) at 24 h before sponge removal; n = 6), GP (GnRH on day 0 and PGF2α on day 5; n = 7), GPG (GnRH on day 0, PGF2α on day 5 and second injection of GnRH on day 7; n = 7) and PsPG (Intravaginal progesterone sponges for 12 days, PGF2α and gonadotropin-releasing hormone (GnRH) respectively, at 24 h before and after sponge removal; n = 8). Results PsPG protocol produced significantly (P < 0.05) higher ERR (87.5%) in ewes as compared to GPG (28.57%) but non-significantly (P > 0.05) higher than GP (85.71%) and PsE (66%). Estrus was compact and more synchronized in PsPG group because 75% of ewes exhibited estrus during the first 48 h. The collective incidence of estrus (78.94%) was also maximum during the first 48 h. Conclusion PsPG estrus induction strategy has a potential to replace eCG based protocols for returning of ovarian cyclicity in infertile Corriedale sheep.

75-79

Authors: Modesta Mitao, Rune Philemon, Joseph Obure, Blandina T. Mmbaga, Sia Msuya, Michael J. Mahande

Objective To determine the risk factors for low birth weight and adverse perinatal outcomes associated with low birth weight in Northern Tanzania. Methods A retrospective cohort study was designed using maternally linked data from Kilimanjaro Christian Medical Centre (KCMC) medical birth registry. A total of 37 799 singleton births delivered from 2000 to 2013 were analyzed. Multiple births, stillbirth and infants with birth defects were excluded. Data analysis was performed using SPSS version 16.0. Chi-square was used to compare difference in proportions between groups. The relative risks (RR) with 95% confidence interval (CIs) for the factors and adverse perinatal outcomes associated with LBW were estimated in a multivariate logistic regression models. Results The incidence of low birth weight was 10.6%. Multivariate logistic regression showed that pre-eclampsia (RR 3.9; 95% CI 3.6–4.2), eclampsia (RR 5.4; 95% CI 4.1–6.9), chronic hypertension (RR 2.8; 95% CI 2.1–3.8), maternal anemia (RR 1.7; 95% CI 1.4–2.2), HIV status (RR 0.8; 95% CI 0.7–0.8), smoking during pregnancy (RR 1.9; 95% CI 1.0–3.5), caesarean section delivery (RR 1.4; 95% CI 1.3–1.5), placental abruption (RR 3.7; 95% CI 1.3–4.7), placenta previa (RR 6.6; 95% CI 4.8–9.3), PROM (RR 2.5; 95% CI 1.9–3.3), maternal underweight (RR 1.3; 95% CI 1.2–1.6), and obesity (RR 1.2; 95% CI 1.1–1.4) and female gender of baby were significantly associated with delivery of low birth weight infants. On the other hand, LBW infants had increased risk of neonatal jaundice (RR 2.7; 95% CI 1.2–6.1), being delivered preterm (RR 2.0; 95% CI 1.8–2.3), Apgar score (<7) at fifth minute (RR 5.5; 95% CI 4.5–6.6) and early neonatal death (RR 3.5; 95% CI 2.6–4.6). Conclusions Low birth weight is associated with adverse perinatal outcomes. Early identification of risk factors for low birth weight through prenatal surveillance of high risk pregnant women may help to prevent these adverse perinatal outcomes.

84-86

Authors: Achour Radhouane, Bouriel Ines, Neji Khaled

The majority of cases of placenta accreta are unanticipated and initially identified intraoperatively. Although color Doppler ultrasound is adequate for the evaluation of placenta accreta in the third trimester, ultrasound diagnosis in the first trimester has never been reported. To our knowledge, this is the first case of placenta accreta detected at 9 weeks' gestation by ultrasound. Placenta accreta with intraplacental lacunae can be identified together with a loss of the hypoechogenic retroplacental myometrial zone. Based on this case, we found that early diagnosis of placenta accreta in the first trimester by ultrasound is possible.

Pages 91-95

Authors: V Vengaiah, A Govardhan Naik, C Changamma

Objective To know the impact of Piper betel leaf stalk (P. betel) extract on Protein and energy metabolism and its role in male albino rats. Methods Healthy adult (3-4 months old) male Wistar strain albino rats were administered with betel leaf stalk extract, at the dose of 50 mg/kg/day through oral gavages for 15 days. Twenty four hours after the last dose, the animals were autopsied. In order to assess antifertility effect in testis, epididymis, seminal vesicle and prostate gland, estimation of total, soluble and structural proteins, free amino acids and DNA, RNA were undertaken. Results The accumulation in proteins indicates the anti-androgenic effect of extract. The reduction in free amino acids will affect the sertoli cell function, results in the damage of spermatogenesis. The significant elevation in testicular DNA content (hyperplasia) was observed. In the present study, P. betel leaf stalk extract decreases the concentration of RNA, in testes, seminal vesicle and prostate gland except in epididymis where it was elevated. It indicates the alterations in rate of protein synthesis and growth rate of tissues due to the administration of P. betel leaf stalk extraction. However, the RNA: DNA ratio was reduced except in prostate. Conclusions P. betel leaf stalk extract exert its anti androgenic effect by alterations in rate of protein synthesis and cellular hypertrophy occur in prostate.

Pages 100-105

Authors: Pallavi Sharma, Manjusha, Sudesh Rani, Hitesh Malhotra, Nitesh, Sandeep Deswal, Surender Singh

Objective To assess antifertility activity of hydroalcoholic extract of Cordia dichotoma G Forst. (C. dichotoma) leaves, which are traditionally used to produce sterility among the tribal women through its abortificient activity. Methods Acute toxicity study was carried out in adult albino rats. The antifertility activity of extract at two dose level (200 and 400 mg/kg, orally) was evaluated in two experimental animal models i.e. anti-implantation and estrogenic/antiestrogenic activity in female rats by observing number of implants, vaginal cornification, body weight, uterus weight and biochemical investigation. Results The extract was found to be safe up to dose of 2 000 mg/kg body weight when administered orally. A good anti-implantation (81.22%) activity in female rats was observed at the tested dose levels (200 and 400 mg/kg, orally). The extract further showed more significant (P<0.01) increase in uterine weight and significant change in biochemical parameters in immature rats. Simultaneous administration of extract along with ethinyl estradiol showed significant estrogenic activity. Conclusions The results suggest that hydro alcoholic extract of C. dichotoma leaves possess significant antifertility activity, which is consistent with the literature report in folk medicine of this plant in fertility regulation.

Pages 112-115

Authors: Deepika Jaiswal, Sameer Trivedi, Neeraj K Agrawal, Kiran Singh

Objective To investigate –844C>T single nucleotide polymorphism (SNP) present in the promoter of cell death pathway gene FASLG with male infertile phenotype. Methods Genotyping for SNP FASLG (rs763110) was done by polymerase chain reaction followed by analysis with specific endonuclease (PCR-RFLP). DNA sequencing was used to ascertain PCR-RFLP results. Results FASLG –844C>T polymorphism, allele and genotype distribution did not differ significantly between patients and controls (OR: 1.03, 95% CI= 0.7638 to 1.3952, P=0.83). Thus SNP-844C>T of the FASLG gene is not associated with male infertility risk in the analyzed patients. Conclusions Human male infertility is a complex disorder and thus other genetic or environmental factors may be contributing to the complex etiology, and further study in other region of Indian populations will verify whether it is associated with male infertility risk.

Pages 123-128

Authors: Soundarrajan Ilavenil, Dhanaraj Karthik, Mariadhas Valan Arasu, Mayakrishnan Vijayakumar, Srisesharam Srigopalram, Selvaraj Arokiyaraj, Sivanesan Ravikumar, Ki Choon

Objective To investigate the differential of protein expression in CCl4 induced mice treated with lycorine. Methods The present study was carried out to identify the differentially expressed protein in carbon tetrachloride (CCl4) induced oxidative stress mice treated with lycorine (5 mg/kg. bw) using 2D gel and MALDI-TOF. Results We observed many kinds of differentially expressed protein in experimental liver. Among these, three are much differently expressed protein which is identified as ATP synthase, regucalcin and HSP60; these proteins are involved in the ATP synthesis, calcium regulation and rescue the integrity cellular proteins respectively. Conclusions This investigation provided a molecular mechanism of the lycorine during CCl4 induced oxidative stress in mice liver.

Pages 135-139

Authors: Baskaran Xavier Ravi, Geo Vigila Antony Varuvel, Rajan kilimas, Jeyachandran Robert

Objective To develop an efficient reproducible protocol for a critically endangered fern, Pteris tripartita (P. tripartita) Sw. through spore culture. Methods The spore derived gametophytes were grown in a half strength MS medium which sub-cultured after three months to MS medium containing growth regulators like BAP, Kinetin and GA3 with 3% sucrose. Results The highest mean number of sporophytes (64.40) induced with 4.16 cm of length and also formed 1.51 cm of root length at 4 mg/L of BAP. On MS medium supplemented with KIN, 56.03 mean numbers of sporophytes with 2.61 cm along with 1.39 cm length of root were obtained in 4 mg/L of KIN. And, the highest mean numbers of juvenile sporophytes (59.66) were induced with 2.64 cm mean lengths of sporophytes with 2.23 cm of root length in 4 mg/L of GA3 in the MS culture medium. The in vitro developed plantlets were washed with distilled water and grown in small cups containing sterilized soil, sand and vermiculite (1:1:1). The acclimatized plantlet showed 90% of a survival rate. Conclusions In this study, reproducible protocol could be useful for their rehabilitation and also to reduce the pressure on natural population in Asian habitat.

Pages 147-152

Authors: Sunghyun Hong, Kandhasamy Sowndhararajan, Taewoo Joo, Chanmook Lim, Haeme Cho, Songmun Kim, Gur-Yoo Kim, Jin-Woo Jhoo

Objective To determine the gene expression of antioxidant enzymes by hemp seed extracts in human hepatoma (HepG2) cells. Methods Ethanol and supercritical fluid (SF) extracts obtained from de-hulled hemp seed were used for the evaluation of in vitro antioxidant activity and gene expression of antioxidant enzymes. In vitro antioxidant activities of the samples evaluated using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) diammonium salt (ABTS) radical scavenging assays. The expression of antioxidant enzymes such as superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) in HepG2 cells was evaluated by real-time PCR. Results In the antioxidant assay, SF extract of hemp seed exhibited higher ABTS and DPPH radical scavenging activities (IC50 of 66.6 µg/mL and 9.2 mg/mL, respectively) than ethanol extract. The results of antioxidant enzyme expression in real-time PCR study revealed the H2O2 (200 µM) challenged HepG2 cells reduced the expression of enzymes such as SOD, GPx and CAT. However, the cells treated with ethanol and SF extracts were up-regulated the expression of antioxidant enzymes in concentration dependent manner. When compared to ethanol extract, the SF extract exhibited higher activity in the expression of all the antioxidant enzymes at the concentration of 500 µg/mL. Conclusions In conclusion, the findings of our study demonstrated that the hemp seed effectively inhibited H2O2 mediated oxidative stress and may be useful as a therapeutic agent in preventing oxidative stress mediated diseases.

Pages 157-165

Authors: J. Simões

Abstract Goats show marked progressive seasonal reproduction at latitude > 25° and reproductive management should be adapted to market demands. The present review aimed to discuss the synchronization of ovulation for timed artificial insemination concerning new insights regarding a clean, green and ethical meat and milk production. Today, the induction of ovulation during breeding season or transitions periods is mainly based on progestagens/ progesterone (P4) devices intravaginally inserted on females, at least during 11 days, plus equine chorionic gonadotropin (eCG) and prostaglandin F2 alfa administration. In last years a reduction to 20 mg of fluorogestone acetate was made and the successful reutilization of devices containing 0.3 g of P4 indicates a possible reduction of their levels. Shortening the period of exogenous progestagens/P4 priming (5 to 7 days) is critical for a rational use of hormones. Moreover, the eCG exchange by socio-sexual cues (male effect) seems to see a great advance, even if a previous photoperiod treatment, or equivalent method, being necessary in high latitudes. Research trends on these subjects are expected in future using different goats breeds in distinct regions of world.