We have recently shown that sperm attachment to bovine endometrial epithelial cells (BEECs) triggers uterine local innate immunity with induction of a pro-inflammatory response in vitro , however details of the mechanism remain unknown. Here, we investigated the involvement of Toll-like receptor 2/4 (TLR2/4) pathway in mediating sperm-BEECs inflammatory process. Immunohistochemistry of the uterine tissue revealed that TLR2 and TLR4 proteins were present in the luminal and glandular epithelia of bovine endometrium. Moreover, BEECs monolayers were treated with TLR2 agonist (Pam; 0, 10, 100, and 1000 ng/ml) or TLR4 agonist (LPS; 0, 0.1, 1, and 10 ng/ml) for 0, 1, 3, or 6 h, followed by evaluating mRNA expression of the pro-inflammatory genes ( TNFA , IL-1B , IL-8 , and PGES ) in BEECs using a real-time PCR. Both Pam and LPS treatments showed a dose-dependent stimulation of mRNA expression of the pro-inflammatory genes. To elucidate the functional role of TLR2/4 in sperm-BEECs interaction, BEECs monolayers were incubated with either TLR2 antagonist or TLR4 antibody for 2 h prior to the co-culture with sperm for 3 h. Importantly, pre-incubation of BEECs with TLR2 antagonist or TLR4 antibody prevented the stimulatory effect of sperm on the transcription of pro-inflammatory genes in BEECs. Furthermore, sperm increased the phosphorylation levels of TLR2/4 downstream targets (p38MAPK and JNK) in BEECs within 1 h of the co-culture. Treatment of BEECs with TLR2 antagonist prior to sperm addition inhibited JNK phosphorylation, while TLR4 antibody inhibited the phosphorylation of both p38MAPK and JNK. In conclusion, the present in vitro findings strongly suggest that bovine endometrial epithelial cells respond to sperm via TLR2/4 signal transduction.

Retained fetal membrane (RFM) is a crucial calving related disorder that affects economic for the farmer and herd health. Retention of Placenta (ROP) is the condition in which the fetal membranes are not expelled within a period of 12 hours after parturition. Buffalo cows (n=34) aged 4-9 years old with 2-6 lactation season were used in the present study. The Experimental buffaloes were divided into either with (n=25) or without (n=9) fetal membrane retention. The objective of this study was to evaluate the effect of oxytetracycline treatment on reproductive performance of buffalo-cows with retained fetal membranes. All buffaloes in treated group received systemic infusion of oxytetracyclin Hcl injection 5% and 10 mg/kg oxytetracycline for 3 days intrauterine. Reproductive performance parameters for all experimental buffaloes were days to first estrous, number of service per conception, days open and pregnancy diagnosis. All reproductive parameters demonstrated significant changes between treated and non-treated animals (P < 0.05). Altogether, the obtained results indicated that retained placenta in buffaloes could be treated successfully by intrauterine infusion of oxytetracycline

The aim of this study was to investigate the interactive and main effects of parity numbers, parity seasonand parity month on postpartum fertility parameters [ovarian rebound (OR), days open (DO) and number of services per conception (S/C

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Key words: D-Penecillamine, Hypotaurine, Epinephrine, In vitro fertilization, BuffaloIn vitro fertilization (IVF) is a necessary procedure in the in vitro production of embryo. Spermatozoa need to undergo capacitation to be able to perform the acrosome reaction which is required for zona pellucida penetration. Different additives are used to aid in that process, such as caffeine and theophylline. In this study, the effects of IVF media supplementation with D-penicillamine, Hypotaurine and epinephrine (PHE) on fertilization and cleavage rates were assessed. Cumulus oocyte complexes (COCs) containing a compact cumulus cells and normal homogenous cytoplasm were selected. In vitro maturation (IVM) was performed in TCM199 supplied with hormones overnight in a culture conditions of 38.5°C and 5% CO 2 in air. After 20-22 hr. of maturation, oocytes were randomly assigned to five groups of sperm-TALP media (control and four different PHE concentrations) then inseminated. The fertilization rate of the expected embryos were assessed after 22 hr. after staining with aceto-orcin 1% or cultured for 48 hr. to estimate the cleavage rate. The current data revealed that, 25 µl/ml PHE significantly (P< 0.05) improved fertilization (55.93 vs 36.96%) and cleavage (46.15 vs 22.22%) rates when compared to control.

The changes in the expression profile of the angiogenicgenes and serum biomarkers were investigated in the cows with the retained placenta after normal parturition. Retained fetal membranes (RFM) are considered one of the main reproductive disorders in dairy cattle. Highlight the effects of Excede® (Ceftiofur Crystalline Free Acid: Zoetis, USA) in combination with Oxytocin (Oxytocin®, each ml contains 10 IU oxytocin, ADWIA, Egypt), Estrumate® (250µg cloprostenol: synthetic analogue of PGF2ά) and Flunixin Meglumine® (nonsteroidal anti-inflammatory drug) on the reproductive performance in dairy cows. A total of 16 cows were allocated into two groups: group 1 (n = 8) the cows that have a normal loosed placenta, and group 2 (n = 8). The cows that have retained fetal membranes for more than 24 h after parturition. Blood with EDTA was collected from the tail vein for studying the expression of angiogenic proteins including vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor1 (VEGFR1), and RANTES genes. Serum was collected at a one-week interval before/after parturition for the study of biochemical changes. The VEGF expression decreased ten folds in cows with retained than normally dropped placentae (p ˂ 0.0001). The vascular endothelial growth factor receptor2 (VEGFR2) showed a threefold increase in cows with normal loosed than retained placentae (p˂0.01). The RANTES was increased in cows with retained rather than normal placentae by about 7 folds (p˂0.01). Phosphorus and total proteins were significantly lower (p˂0.05) in RFM cows one week after parturition. The serum albumin, TG, HDL, and cholesterol were significantly lower (p˂0.05) in RFM before, at, and after parturition. CPK was significantly higher in RFM before, at, and after parturition (p˂0.05). Increased values of MDA and decreased values of CAT, SOD, and R-GSH were detected in the blood of cows suffering from RFM (p˂0.05). Reproductive performance parameters for all studied cows were days to first estrous, number of service per conception and pregnancy rate. All reproductive parameters demonstrated significant (P < 0.05) changes between treated animals. The translation of VEGF and its receptor and RANTESmRNA at the time of parturition in dairy cows could be proxy biomarkers for the prediction of retained fetal membranes (RFM). In the same line, the activity of total proteins, serum albumin, TG, HDL, cholesterol, and CPK would be useful in the prediction of the RFM in dairy cows. Altogether, the obtained results indicated that retained fetal membranes in cows could be treated successfully without any interference by systemic injection of cephalosporin with caring of temperature. The reproductive performance criteria for all cows studied were the time of first estrus, the number of services per conception, and pregnancy rates. All parameters of reproductive performance showed significant changes (P < 0.05) among the treated animals. The results also indicated that cephalosporin can be used by systemic injection to treat placental retention with caring of the animal's temperature.

Aims: We investigated the effect of intraperitoneal titanium dioxide nanoparticles (TiO2 NPs, 100 mg/kg) for 5 consecutive days on the developmental competence of murine oocytes via its effects on the antioxidant and oxidative stress biomarkers as well as their effects on mRNAs expression for BAX, Caspase 3, P53 and hypoxia inducing factor-1α (HIF- 1). Moreover, the possible ameliorating effect of fructose (2.75 mM/ml) was examined. Materials and methods: Thirty sexually mature (8-12 weeks old; ~ 25 g body weight) female mice were used for the current study. The female mice were divided into three groups as follows: Group1 (G1): Female mice received deionized water and the collected oocytes were cultured in global® total media (control group). Group 2 (G2): Female mice received TiO2 NPs at a dose of 100 mg/kg BW by i/p injection for 5 consecutive days, and the collected oocytes were cultured in global® total media. Group 3 (G3): Female mice received TiO2 NPs at a dose of 100 mg/kg BW + fructose 2.75 mM by i/p injection for 5 consecutive days, and the collected oocytes were cultured in global® total media. Results: Nano-titanium significantly decreased GSH, GPx, and NO while MDA and TAC were increased. The rate of MI, MII, GVBD and degenerated oocytes were significantly decreased. Meanwhile, the rate of activated oocytes was significantly increased. TiO2 NPs significantly upregulated the relative expression of apoptotic genes (BAX, Caspase 3 and P53) and HIF-1. Intraperitoneal fructose (2.75 mM/kg) significantly alleviated the detrimental effects of TiO2 NPs. The transmission electron microscopy showed the ability of the fructose to maintain the surface of murine oocytes. Conclusion: Collectively, these results suggest that the i/p infusion of fructose for consecutive 5 days promote the oocyte development and decrease the toxic effects of TiO2 NPs by changing the oxidative and antioxidant biomarkers secreted by COCs and affects the mRNAs expression of apoptotic and hypoxia inducing factor.

In the current, study the effect of the addition of L-carnitine (LC) in in vitro maturation (IVM) medium for buffalo oocytes and different cryo-devices on developmental competence. They were matured in IVM medium supplemented with 0.0, 0.3, 0.6 and 1.2 mg/mL of LC and vitrified by using either straw (ST), open pulled-straw (OPS), solid surface vitrification (SSV). The effects of LC during invitro fertilization and invitro culture on the developmental potential were examined. ST showed a higher recovery rate when using 0.6 mg/ml; viability rate (VR) when using 0.3mg/ml and lower zona pellucida (ZP) and cytoplasmic abnormalities when using 1.2 mg/ml LC (P<0.05). OPS showed a higher recovery and viability rates when using 0.3 mg (P<0.05). SSD showed a higher recovery and VR when using 0.6 mg/ml LC (P<0.05). Maturation and fertilization rates were increased when using 0.3 mg LC and OPS (P<0.05). Cleavage was increased in 0.3 (OPS) and 0.6 mg LC (SSD). The blastocyst rate was increased in 0.3 (OPS) and 0.6 mg/ml (SSD) (P< 0.05). In conclusion, 0.3 mg/ml LC and OPS gives a higher recovery and viability rates but 0.6 mg/ml LC and of both SSD and ST gives a higher recovery and viability rates.