Przemysław Gilun

Antioxidant Enzyme Activity and mRNA Expression in Reproductive Tract of Adult Male European Bison (Bison bonasus, Linnaeus 1758)

Antioxidants in the male reproductive tract are the main defence factors against oxidative stress caused by reactive oxygen species production, which compromises sperm function and male fertility. This study was designed to determine the activity of antioxidant enzymes, such as superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx), in the testicular and epididymidal tissues of adult male European bison (Bison bonasus). The reproductive tract tissues were subjected to real-time reverse transcriptase-polymerase chain reaction (RT-PCR) analysis to quantify mRNA expression levels of five antioxidant enzymes: copper/zinc SOD (Cu/Zn SOD), secretory extracellular SOD (Ec-SOD), CAT, phospholipid hydroperoxide glutathione peroxidase (PHGPx) and GPx5. The corpus and cauda epididymidal tissues displayed greater (p < 0.05) SOD activity compared with the testicular tissue. It was found that CAT activity was lowest (p < 0.05) in the cauda epididymidis, whereas negligible GPx activity was detected in the reproductive tract tissues. There were no detectable differences in the mRNA expression level of Cu/Zn SOD among the different reproductive tract tissues. Small amounts of Ec-SOD mRNA were found in the reproductive tract, particularly in the epididymides. The caput and cauda epididymides exhibited greater (p < 0.05) level of CAT mRNA expression, whereas PHGPx mRNA was more (p < 0.05) expressed in the testis. Furthermore, extremely large amounts of GPx5 mRNA were detected in the caput epididymidal tissue compared with other tissues of the reproductive tract. It can be suggested that the activity of the antioxidant enzymes and the relative gene expression of the enzymes confirm the presence of tissue-specific antioxidant defence systems in the bison reproductive tract, which are required for spermatogenesis, epididymal maturation and storage of spermatozoa.

Pattern of expression of vascular endothelial growth factor and its receptors in the ovine choroid plexus during long and short photoperiods

Vascular endothelial growth factor (VEGF-A) plays an important role in maintaining cerebrospinal fluid (CSF) homeostasis and the function of the choroid plexuses (CPs). The objective of the study was to determine the expression of vascular endothelial growth factor (VEGF-A), tyrosine kinase receptors Flt-1 and KDR and KDR co-receptor neuropilin 1 (NRP-1) in ovine CPs during different photoperiods. CPs were collected from the lateral brain ventricles from ovariectomized, estradiol-treated ewes during long day (LD; 16L:8D, n = 5) and short day (SD; 8L:16D, n = 5) photoperiods. We analyzed mRNA expression levels of two VEGF-A isoforms, VEGF-A120 and VEGF-A164 and our results indicate that VEGF-A164 was the predominant isoform. Expression levels of VEGF-A and Flt-1 were similar during the SD and LD photoperiods. There were significant increases in KDR mRNA and protein expression (p < 0.05) and NRP-1 mRNA expression (p < 0.05) during SD. These data show that expression of KDR and its co-receptor NRP-1 are up-regulated by short photoperiod and that this effect is not dependent on ovarian steroids. Our results suggest that the VEGF-A-system may be involved in photoperiodic plasticity of CP capillaries and may therefore be responsible for photoperiodic changes in the CSF turnover rate in ewes.

Local transfer of testosterone and aromatase activity in the spermatic cord in wild boar/pig hybrids in short-daylight and long-daylight periods

The local transfer of testosterone (T) and immunolocalization of cytochrome P450 aromatase (P450arom) in the spermatic cord vessels of ten male wild boar×domestic pig hybrids were examined in December (short-daylight period) and June (long-daylight period). Total T concentration was determined in the jugular vein (JV) and free T concentration was estimated in the common carotid artery (CA), branches of the testicular artery supplying the testis (TA) and epididymis (EA), as well as in testicular veins draining blood from the testis (TV) and spermatic cord (SV). P450arom was immunolocalized in the arterial and venous vessels of the spermatic cord. The concentrations of total T in the JV and free T in the CA did not differ between the examined periods. However, in December, free T concentrations in the TA and EA were higher (p<0.01-0.001) than in the CA. In June, free T concentration was higher (p<0.01) in EA than in CA and TA. The concentrations of free T in the TV and SV were higher (p<0.001) than in the JV regardless of the period. Also, free T concentration in the SV was higher (p<0.05) in June than in December. P450arom was expressed in all layers of the arterial and venous vessels of the spermatic cord. In June, the intensity of the P450arom staining was higher than in December. The results suggest that the local supply of the male reproductive organs with steroid hormones operate in the hybrids of wild boar×domestic pig. This supply includes the local transfer of testosterone and the P450arom action.

Relationship Between Contractions of the Uterus and Concentration of PGF2α in Uterine Venous Blood after Luteolysis in Gilts

The origin and physiological significance of high pulses of prostaglandin F2α (PGF2α) in uterine venous blood that occur 2-3 days after luteolysis are not well understood. We studied the relationship between contractions of the uterus evoked by exogenous oxytocin (OT) and PGF2α concentration in uterine venous blood on day 17 of the porcine oestrous cycle. The infusion of OT into the uterine artery produced an immediate increase in the uterine intraluminal pressure (UIP) (p < 0.001) and a simultaneous elevation in PGF2α concentration in uterine venous blood (p < 0.0001). The infusion of indomethacin (IND) into the uterine artery slightly decreased PGF2α concentration in uterine venous blood, but it did not suppress uterine contraction or the rapid increase in PGF2α concentration in uterine venous blood just after OT infusion (p < 0.0001), which was lower that in gilts not treated with IND. We conclude that the spikes of PGF2α concentration in uterine venous blood occurring after OT infusion on day 17 of the porcine oestrous cycle are mainly caused by the excretion with venous blood from the remodelled uterus and that PGF2α synthesis may contribute to this. These results suggest that the high spikes in PGF2α concentration that occur 2-3 days after luteolysis in pigs, sheep, cows and mares all have a similar origin.

Determination of the activity and relative abundance of mRNA for antioxidant enzymes in stallion testicular and epididymal tissues: A comparison between two breeding seasons

The key prerequisite for successful insemination is sperm characterized to have positive values for morphological and biological variables which are determined by, among others, effective antioxidant protection during the lifespan of sperm cells. This study evaluated the activity and relative abundance of mRNA for antioxidant enzymes in stallion testicular and epididymal tissues during breeding (n = 5) and non-breeding (n = 5) seasons. The activity of superoxide dismutase (SOD) was greater (P < 0.05) during the breeding season, in particular in the testes and the caput epididymis, and SOD1 was the predominant isoform of the enzyme. The expression of the SOD3 gene was markedly less in the analyzed tissues, which indicates that this enzyme contributes to the antioxidant protection of the stallion reproductive tract. The activity of catalase (CAT) was less (P < 0.05) in the testes during both seasons while its relative abundances only during the breeding season. The greatest CAT activity was noted in the cauda epididymis during the breeding season. The activity of glutathione peroxidases (GPx) was greater (P < 0.05) in the testes than in other tissues and 10-fold greater during the breeding season. Similarly, relative abundance of GPx5 mRNA was greater (P < 0.05) in the caput epididymis than in the remaining tissues during both seasons. This study demonstrated that season has an ambiguous influence on the antioxidant defense system in stallion reproductive tissues. Seasonal differences in the present study, however, indicate that the reproductive system of stallions adapts well to environmental seasonal changes.

Expression of hypoxia inducible factor 1α and antioxidant enzymes: Superoxide dismutases-1 and -2 in ischemic porcine endometrium

The aim of this study was to determine how 60-min ischemia changes the expression of hypoxia inducible factor 1α (HIF-1α) and superoxide dismutases (SOD)-1 and -2 in selected regions of porcine uterine horns. The results showed that 60-min ischemia of the porcine uterus conducted at the mid-secretory estrous phase caused decreased HIF-1α and increased SOD-2 gene expression. Higher expression of SOD-2 suggests that this enzyme may play an important role in the suppression of HIF-1α accumulation in an ischemic endometrium.

Expression of Hypoxia-Inducible Factor-1α and association with Cu/Zn superoxide dismutase in the reproductive tract tissues of adult male roe deer (Capreolus capreolus) during the reproduction season

Abstract This study aimed to determine the mRNA and protein expressions of HIF-1α and SOD-1 in testes and epididymis of adult male roe deer during the periods of reproductive season. The mRNA expression levels of investigated factors were examined using RT-PCR, while protein expression levels were evaluated using Western blot analysis. Differences between tissues within this period and between periods were evaluated. Testicular tissue exhibited lower HIF-1α and SOD-1 mRNA expression levels during all periods as compared to epididymal tissues. Among the epididymal tissues, cauda epididymis showed higher HIF-1α mRNA expression levels in comparison with caput and corpus tissues in the pre-rut period, and not much lower SOD-1 mRNA expression levels in comparison with caput in all periods. Protein expression of HIF-1α was higher in the epididymis than in the testicular tissues in the pre-rut and rut periods. Its expression was the highest in caput epididymal tissue compared with either the testicular, corpus and cauda epididymal tissue during rut and higher than corpus and testis in pre- and post-rut periods. Testicular tissue was characterized by lower SOD-1 protein expression levels than in epididymis in pre-rut and rut periods. This study indicates that the reproductive tract of male roe deer possesses a tissue-specific defense system which protects the tissues from oxygen deficiency damage, which is important during cyclic changes occurring in the reproductive tissues during the reproductive season.