Long form of leptin receptor gene and protein expression in the porcine trophoblast and uterine tissues during early pregnancy and the oestrous cycle

Kamiński

Siawrys

et al. 2013

Leptin is a polypeptide hormone produced primarily by adipocytes. It has been implicated in the regulation of satiety and energy homeostasis. Leptin has been suggested to play a role in reproduction based on its involvement in the regulation of the hypothalamic-pituitary-gonadal axis via endocrine, paracrine and/or autocrine pathways. The aim of the present study was to localize the cellular distribution of leptin and the long isoform of leptin receptor (OB-Rb) genes in porcine ovarian antral follicles and to compare the expression levels of leptin and OB-Rb mRNAs in porcine granulosa cells (GC), theca interna (TIC) and theca externa (TEC) cells during the luteal phase of the estrous cycle and in early pregnancy. The expression of leptin and OB-Rb genes was detected in GC, TIC and TEC. Significantly higher levels of leptin gene expression in GC were observed during the mid-and late-luteal phases of the cycle than on days 30 to 32 of pregnancy. On days 14 to 16 of pregnancy, leptin mRNA expression was higher than that on days 14 to 16 of the cycle. The expression of the OB-Rb gene in GC and TEC increased during pregnancy in comparison with the analyzed luteal phases of the cycle. Our results validate the hypothesis that locally produced leptin plays a role in the regulation of porcine reproduction at the ovarian level and exerts a direct effect on porcine follicles. The differences in OB-Rb gene expression in porcine GC and theca cells also suggest that their sensitivity to leptin varies in the ovaries of pregnant and cyclic pigs.

Section: Introductionmentioning

confidence: 99%

Expression of leptin and its receptor genes in the ovarian follicles of cycling and early pregnant pigs

Kamiński

Siawrys

et al. 2013

“…Western blotting Western blotting analysis was performed essentially as described by Smolinska et al (2007). Briefly, equal amounts of porcine uterine lysates (endometrium and myometrium parts separately, 10 µg of total proteins) were resolved by SDS-PAGE (12.5% gel) for separating PPO, OX1R, OX2R and actin, and then transferred to nitrocellulose membranes (Whatman, Piscataway, NJ, USA).…”

Section: Experimental Animalsmentioning

confidence: 99%

Expression of the orexin system in the porcine uterus, conceptus and trophoblast during early pregnancy

Kieżun

Szeszko

et al. 2015

Orexin A and B are hypothalamic peptides derived from the prepro-orexin (PPO) precursor. Orexins stimulate food intake and arousal. Those peptides bind and activate two G protein-coupled receptors: orexin receptor 1 (OX1R) and orexin receptor 2 (OX2R). Numerous authors have suggested that orexins play an important role in the regulation of the reproductive functions. The objective of the present study was to analyse the presence of and changes in the gene and protein expression pattern of the orexin system in the porcine uterus, conceptus and trophoblast (chorioallantois) during early pregnancy. In the endometrium, the highest PPO and OX1R gene expression was detected on days 15 to 16 of gestation. The OX2R mRNA content in the endometrium was higher on days 10 to 11 and 15 to 16 than on days 12 to 13 and 27 to 28. In the trophoblasts, PPO gene expression was higher on days 30 to 32 than on days 27 to 28. The highest PPO protein content in the endometrium was noted on days 12 to 13. The highest OX1R protein content in the endometrium was detected on days 10 to 11, whereas OX2R protein on days 15 to 16. In the trophoblasts, PPO and OX1R protein levels were more pronounced on days 27 to 28 than on days 30 to 32, but OX2R expression was higher on days 30 to 32. The expression of PPO, OX1R and OX2R was different in the conceptuses and trophoblasts during early pregnancy. Local orexin production and the presence of the specific orexin receptors suggest that the orexin system may participate in the control of porcine reproductive functions by exerting endocrine and auto/paracrine effects on the uterus, conceptuses and trophoblasts during early pregnancy. This study provides the first evidence for the presence of orexins and their receptors in the uteri, conceptuses and trophoblasts in pigs during early pregnancy. The local orexin system is dependent on the stage of pregnancy.Keywords: orexin A and B, orexin receptors 1 and 2, uterus, pregnancy, pig ImplicationsThere is a growing body of evidence to implicate the existence of a common endocrine system that controls metabolism and the reproductive system. Orexin A and orexin B, the hormones that control energy homoeostasis, may be a part of the above system. This paper provides the pioneering information about the presence of orexins and orexin receptors in the porcine uterus, conceptus and trophoblast during early pregnancy, the crucial period for reproductive success. The expression levels of orexins and their receptors were found to be correlated with the stages of early pregnancy. A sound knowledge of mechanisms that control energy homoeostasis and reproduction is needed in animal breeding practice.

“…Western blotting Western blotting analysis was performed as described by Smolinska et al (2007). Briefly, equal amounts of porcine pituitary (anterior and posterior parts, 10 μg) were resolved by SDS-PAGE (12.5%) for separating PPO and actin and transferred to nitrocellulose membranes (Whatman, Piscataway, NJ, USA).…”

Section: Measurement Of Plasma Progesterone Levelmentioning

confidence: 99%

“…After isolation from gel, DNA was sequenced (ABI Prism TM BigDye TM Terminator Cycle Sequencing kit, ABI Prism 3777 DNA sequencer, USA) in both directions to confirm the accuracy of amplification. Comparison of the PCR-amplified DNA sequences to those in the database indicated 100% homology at the nucleotide level.Western blotting Western blotting analysis was performed as described by Smolinska et al (2007). Briefly, equal amounts of porcine pituitary (anterior and posterior parts, 10 μg) were resolved by SDS-PAGE (12.5%) for separating PPO and actin and transferred to nitrocellulose membranes (Whatman, Piscataway, NJ, USA).…”

mentioning

confidence: 99%

The effect of the estrous cycle on the expression of prepro-orexin gene and protein and the levels of orexin A and B in the porcine pituitary

Nitkiewicz

Maleszka

et al. 2014

Hypothalamic peptides orexin A (OXA) and orexin B (OXB) are derived from the proteolytic cleavage of a common precursor molecule, prepro-orexin (PPO). They act via two orexin receptors (OX1R and OX2R), which belong to the G-protein coupled receptor superfamily. Orexins are implicated in the regulation of arousal states, energy homeostasis and reproductive neuroendocrine function. The objective of this study was to investigate the presence and changes in orexin expression in the porcine pituitary during the estrous cycle. Adenohypophysis (AP) and neurohypophysis (NP) tissue samples were harvested on days 2 to 3, 10 to 12, 14 to 16, and 17 to 19 of the estrous cycle. The expression of the PPO gene increased in AP and NP during the estrous cycle. The highest PPO protein concentrations in AP were reported on days 2 to 3 (P < 0.05), and in NP -on days 10 to 12 and 17 to 19 ( P < 0.05). The expression of PPO mRNA was lower in AP than in NP, but PPO protein levels were higher in AP. In AP, OXA immunoreactivity was higher ( P < 0.05) on days 10 to 12 and 14 to 16. In NP, the highest ( P < 0.05) content of the analyzed protein was observed on days 10 to 12 and the lowest ( P < 0.05) -on days 14 to 16 and 17 to 19. OXB immunoreactivity in AP reached the highest level ( P < 0.05) on days 2 to 3, and the lowest level ( P < 0.05) was determined on days 10 to 12 and 17 to 19. OXB protein concentrations in NP peaked ( P < 0.05) on days 10 to 12 of the cycle. Our study was the first experiment to demonstrate the expression of the orexin gene and orexin proteins in the porcine pituitary and the correlations between expression levels and the phase of the estrous cycle. Keywords: prepro-orexin, orexin A and B, pituitary, estrous cycle, pig ImplicationsThe relationship between nutritional status and reproductive success in animals has been studied for many years. Considerable evidence has been accumulated to implicate the existence of a common endocrine system that controls metabolism and the reproductive system. It is suggested that orexins belong to the group of hormones, which regulate metabolic status and reproduction. In this study, we demonstrated differences in orexin gene and orexin proteins in the porcine pituitary during the estrous cycle. The results imply the impact of hormonal milieu on orexins' expression in the porcine pituitary. A sound knowledge of mechanisms that control energy homeostasis and reproduction is needed in animal breeding practice. IntroductionOrexin A and B (OXA, OXB), also known as hypocretin 1 and 2, are hypothalamic peptides with a 46% sequence homology. OXA is a 33-amino acid peptide with a molecular mass of~3.5 kDa, which has an N-terminal pyroglutamyl residue and C-terminal amidation. OXB is a 28-amino acid peptide with a molecular mass of~2.9 kDa and C-terminal amidation. Orexins are cleaved from a 130-amino acid common precursor molecule, prepro-orexin (PPO), which is synthesized mainly in neurons inside and in the area of the lateral and posterior hypothalamus. Orexin action is mediated ...