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1.
Cell-to-cell interaction via cell contact-dependent pathway is essentially important for maintenance and regulation of corpus luteum (CL) integrity and its physiological actions. The objective of the present study was to evaluate the mRNA expression of the cell adhesion molecules (CAMs) that are constituent factors of gap junctions [connexin (Cx) 43] and adherence junctions (VE-, E-, N-cadherin) in two types of endothelial cells from the mid CL and in CL tissue during the estrous cycle and PGF(2alpha)-induced luteolysis in the cow. Specific mRNA expression for Cx43 and N-cadherin was detected in cytokeratin-positive (CK+) and cytokeratin-negative (CK-) luteal endothelial cells (EC) and fully luteinized granulosa cells (LGC). E-cadherin mRNA was expressed in CK+EC and LGC, but not in CK-EC. VE-cadherin mRNA was expressed in both CK+ and CK-EC. During the estrous cycle, Cx43 mRNA expression was significantly lower in the regressing CL. VE-cadherin expression also tended to increase in the mid CL and increased significantly in the regressing CL. E-cadherin mRNA expression was higher in the early and late CL than in the mid- and regressing CL. N-cadherin mRNA expression gradually increased from the early to late CL followed by a decrease in the regressing CL. During PGF(2alpha)-induced luteolysis, Cx43 mRNA expression appeared to increase, and VE-cadherin and E-cadherin mRNA significantly increased at 24 h. N-cadherin mRNA expression decreased 2 and 4 h after PGF(2alpha) administration. Collectively, expression of the mRNAs for CAMs was different in the two types of luteal endothelial cells and fully luteinized granulosa cells and changed independently in the CL during the estrous cycle and PGF(2alpha)-induced luteolysis in the cow. The results suggest that CAMs play physiological roles in cell-to-cell communication to regulate both gap and adherence junctions during CL development and regression in the cow.  相似文献   

2.
The aim of this study was to evaluate mRNA expression, protein concentration and localization of the assumedly important lymphangiogenic factors VEGFC and VEGFD and the receptor FLT4 in bovine corpora lutea (CL) during different physiological stages. In experiment 1, CL were collected in a slaughterhouse and stages (days 1–2, 3–4, 5–7, 8–12, 13–16, >18) of oestrous cycle and month <3, 3–5, 6–7 and >8 of pregnancy. In experiment 2, prostaglandin F2α (PGF)‐induced luteolysis was performed in 30 cows, which were injected with PGF analogue on day 8–12 (mid‐luteal phase), and CL were collected before and 0.5, 2, 4, 12, 24, 48 and 64 h after PGF injection. The mRNA expression was characterized by RT‐qPCR. All three factors were clearly expressed and showed significant changes during different groups and periods examined in both experiments. Protein concentrations of VEGFD and FLT4 measured by ELISA were not detectable in early cyclic CL but increased to higher plateau levels during pregnancy. After PGF‐induced luteolysis FLT4 protein showed an increase within 2–24 h after the injection. FLT4 localization by immunohistochemistry in the cytoplasm of luteal cells was relatively weak in early CL. It increased in late CL and especially in CL during pregnancy. During pregnancy, a positive FLT4 staining in both the nucleus and cytoplasm of lymphatic endothelial cells in peripheral tissue was observed. In conclusion, our results lead to the assumption that lymphangiogenic factors are produced and regulated in CL and may be involved in mechanisms regulating CL function, especially during pregnancy.  相似文献   

3.
Luteolytic capacity is defined as the ability of corpora lutea (CL) to undergo luteolysis after prostaglandin (PG) F2alpha treatment. The mechanisms causing acquisition of luteolytic capacity are not yet identified but CL without luteolytic capacity have PGF2alpha receptors and respond to PGF2alpha with some changes in gene expression. Inhibition of progesterone biosynthesis is a key feature of luteolysis and therefore we postulated that genes involved in progesterone biosynthesis would be regulated by PGF2alpha differently in CL with or without luteolytic capacity. Gilts on day 9 after estrus (lack luteolytic capacity) or day 17 of pseudopregnancy (with luteolytic capacity) were treated with saline or a PGF2alpha analog (cloprostenol) and CL were collected 0.5 (Experiment I) or 10 h (Experiment II) later. In Experiment III, large luteal cells from CL on day 9 or 17 were cultured for 1, 12 and 24h with or without PGF2alpha. PGF2alpha decreased LDL receptor mRNA (27%), steroidogenic acute regulatory protein (StAR) mRNA (41%), StAR protein (75%), LH receptor mRNA (55%), and LH receptor protein (45%) at 10 h after treatment in day 17 but not day 9 CL. PGF2alpha increased DAX-1 mRNA at 0.5 h (43%) and 10 h (46%) after PGF2alpha in day 17 but not day 9 CL but decreased 3betaHSD mRNA ( approximately 20% at 10 h) in both days 9 and 17 CL. In vitro, PGF2alpha decreased StAR mRNA at 12 h only in day 17 luteal cells; however, continuous treatment with PGF2alpha for 24 h decreased StAR mRNA in both days 9 and 17 luteal cells. Thus, luteolytic capacity involves a critical change in responsiveness of DAX-1, StAR, and LH receptor to PGF2alpha that results in inhibition of luteal progesterone biosynthesis.  相似文献   

4.
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5.
Prostaglandin F(2α) (PGF(2α)) induces luteolysis via a specific receptor, PTGFR. Although PTGFR mRNA expression in the bovine corpus luteum (CL) has been studied previously, changes in PTGFR protein and its localization are not fully understood during the life span of the CL. In addition to full-length PTGFR, several types of PTGFR isoforms, such as PTGFRα (type I) and PTGFRζ (type II), were reported in the bovine CL, suggesting isoform-specific luteal action. Full-length PTGFR mRNA in the bovine CL increased from the early to the mid-luteal phase and decreased during luteolysis, whereas PTGFR protein remained stable. PTGFR protein was localized to both luteal and endothelial cells and was expressed similarly during the life span of the CL. Like full-length PTGFR mRNA, PTGFRα and PTGFRζ mRNA also increased from the early to mid-luteal phases, and mRNA of PTGFRζ, but not PTGFRα, decreased in the regressing CL. During PGF(2α)-induced luteolysis, the mRNAs of full-length PTGFR, PTGFR,α and PTGFRζ decreased rapidly (from 5 or 15 min after PGF(2α) injection), but PTGFR protein decreased only 12 h later. Silencing full-length PTGFR using small interfering RNA prevented PGF(2α)-stimulated cyclooxygenase-2 (PTGS2) mRNA induction. By contrast, PGF(2α) could stimulate vascular endothelial growth factor A (VEGFA) mRNA even when full-length PTGFR was knocked down, thus suggesting that PGF(2α) may stimulate PTGS2 via full-length PTGFR, whereas VEGFA is stimulated via other PTGFR isoforms. Collectively, PTGFR protein was expressed continually in the bovine CL during the estrous cycle, implying that PGF(2α) could function throughout this period. Additionally, the bovine CL expresses different PTGFR isoforms, and thus PGF(2α) may have different effects when acting via full-length PTGFR or via PTGFR isoforms.  相似文献   

6.
The aim of this study was to characterize expression patterns of hypoxia‐inducible factor‐1alpha (HIF1A) and vasohibin family members (VASH1 and VASH2) during different stages of ovarian function in cow. Experiment 1: Antral follicle classification occurred by follicle size and estradiol‐17beta (E2) concentration in the follicular fluid into 5 groups (<0.5, 0.5–5, 5–40, 40–180 and >180 E2 ng/ml). Experiment 2: Corpora lutea (CL) were assigned to the following stages: days 1–2, 3–4, 5–7, 8–12, 13–16 and >18 (after regression) of oestrous cycle and of pregnancy (months 1–2, 3–4, 6–7, >8). Experiment 3: Cows on days 8–12 were injected with a prostaglandin F2alpha (PGF) analogue and CL were collected before and 0.5, 2, 4, 12, 24, 48 and 64 hr after PGF injection. Expression of mRNA was measured by qPCR, steroid hormone concentration by EIA and localization by immunohistochemistry. HIF1A mRNA expression in our study increases significantly in follicles during final maturation. The highest HIF1A mRNA expression was detected during the early luteal phase, followed by a significant decrease afterwards. In contrast, the mRNA of vasohibins in small follicle was high, followed by a continuous and significant downregulation in preovulatory follicles. The obtained results show a remarkable inverse expression and localization pattern of HIF1A and vasohibins during different stages of ovarian function in cow. These results lead to the assumption that the examined factors are involved in the local mechanisms regulating angiogenesis and that the interactions between proangiogenic (HIF1A) and antiangiogenic (vasohibins) factors impact all stages of bovine ovary function.  相似文献   

7.
Prostaglandin F2alpha (PGF2alpha) is the primary luteolysin in the cow. During the early luteal phase, the corpus luteum (CL) is resistant to the luteolytic effect of PGF2alpha. Once mature, the CL becomes responsive to PGF2alpha and undergoes luteal regression. These actions of PGF2alpha coincide with changes in luteal blood flow (BF): PGF2alpha has no effect on BF in the early CL, but acutely increases BF in the peripheral vasculature of the mature CL within 30 min of PGF2alpha injection. During spontaneous luteolysis, luteal BF increases on Days 17-18 of the estrous cycle, prior to any decrease in plasma progesterone (P). The increase in luteal BF is synchronous with an increase in plasma PGFM levels, suggesting that pulsatile release of PGF2alpha from uterus stimulates the increase in luteal BF. Serial biopsies of these CL showed that mRNA expression for endothelial nitric oxide synthase (eNOS) together with endothelin-1 (ET-1) and angiotensin converting enzyme (ACE) increases on Days 17-18 when the luteal BF is elevated. On Day 19 when plasma P level firstly decreases, eNOS mRNA returns to the basal level whereas ET-1 and ACE mRNA remains elevated. Cyclooxygenase-2 (COX-2) mRNA expression increases on Day 19. In support of these data, an in vivo microdialysis study revealed that luteal ET-1 and angiotensin II (Ang II) secretion increases and precedes PGF2alpha secretion during spontaneous luteolysis. In conclusion, we show for the first time that an acute increase of BF occurs in the peripheral vasculature of the mature CL together with increases in eNOS expression and ET-1 and Ang II secretion in the CL during the early stages of luteolysis in the cow. We propose that the increase in luteal BF may be induced by NO from large arterioles surrounding the CL, and simultaneously uterine or exogenous PGF2alpha directly increases ET-1 and Ang II secretion from endothelial cells of microcapillary vessels within the CL, thereby suppressing P secretion by luteal cells. Taken together, our results indicate that an acute increase in luteal BF occurs as a first step of luteolysis in response to PGF2alpha. Therefore, local BF plays a key role to initiate luteal regression in the cow.  相似文献   

8.
Although prostaglandin (PG) F2alpha is known to be a principal luteolytic factor, its action on the bovine corpus luteum (CL) is mediated by other intra-ovarian factors. Tumor necrosis factor-alpha (TNFalpha) and its specific receptors are present in the bovine CL with the highest expressions at luteolysis. TNFalpha in combination with interferon-gamma reduced progesterone (P4) secretion, increased PGF2alpha and leukotriene C4 (LTC4) production, and induced apoptosis of the luteal cells in vitro. Low concentrations of TNFalpha caused luteolysis, which resulted in a decreased level of P4, and increased levels of PGF2alpha, LTC4 and nitrite/nitrate (stable metabolites of nitric oxide-NO) in the blood. Inhibition of local NO production counteracts spontaneous and PGF2alpha-induced luteolysis. Therefore, NO is a likely candidate for the molecule that mediates PGF2alpha and TNFalpha actions during luteolysis. Both PGF2alpha and TNFalpha increase NO concentrations in blood, and stimulate NO synthase expression on protein level in the bovine CL cells. NO stimulates PGF2alpha and LTC4 secretion, inhibits P4 production and reduces the number of viable luteal cells. TNFalpha and NO induce apoptotic death of the CL by modulating expression of bcl-2 family genes and by stimulating expression and activity of caspase-3. The above findings indicate that TNFalpha and NO play crucial roles in functional and structural luteolysis in cattle.  相似文献   

9.
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10.
The components of the IGF-system were shown to be differentially regulated in bovine antral follicles and corpora lutea (CL) during different stages of the estrous cycle, and to have important functions for specific stages. The aim of this study was to investigate the detailed pattern of mRNA expression of most constituents of the IGF-system and their possible involvement in prostaglandin (PG)F2-induced luteolysis in the bovine CL. Therefore, cows in the mid-luteal phase (days 8–12) were injected with the PGF2-analogue Cloprostenol, and CL were collected by transvaginal ovariectomy at 2, 4, 12, 48 and 64 h after PGF2-injection. Real-time RT-PCR using SYBR Green I detection was employed to determine mRNA expressions of the following factors: ubiquitin (UBQ), insulin-like growth factor I (IGF I), IGF II, IGF-receptor type 1 (IGFR-1), growth hormone receptor (GH-R) and IGF-binding proteins-1–6 (IGFBP-1–6). Total extractable RNA decreased with ongoing luteolysis. IGFBP-1 mRNA was significantly up-regulated at 2 h after PGF2 and maximal at 4 h with a 34-fold increase. IGFBP-5 mRNA was significantly up-regulated after 12 h with a maximum of an 11-fold increase at 64 h. For GH-R, IGFR-1, IGF II, IGFBP-3 and -4 mRNA expression, we found a significant down-regulation in certain stages. There was a significant up-regulation for IGFBP-2 and -6 mRNA at 64 h after induced luteolysis. There were no significant changes in IGF I mRNA expression. In conclusion, the IGF-system with all its components seems to play an important role in the very complex process of PGF2-induced luteolysis in bovine CL.  相似文献   

11.
Endothelin-1 (ET-1) is a luteolytic mediator in the bovine corpus luteum (CL), and its action appears to be via endothelin type A receptor (ETR-A). Thus, the aim of the present study was to determine the effect of ETR-A antagonist on PGF2alpha-induced luteolysis in the cow. Cows on days 10-12 of the estrous cycle were subjected to five intraluteal injections of the ETR-A antagonist LU 135252 in saline or only saline at -0.5, 2, 4, 6, and 8 h after PGF2alpha administration (=0 h). Serial luteal biopsies were conducted to determine the expression of mRNA in the luteal tissue. There were no significant differences in the decrease in plasma progesterone (P) concentrations and the mRNA expressions of steroidogenic acute regulatory protein and 3beta-hydroxysteroid dehydrogenase/Delta5, Delta4-isomerase between the ETR-A antagonist-treated group and the control group. However, the start of the decline in CL volume and blood flow area surrounding the CL was delayed for almost two days in the ETR-A antagonist-treated group compared to the control group. The mRNA expression of preproET-1 and endothelin type B receptor increased in both groups, while the ETR-A mRNA remained unchanged. In addition, caspase-3 mRNA expression increased significantly at 24 h in the control group only and its level was higher than that of the ETR-A antagonist-treated group. Thus, the present study suggests that ET-1 regulates structural luteolysis via ETR-A by controlling blood vessel contraction in the CL of the cow.  相似文献   

12.
The corpus luteum (CL) undergoes regression by prostaglandin (PG)F(2alpha) from uterus and endothelin-1 (ET-1) plays an important role during luteolysis as a local mediator of PGF(2alpha) in the cow. Endothelial cells (EC) and luteal cells are main cell types making up the CL and their interactions are vital for CL function. We aimed to examine the relevance of interactions between EC and luteal cells on stimulation of genes which involved ET-1 synthesis by PGF(2alpha). We further focused the impact of maturity of luteal cells on the stimulation of the genes. To make a microenvironment which resembles the CL, we used bovine aortic endothelial cells (BAEC) and luteinizing or fully-luteinized granulosa cells (GC) and evaluated the effect of PGF(2alpha) on the expression for mRNA of ET-1 system by using real-time RT-PCR. PGF(2alpha) stimulated the expression of preproET-1 and endothelin converting enzyme-1 mRNA only in the co-cultures of BAEC with fully-luteinized GC, but not with luteinizing GC. The data suggest that interactions between BAEC and fully-luteinized GC enhance the capability of BAEC to produce ET-1 in response to PGF(2alpha). This mechanism may contribute to the local induction of luteolytic action of PGF(2alpha) which is dependent on the age/maturation of the CL.  相似文献   

13.
Endothelin-1 (ET-1), a 21-amino acid peptide was initially identified as a potent vasoconstrictor, ET-1 plays an important role in the female reproductive cycle: its quick ascent during luteal regression, ability to inhibit steroidogenesis in vitro and in vivo, combined with the observation that the luteolytic effects of prostaglandin F2alpha (PGF2alpha) were delayed by pretreatment with ET-1 receptors type A (ETA) antagonists suggest that this peptide functions as an important element of the luteolytic cascade. The observation that ETA receptor expression was inversely correlated with steroidogenesis in luteal cells; namely factors which stimulated steroidogenesis inhibited ETA receptor levels is also in accord with the inhibitory role of ET-1 in corpus luteum (CL) function. Contrary to the mature mid cycle CL, the CL of early cycle is refractory to PGF2alpha-induced luteolysis. PGF2alpha administered at early luteal phase (day 4 of the cycle) failed to increase luteal ET-1 gene expression or its ETA receptors. In contrast, both genes were markedly induced in mid cycle CL exposed to PGF2alpha. ET-1 gene is transcribed as prepro ET-1 (ppET-1) and the active form of peptide is derived from the inactive intermediate big ET-1, by endothelin-converting enzyme-1 (ECE-1), therefore alterations in mature ET-1 levels can be achieved by modulating the expression of ppET-1 and/or ECE-1. Analysis using in situ hybridization and enriched luteal cell subpopulations showed that both steroidogenic and endothelial cells of the CL expressed high levels of ECE-1 mRNA. The ppET-1 mRNA, on the other hand, was only expressed by resident endothelial cells, suggesting that luteal parenchymal and endothelial cells cooperate in the biosynthesis of mature bioactive ET-1. A significant, four-fold elevation in ECE-1 expression (mRNA and protein levels) occurred during the transition of the CL from early to mid luteal phase. This increase was accompanied by a significant rise in ET-1 peptide. Surprisingly however, ppET-1 mRNA levels remained similar during early and mid luteal phase. Collectively, these studies demonstrate that: (a) the various components of ET-1 system (ET-1/ECE-1/ETA) are dynamically and independently regulated during bovine luteal life span. (b) The CL becomes PGF2alpha-responsive only when both ppET-1 and ECE-1 genes are expressed at a level which enable an uninterrupted ET-1 biosynthesis.  相似文献   

14.
Estrogen (E) exerts its function by binding to two intracellular estrogen receptors, ERalpha and ERbeta. Although ERs have been reported to be expressed in the bovine corpus luteum (CL), the mechanisms that control ER expression in the bovine CL are not fully understood. To determine the possible regulatory mechanisms of ERalpha and ERbeta that meditate distinct E functions, we examined 1) the changes in the protein expressions of ERs in the CL throughout the luteal phase and 2) the effects of prostaglandin (PG) F2alpha, tumor necrosis factor-alpha (TNFalpha) and interferon-gamma (IFNgamma) on the expressions of ERs in cultured bovine luteal cells. Western blot analyses revealed that ERalpha and ERbeta proteins were expressed throughout the luteal phase. The ERalpha protein level was high at the early luteal (Days 2-3 after ovulation) and mid-luteal stages (Days 8-12) and was extremely low at the regressed luteal stage (Days 19-21). The ERbeta protein level increased from the early to developing luteal stage, remained at the same level at the mid-luteal stage and decreased thereafter. The ratio of ERbeta to ERalpha was higher in the regressed stage than in the other stages. Luteal cells obtained from mid-stage CLs (Days 8-12) were incubated with PGF2alpha (0.01-1 microM), TNFalpha (0.0145-1.45 nM) or IFNgamma (0.0125-1.25 nM) for 24 h. PGF2alpha and TNFalpha inhibited ERa and ERbeta mRNA expressions. IFNgamma suppressed ERbeta mRNA expression but did not affect the expression of ERalpha mRNA. However, the ERalpha and ERbeta protein levels were not affected by any of the above treatments. These data indicate that PGF2alpha, TNFalpha and IFNgamma regulate ERalpha and ERbeta mRNA expressions in bovine luteal cells. Moreover, the changes in the ERbeta/ERalpha ratio throughout the luteal phase suggest that ERalpha is associated with luteal maintenance. Therefore, a dramatic decrease in ERalpha at the regressed luteal stage could result in progression of structural luteolysis in the bovine CL.  相似文献   

15.
Improving our understanding of the mechanisms controlling the corpus luteum (CL) and its role in regulating the reproductive cycle should lead to improvements in the sustainability of today's global animal industry. The corpus luteum (CL) is a transient endocrine organ composed of a heterogeneous mixture steroidogenic, endothelial and immune cells, and it is becoming clear that immune mechanisms play a key role in CL regulation especially in luteolysis. Toll‐like receptors (TLR) mediate innate immune mechanisms via the production of pro‐inflammatory cytokines, especially within various tissues, although the role of TLR within CL remains unknown. Thus, the objectives of this study were to characterize TLR mRNA expression in the CL during the oestrous cycle and in pregnancy (day 30–50), and to examine the role of TLR signalling in luteal cells. Corpora lutea were collected at various stages of the cycle and pregnancy and analysed for TLR and cytokine mRNA expression. In addition, luteal cells were cultured with the TLR4 ligand (lipopolysaccharide, LPS) for 24 h to evaluate the role of TLR4 in regulating luteal function. Toll‐like receptors 1, 2, 4, 6, tumour necrosis factor alpha (TNF), interferon gamma (IFN‐G), and interleukin (IL)‐12, mRNA expressions were greatest in regressing CL compared with earlier stages (p < .05), whereas no change was observed for IL‐6 mRNA expression. Cytokine mRNA expression in cultured luteal cells was not altered by LPS. Based on these data, one or more of the TLRs found within the CL may play a role in luteolysis, perhaps via pro‐inflammatory cytokine mRNA expression.  相似文献   

16.
We recently demonstrated that luteal cells flow out from the ovary via lymphatic vessels during luteolysis. However, the regulatory mechanisms of the outflow of luteal cells are not known. Matrix metalloproteinases (MMPs) can degrade the extracellular matrix and basal membrane, and tissue inhibitors of matrix metalloproteinases (TIMPs) inhibit the activity of MMPs. To test the hypothesis that MMP expression in luteal cells is regulated by luteolytic factors, we investigated the effects of prostaglandin F2α (PGF), interferon γ (IFNG) and tumor necrosis factor α (TNF) on the mRNA expression of MMPs and TIMPs in cultured luteal cells. Luteal cells obtained from the CL at the mid-luteal stage (days 8–12 after ovulation) were cultured with PGF (0.01, 0.1, 1 μM), IFNG (0.05, 0.5, 5 nM) and TNF (0.05, 0.5, 0.5 nM) alone or in combination for 24 h. PGF and IFNG significantly increased the expression of MMP-1 mRNA. In addition, 1 μM PGF in combination with 5 nM IFNG stimulated MMP-1 and MMP-9 mRNA expression significantly more than either treatment alone. In contrast, IFNG significantly decreased the level of MMP-14 mRNA. The mRNA expression of TIMP-1, which preferentially inhibits MMP-1, was suppressed by 5 nM INFG. One μM PGF and 5 nM IFNG suppressed TIMP-2 mRNA expression. These results suggest a new role of MMPs: luteal MMPs stimulated by PGF and IFNG break down the extracellular matrix surrounding luteal cells, which accelerates detachment from the CL during luteolysis, providing an essential prerequisite for outflow of luteal cells from the CL to lymphatic vessels.  相似文献   

17.
This study investigated the chemotactic activity of equine CL at different stages of the oestrous cycle. The purpose of this was to ascertain whether luteal tissue itself contributes to the massive influx of leucocytes around the time of natural and induced luteal regression. Corpora lutea were collected at different stages of dioestrus and after treatment with PGF2alpha. Culture medium harvested after incubation of luteal tissue for 20 h was chemotactic for both polymorphonuclear and mononuclear cells in late dioestrus (before functional regression) as well as after natural and induced luteal regression. By contrast, midluteal tissue showed no chemotactic activity. This is the first report of the ability of equine luteal tissue actively to recruit inflammatory cells in vitro and supports our earlier findings that this infiltration starts prior to functional luteolysis. We hypothesise that this early influx of inflammatory cells may play an active role in luteal regression. Further research is needed to identify the specific chemotactic factor(s).  相似文献   

18.
Nitric oxide induces apoptosis in bovine luteal cells   总被引：1，自引：0，他引：1  
We previously showed in in vivo and in vitro studies that nitric oxide (NO) is engaged in luteolysis in cattle. Nitric oxide produced locally in the bovine corpus luteum (CL) inhibits progesterone (P4) synthesis and is suggested to be a component of the luteolytic cascade induced by uterine prostaglandin (PG) F2alpha. In the present study, the molecular mechanisms of NO action during structural luteolysis were studied in cultured bovine luteal cells (Days 15-17 of the estrous cycle). The effects of the NO donor (NONOate; 10(-4)M) on DNA fragmentation, cell viability, P4 production and caspase-3 activity were compared with those of PGF2alpha (10(-6)M). Moreover, mobilization of intracellular calcium [Ca2+]i and gene expressions of Fas-L, Fas, bcl-2, bax, and caspase-3 in the cells were determined by semi-quantitative RT-PCR after NONOate treatment. Caspase-3 activity was examined calorimetrically. Contrary to PGF2alpha NONOate decreased cell viability. DNA fragmentation after NONOate treatment increased by more than with PGF22alpha. NONOate increased mobilization of [Ca2+]i in the cells. Although the NO donor did not affect Fas-L and bcl-2 gene expression, it stimulated Fas and bax mRNA and caspase-3 expression. The ratio of bcl-2 to bax mRNA level decreased in the cells treated with NONOate. Moreover, NONOate stimulated caspase-3 activity more effectively than PGF2alpha. The overall results suggest that NO is a luteolytic factor that plays a crucial role in regulation of the estrous cycle in structural luteolysis by inducing apoptosis of luteal cells in cattle.  相似文献   

19.
Structural luteolysis occurs by apoptosis of luteal cells. The present study examined the effects of activators of well-characterized second messengers on Fas and caspase-3 mRNA expression and on P4 production in luteal cells in order to trace the pro- and anti-apoptotic factors in the bovine corpus luteum (CL). Cultured bovine mid luteal cells were treated for 24 h with a cyclic AMP analogue (8-bromo cyclic AMP; 8br-cAMP; 2.5 mM), a protein kinase C (PKC) activator (phorbol 12-myristate 13-acetate; PMA; 10 microM), or calcium ionophore (A23187; 10 microM). Fas and caspase-3 mRNA expression was inhibited by 8br-cAMP and PMA but was increased by A23187 (P<0.05). In addition, P4 production by bovine luteal cells was stimulated by 8br-cAMP and PMA, whereas it was inhibited by A23187, compared with untreated controls (P<0.05). The overall results suggest that cAMP and PKC suppress apoptosis in bovine luteal cells through inhibition of Fas and caspase-3 mRNA expression and through stimulation of P4 production. Therefore, substances that activate cAMP or PKC may act as survival factors in the bovine CL. Furthermore, substances that mobilize Ca2+ may act as apoptotic factors by stimulating Fas and caspase-3 expression in the bovine luteal cells.  相似文献   

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