Novel Insights on the Corpus Luteum Function: Role of Vaspin on Porcine Luteal Cell Angiogenesis, Proliferation and Apoptosis by Activation of GRP78 Receptor and MAP3/1 Kinase Pathways

Abstract

Formation and limited lifespan of corpus luteum (CL) are important for proper ovarian periodicity and fertility. Failed vascularization, imbalance between proliferation and apoptosis leads to luteal phase deficiency and infertility. The aim of this study was to examine the effect of vaspin on angiogenesis, apoptosis and proliferation as well as the involvement of 78-kDa glucose-regulated protein receptor (GRP78) and mitogen-activated kinase (MAP3/1) in these processes. Porcine luteal cells were incubated with vaspin (0.1–10 ng/mL) for 24 h to 72 h and then mRNA and protein expression of angiogenesis: vascular endothelial growth factor (VEGFA), fibroblast growth factor 2 (FGF2), angiopoietin 1 (ANGPT1), VEGFA receptors (VEGFR1, VEGFR2), apoptosis: caspase 3, bcl-2-like protein 4 (BAX), B-cell lymphoma (BCL2), and proliferation: proliferating cells nuclear antigen (PCNA), cyclin A factors as well as secretion of VEGFA, FGF2, ANGT1 were measured by real-time polymerase chain reaction (PCR), immunoblotting and enzyme-linked immunosorbent assay (ELISA), respectively. Moreover, apoptosis was assessed by caspase activity using the Caspase-Glo 3/7 assay, while proliferation was by alamarBlue. We found that vaspin enhanced luteal cell angiogenesis, proliferation, and significantly decreased apoptosis. Additionally, using GRP78 siRNA and the pharmacological inhibitor of MAP3/1 (PD98059), we observed that the effect of vaspin was reversed to the control level in all investigated processes. Taken together, our results suggest that vaspin is a new regulator of female fertility by direct regulation of CL formation and maintenance of luteal cell function.

Review: Vaspin (SERPINA12) Expression and Function in Endocrine Cells

“…Inappropriate vascularization leads to luteal phase deficiency and consequently miscarriages. The stimulatory effect of vaspin on VEGFA, FGF2, and ANGPT1 secretion and mRNA expression, as well as VEGFA receptor protein expression, was previously demonstrated [ 81 ]. As a molecular mechanism of vaspin regulatory action, prostaglandin synthesis and angiogenesis were shown to activate GRP78 and MAP3/1.…”

“…Additionally, angiogenesis is another important process for CL function maintenance. The development of capillaries from pre-existing blood vessels is essential for the formation, function, and establishment of CL as an endocrine gland by supplying cholesterol for P4 production and regulation by vascular endothelial growth factor (VEGFA), fibroblast growth factor 2 (FGF2), and angiopoietin 1 (ANGPT1) [ 81 ]. Inappropriate vascularization leads to luteal phase deficiency and consequently miscarriages.…”

“…As a molecular mechanism of vaspin regulatory action, prostaglandin synthesis and angiogenesis were shown to activate GRP78 and MAP3/1. Additionally, vaspin was reported to be a pro-survival factor for granulosa and luteal cells by increasing the ratio of B-cell lymphoma 2/ BCL2-associated X (BCL2/BAX), proliferating cell nuclear antigen (PCNA), cyclin D, and A protein and mRNA expression, and decreasing caspases 3, 8, and 9 mRNA, protein expression, and activity [ 59 , 81 ]. Endocrine function maintenance and regression and ovarian follicle growth and their ovulation are necessary processes for CL formation.…”

“…Thus, vaspin could be used to improve oestrus/menstrual cycle synchronization and to enable reproductive optimization [ 59 , 80 ]. Additionally, it is a luteotropic factor in luteal cells, which may prevent luteolysis of CL and luteal phase deficiency, causes of infertility and miscarriages [ 58 , 81 ]. Vaspin also improved oocyte maturation [ 78 ], which in the future may refine in vitro fertilization protocols.…”

Expression and Impact of Vaspin on In Vitro Oocyte Maturation through MAP3/1 and PRKAA1 Signalling Pathways

“…In porcine CL cells, vaspin enhances steroidogenesis by increasing level of P4 and oestradiol, as well as enzymes participating in its synthesis via 78-kDa glucose-regulated protein (GRP78) receptors and the PKA signalling pathway. Moreover, vaspin exerts a positive impact on CL angiogenesis connected with upregulation in mRNA expression and the secretion of vascular and endothelial growth factors and angiopoietin 1 by GRP78 and MAP3/1 activation [24]. Importantly, the effect of vaspin on oocyte maturation has not been studied in pigs or other species.…”