The In Vitro Effect of Steroid Hormones, Arachidonic Acid, and Kinases Inhibitors on Aquaporin 1, 2, 5, and 7 Gene Expression in the Porcine Uterine Luminal Epithelial Cells during the Estrous Cycle.

Damian Tanski,Malgorzata Tanska,Mariusz T Skowronski,Ewa Lepiarczyk,Agnieszka Skowronska

doi:10.3390/cells10040832

Damian Tanski, Malgorzata Tanska + Show 3 more

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https://doi.org/10.3390/cells10040832

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Abstract

Aquaporins (AQPs) are integral membrane proteins, which play an important role in water homeostasis in the uterus. According to the literature, the expression of aquaporins in reproductive structures depends on the local hormonal milieu. The current study investigated the effect of selected PKA kinase inhibitor H89 and MAPK kinase inhibitor PD98059, on the expression of AQP1, 2, 5, and 7, and steroid hormones (E2), progesterone (P4), and arachidonic acid (AA) in the porcine endometrium on days 18–20 and 2–4 of the estrous cycle (the follicular phase where estrogen and follicle-stimulating hormone (FSH) are secreted increasingly in preparation for estrus and the luteal phase where the ovarian follicles begin the process of luteinization with the formation of the corpus luteum and progesterone secretion, respectively). The luminal epithelial cells were incubated in vitro in the presence of the aforementioned factors. The expression of mRNA was determined by the quantitative real-time PCR technique. In general, in Experiment 1, steroid hormones significantly increased expression of AQP1, 2, and 5 while arachidonic acid increased expression of AQP2 and AQP7. On the other hand, MAPK kinase inhibitor significantly decreased the expression of AQP1 and 5. In Experiment 2, E2, P4, or AA combined with kinase inhibitors differentially affected on AQPs expression. E2 in combination with PKA inhibitor significantly decreased expression of AQP1 but E2 or P4 combined with this inhibitor increased the expression of AQP5 and 7. On the contrary, E2 with PD98059 significantly increased AQP5 and AQP7 expression. Progesterone in combination with MAPK kinase inhibitor significantly downregulated the expression of AQP5 and upregulated AQP7. Arachidonic acid mixed with H89 or PD98059 caused a decrease in the expression of AQP5 and an increase of AQP7. The obtained results indicate that estradiol, progesterone, and arachidonic acid through PKA and MAPK signaling pathways regulate the expression of AQP1 and AQP5 in the porcine luminal epithelial cells in the periovulatory period.

Highlights

The endometrium undergoes diverse cell proliferation, growth, and apoptosis cycles, as a function of the estrous cycle and pregnancy
The mRNA expression of AQP2 in the porcine uterine luminal epithelial cells on days 18–20 and days 2–4 of the estrous cycle was significantly upregulated by P4, arachidonic acid (AA) (Figure 2A, p < 0.05) and E2, P4 as well as AA (Figure 2B, p < 0.05), respectively
Progesterone significantly increased the expression of AQP5 mRNA in the porcine luminal epithelial cells during the follicular phase of the estrous cycle (Figure 3A, p < 0.05)

Summary

Introduction

The endometrium undergoes diverse cell proliferation, growth, and apoptosis cycles, as a function of the estrous cycle and pregnancy. Aquaporins (AQPs) are considered to be important regulators of water homeostasis for normal uterus function, participating in water movement at an intraluminal, interstitial, and capillary level during the estrous cycle, implantation period, and parturition, creating the proper fluid microenvironment in the uterus [2,3,4]. Since their discovery in the uterus of mammals, they have been intensively studied, using molecular and pharmacological methods [5,6,7]. While it is known that some of the uterine AQP genes and proteins are regulated by E2 and/or P4 or other factors, [9,10,11] much remains to be learned about how different AQPs can be regulated

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