Peroxisome proliferator‐activated receptor gamma ligands affect NF‐kB and cytokine synthesis in the porcine endometrium—An in vitro study

Abstract

Problem Cytokines, mediators of the immune response, are involved in the regulation of female reproductive processes during the estrous cycle and pregnancy. The present study aimed to investigate the effect of selected peroxisome proliferator‐activated receptor gamma (PPARγ) ligands on the expression of nuclear factor kappa B (NF‐κB) and selected cytokines, such as interleukin (IL)‐1β, ‐4, ‐6, ‐8, ‐10, and the leukemia inhibitory factor, in the porcine endometrium on days 10‐12 and 14‐16 of the estrous cycle (mid‐ and late luteal phase, respectively) or pregnancy (maternal recognition of pregnancy and beginning of implantation, respectively). Method of study Endometrial slices were incubated in vitro in the presence of PPARγ agonists, 15‐deoxy‐Δ12, 14‐prostaglandin J2 or rosiglitazone, and PPARγ antagonist T0070907. mRNA and protein levels in tissues were determined by real‐time PCR and Western blot. Results On days 10‐12 of the estrous cycle and days 14‐16 of pregnancy, PPARγ ligands enhanced the expression of NF‐κB, mRNA cytokines, and/or proteins. During the late luteal phase of the estrous cycle (days 14‐16) and maternal recognition of pregnancy (days 10‐12), PPARγ ligands inhibited the expression of NF‐κB, and they differentially affected the expression of mRNA and proteins of cytokines. Conclusion Our results indicate that PPARγ is engaged in the endometrial synthesis of NF‐κB and selected cytokines in pigs. The influence of PPARγ ligands on the tested components of the immune system varied subject to the physiological status of females, and it could be associated with differences in endometrial receptivity.

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“…This shows that MMP-2 was upregulated via the increased activity of the NF-kB. This result is consistent with the previous investigations, which showed that the NF-kB activation is increased proliferation, angiogenesis, and apoptosis in endometriosis (Kunicka et al, 2019, Calibasi-Kocal et al, 2019. It has been shown that NF-βB acts via regulation of the MMPs expression in the adhesion and invasion of endometriosis cells on the peritoneal surface (Lee et al, 2010, Gottschal et al, 2002.…”

Peroxisome proliferator‐activated receptor alpha regulates the expression of the immune response mediators in the porcine endometrium during the estrous cycle and early pregnancy

“…Furthermore, the ligands did not change IL‐6 mRNA expression, but the PPARα agonist increased IL‐6 protein expression in the mid‐luteal phase of the estrous cycle and at the beginning of implantation. The absence of any associations between mRNA and protein expression in the current study and in our previous research could be attributed to various biological and technical factors, such as differences in mRNA and protein stability, transcriptional and posttranscriptional regulation, and existing feedback, which can suppress mRNA expression through high protein expression and attenuate post‐transcriptional processes through high mRNA content . Further research is needed to determine biological factors that exert the greatest influence on the relationship between mRNA and protein expression …”

“…The procedures for tissue harvesting and culture were described previously . Briefly, tissue explants (200 ± 10 mg) were incubated in the M199 medium (Sigma Aldrich) supplemented with 0.1% BSA fraction V and antibiotics: nystatin (120 IU/mL; Sigma Aldrich) and gentamicin (40, 40 mg/mL; Sigma Aldrich).…”

“…After incubation, endometrial explants were homogenized in ice with a cold isolation buffer (Tris‐HCl [10 mmol/L], EDTA [5 mmol/L], NaCl [50 mmol/L], sodium pyrophosphate [30 mmol/L], sodium fluoride [50 mmol/L], sodium azide [0.02%], sodium orthovanadate [100 µmol/L], Triton X‐100 [1%], phenylmethanesulfonyl fluoride [1 mmol/L], leupeptin [2 µg/mL], aprotinin [2 µg/mL], and pepstatin [1 µg/mL]) according to a previously described procedure . The samples were tested individually in the Western blot analysis.…”

“…We have found that interferon γ (IFNγ) increases mRNA expression of PPARα in the porcine endometrium during early pregnancy (day 14) . Our recent research has shown that two other isoforms of PPAR—beta and gamma—regulate the mRNA expression and protein secretion of NF‐κB and selected pro‐ and anti‐inflammatory cytokines in the in vitro incubation of porcine endometrial explants during the mid‐ and late‐luteal phases of the estrous cycle as well as during early pregnancy . Taking into account previous reports, we hypothesized that PPARα changes the gene and protein expression of immune response mediators in porcine endometrial explants.…”