SIRT6 Is Decreased with Preterm Labor and Regulates Key Terminal Effector Pathways of Human Labor in Fetal Membranes1

Abstract

Preterm birth is a major determinant of neonatal mortality and morbidity, affecting approximately one-third of preterm births as a result of prelabor rupturing of membranes. Infection and inflammation have strong causal links to preterm delivery, resulting in the activation of nuclear factor-kappaB (NFKB) and its downstream targets. Human sirtuin (SIRT) 6, which has ADP-ribosyl transferase and deacetylase activity, exhibits anti-inflammatory actions. The aims of this study were to determine the effect of 1) human preterm labor on SIRT6 expression in human gestational tissue and 2) the effect in primary amnion cells of SIRT6 inhibition, using small interfering RNA (siRNA) on prolabor mediators. To determine the effect of human preterm labor on SIRT6 expression, human fetal membranes were collected from women at preterm at the time of Cesarean section (no labor; n = 9) and from women after spontaneous labor and delivery (n = 9). SIRT6 mRNA and protein expression were significantly lower in fetal membranes after spontaneous preterm labor. Transfection of primary amnion cells with SIRT6 siRNA was associated with an increase in IL-1beta-induced proinflammatory cytokine gene expression and release (IL6, IL8, TNF [TNF-alpha]), cyclooxygenase ([COX]-2; official symbol PTGS2) expression and subsequent prostaglandin (PGE(2) and PGF(2alpha)) release, and MMP9 gene expression and release of pro-MMP9. To determine whether SIRT6 affects NFKB transcriptional activity, primary amnion cells were transfected with NFKB tagged with luciferase and stimulated with IL1B. As expected, IL1B induced NFKB transcriptional activity. However, when cells were also cotransfected with a vector expressing SIRT6, there was a decrease in NFKB transcriptional activity. In conclusion, SIRT6 plays a role in regulating the terminal effector pathways of human labor and delivery via the NFKB pathway.

Sirtuin-6 Preserves R-spondin-1 Expression and increases Resistance of intestinal Epithelium to injury in Mice

“…Thus, the effect Evidence suggests that Sirt6 expression is inhibited in numerous pathological conditions associated with inflammation. For example, decreased Sirt6 expression in fetal membranes occurs in preterm labor (37 in each group) with DSS-induced colitis. Colonic tissues were collected on d 7 after induction of colitis with 3.5% DSS drinking water.…”

Human cathelicidin antimicrobial protein 18 (hCAP18/LL-37) is increased in foetal membranes and myometrium after spontaneous labour and delivery

“…To determine the expression of hCAP18 in foetal membranes and myometrium, immunohistochemistry (IHC) was performed on paraffin-embedded sections as described previously (Lim et al, 2013) using the IHC Select ® HRP Detection Set (Merck Millipore; Billerica, MA, USA). Briefly, sections were deparaffinised, followed by an antigen retrieval step (boiled in 10 mM of Tris, 1 mM of EDTA, pH 9.0 for 10 min followed by 20-min incubation) and then endogenous peroxidases were inactivated by adding 3% hydrogen peroxide in methanol for 10 min.…”

“…Western blotting was performed as we have previously described (Lim et al, 2013). Forty micrograms of protein was separated on 8-16% polyacrylamide gels (Bio-Rad Laboratories, Hercules, CA, USA) and transferred to nitrocellulose.…”

“…Extraction of RNA and quantitative RT-PCR (qRT-PCR) was performed as previously described (Lim et al, 2013). Total RNA was extracted from cells and tissues using TRIsure reagent according to the manufacturer's instructions (Bioline, Alexandria, NSW, Australia).…”

Class I to III Histone Deacetylases Differentially Regulate Inflammation-Induced Matrix Metalloproteinase 9 Expression in Primary Amnion Cells

“…20,22 Cells were transfected with 100 nmol/L SIRT1 or NS siRNA (Ambion, Austin, Texas) in DMEM/F-12 for 48 hours. The medium was then replaced with DMEM/F-12 with or without 1 ng/mL IL1b, and the cells were incubated at 37 C for an additional 20 hours.…”