Sirtuin 6 (SIRT6) Content and Hypoxia‐Inducible Factor 1‐Alpha (HIF‐1α) Expression in Human Umbilical Vein Endothelial Cells (HUVECs) in Response to Cyanidin‐3‐O‐ β‐glucoside (C3G): Pregnancy Complicated by Diabetes (PCD) vs. Normoglycemic Pregnancy (NP)

Abstract

Diabetes during pregnancy induces chronic hypoxia stress and excessive inflammatory response in placental tissue, particularly evident at the endothelial level. Hypoxia‐inducible factor 1‐alpha (HIF‐1α) is the oxygen‐sensitive subunit of the transcription factor HIF‐1, and its expression is increased in placentas from PCD. SIRT6, a chromatin‐associated protein with both histone deacetylase and mono‐ADP ribosyltransferase enzymatic activities, plays an important role in placental development and homeostasis. SIRT6 is believed to protect cells against oxidative stress, however, this mechanism of action in the human placenta has not been clarified enough. According to recent studies, SIRT6 may act as as a corepressor of the transcription factor HIF1α to control the expression of multiple glycolytic genes to regulate glucose homeostasis. A typical anthocyanin, C3G, a naturally occuring pigment belonging to the group of flavonoids (a subclass of the polyphenol family) was found to be potent SIRT6 activator. The aim of this study was to examine comparatively the effect of C3G on SIRT6 level in HUVECs isolated after PCD (White’s class C) vs. NP. In addition, HIF‐1α expressions were compared. HUVECs were isolated immediately after cesarean section delivery from PCD (N = 24) and NP (N = 24) patients (group I and group II, respectively). The umbilical cord segments was subjected to collagenase digestion and the HUVECs were cultured in vitro under normoxia. Upon reaching confluence, HUVECs were exposed to C3G (0.1, 1.0 and 10 μM) administered into the culture media in the respective subgroups A, B and C during the 7 day period. SIRT6 levels were determined in HUVECs culture lysates obtained in the days 1,3 and 7, using ELISA. After termination of the cultures immunostaining for HIF1α was performer with polyclonal antibody in formalin fixed and paraffin embedded sections. To estimate the mean expression of HIF1α, quantitative immunohostochemistry was applied. The initial HIF1α expression was estimated in the respective controls. After 3 and 7 days of the culture, in both groups the mean SIRT6 levels were significantly (p < 0.05) higher in the subgroups B and C, compared to the initial levels at day 1. However, the mean SIRT6 levels in group I were 5.7‐fold and 2.9‐fold lower (the subgroups B and C, respectively) than observed in euglycemic group II. The initial expression of HIF1α was significantly higher in PCD and amounted to 265% of the average value determned in NP (group II). The mean expression of HIF1α was significantly (p < 0.05) downregulated in group I (the subgroups B and C), whereas in group II was nearly unaffected. These results may suggest potential beneficial effect of SIRT6 activation by C3G in PCD with regard to counteracting oxidative stress and modulation of placental metabolic changes at endothelial level. It is likely that by attenuating HIF1α transcriptional output, SIRT6 promotes mitochondrial oxidative metabolism.Support or Funding InformationWUM grant: 2M2‐W1‐19