Bardet-Biedl syndrome obesity: BBS4 regulates cellular ER stress in early adipogenesis.

BACKGROUND: Bardet-Biedl syndrome (BBS) is an autosomal recessive ciliopathy, presenting with early obesity onset. The etiology of BBS obesity involves both central and peripheral defects, through mechanisms mostly yet to be deciphered. We previously showed BBS4 expression in adipogenesis, peaking at day 3 of differentiation. Obesity is characterized by cellular stress which promotes pathological consequences. AIMS: We set out to test a possible role of BBS4 in adipocyte endoplasmic reticulum (ER) stress-induced unfolding protein response (UPR). METHODS: BBS4 silenced (SiBBS4) and overexpressing (OEBBS4) pre-adipocyte murine cell lines were subjected to ER-stress induction (Tunicamycin, TM) during adipogenesis. ER-stress UPR was analyzed at the transcript, protein and biochemical levels (microscopy, immunocytochemistry, western blotting, quantitative RT-PCR and X-box binding protein 1 (XBP-1) splicing). RESULTS: In silico analysis showed that BBS4 harbors an ER localization sequences indicative of ER localization. We verified BBS4's ER localization in adipocytes by immunocytochemistry and cellular protein fractionation. Furthermore, we demonstrated that BBS4 expression is significantly up-regulated by ER-stress, as indicated by protein and transcript levels. SiBBS4 adipocytes exhibited swollen ER typical to ER-stress and significant XBP-1 down-regulation at day 3 of differentiation. Following ER-stress, SiBBS4 adipocytes exhibited XBP-1 ER retention, failure to translocate to the nucleus and depletion of the nuclear active cleaved ATF6α. BBS4 did not alter ATF6α processing by S1P and S2P in the Golgi. Notably, SiBBS4 cells demonstrated significant reduction in the downstream activated phospho-IRE1α, independent of ER-stress. CONCLUSIONS: At day 3 of adipogenesis, coinciding with the timing of its peak expression, BBS4 is localized to the ER and is involved in the ER stress response and trafficking. BBS4 depletion results in swollen ER with impaired intracellular nucleus translocation of XBP-1 and ATF6α. Thus, BBS4 affects the ER stress response in early adipogenesis, altering ER stress responsiveness and the adipocyte ER phenotype.

Maternal and neonatal irisin precursor gene FNDC5 polymorphism is associated with preterm birth.

Irisin is a novel secreted myokine, encoded by the fibronectin type III domain-containing protein 5 (FNDC5) precursor gene. Irisin plays a role in the female reproductive system in pregnancy and in embryonic development, and is associated with fetal size. It is expressed in the ovary, placenta and neonatal cord serum. We studied whether maternal and neonatal FNDC5 genetic polymorphisms are associated with preterm birth (PTB). Blood for DNA analysis was collected from Israeli mothers (n = 315) and from umbilical veins of their respected idiopathic preterm (24-36 weeks) and control term (>37 weeks) newborns (n = 161). Genotypes of maternal and neonatal FNDC5 polymorphisms (rs726344 and rs1746661) were determined by restriction fragment length polymorphism analysis. Genotype-phenotype associations were analyzed using SPSS program. The Frequency of FNCD5 rs726344 G alleles in the Israeli cohort is 82%. We found significant FNCD5 rs726344 genotype frequencies control and PTB groups. Women bearing the FNDC5 rs726344 GG genotype had 2.18 fold ([CI] 1.193-4.008, p = 0.01) higher chance to deliver at term compared to both AG and AA genotypes (adjusting to age, gravidity, parity, weight percentile per gestational age and gender of newborn). Neonates carrying the FNDC5 rs726344 GG genotype had 2.24 fold ([CI] 0.979-5.134, p = 0.05) higher chance to be born at term compared to either AG or AA genotypes (adjusting to parity, previous abortions and weight percentile per gestational age). There was no significant association of the rs1746661 polymorphism with PTB. Thus, we determined FNDC5 polymorphisms frequencies in the Israeli population and demonstrated that maternal and neonatal FNDC5 rs726344 polymorphism is significantly associated with increased risk for PTB.

Pancreatic stellate cell activation is regulated by fatty acids and ER stress.

INTRODUCTION: Pancreatic pathologies are characterized by a progressive fibrosis process. Pancreatic stellate cells (PSC) play a crucial role in pancreatic fibrogenesis. Endoplasmic reticulum (ER) stress emerges as an important determinant of fibrotic remodeling. Overload of fatty acids (FA), typical to obesity, may lead to lipotoxic state and cellular stress. AIM: To study the effect of different lipolytic challenges on pancreatic ER stress and PSC activation. METHODS: Primary PSCs were exposed to different FAs, palmitate (pal) and oleate (ole), at pathophysiological concentrations typical to obese state, and in acute caerulein-induced stress (cer). PSC activation and differentiation were analyzed by measuring fat accumulation (oil-red staining and quantitation), proliferation (cells count) and migration (wound- healing assay). PSC differentiation markers (α-sma, fibronectin, tgf-ß and collagen secretion), ER stress unfolded protein response and immune indicators (Xbp1, CHOP, TNF-α, IL-6) were analyzed at the transcript and protein expression levels (quantitative RT-PCR and western blotting). RESULTS: PSC exposure to pal and ole FAs (500µM) increased significantly fat accumulation. Proliferation and migration analysis demonstrated that ole FA retained PSC activation, while exposure to pal FA significantly halted proliferation rate and delayed migration. Cer significantly augmented PSC differentiation markers α- sma, fibronectin and collagen, and ER stress and inflammation markers including Xbp1, CHOP, TNF-α and IL-6. The ole FA treatment significantly elevated PSC differentiation markers α-sma, fibronectin and collagen secretion. PSC ER stress was demonstrated following pal treatment with significant elevation of Xbp1 splicing and CHOP levels. CONCLUSION: Exposure to pal FA halted PSC activation and differentiation and elevated ER stress markers, while cer and ole exposure significantly induced activation, differentiation and fibrosis. Thus, dietary FA composition should be considered and optimized to regulate PSC activation and differentiation in pancreatic pathologies.