Palmitate increases Nur77 expression by modulating ZBP89 and Sp1 binding to the Nur77 proximal promoter in pancreatic β-cells

Nur77 is a stress sensor in pancreatic β-cells, which negatively regulates glucose-stimulated insulin secretion. We recently showed that a lipotoxic shock caused by exposure of β-cells to the saturated fatty acid palmitate strongly increases Nur77 expression. Here, using dual luciferase reporter assays and Nur77 promoter deletion constructs, we identified a regulatory cassette between −1534 and −1512 bp upstream from the translational start site mediating Nur77 promoter activation in response to palmitate exposure. Chromatin immunoprecipitation, transient transfection and siRNA-mediated knockdown assays revealed that palmitate induced Nur77 promoter activation involves Sp1 recruitment and ZBP89 release from the gene promoter.     

Two new polymorphic markers in the human proα2(1) collagen gene

Summary Structural defects in the human type 1 collagen genes are known to be the cause of several inherited disorders of connective tissue, such as osteogenesis imperfecta. The analysis and prenatal diagnosis of these disorders would be facilitated by establishing a set of polymorphic markers at these gene loci. We have previously reported the presence of an Msp 1 restriction fragment length polymorphism in the pro2(1) collagen genes of several Southern African populations (Grobler-Rabie et al., in press). This report describes the detection of a Bgl II and an EcoRI polymorphism in the pro2 gene of South African Blacks.     

Cyclic stretch induces cyclooxygenase-2 gene expression in vascular endothelial cells via activation of nuclear factor kappa-β

Vascular endothelial cells respond to biomechanical forces, such as cyclic stretch and shear stress, by altering gene expression. Since endothelial-derived prostanoids, such as prostacyclin and thromboxane A₂, are key mediators of endothelial function, we investigated the effects of cyclic stretch on the expression of genes in human umbilical vein endothelial cells controlling prostanoid synthesis: cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2), prostacyclin synthase (PGIS) and thromboxane A₂ synthase (TXAS). COX-2 and TXAS mRNAs were upregulated by cyclic stretch for 24 h. In contrast, PGIS mRNA was decreased and stretch had no effect on COX-1 mRNA expression. We further show that stretch-induced upregulation of COX-2 is mediated by activation of the NF-κβ signaling pathway.     

Palmitic acid suppresses apolipoprotein M gene expression via the pathway of PPARβ/δ in HepG2 cells

It has been demonstrated that apolipoprotein M (APOM) is a vasculoprotective constituent of high density lipoprotein (HDL), which could be related to the anti-atherosclerotic property of HDL. Investigation of regulation of APOM expression is of important for further exploring its pathophysiological function in vivo. Our previous studies indicated that expression of APOM could be regulated by platelet activating factor (PAF), transforming growth factors (TGF), insulin-like growth factor (IGF), leptin, hyperglycemia and etc., in vivo and/or in vitro. In the present study, we demonstrated that palmitic acid could significantly inhibit APOM gene expression in HepG2 cells. Further study indicated neither PI-3 kinase (PI3K) inhibitor LY294002 nor protein kinase C (PKC) inhibitor GFX could abolish palmitic acid induced down-regulation of APOM expression. In contrast, the peroxisome proliferator-activated receptor beta/delta (PPAR_β/δ) antagonist GSK3787 could totally reverse the palmitic acid-induced down-regulation of APOM expression, which clearly demonstrates that down-regulation of APOM expression induced by palmitic acid is mediated via the PPAR_β/δ pathway.     

Feedback regulation of the α2(1) collagen gene via the Mek-Erk signaling pathway

The extracellular matrix (ECM) provides the microenvironment that is pivotal for cell growth, motility, attachment, and differentiation. Advances in cell culture techniques have led to the development of cell-derived ECM model systems that are more reflective of the in vivo architecture of the ECM in tissue. In this study, a fibroblast-derived ECM (fd-ECM) was used to study the feedback regulation of type I collagen synthesis in fibroblasts. Fibroblasts plated on a preformed fd-ECM showed a significant decrease in the production of type I collagen and pro-α2(1) collagen mRNA compared to cells grown in the absence of a matrix. Function-blocking antibodies showed that this downregulation of type I collagen gene expression is mediated via α2β1 integrin. The use of several kinase inhibitors and a dominant negative ras construct (N17Ras) showed that the matrix-mediated downregulation of COL1A2 occurs via Ras-dependent activation of the MEK/ERK signaling pathway. Deletion analysis of the COL1A2 promoter implicated the region between -375 and -107 as containing a potential matrix responsive element. The use of Sp1 siRNA demonstrated that Sp1 is an important mediator of this feedback inhibition. This study provides some new insights into the feedback regulation of COL1A2 gene expression.     

TGF-β1 induces type I collagen deposition in granulosa cells via the AKT/GSK-3β signaling pathway-mediated MMP1 down-regulation

Type I collagen is the most abundant extracellular matrix (ECM) protein in the mammalian ovary, and comprises two COL1A1 subunits and one COL1A2 subunit. Matrix metalloproteinase 1 (MMP1) is a typical collagenase of type I collagen, that can be detected in ovarian follicles and early corpus luteum. Previous studies demonstrated that MMP1-mediated degradation of type I collagen plays a functional role in regulating corpus luteum formation, and transforming growth factor β1 (TGF-β1) inhibits luteinization and progesterone production in granulosa cells (GCs). Whether TGF-β1 regulates the expression of MMP1, COL1A1, or the deposition of type I collagen during corpus luteum formation remains to be elucidated. This study aimed to investigate the molecular mechanisms through which TGF-β1 regulates MMP1 expression and type I collagen deposition in GCs. Our results show that TGF-β1 upregulates COL1A1 expressions and downregulates MMP1 expression. Inhibition approaches, including pharmacological inhibitors such as p38 inhibitor (SB203580), ERK1/2 inhibitor (U0126), AKT inhibitor (LY294002), and GSK-3β inhibitor (LiCl), as well as knockdown using siRNA specific to these genes, were used. Our results suggest that TGF-β1 decreases MMP1 production via an ALK5-mediated AKT/GSK-3β-dependent signaling pathway, and a decrease in MMP1 levels and an increase in COL1A1 levels synergistically promote type I collagen deposition in GCs. Collectively, these findings provide novel insights into the underlying molecular mechanisms by which TGF-β1 upregulates type I collagen deposition in GCs.     

Megalin is downregulated via LPS-TNF-α-ERK1/2 signaling pathway in proximal tubule cells

Expression and function of megalin, an endocytic receptor in proximal tubule cells (PTCs), are reduced in diabetic nephropathy, involved in the development of proteinuria/albuminuria. Lipopolysaccharide (LPS) is chronically increased in diabetic sera, by the mechanism called metabolic endotoxemia. We investigated low-level LPS-mediated signaling that regulates megalin expression in immortalized rat PTCs (IRPTCs). Incubation of the cells with LPS (10 ng/ml) for 48 h suppressed megalin protein expression and its endocytic function. TNF-α mRNA expression was increased by LPS treatment, and knockdown of the mRNA with siRNA inhibited LPS-mediated downregulation of megalin mRNA expression at the 24-h time point. Incubation of IRPTCs with exogenous TNF-α also suppressed megalin mRNA and protein expression at the 24- and 48-h time points, respectively. MEK1 inhibitor PD98059 competed partially but significantly TNF-α-mediated downregulation of megalin mRNA expression. Collectively, low-level LPS-mediated TNF-α-ERK1/2 signaling pathway is involved in downregulation of megalin expression in IRPTCs.     

Length polymorphism in the pro α2(I) collagen gene: an alternative explanation in a case of Marfan syndrome

Summary A 38 base pair (bp) insertion in the pro 2(I) collagen gene (COL1A2) of a patient with Marfan syndrome has been proposed to be the possible cause of the disease (Henke et al. 1985). However, analysis of this insertion in DNA from the patient in question and from random normal individuals reveals it to be a common polymorphism. We suggest that the 38 bp insertion is not related to the primary defect in this case of Marfan syndrome.     

SpαI/65 hereditary elliptocytosis in Calabria (southern Italy)

The ^I/65 variant of spectrin has been described in black people, in North Africans and recently in two southern Italian families. This variant is associated in the heterozygous state with mild Hereditary Elliptocytosis (HE) and the molecular basis of the defect is invariably the duplication of TTG at codon 154 of the spectrin gene. The present study reports the identification of five Calabrian families with SP^I/65 HE and their distribution in the population.     

The α2(XI) collagen gene lies within 8 kb of Pb in the proximal portion of the murine major histocompatibility complex

A number of serious hereditary disorders are now known to be associated with defective expression of collagen genes, and these findings have underscored the important and varied roles that the collagen family of genes must play during normal mammalian development. Although the activities of genes encoding the quantitatively major types of collagen are fairly well characterized, functions of the many minor types of collagen remain a matter of speculation. As a first step toward a functional analysis of type XI collagen, a member of this class of poorly understand minor collagen proteins which is expressed primarily in hyaline cartilage, we have used human probes for the gene encoding the protein's 2-subunit (COL11A2) to isolate and map homologous murine DNA sequences. Our results demonstrate that Col11a-2 is embedded within the major histocompatibility complex (MHC), within 8.4 kb of the class II pseudogene locus, Pb, and confirm that human and murine 2(XI) collagen genes are located in very similar genomic environments. The conserved location of these genes raises the possibility that type XI collagen genes may contribute to one or more of the diverse hereditary disorders known to be linked to the MHC in mouse and human.