TCF7L2 (Transcription Factor 7-Like 2) Regulation of GATA6 (GATA-Binding Protein 6)-Dependent and -Independent Vascular Smooth Muscle Cell Plasticity and Intimal Hyperplasia.

Objective- TCF7L2 (transcription factor 7-like 2) is a Wnt-regulated transcription factor that maintains stemness and promotes proliferation in embryonic tissues and adult stem cells. Mice with a coronary artery disease-linked mutation in Wnt-coreceptor LRP6 (LDL receptor-related protein 6) exhibit vascular smooth muscle cell dedifferentiation and obstructive coronary artery disease, which are paradoxically associated with reduced TCF7L2 expression. We conducted a comprehensive study to explore the role of TCF7L2 in vascular smooth muscle cell differentiation and protection against intimal hyperplasia. Approach and Results- Using multiple mouse models, we demonstrate here that TCF7L2 promotes differentiation and inhibits proliferation of vascular smooth muscle cells. TCF7L2 accomplishes these effects by stabilization of GATA6 (GATA-binding protein 6) and upregulation of SM-MHC (smooth muscle cell myosin heavy chain) and cell cycle inhibitors. Accordingly, TCF7L2 haploinsufficient mice exhibited increased susceptibility to injury-induced hyperplasia, while mice overexpressing TCF7L2 were protected against injury-induced intimal hyperplasia compared with wild-type littermates. Consequently, the overexpression of TCF7L2 in LRP6 mutant mice rescued the injury-induced intimal hyperplasia. Conclusions- Our novel findings imply cell type-specific functional role of TCF7L2 and provide critical insight into mechanisms underlying the pathogenesis of intimal hyperplasia.

Cholesterol-Independent Suppression of Lymphocyte Activation, Autoimmunity, and Glomerulonephritis by Apolipoprotein A-I in Normocholesterolemic Lupus-Prone Mice.

Apolipoprotein (Apo)A-I, the major lipid-binding protein of high-density lipoprotein, can prevent autoimmunity and suppress inflammation in hypercholesterolemic mice by attenuating lymphocyte cholesterol accumulation and removing tissue-oxidized lipids. However, whether ApoA-I mediates immune-suppressive or anti-inflammatory effects under normocholesterolemic conditions and the mechanisms involved remain unresolved. We transferred bone marrow from systemic lupus erythematosus (SLE)-prone Sle123 mice into normal, ApoA-I-knockout (ApoA-I(-/-)) and ApoA-I-transgenic (ApoA-I(tg)) mice. Increased ApoA-I in ApoA-I(tg) mice suppressed CD4(+) T and B cell activation without changing lymphocyte cholesterol levels or reducing major ApoA-I-binding oxidized fatty acids. Unexpectedly, oxidized fatty acid peroxisome proliferator-activated receptor γ ligands 13- and 9-hydroxyoctadecadienoic acid were increased in lymphocytes of autoimmune ApoA-I(tg) mice. ApoA-I reduced Th1 cells independently of changes in CD4(+)Foxp3(+) regulatory T cells or CD11c(+) dendritic cell activation and migration. Follicular helper T cells, germinal center B cells, and autoantibodies were also lower in ApoA-I(tg) mice. Transgenic ApoA-I also improved SLE-mediated glomerulonephritis. However, ApoA-I deficiency did not have the opposite effects on autoimmunity or glomerulonephritis, possibly as the result of compensatory increases in ApoE on high-density lipoprotein. We conclude that, although compensatory mechanisms prevent the proinflammatory effects of ApoA-I deficiency in normocholesterolemic mice, increasing ApoA-I can attenuate lymphocyte activation and autoimmunity in SLE independently of cholesterol transport, possibly through oxidized fatty acid peroxisome proliferator-activated receptor γ ligands, and it can reduce renal inflammation in glomerulonephritis.

Nonalcoholic fatty liver disease induced by noncanonical Wnt and its rescue by Wnt3a.

Nonalcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease, which begins with isolated steatosis and advances to nonalcoholic steatohepatitis (NASH), steatofibrosis, and cirrhosis. The pathways involved in disease progression are not understood. Loss-of-function mutations in Wnt coreceptor LDL receptor-related protein 6 (LRP6) underlie early-onset atherosclerosis, metabolic risk factors, and NAFLD in humans by unknown mechanisms. We generated mice with the human disease-associated LRP6(R611C) mutation and phenotypically characterized their liver. Homozygote LRP6(R611C) (LRP6(mut/mut)) mice exhibited both steatohepatitis and steatofibrosis. These traits were associated with increased activity of the noncanonical Wnt/Ras homolog family member A, Rho-associated protein kinase 2, and PKC-α/-µ pathways. Accordingly, there was increased TGF-ß1 activity, coupled with enhanced expression of smooth muscle α-actin and vimentin that colocalized with albumin in LRP6(mut/mut) mouse liver. LRP6 knockdown reprogramed HepG2 cells to express both these markers, linking impaired Wnt signaling with hepatocyte transdifferentiation. The causal link between altered Wnt signaling and NASH was established by normalization of the disease pathways and rescue of the liver traits by Wnt3a administration to LRP6(mut/mut) mice. Thus, this study identifies diverse disease pathways that underlie a spectrum of NASH-related liver diseases and are linked by a single human genetic variant. LRP6 and noncanonical Wnt pathways are important potential therapeutic targets against NASH.

Autoimmune-mediated reduction of high-density lipoprotein-cholesterol and paraoxonase 1 activity in systemic lupus erythematosus-prone gld mice.

OBJECTIVE: To characterize modifications of high-density lipoprotein (HDL) in autoimmune gld mice that may be relevant to premature atherosclerosis in systemic lupus erythematosus, and to assess their relationship to specific aspects of autoimmune disease. METHODS: HDL cholesterol (HDL-C), apolipoprotein A-I (Apo A-I), paraoxonase 1 (PON1) activity, hepatic gene expression, and HDL biogenesis were measured in aging female gld and wild-type congenic mice. Autoantibodies, lymphoid organs, and cytokines were analyzed by enzyme-linked immunosorbent assay, flow cytometry, and multiplex assay, respectively. RESULTS: Plasma HDL-C, HDL Apo A-I, and HDL-associated PON1 activity were reduced in aging gld mice in association with the development of autoimmunity, independent of changes in hepatic Apo A-I and PON1 expression or HDL biogenesis. Hepatic induction of the acute-phase reactant serum amyloid A1 resulted in its incorporation into HDL in gld mice. Deletion of the lipid-sensitive receptor G2A in gld mice (G2A-/- gld) attenuated reductions in HDL-C and PON1 activity without altering hepatic Apo A-I and PON1 expression, HDL biogenesis, or levels of acute-phase proinflammatory cytokines. Plasma anti-Apo A-I autoantibodies were elevated in aging gld mice commensurate with detectable increases in Apo A-I immune complexes. Autoantibody levels were lower in aging G2A-/- gld mice compared with gld mice, and anti-Apo A-I autoantibody levels were significantly related to HDL-C concentrations (r=-0.645, P<0.00004) and PON1 activity (r=-0.555, P<0.0007) among autoimmune gld and G2A-/- gld mice. CONCLUSION: Autoantibodies against Apo A-I contribute to reducing HDL-C and PON1 activity in autoimmune gld mice independently of hepatic HDL biogenesis, suggesting that functional impairment and premature clearance of HDL immune complexes may be principal mechanisms involved.

ApoE-dependent modulation of HDL and atherosclerosis by G2A in LDL receptor-deficient mice independent of bone marrow-derived cells.

OBJECTIVE: Deletion of the lysophospholipid-sensitive receptor, G2A, in low-density lipoprotein receptor knockout (LDLR(-/-)) mice elevates plasma high-density lipoprotein (HDL) cholesterol and suppresses atherosclerosis. However, chemotactic action of G2A in monocytes/macrophages, in addition to its modulatory effect on HDL, may contribute to the proatherogenic action of G2A. METHODS AND RESULTS: We determined that deletion of G2A in LDLR(-/-) mice increases the ApoA1, ApoE, and cholesterol content of plasma HDL fractions. Hepatocytes were shown to express G2A and hepatocytes from G2A-deficient LDLR(-/-) mice secreted more ApoA1 and ApoE in HDL fractions compared to their G2A-sufficient counterparts. The atheroprotective and HDL modulatory effects of G2A deficiency were dependent on the presence of ApoE, as deletion of G2A in ApoE(-/-) and ApoE(-/-)LDLR(-/-) mice failed to raise HDL and did not suppress atherosclerosis. G2A deficiency in bone marrow-derived cells of LDLR(-/-) mice had no effect on atherosclerosis or HDL, whereas G2A deficiency in resident tissues was sufficient to raise HDL and suppress atherosclerosis. CONCLUSIONS: These data demonstrate that the chemotactic function of G2A in bone marrow-derived monocytes does not modulate atherosclerosis in LDLR(-/-) mice and suggest an ApoE-dependent function for G2A in the control of hepatic HDL metabolism that might contribute to its proatherogenic action.

Deletion of the G2A receptor fails to attenuate experimental autoimmune encephalomyelitis.

Lysophosphatidylcholine (LPC) is a chemotactic lysolipid produced during inflammation by the hydrolytic action of phospholipase A(2) enzymes. LPC stimulates chemotaxis of T cells in vitro through activation of the G protein-coupled receptor, G2A. This has led to the proposition that G2A contributes to the recruitment of T cells to sites of inflammation and thus promotes chronic inflammatory autoimmune diseases associated with the generation and subsequent tissue infiltration of auto-antigen-specific effector T cells. However, one study suggests that G2A may negatively regulate T cell proliferative responses to antigen receptor engagement and thereby attenuates autoimmunity by reducing the generation of autoreactive T cells. To address the relative contribution of these G2A-mediated effects to the pathophysiology of T cell-mediated autoimmune disease, we examined the impact of G2A inactivation on the onset and severity of murine experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis (MS). Wild type (G2A(+/+)) and G2A-deficient (G2A(-/-)) C57BL/6J mice exhibited a similar incidence and onset of disease following immunization with MOG(35-55) peptide. Disease severity was only moderately reduced in G2A(-/-) mice. Similar numbers of MOG(35-55) specific T cells were generated in secondary lymphoid organs of MOG(35-55)-immunized G2A(+/+) and G2A(-/-) mice. Comparable numbers of T cells were detected in spinal cords of G2A(+/+) and G2A(-/-) mice. We conclude that the proposed anti-proliferative and chemotactic functions of G2A are not manifested in vivo and therefore therapeutic targeting of G2A is unlikely to be beneficial in the treatment of MS.

Distribution pattern of phthirapterans infesting certain common Indian birds.

The prevalence and frequency distribution patterns of 10 phthirapteran species infesting house sparrows, Indian parakeets, common mynas, and white breasted kingfishers were recorded in the district of Rampur, India, during 2004-05. The sample mean abundances, mean intensities, range of infestations, variance to mean ratios, values of the exponent of the negative binomial distribution, and the indices of discrepancy were also computed. Frequency distribution patterns of all phthirapteran species were skewed, but the observed frequencies did not correspond to the negative binomial distribution. Thus, adult-nymph ratios varied in different species from 1:0.53 to 1:1.25. Sex ratios of different phthirapteran species ranged from 1:1.10 to 1:1.65 and were female biased.

Proteomics data on MAP Kinase Kinase 3 knock out bone marrow derived macrophages exposed to cigarette smoke extract.

This data article reports changes in the phosphoproteome and total proteome of cigarette smoke extract (CSE) exposed WT and MAP Kinase Kinase 3 knock out (MKK3-/-) bone marrow derived macrophages (BMDM). The dataset generated is helpful for understanding the mechanism of CSE induced inflammation and the role of MAP kinase signaling pathway. The cellular proteins were labeled with isobaric tags for relative and absolute quantitation (iTRAQ®) reagents and analyzed by LC-MS/MS. The standard workflow module for iTRAQ® quantification within the Proteome Discoverer was utilized for the data analysis. Ingenuity Pathway Analysis (IPA) software and Reactome was used to identify enriched canonical pathways and molecular networks (Mannam et al., 2016) [1]. All the associated mass spectrometry data has been deposited in the Yale Protein Expression Database (YPED) with the web-link to the data: http://yped.med.yale.edu/repository/ViewSeriesMenu.do;jsessionid=6A5CB07543D8B529FAE8C3FCFE29471D?series_id=5044&series_name=MMK3+Deletion+in+MEFs.

The Protective Effect of Transcription Factor 7-Like 2 Risk Allele rs7903146 against Elevated Fasting Plasma Triglyceride in Type 2 Diabetes: A Meta-Analysis.

BACKGROUND: The results from published studies regarding association of transcription factor 7-like 2 (TCF7L2) variant rs7903146 with dyslipidemia have been conflicting and inconclusive. METHODS: We carried out a meta-analysis that aimed to investigate the association of the rs7903146 variant with plasma lipid levels using electronic database and published studies. Data was extracted by a standard algorithm. Dominant, recessive, homozygote, and heterozygote comparison models were utilized. RESULTS: 24 studies incorporating 52,785 subjects were included in this meta-analysis. Overall, the minor allele (T) was associated with lower risk for hypertriglyceridemia in subjects with type 2 diabetes (dominant model: SMD = -0.04, 95% CI (-0.08, 0.00), P = 0.048, P heterogeneity = 0.47; recessive model: SMD = -0.10, 95% CI (-0.18, -0.02), P = 0.01, P heterogeneity = 0.56). No association was found between minor (T) allele and plasma TC, LDL-c, or HDL-c levels in subjects with type 2 diabetes or metabolic syndrome (MetS) and no association was found between minor (T) allele and plasma TG levels in nondiabetic subjects. CONCLUSIONS: Our meta-analysis indicated the association between TCF7L2 rs7903146 polymorphism and low plasma triglyceride (TG) level in subjects with type 2 diabetes. No association was found between rs7903146 variant and plasma lipids in nondiabetic subjects.

Impaired LRP6-TCF7L2 Activity Enhances Smooth Muscle Cell Plasticity and Causes Coronary Artery Disease.

Mutations in Wnt-signaling coreceptor LRP6 have been linked to coronary artery disease (CAD) by unknown mechanisms. Here, we show that reduced LRP6 activity in LRP6(R611C) mice promotes loss of vascular smooth muscle cell (VSMC) differentiation, leading to aortic medial hyperplasia. Carotid injury augmented these effects and led to partial to total vascular obstruction. LRP6(R611C) mice on high-fat diet displayed dramatic obstructive CAD and exhibited an accelerated atherosclerotic burden on LDLR knockout background. Mechanistically, impaired LRP6 activity leads to enhanced non-canonical Wnt signaling, culminating in diminished TCF7L2 and increased Sp1-dependent activation of PDGF signaling. Wnt3a administration to LRP6(R611C) mice improved LRP6 activity, led to TCF7L2-dependent VSMC differentiation, and rescued post-carotid-injury neointima formation. These findings demonstrate the critical role of intact Wnt signaling in the vessel wall, establish a causal link between impaired LRP6/TCF7L2 activities and arterial disease, and identify Wnt signaling as a therapeutic target against CAD.

Plasma cardiotrophin-1 levels are associated with hypertensive heart disease: a meta-analysis.

Cardiotrophin-1 (CT-1) is a member of the interleukin 6 cytokine superfamily. Plasma CT-1 levels have been associated with heart failure and hypertension in small independent studies. Whether plasma CT-1 levels are associated with progression of hypertensive heart disease is poorly understood. The authors carried out a meta-analysis using published studies and electronic databases. Relevant data were extracted using standardized algorithms. Additional data were obtained directly from investigators when indicated. A total of 18 studies were included that reported on association between CT-1 level and hypertension (n=8), cardiac hypertrophy (n=9), and heart failure (HF) (n=10). The serum levels of CT-1 were significantly higher in patients with hypertension (standard mean difference [SMD], 0.85; 95% confidence interval [CI], 0.64-1.06 fmol/mL), left ventricular hypertrophy (SMD, 0.88; 95% CI 0.60-1.17 fmol/mL), or HF (SMD, 0.66; 95% CI, 0.51-0.80 fmol/mL) compared with controls. Subgroup analysis revealed CT-1 levels to be highest in patients with hypertension-induced hypertrophy with HF, followed by patients with hypertension-induced left ventricular hypertrophy without HF (SMD, 0.52; 95% CI, 0.30-0.75 fmol/mL), patients with hypertension without left ventricular hypertrophy (SMD, 0.67; 95% CI, 0.46-0.88 fmol/mL) as compared with normotensive patients (SMD, 0.74; 95% CI, 10.45-1.04 fmol/mL). Increased plasma CT-1 levels are associated with risk for HF in hypertensive patients. CT-1 may serve as a novel biomarker in determining prognosis in hypertensive patients.

The combined hyperlipidemia caused by impaired Wnt-LRP6 signaling is reversed by Wnt3a rescue.

The underlying molecular genetic basis of combined hyperlipidemia, the most common atherogenic lipid disorder, is poorly characterized. Rare, nonconservative mutations in the Wnt coreceptor, LRP6, underlie autosomal dominant atherosclerosis, combined hyperlipidemia, and fatty liver disease. Mice with LRP6(R611C) mutation similarly developed elevated plasma LDL and TG levels and fatty liver. Further investigation showed that LRP6(R611C) mutation triggers hepatic de novo lipogenesis, lipid and cholesterol biosynthesis, and apoB secretion by an Sp1-dependent activation of IGF1, AKT, and both mTORC1 and mTORC2. These pathways were normalized after in vitro treatment of primary hepatocytes from LRP6(R611C) mice with either the IGF1R antagonist PPP, rapamycin, or rmWnt3a. Strikingly, in vivo administration of rmWnt3a to LRP6(R611C) mice normalized the altered expression of enzymes of DNL and cholesterol biosynthesis, and restored plasma TG and LDL levels to normal. These findings identify Wnt signaling as a regulator of plasma lipids and a target for treatment of hyperlipidemia.

Iron-ion radiation accelerates atherosclerosis in apolipoprotein E-deficient mice.

Radiation exposure from a number of terrestrial sources is associated with an increased risk for atherosclerosis. Recently, concern over whether exposure to cosmic radiation might pose a similar risk for astronauts has increased. To address this question, we examined the effect of 2 to 5 Gy iron ions ((56)Fe), a particularly damaging component of cosmic radiation, targeted to specific arterial sites in male apolipoprotein E-deficient (apoE(-/-)) mice. Radiation accelerated the development of atherosclerosis in irradiated portions of the aorta independent of any systemic effects on plasma lipid profiles or circulating leukocytes. Further, radiation exposure resulted in a more rapid progression of advanced aortic root lesions, characterized by larger necrotic cores associated with greater numbers of apoptotic macrophages and reduced lesional collagen compared to sham-treated mice. Intima media thickening of the carotid arteries was also exacerbated. Exposure to (56)Fe ions can therefore accelerate the development of atherosclerotic lesions and promote their progression to an advanced stage characterized by compositional changes indicative of increased thrombogenicity and instability. We conclude that the potential consequences of radiation exposure for astronauts on prolonged deep-space missions are a major concern. Knowledge gained from further studies with animal models should lead to a better understanding of the pathophysiological effects of accelerated ion radiation to better estimate atherogenic risk and develop appropriate countermeasures to mitigate its damaging effects.