Human collagen XV is a prominent histopathological component of sinusoidal capillarization in hepatocellular carcinogenesis.

BACKGROUND: Increased expression of collagen XV has been reported in hepatocellular carcinogenesis in mice. The aim of this study was to confirm the previous murine findings in human hepatocellular carcinoma (HCC) specimens, along with the histopathological distribution of collagen XV in tumoral tissues. METHODS: Sixty-three primary HCC specimens were examined. Immunostaining of collagen XV and quantitative reverse transcriptional PCR of COL15A1, which encodes collagen XV, were performed. RESULTS: Positive staining of collagen XV was observed in all tumoral regions, regardless of differentiation level or pathological type of HCC, along the sinusoid-like endothelium, whereas collagen XV was not expressed in any non-tumoral region. The intensity score of collagen XV immunostaining and the mRNA value of COL15A1 were significantly correlated. COL15A1 expression in tumors was 3.24-fold higher than in non-tumoral regions. Multivariate analysis showed that COL15A1 expression was significantly higher in the absence of hepatitis virus and moderately differentiated HCC. CONCLUSIONS: COL15A1 mRNA was up-regulated in HCC and collagen XV was expressed along the sinusoid-like endothelium of HCC but not in non-tumoral regions, which implies that collagen XV contributes to the capillarization of HCC.

Eosinophil Cationic Protein Shows Survival Effect on H9c2 Cardiac Myoblast Cells with Enhanced Phosphorylation of ERK and Akt/GSK-3ß under Oxidative Stress.

Eosinophil cationic protein (ECP) is well known as a cationic protein contained in the basic granules of activated eosinophils. Recent studies have reported that ECP exhibits novel activities on various types of cells, including rat neonatal cardiomyocytes. Here we evaluated the effects of ECP on rat cardiac myoblast H9c2 cells. Our results showed that ECP enhanced the survival of the cells, in part by promoting the ERK and Akt/GSK-3ß signaling pathways. ECP attenuated the cytotoxic effects of H2O2 on H9c2 cells as well as the production of reactive oxygen species, the number of apoptotic cells and caspase 3/7 activity in the cells. In conclusion, ECP activated the ERK and Akt/GSK-3ß pathways, resulting in anti-oxidative effects on H9c2 cells that attenuated apoptosis.

ADAMTS1 inhibits lymphangiogenesis by attenuating phosphorylation of the lymphatic endothelial cell-specific VEGF receptor.

Angiogenesis and lymphangiogenesis play roles in malignant tumor progression, dissemination, and metastasis. ADAMTS1, a member of the matrix metalloproteinase family, is known to inhibit angiogenesis. Recombinant ADAMTS1 was shown to strongly inhibit angiogenesis. We investigated whether ADAMTS1 inhibited lymphangiogenesis in the present study. We examined cell proliferation and cell migration in normal human dermal lymphatic microvascular endothelial cells (HMVEC-dLy) transduced with or without adenoviral human ADAMTS1 gene therapy. We then examined the VEGFC/VEGFR3 signal transduction pathway in ADAMTS1-transduced HMVEC-dLy. Cell proliferation and tube formation in Matrigel were significantly lower with transduced ADAMTS1 than with control (non-transduced HMVEC-dLy). The phosphorylation of VEGFR3 was also attenuated by ADAMTS1 gene therapy in HMVEC-dLy. Immunoprecipitation assays revealed that ADAMTS1 formed a complex with VEGFC. Our results demonstrated that ADAMTS1 inhibited lymphangiogenesis in vitro. The data highlight the new function of ADAMTS1 in the regulation of lymphangiogenesis and the therapeutic potential of ADAMTS1 in cancer therapy.

Light and electron microscopic detection of inflammation-targeting liposomes encapsulating high-density colloidal gold in arthritic mice.

OBJECTIVE: We have previously demonstrated the efficient and time-dependent transvascular localization of Sialyl Lewis X (SLX)-liposomes to inflammatory sites, but the final target of the SLX-liposomes remained uncertain. The aim of this study was to identify the target cells of the liposomes within the inflamed joints of collagen antibody-induced arthritis (CAIA) model mice. METHODS: SLX-liposomes and unlabeled liposomes encapsulating high-density colloidal gold were administered intravenously into the caudal vein of CAIA mice on day 5 after induction of arthritis when the inflammatory score was maximal (n = 6 per group). Six hours or 24 h after liposome administration, animals were euthanized and hind limbs and ankles were excised without perfusion. After fixation, synovial tissues were examined by light microscopy after silver enhancement of colloidal gold or by transmission electron microscopy. RESULTS: Silver-enhanced signals were detected within the cells around E-selectin-positive blood vessels in the synovium of the SLX-liposome group. These cells were positive for the macrophage/monocyte marker F4/80 or neutrophil marker Ly-6G. Transmission electron microscopy detected the colloidal gold signals together with liposome-like structures within the phagosomes of synovial macrophages. Transmission electron microscopy and energy dispersive X-ray spectrometry could determine gold elements in the lysosomes of synovial macrophages. CONCLUSIONS: The results of the current study demonstrate that SLX-liposomes primarily targeting E-selectin in activated endothelial cells could potentially deliver their contents into inflammatory cells around synovial blood vessels in arthritic joints.

ADAMTS1, ADAMTS5, ADAMTS9 and aggrecanase-generated proteoglycan fragments are induced following spinal cord injury in mouse.

ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) proteinases are involved in a variety of biological processes such as angiogenesis, cancer and arthritis. ADAMTSs appears to be responsible for the cleavage of proteoglycans in several tissues including brain and cartilage. Chondroitin sulfate proteoglycans (CSPGs) maintains the integrity of the brain extracellular matrix and major inhibitory contributors for glial scar and neural plasticity. The activity of aggrecanases in the central nervous system (CNS) has been reported. ADAMTSs are an enzyme degrading CSPGs in the brain. However, there is a little knowledge regarding ADAMTSs in the CNS. We investigated the expression levels of ADAMTSs mRNAs by RT-PCR after spinal cord injury in mouse. Transcripts encoding 4 of the 19 known ADAMTSs were evaluated in the mouse spinal cord following injury. ADAMTS1, -5 and -9 expression levels were found to be upregulated. No change was observed in ADAMTS4 expression. By means of immunohistochemistry, ADAMTSs were detected in the astrocytes implying its cellular source in SCI. Western blot analyses indicated that aggrecanase-generated proteoglycan fragments are produced after SCI.

Drosophila type XV/XVIII collagen mutants manifest integrin mediated mitochondrial dysfunction, which is improved by cyclosporin A and losartan.

Vertebrate collagen types XV and XVIII are broadly distributed basement membrane components, classified into a structurally distinct subgroup called "multiplexin collagens". Mutations in mammalian multiplexins are identified in some degenerative diseases such as Knobloch syndrome 1 (KNO1) or skeletal/cardiac myopathies, however, these progressive properties have not been elucidated. Here we investigated Drosophila mutants of Multiplexin (Mp), the only orthologue of vertebrate collagen types XV and XVIII, to understand the pathogenesis of multiplexin-related diseases. The mp mutants exhibited morphological changes in cardiomyocytes and progressive dysfunction of the skeletal muscles, reminiscent phenotypes observed in Col15a1-null mice. Ultrastructural analysis revealed morphologically altered mitochondria in mutants' indirect flight muscles (IFMs), resulting in severely attenuated ATP production and enhanced reactive oxygen species (ROS) production. In addition, mutants' IFMs exhibited diminished ßPS integrin clustering and abolished focal adhesion kinase (FAK) phosphorylation. Furthermore, mutants' defective IFMs are improved by the administrations of cyclosporin A, an inhibitor against mitochondrial permeability transition pore (mPTP) opening or losartan, an angiotensin II type 1 receptor (AT1R) blocker. Thus, our results suggest that Mp modulates mPTP opening and AT1R activity through its binding to integrin and that lack of Mp causes unregulated mPTP opening and AT1R activity, leading to mitochondrial dysfunctions. Hence, our results provide new insights towards the roles of multiplexin collagens in mitochondrial homeostasis and may serve as pharmacological evidences for the potential use of cyclosporin A or losartan for the therapeutic strategies.

A new model of development of the mammalian ovary and follicles.

Ovarian follicular granulosa cells surround and nurture oocytes, and produce sex steroid hormones. It is believed that during development the ovarian surface epithelial cells penetrate into the ovary and develop into granulosa cells when associating with oogonia to form follicles. Using bovine fetal ovaries (n = 80) we identified a novel cell type, termed GREL for Gonadal Ridge Epithelial-Like. Using 26 markers for GREL and other cells and extracellular matrix we conducted immunohistochemistry and electron microscopy and chronologically tracked all somatic cell types during development. Before 70 days of gestation the gonadal ridge/ovarian primordium is formed by proliferation of GREL cells at the surface epithelium of the mesonephros. Primordial germ cells (PGCs) migrate into the ovarian primordium. After 70 days, stroma from the underlying mesonephros begins to penetrate the primordium, partitioning the developing ovary into irregularly-shaped ovigerous cords composed of GREL cells and PGCs/oogonia. Importantly we identified that the cords are always separated from the stroma by a basal lamina. Around 130 days of gestation the stroma expands laterally below the outermost layers of GREL cells forming a sub-epithelial basal lamina and establishing an epithelial-stromal interface. It is at this stage that a mature surface epithelium develops from the GREL cells on the surface of the ovary primordium. Expansion of the stroma continues to partition the ovigerous cords into smaller groups of cells eventually forming follicles containing an oogonium/oocyte surrounded by GREL cells, which become granulosa cells, all enclosed by a basal lamina. Thus in contrast to the prevailing theory, the ovarian surface epithelial cells do not penetrate into the ovary to form the granulosa cells of follicles, instead ovarian surface epithelial cells and granulosa cells have a common precursor, the GREL cell.

Differential expression of basement membrane type IV collagen α2 and α6 chains as a prognostic factor in patients with extrahepatic bile duct carcinoma.

BACKGROUND: The destruction of the basement membrane (BM) is the first step in cancer invasion and metastasis. Type IV collagen is a major component of the BM, and is composed of six genetically distinct α(IV) chains; α1(IV) to α6(IV). The loss of α5(IV) and α6(IV) chains from the epithelial BM at the early stage of cancer invasion has been reported in several types of cancers. However, the expression of α5(IV) and α6(IV) chains in extrahepatic bile duct carcinoma (EBDC) remains unclear. METHODS: We examined the expression of α(IV) chains by immunohistochemistry using 71 resected EBDC specimens. Prognostic significance of α(IV) chains was examined by Cox regression and Kaplan-Meier analyses. RESULTS: In the invasive cancer, the expression of α6(IV) chain in the BM was lost partially or completely preceded by the loss of α2(IV) chain. The loss of α6(IV) chain in the BM of the invasive cancer was related to the tumor classification, TNM stages, and the expression of α2(IV) chain. The patients with α2(IV)-negative and α6(IV)-negative chains had significantly poorer prognosis than those with α2(IV)-positive and α6(IV)-positive/negative chains (P = 0.04). CONCLUSIONS: The loss of α2(IV) and α6(IV) chains might be a useful prognostic factor in patients with EBDC.

Tumor growth inhibitory effect of ADAMTS1 is accompanied by the inhibition of tumor angiogenesis.

Angiogenesis plays an important role in tumor progression. Several reports have demonstrated that a disintegrin and metalloproteinase with thrombospondin motifs1 (ADAMTS1) inhibited angiogenesis via multiple mechanisms. The aim of this study was to investigate the effect of ADAMTS1 on endothelial cells in vitro and on tumor growth with regard to angiogenesis in vivo. We examined the effects of the transfection of ADAMTS1 using two constructs, full-length ADAMTS1 (full ADAMTS1) and catalytic domain-deleted ADAMTS1 (delta ADAMTS1). Transfection of both the full ADAMTS1 and delta ADAMTS1 gene constructs demonstrated the secretion of tagged-ADAMTS1 protein into the conditioned medium, so we examined the effects of ADAMTS1-containing conditioned medium on endothelial cells. Both types of conditioned media inhibited endothelial tube formation, and this effect was completely abolished after immunoprecipitation of the secreted protein from the medium. Both types of conditioned media also inhibited endothelial cell migration and proliferation. We then examined the impact of ADAMTS1 on endothelial cell apoptosis. Both conditioned media increased the number of Annexin V-positive endothelial cells and caspase-3 activity and this effect was attenuated when z-vad was added. These results indicated that ADAMTS1 induced endothelial cell apoptosis. We next examined the effects of ADAMTS1 gene transfer into tumor-bearing mice. Both full ADAMTS1 and delta ADAMTS1 significantly inhibited the subcutaneous tumor growth. Collectively, our results demonstrated that ADAMTS1 gene transfer inhibited angiogenesis in vitro and in vivo, likely as a result of the induction of endothelial cell apoptosis by ADAMTS1 that occurs independent of the protease activity.

Bral2 is indispensable for the proper localization of brevican and the structural integrity of the perineuronal net in the brainstem and cerebellum.

Perineuronal nets (PNNs) are pericellular coats of condensed matrix that enwrap the cell bodies and dendrites of many adult central nervous system (CNS) neurons. These extracellular matrices (ECMs) play a structural role as well as instructive roles in the control of CNS plasticity and the termination of critical periods. The cartilage link protein Crtl1/Hapln1 was reported to be a trigger for the formation of PNNs in the visual cortex. Bral2/Hapln4 is another link protein that is expressed in PNNs, mainly in the brainstem and cerebellum. To assess the role of Bral2 in PNN formation, we examined the expression of PNN components in targeted mouse mutants lacking Bral2. We show here that Bral2-deficient mice have attenuated PNNs, but the overall levels of chondroitin sulfate proteoglycans, lecticans, are unchanged with the exception of neurocan. Bral2 deficiency markedly affected the localization of brevican in all of the nuclei tested, and neurocan concomitant with Crtl1 in some of the nuclei, whereas no effect was seen on aggrecan even with the attenuation of Crtl1. Bral2 may have a role in the organization of the PNN, in association with brevican, that is independent of aggrecan binding. There was a heterogenous attenuation of PNN components, including glycosaminoglycans, indicating the elaborate molecular organization of the PNN components. Strikingly, a slight decrease in the number of synapses in deep cerebellar nuclei neurons was found. Taken together, these results imply that Bral2-brevican interaction may play a key role in synaptic stabilization and the structural integrity of the PNN.

Mechanical stretch enhances COL2A1 expression on chromatin by inducing SOX9 nuclear translocalization in inner meniscus cells.

The meniscus plays an important role in controlling the biomechanics of the knee. However, the mechanical stress-related response in meniscus cells remains unclear. We investigated mechanical stretch-regulated gene expression in human meniscus cells. Human inner and outer meniscus cells were prepared from the inner and outer halves of the lateral meniscus. The gene expressions of Sry-type HMG box (SOX) 9 and α1(II) collagen (COL2A1) were assessed by real-time PCR analyses after cyclic tensile strain (CTS) treatment (0.5 Hz, 5% stretch). The localization and phosphorylation of SOX9 were evaluated by immunohistochemical and Western blot (WB) analyses. Chromatin immunoprecipitation (IP) analysis was performed to assess the stretch-related protein-DNA complex formation between SOX9 and the COL2A1 enhancer on chromatin. Type II collagen deposition and SOX9 production were detected only in inner menisci. CTS treatments increased expression of the COL2A1 and SOX9 genes in inner meniscus cells, but not in outer meniscus cells. In addition, CTS treatments stimulated nuclear translocalization and phosphorylation of SOX9 in inner meniscus cells. Chromatin IP analyses revealed that CTS increased the association between SOX9 and its DNA-binding site, included in the COL2A1 enhancer, on chromatin. Our results indicate that inner and outer meniscus cells have different properties in mechanical stretch-induced COL2A1 expression. In inner meniscus cells, mechanical stretch may have an essential role in the epigenetic regulation of COL2A1 expression.

Drosophila type XV/XVIII collagen, Mp, is involved in Wingless distribution.

Multiplexin (Mp) is the Drosophila orthologue of vertebrate collagens XV and XVIII. Like them, Mp is widely distributed in the basement membranes of the developing embryos, including those of neuroblasts in the central and peripheral nervous systems, visceral muscles of the gut, and contractile cardioblasts. Here we report the identification of mutant larvae bearing piggyBac transposon insertions that exhibit decrease Mp production associated with abdominal cuticular and wing margin defects, malformation of sensory organs and impaired sensitivity to physical stimuli. Additional findings include the abnormal ultrastructure of fatbody associated with abnormal collagen IV deposition, and reduced Wingless deposition. Collectively, these findings are consistent with the notion that Mp is required for the proper formation and/or maintenance of basement membrane, and that Mp may be involved in establishing the Wingless signaling gradients in the Drosophila embryo.

Lack of collagen XVIII long isoforms affects kidney podocytes, whereas the short form is needed in the proximal tubular basement membrane.

Collagen XVIII is characterized by three variant N termini, an interrupted collagenous domain, and a C-terminal antiangiogenic domain known as endostatin. We studied here the roles of this collagen type and its variant isoforms in the mouse kidney. Collagen XVIII appeared to be in a polarized orientation in the tubular basement membranes (BMs), the endostatin domain embedded in the BM, and the N terminus residing at the BM-fibrillar matrix interface. In the case of the glomerular BM (GBM), collagen XVIII was expressed in different isoforms depending on the side of the GBM. The orientation appeared polarized here, too, both the endothelial promoter 1-derived short variant of collagen XVIII and the epithelial promoter 2-derived longer variants having their C-terminal endostatin domains embedded in the BM and the N termini at the respective BM-cell interfaces. In addition to loosening of the proximal tubular BM structure, the Col18a1(-/-) mice showed effacement of the glomerular podocyte foot processes, and microindentation studies showed changes in the mechanical properties of the glomeruli, the Col18a1(-/-) glomeruli being ∼30% softer than the wild-type. Analysis of promoter-specific knockouts (Col18a1(P1/P1) and Col18a1(P2/P2)) indicated that tubular BM loosening is due to a lack of the shortest isoform, whereas the glomerular podocyte effacement was due to a lack of the longer isoforms. We suggest that lack of collagen XVIII may also have disparate effects on kidney function in man, but considering the mild physiological findings in the mutant mice, such effects may manifest themselves only late in life or require other compounding molecular changes.

Olmesartan reduces arterial stiffness and serum adipocyte fatty acid-binding protein in hypertensive patients.

Adipocyte fatty acid binding protein (A-FABP) has been reported to be involved in insulin resistance, lipid metabolism, and atherosclerosis; however, little is known about the effect of medication on the change in circulating A-FABP in human subjects. We evaluated the effects of angiotensin II type 1 receptor blocker (ARB) on arterial stiffness and its association with serum A-FABP in patients with hypertension. Thirty patients newly diagnosed with essential hypertension were treated with olmesartan (20 mg/day), an ARB, for 6 months. Serum levels of A-FABP and high-sensitivity C-reactive protein (hsCRP) were examined and the cardio-ankle vascular index (CAVI), which is a marker of arterial stiffness, was also determined. Serum A-FABP at baseline was significantly correlated with the body mass index (r = 0.45, P = 0.01), homeostasis model assessment as a marker of insulin resistance (r = 0.53, P < 0.01), and systolic blood pressure (r = 0.37, P = 0.047), and tended to be correlated with low-density lipoprotein cholesterol, triglyceride, and CAVI. Olmesartan treatment resulted in a significant decrease in CAVI, serum A-FABP levels, and hsCRP, besides a significant reduction of blood pressure. Multiple regression analysis revealed that the change in CAVI was independently correlated with the change in serum A-FABP. Olmesartan ameliorated arterial stiffness in patients with hypertension, which may be involved in the reduction of serum A-FABP.

AHR, a novel acute hypoxia-response sequence, drives reporter gene expression under hypoxia in vitro and in vivo.

ADAMTS1 (a disintegrin and metalloproteinase with thrombospondin motifs 1) is an early immediate gene. We have previously reported that ADAMTS1 was strongly induced by hypoxia. In this study, we investigated whether ADAMTS1 promoter-driven reporter signal is detectable by acute hypoxia. We constructed the GFP (green fluorescent protein) expression vector [AHR (acute hypoxia-response sequence)-GFP] under the control of ADAMTS1 promoter and compared it with the constitutive GFP-expressing vector under the control of CMV (cytomegalovirus promoter-GFP). We transduced AHR-GFP and examined whether GFP signals can be detected under the acute hypoxia. When the human umbilical vein [HUVEC (human umbilical vein endothelial cells)] was transduced under normoxia, there were few GFP signals, while CMV-GFP showed considerable GFP signals. When HUVEC was stimulated with hypoxia, GFP signals from AHR-GFP gene were induced under hypoxic conditions. Notably, the GFP signals peaked at 3 h under hypoxia. In ischaemic hind limb model, transduced AHR-GFP showed hypoxic induction of GFP signals. In summary, we have demonstrated that the AHR system induced the reporter gene expression by acute hypoxia, and its induction is transient. This is the first report showing the unique acute hypoxia-activated gene expression system.

Distribution of α(IV) collagen chains in the ocular anterior segments of adult mice.

The distribution of the collagen chains from α1(IV) to α6(IV) could serve as a basis for the characterization of type IV collagen. In this study, immunohistochemistry of the ocular anterior segment of adult mice was performed using specific monoclonal antibodies against each chain in the series from α1(IV) to α6(IV). The results show that the components of type IV collagen in vascular basement membranes are α1(IV) and α2(IV) with or without α5(IV) and α6(IV) chains and those in epithelium and muscle basement membranes are α1(IV), α2(IV), α5(IV), and α6(IV) chains. In corneal endothelium, pigmented epithelium of iris and ciliary body, and trabecular meshwork, α3(IV) and α4(IV) chains are also expressed in addition to α1(IV), α2(IV), α5(IV), and α6(IV) chains. Moreover, we investigated the change in molecular composition in ciliary body during postnatal development. α3(IV) and α4(IV) chains were also expressed in addition to α1(IV), α2(IV), α5(IV), and α6(IV) chains in ciliary pigmented epithelium basement membrane from 7 days after birth. This result suggests that the basement membranes gradually change their biochemical features owing to temporal regulation. Taken together, these findings suggest that the different distribution and the developmental expression of α1(IV) to α6(IV) chains are associated with the tissue-specific function of type IV collagen in basement membranes.

Clonal overgrowth of esophageal smooth muscle cells in diffuse leiomyomatosis-Alport syndrome caused by partial deletion in COL4A5 and COL4A6 genes.

This is a study of a patient who manifests all of the features of a diffuse leiomyomatosis-Alport syndrome (DL-ATS), and her two-year-old son who has already been diagnosed with Alport syndrome. Fourteen years ago, the patient underwent a partial esophageal resection followed by a replacement with jejunum. Recently, she underwent a surgical resection of the esophagus due to esophageal dysfunction. Genetic analyses of COL4A5 and COL4A6 on the X-chromosome were efficiently performed using the genomic DNA of her son. We have identified a novel deletion of 194-kb in length, encompassing COL4A5-COL4A6 promoters as well as nearly the entire large intron 1 of COL4A5 and intron 2 of COL4A6. To uncover the relationship of the esophagus-specific occurrence of the tumor and the expression of those genes, immunohistochemical analyses of type IV collagen α chains were conducted in the non-affected individuals. The esophageal smooth muscle-specific expression of α5(IV) and α6(IV) chains in the gastrointestinal tract was observed. Moreover, CAG repeat analysis of the androgen receptor gene and an immunohistochemical analysis in the leiomyoma revealed clonal overgrowth of the cells which received X-inactivation on the non-affected allele. These results may suggest that the dominant effect was caused by the partial deletion of the esophageal smooth muscle-specific genes, COL4A5 and COL4A6.

Connective tissue growth factor induction in a pressure-overloaded heart ameliorated by the angiotensin II type 1 receptor blocker olmesartan.

Connective tissue growth factor (CTGF) is a secreted protein that regulates fibrosis. We hypothesized that CTGF is induced in a pressure-overloaded (PO) heart and that blocking the angiotensin II type 1 receptor would reduce CTGF expression. Accordingly, we administered olmesartan and compared its effects with other antihypertensive drugs in a PO heart. CTGF induction was determined in a rat PO model, and olmesartan, hydralazine or saline was continuously administered. The effects of olmesartan on CTGF induction, myocyte hypertrophy and fibrosis were evaluated. The effect of olmesartan on cardiac function was also examined in CTGF- and transforming growth factor-beta 1 (TGF-ß1)-infused rats. CTGF was increased in the PO heart 3 days after aortic banding and was markedly distributed around the perivascular fibrotic area. After 28 days, blood pressure was not significantly different in the olmesartan and hydralazine groups, but olmesartan treatment reduced CTGF distribution in PO hearts. Olmesartan was associated with a significantly reduced myocyte hypertrophy index (4.77±0.48 for olmesartan and 6.05±1.45 for saline, P<0.01), fibrosis area (32.0±15.5% compared with the saline group, P<0.05) and serum TGF-ß1 level (62.6±10.6 ng ml⁻¹ for olmesartan and 84.4±7.2 ng ml⁻¹ for hydralazine, P<0.05). In addition, cardiac function was significantly preserved in the olmesartan group compared with the saline group. Finally, olmesartan ameliorated the cardiac dysfunction in CTGF- and TGF-ß1-infused rats. Olmesartan attenuated CTGF induction, reduced perivascular fibrosis and ameliorated cardiac dysfunction in a PO heart. Our results provide insight into the beneficial effects of olmesartan on PO hearts, independent of blood-pressure lowering.

Increased activity and expression of histone deacetylase 1 in relation to tumor necrosis factor-alpha in synovial tissue of rheumatoid arthritis.

INTRODUCTION: The purpose of this study was to investigate the profile of histone deacetylase (HDAC) expression in the synovial tissue of rheumatoid arthritis (RA) compared with that of normal control and osteoarthritis (OA), and to examine whether there is a link between HDAC activity and synovial inflammation. METHODS: HDAC activity and histone acetyltransferase (HAT) activity were determined in nuclear extracts of total synovial tissue surgically obtained from normal, OA and RA joints. The level of cytoplasmic tumor necrosis factor a (TNFα) fraction was measured by ELISA. Total RNA of synovial tissue was used for RT-PCR of HDAC1-8. In synovial fibroblasts from RA (RASFs), the effects of TNFα on nuclear HDAC activity and class I HDACs (1, 2, 3, 8) mRNA expressions were examined by quantitative real-time PCR. The protein expression and distribution of class I HDACs were examined by Western blotting. RESULTS: Nuclear HDAC activity was significantly higher in RA than in OA and normal controls and correlated with the amount of cytoplasmic TNFα. The mRNA expression of HDAC1 in RA synovial tissue was higher than in OA and normal controls, and showed positive correlation with TNFα mRNA expression. The protein level of nuclear HDAC1 was higher in RA synovial tissue compared with OA synovial tissue. Stimulation with TNFα significantly increased the nuclear HDAC activity and HDAC1 mRNA expression at 24 hours and HDAC1 protein expression at 48 hours in RASFs. CONCLUSIONS: Our results showed nuclear HDAC activity and expression of HDAC1 were significantly higher in RA than in OA synovial tissues, and they were upregulated by TNFα stimulation in RASFs. These data might provide important clues for the development of specific small molecule HDAC inhibitors.

Serum adipocyte fatty acid-binding protein is independently associated with coronary atherosclerotic burden measured by intravascular ultrasound.

OBJECTIVES: Adipocyte fatty acid-binding protein (A-FABP) has been shown to have an effect on insulin resistance, lipid metabolism, and atherosclerosis in animals. We therefore investigated the association between the serum A-FABP level and coronary atherosclerosis. METHODS: One hundred twenty-five consecutive patients with coronary artery disease (CAD) were enrolled after coronary angiography. Plaque volume in non-culprit coronary arteries was determined using intravascular ultrasound and expressed as percent plaque volume (%PV). Voluntary blood donors (n=120), matched for age and gender, served as controls. Serum levels of A-FABP, adiponectin, and inflammatory markers were measured by enzyme-linked immunosorbent assay. RESULTS: The serum A-FABP level in CAD patients was significantly higher than in control subjects (median [25th-75th percentiles], 27.2 [20.5-37.1] ng/mL vs. 18.9 [14.6-24.5] ng/mL) (p<0.01). Serum A-FABP showed 0.74 of the area under the curve in the receiver operating characteristic curve for the detection of CAD, with 76% specificity and 65% sensitivity with a cut-off value of 20.1 ng/mL. Further, in CAD patients, serum A-FABP had a significant correlation with %PV in all subjects (r=0.33, p<0.01). Serum A-FABP was positively correlated with the body mass index, serum interleukin-6 and high-sensitive CRP, and negatively correlated with HDL-cholesterol and serum adiponectin in CAD patients. Stepwise regression analysis revealed that serum A-FABP was independently associated with %PV. CONCLUSION: Increased serum A-FABP was significantly associated with a greater coronary plaque burden. Our findings revealed that the measurement of serum A-FABP could be utilized for the evaluation of the extent of coronary atherosclerosis.