BBA - Molecular Basis of Disease (v.1842, #5)
Editorial Board (i).
Development and pathomechanisms of cardiomyopathy in very long-chain acyl-CoA dehydrogenase deficient (VLCAD−/−) mice by Sara Tucci; Ulrich Flögel; Sven Hermann; Marga Sturm; Michael Schäfers; Ute Spiekerkoetter (677-685).
Hypertrophic cardiomyopathy is a typical manifestation of very long-chain acyl-CoA dehydrogenase deficiency (VLCADD), the most common long-chain β-oxidation defects in humans; however in some patients cardiac function is fully compensated. Cardiomyopathy may also be reversed by supplementation of medium-chain triglycerides (MCT). We here characterize cardiac function of VLCAD-deficient (VLCAD−/−) mice over one year. Furthermore, we investigate the long-term effect of a continuous MCT diet on the cardiac phenotype. We assessed cardiac morphology and function in VLCAD−/− mice by in vivo MRI. Cardiac energetics were measured by 31P-MRS and myocardial glucose uptake was quantified by positron-emission-tomography (PET). Metabolic adaptations were identified by the expression of genes regulating glucose and lipid metabolism using real-time-PCR. VLCAD−/− mice showed a progressive decrease in heart function over 12 months accompanied by a reduced phosphocreatine-to-ATP-ratio indicative of chronic energy deficiency. Long-term MCT supplementation aggravated the cardiac phenotype into dilated cardiomyopathy with features similar to diabetic heart disease. Cardiac energy production and function in mice with a β-oxidation defect cannot be maintained with age. Compensatory mechanisms are insufficient to preserve the cardiac energy state over time. However, energy deficiency by impaired β-oxidation and long-term MCT induce cardiomyopathy by different mechanisms. Cardiac MRI and MRS may be excellent tools to assess minor changes in cardiac function and energetics in patients with β-oxidation defects for preventive therapy.
Keywords: VLCAD-deficiency; Dilated cardiomyopathy; MRI; PET; MCT-supplementation; Energy metabolism;
Piwi-like 1 and 4 gene transcript levels are associated with clinicopathological parameters in renal cell carcinomas by Omar Al-Janabi; Sven Wach; Elke Nolte; Katrin Weigelt; Tilman T. Rau; Christine Stöhr; Wolfgang Legal; Stefan Schick; Thomas Greither; Arndt Hartmann; Bernd Wullich; Helge Taubert (686-690).
Piwi-like gene family members (Piwil 1–4) are considered stem cell-associated genes/proteins. These are expressed predominantly in germline cells, but are re-expressed in different tumors. Piwil 1–4 gene expression has not previously been studied and correlated with clinicopathological parameters in renal cell carcinomas (RCC). The Piwil 1–4 transcript levels were analyzed by quantitative real-time PCR in 73 clear cell RCC (ccRCC) tissues and corresponding normal tissues. The transcript levels of Piwil 1, 2 and 4 were strongly and significantly correlated with each other, in both the tumor tissues and the normal tissues (P < 0.001; Spearman's rank test). Piwil 4 gene expression was significantly higher in the ccRCC tissues than that in the corresponding normal renal tissues (P < 0.001; Wilcoxon signed-rank test). When the ccRCC patient cohort was divided according to the median Piwil 1–4 expression into low- and high-expression groups and according to age into younger (≤ 64 years) and older patient groups (> 64 years), the younger patients displayed significantly higher levels of Piwil 1 mRNA in comparison to the older patients (P = 0.010; Fisher's exact test). Interestingly, Piwil 1 expression was left–right polarized in the normal tissues but not in the tumor tissues (P = 0.004; Fisher's exact test). Altogether, associations were determined between the Piwi-like family member expression levels and clinicopathological parameters of ccRCC, suggesting a potential role for these genes/proteins in ccRCC diagnostics and tumorigenesis as well as in renal tissue embryology.
Keywords: Piwi-like genes; Clear cell renal cell carcinoma; Quantitative RT-PCR; Age; Normal renal tissue; Tumorigenesis;
MDMA induces cardiac contractile dysfunction through autophagy upregulation and lysosome destabilization in rats by Kaori Shintani-Ishida; Kanju Saka; Koji Yamaguchi; Makiko Hayashida; Hisashi Nagai; Genzou Takemura; Ken-ichi Yoshida (691-700).
The underlying mechanisms of cardiotoxicity of 3,4-methylenedioxymethylamphetamine (MDMA, “ecstasy”) abuse are unclear. Autophagy exerts either adaptive or maladaptive effects on cardiac function in various pathological settings, but nothing is known on the role of autophagy in the MDMA cardiotoxicity. Here, we investigated the mechanism through which autophagy may be involved in MDMA-induced cardiac contractile dysfunction. Rats were injected intraperitoneally with MDMA (20 mg/kg) or saline. Left ventricular (LV) echocardiography and LV pressure measurement demonstrated reduction of LV systolic contractility 24 h after MDMA administration. Western blot analysis showed a time-dependent increase in the levels of microtubule-associated protein light chain 3-II (LC3-II) and cathepsin-D after MDMA administration. Electron microscopy showed the presence of autophagic vacuoles in cardiomyocytes. MDMA upregulated phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) at Thr172, mammalian target of rapamycin (mTOR) at Thr2446, Raptor at Ser792, and Unc51-like kinase (ULK1) at Ser555, suggesting activation of autophagy through the AMPK-mTOR pathway. The effects of autophagic inhibitors 3-methyladenine (3-MA) and chloroquine (CQ) on LC3-II levels indicated that MDMA enhanced autophagosome formation, but attenuated autophagosome clearance. MDMA also induced release of cathepsins into cytosol, and western blotting and electron microscopy showed cardiac troponin I (cTnI) degradation and myofibril damage, respectively. 3-MA, CQ, and a lysosomal inhibitor, E64c, inhibited cTnI proteolysis and improved contractile dysfunction after MDMA administration. In conclusion, MDMA causes lysosome destabilization following activation of the autophagy-lysosomal pathway, through which released lysosomal proteases damage myofibrils and induce LV systolic dysfunction in rat heart.
Keywords: MDMA; Cardiac contractile dysfunction; Autophagy; Lysosome; Cathepsin;
Loss of duplexmiR-223 (5p and 3p) aggravates myocardial depression and mortality in polymicrobial sepsis by Xiaohong Wang; Wei Huang; Yang Yang; Yigang Wang; Tianqing Peng; Jiang Chang; Charles C. Caldwell; Basilia Zingarelli; Guo-Chang Fan (701-711).
Sepsis is the leading cause of death in critically ill patients. While myocardial dysfunction has been recognized as a major manifestation in severe sepsis, the underlying molecular mechanisms associated with septic cardiomyopathy remain unclear. In this study, we performed a miRNA array analysis in hearts collected from a severe septic mouse model induced by cecal ligation and puncture (CLP). Among the 19 miRNAs that were dys-regulated in CLP-mouse hearts, miR-223(3p) and miR-223*(5p) were most significantly downregulated, compared with sham-operated mouse hearts. To test whether a drop of miR-223 duplex plays any roles in sepsis-induced cardiac dysfunction and inflammation, a knockout (KO) mouse model with a deletion of the miR-223 gene locus and wild-type (WT) mice were subjected to CLP or sham surgery. We observed that sepsis-induced cardiac dysfunction, inflammatory response and mortality were remarkably aggravated in CLP-treated KO mice, compared with control WTs. Using Western-blotting and luciferase reporter assays, we identified Sema3A, an activator of cytokine storm and a neural chemorepellent for sympathetic axons, as an authentic target of miR-223* in the myocardium. In addition, we validated that miR-223 negatively regulated the expression of STAT-3 and IL-6 in mouse hearts. Furthermore, injection of Sema3A protein into WT mice revealed an exacerbation of sepsis-triggered inflammatory response and myocardial depression, compared with control IgG1 protein-treated WT mice following CLP surgery. Taken together, these data indicate that loss of miR-223/-223* causes an aggravation of sepsis-induced inflammation, myocardial dysfunction and mortality. Our study uncovers a previously unrecognized mechanism underlying septic cardiomyopathy and thereby, may provide a new strategy to treat sepsis.
Keywords: Sepsis; MicroRNA; Myocardial depression; Sema3A; Inflammation;
Role of microRNA-27a in down-regulation of angiogenic factor AGGF1 under hypoxia associated with high-grade bladder urothelial carcinoma by Yan Xu; Ming Zhou; Jingjing Wang; Yuanyuan Zhao; Sisi Li; Bisheng Zhou; Zhenhong Su; Chengqi Xu; Yue Xia; Huijun Qian; Xin Tu; Wei Xiao; Xiaoping Chen; Qiuyun Chen; Qing K. Wang (712-725).
Hypoxia stimulates angiogenesis under a variety of pathological conditions, including malignant tumors by inducing expression of angiogenic factors such as VEGFA. Surprisingly, here we report significant association between down-regulation of a new angiogenic factor AGGF1 and high-grade urothelial carcinoma. The proportion of strong AGGF1 expression cases was significantly lower in the high-grade urothelial carcinoma group than that in the low-grade urothelial carcinoma group (P = 1.40 × 10− 5) or than that in the normal urothelium tissue group (P = 2.11 × 10− 4). We hypothesized that tumor hypoxia was responsible for differential expression of the AGGF1 protein in low- and high-grade urothelial carcinomas, and therefore investigated the molecular regulatory mechanism for AGGF1 expression under hypoxia. Under hypoxic conditions, AGGF1 protein levels declined without any change in mRNA levels and protein stability. Hypoxia-induced down-regulation of AGGF1 was mediated by miR-27a. Overexpression of miR-27a suppressed AGGF1 expression through translational inhibition, but not by RNA degradation. Moreover, the hypoxia-induced decrease of AGGF1 expression disappeared after miR-27a expression was inhibited. Furthermore, down-regulation of AGGF1 reduced hypoxia-induced apoptosis in cancer cells. Taken together, the results of this study indicate that (1) hypoxia down-regulates expression of the AGGF1 protein, but not AGGF1 mRNA, by inducing expression of miR-27a; (2) Down-regulation of AGGF1 had an apparent protective role for cancer cells under hypoxia; (3) Down-regulation of the AGGF1 protein confers a significant risk of high-grade human urothelial bladder carcinoma.
Keywords: Bladder urothelial carcinoma; AGGF1; Hypoxia; MiR-27a;
Cardiolipin content is involved in liver mitochondrial energy wasting associated with cancer-induced cachexia without the involvement of adenine nucleotide translocase by Cloé Mimsy Julienne; Marine Tardieu; Stéphan Chevalier; Michelle Pinault; Philippe Bougnoux; François Labarthe; Charles Couet; Stéphane Servais; Jean-François Dumas (726-733).
Cancer-induced cachexia describes the progressive skeletal muscle wasting associated with many cancers leading to shortened survival time in cancer patients. We previously reported that cardiolipin content and energy-wasting processes were both increased in liver mitochondria in a rat model of peritoneal carcinosis (PC)-induced cachexia. To increase the understanding of the cellular biology of cancer cachexia, we investigated the involvement of adenine nucleotide translocator (ANT) in mitochondrial energy-wasting processes in liver mitochondria of PC and pair-fed control rats and its interactions with cardiolipin in isolated liver mitochondria from healthy rats exposed to cardiolipin-enriched liposomes. We showed in this study that functional ANT content was decreased in liver mitochondria from PC rats but without any effects on the efficiency of ATP synthesis. Moreover, non-phosphorylating energy wasting was not affected by saturating concentrations of carboxyatractylate (CAT), a potent inhibitor of ANT, in liver mitochondria from PC rats. Decreased efficiency of ATP synthesis was found in normal liver mitochondria exposed to cardiolipin-enriched liposomes, with increased non-phosphorylating energy wasting, thus mimicking mitochondria from PC rats. However, the functional ANT content in these cardiolipin-enriched mitochondria was unchanged, although non-phosphorylating energy wasting was reduced by CAT-induced inhibition of ANT. Finally, non-phosphorylating energy wasting was increased in cardiolipin-enriched mitochondria with substrates for complexes 1 and 2, but not for complex 4. In conclusion, increased energy wasting measured in liver mitochondria from rats with cancer cachexia is dependent on cardiolipin but independent of ANT. Interactions between ANT and cardiolipin are modified when cancer cachexia occurs.
Keywords: ANT; ATP synthesis; Cardiolipin; Oxidative phosphorylation; Peritoneal carcinosis; Undernutrition;
Psoriasis pathogenesis — Pso p27 is generated from SCCA1 with chymase by Hilde Lysvand; Lars Hagen; Lidija Klubicka; Geir Slupphaug; Ole-Jan Iversen (734-738).
Psoriasis is a chronic inflammatory skin disease with unknown aetiology. Infiltration of inflammatory cells as the initial event in the development of new psoriatic plaques together with the defined inflamed areas of such lesions argues for an immunological disease with a local production of a causal antigen. The auto-antigen Pso p27 is a protein expressed in the skin lesions. We recently demonstrated that Pso p27 is homologous to the core amino acid sequences of squamous cell carcinoma antigens 1 and 2 (SCCA1/2) and it is apparently generated from SCCA molecules by digestion with highly specific endoproteases. In this communication we demonstrate the generation of Pso p27 from SCCA1 with extracts from psoriatic scale and even more remarkably, the generation of Pso p27 from SCCA1 in the presence of mast cell associated chymase. These findings open up for new therapeutic strategies in psoriasis and probably also in other autoimmune diseases as Pso p27 epitopes have been detected in diseased tissues from patients with various chronic inflammatory diseases.
Keywords: Autoimmunity; Chymase; Pathogenesis; Pso p27; Psoriasis; SCCA1;
FXR-dependent reduction of hepatic steatosis in a bile salt deficient mouse model by Cindy Kunne; Alexandra Acco; Suzanne Duijst; Dirk R. de Waart; Coen C. Paulusma; Ingrid Gaemers; Ronald P.J. Oude Elferink (739-746).
It has been established that bile salts play a role in the regulation of hepatic lipid metabolism. Accordingly, overt signs of steatosis have been observed in mice with reduced bile salt synthesis. The aim of this study was to identify the mechanism of hepatic steatosis in mice with bile salt deficiency due to a liver specific disruption of cytochrome P450 reductase.In this study mice lacking hepatic cytochrome P450 reductase (Hrn) or wild type (WT) mice were fed a diet supplemented with or without either 0.1% cholic acid (CA) or 0.025% obeticholic acid, a specific FXR-agonist.Feeding a CA-supplemented diet resulted in a significant decrease of plasma ALT in Hrn mice. Histologically, hepatic steatosis ameliorated after CA feeding and this was confirmed by reduced hepatic triglyceride content (115.5 ± 7.3 mg/g liver and 47.9 ± 4.6 mg/g liver in control- and CA-fed Hrn mice, respectively). The target genes of FXR-signaling were restored to normal levels in Hrn mice when fed cholic acid. VLDL secretion in both control and CA-fed Hrn mice was reduced by 25% compared to that in WT mice. In order to gain insight in the mechanism behind these bile salt effects, the FXR agonist also was administered for 3 weeks. This resulted in a similar decrease in liver triglycerides, indicating that the effect seen in bile salt fed Hrn animals is FXR dependent.In conclusion, steatosis in Hrn mice is ameliorated when mice are fed bile salts. This effect is FXR dependent. Triglyceride accumulation in Hrn liver may partly involve impaired VLDL secretion.
Keywords: Bile acid; Fatty liver; Hrn; CYP450-reductase; FXR;
Thromboxane synthase expression and correlation with VEGF and angiogenesis in non-small cell lung cancer by Mary Clare Cathcart; Kathy Gately; Robert Cummins; Clive Drakeford; Elaine W. Kay; Kenneth J. O'Byrne; Graham P. Pidgeon (747-755).
Background: Thromboxane synthase (TXS) metabolizes prostaglandin H2 into thromboxanes, which are biologically active on cancer cells. TXS over-expression has been reported in a range of cancers, and associated with angiogenesis and poor outcome. TXS has been identified as a potential therapeutic target in NSCLC. This study examines a link between TXS expression, angiogenesis, and survival in NSCLC. Methods: TXS and VEGF metabolite levels were measured in NSCLC serum samples (n = 46) by EIA. TXB2 levels were correlated with VEGF. A 204-patient TMA was stained for TXS, VEGF, and CD-31 expression. Expression was correlated with a range of clinical parameters, including overall survival. TXS expression was correlated with VEGF and CD-31. Stable TXS clones were generated and the effect of overexpression on tumor growth and angiogenesis markers was examined in-vitro and in-vivo (xenograft mouse model). Results: Serum TXB2 levels were correlated with VEGF (p < 0.05). TXS and VEGF were expressed to a varying degree in NSCLC tissue. TXS was associated with VEGF (p < 0.0001) and microvessel density (CD-31; p < 0.05). TXS and VEGF expression levels were higher in adenocarcinoma (p < 0.0001) and female patients (p < 0.05). Stable overexpression of TXS increased VEGF secretion in-vitro. While no significant association with patient survival was observed for either TXS or VEGF in our patient cohort, TXS overexpression significantly (p < 0.05) increased tumor growth in-vivo. TXS overexpression was also associated with higher levels of VEGF, microvessel density, and reduced apoptosis in xenograft tumors. Conclusion: TXS promotes tumor growth in-vivo in NSCLC, an effect which is at least partly mediated through increased tumor angiogenesis.
Keywords: Non-small cell lung cancer; Thromboxane synthase; Angiogenesis; VEGF; CD-31;