GABA receptors ameliorate Hcy-mediated integrin shedding and constrictive collagen remodeling in microvascular endothelial cells

Mammalian endothelial cells are deficient in cystathionine β synthetase (CBS) activity, which is responsible for homocysteine (Hcy) clearance. This deficiency makes the endothelium theprime target for Hcy toxicity. Hcy induces integrin shedding in microvascular endothelial cells (MVEC) by increasing matrix metalloproteinase (MMP). Hcy competes with inhibitory neurotransmitter γ aminobutyric acid (GABA)-A receptor. We hypothesized that Hcy transduces MVEC remodeling by increasing metalloproteinase activity and shedding β-1 integrin by inactivating the GABA-A/B receptors, thus behaving as an excitatory neurotransmitter. MVEC were isolated from mouse brain. The presence of GABA-A receptor was determined by immunolabeling. It was induced by muscimol, an agonist of GABA-A receptors as measured by Western blot analysis. Hcy induced MMP-2 activity in a dose- and time-dependent maner, measured by zymography. GABA-A/B receptors ameliorated the Hcy-mediated MMP-2 activation. Hcy selectively increased the levels of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-3 but decreased the levels of TIMP-4. Treatment with muscimol decreased the levels of TIMP-1 and TIMP-3 and increased the levels of TIMP-4 to control. Hcy caused a robust increae in the levels of a disintegrin and metalloproteinase (ADAM)-12. In the medium of MVEC reated with Hcy, the levels of β-1 integrin were significantly increased. Treatment with muscimol or baclofen (GABA-B receptor agonist) ameliorated the levels significantly increased. Treatment with muscimol or baclofen (GABA-B receptor agonist) ameliorated the levels of β-1 integrin in the medium. These results suggested that Hcy induced DAM-12. Significantly, Hcy facilitated the β-1 integrin shedding. Treatment of MVEC with muscimole or baclofen during Hcy administration ameliorated the expression of metalloproteinase, integrin-shedding, and constrictive collagen remodeling, suggesting a role of Hcy in GABA receptor-mediated cerebrovascular remodeling.     

Antibacterial Co(II), Ni(II), Cu(II) and Zn(II) complexes of Schiff bases derived from fluorobenzaldehyde and triazoles

Antibacterial Schiff bases derived from 1,2,4-triazoles as well as their metal complexes incorporating cobalt(II), nickel(II), copper(II) and zinc(II) have been synthesized and characterized. Physico-chemical studies suggest that an octahedral geometry for the cobalt(II), nickel(II) and zinc(II)and square-planer geometry for the copper(II) complexes. These complexes have been screened for antibacterial activity against three Gram-positive (Staphylococcus aureus, Staphylococcus epidermidis and Bacillus subtilis) and two Gram-negative (Salmonella typhi and Pseudomonas aeruginosa) bacterial strains, and results compared with the activity of the free ligands. The metal complexes were found to be more potent against one or more bacterial strains than the free ligands.     

Analysis, expression and prevalence of the Mycobacterium tuberculosis homolog of bacterial virulence regulating proteins

We have previously reported the identification of a gene from Mycobacterium tuberculosis, H37Rv, which on the basis of its nucleotide sequence encoded a protein product of 38 kDa. This 38-Kda mycobacterial protein designated as VirS exhibits homology with the VirF protein of Shigella, the VirFy protein of Yersinia and the Cfad, Rns and FapR proteins from various enterotoxigenic Escherichia coli strains. In this communication, we show the close sequence and structural similarities of the VirS protein with VirF, VirFy, Cfad, Rns and FapR and describe the results of our studies on the characterization of the virS gene promoter and its expression in E. coli and mycobacteria. virS was present exclusively in the species belonging to the M. tuberculosis complex as revealed by Southern blot and PCR analysis. Our findings suggest the involvement of virS in the regulation of pathogenesis of M. tuberculosis.     

Molecular mechanism of the anti-inflammatory activity of a natural diarylnonanoid, malabaricone C

The spice-derived phenolic, malabaricone C (mal C), has recently been shown to accelerate healing of the indomethacin-induced gastric ulceration in mice. In this study, we explored its anti-inflammatory activity and investigated the underlying mechanism of the action. Mal C suppressed the microvascular permeability and the levels of tumor necrosis factor-α, interleukin-1β, and nitric oxide in the lipopolysaccharide (LPS)-administered mice. At a dose of 10 mg/kg, it showed anti-inflammatory activity comparable to that of omeprazole (5 mg/kg) and dexamethasone (50 mg/kg). It also reduced the expression and activities of inducible nitric oxide synthase, cyclooxygenase-2, as well as the pro- vs anti-inflammatory cytokine ratio in the LPS-treated RAW macrophages. Mal C was found to inhibit LPS-induced NF-kB activation in RAW 264.7 cells by blocking the MyD88-dependent pathway. Mal C suppressed NF-κB activation and iNOS promoter activity, which correlated with its inhibitory effect on IκB phosphorylation and degradation, and NF-κB nuclear translocation, in the LPS-stimulated macrophages. It also inhibited LPS-induced phosphorylation of p38 and JNK, which are also upstream activators of NF-κB, without affecting Akt phosphorylation. Mal C also effectively blocked the PKR-mediated activation of NF-κB. These findings indicate that mal C exerts an anti-inflammatory effect through NF-κB-responsive inflammatory gene expressions by inhibiting the p38 and JNK-dependent canonical NF-κB pathway as well as the PKR pathway, and is a potential therapeutic agent against acute inflammation.     

Increased endogenous H2S generation by CBS, CSE, and 3MST gene therapy improves ex vivo renovascular relaxation in hyperhomocysteinemia

Hydrogen sulfide (H(2)S) has recently been identified as a regulator of various physiological events, including vasodilation, angiogenesis, antiapoptotic, and cellular signaling. Endogenously, H(2)S is produced as a metabolite of homocysteine (Hcy) by cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (3MST). Although Hcy is recognized as vascular risk factor at an elevated level [hyperhomocysteinemia (HHcy)] and contributes to vascular injury leading to renovascular dysfunction, the exact mechanism is unclear. The goal of the current study was to investigate whether conversion of Hcy to H(2)S improves renovascular function. Ex vivo renal artery culture with CBS, CSE, and 3MST triple gene therapy generated more H(2)S in the presence of Hcy, and these arteries were more responsive to endothelial-dependent vasodilation compared with nontransfected arteries treated with high Hcy. Cross section of triple gene-delivered renal arteries immunostaining suggested increased expression of CD31 and VEGF and diminished expression of the antiangiogenic factor endostatin. In vitro endothelial cell culture demonstrated increased mitophagy during high levels of Hcy and was mitigated by triple gene delivery. Also, dephosphorylated Akt and phosphorylated FoxO3 in HHcy were reversed by H(2)S or triple gene delivery. Upregulated matrix metalloproteinases-13 and downregulated tissue inhibitor of metalloproteinase-1 in HHcy were normalized by overexpression of triple genes. Together, these results suggest that H(2)S plays a key role in renovasculopathy during HHcy and is mediated through Akt/FoxO3 pathways. We conclude that conversion of Hcy to H(2)S by CBS, CSE, or 3MST triple gene therapy improves renovascular function in HHcy.     

Overexpression of hyaluronan-binding protein 1 (HABP1/p32/gC1qR) in HepG2 cells leads to increased hyaluronan synthesis and cell proliferation by up-regulation of cyclin D1 in AKT-dependent pathway

Overexpression of the mature form of hyaluronan-binding protein 1 (HABP1/gC1qR/p32), a ubiquitous multifunctional protein involved in cellular signaling, in normal murine fibroblast cells leads to enhanced generation of reactive oxygen species (ROS), mitochondrial dysfunction, and ultimately apoptosis with the release of cytochrome c. In the present study, human liver cancer cell line HepG2, having high intracellular antioxidant levels was chosen for stable overexpression of HABP1. The stable transformant of HepG2, overexpressing HABP1 does not lead to ROS generation, cellular stress, and apoptosis, rather it induced enhanced cell growth and proliferation over longer periods. Phenotypic changes in the stable transformant were associated with the increased "HA pool," formation of the "HA cable" structure, up-regulation of HA synthase-2, and CD44, a receptor for HA. Enhanced cell survival was further supported by activation of MAP kinase and AKT-mediated cell survival pathways, which leads to an increase in CYCLIN D1 promoter activity. Compared with its parent counterpart HepG2, the stable transformant showed enhanced tumorigenicity as evident by its sustained growth in low serum conditions, formation of the HA cable structure, increased anchorage-independent growth, and cell-cell adhesion. This study suggests that overexpression of HABP1 in HepG2 cells leads to enhanced cell survival and tumorigenicity by activating HA-mediated cell survival pathways.     

Molecular cloning and expression profile of snow trout GPDH gene in response to abiotic stress

Glycerol-3-phosphate dehydrogenase (GPDH) gene possibly plays a key role for cold acclimation process in snow trout during winter months when water temperature goes down to 4–5?°C. In this study, 1,012?bp nucleotide fragment of GPDH gene was obtained from two snow trout species (Schizothorax richardsonii and S. niger; family: Cyprinidae), distributed in several Himalayan rivers. The gene encoded a protein of 334 amino acids. The encoded protein sequence was very similar to GPDH of Danio rerio (94.36?%) using BLASTx searches. In S. richardsonii the qRT-PCR showed highest expression in muscle tissue followed by liver and also revealed 19 fold gene expression in liver tissue under cold (5?°C) in comparison with warm (15?°C) condition. The elevated expression levels of GPDH cDNA on cold treatment furthermore suggest that GPDH plays a role in stress related responses in S. richardsonii. The phylogenetic analysis showed that the two snow trout species GPDH share the same clade with characterized GPDHs from other teleost fishes suggesting a common evolutionary origin and a similar catalytic function. In addition, the Ka/Ks ratios of these sequences suggested that they are under purifying selection. Moreover, the expression profile of GPDH gene among co generic species of genus Schizothorax showed that GPDH cDNA expression was highest in S. richardsonii and lowest in S. esocinus which gives an indication of species specific adaptation in relation to different geographical areas.     

Berger C. Mayne (1920–2011): a friend and his contributions to photosynthesis research

We provide here insights on the life and work of Berger C. Mayne (1920-2011). We remember and honor Berger, whose study of photosynthesis began with the most basic processes of intersystem electron transport and oxygen evolution, continued with application of fluorescence techniques to the study of photophosphorylation and the unique features of photosystems in specialized cells, and concluded with collaborative study of photosynthesis in certain nitrogen fixing symbioses. Berger loved the outdoors and was dedicated to preserving the environment and to social justice, and was a wonderful friend.