Role of micro-RNA132 and its long non coding SOX2 in diagnosis of lupus nephritis.

BACKGROUND: The skin and the kidney are commonly affected in systemic lupus erythematosus (SLE) with similar molecular mechanisms. Although clinical indicators of renal injury in SLE are fairly uncontroversial, few biomarkers are reliable. The role of micro-RNAs (mi-RNAs) in lupus nephritis (LN) pathogenesis has been investigated to help in early diagnosis. PURPOSE: The aim of work is to evaluate miRNA132 and SOX2 expressions in SLE Egyptian patients; with and without nephritis, and the relation between miRNA132 and its long non-coding gene SOX2 in both patients groups. RESEARCH DESIGN: This is a case-control study involving 100 SLE patients with and without LN (LN and non-LN groups), and 50 age-and sex-matched healthy controls. The study was carried out to detect miRNA132 and SOX2 expression by quantitative Real-Time Polymerase chain reaction methods. The SLE disease activity index (SLEDAI) was assessed. RESULTS: SLEDAI increased in LN compared to non-LN. Micro-RNA132 expression was significantly increased in patient groups compared to controls (p<0.01) and increased in LN more than non-LN group (p<0.001). SOX2 significantly decreased in patient groups compared to controls (p<0.001), and was more in LN compared to non-LN group (p<0.001). There was a negative correlation between miRNA132 and SOX2 expression in both patient groups (p<0.001). CONCLUSION: miRNA132 and SOX2 may play a role in SLE activity and help in the early non-invasive diagnosis of LN.

Effect of histone deacetylase inhibitor on epithelial-mesenchymal transition of liver fibrosis.

Liver fibrosis is an excessively reversible wound healing process and the fibrotic disorder is the activation of hepatic stellate cell that requires extensive alterations in gene expression. As reversible deacetylation of histone proteins modulate gene expression, we examined the effect of valproic acid (VPA) as selective histone deacetylase inhibitor on CCl-4 induced liver fibrosis. Thirty rats were divided into three equal groups; control group, fibrotic group and VPA-treated group. The rats were sacrificed after 6 weeks of liver fibrosis induction. The histopathological effect on liver tissue was examined. The expression of α-SMA and Smad-4 mRNA and serum levels of TGF-ß1, alanine aminotransferase, and aspartate aminotransferase were determined. Treatment of rats with VPA attenuated carbon tetrachloride-induced liver fibrosis. Moreover, α-SMA and Smad-4 expression was repressed under VPA treatment and both serum TGF-ß1 and liver enzymes were significantly decreased. The histone deacetylase inhibitor-1 VPA inhibits the epithelial-mesenchymal transition and affects hepatic stellate cell activation during liver fibrosis through downregulation of Smad4 and α-SMA expression which may serve as a promising agent in liver fibrosis treatment. © 2018 IUBMB Life, 70(6):511-518, 2018.

Pyrazole derivatives ameliorate synovial inflammation in collagen-induced arthritis mice model via targeting p38 MAPK and COX-2.

The type II collagen-induced arthritis (CIA) model and human rheumatoid arthritis exhibit similar characteristics. Both diseases involve the production of inflammatory cytokines and other mediators, triggering an inflammatory cascade linked to bone and cartilage damage. Recently, new pyrazole compounds with various pharmacological activities, including antimicrobial, anticancer, anti-inflammatory, and analgesic agents, have been reported. Our aim is to evaluate the therapeutic effectiveness of two newly synthesized pyrazole derivatives, M1E and M1G, in reducing inflammation and oxidative stress in a mouse model of collagen-induced arthritis. Arthritis was induced in DBA/1J mice, and the therapeutic effect of the M1E and M1G is assessed by measuring the arthritic index, quantifying the expression of inflammatory genes such as p38 MAPK, COX-2, IL1ß, MMP3, and TNF-α using real-time PCR and analyzing protein expression using western blotting for phosphorylated p38 MAPK and COX-2. Oxidative stress markers and hind paws joint histopathology were also evaluated. Treatment with the two pyrazole derivatives significantly (p < 0.001) improved the arthritic score; downregulated the expression of inflammatory genes p38 MAPK, COX-2, IL1ß, MMP3, and TNF-α; and reduced the protein expression of phosphorylated p3  MAPK and COX-2. In addition, both compounds ameliorated oxidative stress by increasing the activities of SOD and reducing the formation of MDA in the paw tissue homogenates. Both M1E and M1G significantly (p < 0.001) improved the pathological features of synovitis. The pyrazole derivatives, M1E and M1G, significantly reduced the arthritic score and the inflammatory cytokine expression, improved synovitis histopathology, and ameliorated oxidative stress in the CIA mice model.

The crystal structure of diadenosine tetraphosphate hydrolase from Caenorhabditis elegans in free and binary complex forms.

The crystal structure of C. elegans Ap(4)A hydrolase has been determined for the free enzyme and a binary complex at 2.0 A and 1.8 A, respectively. Ap(4)A hydrolase has a key role in regulating the intracellular Ap(4)A levels and hence potentially the cellular response to metabolic stress and/or differentiation and apoptosis via the Ap(3)A/Ap(4)A ratio. The structures reveal that the enzyme has the mixed alpha/beta fold of the Nudix family and also show how the enzyme binds and locates its substrate with respect to the catalytic machinery of the Nudix motif. These results suggest how the enzyme can catalyze the hydrolysis of a range of related dinucleoside tetraphosphate, but not triphosphate, compounds through precise orientation of key elements of the substrate.

Analysis of the catalytic and binding residues of the diadenosine tetraphosphate pyrophosphohydrolase from Caenorhabditis elegans by site-directed mutagenesis.

The contributions to substrate binding and catalysis of 13 amino acid residues of the Caenorhabditis elegans diadenosine tetraphosphate pyrophosphohydrolase (Ap(4)A hydrolase) predicted from the crystal structure of an enzyme-inhibitor complex have been investigated by site-directed mutagenesis. Sixteen glutathione S-transferase-Ap(4)A hydrolase fusion proteins were expressed and their k(cat) and K(m) values determined after removal of the glutathione S-transferase domain. As expected for a Nudix hydrolase, the wild type k(cat) of 23 s(-1) was reduced by 10(5)-, 10(3)-, and 30-fold, respectively, by replacement of the conserved P(4)-phosphate-binding catalytic residues Glu(56), Glu(52), and Glu(103) by Gln. K(m) values were not affected, indicating a lack of importance for substrate binding. In contrast, mutating His(31) to Val or Ala and Lys(83) to Met produced 10- and 16-fold increases in K(m) compared with the wild type value of 8.8 microm. These residues stabilize the P(1)-phosphate. H31V and H31A had a normal k(cat) but K83M showed a 37-fold reduction in k(cat). Lys(36) also stabilizes the P(1)-phosphate and a K36M mutant had a 10-fold reduced k(cat) but a relatively normal K(m). Thus both Lys(36) and Lys(83) may play a role in catalysis. The previously suggested roles of Tyr(27), His(38), Lys(79), and Lys(81) in stabilizing the P(2) and P(3)-phosphates were not confirmed by mutagenesis, indicating the absence of phosphate-specific binding contacts in this region. Also, mutating both Tyr(76) and Tyr(121), which clamp one substrate adenosine moiety between them in the crystal structure, to Ala only increased K(m) 4-fold. It is concluded that interactions with the P(1)- and P(4)-phosphates are minimum and sufficient requirements for substrate binding by this class of enzyme, indicating that it may have a much wider substrate range then previously believed.

Crystallization of a complex of Caenorhabditis elegans diadenosine tetraphosphate hydrolase and a non-hydrolysable substrate analogue, AppCH2ppA.

The molecule diadenosine tetraphosphate (Ap(4)A) has been suggested to be a component of the cellular response to metabolic stress and/or, via the intracellular Ap(3)A/Ap(4)A ratio, to be involved in differentiation and apoptosis. Thus, the enzyme Ap(4)A hydrolase has a key metabolic role through regulation of the intracellular Ap(4)A levels. Crystals of this enzyme from the nematode Caenorhabditis elegans have been obtained in the presence of a non-hydrolysable substrate analogue, AppCH(2)ppA. The crystals belong to space group P2(1), unit-cell parameters a = 57.6, b = 36.8, c = 68.9 A, beta = 114.2 degrees, and diffract to approximately 2.0 A. Determination of the structure of this complex will provide insights into the substrate specificity and catalytic activity of this class of enzymes.