Shafaq Rasool

Cloning and expression analysis of chalcone synthase gene from Coleus forskohlii

Flavonoids are an important class of secondary metabolites that play various roles in plants such as mediating defense, floral pigmentation and plant–microbe interaction. Flavonoids are also known to possess antioxidant and antimicrobial activities. Coleus forskohlii (Willd.) Briq. (Lamiaceae) is an important medicinal herb with a diverse metabolic profile, including production of a flavonoid, genkwanin. However, components of the flavonoid pathway have not yet been studied in this plant. Chalcone synthase (CHS) catalyses the first committed step of flavonoid biosynthetic pathway. Full-length cDNA, showing homology with plant CHS gene was isolated from leaves of C. forskohlii and named CfCHS (GenBank accession no. KF643243). Theoretical translation of CfCHS nucleotide sequence shows that it encodes a protein of 391 amino acids with a molecular weight of 42.75 kDa and pI 6.57. Expression analysis of CfCHS in different tissues and elicitor treatments showed that methyl jasmonate (MeJA) strongly induced its expression. Total flavonoids content and antioxidant activity of C. forskohlii also got enhanced in response to MeJA, which correlated with increased CfCHS expression. Induction of CfCHS by MeJA suggest its involvement in production of flavonoids, providing protection from microbes during herbivory or mechanical wounding. Further, our in silico predictions and experimental data suggested that CfCHS may be posttranscriptionally regulated by miR34.

Anti-asthmatic activity of azepino [2, 1-b] quinazolones, synthetic analogues of vasicine, an alkaloid from Adhatoda vasica

Asthma is characterized by persistent airway inflammation caused by over expression of pro-inflammatory immune response, predominantly by eosinophils and lymphocytes. Lymphocytes (CD4+ Th2) have been documented to be responsible for the pathogenesis of asthma by secreting Th2 cytokines and activating eosinophils, leading to airway hypersensitivity. Secretion of Th2 cytokines has been shown critical for the induction of the characteristic airway inflammation in humans and animal models of asthma. These cytokines influence the inflammatory response and lead to the pathological changes associated with asthma. In the present study, 10 azepino [2,1-b] quinazolone derivatives (R1 to R10) were synthesised and evaluated for their anti-asthmatic activity using a murine model of asthma. The compounds R2, R4, R6, R7 and R8 caused a notable decrease Th2 cytokine secretion and eosinophilia in asthma-induced animals. However, the decrease was highly significant in case of R8-treated animals. Crystal structure of R8 was made by X-ray crystallography. Molecular modelling studies were done for the compound R8 with transcription factors STAT6 and GATA3 which are the main transcription factors responsible for Th2 cell differentiation. Also the pharmacokinetics of R8 was carried out in mice after oral and intravenous administrations.

Ethanolic Extract of Alternanthera sessilis (AS-1) Inhibits IgE-mediated Allergic Response in RBL-2H3 Cells

The present study was designed to investigate the anti-allergic effects of ethanolic extract of Alternanthera sessilis (AS-1) in rat basophilic leukemia (RBL-2H3) cells. It significantly reduced the β-hexosaminidase release from anti-DNP-IgE sensitized RBL-2H3 cells. AS-1also inhibited the IgE antibody-induced increase in Interleukin-6 (IL-6), TNF-α, IL-13 and IL-4 production in these cells. The inhibitory effect of AS-1 on these cytokine was found to be nuclear factor-KB (NF-kB) dependent, as it attenuated the degradation of IKBa and nuclear translocation of NFkB. In addition, AS-1 significantly attenuated the DNP HAS-induced intracellular Ca2+ release from these cells, which makes us speculate strongly that the decreased intracellular Ca2+ is involved in the inhibitory effect of AS-1 on β-hexoaminidase release. Taken together, anti-allergic effects of AS-1 suggest possible therapeutic application of this extract in allergic diseases.

Evaluation of cellobiose dehydrogenase and laccase containing culture fluids of Termitomyces sp. OE147 for degradation of Reactive blue 21

Highlights • Use of culture filtrates of Termitomyces sp. OE147.• Decolourization and degradation of Reactive blue 21.• Cellobiose dehydrogenase and laccase as effective redox couple.

Homology modeling and docking study of chalcone synthase gene (CfCHS) from Coleus forskohlii

Abstract Flavonoids are important secondary metabolites in plants. Chalcone synthase (CHS) catalyzes the first committed step in the flavonoids biosynthesis. CHS belongs to type ΙΙΙ polyketide synthases that are known for their broad substrate specificity and catalytic potential toward a wide range of thioesters, to produce diverse novel polyketides of pharmaceutical importance. In this study, an in silico approach was used to understand the structure and function of CHS protein from an important medicinal plant Coleus forskohlii. A homology model of CfCHS was built and docking studies were carried out using 25 ligands. Best four docked ligands in proposed binding pocket of CfCHS were: Cinnamoyl CoA, 2-Carbamoylbenzoyl CoA, Benzoyl CoA and p-Coumaroyl CoA. Cys 164, His 304, and Asn 337 were found to be catalytic residues of CfCHS. Further two important residues, Phe 216 and Phe 266, were found to be the gatekeeper residues involved in π–π interaction with ligands. Present study revealed broad spectrum substrate profile of CfCHS and important key residues involved in substrate binding. This is the first report of homology modeling and docking analysis of CHS from C. forskohlii.

Therapeutic effects of R8, a semi-synthetic analogue of Vasicine, on murine model of allergic airway inflammation via STAT6 inhibition

This is a follow-up study of our previous work in which we screened a series of Vasicine analogues for their anti-inflammatory activity in a preventive OVA induced murine model of asthma. The study demonstrated that R8, one of the analogues, significantly suppressed the Th2 cytokine production and eosinophil recruitment to the airways. In the present study, we have been using two standard experimental murine models of asthma, where the mice were treated with R8 either during (preventive use) or after (therapeutic use) the development of asthma features. In the preventive model, R8 reduced inflammatory cell infiltration to the airways, OVA specific IgE and Th2 cytokine production. Also, the R8 treatment in the therapeutic model decreased methacholine induced AHR, Th2 cytokine release, serum IgE levels, infiltration of inflammatory cells into the airways, phosphorylation of STAT6 and expression of GATA3. Moreover, R8 not only reduced goblet cell metaplasia in asthmatic mice but also reduced IL-4 induced Muc5AC gene expression in human alveolar basal epithelial cells. Further, R8 attenuated IL-4 induced differentiation of murine splenocytes into Th2 cells in vitro. So, we may deduce that R8 treatment profoundly reduced asthma features by attenuating the differentiation of T cells into Th2 cells by interfering with the binding of IL-4 to its receptor in turn decreasing the phosphorylation of STAT6 and expression of GATA3 in murine model of asthma. These preclinical findings suggest a possible therapeutic role of R8 in allergic asthma.