Genetic transformation of Indian isolate of Lemna minor mediated by Agrobacterium tumefaciens and recovery of transgenic plants

Transgenic plants of an Indian isolate of Lemna minor have been developed for the first time using Agrobacterium tumefaciens and hard nodular cell masses ‘nodular calli’ developed on the BAP - pretreated daughter frond explants in B₅ medium containing sucrose (1.0 %) with 2,4-D (5.0 μM) and 2-iP (50.0 μM) or 2,4-D (50.0 μM) and TDZ (5.0 μM) under light conditions. These calli were co-cultured with A. tumefaciens strain EHA105 harboring a binary vector that contained genes for β-glucuronidase with intron and neomycin phosphortransferase. Transformed cells selected on kanamycin selection medium were regenerated into fronds whose transgenic nature was confirmed by histochemical assay for GUS activity, PCR analysis and Southern hybridization. The frequency of transformation obtained was 3.8 % and a period of 11–13 weeks was required from initiation of cultures from explants to fully grown transgenic fronds. The pretreatment of daughter fronds with BAP, use of non-ionic surfactant, presence of acetosyringone in co-cultivation medium, co-culture duration of 3 d and 16 h photoperiod during culture were found crucial for callus induction, frond regeneration and transformation of L. minor. This transformation system can be used for the production of pharmaceutically important protein and in bioremediation.     

Ligand-independent dimer formation of epidermal growth factor receptor (EGFR) is a step separable from ligand-induced EGFR signaling

Dimerization and phosphorylation of the epidermal growth factor (EGF) receptor (EGFR) are the initial and essential events of EGF-induced signal transduction. However, the mechanism by which EGFR ligands induce dimerization and phosphorylation is not fully understood. Here, we demonstrate that EGFRs can form dimers on the cell surface independent of ligand binding. However, a chimeric receptor, comprising the extracellular and transmembrane domains of EGFR and the cytoplasmic domain of the erythropoietin receptor (EpoR), did not form a dimer in the absence of ligands, suggesting that the cytoplasmic domain of EGFR is important for predimer formation. Analysis of deletion mutants of EGFR showed that the region between (835)Ala and (918)Asp of the EGFR cytoplasmic domain is required for EGFR predimer formation. In contrast to wild-type EGFR ligands, a mutant form of heparin-binding EGF-like growth factor (HB2) did not induce dimerization of the EGFR-EpoR chimeric receptor and therefore failed to activate the chimeric receptor. However, when the dimerization was induced by a monoclonal antibody to EGFR, HB2 could activate the chimeric receptor. These results indicate that EGFR can form a ligand-independent inactive dimer and that receptor dimerization and activation are mechanistically distinct and separable events.     

On the interaction of UV-B radiation (280–315 nm) with water stress in crop plants

Cowpea ( Vigna unguiculata L. Walp.) seedlings (3-day-old) were subjected to 4 kinds of experimental treatments: (1) control without exposure to any stress (−D-UV), (2) moderate water stress with no UV-B irradiation (+D-UV), (3) no water stress but to UV-B radiation (−D+UV). and (4)moderate water stress and exposure to UV-B (+D+UV). UV-B and drought stress in the combined form elicited beneficial effects on the morphological and growth characteristics, and a few additive inhibitory effects in some functional processes. An increase in the specific leaf weight (SLW) was observed in the combination of stresses, which could be a defence mechanism against UV-B. The combination of stresses promoted the synthesis of anthocyanins and phenolic compounds. The responses of plants to the combination of stresses indicate that during simultaneous exposure of plants to multiple stresses, one form of stress could minimize the damage by the other. The enhancement of superoxide dismutase (SOD) and catalase activities appear to serve as acclimation mechanisms to scavenge the toxic, free radicals of oxygen produced under stress conditions. However, the inhibition in nitrate metabolism was greater in the combined stresses than in either of the stresses imposed separately. The results of this study illustrate that the interaction of stresses during simultaneous multiple stress conditions brings out certain beneficial effects.     

Detection of ischemia-reperfusion cardiac injury by cardiac muscle chemiluminescence

Various methods have been used in the past to assess the implication of oxygen free radicals (OFR) in ischemia-reperfusion-induced cardiac injury. Luminol-enhanced tert-butyl-initiated chemiluminescence in cardiac tissue reflects oxidative stress and is a very sensitive method. It was used to elucidate the role of OFR in cardiac injury due to ischemia and reperfusion. Studies were conducted on perfused isolated rabbit hearts in three groups (n = 8 in each): I, control; II, submitted to global ischemia for 30 min; III, submitted to ischemia for 30 min followed by reperfusion for 60 min. The heart tissue was then assayed for chemiluminescence (CL); content of malondialdehyde (MDA), an indicator of OFR-induced cardiac injury; and activity of tissue levels of antioxidants [superoxide dismutase (SOD), catalase, glutathione peroxidase (GSH-Px)].The control values for left and right ventricular CL and malondialdehyde were 81.1 ± 15.4 (S.E.) and 182.4 ± 50.3 (S.E.), mv-min-mg protein^–1; and 0.024 ± 0.006 (S.E.) and 0.324 ± 0.005 (S.E.) nmoles-mg protein^–1 respectively. Ischemia produced an increase in the cardiac CL (3.3 to 4.4 fold) and MDA content (2 to 2.6 fold). Reperfusion following ischemia also produced similar changes in CL and MDA content. The control values for activity of left ventricular SOD, catalase, and GSH-Px were 45.77 ± 1.73 (S.E.) U-mg protein^–1 5.35 ± 0.51 (S.E.) K-10^–3-sec^–1-mg protein^–1, and 77.50 ± 7.70 (S.E.) nmoles NADPH-min^–1-mg protein^–1 respectively. Activities of SOD and catalase decreased during ischemia but were similar to control values in ischemic-reperfused hearts. The GSH-Px activity of left ventricle was unaffected by ischemia, and ischemia-reperfusion. GSH-Px activity of the right ventricle increased with ischemia, and ischemic-reperfusion.These results indicate that cardiac tissue chemiluminescence would be a useful and sensitive tool for the detection of oxygen free radical-induced cardiac injury.     

Design,Synthesis and Pharmacological Screening of Novel Antihypertensive Agents Using Hybrid Approach

Eight derivatives of general formula 2-(2-(4-(3-((5-substituted methylene)-4-oxo-2-(phenylimino)thiazolidin-3-yl)-2-hydroxypropylamino)benzoyl)hydrazinyl)-2-oxoethyl nitrate were synthesized and tested for electrocardiographic, antiarrhythmic, vasorelaxing and antihypertensive activity as well as for in-vitro nitric oxide (NO) releasing ability. Compound 8b 2-(2-(4-(3-(5-benzyliden-4-oxo-2-(phenylimino)thiazolidin-3-yl)-2-hydroxypropylamino)benzoyl)hydrazinyl)-2-oxoethyl nitrate, was the most potent in this series. The pharmacological results suggested that the antiarrhythmic effects of these compounds were related to their adrenolytic properties which are believed to be due to the presence of the 5-(substituted)methylen-2-(phenylimino)thiazolidin-4-one moiety with less bulky, electron donating substituent on the phenyl ring at 5th position of the thiazolidin-4-one. In conclusion, most of the synthesized compounds were significantly potent as antiarrhythmic and antihypertensive; this might be due to the presence of different pharmacopores which might act at different locations with different mode of action. Further insights of the same can be obtained by doing investigation at receptor level. The potency of compounds 8a–8h were promising enough to continue further experiments.