Dipeptidyl Peptidase IV Inhibition Activates CREB and Improves Islet Vascularization through VEGF-A/VEGFR-2 Signaling Pathway

Substitution of pancreatic islets is a potential therapy to treat diabetes and it depends on reconstitution of islet’s capillary network. In this study, we addressed the question whether stabilization of Glucagon-Like-Peptide-1 (GLP-1) by inhibiting Dipeptidyl Peptidase-IV (DPP-IV) increases β-cell mass by modulating vascularization. Mouse or porcine donor islets were implanted under kidney capsule of diabetic mice treated with DPP-IV inhibitor sitagliptin. Grafts were analyzed for insulin production, β-cell proliferation and vascularization. In addition, the effect of sitagliptin on sprouting and Vascular Endothelial Growth Factor (VEGF)-A expression was examined ex vivo. The cAMP response element-binding (CREB) and VEGF-A/ Vascular Endothelial Growth Factor Receptor (VEGFR)-2 signaling pathway leading to islet vascularization was explored. Sitagliptin increased mean insulin content of islet grafts and area of insulin-positive tissue as well as β-cell proliferation. Interestingly, sitagliptin treatment also markedly increased endothelial cell proliferation, microvessel density and blood flow. Finally, GLP-1 (7-36) stimulated sprouting and VEGF expression, which was significantly enhanced by sitagliptin- mediated inhibition of DPP-IV. Our in vivo data demonstrate that sitagliptin treatment phosphorylated CREB and induced islet vascularization through VEGF-A/VEGFR-2 signaling pathway. This study paves a new pathway for improvement of islet transplantation in treating diabetes mellitus.     

Synthesis and characterization of biologically significant 5,5′-(1,4-phenylene)bis(1-N-alkoxyphthalimido-3-aryl-2-pyrazoline) derivatives

A facile synthesis of 5,5′-(1,4-phenylene)bis(3-aryl-2-pyrazolines) 4a-g has been achieved by the cyclo-addition reaction of hydrazine hydrate with bis-substituted chalcones 3a-g, which in turn were prepared by the Clasien-Schmidt condensation of p-substituted acetophenones 1a-g with terephthaldehyde. Condensation of 4a-g with ω-bromoalkoxyphthalimides 5a-b afforded the titled compounds 6a-n, some of which exhibited significant antimalarial as well as antimicrobial activity.     

The Role of Cross-Chain Ionic Interactions for the Stability of Collagen Model Peptides

The contribution of ionic interactions to the stability of the collagen triple helix was studied using molecular dynamics (MD) simulations and biophysical methods. To this end, we examined the stability of a host-guest collagen model peptide, Ac-GPOGPOGPYGXOGPOGPO-NH₂, substituting KGE, KGD, EGK, and DGK for the YGX sequence. All-atom, implicit solvent MD simulations show that the fraction of cross-chain ionic interactions formed is different, with the most pronounced in the KGE and KGD sequences, and the least in the DGK sequence. To test whether the fraction of cross-chain ionic interactions correlates with the stability, experimental measurements of thermostability were done using differential scanning calorimetry and circular dichroism spectroscopy. It was found that the melting temperature is very similar for KGE and KGD peptides, whereas the EGK peptide has lower thermostability and the DGK peptide is the least thermostable. A novel, to our knowledge, computational protocol termed temperature-scan MD was applied to estimate the relative stabilities of the peptides from MD simulations. We found an excellent correlation between transition temperatures obtained from temperature-scan MD and those measured experimentally. These results suggest the importance of cross-chain ionic interactions for the stability of collagen triple helix and confirm the utility of MD simulations in predicting interactions and stability in this system.     

Cloning and functional characterization of 3-hydroxy-3-methylglutaryl coenzyme A reductase gene from Withania somnifera: an important medicinal plant

Withania somnifera (L.) Dunal is one of the most valuable medicinal plants synthesizing a large number of pharmacologically active secondary metabolites known as withanolides, the C₂₈-steroidal lactones derived from triterpenoids. Though the plant has been well characterized in terms of phytochemical profiles as well as pharmaceutical activities, not much is known about the biosynthetic pathway and genes responsible for biosynthesis of these compounds. In this study, we have characterized the gene encoding 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR; EC 1.1.1.34) catalyzing the key regulatory step of the isoprenoid biosynthesis. The 1,728-bp full-length cDNA of Withania HMGR (WsHMGR) encodes a polypeptide of 575 amino acids. The amino acid sequence homology and phylogenetic analysis suggest that WsHMGR has typical structural features of other known plant HMGRs. The relative expression analysis suggests that WsHMGR expression varies in different tissues as well as chemotypes and is significantly elevated in response to exposure to salicylic acid, methyl jasmonate, and mechanical injury. The functional color assay in Escherichia coli showed that WsHMGR could accelerate the biosynthesis of carotenoids, establishing that WsHMGR encoded a functional protein and may play a catalytic role by its positive influence in isoprenoid biosynthesis.     

Molecular breeding of ameliorating commercial pearl millet hybrid for downy mildew resistance

Downy mildew (DM) caused by Sclerospora graminicola is the most calamitous disease of pearl millet. Therefore, for introgression of DM resistance (DMR) in HHB 197 (MH-1302), an elite pearl millet hybrid, a marker-assisted breeding was undertaken by targeting three DMR loci on linkage groups (LGs) 1, 2 and 4. Breeding programme was initiated by crossing HBL 11 (DM susceptible), male parent of HHB 197 hybrid with ICMP 451 (DM-resistant) to produce true \(\hbox {F}_{1}\) plants. By conducting three rounds of backcrossing and selection, \(\hbox {BC}_{3}\hbox {F}_{1}\) lines were generated. Foreground selection was employed using six polymorphic simple sequence repeat (SSR) markers of the 18 total selected markers. Four of these markers were linked to LG 1, five to LG 2 and nine to LG 4. Background selection was performed in \(\hbox {BC}_{3}\hbox {F}_{1}\) generation using 33 polymorphic SSR markers of a total of 56 evenly spread SSR markers in the pearl millet genome to check recovery of recurrent parent genome. On the basis of genotypic selection (foreground as well as background) using selected SSR markers, agronomic performance in field and DM screening in greenhouse; 10 improved HBL 11 lines were selected and crossed with ICMA 97111 to produce DM-resistant HHB 197 hybrid versions. Six putatively improved HHB 197 hybrids were successfully tested in first year trials at Hisar and Bawal locations of Haryana and two selected versions with higher yield and zero DM incidence will be further tested in multilocation trials.     

Incidence of metallo-beta-lactamase-producing <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis> isolates from hospital setting in Pakistan

The aim of this study was monitoring and surveillance in different wards of the PIMS hospital, Islamabad, to understand emerging challenges of antibiotic resistance in particular association with most virulent serotypes of Klebsiella pneumoniae. The study was conducted during March 2015 to September 2015. The study showed that rate of isolation of K. pneumoniae was 37% (103 positives out of a total of 277 clinical samples) and 7.7% (8) were phenotypically and genotypically confirmed to be metallo-β-lactamase resistant (carbapenem resistant) and all of them were multidrug resistant (MDR). These carbapenem-resistant isolates were isolated from blood, endotracheal tubes, and pus. Molecular screening for the presence of integrons indicated that distribution of class I integrons (87.5% of carbapenem-resistant K. pneumoniae isolates) was higher than class II integrons (1.25%) among given isolates. The study indicated that exposure of metallo-beta-lactamase-producing strains through hospitalizations increases the chances of spread of MDR pathogens. There is an urgent need for effective surveillance and monitoring strategies to control the spread of extremely resistant K. pneumoniae implicated in nosocomial infections leading to the increased health burden and enforcement of policy guideline on appropriate antibiotics usage.