Synthesis and pharmacological study of novel pyrido-quinazolone analogues as anti-fungal, antibacterial, and anticancer agents

A versatile method for novel pyrido-quinazolones was described here and tested for anti-fungal, antibacterial, and anticancerous activities. These synthesized compounds were characterized on the basis of spectroscopic techniques and evaluated for specific radiopharmaceuticals. Preliminary radiolabeling results with (99m)Tc and biological evaluation studies showed promising results for further evaluation in vivo. The efficiency of labeling was more than 98% and complexes were stable for about 18 h at 25 degrees C in the presence of serum.     

Magnesium-dependent phosphatase-1 is a protein-fructosamine-6-phosphatase potentially involved in glycation repair

Fructosamine-3-kinase (FN3K) is a recently described protein-repair enzyme responsible for the removal of fructosamines, which are the products of a spontaneous reaction of glucose with amines. We show here that, compared with glucose, glucose 6-phosphate (Glu-6-P) reacted 3-6-fold more rapidly with proteins and 8-fold more rapidly with N-alpha-t-Boc-lysine, being therefore a more significant intracellular glycating agent than glucose in skeletal muscle and heart. Fructosamine 6-phosphates, which result from the reaction of amines with Glu-6-P, were not substrates for FN3K. However, a phosphatase that dephosphorylates protein-bound fructosamine 6-phosphates was found to be present in rat tissues. This enzyme was purified to near homogeneity from skeletal muscle and was identified as magnesium-dependent phosphatase-1 (MDP-1), an enzyme of the haloacid dehalogenase family with a putative protein-tyrosine phosphatase function. Human recombinant MDP-1 acted on protein-bound fructosamine 6-phosphates with a catalytic efficiency >10-fold higher than those observed with its next best substrates (arabinose 5-phosphate and free fructoselysine 6-phosphate) and >100-fold higher than with protein-phosphotyrosine. It had no detectable activity on fructosamine 3-phosphates. MDP-1 dephosphorylated up to approximately 75% of the fructosamine 6-phosphates that are present on lysozyme after incubation of this protein with Glu-6-P. Furthermore, lysozyme glycated with Glu-6-P was converted by MDP-1 to a substrate for FN3K. We conclude that MDP-1 may act physiologically in conjunction with FN3K to free proteins from the glycation products derived from Glu-6-P.     

A novel dermal tissue construct: Development and in vitro characterization

A dermal tissue construct composed of human dermal fibroblasts and a chitosan sponge has been developed, targeted towards the treatment of diabetic nonhealing ulcers. The construct has been designed in a way that the dermal fibroblasts are arranged as a three‐dimensional sheet adhered entirely on one side of the chitosan sponge. This design would allow maximal diffusion of growth factors from the cells to the wound bed when the construct is applied on the wound with the cellular sheet side making contact with the wound bed. The diffusion of secreted growth factors would take place directly from cells to the wound bed without being impeded by a matrix. The cells are present at a high density in the dermal construct, which would aid in accelerated wound healing. The construct has a porous chitosan sponge base, which would allow gas exchange, and renders the dermal construct very flexible so that it would take the shape of the wound contours well, while having mechanical integrity. The viability of cells in the construct is greater than 90%. The dermal construct produces a high amount of vascular endothelial growth factor, from 42 ng to 31 ng in 24 h. The construct also produces high amounts of Interleukin‐8 (IL‐8), from 375 ng to 1065 ng in the first 24 h. Both VEGF and IL‐8 have important roles in the healing of chronic diabetic ulcers. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010     

Altered chemokine production and accumulation of regulatory T cells in intestinal adenomas of APCMin/+ mice

Tumor progression in the colon moves from aberrant crypt foci to adenomatous polyps to invasive carcinomas. The composition of the tumor-infiltrating leukocyte population affects the ability of the immune system to fight the tumor. T cell infiltration into colorectal adenocarcinomas, particularly T helper 1 (Th1) type T cells as well as increased regulatory T cell (Treg) frequencies, is correlated with improved prognosis. However, whether Th1 cells and Tregs are already present at the adenoma stage is not known. In this study, the APC^Min/+ mouse model of intestinal adenomatous polyposis was used to investigate tumor-associated lymphocyte subsets and the mechanisms of their accumulation into gastrointestinal adenomas. Compared to unaffected tissue, adenomas accumulated CD4⁺FoxP3⁺ putative Treg in parallel with lower frequencies of conventional T cells and B cells. The accumulation of Treg was also observed in human adenomatous polyps. Despite high Treg numbers, the function of conventional T cells present in the APC^Min/+ adenomas was not different from those in the unaffected tissue. Adenomas displayed an altered chemokine balance, with higher CCL17 and lower CXCL11 and CCL25 expression than in the unaffected tissue. In parallel, CXCR3⁺ Tregs were largely absent from adenomas. The data indicate that already in early stages of tumor development, the balance of lymphocyte-recruiting chemokines is altered possibly contributing to the observed shift toward higher frequencies of Treg.     

Allergenic pollen in the atmosphere of Rohtak city,Haryana (India): a pioneer study

An attempt has been made in the present study to assess the allergenicity of dominant pollen types recorded from the atmosphere of Rohtak city. Skin prick test was performed with the antigenic extracts of 22 pollen types on 150 local patients who visited Asthma Clinic, University of Health Sciences, Rohtak. Markedly positive skin reactions (2+ and above) varied from 4.6 to 20.6 % to various pollen antigens. Cenchrus ciliaria (20.6 %), Zea mays (20 %) and Pennisetum typhoides (19.3 %) were the pollen allergens exhibiting maximum sensitivity. Antigenic extract of Cassia occidentalis, Cynodon dactylon and Ricinus communis showed marked skin reactivity in 18.6 % of patients. Prosopis juliflora, Chenopodium murale, Amaranthus spinosus, Cassia fistula and Cassia siamea showed 2+ and above reactions in 16.6, 15.3, 14.6 and 14.0 % of the local patients, respectively. Least reactivity (4.6 %) was shown to the antigenic extract of Cyperus rotundus. Out of 52 sera screened for the presence of specific IgE antibodies against different antigenic extracts, only 5.5 % showed >60 % binding. About 30 % and above binding was shown to the antigenic extracts of Z. mays, A. spinosus, R. communis and Xanthium strumarium. The concordance between positive skin reaction and serum-specific IgE antibodies ranged from 15 to 69 %.     

A Highlights from MBoC Selection: Role of the loop L4,5 in allosteric regulation in mtHsp70s: in vivo significance of domain communication and its implications in protein translocation

Mitochondrial Hsp70 (mtHsp70) is essential for a vast repertoire of functions, including protein import, and requires effective interdomain communication for efficient partner-protein interactions. However, the in vivo functional significance of allosteric regulation in eukaryotes is poorly defined. Using integrated biochemical and yeast genetic approaches, we provide compelling evidence that a conserved substrate-binding domain (SBD) loop, L_4,5, plays a critical role in allosteric communication governing mtHsp70 chaperone functions across species. In yeast, a temperature-sensitive L_4,5 mutation (E467A) disrupts bidirectional domain communication, leading to compromised protein import and mitochondrial function. Loop L_4,5 functions synergistically with the linker in modulating the allosteric interface and conformational transitions between SBD and the nucleotide-binding domain (NBD), thus regulating interdomain communication. Second-site intragenic suppressors of E467A isolated within the SBD suppress domain communication defects by conformationally altering the allosteric interface, thereby restoring import and growth phenotypes. Strikingly, the suppressor mutations highlight that restoration of communication from NBD to SBD alone is the minimum essential requirement for effective in vivo function when primed at higher basal ATPase activity, mimicking the J-protein–bound state. Together these findings provide the first mechanistic insights into critical regions within the SBD of mtHsp70s regulating interdomain communication, thus highlighting its importance in protein translocation and mitochondrial biogenesis.     

Central leptin gene therapy blocks ovariectomy-induced adiposity

Objective: In this study, we tested the hypothesis that insufficiency of leptin restraint in the hypothalamus is responsible for promoting weight gain and adiposity after ovariectomy (ovx). Whether increasing leptin transgene expression can overcome the diminution in leptin restraint was evaluated in ovx rats. Research Methods and Procedures: Enhanced leptin or green fluorescent protein (GFP; control) transgene expression was induced by a single intracerebroventricular injection of recombinant adeno‐associated viral vector encoding either leptin gene (rAAV‐lep) or GFP gene (rAAV‐GFP; control) in acutely and chronically ovx rats. Body weight and food intake responses were monitored weekly. White adipose tissue (WAT) mass and serum levels of WAT‐derived hormones, leptin, and adiponectin were analyzed at termination of the experiments. Results and Discussion: An increase in leptin transgene expression in the hypothalamus initiated soon after ovx blocked hyperphagia and body weight gain and markedly suppressed WAT mass and adipokines, leptin, and adiponectin. Similar suppression of weight gain and adiposity and serum leptin and adiponectin levels after intracerebroventricular rAAV‐lep injection in chronically ovx rats were observed concomitant with unchanged daily food intake. These findings are consistent with the hypothesis that in the absence of ovarian steroids, the existent insufficiency of leptin restraint at the hypothalamic level can be overcome with ectopic leptin expression, thereby reinstating central control on weight and adiposity.     

Changes in airborne pollen concentrations in Delhi,India

In order to find the qualitative and quantitative changes in airborne pollen concentrations in Delhi metropolis area an aerobiological survey was undertaken from September 1990 to August 1997. Air samples were collected daily using a Rotorod Aeroallergen Sampler at 10?m above the ground level. Ninety-four pollen types were recorded and the major contributors include Morus, Cannabis, Chenopod/Amaranth, Prosopis, Artemisia, and Eucalyptus. Ten pollen types contributed 90% of the total pollen load. Two major pollen seasons were recorded each year (February–April and September–November), although pollen grains in low frequency were recorded throughout the whole year. A significant reduction in pollen concentration was observed in subsequent years. The number of Morus, Cannabis, Prosopis, and Artemisia pollen decreased considerably while the number of Ricinus communis pollen did not show any considerable change during the study period. It is suggested that the reduction in pollen numbers from 1990 to 1997 in Delhi is due to massive clearing of vegetation for developmental activities of the city.     

TIM3 Mediates T Cell Exhaustion during Mycobacterium tuberculosis Infection

While T cell immunity initially limits Mycobacterium tuberculosis infection, why T cell immunity fails to sterilize the infection and allows recrudescence is not clear. One hypothesis is that T cell exhaustion impairs immunity and is detrimental to the outcome of M. tuberculosis infection. Here we provide functional evidence for the development T cell exhaustion during chronic TB. Second, we evaluate the role of the inhibitory receptor T cell immunoglobulin and mucin domain–containing-3 (TIM3) during chronic M. tuberculosis infection. We find that TIM3 expressing T cells accumulate during chronic infection, co-express other inhibitory receptors including PD1, produce less IL-2 and TNF but more IL-10, and are functionally exhausted. Finally, we show that TIM3 blockade restores T cell function and improves bacterial control, particularly in chronically infected susceptible mice. These data show that T cell immunity is suboptimal during chronic M. tuberculosis infection due to T cell exhaustion. Moreover, in chronically infected mice, treatment with anti-TIM3 mAb is an effective therapeutic strategy against tuberculosis.