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Nanoparticle-reinforced polymer-based scaffolding matrices as artificial bone-implant materials are potential suitors for bone regenerative medicine as they simulate the native bone. In the present work, a series of bioinspired, osteoconductive tricomposite scaffolds made up of nano-hydroxyapatite (NHA) embedded xanthan gum-chitosan (XAN-CHI) polyelectrolyte complex (PEC) are explored for their bone-regeneration potential. The Fourier transform infrared spectroscopy studies confirmed complex formation between XAN and CHI and showed strong interactions between the NHA and PEC matrix. The X-ray diffraction studies indicated regulation of the nanocomposite (NC) scaffold crystallinity by the physical cues of the PEC matrix. Further results exhibited that the XAN-CHI/NHA5 scaffold, with a 50/50 (polymer/NHA) ratio, has optimized porous structure, appropriate compressive properties, and sufficient swelling ability with slower degradation rates, which are far better than those of CHI/NHA and other XAN-CHI/NHA NC scaffolds. The simulated body fluid studies showed XAN-CHI/NHA5 generated apatite-like surface structures of a Ca/P ratio ∼1.66. Also, the

Zinc oxide nanoparticles (ZnO NPs) were synthesized by a green method using Azadirachta indica leaf extract. The structure of the prepared ZnO (NPs) were characterized by FT-IR, XRD, SEM-EDX and TEM analyses. The biosynthesized ZnO (NPs) were then used as a catalyst for the synthesis of steroidal dihydropyrazole derivatives through a one-pot multicomponent reaction involving phenyl acetylene and hydrazine derivatives. The anticancer activity of newly synthesized compounds were evaluated against three cancer cell lines namely HeLa (human cervical carcinoma), Hep3B (human hepatocellular carcinoma) and MCF7 (human breast adenocarcinoma) by MTT assay. The tested compounds were found to be active against all cancer cell lines and less toxic towards normal peripheral blood mononuclear cells (PBMCs). Antioxidant activity have also been investigated via free radical scavenging ability using DPPH, FRAP and ABTS assay. The tested compounds were found to exhibit moderate to good antioxidant activity which increases with increase in the concentration of steroidal dihydropyrazoles. Among all the tested steroidal dihydropyrazoles, compound 17 is found to be most active.

Tuberculosis (TB) is a significant and continuing problem worldwide, with a death toll of around 1.5 million human lives annually. BCG, the only vaccine against TB, offers a varied degree of protection among human subjects in different regions and races of the world. The majority of the population living near the tropics carries a varying degree of tolerance against BCG due to the widespread prevalence of non-tuberculous mycobacteria (NTM). Interestingly, ≈90% of the Mycobacterium tuberculosis (Mtb) infected population restrain the bacilli on its own, which strengthens the notion of empowering the host immune system to advance the protective efficacy of existing mycobacterial vaccines. In general, Mtb modulates IL-10/STAT3 signaling to skew host mononuclear phagocytes toward an alternatively activated, anti-inflammatory state that helps it thrive against hostile immune advances. We hypothesized that modulating the IL-10/STAT3 driven anti-inflammatory effects in mononuclear cells may improve the prophylactic ability of TB vaccines. This study investigated the immunotherapeutic ability of a porphyrin based small molecule inhibitor of IL-10/STAT3 axis, 5, 15-diphenyl porphyrin (DPP), in improving anti-TB immunity offered by second generation recombinant BCG30 (rBCG30-ARMF-II®) vaccine in mice. The DPP therapy potentiated vaccine induced anti-TB immunity by down-modulating anti-inflammatory responses, while simultaneously up-regulating pro-inflammatory immune effector responses in the immunized host. The employed DPP based immunotherapy led to the predominant activation/proliferation of pro-inflammatory monocytes/macrophages/DCs, the concerted expansion of CD4+/CD8+ effector and central memory T cells, alongside balanced Th17 and Treg cell amplification, and conferred augmented resistance to aerosol Mtb challenge in rBCG30 immunized BALB/c mice.

Phosphopantetheine adenylyltransferase (PPAT, EC. 2.7.7.3) catalyzes an essential step in the reaction that transfers an adenylyl group from adenosine tri phosphate (ATP) to 4′-phosphopantetheine (pPant) yielding 3′- dephospho-coenzyme A (dPCoA) and pyrophosphate (PP) in the coenzyme A (CoA) biosynthesis pathway. The enzyme PPAT from Acinetobacter baumannii (AbPPAT) was cloned, expressed and purified. The binding studies of AbPPAT were carried out with two compounds, tri‑sodium citrate (TSC) and l -ascorbic acid (LAA, vitamin-C) using fluorescence spectroscopic (FS) and surface Plasmon resonance (SPR) methods. Both methods provided similar values of dissociation constants for TSC and LAA which were of the order of 10−8 M and 10−5 M respectively. The computer aided docking studies indicated fewer interactions of LAA with AbPPAT as compared to those of TSC. The freshly purified samples of AbPPAT were crystallized. The crystals of AbPPAT were soaked in the solutions containing TSC and LAA. However, the crystals of the complex of AbPPAT with LAA did not diffract well and hence the structure of the complex of AbPPAT with LAA could not be determined. On the other hand, the crystals of the complex of AbPPAT with TSC diffracted well and the structure was determined at 1.76 A resolution. It showed that TSC bound to AbPPAT at the ATP binding site and formed several intermolecular contacts including 12 hydrogen bonds. The results of binding studies for both TSC and LAA and the structure of the complex of AbPPAT with TSC clearly indicated a potential role of TSC and LAA as antibacterial agents.

Among various metal-based nanoparticles, silver nanoparticles (AgNPs) manifest superior inhibitory effect against several microorganisms. In fact, the AgNPs based therapy has been reported to inhibit both sensitive as well resistant isolates of bacteria and other disease causing microbes with equal propensity. Keeping this fact into consideration, we executed bio-mediated synthesis of AgNPs employing Hibiscus rosa sinensis flower extract. The as-synthesized AgNPs were evaluated for their potential to inhibit Streptococcus mutans (S. mutans), one of the main causative bacteria for dental caries. Beside several other reasons, orthodontic appliances have also been reported to facilitate infliction of oral cavity with a range of microbes including S. mutans. To determine the growth inhibitory and anti-adherence activity of AgNPs on orthodontic appliance, we performed microbiological assays employing AgNPs adsorbed on to the orthodontic wires. Topographic analysis of orthodontic wires was executed by scanning electron microscopy. In addition to antimicrobial and antibiofilm activity against oral S. mutans, the as-fabricated AgNPs demonstrated significant inhibitory and anti-biofilm properties against other biofilm forming bacteria such as Escherichia coli and Listeria monocytogenes as well.

Visceral leishmaniasis (VL)-related mortality and morbidity imposes a great deal of health concern across the globe. The existing anti-leishmanial drug regimen generally fails to eliminate newly emerging resistant isolates of this dreadful parasite. In such circumstances, the development of a prophylactic strategy to impart protection against the disease is likely to take center stage. In order to develop a promising prophylactic vaccine, it is desirable to identify an adequately potential vaccine candidate. In silico analysis of Leishmania tubulin folding cofactor D protein predicted its potential to activate both B- and T-cell repertoires. Furthermore, the ELISA employing anti-peptide27 (a segment of tubulin folding cofactor D) antibody revealed its proficiency in VL diagnosis and treatment monitoring. The peptide27 and its cocktail with another Leishmania peptide (peptide23) prompted the up-regulation of pro-inflammatory cytokines, such as IFN-γ, TNF-α, IL-2, IL-17, etc., and the down-regulation of immune-regulatory cytokines, such as IL-10, in the immunized BALB/c mice. Coherent to the consequence of peptide-specific humoral immune response, peptide cocktail-based immunization ensued in the predominant amplification of pathogen-specific IgG2a over the IgG1 isotype, up-regulated proliferation of T lymphocytes, and enhanced production of nitric oxide, reactive oxygen species, etc. We also established that the peptide cocktail modulated host MAPK signaling to favor the amplification of Th1-dominated immune response in the host. The peptide cocktail mediated the activation of the host immune armory, which was eventually translated into a significant decline in parasitic load in the visceral organs of experimental animals challenged with Leishmania donovani.

Aims and Objectives: In the present study we tried to improve anti-TB immunity offered by 2nd generation rBCG30 vaccine. We attempted to bolster rBCG30 invoked anti-TB immunity in mice through modulation of the IL-10/STAT3 interaction mediated anti-inflammatory program. The study was aimed towards generating optimum activated APCs, early either after vaccination (with rBCG30) or infection (with Mtb), which can process and present mycobacterial antigens more efficiently to T cells. It was envisage that downmodulating IL-10/STAT3 signaling may improve immunity and protection offered by rBCG30 vaccine. Methods: A small molecule immunomodulator aimed at disrupting anti-inflammatory signaling orchestrated by IL-10/STAT3 axis was administered in rBCG30 immunized mice. The comparative expansion of pathogen permissive Alternatively Activated Monocytes/Macrophages/DCs (AAMs/AADCs) and Classically Activated Monocytes/Macrophages/DCs (CAMs/CADCs), the cells that resist establishment of successful Mtb infection, was profiled in both peritoneal and splenic compartment. To assess innate and adaptive T cell responses, splenocytes cytokine production was checked ex vivo using sandwich ELISA. We also checked polyfunctional T cell response, T cell memory response and comparative Th17/Treg cells expansion using flow cytometry. Finally, to assess the real time efficacy of the employed immunotherapeutic strategy, in vivo protection study was undertaken in BALB/c mice challenge with aerosolized Mtb (H37Rv). Results: The small molecule mediated inhibition of IL-10/STAT3 signaling was resulted in improved pro inflammatory immune responses in both post-vaccination and post-infection immunotherapy schedules. The improved immune profile in rBCG30 vaccinated and immunotherapy administered mice was accompanied with enhanced expansion of CAMs/CADCs, elevated production of polyfunctional T cells producing signature Th1 cytokines, long lasting CD4+ and CD8+ memory T cell response and balanced Th17/Treg cells ratio. Interestingly, post-infection, and not post-vaccination immunotherapy, was able to significantly reduce mycobacterial loads in spleen and lungs of experimentally infected mice. Conclusions: Small molecule mediated transient disruption of anti-inflammatory effectors, such as IL-10/STAT3, during vaccination or post infection with Mtb, may serve to enhance the efficacy of existing and forthcoming TB vaccines. The strategy can also be tested concomitantly with standard chemotherapy (either in vaccinated or unvaccinated host) to enhance the efficacy of therapeutic regimen and to reduce the treatment length.

Several recent studies have contributed to our understanding of the need for host directed immunotherapeutic strategies against Mycobacterium tuberculosis (Mtb), either concomitantly with classical chemotherapy or as standalone intervention. Nevertheless, most of these studies were focused on enhancing chemotherapeutic potential of standard anti-tuberculosis chemotherapy and only few were aimed towards improving prophylactic immunity offered either by BCG or its potential replacements. In the present study we tried to address the potential gap with the aim to improve anti-TB immunity offered by 2nd generation rBCG30 vaccine. We attempted to bolster rBCG30 invoked anti-TB immunity in mice through modulation of the IL-10/STAT3 interaction mediated anti inflammatory program. An immunomodulator of IL-10/STAT3 signaling axis was administered in immunized mice to reinforce the anti-pathogen capabilities of mononuclear phagocyte system with expectation to improve T cell mediated secondary immune responses. We envisage that modulation of this crucial praxis may result in reduced expansion of pathogen permissive AAMs/AADCs (Alternatively Activated Monocytes/Macrophages/DCs) and augmented abundance of pro-inflammatory CAMs/CADCs (Classically Activated Monocytes/Macrophages/DCs), the cells that resist establishment of successful Mtb infection. The study was also aimed towards generating optimum activated APCs early either after vaccination or infection, which can process and present mycobacterial antigens more efficiently to T cells.

In the present study, we investigated potential of chitosan-based nanoparticles (CNPs) to deliver loaded therapeutic molecules to pathogen harboring macrophages. We fabricated stable CNPs employing ionic cross-linking method and evaluated their potential to target RAW 264.7 cells. The physicochemical characterization of as-synthesized CNPs was determined using electron microscopy, infrared microscopy and zeta potential measurement. Next, cellular uptake and intracellular localization studies of CNPs were followed in living RAW264.7 cells using confocal microscopy. We found that both Acr-1 loaded (CNP-A) and 4-SO4-GalNAc ligand harboring (CNP-L) chitosan nanoparticle experience increased cellular uptake by Mycobacterium smegmatis infected RAW cells. Following cellular digestion in model macrophage cell line (RAW), CNPs provide an increased immune response. Further, 4-SO4-GalNAc bearing CNP-L exhibits high binding affinity as well as antibacterial efficacy toward M. smegmatis. The data of the present study suggest that CNP-based nanoparticle offer a promising delivery strategy to target infected macrophages for prevention and eradication of intracellular pathogens such as M. smegmatis.

Amyloids are highly organized protein aggregates that arise from inappropriately folded versions of proteins or polypeptides under both physiological as well as simulated ambiences. Once thought to be irreversible assemblies, amyloids have begun to expose their more dynamic and reversible attributes depending upon the intrinsic properties of the precursor protein/peptide and experimental conditions such as temperature, pressure, structural modifications in proteins, or presence of chemicals in the reaction mixture. It has been repeatedly proposed that amyloids undergo transformation to the bioactive peptide/protein forms under specific conditions. In the present study, amyloids assembled from the model protein ovalbumin (OVA) were found to release the precursor protein in a slow and steady manner over an extended time period. Interestingly, the released OVA from amyloid depot was found to exhibit biophysical characteristics of native protein and reacted with native-OVA specific monoclonal as well as polyclonal antibodies. Moreover, antibodies generated upon immunization of OVA amyloidal aggregates or fibrils were found to recognize the native form of OVA. The study suggests that amyloids may act as depots for the native form of the protein and therefore can be exploited as vaccine candidates, where slow antigen release over extended time periods is a pre-requisite for the development of desired immune response.

Nano-sized drug delivery systems (NDDS) have been widely exploited to achieve targeted delivery of pharmaco-materials. Traditional pharmaceutical approaches, implied in the synthesis of nano-formulations, are obscure owing to the incompatible physico-chemical properties of the core drug as well as some other factors crucial in development of NDDS. Infact, most of the existing methods used in development of NDDS rely on usage of additives or excipients, a special class of chemicals. Barring few exceptions, the usage of synthetic excipients ought to be curtailed because of several associated undesirable features. Such issues necessitate strategies that lead to development of the synthetic excipient free drug delivery system. Plant based extracts have great potential to induce synthesis of nano-sized particles. Considering this fact, here we propose a prototype employing orange fruit juice (OJ) to facilitate bio-mediated synthesis of nano-sized supra-molecular assemblies of 5-fluorouracil (5-FU), a potent anticancer drug. The as-synthesized 5-FU Nanoparticles (NPs) retained the anti-neoplastic efficacy of the parent compound and induced apoptosis in cancer cells. The novel 5-FU NPs formulation demonstrated enhanced efficacy against DMBA induced experimental fibrosarcoma in the mouse model when compared to the micro-sized crystals of parent 5-FU drug.

Amyloids are highly organized protein aggregates that arise from inappropriately folded versions of proteins or polypeptides under both physiological as well as simulated ambiences. Once thought to be irreversible assemblies, amyloids have begun to expose their more dynamic and reversible attributes depending upon the intrinsic properties of the precursor protein/peptide and experimental conditions such as temperature, pressure, structural modifications in proteins, or presence of chemicals in the reaction mixture. It has been repeatedly proposed that amyloids undergo transformation to the bioactive peptide/protein forms under specific conditions. In the present study, amyloids assembled from the model protein ovalbumin (OVA) were found to release the precursor protein in a slow and steady manner over an extended time period. Interestingly, the released OVA from amyloid depot was found to exhibit biophysical characteristics of native protein and reacted with native-OVA specific monoclonal as well as polyclonal antibodies. Moreover, antibodies generated upon immunization of OVA amyloidal aggregates or fibrils were found to recognize the native form of OVA. The study suggests that amyloids may act as depots for the native form of the protein and therefore can be exploited as vaccine candidates, where slow antigen release over extended time periods is a pre-requisite for the development of desired immune response.

The EPR study of Cu2+ doped potassium oxalate monoperhydrate is carried out at room temperature. Two magnetically inequivalent interstitial sites are found embedded in orthorhombic crystal fields. The ground state wave functions are constructed. The obtained results are compared with that of isotypic crystal Cu2+ doped potassium oxalate monohydrate to explore the difference arising due to the presence of H2O2 in the former and H2O in the latter crystal. The spin-Hamiltonian parameters (g- and A-values) covalency parameter (α′2), the mixing coefficients (α, β) and Fermi contact term (k) are calculated for both the complexes.

EPR spectra of (i) CuO annealed in air at different temperatures up to 900 \ifmmode^\circ\else\textdegree\fi{}C and (ii) CuO reacted with BaO in the ratio 3:2 at different temperatures up to 900 \ifmmode^\circ\else\textdegree\fi{}C were studied. In (i), evidence for the formation of monomers, dimers, and tetramers of copper ions was found. In (ii), in addition to those spectra obtained in (i), aggregates of four copper ions coupled through cyclic exchange with resultant electronic spin S=2 and affected by the immediate barium neighbors were obtained. It is found that these copper aggregates giving the EPR signals are magnetically isolated from the main CuO cluster.

Results of EPR and optical studies of vanadyl ion (VO2+) doped in single crystals of sodium formate are reported. Impurity sites in the unit cell and the molecular structure of the vanadyl complexes are discussed. The spin Hamiltonian parameters are determined. From optical and EPR data, crystal field parameters are calculated and discussed. Two complexes at substitutional sites at two different levels of doping have been identified.

We report the observation of an EPR spectrum attributable to Cu2+Cu2+ pairs with resultant electronic spin S = 1, from cupric oxide annealed in the temperature range from 700 to 900°C. The spectrum consists of only a single line at g = 3.12 with seven hyperfine components. From the weight loss and oxygen content measurements, it was found that the observation of such a signal strongly depends on the oxygen content of the sample.

A systematic Electron Spin Resonance investigation is carried out in CuO bulk powder and thin films, after calcining them to different temperatures ranging from 100°C to 1000°C. In CuO powder, two separate signals (one broad and one sharp) were observed at calcination temperature of 500°C, for the fist time. They got intensified, came closer and amalgamated on further heating upto 800°C, indicating complete destruction of antiferromagnetic ordering in this powdered compound. In thin films of CuO, the first appearance of the signal at 200°C and its intensification and shifting on further heating indicates that in thin film form, the destruction of antiferromagnetism takes place much earlier.