Dual template (epitope) imprinted electrode for sensing bacterial protein with high selectivity.

Epitope imprinting has shown better prospects to synthesize synthetic receptors for proteins. Here, dual epitope imprinted polymer electrode (DEIP) matrix was fabricated on gold surface of electrochemical quartz crystal microbalance (EQCM) for recognition of target epitope sequence in blood samples of patients suffering from brain fever. Epitope sequences from outer membrane protein Por B of Neisseria meningitidis (MC58) bacteria predicted through immunoinformatic tools were chosen for imprinting. Self-assembled monolayers (SAM) of cysteine appended epitope sequences on gold nanoparticles were subjected to polymerization prior to electrodeposition on gold coated EQCM electrode. The polymeric matrix was woven around the cysteine appended epitope SAMs through multiple monomers (3-sulfo propyl methacrylate potassium salt (3-SPMAP), benzyl methacrylate (BMA)) and crosslinker (N, N'-methylene-bis-acrylamide). On extraction of the peptide sequences, imprinted cavities were able to selectively and specifically bind targeted epitope sequences in laboratory samples as well as 'real' samples of patients. Selectivity of sensor was examined through mismatched peptide sequences and certain plasma proteins also. The sensor was able to show specific binding towards the blood samples of infected patients, even in the presence of 'matrix' and other plasma proteins such as albumin and globulin. Even other peptide sequences, similar to epitope sequences only with one or two amino acid mismatches were also unable to show any binding. The analytical performance of DEIP-EQCM sensor was tested through selectivity, specificity, matrix effect, detection limit (0.68-1.01 nM), quantification limit (2.05-3.05 nM) and reproducibility (RSD ~ 5%). Hence, a diagnostic tool for bacterium causing meningitis is successfully fabricated in a facile manner which will broaden the clinical access and make efficient population screening feasible.

One-pot Synthesis of 2-Aminoindole through SET Oxidative Cyclization: Concise Synthesis of Tryptanthrin and Phaitanthrin E.

An efficient domino approach for the synthesis of biologically important 2-aminoindole derivatives has been developed using CuBr2 -mediated SET oxidative cyclization as a key step. This one-pot multicomponent strategy utilizes readily available ethyl propiolate, tosyl azide, and substituted aryl amines as starting materials. The generality and scope of this mild method are demonstrated with a wide variety of substrates to furnish functionalized 2-aminoindoles in good yields. The synthetic power of this strategy is further exemplified in the concise synthesis of biologically important alkaloids, Phaitanthrin E and Tryptanthrin.

Role of bone marrow-derived macrophages (BMDMs) in neurovascular interactions during stroke.

Stroke is a leading cause of disability worldwide and hence remains a major medical concern. Besides several pathological features, such as excitotoxicity, peri-infarct depolarization, acidosis, reactive oxygen species generation, apoptosis, and necrosis, dysregulation of the immune system severely affects stroke outcomes. After stroke onset, microglia - the brain-resident macrophage immune cells - and peripheral immune cells affect stoke injury/recovery by releasing pro-inflammatory and/or anti-inflammatory cytokines depending on their microenvironment. These pro- or anti-inflammatory cytokines further affect integrity of the blood brain barrier (BBB) and modulate immune infiltration after stroke. Among peripheral immune cells, bone marrow-derived macrophages (BMDMs) play a critical role in stroke pathology which peaks between three and seven days post-stroke. BMDMs have been extensively studied for their role in exacerbation of stroke injury, however they have rarely been studied for their role in tissue repair. Nonetheless, these reparative roles are gaining attention since recent studies have shown either failure or worsening of long-term post-stroke recovery after blockade of peripheral immune infiltration. These diverse but paradoxical effects of infiltrating monocytes/macrophages encouraged us to summarize the latest findings in neuro-immune and immune-vascular interactions. This review highlights the multifaceted role of BMDMs in stroke onset and resolution, and emphasizes the significance of tapping the potential of these cells to gain better insight into disease progression and therapy.

Identification and functional characterization of a bacterial homologue of Zeta toxin in Leishmania donovani.

Zeta-toxin is a cognate toxin of epsilon antitoxin of prokaryotic Type II toxin-antitoxin system (TA) and play an important role in cell death. An orthologue of bacterial-zeta-toxin (BzT) was identified in Leishmania donovani with similar structural and functional features. Leishmania zeta-toxin (named Ld_ζ1) harboring similar UNAG and ATP-binding pockets showed UNAG kinase and ATP-binding activity. An active Ld_ζ1 was found to express in infective extracellular promastigotes stage of L. donovani and episomal overexpression of an active Ld_ζ1domain-triggered cell death. This study demonstrates the presence of prokaryotic-like-zeta-toxin in eukaryotic parasite Leishmania and its association with cell death. Conceivably, phosphorylated UNAG or analogues, the biochemical mimics of zeta-toxin function mediating cell death can act as a novel anti-leishmanial chemotherapeutics.

Integrative natural medicine inspired graphene nanovehicle-benzoxazine derivatives as potent therapy for cancer.

Natural products from medicinal plants have always attracted a lot of attention due to their diverse and interesting therapeutic properties. We have employed the principles of green chemistry involving isomerization, coupling and condensation reaction to synthesize a class of compounds derived from eugenol, a naturally occurring bioactive phytophenol. The compounds were characterized structurally by 1H-, 13C-NMR, FT-IR spectroscopy and mass spectrometry analysis. The purity of compounds was detected by HPLC. The synthesized compounds exhibited anti-cancer activity. A 10-12-fold enhancement in efficiency of drug molecules (~ 1 µM) was observed when delivered with graphene oxide (GO) as a nanovehicle. Our data suggest cell death via apoptosis in a dose-dependent manner due to increase in calcium levels in specific cancer cell lines. Interestingly, the benzoxazine derivatives of eugenol with GO nanoparticle exhibited enhanced therapeutic potential in cancer cells. In addition to anti-cancer effect, we also observed significant role of these derivatives on parasite suggesting its multi-pharmacological capability.

PI3K/Akt/GSK3ß induced CREB activation ameliorates arsenic mediated alterations in NMDA receptors and associated signaling in rat hippocampus: Neuroprotective role of curcumin.

Protective efficacy of curcumin in arsenic induced NMDA receptor dysfunctions and PI3K/Akt/ GSK3ß signalling in hippocampus has been investigated in vivo and in vitro. Exposure to sodium arsenite (in vivo - 20 mg/kg, body weight p.o. for 28 days; in vitro - 10 µM for 24 h) and curcumin (in vivo - 100 mg/kg body weight p.o. for 28 days; in vitro - 20 µM for 24 h) was carried out alone or simultaneously. Treatment with curcumin ameliorated sodium arsenite induced alterations in the levels of NMDA receptors, its receptor subunits and synaptic proteins - pCaMKIIα, PSD-95 and SynGAP both in vivo and in vitro. Decreased levels of BDNF, pAkt, pERK1/2, pGSK3ß and pCREB on sodium arsenite exposure were also protected by curcumin. Curcumin was found to decrease sodium arsenite induced changes in hippocampus by modulating PI3K/Akt/GSK3ß neuronal survival pathway, known to regulate various cellular events. Treatment of hippocampal cultures with pharmacological inhibitors for ERK1/2, GSK3ß and Akt individually inhibited levels of CREB and proteins associated with PI3K/Akt/GSK3ß pathway. Simultaneous treatment with curcumin was found to improve sodium arsenite induced learning and memory deficits in rats assessed by water maze and Y-maze. The results provide evidence that curcumin exercises its neuroprotective effect involving PI3K/Akt pathway which may affect NMDA receptors and downstream signalling through TrKß and BDNF in arsenic induced cognitive deficits in hippocampus.

Neurotrophic factor mediated neuronal differentiation of human cord blood mesenchymal stem cells and their applicability to assess the developmental neurotoxicity.

Plasticity and developmental capacity of stem cells have now been established as a promising tool to restore the degenerative disorders. The linearity differentiation of human mesenchymal stem cells (hMSCs) into adipogenic, chondrogenic, osteogenic and even in neuronal subtypes has been demonstrated. The number of xenobiotics such as dexamethasone, insulin, isobutyl 1-methyle xanthine and retinoic acid has been reported for the potential to differentiate hMSCs into neuronal subtypes. But, the applicability of indigenous neurotrophic factor-nerve growth factor (NGF) has not been explored for the purpose. Thus, the present investigations were carried out to study the NGF induced neuronal differentiation of hMSCs. Following the isolation, purification and characterization of hMSCs were allowed to differentiate into neuronal subtypes under the influence of NGF (50 ng/mL). At various concentrations of NGF, the neuronal makers were analysed at both mRNA and protein levels. Cells, exposed with NGF were showing the significant and gradual increase in the neuronal markers in differentiating cells. The magnitude of expression of markers was maximum at day 4 of differentiation. NGF at 50 ng/mL concentration was found to induce neuronal differentiation of hMSCs into neuronal subtypes.

Advances in Stem Cell Research- A Ray of Hope in Better Diagnosis and Prognosis in Neurodegenerative Diseases.

Neurodegeneration and neurodegenerative disorders have been a global health issue affecting the aging population worldwide. Recent advances in stem cell biology have changed the current face of neurodegenerative disease modeling, diagnosis, and transplantation therapeutics. Stem cells also serve the purpose of a simple in-vitro tool for screening therapeutic drugs and chemicals. We present the application of stem cells and induced pluripotent stem cells (iPSCs) in the field of neurodegeneration and address the issues of diagnosis, modeling, and therapeutic transplantation strategies for the most prevalent neurodegenerative disorders. We have discussed the progress made in the last decade and have largely focused on the various applications of stem cells in the neurodegenerative research arena.

Differential responses of Trans-Resveratrol on proliferation of neural progenitor cells and aged rat hippocampal neurogenesis.

The plethora of literature has supported the potential benefits of Resveratrol (RV) as a life-extending as well as an anticancer compound. However, these two functional discrepancies resulted at different concentration ranges. Likewise, the role of Resveratrol on adult neurogenesis still remains controversial and less understood despite its well documented health benefits. To gather insight into the biological effects of RV on neurogenesis, we evaluated the possible effects of the compound on the proliferation and survival of neural progenitor cells (NPCs) in culture, and in the hippocampus of aged rats. Resveratrol exerted biphasic effects on NPCs; low concentrations (10 µM) stimulated cell proliferation mediated by increased phosphorylation of extracellular signal-regulated kinases (ERKs) and p38 kinases, whereas high concentrations (>20 µM) exhibited inhibitory effects. Administration of Resveratrol (20 mg/kg body weight) to adult rats significantly increased the number of newly generated cells in the hippocampus, with upregulation of p-CREB and SIRT1 proteins implicated in neuronal survival and lifespan extension respectively. We have successfully demonstrated that Resveratrol exhibits dose dependent discrepancies and at a lower concentration can have a positive impact on the proliferation, survival of NPCs and aged rat hippocampal neurogenesis implicating its potential as a candidate for restorative therapies against age related disorders.

Endoplasmic Reticulum Stress Induced Synthesis of a Novel Viral Factor Mediates Efficient Replication of Genotype-1 Hepatitis E Virus.

Hepatitis E virus (HEV) causes acute hepatitis in many parts of the world including Asia, Africa and Latin America. Though self-limiting in normal individuals, it results in ~30% mortality in infected pregnant women. It has also been reported to cause acute and chronic hepatitis in organ transplant patients. Of the seven viral genotypes, genotype-1 virus infects humans and is a major public health concern in South Asian countries. Sporadic cases of genotype-3 and 4 infection in human and animals such as pigs, deer, mongeese have been reported primarily from industrialized countries. Genotype-5, 6 and 7 viruses are known to infect animals such as wild boar and camel, respectively. Genotype-3 and 4 viruses have been successfully propagated in the laboratory in mammalian cell culture. However, genotype-1 virus replicates poorly in mammalian cell culture and no other efficient model exists to study its life cycle. Here, we report that endoplasmic reticulum (ER) stress promotes genotype-1 HEV replication by inducing cap-independent, internal initiation mediated translation of a novel viral protein (named ORF4). Importantly, ORF4 expression and stimulatory effect of ER stress inducers on viral replication is specific to genotype-1. ORF4 protein sequence is mostly conserved among genotype-1 HEV isolates and ORF4 specific antibodies were detected in genotype-1 HEV patient serum. ORF4 interacted with multiple viral and host proteins and assembled a protein complex consisting of viral helicase, RNA dependent RNA polymerase (RdRp), X, host eEF1α1 (eukaryotic elongation factor 1 isoform-1) and tubulinß. In association with eEF1α1, ORF4 stimulated viral RdRp activity. Furthermore, human hepatoma cells that stably express ORF4 or engineered proteasome resistant ORF4 mutant genome permitted enhanced viral replication. These findings reveal a positive role of ER stress in promoting genotype-1 HEV replication and pave the way towards development of an efficient model of the virus.

Molecular Mechanism of Switching of TrkA/p75(NTR) Signaling in Monocrotophos Induced Neurotoxicity.

We demonstrate the role of molecular switching of TrkA/p75(NTR) signaling cascade in organophosphate pesticide-Monocrotophos (MCP) induced neurotoxicity in stem cell derived cholinergic neurons and in rat brain. Our in-silico studies reveal that MCP followed the similar pattern of binding as staurosporine and AG-879 (known inhibitors of TrkA) with TrkA protein (PDB ID: 4AOJ) at the ATP binding sites. This binding of MCP to TrkA led to the conformational change in this protein and triggers the cell death cascades. The in-silico findings are validated by observing the down regulated levels of phosphorylated TrkA and its downstream molecules viz., pERK1/2, pAkt and pCREB in MCP-exposed cells. We observe that these MCP induced alterations in pTrkA and downstream signaling molecules are found to be associated with apoptosis and injury to neurons. The down-regulation of TrkA could be linked to increased p75(NTR). The in-vitro studies could be correlated in the rat model. The switching of TrkA/p75(NTR) signaling plays a central role in MCP-induced neural injury in rBNSCs and behavioral changes in exposed rats. Our studies significantly advance the understanding of the switching of TrkA/p75(NTR) that may pave the way for the application of TrkA inducer/p75(NTR) inhibitor for potential therapeutic intervention in various neurodegenerative disorders.