Metallic silver nanoparticle: a therapeutic agent in combination with antifungal drug against human fungal pathogen

Silver nanoparticles (Ag-NPs) are known to have inhibitory and fungicidal effects. Resistance against fungal infection has emerged as a major health problem in recent years, which needs great and immediate concern. Here, we report the extracellular biological synthesis of silver nanoparticles through a simple green route approach using a marine mangrove (Rhizophora mucronata) and silver nitrate. Aqueous extract of marine mangrove helped in reduction and was used as capping agent in biological synthesis. Nanoparticles were characterized using microscopy and spectroscopy techniques such as HRTEM, UV–Vis absorption spectroscopy and FTIR spectroscopy. X-ray diffraction analysis showed that the nanoparticles had face centered cubic structure with crystalline nature. FTIR spectroscopy showed the presence of different functional groups, such as hydroxyl and carbonyl, involved in the synthesis of nanoparticles. The antifungal activity of fluconazole and itraconazole was enhanced against the tested pathogenic fungi in the presence of Ag-NP and confirmed from increase in fold area of inhibition. This environmentally friendly method of biological synthesis can be easily integrated for various medical applications.     

Herpetic stromal keratitis in the absence of viral antigen recognition

Herpetic stromal keratitis (HSK), resulting from ocular infection with herpes simplex virus (HSV), is thought to represent a T cell mediated immunopathologic lesion. Antigens recognized by the inflammatory T cells remain unresolved and non-TCR mediated activation of T cells (bystander activation) is considered as also involved. This report documents further evidence for the bystander activation mechanisms using three T cell transgenic RAG-/- mouse strains. Accordingly HSK occurred in PCC RAG-/-, P14 RAG-/-, and OT-1 RAG-/- mice. In none of the models could HSV specific T cell reactivity be demonstrated and animals were unprotected from lesion development by immunization prior to HSV ocular infection. The results support the role of bystander activation as a mechanism of T cell mediated immunopathology and show that CD8(+) as well as CD4(+) T cells can participate in HSK lesion development.     

Necrostatin-1 protects against glutamate-induced glutathione depletion and caspase-independent cell death in HT-22 cells

Glutamate, a major excitatory neurotransmitter in the CNS, plays a critical role in neurological disorders such as stroke and Parkinson's disease. Recent studies have suggested that glutamate excess can result in a form of cell death called glutamate-induced oxytosis. In this study, we explore the protective effects of necrostatin-1 (Nec-1), an inhibitor of necroptosis, on glutamate-induced oxytosis. We show that Nec-1 inhibits glutamate-induced oxytosis in HT-22 cells through a mechanism that involves an increase in cellular glutathione (GSH) levels as well as a reduction in reactive oxygen species production. However, Nec-1 had no protective effect on free radical-induced cell death caused by hydrogen peroxide or menadione, which suggests that Nec-1 has no antioxidant effects. Interestingly, the protective effect of Nec-1 was still observed when cellular GSH was depleted by buthionine sulfoximine, a specific and irreversible inhibitor of glutamylcysteine synthetase. Our study further demonstrates that Nec-1 significantly blocks the nuclear translocation of apoptosis-inducing factor (a marker of caspase-independent programmed cell death ) and inhibits the integration of Bcl-2/adenovirus E1B 19 kDa-interacting protein 3 (a pro-death member of the Bcl-2 family) into the mitochondrial membrane. Taken together, these results demonstrate for the first time that Nec-1 prevents glutamate-induced oxytosis in HT-22 cells through GSH related as well as apoptosis-inducing factor and Bcl-2/adenovirus E1B 19 kDa-interacting protein 3-related pathways.     

Immobilized pH gradient-driven paper-based IEF: a new method for fractionating complex peptide mixtures before MS analysis

Introduction  

The vast difference in the abundance of different proteins in biological samples limits the determination of the complete proteome of a cell type, requiring fractionation of proteins and peptides before MS analysis.     

Direct and circulatory paths of Permethrin (NRDC-143) causing histopathological changes in the gills of rainbow trout, Salmo gairdneri Richardson

Rainbow trout were exposed to sublethal concentrations of Permethrin in water (0.09, 0.18 and 0.35 μg l⁻¹) and in food (85, 180 and 350 μg kg⁻¹ dry diet) in 20–0 day experiments. Histological changes in the gills included epithelial separation or necrosis, mucus cell hyperplasia, clubbing of epithelial cells or hyperplasia and fusion of adjacent secondary lamellae. These changes were noted in all fish, suggesting that the pesticide or its metabolites reached the gills not only directly through the water, but also indirectly via the circulation.     

Bystander activation involving T lymphocytes in herpetic stromal keratitis

Herpes simplex virus infection of mouse corneas can lead to the development of an immunopathological lesion, termed herpetic stromal keratitis (HSK). Such lesions also occur in TCR-transgenic mice backcrossed to SCID (TgSCID) that are unable to mount detectable HSV-specific immune responses. The present study demonstrates that lesion expression in such mice depends on continuous viral replication, whereas in immunocompetent mice, lesions occurred even if virus replication was terminated at 4 days after infection. The continuous replication in TgSCID mice was considered necessary to produce an activating stimulus to CD4(+) T cells that invade the cornea. Lesions in TgSCID were resistant to control by cyclosporin A, but were inhibited by treatment with rapamycin. This result was interpreted to indicate that T cell activation involved a non-TCR-mediated cytokine-driven bystander mechanism. Bystander activation was also shown to play a role in HSK lesions in immunocompetent mice. Accordingly, in immunocompetent DO11.10 mice, lesions were dominated by KJ1.26(+) OVA-specific CD4(+) T cells that were unreactive with HSV. In addition, KJ1.26(+) HSV nonimmune cells parked in ocularly infected BALB/c mice were demonstrable in HSK lesions. These results provide insight for the choice of new strategies to manage HSK, an important cause of human blindness.     

Cancerous cell targeting and destruction using pH stabilized amperometric bioconjugated gold nanoparticles from marine macroalgae,Padina gymnospora

The main aim of this study was, using biomechanistic approach, to synthesize and characterize amperometric stable gold nanoparticles (AuNPs) under different pH conditions using UV Spec, dynamic light scattering and TEM with energy dispersive X-ray analysis. The biomolecules involved in conjugation and reduction were further characterized by Fourier transform infrared analysis. The pH stabilized nanoparticles were studied to determine the functional and molecular mechanism of cell death on liver cancer (HepG2) cell line and gastric cancer (YCC3) cell line. The zeta potential and TEM imaging demonstrated that AuNPs were spherical in nature and can pass through the cellular membrane because of their intrinsic properties of AuNPs to bind to carbon-bonded sulfhydryl (–C–SH or R–SH) group and, therefore, could interact with intracellular components of the cell which was confirmed through phase contrast microscopy. Altered molecular mechanism and cellular effects in different cancer cell suggest a potential for in vivo applications of gold nanomaterials.     

PD-1 modulates regulatory T cells and suppresses T-cell responses in HCV-associated lymphoma

T regulatory (T(R)) cells suppress T-cell responses that are critical in the development of chronic viral infection and associated malignancies. Programmed death-1 (PD-1) also has a pivotal role in regulation of T-cell functions during chronic viral infection. To examine the role of PD-1 pathway in regulating T(R)-cell functions that inhibit T-cell responses during virus-associated malignancy, T(R) cells were investigated in the setting of hepatitis C virus-associated lymphoma (HCV-L), non-HCV-associated lymphoma (non-HCV-L), HCV infection alone and healthy subjects (HS). Relatively high numbers of CD4(+)CD25(+) and CD8(+)CD25(+) T(R) cells, as well as high levels of PD-1 expressions on these T(R) cells were found in the peripheral blood of subjects with HCV-L compared with those from non-HCV-L or HCV alone or HS. T(R) cells from the HCV-L subjects were capable of suppressing the autogeneic lymphocyte response, and depletion of T(R) cells in peripheral blood mononuclear cells from HCV-L improved T-cell proliferation. Additionally, the suppressed T-cell activation and proliferation in HCV-L was partially restored by blocking the PD-1 pathway ex vivo, resulting in both a reduction in T(R)-cell number and the ability of T(R) to suppress the activity of effector T cells. This study suggests that the PD-1 pathway is involved in regulating T(R) cells that suppress T-cell functions in the setting of HCV-associated B-cell lymphoma.     

Natural killer cells as novel helpers in anti-herpes simplex virus immune response

Innate defenses help to eliminate infection, but some of them also play a major role in shaping the magnitude and efficacy of the adaptive immune response. With regard to influencing subsequent adaptive immunity, NK cells aided by dendritic cells may be the most relevant components of the innate reaction to herpes simplex virus (HSV) infection. We confirm that mice lacking or depleted of NK cells are susceptible to HSV-induced lesions. The quantity and quality of CD8(+) cytotoxic T lymphocytes generated in the absence of NK cells were diminished, thereby contributing to susceptibility to HSV-induced encephalitis. We demonstrate a novel helper role for NK cells, in that NK cells compensate for the loss of CD4 helper T cells and NK cell supplementation enhances the function of wild type anti-HSV CD8 T cells. In addition, NK cells were able to partially rescue the dysfunctional CD8(+) T cells generated in the absence of CD4 T helper cells, thereby performing a novel rescue function. Hence, NK cells may well be exploited for enhancing and rescuing the T-cell response in situations where the CD4 helper response is affected.