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1.
PI Kinase-EhGEF2-EhRho5 axis contributes to LPA stimulated macropinocytosis in Entamoeba histolytica
Entamoeba histolytica is a protozoan responsible for several pathologies in humans. Trophozoites breach the intestinal site to enter the bloodstream and thus traverse to a secondary site. Macropinocytosis and phagocytosis, collectively accounting for heterophagy, are the two major processes responsible for sustenance of Entamoeba histolytica within the host. Both of these processes require significant rearrangements in the structure to entrap the target. Rho GTPases play an indispensable role in mustering proteins that regulate cytoskeletal remodelling. Unlike phagocytosis which has been studied in extensive detail, information on machinery of macropinocytosis in E. histolytica is still limited. In the current study, using site directed mutagenesis and RNAi based silencing, coupled with functional studies, we have demonstrated the involvement of EhRho5 in constitutive and LPA stimulated macropinocytosis. We also report that LPA, a bioactive phospholipid present in the bloodstream of the host, activates EhRho5 and translocates it from cytosol to plasma membrane and endomembrane compartments. Using biochemical and FRAP studies, we established that a PI Kinase acts upstream of EhRho5 in LPA mediated signalling. We further identified EhGEF2 as a guanine nucleotide exchange factor of EhRho5. In the amoebic trophozoites, EhGEF2 depletion leads to reduced macropinocytic efficiency of trophozoites, thus phenocopying its substrate. Upon LPA stimulation, EhGEF2 is found to sequester near the plasma membrane in a wortmannin sensitive fashion, explaining a possible mode for activation of EhRho5 in the amoebic trophozoites. Collectively, we propose that LPA stimulated macropinocytosis in E. histolytica is driven by the PI Kinase-EhGEF2-EhRho5 axis. 相似文献
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Chandima Jeewandara Thiruni N. Adikari Laksiri Gomes Samitha Fernando R. H. Fernando M. K. T. Perera Dinuka Ariyaratne Achala Kamaladasa Maryam Salimi Shamini Prathapan Graham S. Ogg Gathsaurie Neelika Malavige 《PLoS neglected tropical diseases》2015,9(4)
Background
Although antibody responses to dengue virus (DENV) in naturally infected individuals have been extensively studied, the functionality of DENV specific memory T cell responses in relation to clinical disease severity is incompletely understood.Methodology/Principal findings
Using ex vivo IFNγ ELISpot assays, and by determining cytokines produced in ELISpot supernatants, we investigated the functionality of DENV-specific memory T cell responses in a large cohort of individuals from Sri Lanka (n=338), who were naturally infected and were either hospitalized due to dengue or had mild or sub clinical dengue infection. We found that T cells of individuals with both past mild or sub clinical dengue infection and who were hospitalized produced multiple cytokines when stimulated with DENV-NS3 peptides. However, while DENV-NS3 specific T cells of those with mild/sub clinical dengue infection were more likely to produce only granzyme B (p=0.02), those who were hospitalized were more likely to produce both TNFα and IFNγ (p=0.03) or TNFα alone.We have also investigated the usefulness of a novel T cell based assay, which can be used to determine the past infecting DENV serotype. 92.4% of DENV seropositive individuals responded to at least one DENV serotype of this assay and none of the seronegatives responded. Individuals who were seronegative, but had received the Japanese encephalitis vaccine too made no responses, suggesting that the peptides used in this assay did not cross react with the Japanese encephalitis virus.Conclusions/significance
The types of cytokines produced by DENV-specific memory T cells appear to influence the outcome of clinical disease severity. The novel T cell based assay, is likely to be useful in determining the past infecting DENV serotype in immune-epidemiological studies and also in dengue vaccine trials. 相似文献3.
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Activation tagging in indica rice identifies ribosomal proteins as potential targets for manipulation of water‐use efficiency and abiotic stress tolerance in plants 下载免费PDF全文
Mazahar Moin Achala Bakshi Anusree Saha M. Udaya Kumar Attipalli R. Reddy K. V. Rao E. A. Siddiq P. B. Kirti 《Plant, cell & environment》2016,39(11):2440-2459
We have generated 3900 enhancer‐based activation‐tagged plants, in addition to 1030 stable Dissociator‐enhancer plants in a widely cultivated indica rice variety, BPT‐5204. Of them, 3000 were screened for water‐use efficiency (WUE) by analysing photosynthetic quantum efficiency and yield‐related attributes under water‐limiting conditions that identified 200 activation‐tagged mutants, which were analysed for flanking sequences at the site of enhancer integration in the genome. We have further selected five plants with low Δ13C, high quantum efficiency and increased plant yield compared with wild type for a detailed investigation. Expression studies of 18 genes in these mutants revealed that in four plants one of the three to four tagged genes became activated, while two genes were concurrently up‐regulated in the fifth plant. Two genes coding for proteins involved in 60S ribosomal assembly, RPL6 and RPL23A, were among those that became activated by enhancers. Quantitative expression analysis of these two genes also corroborated the results on activating–tagging. The high up‐regulation of RPL6 and RPL23A in various stress treatments and the presence of significant cis‐regulatory elements in their promoter regions along with the high up‐regulation of several of RPL genes in various stress treatments indicate that they are potential targets for manipulating WUE/abiotic stress tolerance. 相似文献
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Gathsaurie Neelika Malavige Louise Jones S. D. Kamaladasa A. Wijewickrama S. L. Seneviratne Antony P. Black Graham S. Ogg 《PloS one》2008,3(11)
Background
In Sri Lanka, varicella zoster virus (VZV) is typically acquired during adulthood with significant associated disease morbidity and mortality. T cells are believed to be important in the control of VZV replication and in the prevention of reactivation. The relationship between viral load, disease severity and cellular immune responses in primary VZV infection has not been well studied.Methodology
We used IFNγ ELISpot assays and MHC class II tetramers based on VZV gE and IE63 epitopes, together with quantitative real time PCR assays to compare the frequency and phenotype of specific T cells with virological and clinical outcomes in 34 adult Sri Lankan individuals with primary VZV infection.Principal Findings
Viral loads were found to be significantly higher in patients with moderate to severe infection compared to those with mild infection (p<0.001) and were significantly higher in those over 25 years of age (P<0.01). A significant inverse correlation was seen between the viral loads and the ex vivo IFNγ ELISpot responses of patients (P<0.001, r = −0.85). VZV-specific CD4+ T cells expressed markers of intermediate differentiation and activation.Conclusions
Overall, these data show that increased clinical severity in Sri Lankan adults with primary VZV infection associates with higher viral load and reduced viral specific T cell responses. 相似文献7.
Biofunctionalization of biomaterials for accelerated in situ endothelialization: a review 总被引:1,自引:0,他引:1
The higher patency rates of cardiovascular implants, including vascular bypass grafts, stents, and heart valves are related to their ability to inhibit thrombosis, intimal hyperplasia, and calcification. In native tissue, the endothelium plays a major role in inhibiting these processes. Various bioengineering research strategies thereby aspire to induce endothelialization of graft surfaces either prior to implantation or by accelerating in situ graft endothelialization. This article reviews potential bioresponsive molecular components that can be incorporated into (and/or released from) biomaterial surfaces to obtain accelerated in situ endothelialization of vascular grafts. These molecules could promote in situ endothelialization by the mobilization of endothelial progenitor cells (EPC) from the bone marrow, encouraging cell-specific adhesion (endothelial cells (EC) and/or EPC) to the graft and, once attached, by controlling the proliferation and differentiation of these cells. EC and EPC interactions with the extracellular matrix continue to be a principal source of inspiration for material biofunctionalization, and therefore, the latest developments in understanding these interactions will be discussed. 相似文献
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Baguneid M de Mel A Yildirimer L Fuller BJ Hamilton G Seifalian AM 《Biotechnology and applied biochemistry》2011,58(1):14-24
Conventionally used vascular grafts such as polyester (Dacron) or expanded polytetrafluoroethylene perform inadequately as small-diameter vascular bypass grafts (SDBGs). SDBGs, which can maintain long-term patency and those that could potentially evolve with the somatic growth, are highly desirable in vascular surgery and thus research into tissue-engineered blood vessels (TEBVs) is of keen interest. A TEBV was developed by seeding endothelial cells onto a collagen matrix that was cross-linked and contracted by smooth muscle cells (SMCs). A polyester graft served as a scaffold. Recovery studies (12 TEBVs and seven controls) were carried out to assess in vivo endothelialization and long-term patency of TEBVs. Hemodynamic observations indicated para-anastomotic turbulences and high shear stress at anastomosis. Recovery studies demonstrated confluent endothelialization, thrombus-free surfaces, and patent TEBVs in all cases. Graft incorporation and neovascularization of the scaffold occurred in both hybrid and control grafts. However, thickened neointima formation occurred in TEBV grafts, which was most likely caused by the rigidity of polyester scaffold. Significant perigraft inflammatory changes could be observed in both TEBVs and control grafts at 1, 4, and 8 weeks. In conclusion, the TEBVs demonstrated satisfactory performance as an infra-renal-aortic graft in a porcine model. The TEBV serves as a promising model and facilitates the development of a TEBV in a clinical setting, potentially with human stem cells and with more biocompatible, biodegradable scaffolds that are mechanically more compliant with natural vessels. 相似文献
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Gathsaurie Neelika Malavige Chandima Jeewandara K. M. Luckmaal Alles Maryam Salimi Laksiri Gomes Achala Kamaladasa S. D. Jayaratne Graham Stuart Ogg 《PLoS neglected tropical diseases》2013,7(9)