Integrative Biology最新文献

In B cells, the chemokine receptor CXCR3 is expressed only by a subset of B cells. However, CXCR3 is highly expressed in a rare type of B-cell lymphoma known as Mantle Cell Lymphoma (MCL) and CXCR3 inhibitor impairs proliferation and induces apoptosis in the MCL cell line JeKo-1. Despite this, the mechanism responsible for maintaining high levels of CXCR3 in MCL cells remains unclear. In this study, we assessed CXCR3 expression and amplification in MCL samples and confirmed that CXCR3 is overexpressed in MCL tissues. We also observed that CXCR3 amplification is present in a small portion of MCL patients and is associated with MCL classification. We then screened ubiquitin-specific proteases (USPs) that might control the degradation of CXCR3 protein. Our investigation revealed that USP35 acts as a potent stabilizer of CXCR3 protein. Knockdown of USP35 substantially reduced the CXCR3 protein levels in JeKo-1 cells, resulting in reduced cell viability, cell cycle arrest, increased apoptosis, and mitigated migration and invasion in these cells. At the molecular level, USP35 deubiquitinates and stabilizes CXCR3. USP35 deficiency attenuated the activation of the JAK1/STAT1 pathway and reduced the expression of β-catenin and c-Myc in JeKo-1 cells. Furthermore, we observed that overexpression of CXCR3 rescued the impaired tumorigenicity of USP35-deficient JeKo-1 cells, and the mechanism may be related to the fact that USP35 promotes CXCR3 deubiquitination to stabilize its expression. These findings collectively demonstrate the oncogenic role of the USP35-CXCR3 axis in JeKo-1 MCL cells.

Immune responses against cancer are inherently stochastic, with small numbers of individual T cells within a larger ensemble of lymphocytes initiating the molecular cascades that lead to tumor cytotoxicity. A potential source of this intra-tumor variability is the differential ability of immune cells to respond to tumor cells. Classical microwell co-cultures of T cells and tumor cells are inadequate for reliably culturing and analyzing low cell numbers needed to probe this variability, and have failed in recapitulating the heterogeneous small domains observed in tumors. Here we leverage a membrane displacement trap array technology that overcomes limitations of conventional microwell plates for immunodynamic studies. The microfluidic platform supports on-demand formation of dense nanowell cultures under continuous perfusion reflecting the tumor microenvironment, with real-time monitoring of T cell proliferation and activation within each nanowell. The system enables selective ejection of cells for profiling by fluorescence activated cell sorting, allowing observed on-chip variability in immune response to be correlated with off-chip quantification of T cell activation. The technology offers new potential for probing the molecular origins of T cell heterogeneity and identifying specific cell phenotypes responsible for initiating and propagating immune cascades within tumors. Insight Box Variability in T cell activation plays a critical role in the immune response against cancer. New tools are needed to unravel the mechanisms that drive successful anti-tumor immune response, and to support the development of novel immunotherapies utilizing rare T cell phenotypes that promote effective immune surveillance. To this end, we present a microfluidic cell culture platform capable of probing differential T cell activation in an array of nanoliter-scale wells coupled with off-chip cell analysis, enabling a high resolution view of variable immune response within tumor / T cell co-cultures containing cell ensembles orders of magnitude smaller than conventional well plate studies.

The standard model of the cell membrane potential Vm describes it as arising from diffusion currents across a membrane with a constant electric field, with zero electric field outside the cell membrane. However, the influence of Vm has been shown to extend into the extracellular space where it alters the cell's ζ-potential, the electrical potential measured a few nm from the cell surface which defines how the cell interacts with charged entities in its environment, including ions, molecules, and other cells. The paradigm arising from surface science is that the ζ-potential arises only from fixed membrane surface charge, and has consequently received little interest. However, if the ζ-potential can mechanistically and dynamically change by alteration of Vm, it allows the cell to dynamically alter cell-cell and cell-molecule interactions and may explain previously unexplained electrophysiological behaviours. Whilst the two potentials Vm and ζ are rarely reported together, they are occasionally described in different studies for the same cell type. By considering published data on these parameters across multiple cell types, as well as incidences of unexplained but seemingly functional Vm changes correlating with changes in cell behaviour, evidence is presented that this may play a functional role in the physiology of red blood cells, macrophages, platelets, sperm, ova, bacteria and cancer. Understanding how these properties will improve understanding of the role of electrical potentials and charges in the regulation of cell function and in the way in which cells interact with their environment. Insight  The zeta (ζ) potential is the electrical potential a few nm beyond the surface of any suspensoid in water. Whilst typically assumed to arise only from fixed charges on the cell surface, recent and historical evidence shows a strong link to the cell's membrane potential Vm, which the cell can alter mechanistically through the use of ion channels. Whilst these two potentials have rarely been studied simultaneously, this review collates data across multiple studies reporting Vm, ζ-potential, electrical properties of changes in cell behaviour. Collectively, this points to Vm-mediated ζ-potential playing a significant role in the physiology and activity of blood cells, immune response, developmental biology and egg fertilization, and cancer among others.

With the expanding ageing population, there is a growing interest in the maintenance of immune health to support healthy ageing. Enthusiasm exists for unravelling the impact of diet on the immune system and its therapeutic potential. However, a key challenge is the lack of studies investigating the effect of dietary patterns and nutrients on immune responses. Thus, we have used an integrative analysis approach to improve our understanding of diet-immune system interactions in older adults. To do so, dietary data were collected in parallel with performing immunophenotyping and functional assays from healthy older (n = 40) participants. Food Frequency Questionnaire (FFQ) was utilised to derive food group intake and multi-colour flow cytometry was performed for immune phenotypic and functional analysis. Spearman correlation revealed the strength of association between all combinations of dietary components, micronutrients, and hallmarks of immunesenescence. In this study, we propose for the first time that higher adherence to the Mediterranean diet is associated with a positive immune-ageing trajectory (Lower IMM-AGE score) in older adults due to the immune protective effects of high dietary fibre and PUFA intake in combating accumulation or pro-inflammatory senescent T cells. Furthermore, a diet rich in Vit A, Vit B6 and Vit B12 is associated with fewer features of immunesenescence [such as accumulation of terminally differentiated memory CD8 T cells] in older adults. Based on our findings we propose a future nutrition-based intervention study evaluating the efficacy of adherence to the MED diet alongside a multi-nutrient supplementation on immune ageing in older adults to set reliable dietary recommendations with policymakers that can be given to geriatricians and older adults. Insight box: There is a growing interest in the maintenance of immune health to boost healthy ageing. However, a key challenge is the lack of studies investigating the effect of dietary patterns and nutrients on immune responses. Thus, to do so we collected dietary data in parallel with performing immunophenotyping and functional assays on healthy older (n = 40) participants, followed by an integrative analysis approach to improve our understanding of diet-immune system interactions in older adults. We strongly believe that these new findings are appropriate for IB and will be of considerable interest to its broad audience.

Plant protease inhibitors (PI's) inhibit the activity of gut proteases and thus provide resistance against insect attack. Previously we have published first report on cloning and characterization of a novel Bowman-Birk protease inhibitor gene (RbTI) from ricebean (Vigna umbellata). In this study, the RbTI gene was further characterized and validated as a potential candidate for transferring insect resistance in economically important crops. We have successfully generated transgenic tobacco plants expressing RbTI gene constitutively under CaMV35S promoter using Agrobacterium transformation. Genomic PCR and GUS analysis confirmed the successful integration of RbTI gene into tobacco plant genome. qRT-PCR analysis revealed highest RbTI gene expression in transformed tobacco leaves nearing maturity. Feeding of transformed tobacco leaf tissue showed prominent effect on larval mortality throughout the larval growth stages mainly during first three days of feeding. For functional analysis of RbTI gene, we estimated the inhibitory activity of protein extracts from normal and transformed tobacco plants against gut proteases of Spodoptera litura and H. armigera larval instars. Maximum inhibition of trypsin (82.42% and 73.25%) and chymotrypsin (69.50% and 60.64%) enzymes was recorded at early larval stages of both insects. The results of this study validated the future use of RbTI gene from ricebean legume as a potential candidate for transferring insect resistance in economically important crops. Insight, innovation, integration: Present study was conducted with the aim to utilize the state of art biotechnological techniques for transferring key pest resistant genes from underutilized promising crop ricebean. The tobacco plant has been utilized as modern plant for proof of concept where a protease inhibitor gene from Ricebean has been transferred to tobacco plant which induced larval mortality within first three days of feeding at all larval developmental stages. The biochemical assays on mid-gut total protein extract showed that the transgenic tobacco leaves have inhibiting effect on trypsin and chymotrypsin enzymes of insect which is otherwise required for digestion of food by them. Hence, we provide a novel gene that could be utilized for pest resistance in other crops different developmental stages.

Shiga toxin (Stx) causes significant renal microvascular injury and kidney failure in the pediatric population, and an effective targeted therapy has yet to be demonstrated. Here we established a human kidney microvascular endothelial cell line for the study of Stx mediated injuries with respect to their morphologic, phenotypic, and transcriptional changes, and modeled Stx induced thrombotic microangiopathy (TMA) in flow-mediated 3D microvessels. Distinct from other endothelial cell lines, both isolated primary and immortalized human kidney microvascular endothelial cells demonstrate robust cell-surface expression of the Stx receptor Gb3, and concomitant dose-dependent toxicity to Stx, with significant contributions from caspase-dependent cell death. Use of a glucosylceramide synthase inhibitor (GCSi) to target disruption of the synthetic pathway of Gb3 resulted in remarkable protection of kidney microvascular cells from Stx injury, shown in both cellular morphologies, caspase activation and transcriptional analysis from RNA sequencing. Importantly, these findings are recapitulated in 3D engineered kidney microvessels under flow. Moreover, whole blood perfusion through Stx-treated microvessels led to marked platelet binding on the vessel wall, which was significantly reduced with the treatment of GCSi. These results validate the feasibility and utility of a bioengineered ex vivo human microvascular model under flow to recapitulate relevant blood-endothelial interactions in STEC-HUS. The profound protection afforded by GCSi demonstrates a preclinical opportunity for investigation in human tissue approximating physiologic conditions. Moreover, this work provides a broad foundation for novel investigation into TMA injury pathogenesis and treatment. Insight Box: Shiga toxin (Stx) causes endothelial injury that results in significant morbidity and mortality in the pediatric population, with no effective targeted therapy. This paper utilizes human kidney microvascular cells to examine Stx mediated cell death in both 2D culture and flow-mediated 3D microvessels, with injured microvessels also developing marked platelet binding and thrombi formation when perfused with blood, consistent with the clinical picture of HUS. This injury is abrogated with a small molecule inhibitor targeting the synthetic pathway of the Shiga toxin receptor. Our findings shed light onto Stx-induced vascular injuries and pave a way for broad investigation into thrombotic microangiopathies.