Response Of Human Cells Research Articles

Monomeric CRP regulates inflammatory responses in human intervertebral disc cells.

CRP is an acute-phase protein that is used as a biomarker to follow severity and progression in infectious and inflammatory diseases. Its pathophysiological mechanisms of action are still poorly defined. CRP in its pentameric form exhibits weak anti-inflammatory activity. The monomeric isoform (mCRP) exerts potent proinflammatory properties in chondrocytes, endothelial cells, and leucocytes. No data exist regarding mCRP effects in human intervertebral disc (IVD) cells. This work aimed to verify the pathophysiological relevance of mCRP in the aetiology and/or progression of IVD degeneration. We investigated the effects of mCRP and the signalling pathways that are involved in cultured human primary annulus fibrosus (AF) cells and in the human nucleus pulposus (NP) immortalized cell line HNPSV-1. We determined messenger RNA (mRNA) and protein levels of relevant factors involved in inflammatory responses, by quantitative real-time polymerase chain reaction (RT-qPCR) and western blot. We also studied the presence of mCRP in human AF and NP tissues by immunohistochemistry. We demonstrated that mCRP increases nitric oxide synthase 2 (NOS2), cyclooxygenase 2 (COX2), matrix metalloproteinase 13 (MMP13), vascular cell adhesion molecule 1 (VCAM1), interleukin (IL)-6, IL-8, and Lipocalin 2 (LCN2) expression in human AF and NP cells. We also showed that nuclear factor-κβ (NF-κβ), extracellular signal-regulated kinase 1/2 (ERK1/2), and phosphoinositide 3-kinase (PI3K) are at play in the intracellular signalling of mCRP. Finally, we demonstrated the presence of mCRP in human AF and NP tissues. Our results indicate, for the first time, that mCRP can be localized in IVD tissues, where it triggers a proinflammatory and catabolic state in degenerative and healthy IVD cells, and that NF-κβ signalling may be implicated in the mediation of this mCRP-induced state.

Induction of adaptive response in E. coli cells alkylating agents depend on their chemical structure

The most well–known alkylating agents (AA) used in mutational breeding of agricultural crops and microorganisms producing biologically active substances are methyl methanesulfonate (MMS) and ethylmethanesulfone (EMS). It is known that both of these substances effectively induce bacterial alkyltransferase, which leads to an adaptive response of bacteria, that is, to the development of resistance to toxic effects. Bacterial alkyltransferase is a homologue of O6-methylguanine-DNA-methyltransferase in human cells. The study of the induction of alkyltransferase in bacteria by various AA allows us to predict their ability to induce an adaptive response in human cells. Lux biosensors pColDp-lux and pAlkA-lux were used to study the ability of MMS, ECG and BPL to induce DNA damage and adaptive response in E.coli cells. MMS effectively elicited an SOS response in the ColD-lux biosensor, characterizing the level of DNA damage, and an adaptive response recorded in the AlkA-lux biosensor by inducing the expression of the alkyltransferase gene. Bifunctional ECG induced both SOS response and adaptive response less actively than monofunctional MMS. The BPL induced an SOS response only at low concentrations, and did not cause an adaptive response at the same concentrations. Thus, bifunctional ECG and BPL, whose activity depends on the reactivity of the products of the disclosure of their cyclic structures, alkylate DNA to a lesser extent than MMC. This is due to the fact that they form electrophilic methyl groups to a lesser extent than MMCs when the cycle is opened. Keywords: lux-biosensor, E.coli, SOS response, adaptive response, methyl methanesulfonate, epichlorohydrin, beta-propiolactone. &nbsp

Multi-range ERK responses shape the proliferative trajectory of single cells following oncogene induction

Oncogene-induced senescence is a phenomenon in which aberrant oncogene expression causes non-transformed cells to enter a non-proliferative state. Cells undergoing oncogenic induction display phenotypic heterogeneity, with some cells senescing and others remaining proliferative. The causes of heterogeneity remain unclear. We studied the sources of heterogeneity in the responses of human epithelial cells to oncogenic BRAFV600E expression. We found that a narrow expression range of BRAFV600E generated a wide range of activities of its downstream effector ERK. In population-level and single-cell assays, ERK activity displayed a non-monotonic relationship to proliferation, with intermediate ERK activities leading to maximal proliferation. We profiled gene expression across a range of ERK activities over time and characterized four distinct ERK response classes, which we propose act in concert to generate the ERK-proliferation response. Altogether, our studies map the input-output relationships between ERK activity and proliferation, elucidating how heterogeneity can be generated during oncogene induction.

EF-24, a Curcumin Analog, Inhibits Cancer Cell Invasion in Human Nasopharyngeal Carcinoma through Transcriptional Suppression of Matrix Metalloproteinase-9 Gene Expression.

Cancer metastasis is a main cause of failure in treating subjects with nasopharyngeal carcinoma (NPC) and is frequently linked to high death rates. EF-24, an analog of curcumin, has exhibited many anti-cancer properties and enhanced bioavailability over curcumin. Nevertheless, the effects of EF-24 on the invasiveness of NPC are poorly understood. In this study, we demonstrated that EF-24 effectively inhibited TPA-induced motility and invasion responses of human NPC cells but elicited very limited cytotoxicity. In addition, the TPA-induced activity and expression of matrix metalloproteinase-9 (MMP-9), a crucial mediator of cancer dissemination, were found to be reduced in EF-24-treated cells. Our reporter assays revealed that such a reduction in MMP-9 expression by EF-24 was transcriptionally mediated by NF-κB via impeding its nuclear translocation. Further chromatin immunoprecipitation assays displayed that the EF-24 treatment decreased the TPA-induced interaction of NF-κB with the MMP-9 promoter in NPC cells. Moreover, EF-24 inhibited the activation of JNK in TPA-treated NPC cells, and the treatment of EF-24 together with a JNK inhibitor showed a synergistic effect on suppressing TPA-induced invasion responses and MMP-9 activities in NPC cells. Taken together, our data demonstrated that EF-24 restrained the invasiveness of NPC cells through the transcriptional suppression of MMP-9 gene expression, implicating the usefulness of curcumin or its analogs in controlling the spread of NPC.

Musculin does not modulate the disease course of Experimental Autoimmune Encephalomyelitis and DSS colitis

Previous evidences show that Musculin (Msc), a repressor member of basic helix-loop-helix transcription factors, is responsible in vitro for the low responsiveness of human Th17 cells to the growth factor IL-2, providing an explanation for Th17 cells rarity in inflammatory tissue. However, how and to what extent Musculin gene can regulate the immune response in vivo in an inflammatory context is still unknown. Here, exploiting two animal models of inflammatory diseases, the Experimental Autoimmune Encephalomyelitis (EAE) and the dextran sodium sulfate (DSS)-induced colitis, we evaluated the effect of Musculin gene knock-out on clinical course, performing also a deep immune phenotypical analysis on T cells compartment and an extended microbiota analysis in colitis-sick mice. We found that, at least during the early phase, Musculin gene has a very marginal role in modulating both the diseases. Indeed, the clinical course and the histological analysis showed no differences between wild type and Msc knock-out mice, whereas immune system appeared to give rise to a regulatory milieu in lymph nodes of EAE mice and in the spleen of DSS colitis-sick mice. Moreover, in the microbiota analysis, we found irrelevant differences between wild type and Musculin knock-out colitis-sick mice, with a similar bacterial strains’ frequency and diversity after the DSS treatment. This work strengthened the idea of a negligible Msc gene involvement in these models.

Transcriptomic Responses to Polymyxin B and Analogues in Human Kidney Tubular Cells.

Polymyxins are last-line antibiotics for the treatment of Gram-negative 'superbugs'. However, nephrotoxicity can occur in up to 60% of patients administered intravenous polymyxins. The mechanisms underpinning nephrotoxicity remain unclear. To understand polymyxin-induced nephrotoxicity, human renal proximal tubule cells were treated for 24 h with 0.1 mM polymyxin B or two new analogues, FADDI-251 or FADDI-287. Transcriptomic analysis was performed, and differentially expressed genes (DEGs) were identified using ANOVA (FDR < 0.2). Cell viability following treatment with polymyxin B, FADDI-251 or FADDI-287 was 66.0 ± 5.33%, 89.3 ± 3.96% and 90.4 ± 1.18%, respectively. Transcriptomics identified 430, 193 and 150 DEGs with polymyxin B, FADDI-251 and FADDI-287, respectively. Genes involved with metallothioneins and Toll-like receptor pathways were significantly perturbed by all polymyxins. Only polymyxin B induced perturbations in signal transduction, including FGFR2 and MAPK signaling. SIGNOR network analysis showed all treatments affected essential regulators in the immune system, autophagy, cell cycle, oxidative stress and apoptosis. All polymyxins caused significant perturbations of metal homeostasis and TLR signaling, while polymyxin B caused the most dramatic perturbations of the transcriptome. This study reveals the impact of polymyxin structure modifications on transcriptomic responses in human renal tubular cells and provides important information for designing safer new-generation polymyxins.

MiR-3646 accelerates inflammatory response of Ang II-induced hVSMCs via CYP2J2/EETs axis in hypertension model

ABSTRACT Background Inflammatory response of human vascular smooth muscle cells (hVSMCs) is a driving factor in hypertension progression. It has been reported that miR-3646 was significantly up-regulated in serum samples from patients with coronary artery disease and acute myocardial infarction mice. However, its role and underlying molecular mechanism related to inflammatory response of angiotensin II (Ang II)-induced hVSMCs remain unclear. Objective We aimed to explore the potential molecular mechanisms related to inflammatory response of angiotensin II (Ang II)-induced hVSMCs. Methods Ang II–induced hypertension model was established after hVSMCs treated with 1 μM Ang II at 24 h. The interaction between microRNA 3646 (miR-3646) and cytochrome P450 2J2 (CYP2J2) was assessed by dual-luciferase reporter gene assay. MTS assay, Lipid Peroxidation MDA Assay Kit, ELISA, Western blot, and qRT-PCR were performed to examine viability, malondialdehyde (MDA) level, inflammatory cytokine levels, and the level of genes and proteins. Results Our findings illustrated that miR-3646 was up-regulated but CYP2J2 was down-regulated in Ang II–induced hVSMCs. Mechanically, miR-3646 negatively targeted to CYP2J2 in Ang II–induced hVSMCs. These findings indicated that miR-3646 regulated inflammatory response of Ang II–induced hVSMCs via targeting CYP2J2. Moreover, functional researches showed that CYP2J2 overexpression alleviated inflammatory response of Ang II–induced hVSMCs via epoxyeicosatrienoic acids/peroxisome proliferator-activated receptor-γ (EETs/PPARγ) axis, and miR-3646 aggravated inflammatory response of Ang II–induced hVSMCs via mediating CYP2J2/EETs axis. Conclusion MiR-3646 accelerated inflammatory response of Ang II–induced hVSMCs via CYP2J2/EETs axis. Our findings illustrated the specific molecular mechanism of miR-3646 regulating hypertension.

The Effects of Mechanical Load on Chondrogenic Responses of Bone Marrow Mesenchymal Stem Cells and Chondrocytes Encapsulated in Chondroitin Sulfate-Based Hydrogel.

Articular cartilage is vulnerable to mechanical overload and has limited ability to restore lesions, which leads to the development of chronic diseases such as osteoarthritis (OA). In this study, the chondrogenic responses of human bone marrow mesenchymal stem cells (BMMSCs) and OA cartilage-derived chondrocytes in 3D chondroitin sulfate-tyramine/gelatin (CS-Tyr)/Gel) hydrogels with or without experimental mechanical load have been investigated. Chondrocytes were smaller in size, had slower proliferation rate and higher level of intracellular calcium (iCa2+) compared to BMMSCs. Under 3D chondrogenic conditions in CS-Tyr/Gel with or without TGF-β3, chondrocytes more intensively secreted cartilage oligomeric matrix protein (COMP) and expressed collagen type II (COL2A1) and aggrecan (ACAN) genes but were more susceptible to mechanical load compared to BMMSCs. ICa2+ was more stably controlled in CS-Tyr/Gel/BMMSCs than in CS-Tyr/Gel/chondrocytes ones, through the expression of L-type channel subunit CaV1.2 (CACNA1C) and Serca2 pump (ATP2A2) genes, and their balance was kept more stable. Due to the lower susceptibility to mechanical load, BMMSCs in CS-Tyr/Gel hydrogel may have an advantage over chondrocytes in application for cartilage regeneration purposes. The mechanical overload related cartilage damage in vivo and the vague regenerative processes of OA chondrocytes might be associated to the inefficient control of iCa2+ regulating channels.

Extra-viral DNA in adeno-associated viral vector preparations induces TLR9-dependent innate immune responses in human plasmacytoid dendritic cells

Adeno-associated viral (AAV) vector suspensions produced in either human derived HEK cells or in Spodoptera frugiperda (Sf9) insect cells differ in terms of residual host cell components as well as species-specific post-translational modifications displayed on the AAV capsid proteins. Here we analysed the impact of these differences on the immunogenic properties of the vector. We stimulated human plasmacytoid dendritic cells with various lots of HEK cell-produced and Sf9 cell-produced AAV-CMV-eGFP vectors derived from different manufacturers. We found that AAV8-CMV-eGFP as well as AAV2-CMV-eGFP vectors induced lot-specific but not production platform-specific or manufacturer-specific inflammatory cytokine responses. These could be reduced or abolished by blocking toll-like receptor 9 signalling or by enzymatically reducing DNA in the vector lots using DNase. Successful HEK cell transduction by DNase-treated AAV lots and DNA analyses demonstrated that DNase did not affect the integrity of the vector but degraded extra-viral DNA. We conclude that both HEK- and Sf9-cell derived AAV preparations can contain immunogenic extra-viral DNA components which can trigger lot-specific inflammatory immune responses. This suggests that improved strategies to remove extra-viral DNA impurities may be instrumental in reducing the immunogenic properties of AAV vector preparations.

Mechanobiology of the Human Intervertebral Disc: Systematic Review of the Literature and Future Perspectives.

Low back pain is an extremely common condition with severe consequences. Among its potential specific causes, degenerative disc disease (DDD) is one of the most frequently observed. Mechanobiology is an emerging science studying the interplay between mechanical stimuli and the biological behavior of cells and tissues. The aim of the presented study is to review, with a systematic approach, the existing literature regarding the mechanobiology of the human intervertebral disc (IVD), define the main pathways involved in DDD and identify novel potential therapeutic targets. The review was carried out in accordance with the Preferential Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines. Studies were included if they described biological responses of human IVD cells under mechanical stimulation or alterations of mechanical properties of the IVD determined by different gene expression. Fifteen studies were included and showed promising results confirming the mechanobiology of the human IVD as a key element in DDD. The technical advances of the last decade have allowed us to increase our understanding of this topic, enabling us to identify possible therapeutic targets to treat and to prevent DDD. Further research and technological innovations will shed light on the interactions between the mechanics and biology of the human IVD.

Reverse vaccinology assisted design of a novel multi-epitope vaccine to target Wuchereria bancrofti cystatin: An immunoinformatics approach.

Proteases are the critical mediators of immunomodulation exerted by the filarial parasites to bypass and divert host immunity. Cystatin is a small (∼15kDa) immunomodulatory filarial protein and known to contribute in the immunomodulation strategy by inducing anti-inflammatory response through alternative activation of macrophages. Recently, Wuchereria bancrofti cystatin has been discovered as a ligand of human toll-like receptor 4 which is key behind the cystatin-induced anti-inflammatory response in major human antigen-presenting cells. Considering the pivotal role of cystatin in the immunobiology of filariasis, cystatin could be an efficacious target for developing vaccine. Herein, we present the design and in-silico analyses of a multi-epitope-based peptide vaccine to target W. bancrofti cystatin through immune-informatics approaches. The 262 amino acid long antigen construct comprises 9 MHC-I epitopes and MHC-II epitopes linked together by GPGPG peptide alongside an adjuvant (50S ribosomal protein L7/L12) at N terminus and 6 His tags at C terminus. Molecular docking study reveals that the peptide could trigger TLR4-MD2 to induce protective innate immune responses while the induced adaptive responses were found to be mediated by IgG, IgM and Th1 mediated responses. Notably, the designed vaccine exhibits high stability and no allergenicity in-silico. Furthermore, the muti epitope-vaccine was also predicted for its RNA structure and cloned in pET30ax for further experimental validation. Taken together, this study presents a novel multi-epitope peptide vaccine for triggering efficient innate and adaptive immune responses against W. bancrofti to intervene LF through immunotherapy.

IL-8 Links NF-κB and Wnt/β-Catenin Pathways in Persistent Inflammatory Response Induced by Chronic Helicobacter pylori Infection

Helicobacter pylori (H. pylori) infection can cause persistent inflammatory response in human gastric mucosal epithelial cells, which may result in the occurrence of cancer. However, the underlying mechanism of carcinogenesis has not been elucidated yet. Herein, we established the models of chronic H. pylori infection in GES-1 cells and C57BL/6J mice. Interleukin 8 (IL-8) level was detected by ELISA. The expression of NF-κB p65, IL-8, Wnt2 and β-catenin mRNA and proteins was evaluated by real-time PCR, Western blotting, immunofluorescence staining, and immunohistochemistry. The infection of H. pylori in mice was evaluated by rapid urease test, H&E staining and Warthin-Starry silver staining. The morphological changes of gastric mucosa were observed by electron microscopy. Our results showed that in H. pylori infected gastric mucosal cells along with activation of NF-κB signaling pathway and increase of IL-8 level, the expression of Wnt2 was also increased significantly, which preliminarily indicates that IL-8 can positively regulate the expression of Wnt2. Studies in chronic H. pylori infected C57BL/6J mice models showed that there was an increased incidence of premalignant lesions in the gastric mucosa tissue. Through comparing changes of gastric mucosal cell ultrastructure and analyzing the relationship between NF-κB signaling pathway and Wnt2 expression, we found that H. pylori infection activated NF-κB signal pathways, and the massive release of IL-8 was positively correlated with the high expression of Wnt2 protein. Subsequently, the activated Wnt/β-catenin signal pathways may be involved in the malignant transformation of gastric mucosal cells. Collectively, H. pylori chronic infection may continuously lead to persistent inflammatory response: activate NF-κB pathway, promote IL-8 release and thereby activate Wnt/β-catenin pathway. IL-8 probably plays an important role of a linker in coupling these two signal pathways.

High extracellular sodium chloride concentrations induce resistance to LPS signal in human dendritic cells.

Recent evidence showed that in response to elevated sodium dietary intakes, many body tissues retain Na+ ions for long periods of time and can reach concentrations up to 200mM. This could modulate the immune system and be responsible for several diseases. However, studies brought contrasted results and the effects of external sodium on human dendritic cell (DC) responses to danger signals remain largely unknown. Considering their central role in triggering T cell response, we tested how NaCl-enriched medium influences human DCs properties. We found that DCs submitted to high extracellular Na+ concentrations up to 200mM remain viable and maintain the expression of specific DC markers, however, their maturation, chemotaxis toward CCL19, production of pro-inflammatory cytokines and ROS in response to LPS were also partially inhibited. In line with these results, the T-cell allostimulatory capacity of DCs was also inhibited. Finally, our data indicate that high NaCl concentrations triggered the phosphorylation of SGK1 and ERK1/2 kinases. These results raised the possibility that the previously reported pro-inflammatory effects of high NaCl concentrations on T cells might be counterbalanced by a downregulation of DC activation.

Magnitude and kinetics of the human immune cell response associated with severe dengue progression by single-cell proteomics

Magnitude and kinetics of the human immune cell response associated with severe dengue progression by single-cell proteomics

Aging alters the cell cycle control and mitogenic signaling responses of human hematopoietic stem cells

AbstractHuman hematopoietic stem cells (HSCs), like their counterparts in mice, comprise a functionally and molecularly heterogeneous population of cells throughout life that collectively maintain required outputs of mature blood cells under homeostatic conditions. In both species, an early developmental change in the HSC population involves a postnatal switch from a state in which most of these cells exist in a rapidly cycling state and maintain a high self-renewal potential to a state in which the majority of cells are in a quiescent state with an overall reduced self-renewal potential. However, despite the well-established growth factor dependence of HSC proliferation, whether and how this mechanism of HSC regulation might be affected by aging has remained poorly understood. To address this knowledge gap, we isolated highly HSC-enriched CD34+CD38−CD45RA−CD90+CD49f+ (CD49f+) cells from cord blood, adult bone marrow, and mobilized peripheral blood samples obtained from normal humans spanning 7 decades of age and then measured their functional and molecular responses to growth factor stimulation in vitro and their regenerative activity in vivo in mice that had undergone transplantation. Initial experiments revealed that advancing donor age was accompanied by a significant and progressively delayed proliferative response but not the altered mature cell outputs seen in normal older individuals. Importantly, subsequent dose-response analyses revealed an age-associated reduction in the growth factor–stimulated proliferation of CD49f+ cells mediated by reduced activation of AKT and altered cell cycle entry and progression. These findings identify a new intrinsic, pervasive, and progressive aging-related alteration in the biological and signaling mechanisms required to drive the proliferation of very primitive, normal human hematopoietic cells.

Baicalin suppresses interleukin-1β-induced apoptosis, inflammatory response, oxidative stress, and extracellular matrix degradation in human nucleus pulposus cells

Objective To explore the effect of baicalin on human nucleus pulposus cells (NPCs) in response to interleukin (IL)-1β stimulation. Methods Viability of NPCs was measured by cell counting kit-8 (CCK-8) assay. TUNEL staining assay and flow cytometry were performed to detect apoptotic cell death of NPCs. Western blot analysis was conducted to detect the expression levels of proteins. Enzyme-linked immunosorbent assay (ELISA) was applied for the determination of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor alpha (TNF-α), and IL-6. Oxidative stress indicators including reactive oxygen species (ROS) production, malondialdehyde (MDA) level, and superoxide dismutase (SOD) activity were measured. Results Baicalin attenuated IL-1β-caused cell viability reduction and apoptosis in NPCs. IL-1β-induced increase in Bax expression and decrease in Bcl-2 expression were attenuated by baicalin treatment. IL-1β-induced production of iNOS, COX-2, IL-6, and TNF-α in NPCs was inhibited by baicalin treatment. Baicalin treatment reversed IL-1β-induced increase in ROS production and MDA level, as well as decrease in SOD activity. Furthermore, baicalin treatment elevated the expression levels of Col II and Aggrecan and downregulated the expression levels of MMP3, MMP13, and ADAMTS5 in IL-1β-induced NPCs. A total of 402 related targets of baicalin and 134 related targets of intervertebral disk degeneration were found, and nine intersection targets were screened out. The Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that mitogen-activated protein kinase (MAPK) pathway was found to be involved in the effects of baicalin. Conclusions Baicalin exhibited protective effects on IL-1β-caused cell viability reduction, apoptosis, oxidative stress, inflammation, and extracellular matrix degradation in NPCs. In addition, we found c-Jun N-terminal kinase (JNK) and p38 MAPK pathways as targets of baicalin through bioinformatic analysis.

Abstract A013: Sex-dependent hematopoietic stem cell aging and leukemogenic potential

Abstract Aging of hematopoietic stem cells (HSCs) is linked to various blood disorders. But how HSC aging and sex co-influence leukemogenesis is poorly understood. By examining HSC aging in young, aging (75% lifespan) and longevity (110% lifespan) mice in males vs females, we have discovered that aging males have a significantly higher HSC frequency than females, and that male HSC aging is characterized by an expansion of Lin-CD150+SP HSCs and a decrease of Lin-CD150-SP HSCs, whereas females show opposite HSC changes towards aging and longevity. In line with HSC changes, aging males exhibit an increase of white blood cell counts that is primarily driven by increased myeloid cells, whereas aging females shows a decrease of white blood cell counts that is primarily driven by reduced lymphoid cells. To understand leukemogenic potential of aging HSCs in the true setting of organismal aging, we generated the first aging mouse model of chronic myeloid leukemia (CML) induced by BCR-ABL1 (Oncogene, 2021, 40: 3152-3163). We found that aging male HSCs are significantly more susceptible to BCR-ABL1 transformation and CML induction in aging male mice than aging female HSCs for CML induction in aging female mice. However, in the absence of oncogenic translocation genes, longevity female but not male mice rapidly develop B/myeloid mixed phenotype acute leukemia, a disease of HSC origin, in primary bone marrow transplantation recipients. Leukemias in our aging mouse models phenocopy those of humans in that the incidence of myeloid leukemia is higher in aging men than in women overall, but longevity women have a higher incidence of myeloid leukemia. Using single cell RNA sequencing analysis of HSCs, we found distinct cell clustering patterns and signaling pathways of aging HSCs between male and female mice. The top pathways seen in male HSC aging involve TGFβ and TNFα signaling that have been linked to poor drug response and persistence of human CML stem cells, while those for female HSC aging involve IFNα/γ signaling pathways that have been associated with good drug response of human CML stem cells. Together, our results demonstrate sex-dependent HSC aging and leukemogenic potential and that molecular changes in aging HSCs can inform drug response of leukemia stem cells. Our findings provide novel insight into the etiology of leukemia in males vs females and suggest potential benefit to use sex-based prevention and therapeutic strategies for leukemia in men vs women. Citation Format: WenYong Chen. Sex-dependent hematopoietic stem cell aging and leukemogenic potential [abstract]. In: Proceedings of the AACR Special Conference: Aging and Cancer; 2022 Nov 17-20; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2022;83(2 Suppl_1):Abstract nr A013.

Atherosclerotic-Derived Endothelial Cell Response Conducted by Titanium Oxide Nanotubes.

Atherosclerosis lesions are described as the formation of an occlusive wall-vessel plaque that can exacerbate infarctions, strokes, and even death. Furthermore, atherosclerosis damages the endothelium integrity, avoiding proper regeneration after stent implantation. Therefore, we investigate the beneficial effects of TiO2 nanotubes (NTs) in promoting the initial response of detrimental human atherosclerotic-derived endothelial cells (AThEC). We synthesized and characterized NTs on Ti6Al4V by anodization. We isolated AThEC and tested the adhesion long-lasting proliferation activity, and the modulation of focal adhesions conducted on the materials. Moreover, ultrastructural cell-surface contact at the nanoscale and membrane roughness were evaluated to explain the results. Our findings depicted improved filopodia and focal adhesions stimulated by the NTs. Similarly, the NTs harbored long-lasting proliferative metabolism after 5 days, explained by overcoming cell-contact interactions at the nanoscale. Furthermore, the senescent activity detected in the AThEC could be mitigated by the modified membrane roughness and cellular stretch orchestrated by the NTs. Importantly, the NTs stimulate the initial endothelial anchorage and metabolic recovery required to regenerate the endothelial monolayer. Despite the dysfunctional status of the AThEC, our study brings new evidence for the potential application of nano-configured biomaterials for innovation in stent technologies.

The Raman Deuterium Isotope Probing method as a new assay for evaluating the cytotoxicity level of the GSK2334470 to the MCF‐7 cells

AbstractThe Raman Deuterium Isotope Probing (Raman‐DIP) method was applied to study the response of human breast adenocarcinoma cells (the MCF‐7 cells) to the GSK2334470 drug in vitro. The cytotoxicity level of the GSK2334470 to the MCF‐7 cell evaluated by the Raman‐DIP method and CCK‐8 assay was highly matched, as both methods suggested that 10 μM was the cytotoxic concentration for inhibition of cell regeneration. In additional, the relationship between drug dose and the cell response (RSI) were fitted, showing that our experimental data were consistent with the dose–response relationship, and the Raman‐DIP could be applied for sensitivity testing of targeted anti‐cancer drugs. The variation of the bio‐macromolecules reflected by the Raman signal in the fingerprint range was closely related to the GSK2334470 concentration. The Raman‐DIP method can monitor the inhibition of the GSK2334470 to the MCF‐7 cell proliferation with high accuracy as the CCK‐8 assay. The combination of Raman‐DIP results and fingerprint range information can effectively improve the evaluation, serving as a supplement for the existing anti‐cancer drug sensibility assay. This method is valuable to reveal the intracellular macromolecules changes related to the drug treatment.

Stress Granules Inhibit Coxsackievirus B3-Mediated Cell Death via Reduction of Mitochondrial Reactive Oxygen Species and Viral Extracellular Release.

Stress granules (SGs) are cytoplasmic aggregates of RNA-protein complexes that form in response to various cellular stresses and are known to restrict viral access to host translational machinery. However, the underlying molecular mechanisms of SGs during viral infections require further exploration. In this study, we evaluated the effect of SG formation on cellular responses to coxsackievirus B3 (CVB3) infection. Sodium arsenite (AS)-mediated SG formation suppressed cell death induced by tumor necrosis factor-alpha (TNF-a)/cycloheximide (CHX) treatment in HeLa cells, during which G3BP1, an essential SG component, contributed to the modulation of apoptosis pathways. SG formation in response to AS treatment blocked CVB3-mediated cell death, possibly via the reduction of mitochondrial reactive oxygen species. Furthermore, we examined whether AS treatment would affect small extracellular vesicle (sEV) formation and secretion during CVB3 infection and modulate human monocytic cell (THP-1) response. CVB3-enriched sEVs isolated from HeLa cells were able to infect and replicate THP-1 cells without causing cytotoxicity. Interestingly, sEVs from AS-treated HeLa cells inhibited CVB3 replication in THP-1 cells. These findings suggest that SG formation during CVB3 infection modulates cellular response by inhibiting the release of CVB3-enriched sEVs.