Effects of substrate stiffness on the polarization states of murine bone marrow-derived neutrophils
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE217379 | GEO | 2022/12/03
REPOSITORIES: GEO
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Project description:We undertook mRNA microarray and gene ontology analyses to screen out substrate stiffness-dependent genes. Total mRNA were extracted from E17 cortical neurons grown on soft or stiff substrates at 5 or 16 hr time points. We identified 114 differentially-expressed mRNA transcripts in cells grown on 0.1 kPa and 20 kPa gels at the 5 hr time-point. Among them, 66 were upregulated in 0.1 kPa gel cultures and the remainder were downregulated (compared to cells grown on stiffer substrates). The expressions of three endocytic genes (Cltc, Dab2, and Myo6) and four adhesion genes(Vcl, Robo2, Nrcam, and Cad11) were confirmed by QGP and smRNA FISH.
2018-01-08 | GSE102350 | GEO
Project description:Elevated intraocular pressure, a major risk factor of glaucoma, is caused by the abnormal function of trabecular outflow pathways. Human trabecular meshwork (HTM) tissue plays an important role in the outflow pathways. However, the molecular mechanisms that how TM cells respond to the elevated IOP are largely unknown. We cultured primary HTM cells on polyacrylamide gels with tunable stiffness corresponding to Young's moduli ranging from 1.1 to 50 kPa. Then next‐generation RNA sequencing (RNA‐seq) was performed to obtain the transcriptomic profiles of HTM cells. Bioinformatics analysis revealed that genes related to glaucoma including DCN, SPARC, and CTGF, were significantly increased with elevated substrate stiffness, as well as the global alteration of HTM transcriptome. Extracellular matrix (ECM)‐related genes were selectively activated in response to the elevated substrate stiffness, consistent with the known molecular alteration in glaucoma. Human normal and glaucomatous TM tissues were also obtained to perform RNA‐seq experiments and supported the substrate stiffness‐altered transcriptome profiles from the in vitro cell model. The current study profiled the transcriptomic changes in human TM cells upon increasing substrate stiffness. Global change of ECM‐related genes indicates that the in vitro substrate stiffness could greatly affect the biological processes of HTM cells. The in vitro HTM cell model could efficiently capture the main pathogenetic process in glaucoma patients, and provide a powerful method to investigate the underlying molecular mechanisms.
2019-12-16 | GSE123100 | GEO
Project description:Extracellular matrix proteomic profiling Mass spectrometry characterization illustrated a reprogrammed proteome in response to matrix stiffness, including a core subset of 84 ECM-specific proteins, including various ECM regulators and tissue-associated ECM-related proteins. Further, we observe remodeling of ECM proteome with softer substrates associated with metabolic/mitochondrial network (178 proteins), while with 40 kPa substrate (92 proteins enriched) resulting in networks associated with enrichment of collagen biosynthesis, integrin cell surface interactions, and extracellular matrix organization networks.
| MSV000092971 | MassIVE
Project description:Cells interact with their mechanical environment and respond in consequence. Mechanical cues can have a wide range of influences on cell behaviour, ranging from guidance of differentiation and cell fate to immune activation. The impact of substrate stiffness on primary macrophages - a key player in innate immunity and inflammation - had not been previously studied. We prepared bone marrow-derived macrophage cultures from adult rat hematopoietic stem cells exposed to M-CSF, and cultured these on polyacrylamide substrates of controlled stiffness (ranging from 50 to 0.1 kPa shear modulus, covering the range found in physiological tissues) for 3 days. The RNA from these cells was then extracted and sequenced.
2023-04-15 | E-MTAB-7900 | biostudies-arrayexpress
Project description:Treating recurrent GBM is a clinical challenge due to its highly resistant and aggressive nature. In order to develop new therapeutic targets for recurrent GBM a better understanding of its molecular landscape is necessary. Here we used a cellular model, developed in our lab which generates paired primary and recurrent samples from GBM cell lines and primary patient samples hence allowing us to compare the molecular differences between the two populations. Total RNA seq analysis of parent and recurrent population of two cell lines and one patient sample revealed a significant upregulation of Extracellular matrix interaction in recurrent population. Since matrix stiffness plays a pivotal role in cell-ECM interaction and downstream signaling, we developed a system that mimicked the brain like substrate stiffness by using collagen coated polyacrylamide-based substrate whose stiffness can be modified from normal brain (0.5kPa) to tumorigenic (10kPa). Using these substrates, we were able to capture the morphological and physiological differences between parent and recurrent GBM which were not evident on plastic surfaces (~1 GPa). On 0.5kPa, unlike circular parent cells, recurrent GBM cells showed two morphologies (circular and elongated). The recurrent cells growing on 0.5kPa also showed higher proliferation, invasion, migration and in-vivo tumorigenicity in orthotropic GBM mouse model, compared to parent cells. Furthermore, recurrent cells exhibited elevated velocity irrespective of substrate stiffness, which indicated that recurrent cells may possess inherent differential mechanosignalling ability which was reflected by higher expression of ECM proteins like Collagen IVA, MMP2 and MMP9. Moreover, mice brain injected with recurrent cells grown on 0.5kPa substrate showed higher Young’s modulus values suggesting that recurrent cells conditioned on 0.5kPa make the surrounding ECM stiffer. Importantly, inhibition of EGFR signaling, that is amplified with tissue stiffening in GBM resulted in decreased invasion, migration and proliferation in 0.5kPa recurrent cells, but interestingly survival remained unaffected, highlighting the importance of mimicking the physiological stiffness of the brain mimicking clinical scenario. Total RNA seq analysis of parent and recurrent cells grown on plastic and 0.5kPa substrate identified PLEKHA7 as significantly upregulated gene specifically in 0.5kPa recurrent sample. Higher protein expression of PLEKHA7 in recurrent GBM as compared to primary GBM was validated in patient biopsies. Accordingly, PLEKHA7 knockdown reduced invasion and survival of recurrent GBM cells. Together, these data provides a model system that captures the differential mechanosensing signals of primary and recurrent GBM cells and identifies a novel potential target specific for recurrent GBM.
2023-06-29 | E-MTAB-11293 | biostudies-arrayexpress
Project description:Purpose: Identify the effect of substrate stiffness on gene expression Methods:Evaluating for differentially expressed mRNAs in the SKOV-3 cells grown on the different substrates via High-throughput sequence Results: We found that the general direction of changes in gene expression of cells grown on the different substrates and the most significant signalling pathways and the expression of gene orthologs broadly involved in platinum drug resistance, apoptosis, cell cycle. Conclusions: Our study represents the first detailed analysis of the effects of substrate stiffness on gene expression of ovarian cancer cells.
2021-06-26 | GSE178888 | GEO
Project description:Substrate elasticity may direct cell-fate decisions of stem cells. However, it is largely unclear how matrix stiffness impacts on differentiation of induced pluripotent stem cells (iPSCs) and if this is also reflected by epigenetic modifications. We have therefore cultured iPSCs on tissue culture plastic (TCP) and polydimethylsiloxane (PDMS) with different Young´s modulus (0.2 kPa, 16 kPa, or 64 kPa) to investigate the sequel on growth and differentiation towards endoderm, mesoderm, and ectoderm. Immunofluorescence and gene expression of canonical differentiation markers was hardly affected by the substrates. Notably, when we analyzed DNA methylation profiles of iPSCs or after three-lineage differentiation, we did not see any significant differences on the three different PDMS elasticities. Only when we compared DNA methylation profiles on PDMS-substrates versus TCP, we observed clear epigenetic differences, particularly upon mesodermal differentiation. Taken together, stiffness of PDMS-substrates did not impact on directed differentiation of iPSCs, whereas the moderate epigenetic differences on TCP might also be attributed to other chemical parameters.
2024-06-12 | GSE253190 | GEO
Project description:In vitro cultures of primary cardiac fibroblasts (CFs), the major extracellular matrix (ECM)-producing cells of the heart, are used to determine molecular mechanisms of cardiac fibrosis. However, the supraphysiologic stiffness of tissue culture polystyrene (TCPS) automatically triggers the conversion of CFs into an activated myofibroblast-like state, and serial passage of the cells results in the induction of replicative senescence. These dramatic phenotypic switches confound interpretation of experimental data obtained with cultured CFs. In an attempt to circumvent TCPS-induced activation and senescence of CFs, we utilized poly (ethylene glycol) (PEG) hydrogels as cell culture platforms with low and high stiffness formulations to mimic healthy and fibrotic cardiac ECM, respectively. As hypothesized, low hydrogel stiffness converted activated CFs into a quiescent state with reduced abundance of a-smooth muscle actin (a-SMA)-containing stress fibers. Unexpectedly, lower substrate stiffness concomitantly augmented CF senescence, marked by elevated senescence-associated b-galactosidase (SA-b-Gal) activity and increased expression of p16 and p21, which are cyclin-dependent kinase (CDK) inhibitors and markers of senescence. Using dynamically stiffening hydrogels with phototunable crosslinking capabilities, we demonstrate that substrate-induced CF senescence is partially reversible. RNA-sequencing analysis revealed widespread transcriptional reprogramming of CFs cultured on low stiffness hydrogels, with a dramatic reduction in the expression of profibrotic genes encoding ECM proteins, and an attendant increase in expression of NF-kB-responsive inflammatory genes that typify the senescence-associated secretory phenotype (SASP). Our findings further demonstrate that alterations in matrix stiffness profoundly impact CF cell state transitions, and suggest mechanisms by which CFs change phenotype in vivo depending on the stiffness of the myocardial microenvironment to which they are exposed.
2023-10-24 | GSE244614 | GEO
Project description:Cell migration is central to many biological processes including embryonic development, wound healing, and cancer progression. Cell migration is sensitive to environmental stiffness, and many cell types exhibit a stiffness optimum at which migration is maximal. Here we present a cell migration simulator that predicts a stiffness optimum that can be shifted by altering the number of active molecular motors and clutches. This prediction is verified experimentally by comparing cell traction and F-actin retrograde flow for two cell types with differing amounts of active motors and clutches: embryonic chick forebrain neurons (ECFNs; optimum ~1 kPa) and U251 glioma cells (optimum ~100 kPa). In addition, the model predicts, and experiments confirm, that the stiffness optimum of U251 glioma cell migration, morphology, and F-actin retrograde flow rate can be shifted to lower stiffness by simultaneous drug inhibition of myosin II motors and integrin-mediated adhesions.
2017-05-23 | GSE95680 | GEO
Project description:Effects of substrate stiffness on the polarization states of murine bone marrow-derived neutrophils
| PRJNA898766 | ENA