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Frontiers in Bioengineering and... 2024The stiffness of the extracellular matrix plays a crucial role in cell motility and spreading, influencing cell morphology through cytoskeleton organization and...
The stiffness of the extracellular matrix plays a crucial role in cell motility and spreading, influencing cell morphology through cytoskeleton organization and transmembrane proteins' expression. In this context, mechanical characterization of both cells and the extracellular matrix gains prominence for enhanced diagnostics and clinical decision-making. Here, we investigate the combined effect of mechanotransduction and ionizing radiations on altering cells' mechanical properties, analysing mammary cell lines (MCF10A and MDA-MB-231) after X-ray radiotherapy (2 and 10 Gy). We found that ionizing radiations sensitively affect adenocarcinoma cells cultured on substrates mimicking cancerous tissue stiffness (15 kPa), inducing an increased structuration of paxillin-rich focal adhesions and cytoskeleton: this process translates in the augmentation of tension at the actin filaments level, causing cellular stiffness and consequently affecting cytoplasmatic/nuclear morphologies. Deeper exploration of the intricate interplay between mechanical factors and radiation should provide novel strategies to orient clinical outcomes.
PubMed: 38903185
DOI: 10.3389/fbioe.2024.1408789 -
CNS Neuroscience & Therapeutics Jun 2024Glioblastoma multiforme (GBM) is the most aggressive primary brain malignancy. Novel therapeutic modalities like tumor electric field therapy (TEFT) have shown promise,...
BACKGROUND
Glioblastoma multiforme (GBM) is the most aggressive primary brain malignancy. Novel therapeutic modalities like tumor electric field therapy (TEFT) have shown promise, but underlying mechanisms remain unclear. The extracellular matrix (ECM) is implicated in GBM progression, warranting investigation into TEFT-ECM interplay.
METHODS
T98G cells were treated with TEFT (200 kHz, 2.2 V/m) for 72 h. Collagen type VI alpha 1 (COL6A1) was identified as hub gene via comprehensive bioinformatic analysis based on RNA sequencing (RNA-seq) and public glioma datasets. TEFT intervention models were established using T98G and Ln229 cell lines. Pre-TEFT and post-TEFT GBM tissues were collected for further validation. Focal adhesion pathway activity was assessed by western blot. Functional partners of COL6A1 were identified and validated by co-localization and survival analysis.
RESULTS
TEFT altered ECM-related gene expression in T98G cells, including the hub gene COL6A1. COL6A1 was upregulated in GBM and associated with poor prognosis. Muti-database GBM single-cell analysis revealed high-COL6A1 expression predominantly in malignant cell subpopulations. Differential expression and functional enrichment analyses suggested COL6A1 might be involved in ECM organization and focal adhesion. Western blot (WB), immunofluorescence (IF), and co-immunoprecipitation (Co-IP) experiments revealed that TEFT significantly inhibited expression of COL6A1, hindering its interaction with ITGA5, consequently suppressing the FAK/Paxillin/AKT pathway activity. These results suggested that TEFT might exert its antitumor effects by downregulating COL6A1 and thereby inhibiting the activity of the focal adhesion pathway.
CONCLUSION
TEFT could remodel the ECM of GBM cells by downregulating COL6A1 expression and inhibiting focal adhesion pathway. COL6A1 could interact with ITGA5 and activate the focal adhesion pathway, suggesting that it might be a potential therapeutic target mediating the antitumor effects of TEFT.
Topics: Collagen Type VI; Humans; Glioblastoma; Brain Neoplasms; Electric Stimulation Therapy; Cell Line, Tumor; Animals; Mice, Nude; Mice
PubMed: 38887185
DOI: 10.1111/cns.14802 -
Nan Fang Yi Ke Da Xue Xue Bao = Journal... May 2024To evaluate the therapeutic effect of normal mouse serum on radiation pneumonitis in mice and explore the possible mechanism.
OBJECTIVE
To evaluate the therapeutic effect of normal mouse serum on radiation pneumonitis in mice and explore the possible mechanism.
METHODS
Mouse models of radiation pneumonitis induced by thoracic radiation exposure were given intravenous injections of 100 μL normal mouse serum or normal saline immediately after the exposure followed by injections once every other day for a total of 8 injections. On the 15th day after irradiation, histopathological changes of the lungs of the mice were examined using HE staining, the levels of TNF-α, TGF-β, IL-1α and IL-6 in the lung tissue and serum were detected using ELISA, and the percentages of lymphocytes in the lung tissue were analyzed with flow cytometry. High-throughput sequencing of exosome miRNA was carried out to explore the changes in the signaling pathways. The mRNA expression levels of the immune-related genes were detected by qRT-PCR, and the protein expressions of talin-1, tensin2, FAK, vinculin, α-actinin and paxillin in the focal adhesion signaling pathway were detected with Western blotting.
RESULTS
In the mouse models of radiation pneumonitis, injections of normal mouse serum significantly decreased the lung organ coefficient, lowered the levels of TNF-α, TGF-β, IL-1α and IL-6 in the serum and lung tissues, and ameliorated infiltration of CD45, CD4 and T lymphocytes in the lung tissue (all <0.05). The expression levels of and genes at both the mRNA and protein levels and the protein expressions of talin-1, tensin2, FAK, vinculin, α‑actinin and paxillin were all significantly down-regulated in the mouse models after normal mouse serum treatment.
CONCLUSION
Normal mouse serum ameliorates radiation pneumonitis in mice by inhibiting the expressions of key proteins in the Focal adhesion signaling pathway.
Topics: Animals; Mice; Signal Transduction; Radiation Pneumonitis; Focal Adhesions; Lung; Interleukin-6; Disease Models, Animal; Tumor Necrosis Factor-alpha; Transforming Growth Factor beta; MicroRNAs; Interleukin-1alpha
PubMed: 38862437
DOI: 10.12122/j.issn.1673-4254.2024.05.01 -
Biomedical Optics Express May 2024Single-molecule super-resolution imaging is instrumental in investigating cellular architecture and organization at the nanoscale. Achieving precise 3D nanometric...
Single-molecule super-resolution imaging is instrumental in investigating cellular architecture and organization at the nanoscale. Achieving precise 3D nanometric localization when imaging structures throughout mammalian cells, which can be multiple microns thick, requires careful selection of the illumination scheme in order to optimize the fluorescence signal to background ratio (SBR). Thus, an optical platform that combines different wide-field illumination schemes for target-specific SBR optimization would facilitate more precise 3D nanoscale studies of a wide range of cellular structures. Here, we demonstrate a versatile multimodal illumination platform that integrates the sectioning and background reduction capabilities of light sheet illumination with homogeneous, flat-field epi- and TIRF illumination. Using primarily commercially available parts, we combine the fast and convenient switching between illumination modalities with point spread function engineering to enable 3D single-molecule super-resolution imaging throughout mammalian cells. For targets directly at the coverslip, the homogenous intensity profile and excellent sectioning of our flat-field TIRF illumination scheme improves single-molecule data quality by providing low fluorescence background and uniform fluorophore blinking kinetics, fluorescence signal, and localization precision across the entire field of view. The increased contrast achieved with LS illumination, when compared with epi-illumination, makes this illumination modality an excellent alternative when imaging targets that extend throughout the cell. We validate our microscopy platform for improved 3D super-resolution imaging by two-color imaging of paxillin - a protein located in the focal adhesion complex - and actin in human osteosarcoma cells.
PubMed: 38855669
DOI: 10.1364/BOE.521362 -
Journal of Oral and Maxillofacial... 2024[This corrects the article on p. 727 in vol. 27, PMID: 38304525.].
[This corrects the article on p. 727 in vol. 27, PMID: 38304525.].
PubMed: 38800422
DOI: 10.4103/jomfp.jomfp_96_24 -
International Journal of Molecular... May 2024Malignant melanoma represents a form of skin cancer characterized by a bleak prognosis and heightened resistance to traditional therapies. Quercetin has demonstrated...
Malignant melanoma represents a form of skin cancer characterized by a bleak prognosis and heightened resistance to traditional therapies. Quercetin has demonstrated notable anti-carcinogenic, anti-inflammatory, anti-oxidant, and pharmacological effects across various cancer types. However, the intricate relationship between quercetin's anti-cancer properties and ganglioside expression in melanoma remains incompletely understood. In this study, quercetin manifests specific anti-proliferative, anti-migratory, and cell-cycle arrest effects, inducing mitochondrial dysfunction and apoptosis in two melanoma cancer cell lines. This positions quercetin as a promising candidate for treating malignant melanoma. Moreover, our investigation indicates that quercetin significantly reduces the expression levels of ganglioside GD3 and its synthetic enzyme. Notably, this reduction is achieved through the inhibition of the FAK/paxillin/Akt signaling pathway, which plays a crucial role in cancer development. Taken together, our findings suggest that quercetin may be a potent anti-cancer drug candidate for the treatment of malignant melanoma.
Topics: Quercetin; Humans; Melanoma; Apoptosis; Mitochondria; Cell Line, Tumor; Gangliosides; Cell Proliferation; Signal Transduction; Down-Regulation; Gene Expression Regulation, Neoplastic; Cell Movement; Skin Neoplasms; Proto-Oncogene Proteins c-akt
PubMed: 38791186
DOI: 10.3390/ijms25105146 -
Biology May 2024Cell-to-cell distant mechanical communication has been demonstrated using in vitro and in vivo models. However, the molecular mechanisms underlying long-range cell...
Cell-to-cell distant mechanical communication has been demonstrated using in vitro and in vivo models. However, the molecular mechanisms underlying long-range cell mechanoresponsive interactions remain to be fully elucidated. This study further examined the roles of α-Catenin and Piezo1 in traction force-induced rapid branch assembly of airway smooth muscle (ASM) cells on a Matrigel hydrogel containing type I collagen. Our findings demonstrated that siRNA-mediated downregulation of α-Catenin or Piezo1 expression or chemical inhibition of Piezo1 activity significantly reduced both directional cell movement and branch assembly. Regarding the role of N-cadherin in regulating branch assembly but not directional migration, our results further confirmed that siRNA-mediated downregulation of α-Catenin expression caused a marked reduction in focal adhesion formation, as assessed by focal Paxillin and Integrin α5 localization. These observations imply that mechanosensitive α-Catenin is involved in both cell-cell and cell-matrix adhesions. Additionally, Piezo1 partially localized in focal adhesions, which was inhibited by siRNA-mediated downregulation of α-Catenin expression. This result provides insights into the Piezo1-mediated mechanosensing of traction force on a hydrogel. Collectively, our findings highlight the significance of α-Catenin in the regulation of cell-matrix interactions and provide a possible interpretation of Piezo1-mediated mechanosensing activity at focal adhesions during cell-cell mechanical communication.
PubMed: 38785839
DOI: 10.3390/biology13050357 -
The Journal of Biological Chemistry May 2024Cancer testis antigens (CTAs) are a collection of proteins whose expression is normally restricted to the gamete but abnormally activated in a wide variety of tumors....
Cancer testis antigens (CTAs) are a collection of proteins whose expression is normally restricted to the gamete but abnormally activated in a wide variety of tumors. The CTA, Testis-specific serine kinase 6 (TSSK6), is essential for male fertility in mice. The functional relevance of TSSK6 to cancer, if any, has not previously been investigated. Here we find that TSSK6 is frequently anomalously expressed in colorectal cancer and patients with elevated TSSK6 expression have reduced relapse-free survival. Depletion of TSSK6 from colorectal cancer cells attenuates anchorage-independent growth, invasion, and growth in vivo. Conversely, overexpression of TSSK6 enhances anchorage independence and invasion in vitro as well as in vivo tumor growth. Notably, ectopic expression of TSSK6 in semi-transformed human colonic epithelial cells is sufficient to confer anchorage independence and enhance invasion. In somatic cells, TSSK6 co-localizes with and enhances the formation of paxillin and tensin-positive foci at the cell periphery, suggesting a function in focal adhesion formation. Importantly, TSSK6 kinase activity is essential to induce these tumorigenic behaviors. Our findings establish that TSSK6 exhibits oncogenic activity when abnormally expressed in colorectal cancer cells. Thus, TSSK6 is a previously unrecognized intervention target for therapy, which could exhibit an exceptionally broad therapeutic window.
PubMed: 38762178
DOI: 10.1016/j.jbc.2024.107380 -
Molecular Human Reproduction Apr 2024Paxillin is a ubiquitously expressed adaptor protein integral to focal adhesions, cell motility, and apoptosis. Paxillin has also recently been implicated as a mediator...
Paxillin is a ubiquitously expressed adaptor protein integral to focal adhesions, cell motility, and apoptosis. Paxillin has also recently been implicated as a mediator of nongenomic androgen receptor (AR) signaling in prostate cancer and other cells. We sought to investigate the relationship between paxillin and AR in granulosa cells (GCs), where androgen actions, apoptosis, and focal adhesions are of known importance, but where the role of paxillin is understudied. We recently showed that paxillin knockout in mouse GCs increases fertility in older mice. Here, we demonstrate that paxillin knockdown in human granulosa-derived KGN cells, as well as knockout in mouse primary GCs, results in reduced AR protein but not reduced mRNA expression. Further, we find that both AR protein and mRNA half-lives are reduced by approximately one-third in the absence of paxillin, but that cells adapt to chronic loss of paxillin by upregulating AR gene expression. Using co-immunofluorescence and proximity ligation assays, we show that paxillin and AR co-localize at the plasma membrane in GCs in a focal adhesion kinase-dependent way, and that disruption of focal adhesions leads to reduced AR protein level. Our findings suggest that paxillin recruits AR to the GC membrane, where it may be sequestered from proteasomal degradation and poised for nongenomic signaling, as reported in other tissues. To investigate the physiological significance of this in disorders of androgen excess, we tested the effect of GC-specific paxillin knockout in a mouse model of polycystic ovary syndrome (PCOS) induced by chronic postnatal dihydrotestosterone (DHT) exposure. While none of the control mice had estrous cycles, 33% of paxillin knockout mice were cycling, indicating that paxillin deletion may offer partial protection from the negative effects of androgen excess by reducing AR expression. Paxillin-knockout GCs from mice with DHT-induced PCOS also produced more estradiol than GCs from littermate controls. Thus, paxillin may be a novel target in the management of androgen-related disorders in women, such as PCOS.
Topics: Animals; Female; Humans; Mice; Focal Adhesions; Gene Expression Regulation; Granulosa Cells; Mice, Knockout; Paxillin; Receptors, Androgen; Signal Transduction
PubMed: 38718206
DOI: 10.1093/molehr/gaae018 -
Biomaterials Advances Jul 2024The muscle tendon junction (MTJ) plays a crucial role in transmitting the force generated by muscles to the tendon and then to the bone. Injuries such as tears and...
The muscle tendon junction (MTJ) plays a crucial role in transmitting the force generated by muscles to the tendon and then to the bone. Injuries such as tears and strains frequently happen at the MTJ, where the regenerative process is limited due to poor vascularization and the complex structure of the tissue. Current solutions for a complete tear at the MTJ have not been successful and therefore, the development of a tissue-engineered MTJ may provide a more effective treatment. In this study, decellularised extracellular matrix (DECM) derived from sheep MTJ was used to provide a scaffold for the MTJ with the relevant mechanical properties and differentiation cues such as the relase of growth factors. Human mesenchymal stem cells (MSCs) were seeded on DECM and 10 % cyclic strain was applied using a bioreactor. MSCs cultured on DECM showed significantly higher gene and protein expression of MTJ markers such as collagen 22, paxillin and talin, than MSCs in 2D culture. Although collagen 22 protein expression was higher in the cells with strain than without strain, reduced gene expression of other MTJ markers was observed when the strain was applied. DECM combined with 10 % strain enhanced myogenic differentiation, while tenogenic differentiation was reduced when compared to static cultures of MSCs on DECM. For the first time, these results showed that DECM derived from the MTJ can induce MTJ marker gene and protein expression by MSCs, however, the effect of strain on the MTJ development in DECM culture needs further investigation.
Topics: Mesenchymal Stem Cells; Tendons; Humans; Animals; Tissue Engineering; Cell Differentiation; Sheep; Tissue Scaffolds; Decellularized Extracellular Matrix; Tensile Strength; Extracellular Matrix; Cells, Cultured
PubMed: 38692180
DOI: 10.1016/j.bioadv.2024.213873