Integrin Receptors Research Articles

Role of TSP-1 and its receptor ITGB3 in the renal tubulointerstitial injury of focal segmental glomerulosclerosis

Focal segmental glomerulosclerosis (FSGS), a common cause of primary glomerulonephritis, has a poor prognosis and is pathologically featured by tubulointerstitial injury. Thrombospondin-1 (TSP-1) is an extracellular matrix protein that acts in combination with different receptors in the kidney. Here, we analyzed the tubular expression of TSP-1 and its receptor integrin β3 (ITGB3) in FSGS. Previously the renal interstitial chip analysis of FSGS patients with tubular interstitial injury showed that the expressions of TSP-1 and ITGB3 were up-regulated. We found that the level of TSP-1 and ITGB3 increased in the tubular cells of FSGS patients. The serum level of TSP-1 increased and was correlated to the degree of tubulointerstitial lesions in FSGS patients. THBS1/ITGB3 signaling induced renal tubular injury in HK-2 cells exposure to BSA and the ADR-induced nephropathy model. THBS1 knockout ameliorated tubular injury and renal fibrosis in ADR-treated mice. THBS1 knockdown decreased the expression of KIM-1 and caspase 3 in the HK-2 cells treated with BSA, while THBS1 overexpression could induce tubular injury. In vivo, we identified cyclo-RGDfK as an agent to block the binding of TSP-1 to ITGB3. Cyclo-RGDfK treatment could alleviate ADR-induced renal tubular injury and interstitial fibrosis in mice. Moreover, TSP-1 and ITGB3 were colocalized in tubular cells of FSGS patients and ADR-treated mice. Taken together, our data showed that TSP-1/ITGB3 signaling contributed to the development of renal tubulointerstitial injury in FSGS, potentially identifying a new therapeutic target for FSGS.

Thrombospondin-4 deletion does not exacerbate muscular dystrophy in β-sarcoglycan-deficient and laminin α2 chain-deficient mice

Muscular dystrophy is a group of genetic disorders that lead to muscle wasting and loss of muscle function. Identifying genetic modifiers that alleviate symptoms or enhance the severity of a primary disease helps to understand mechanisms behind disease pathology and facilitates discovery of molecular targets for therapy. Several muscular dystrophies are caused by genetic defects in the components of the dystrophin-glycoprotein adhesion complex (DGC). Thrombospondin-4 overexpression has been shown to mitigate dystrophic disease in mouse models for Duchenne muscular dystrophy (dystrophin deficiency) and limb-girdle muscular dystrophy type 2F (LGMD2F, δ-sarcoglycan deficiency), while deletion of the thrombospondin-4 gene exacerbated the diseases. Hence, thrombospondin-4 has been considered a candidate molecule for therapy of muscular dystrophies involving the DGC. We have investigated whether thrombospondin-4 could act as a genetic modifier for other DGC-associated diseases: limb-girdle muscular dystrophy type 2E (LGMD2E, β-sarcoglycan deficiency) and laminin α2 chain-deficient muscular dystrophy (LAMA2-RD). Deletion of the thrombospondin-4 gene in mouse models for LGMD2E and LAMA2-RD, respectively, did not result in worsening of the dystrophic phenotype. Loss of thrombospondin-4 did not enhance sarcolemma damage and did not impair trafficking of transmembrane receptors integrin α7β1 and dystroglycan in double knockout muscles. Our results suggest that thrombospondin-4 might not be a relevant therapeutic target for all muscular dystrophies involving the DGC. This data also demonstrates that molecular pathology between very similar diseases like LGMD2E and 2F can differ significantly.

Integrin receptor-targeted, doxorubicin-loaded cerium oxide nanoparticles delivery to combat glioblastoma.

Aim: To assess the chemo-immunomodulatory effects of doxorubicin-loaded cerium oxide nanoparticles coated with oleyl amine-linked cyclic RGDfK peptide (CeNP+Dox+RGD) to target both gliomas and its tumor microenvironment (TME) via integrin receptors. Materials& methods: CeNP+Dox+RGD nanoparticles are synthesized by the sequential addition of cerium III chloride heptahydrate, beta-cyclodextrin, oleic acid, and F127 micelle (CeNP). Doxorubicin was then loaded into CeNPs and coated with oleyl amine-linked cyclic RGDfK peptide to form stable CeNP+Dox+RGD nanoparticles. Results: CeNP+Dox+RGD nanoparticles crossed blood-brain barrier (BBB) effectively and demonstrated threefold enhanced survivability in glioma-bearing mice. The IHC profiling of glial tumor cross-sections showed increased CD80 expression (M1 TAMs) and decreased arginase-1 expression (M2 TAMs). Conclusion: CeNP+Dox+RGD can be an immunotherapeutic treatment option to combat glioblastoma.

Photosensitive and dual-targeted chromium nanoparticle delivering small interfering RNA YTHDF1 for molecular-targeted immunotherapy in liver cancer

Tumor-associated macrophages (TAMs) are a promising target for cancer immunotherapy, but delivering therapeutic agents to TAMs within the tumor microenvironment (TME) is challenging. In this study, a photosensitive, dual-targeting nanoparticle system (M.RGD@Cr-CTS-siYTHDF1 NPs) was developed. The structure includes a shell of DSPE-modified RGD peptides targeting integrin receptors on tumor cells and carboxymethyl mannose targeting CD206 receptors on macrophages, with a core of chitosan adsorbing m6A reading protein YTHDF1 siRNA and chromium nanoparticles (Cr NPs). The approach is specifically designed to target TAM and cancer cells, utilizing the photothermal effect of Cr NPs to disrupt the TME and deliver siYTHDF1 to TAM. In experiments with tumor-bearing mice, M.RGD@Cr-CTS-siYTHDF1 NPs, when exposed to laser irradiation, effectively killed tumor cells, disrupted the TME, delivered siYTHDF1 to TAMs, silenced the YTHDF1 gene, and shifted the STAT3-STAT1 equilibrium by reducing STAT3 and enhancing STAT1 expression. This reprogramming of TAMs towards an anti-tumor phenotype led to a pro-immunogenic TME state. The strategy also suppressed immunosuppressive IL-10 production, increased expression of immunostimulatory factors (IL-12 and IFN-γ), boosted CD8 + T cell infiltration and M1-type TAMs, and reduced Tregs and M2-type TAMs within the TME. In conclusion, the dual-targeting M.RGD@Cr-CTS-siYTHDF1 NPs, integrating dual-targeting capabilities with photothermal therapy (PTT) and RNA interference, offer a promising approach for molecular targeted cancer immunotherapy with potential for clinical application.Graphical

Abstract PO-035: Genome-wide CRISPR knockout screen identifies sialylation as a tumor suppressive mechanism in B-cell malignancies

Abstract Ba/F3 is a murine pro-B cell line that is dependent on interleukin-3 (IL-3) for its survival and proliferation. It has been widely used for identifying oncogenes and mechanisms of drug-resistance. Using this cell line, we conducted a genome-wide CRISPR knockout screen to identify genes whose loss can confer IL-3 independence, hence may be implied as tumor suppressor genes (TSGs). We identified several genes that map to human chromosome 6q, a frequent deletion hotspot in B-cell malignancies, that may act as TSGs. One of these putative novel TSGs is Slc35a1, which encodes the membrane solute carrier that facilitates the transport of nucleotide sugar (CMP-sialic acid) into the Golgi apparatus for subsequent sialylation. Interestingly, in addition to Slc35a1, pathway analysis of hits from the CRISPR screen identified several genes in the sialic acid metabolism pathway, Nans, Cmas, and St3gal5. We confirmed that their silencing led to decreased protein sialyation and could confer IL-3-independent cell survival and growth in Ba/F3 cells. We continued to characterize Slc35a1 since its homozygous and heterozygous deletions are seen in 7-40% of different types of B-cell malignancies, including approximately 40% of patients with Diffuse Large B-cell Lymphoma. Slc35a1 deletion in murine and human cell-lines conferred cytokine-independent growth. Tail-vein injection of Slc35a1-silenced Ba/F3 cells resulted in allograft tumor growth presenting with splenomegaly and lymphadenopathy. Using Maackia amurensis lectin II (MAL II) to pull down sialylated proteins for mass spectrometry, we identified proteins that were differentially sialylated in Slc35a1 wild-type and knockout cells, among which are the α and β subunits of the integrin receptor, lymphocyte functional antigen 1 (LFA-1). As integrin desialylation can promote its constitutive activation, we are currently studying the impact of LFA-1 sialylation on B-cell function and transformation. We have also developed mouse models with B-cell specific ablation of Nans and Slc35a1 to further evaluate the role of sialylation on B-cell development and tumorigenesis in vivo. Overall, our study demonstrates that sialylation may function as a tumor suppressive mechanism and serve as a therapeutic target in B-cell cancers. Citation Format: Namratha Sheshadri, Rongrong Li, Muhammad Usama Tariq, Jianliang Shen, Jaeyong Jung, Junrong Yan, Kevin Lu, Zhiyuan Shen, Ping Xie, Wei-Xing Zong. Genome-wide CRISPR knockout screen identifies sialylation as a tumor suppressive mechanism in B-cell malignancies [abstract]. In: Proceedings of the Fourth AACR International Meeting on Advances in Malignant Lymphoma: Maximizing the Basic-Translational Interface for Clinical Application; 2024 Jun 19-22; Philadelphia, PA. Philadelphia (PA): AACR; Blood Cancer Discov 2024;5(3_Suppl):Abstract nr PO-035.

Tumour-specific activation of a tumour-blood transport improves the diagnostic accuracy of blood tumour markers in mice

The accuracy of blood-based early tumour recognition is compromised by signal production at non-tumoral sites, low amount of signal produced by small tumours, and variable tumour production. Here we examined whether tumour-specific enhancement of vascular permeability by the particular tumour homing peptide, iRGD, which carries dual function of binding to integrin receptors overexpressed in the tumour vasculature and is known to promote extravasation via neuropilin-1 receptor upon site-specific cleavage, might be useful to improve blood-based tumour detection by inducing a yet unrecognised vice versa tumour-to-blood transport. To detect an iRGD-induced tumour-to-blood transport, we examined the effect of intravenously injected iRGD on blood levels of α-fetoprotein (AFP) and autotaxin in several mouse models of hepatocellular carcinoma (HCC) or in mice with chronic liver injury without HCC, and on prostate-specific antigen (PSA) levels in mice with prostate cancer. Intravenously injected iRGD rapidly and robustly elevated the blood levels of AFP in several mouse models of HCC, but not in mice with chronic liver injury. The effect was primarily seen in mice with small tumours and normal basal blood AFP levels, was attenuated by an anti-neuropilin-1 antibody, and depended on the concentration gradient between tumour and blood. iRGD treatment was also able to increase blood levels of autotaxin in HCC mice, and of PSA in mice with prostate cancer. We conclude that iRGD induces a tumour-to-blood transport in a tumour-specific fashion that has potential of improving diagnosis of early stage cancer. Deutsche Krebshilfe, DKTK, LOEWE-Frankfurt Cancer Institute.

Membrane-induced 2D phase separation of the focal adhesion protein talin

Focal adhesions form liquid-like assemblies around activated integrin receptors at the plasma membrane. How they achieve their flexible properties is not well understood. Here, we use recombinant focal adhesion proteins to reconstitute the core structural machinery in vitro. We observe liquid-liquid phase separation of the core focal adhesion proteins talin and vinculin for a spectrum of conditions and interaction partners. Intriguingly, we show that binding to PI(4,5)P2-containing membranes triggers phase separation of these proteins on the membrane surface, which in turn induces the enrichment of integrin in the clusters. We suggest a mechanism by which 2-dimensional biomolecular condensates assemble on membranes from soluble proteins in the cytoplasm: lipid-binding triggers protein activation and thus, liquid-liquid phase separation of these membrane-bound proteins. This could explain how early focal adhesions maintain a structured and force-resistant organization into the cytoplasm, while still being highly dynamic and able to quickly assemble and disassemble.

Tuning of Ultrasmall Gold Nanoparticles Surface Properties Affect Their Biological Fate

AbstractUltrasmall nanoparticles of 10 nm or less in size have been shown to have great potential in the biomedical field due to their high surface area and strong tissue penetration. Their easy functionalization and unique behavior at the nanoscale, such as the reduced corona formation and lower liver retention allow them to be a potential tool for precision targeting. In this study, PEGylated ultrasmall gold nanoparticles (GNPs) with a 4 nm core size are developed. They are functionalized with the cyclic RGD (cRGD) targeting peptide, which provides high binding affinity toward αVβ3 integrin receptor, often overexpressed in solid tumors. Further evidence is presented that cRGD functionalized GNPs partially escape lysosomes while penetrating deeper into the liver parenchyma. These particles provide a potential future strategy for specific αVβ3 integrin targeting.

Fibronectin/α5 Integrin Contribute to Hypertension-Associated Arterial Ageing and Calcification through Affecting BMP2/MGP Imbalance and Enhancing Vascular Smooth Muscle Cell Phenotypic Transformation

Introduction: Hypertension can accelerate and aggravate the process of arterial ageing and calcification. However, the mechanism behind has yet to be well elucidated. Methods: Here, we monitored the dynamic changes of fibronectin (FN)/α5 integrin, bone morphogenetic protein 2/matrix Gla protein (BMP2/MGP), and Runx2 in the aorta of spontaneously hypertensive rats (SHRs) and thoracic aortic vascular smooth muscle cells (VSMCs), also the phenotypic transformation of VSMCs during the process of arterial ageing and calcification. Further, study on arterial ageing and calcification through antagonist experiments at the molecular level was explored. Results: We found extracellular FN and its α5 integrin receptor expressions were positively associated with arterial ageing and calcification in SHR during ageing, as well in VSMCs from SHR in vitro. Integrin receptor inhibitor of GRGDSP would delay this arterial ageing and calcification process. Moreover, the elevated FN and α5 integrin receptor expression evoked the disequilibrium of BMP2/MGP, where the expression of BMP2, a potent osteogenic inducer, increased while MGP, a calcification inhibitor, decreased. Furthermore, it was followed by the upregulation of Runx2 and the phenotypic transformation of VSMCs from the contractile phenotype into the osteoblast-like cells. Notably, BMP2 antagonist of rmNoggin was sufficient to ameliorate the ageing and calcification process of VSMCs and exogenous BMP2-adding accelerate and aggregate the process. Conclusion: Our study revealed that hypertension-associated arterial ageing and calcification might be a consequence that hypertension up-regulated FN and its high binding affinity integrin α5 receptor in the aortic wall, which in turn aggravated the imbalance of BMP2/MGP, promoted the transcription of Runx2, and induced the phenotypic transformation of VSMCs from the contractile phenotype into the osteoblast-like cells. Our study would provide insights into hypertension-associated arterial ageing and calcification and shed new light on the control of arterial calcification, especially for those with hypertension.

Anticancer Properties of Different Varieties of Date Palm (Phoenix dactylifera L.) Leaf Extracts in Human Tumor Cells: a Comparative Study.

Plant polyphenols are nutraceutical components with relevant biological effects on human health. They act against development of several diseases including cancer. In this study, the methanolic extracts of four date palm Phoenix dactylifera leaves (Deglet Noor (DN), Barhee (B), Khalas (KS) and Khunezi (KZ)) collected from south Tunisia were preliminary analyzed for their effects against U87 (human glioblastoma) and MDA-MB-231 (human breast cancer) cell line development. Results showed that Barhee extract (30μg/mL) was the most efficient to reduce the growth of both tumor cells to about 40% (p < 0.05) without inducing cytotoxicity. Significantly, KS, KZ, DN and B extracts (30μg/mL) decreased MDA-MB-231 and U87 cell adhesion towards fibrinogen and fibronectin. Using integrin blocking antibodies, leaf extracts competitively decreased human glioblastoma cell attachment to immobilized antibodies by interfering to αvβ3 and α5β1 integrin receptors. At the same concentration, extracts decreased MDA-MB-23 and U87 cell migration performed with wound healing assay. Particularly, Barhee and Deglet Noor leaf extracts (30μg/mL) significantly reduced U87 cell invasion by 52.92% (p < 0.01) and 74.56% (p < 0.01), respectively. Collegially, our findings revealed beneficial proprieties of four varieties of date palm leaf especially those displayed by DN and B extracts that may serve as active candidates against human glioblastoma and breast cancer progression.

Systematic proteomics analysis revealed different expression of laminin interaction proteins in breast cancer: lower in luminal subtype and higher in claudin-low subtype.

Breast cancer is a major public health concern. Proteomics enables identification of proteins with aberrant properties. Here, we identified proteins with abnormal expression levels in breast cancer tissues and systematically analyzed and validated the data to locate potential diagnostic and therapeutic targets. Protein expression level in breast cancer tissues and para-carcinoma tissues were detected by Isobaric Tags for Relative and Absolute Quantification (iTRAQ) technology and further screened through Gene Expression Profiling Interactive Analysis (GEPIA) database. Cellular components, protein domain and Reactome pathway analysis were performed to screen functional targets. Abnormal expression levels of functional targets were validated by Oncomine database, quantitative real time polymerase chain reaction (qRT-PCR) and proteomics detection. Protein correlation analysis was performed to explain the abnormal expression levels of potential targets in breast cancer. Overall, 207 and 207 proteins were up- and down-regulated, respectively, in breast cancer tissues, and approximately 50% were also detected in the GEPIA database. The overlapping proteins were mainly extracellular proteins containing epidermal growth factor-like domain in leukocyte adhesion molecule (EGF-Lam) domain and enriched in laminin interaction pathway. Moreover, the downregulated laminin interaction proteins could be functional targets, which were also validated through Oncomine-Richardson and Oncomine-Curtis database. However, the lower expression level of laminin interaction proteins only fit for luminal breast cancer cells with no or low metastasis ability because the proteins achieved higher expression level in more invasive claudin-low breast cancer cells. In addition, when compared with corresponding in situ carcinoma tissues, above-mentioned proteins also showed higher expression levels in invasive carcinoma tissues. Finally, we have revealed the negative correlation between the laminin interaction proteins and the claudins. The laminin interaction protein, especially for laminins with β1 and γ1 subunits and their integrin receptors with α1 and α6 subunits, showed lower expression levels in luminal breast cancer with no or lower metastatic ability, but showed higher expression levels in claudin-low breast cancer with higher metastatic ability; and their higher expression could be related to the low claudin expression.

Stem-Cell-Regenerative and Protective Effects of Squid (Symplectoteuthis oualaniensis) Skin Collagen Peptides against H2O2-Induced Fibroblast Injury.

Recently, there has been a growing interest in collagen peptides derived from marine sources for their notable ability to protect skin cells against apoptosis induced by oxidants. Therefore, the current study aimed to investigate the fundamental properties of collagen peptides, including their physicochemical, thermal, structural, stem-cell-regenerative, and skin-cell-protective effects, in comparison to commercial collagen peptides. The acid-soluble (ASC) and pepsin-soluble (PSC) collagens exhibited three distinct bands on SDS-PAGE, namely α (α1 and α2), β, and γ chains, confirming a type I pattern. The thermal profiles obtained from TG and DSC analyses confirmed the denaturation of PSC and ASC at temperatures ranging from 51.94 to 56.4 °C and from 52.07 to 56.53 °C, respectively. The purified collagen peptides were analyzed using SDS-PAGE and MALDI-TOF mass spectrometry, revealing a mass range of 900-15,000 Da. Furthermore, the de novo peptide sequence analysis confirmed the presence of the Gly-X-Y repeating sequence in collagen peptides. Collagen peptide treatments significantly enhanced HFF-1 cell proliferation and migration compared to the control group. ELISA results confirmed the potential interactions between collagen peptides and HFF-1 cells through α2β1, α10β1, and α11β1 integrin receptors. Notably, collagen peptide treatment effectively restored the proliferation of HFF-1 cells damaged by H2O2. Consequently, the advantageous characteristics of squid skin collagen peptides highlight their promising role in regenerative medicine.

Inhibition of alphaV integrins with cilengitide downregulates in vitro pro-fibrotic events in aortic smooth muscle cells and limits in vivo thoracic aortic aneurysm progression

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Italian Ministry of Health "Ricerca Finalizzata - Giovani Ricercatori" Background The ‘greatest artery’, i.e., the aorta, can be affected by either acute conditions or chronic disorders, such as the aneurysm, which is the segmental aortic dilation with at least 50% increase in the normal diameter. Aortic dilation can often lead to lethal events, such as aortic dissection and/or vessel rupture. The aneurysms occurring in the thoracic trait of the aorta (TAA) may depend on comorbidities, such as hypertension (i.e., sporadic non-syndromic TAA, sTAA), or on genetically determined conditions, such as Marfan syndrome (MFS). To date, the surgical replacement of aorta represents the most effective therapeutic approach in all TAA patients, since an etiologic pharmacological treatment is still lacking. Thus, the identification of novel disease targets is mandatory. Integrin receptors are key players in cellular mechanotransduction responses. Specifically, integrins containing αV subunit have been described as important mechanotransduction mediators and responsible for pro-fibrotic molecular events. Recently, it has been reported the effectiveness of a broad αV integrin inhibitor in limiting the TAA progression by acting on FAK/AKT/mTORC signaling pathway in a MFS in vivo model. However, the specific αV integrin subgroup directly involved in this detrimental process is yet not known. Purpose We evaluated the effectiveness of a specific αVβ3 and αVβ5 integrin inhibitor (i.e., cilengitide) in downregulating pro-fibrotic in vitro events on vascular smooth muscle cells (VSMC) isolated from TAA patients’ aortic wall, and in limiting in vivo thoracic aortic dilation on a MFS mice model. Results By comparison with healthy control (HC)-VSMC, both sTAA-VSMC and MFS-VSMC showed higher levels of αVβ3 and αVβ5 integrins as well as pro-fibrotic collagen I. The in vitro inhibition of these two αV integrins mediated by cilengitide resulted more effective in collagen I downregulation when compared with GLPG0187 (pan αV inhibitor). Noteworthy, the cilengitide-mediated αV integrin inhibition consistently tempered pro-fibrotic phenotype of sTAA- and MFS-VSMC by directly acting on both canonical (p-SMAD2/3) and non-canonical (p-ERK1/2) TGF-β downstream pathways and by downregulating mechanotransduction mediators (e.g., RhoA GTPase, ROCK1, talin-1). The 2D-echocardiographic results on MFS mice treated with cilengitide confirmed the greater effectiveness of this drug in limiting the TAA progression, in comparison not only with vehicle but also with the broad integrin inhibitor GLPG0187. Conclusion Altogether, our results improve the know-how on αV integrin inhibition, highlighting the role of cilengitide as potential target for future therapeutic approaches to limit TAA progression, both in sporadic and in MFS patients.

Deep proteome profiling of reticulated platelets in patients with chronic coronary syndrome using mass cytometry

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Else Körner Fresenius Stiftung (EKFS) Background Immature or reticulated platelets (RPs), a subgroup of larger and RNA-rich platelets, which are newly released by megakaryocytes from the bone marrow into blood, have gained rising interest because of the further decryption of RPs’ prothrombotic biology and the potential as biomarkers for predicting adverse events in cardiovascular disease. Purpose The biological features of RPs in patients with chronic coronary syndrome (CCS) are largely unknown. With this project, we aimed to perform the first high resolution proteomic characterisation of reticulated platelets using mass cytometry(CyTOF). Method Two samples of thrombocytes from peripheral blood (non-stimulated, TRAP-stimulated) were prepared from each patient with a custom-made CyTOF-antibody panel of 20 antibodies targeting essential transmembrane proteins(anti-CD9, anti-CD29, anti-CD31, anti-CD36-, anti-CD40, anti-CD41, anti-CD42a, anti-CD42b-, anti-CD47, anti-CD61, anti-CD62P-, anti-CD63, anti-CD69, anti-CD107a, anti-CD154, anti-GPVI, anti-GPIIbIIa complex, anti-Par1, anti-PEAR-1 and the negative control anti-CD3 coupled with different metal isotopes). RPs and mature platelets (MPs) were detected based on their RNA content. Earth mover’s distance (EMD) was assessed as a measure of differential expression. Results CyTOF detected an upregulation of important transmembrane receptors in RPs compared to MPs in quiescent platelets (Fig.1A and B). The collagen receptor GPVI (EMD 33.95), the collagen integrin receptor unit ITGB1 (EMD 32.37), the adhesion protein CD9 (EMD 31.78) and the von Willebrand receptor unit CD42b (EMD 30.08) were upregulated in RPs which all serve as crucial regulators of platelet activation and aggregation. RPs show higher median signal intensities of typical activation markers compared to MPs upon TRAP stimulation (Fig.1C). Conclusion We provide the first proteomic characterization of RPs’ pro-thrombotic profile offering the first biomolecular explanation of their hyperactivity and detrimental role in CCS patients.

Hierarchical Chiral Calcium Silicate Hydrate Films Promote Vascularization for Tendon-to-Bone Healing.

Revascularization after rotator cuff repair is crucial for tendon-to-bone healing. The chirality of materials has been reported to influence their performance in tissue repair. However, data on the use of chiral structures to optimize biomaterials as a revascularization strategy remain scarce. Here, calcium silicate hydrate (CSO) films with hierarchical chirality on the atomic to micrometer scale are developed. Interestingly, levorotatory CSO (L-CSO) films promote the migration and angiogenesis of endothelial cells, whereas dextral and racemic CSO films do not induce the same effects. Molecular analysis demonstrates that L-chirality can be recognized by integrin receptors and leads to the formation of focal adhesion, which activates mechanosensitive ion channel transient receptor potential vanilloid 4 to conduct Ca2+ influx. Consequently, the phosphorylation of serum response factor is biased by Ca2+ influx to promote the vascular endothelial growth factor receptor 2 signaling pathway, resulting in enhanced angiogenesis. After implanted in a rat rotator cuff tear model, L-CSO films strongly enhance vascularization at the enthesis, promoting collagen maturation, increasing bone and fibrocartilage formation, and eventually improving the biomechanical strength. This study reveals the mechanism through which chirality influences angiogenesis in endothelial cells and provides a critical theoretical foundation for the clinical application of chiral biomaterials.

Early Stages of Ex Vivo Collagen Glycation Disrupt the Cellular Interaction and Its Remodeling by Mesenchymal Stem Cells-Morphological and Biochemical Evidence.

Mesenchymal stem cells (MSCs), pivotal for tissue repair, utilize collagen to restore structural integrity in damaged tissue, preserving its organization through concomitant remodeling. The non-enzymatic glycation of collagen potentially compromises MSC communication, particularly upon advancing the process, underlying various pathologies such as late-stage diabetic complications and aging. However, an understanding of the impact of early-stage collagen glycation on MSC interaction is lacking. This study examines the fate of in vitro glycated rat tail collagen (RTC) upon exposure to glucose for 1 or 5 days in contact with MSCs. Utilizing human adipose tissue-derived MSCs (ADMSCs), we demonstrate their significantly altered interaction with glycated collagen, characterized morphologically by reduced cell spreading, diminished focal adhesions formation, and attenuated development of the actin cytoskeleton. The morphological findings were confirmed by ImageJ 1.54g morphometric analysis with the most significant drop in the cell spreading area (CSA), from 246.8 μm2 for the native collagen to 216.8 μm2 and 163.7 μm2 for glycated ones, for 1 day and 5 days, respectively, and a similar trend was observed for cell perimeter 112.9 μm vs. 95.1 μm and 86.2 μm, respectively. These data suggest impaired recognition of early glycated collagen by integrin receptors. Moreover, they coincide with the reduced fibril-like reorganization of adsorbed FITC-collagen (indicating impaired remodeling) and a presumed decreased sensitivity to proteases. Indeed, confirmatory assays reveal diminished FITC-collagen degradation for glycated samples at 1 day and 5 days by attached cells (22.8 and 30.4%) and reduced proteolysis upon exogenous collagenase addition (24.5 and 40.4%) in a cell-free system, respectively. The mechanisms behind these effects remain uncertain, although differential scanning calorimetry confirms subtle structural/thermodynamic changes in glycated collagen.

Peptide-Directed Synthesis of Aggregation-Induced Emission Enhancement-Active Gold Nanoclusters for Single- and Two-Photon Imaging of Lysosome and Expressed αvβ3 Integrin Receptors.

This study explores the synthesis and characterization of aggregation-induced emission enhancement (AIEE)-active gold nanoclusters (AuNCs), focusing on their near-infrared luminescence properties and potential applications in biological imaging. These AIEE-active AuNCs were synthesized via the NaBH4-mediated reduction of HAuCl4 in the presence of peptides. We systematically investigated the influence of the peptide sequence on the optical features of the AuNCs, highlighting the role of glutamic acid in enhancing their quantum yield (QY). Among the synthesized peptide-stabilized AuNCs, EECEE-stabilized AuNCs exhibited the maximum QY and a pronounced AIEE effect at pH 5.0, making them suitable for the luminescence imaging of intracellular lysosomes. The AIEE characteristic of the EECEE-stabilized AuNCs was demonstrated through examinations using transmission electron microscopy, dynamic light scattering, zeta potential analysis, and single-particle imaging. The formation of the EECEE-stabilized AuNCs was confirmed by size-exclusion chromatography and mass spectrometry. Spectroscopic and electrochemical examinations uncover the formation process of EECEE-stabilized AuNCs, comprising EECEE-mediated reduction, NaBH4-induced nucleation, complex aggregation, and subsequent cluster growth. Furthermore, we demonstrated the utility of these AuNCs as luminescent probes for intracellular lysosomal imaging, leveraging their pH-responsive AIEE behavior. Additionally, cyclic arginylglycylaspartic acid (RGD)-modified AIEE dots, derived from cyclic RGD-linked peptide-induced aggregation of EECEE-stabilized AuNCs, were developed for single- and two-photon luminescence imaging of αvβ3 integrin receptor-positive cancer cells.

Overcoming Nutrient Stress: Integrin αvβ3-Driven Metabolic Adaptation Supports Tumor Initiation.

Nutrient stress accompanies several stages of tumor progression, including metastasis formation. Metabolic reprogramming is a hallmark of cancer, and it has been associated with stress tolerance and anchorage-independent cell survival. Adaptive responses are required to support cancer cell survival under these conditions. In this issue of Cancer Research, Nam and colleagues showed that the extracellular matrix (ECM) receptor integrin β3 was upregulated in lung cancer cells in response to nutrient starvation, resulting in increased cell survival that was independent from ECM binding. Delving into the molecular mechanisms responsible for this, the authors found that integrin β3 promoted glutamine metabolism and oxidative phosphorylation (OXPHOS) by activating a Src/AMPK/PGC1α signaling pathway. Importantly, in vivo experiments confirmed that OXPHOS inhibition suppressed tumor initiation in an orthotopic model of lung cancer, while β3 knockout completely abrogated tumor initiation. These observations indicate that targeting signaling pathways downstream of αvβ3 could represent a promising therapeutic avenue to prevent lung cancer progression and metastasis. See related article by Nam et al., p. 1630.

Liquid crystal monomers disrupt photoreceptor patterning of zebrafish larvae via thyroid hormone signaling

Liquid crystal monomers (LCMs) are the raw material for liquid crystal displays, and their use is steadily increasing in electronic products. Recently, LCMs have been reported to be novel endocrine disrupting chemicals, however, the mechanisms underlying their potential for thyroid hormone disruption and visual toxicity are not well understood. In this study, six widely used fluorinated LCMs (FLCMs) were selected to determine putative mechanisms underlying FLCM-induced toxicity to the zebrafish thyroid and visual systems. Exposure to FLCMs caused damage to retinal structures and reduced cell density of ganglion cell layer, inner nuclear layer, and photoreceptor layer approximately 12.6–46.1%. Exposure to FLCMs also disrupted thyroid hormone levels and perturbed the hypothalamic-pituitary-thyroid axis by affecting key enzymes and protein in zebrafish larvae. A thyroid hormone-dependent GH3 cell viability assay supported the hypothesis that FLCMs act as thyroid hormone disrupting chemicals. It was also determined that FLCMs containing aliphatic ring structures may have a higher potential for T3 antagonism compared to FLCMs without an aliphatic ring. Molecular docking in silico suggested that FLCMs may affect biological functions of thyroxine binding globulin, membrane receptor integrin, and thyroid receptor beta. Lastly, the visual motor response of zebrafish in red- and green-light was significantly inhibited following exposure to FLCMs. Taken together, we demonstrate that FLCMs can act as thyroid hormone disruptors to induce visual dysfunction in zebrafish via several molecular mechanisms.

Looking both ways: Electroactive biomaterials with bidirectional implications for dynamic cell-material crosstalk.

Cells exist in natural, dynamic microenvironmental niches that facilitate biological responses to external physicochemical cues such as mechanical and electrical stimuli. For excitable cells, exogenous electrical cues are of interest due to their ability to stimulate or regulate cellular behavior via cascade signaling involving ion channels, gap junctions, and integrin receptors across the membrane. In recent years, conductive biomaterials have been demonstrated to influence or record these electrosensitive biological processes whereby the primary design criterion is to achieve seamless cell-material integration. As such, currently available bioelectronic materials are predominantly engineered toward achieving high-performing devices while maintaining the ability to recapitulate the local excitable cell/tissue microenvironment. However, such reports rarely address the dynamic signal coupling or exchange that occurs at the biotic-abiotic interface, as well as the distinction between the ionic transport involved in natural biological process and the electronic (or mixed ionic/electronic) conduction commonly responsible for bioelectronic systems. In this review, we highlight current literature reports that offer platforms capable of bidirectional signal exchange at the biotic-abiotic interface with excitable cell types, along with the design criteria for such biomaterials. Furthermore, insights on current materials not yet explored for biointerfacing or bioelectronics that have potential for bidirectional applications are also provided. Finally, we offer perspectives aimed at bringing attention to the coupling of the signals delivered by synthetic material to natural biological conduction mechanisms, areas of improvement regarding characterizing biotic-abiotic crosstalk, as well as the dynamic nature of this exchange, to be taken into consideration for material/device design consideration for next-generation bioelectronic systems.