ITGB1 Expression Research Articles

Integrin β1 regulates the perineural invasion and radioresistance of oral squamous carcinoma cells by modulating cancer cell stemness

Perineural invasion and radioresistance are the main determinants of treatment outcomes in oral squamous cell carcinoma (OSCC), but the exact mechanism is still unknown. We conducted an in vitro experiment to evaluate the role of integrin β1 (ITGB1) in the perineural invasion, radioresistance, and tumor aggressiveness of OSCC. Two OSCC cell lines (SCC25, SCC15) and radiation-induced radioresistant OSCC cell lines were used in this study. The expression of ITGB1 was compared between control radiosensitive and radioresistant OSCC cell lines. ITGB1 was inhibited by small hairpin RNA, and then the adhesion to neuronal cells, responsiveness to radiation, and aggressiveness of both OSCC cell lines were evaluated. Expression of ITGB1 and adhesion to neuronal cells were increased in radioresistant OSCC compared with control radiosensitive OSCC, and increased ITGB1 expression was more prominent in cancer stem cell-like cells. When the expression of ITGB1 was inhibited, the adhesion to neuronal cells, resistance to radiation, and invasion and migration of radioresistant OSCC were significantly reduced. Moreover, the expression of cancer stem cell markers and size of spheroid formations were also significantly attenuated by inhibiting ITGB1. These findings suggest that ITGB1 may be a significant contributor to perineural invasion and the maintenance of radioresistance in OSCC cells, and is associated with cancer stem cell-like cells. Furthermore, our results suggest a possible relationship between perineural invasion and radioresistance of OSCC. More detailed research is warranted to evaluate the role of ITGB1 as a novel emerging therapeutic target for radioresistant OSCC.

Liraglutide intervention improves high-glucose-induced reactive gliosis of Müller cells and ECM dysregulation

Reactive gliosis of Müller cells plays an important role in the pathogenesis of diabetic retinopathy (DR). Liraglutide, a glucagon-like peptide-1 receptor (GLP-1R) agonist, has been shown to improve DR by inhibiting reactive gliosis. However, the mechanism of inhibition has yet to be elucidated. This study investigated the effects of liraglutide on Müller glia reactivity in the early stages of DR and the underlying mechanisms. Proteomics combined with bioinformatics analysis, HE staining, and immunofluorescence staining revealed ganglion cell loss, reactive gliosis of Müller cells, and extracellular matrix (ECM) imbalance in rats with early stages of DR. High glucose (HG) exposure up-regulated GFAP and TNF-α expression and down-regulated ITGB1 expression and FN1 content in extracellular fluid in rMC1 cells, thereby promoting reactive gliosis. GLP-1R knockdown and HG+DAPT inhibition experiments show that liraglutide balances ECM levels by inhibiting activation of the Notch1/Hes1 pathway and ameliorates high-glucose-induced Müller glia reactivity. Thus, the study provides new targets and ideas for improvement of DR in early stages.

Targeting ITGB4/SOX2-driven lung cancer stem cells using proteasome inhibitors

This study investigates the role of integrin β4 (ITGB4) and stemness-associated factor SOX2 in platinum resistance in lung squamous cell carcinoma (LUSC). The expression of SOX2 and ITGB4 is found to be high in all LUSC subtypes, but the impact of ITGB4 expression on overall patient survival varies by subtype. Cancer stem cells (CSCs) isolated from LUSC patients were found to be resistant to cisplatin, but knocking down ITGB4 or SOX2 sensitized them to cisplatin. Carfilzomib (CFZ) synergized with cisplatin and suppressed CSC growth by inhibiting ITGB4 and SOX2 expression. Additionally, CFZ was found to inhibit SOX2 expression epigenetically by inhibiting histone acetylation at the SOX2 promoter site. CFZ also suppressed the growth of SOX2-dependent small cell lung cancer cells invitro and invivo. The study highlights the unique function of CFZ as a transcriptional suppressor of SOX2, independent of its proteasome inhibitory function.

LSD1 knockdown confers protection against osteoclast formation by reducing histone 3 lysine 9 monomethylation and dimethylation in ITGB3 promoter

ITGB3, an osteoclast marker, is involved in osteoclast formation. Nevertheless, its related mechanism remains poorly characterized. Herein, this study examines the mechanisms affecting osteoclast formation with the involvement of ITGB3. Osteoclast formation was induced with macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-kappa B ligand (RANKL), followed by measurement of the mRNA and protein expression of ITGB3 and LSD1. After gain- and loss-of-function assays, cell viability and the expression of osteoclast marker genes (NFATc1, ACP5, and CTSK) were assessed, and osteoclast formation was evaluated with TRAP staining. ChIP assays were used to examine histone 3 lysine 9 (H3K9) monomethylation (H3K9me1) and H3K9 dimethylation (H3K9me2) modifications and LSD1 protein enrichment in the ITGB3 promoter. During osteoclast formation, ITGB3 and LSD1 were gradually augmented. Knockdown of LSD1 or ITGB3 curbed cell viability, the expression of osteoclast marker genes, and osteoclast formation. Moreover, overexpression of ITGB3 nullified the suppressive impact of LSD1 knockdown on osteoclast formation. Mechanistically, LSD1 promoted ITGB3 expression by reducing H3K9 levels in the ITGB3 promoter. LSD1 enhanced ITGB3 expression by decreasing H3K9me1 and H3K9me2 levels in ITGB3 promoter to boost osteoclast formation.

Identification of a Favorable Prognostic Subgroup in Oral Squamous Cell Carcinoma: Characterization of ITGB4/PD-L1high with CD8/PD-1high

Integrin β4 (ITGB4) is a member of the integrin family, which plays a crucial role in mediating cell adhesion to the extracellular matrix. Recent studies have demonstrated that ITGB4 is involved in tumorigenesis and metastasis during the development of cancer. However, the role of ITGB4 in oral squamous cell carcinoma (OSCC) remains unclear. A Multiplex immunohistochemistry (OPAL™, mIHC) assay was employed to stain ITGB4, ALDH1, PD-L1, cytokeratin (CK), CD8 and PD-1 in a human OSCC tissue microarray, containing 26 normal oral epithelium samples, 21 oral epithelium dysplasia samples and 76 OSCC samples. The expression pattern and clinicopathological characteristics of ITGB4 were analyzed and compared with those of PD-1, PD-L1, ALDH1 and CD8. The correlation between subgroups of tumor cells, including ITGB4+PD-L1+ and ITGB4+ALDH1+, and subgroups of T cells, including CD8+ and CD8+PD-1+, was evaluated using two-tailed Pearson’s statistics. A Kaplan–Meier curve was built, and a log-rank test was performed to analyze the survival rate of different subgroups. The mIHC staining results show that ITGB4 was mostly expressed in the tumor cells, with a significant increase in the OSCC specimens compared with normal oral epithelium and oral epithelium dysplasia. The paired analysis, conducted between the OSCC tumor tissue and normal paracancer mucosa, confirmed the results. The study further revealed that ITGB4+PD-L1+ cancer cells, but not ITGB4+ALDH1+ cancer cells, were significantly associated with the infiltration of CD8+ T cells (positivity p = 0.005, positive number p = 0.03). Additionally, ITGB4+PD-L1+ tumor cells were positively correlated with CD8+PD-1+ T cells (positivity p = 0.02, positive number p = 0.03). Most intriguingly, the subgroup of ITGB4/PD-L1high with CD8/PD-1high displayed the best prognosis compared with the other considered subgroups. The results show that the expression of ITGB4 was increased in OSCC compared with normal oral mucosa. Furthermore, a specific subgroup with high levels of expression of ITGB4/PD-L1 and CD8/PD-1 was found to have a relatively better prognosis compared with the other subgroups. Ultimately, this study sheds light on the potential role of ITGB4 in OSCC and provides a basis for further investigation.

Identification and Validation of Anoikis-Related Signatures for Predicting Prognosis in Lung Adenocarcinoma with Machine Learning.

Lung adenocarcinoma (LUAD) is an aggressive cancer that has an extremely poor prognosis. As well as facilitating the detachment of cancer cells from the primary tumor site, anoikis plays an important role in cancer metastasis. Few studies to date, however, have examined the role of anoikis in LUAD, in patient prognosis. A total of 316 anoikis-related genes (ANRGs) integrated from Genecards and Harmonizome portals. LUAD transcriptome data were retrieved from the Genotype-Tissue Expression Project (GEO) and The Cancer Genome Atlas (TCGA). Anoikis-related prognostic genes (ANRGs) were primarily screened by univariate Cox regression. All ANRGs were included in the Least Absolute Shrinkage and Selection Operator (LASSO) Cox regression model to construct the powerful prognostic signature. This signature was validated and assessed using the Kaplan-Meier method as well as univariate and multivariate Cox regression analyses. Anoikis-related regulators of risk score were identified using a XG-boost machine learning model. The expression of ITGB4 protein was examined in a ZhengZhou University (ZZU) tissue cohort by immunohistochemistry, and the potential mechanisms of action of ITGB4 in LUAD were explored by GO, KEGG, and ingenuity pathway analyses and by GSEA. A risk score signature was constructed based on eight ANRGs, with high risk scores found to closely correlate with unfavorable clinical features. ITGB4 expression may be associated with 5-year over survival, with immunohistochemistry showed that the expression of ITGB4 was higher in LUAD than in nontumor tissues. Enrichment analysis suggested that ITGB4 may promote LUAD development by targeting E2F, MYC, and oxidative phosphorylation signaling pathways. Our anoikis-related signature from RNA-seq data may be a novel prognostic biomarker in patients with LUAD. It may help physicians develop personalized LUAD treatments in clinical practice. Moreover, ITGB4 may affect the development of LUAD through the oxidative phosphorylation pathway.

USP18 promotes endometrial receptivity via the JAK/STAT1 and the ISGylation pathway.

Interferon-tau (IFNT), a pregnancy recognition signal in ruminants, promotes the establishment of endometrial receptivity by inducing the expression of interferon-stimulated genes (ISGs) via the Janus kinase/signal transducer and activator of transcription (JAK/STATs) signaling pathway. However, the precise mechanisms remain largely unknown. Ubiquitin-specific protease 18 (USP18) acts specifically on the ISGylation modification system to exert deubiquitination and participates in the regulation of the type I IFN signaling pathway. The purpose of this study was to determine the role and mechanism of USP18 on endometrial receptivity in goat. USP18 was mainly localized in the uterine luminal and glandular epithelium, and its expression levels were significantly increased from days 5-18 of early pregnancy. Progesterone (P4), estradiol (E2), and IFNT significantly stimulated USP18 expression in goat endometrial epithelial cells (gEECs) cultured invitro. Meanwhile, the markers of endometrial receptivity HOXA11, ITGB1, ITGB3, and ITGB5 were significantly upregulated after USP18 overexpression in gEECs. However, USP18 interference significantly inhibited the expression of HOXA10, ITGB1, ITGB3, and ITGB5 in gEECs. In addition, both the phosphorylation levels of STAT1 and the expression of ISGylation-modified proteins were significantly increased after USP18 silencing in gEECs. Furthermore, pretreatment with the STAT1 inhibitor Fludara markedly restored the effect of USP18 interference in gEECs. In summary, USP18 may play an important role in promoting goat endometrial receptivity by regulating the JAK/STAT1 pathway and ISGylation.

Reduced expression of Il10, Stat3, Hoxa10, and Itgb3 in the embryo implantation site of rat model with prenatal androgen-induced polycystic ovary syndrome.

Impaired implantation due to the reduced endometrial receptivity considers an etiology for infertility in polycystic ovary syndrome (PCOS). In this context, we aimed to compare the expression of interleukin 10 (Il10), homeobox A10 (Hoxa10), signal transducer and activator of transcription 3 (Stat3), and β3-integrin (Itgb3) in the embryo implantation site of a prenatally-androgenized rat model of PCOS before and during gestation. PCOS rat model was created by the injection of testosterone prenatally. The uterine tissues were collected before pregnancy (day 0) and on days 0.5, 4.5, 5.5, and 8.5 of gestation in the PCOS rat model and controls (n=6; each group). RNA was extracted from the uterine samples and reverse transcribed to cDNA. Expression levels of Il10, Stat3, Hoxa10, and Itgb3 were measured using SYBR Green real-time RT-PCR and compared between the two groups. PCOS rats showed decreased expression levels of the Il10 on day 8.5 compared to control rats. The mRNA levels of Hoxa10, Itgb3, and Stat3 were significantly decreased in the PCOS group on day 0 as well as on days 0.5, 4.5, 5.5, and 8.5 for Hoxa10, Itgb3, and Stat3. The decreased gene expression of Il10, Hoxa10, Stat3, and Itgb3 in the PCOS rat model indicates the importance of the Il10 signaling axis as one of the possible disrupted mechanisms of endometrial receptivity in PCOS.

Novel ITGB6 variants cause hypoplastic-hypomineralized amelogenesis imperfecta and taurodontism: characterization of tooth phenotype and review of literature

ObjectivesTo characterize phenotype and genotype of amelogenesis imperfecta (AI) in a Thai patient, and review of literature.Materials and methodsVariants were identified using trio-exome and Sanger sequencing. The ITGB6 protein level in patient’s gingival cells was measured. The patient’s deciduous first molar was investigated for surface roughness, mineral density, microhardness, mineral composition, and ultrastructure.ResultsThe patient exhibited hypoplastic-hypomineralized AI, taurodontism, and periodontal inflammation. Exome sequencing identified the novel compound heterozygous ITGB6 mutation, a nonsense c.625 G > T, p.(Gly209*) inherited from mother and a splicing c.1661-3 C > G from father, indicating AI type IH. The ITGB6 level in patient cells was significantly reduced, compared with controls. Analyses of a patient’s tooth showed a significant increase in roughness while mineral density of enamel and microhardness of enamel and dentin were significantly reduced. In dentin, carbon was significantly decreased while calcium, phosphorus, and oxygen levels were significantly increased. Severely collapsed enamel rods and a gap in dentinoenamel junction were observed. Of six affected families and eight ITGB6 variants that have been reported, our patient was the only one with taurodontism.ConclusionWe report the hypoplasia/hypomineralization/taurodontism AI patient with disturbed tooth characteristics associated with the novel ITGB6 variants and reduced ITGB6 expression, expanding genotype, phenotype, and understanding of autosomal recessive AI.

Abstract 2927: ITGB6 ProTriTAC™, a protease-activated T cell engager prodrug targeting Integrin-β6 for the treatment of solid tumors

Abstract Integrin-β6 (ITGB6) is a transmembrane protein that partners exclusively with the integrin-αV subunit to form the integrin-αVβ6 heterodimer. Due to its roles in tissue remodeling and cancer pathogenesis, therapeutic targeting of integrin-αVβ6 has been investigated for both fibrotic diseases and cancers. ITGB6 is homogenously overexpressed in numerous solid tumor types, including breast, head and neck, pancreatic, and non-small cell lung cancers. Additionally, its expression is an unfavorable prognostic marker in multiple cancers. However, past attempts to therapeutically target ITGB6 have been hindered by its expression in several normal tissues, including bladder, skin, lung, kidney, and muscle. For example, clinical testing of an integrin αVβ6-targeting monoclonal antibody was discontinued reportedly due to toxicity concerns. To reduce on-target/off-tumor toxicities from ITGB6 expression in normal tissues and to enable safe targeting of ITGB6 in solid tumors, we developed an ITGB6-targeting ProTriTAC™ (Protease-activated Tri-specific T cell Activating Construct), a T cell engager prodrug which is engineered to be preferentially activated in tumor tissue. ProTriTACs consists of three humanized antibody-derived binding domains on a single polypeptide chain: anti-albumin for half-life extension, anti-CD3 for T cell engagement, and anti-target antigen for tumor cell engagement. The anti-albumin domain, bearing a masking moiety and a protease-cleavable linker, keeps the prodrug inert by inhibiting binding of the adjacent anti-CD3 domain to T cells. Cleavage of the linker by tumor-associated proteases removes the anti-albumin domain along with the masking moiety to reveal a potent active drug that directs T cell killing within the tumor. As designed, in vitro binding and functional assays show that the protease-activated T cell engager has >1000-fold improved binding to human T cells and ~100-fold improved T cell killing compared to the intact masked prodrug. In immunodeficient mice reconstituted with human T cells, anti-ITGB6 ProTriTAC demonstrated robust anti-tumor activity in multiple ITGB6-expressing xenograft tumor models, including HCC70 (breast), CAL27 (head and neck), and HPAFII (pancreatic), with complete tumor regression as low as 30 μg/kg. To determine the potential toxicity and toxicokinetic characteristics of anti-ITGB6 ProTriTAC, an intra-animal dose escalation was conducted in cynomolgus monkeys. Starting at 20 μg/kg with weekly 3-fold dose escalations, anti-ITGB6 ProTriTAC was well tolerated up to 540 μg/kg, which was the highest dose tested. The preclinical activity in rodent tumor models, coupled to its tolerability in cynomolgus monkeys, support the development of anti-ITGB6 ProTriTAC as a therapeutic in a broad range of ITGB6-expressing solid tumors. Citation Format: Regina J. Lin, Sony S. Rocha, Maria R. Dayao, Subramanian Thothathri, Raphaela Rose Banzon, Kevin J. Wright, Wade Aaron, Yinghua Xiao, Nick Bergo, Linh To, Mabel Bush, Manasi Barath, Yi Yang, Timothy Yu, Willis Kwong, Hubert Situ, Eric Bragg, Jessica O'Rear, Kevin Carlin, Stephen Yu, Maritza Solorio, Bryan Lemon, Richard Austin, Holger Wesche, S. Jack Lin. ITGB6 ProTriTAC™, a protease-activated T cell engager prodrug targeting Integrin-β6 for the treatment of solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2927.

N-TELOPEPTIDE DERIVED FROM TYPE II COLLAGEN UPREGULATES EXPRESSION OF BETA1 INTEGRIN AND MMPS IN CARTILAGE OF PATIENTS RECEIVING HIP OR KNEE ARTHROPLASTY

Previous studies showed that telo-peptides degraded from type II collagen, a type of collagen fragments, could induce cartilage damage in bovine stifle joints. We aim to investigate the role of integrins (ITGs) and matrix metalloproteinases (MMPs) in collagen fragment-induced human cartilage damage that is usually observed in osteoarthritis (OA). We hypothesized that N-telopeptide (NT) derived from type II collagen could up-regulate the expression of β1 integrin (ITGB1) and then MMPs that may lead to osteoarthritic cartilage damage.Human chondrocytes were isolated from femoral head or tibial plateau of patients receiving arthroplasty (N = 24). Primary chondrocyte cultures were either treated with 30 µM NT, or 30 µM scrambled NT (SN), or PBS, or left untreated for 24 hrs. Total proteins and RNAs were extracted for examination of expression of ITGB1 and MMPs-3&13 with Western blotting and quantitative real-time PCR.Compared to untreated or PBS treated chondrocytes, NT-treated chondrocytes expressed significantly higher levels of ITGB1 and MMPs-3&-13. However, SN also up-regulated expression of ITGB1 and MMP-13.ITGB1 and MMPs-3&-13 might mediate the catalytic effect of NT, a type of collagen fragments, on human cartilage damage that is a hallmark of OA.

Abstract 3531: Discrete extracellular matrix-secreting tumor cell subpopulations remodel the Ewing sarcoma tumor microenvironment to promote invasion

Abstract Ewing sarcoma (EwS) cells exist along a neuro-mesenchymal transcriptional continuum that is largely determined by transcriptional activity of the EWS::FLI1 fusion oncoprotein. Although EWS::FLI1-low state cells are more metastatic in experimental models, the role of these cells in established EwS tumor ecosystems is unknown. Here we have leveraged multimodal single-cell sequencing, spatial transcriptomics, and immunohistochemical profiling to characterize EwS cell subpopulations in cell lines, PDX models, and patient tumors. We identify CD73 as a marker of mesenchymal-high state EwS cells and show that CD73+ tumor cells share properties of experimentally-induced EWS::FLI1-low cells including altered cytoskeletons, enhanced migration and invasion, and increased expression of EWS::FLI1-repressed genes. However, CD73+ cells retain proliferative capacity and do not down-regulate the fusion or the EWS::FLI1-activated signature. Instead, CD73+ EwS cells selectively express a gene signature that is enriched in extracellular matrix (ECM) genes. Examination of xenograft and EwS patient biopsies confirms the existence of subpopulations of spatially and transcriptionally distinct ECM-producing tumor cells and shows them to be more prevalent in infiltrating regions. Importantly, the local tumor microenvironment (TME) surrounding these cells is rich in tumor-derived TNC, SPARC, and biglycan, ECM proteins that have been widely implicated in cancer metastatic progression. Significantly, CD73+ EwS cells also upregulate expression of ECM-sensing genes, including ITGA5 and ITGB1, suggesting potential to activate autocrine feedback loops. In support of this, interrupting outside-in ECM:integrin signaling cascades, through pharmacologic inhibition of FAK or SRC pathways, strongly reduces the invasive properties of CD73+ tumor spheroids in 3D collagen. Together these studies confirm the presence and functional importance of transcriptionally distinct tumor cell subpopulations in established EwS tumor ecosystems. In particular, our data reveal that highly mesenchymal tumor cells both generate and respond to pro-metastatic ECM proteins to support invasion. Studies are ongoing to determine the role of tumor cell-derived ECM on EwS progression and treatment resistance. Citation Format: Emma D. Wrenn, April A. Apfelbaum, Shireen Ganapathi, Erin R. Rudzinski, Xuemei Deng, Nicolas Garcia, Katherine Braun, Trisha Lipson, Shruti Bhise, Sami B. Kanaan, Olivia Waltner, Erika Newman, Scott Furlan, Elizabeth R. Lawlor. Discrete extracellular matrix-secreting tumor cell subpopulations remodel the Ewing sarcoma tumor microenvironment to promote invasion. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3531.

PD17-06 WILMS TUMOR: INSIGHTS INTO THE ROLE OF INTEGRINS DRIVING CANCER STEM CELLS SELF-RENEWAL VS. DIFFERENTIATION

You have accessJournal of UrologyCME1 Apr 2023PD17-06 WILMS TUMOR: INSIGHTS INTO THE ROLE OF INTEGRINS DRIVING CANCER STEM CELLS SELF-RENEWAL VS. DIFFERENTIATION Astgik Petrosyan, Valentina Villani, Paola Aguiari, Min Madhi, Roger De Filippo, Gregory Shackleford, Anat Erdreich-Epstein, Stefano Da Sacco, and Laura Perin Astgik PetrosyanAstgik Petrosyan More articles by this author , Valentina VillaniValentina Villani More articles by this author , Paola AguiariPaola Aguiari More articles by this author , Min MadhiMin Madhi More articles by this author , Roger De FilippoRoger De Filippo More articles by this author , Gregory ShacklefordGregory Shackleford More articles by this author , Anat Erdreich-EpsteinAnat Erdreich-Epstein More articles by this author , Stefano Da SaccoStefano Da Sacco More articles by this author , and Laura PerinLaura Perin More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003272.06AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Wilms Tumor (WT) is the most common pediatric renal cancer. Growing evidence links WT to aberrant nephrogenesis. While studies highlighted the genetic complexity of WT, little is known about the molecular mechanisms that regulate WT development. Here we report that uncommitted nephrogenic progenitors (NPs) expressing SIX2 and CITED1 (the master regulators of renal development) present characteristics of cancer stem cells (CSCs) and are the ones driving WT. We have also studied the role of integrins in these NPs in regulating WT development. METHODS: WT and human fetal kidney (hFK) samples were histologically analyzed, digested to single-cell suspension, incubated with Smartflare-probe to isolate SIX2+CITED1+ cells, and processed for RNA-seq, single-cell RNA-seq and spatial transcriptomics. Xenografts of WT-NPs and hFK-NPs were generated and tumor formation was assessed. Analyses of mechanisms that regulate self-renewal vs. differentiation were performed in vitro and in vivo. Knockdown with miREs against SIX2 and CITED1 was performed on WT-NPs and processed for RNA-seq. RESULTS: By comparing NPs from different WT subtypes and NPs from hFK we identified that cells expressing SIX2 and CITED1 fulfill CSC criteria, reliably recapitulating WT in transplantation studies. We showed that self-renewal vs. differentiation of SIX2+CITED1+ WT CSCs is regulated by the interplay between integrins ITGB1 and ITGB4. WT transplantation studies show that blocking ITGB1 or ITGB4 leads to higher number of SIX2+CITED1+ cells in the xenografts. Knockdown of SIX2 and CITED1 increased expression of kidney differentiation markers LHX1, WNT7B, PODXL, MECOM, reduced expression of nephrogenic markers MEOX1, TMEM100, EYA1, MAYFB, and increased expression of ITGB1, ITGB4, and LAMA5. CONCLUSIONS: These studies define SIX2+CITED1+ cells as the nephrogenic CSCs of WT, where ITGB1 and ITGB4 interplay may play a role in self-renewal vs. differentiation and serve as a potential target for new strategies to treat WT. Source of Funding: GOFARR Fund © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e497 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Astgik Petrosyan More articles by this author Valentina Villani More articles by this author Paola Aguiari More articles by this author Min Madhi More articles by this author Roger De Filippo More articles by this author Gregory Shackleford More articles by this author Anat Erdreich-Epstein More articles by this author Stefano Da Sacco More articles by this author Laura Perin More articles by this author Expand All Advertisement PDF downloadLoading ...

Psoralidin inhibits osteosarcoma growth and metastasis by downregulating ITGB1 expression via the FAK and PI3K/Akt signaling pathways

BackgroundPsoralea corylifolia is a medicinal leguminous plant that has long been used to treat various diseases. Psoralidin (PSO) is the main extract compound of P. corylifolia and exhibits antibacterial, antitumor, anti-inflammatory, antioxidant, and other pharmacological activities. PSO has demonstrated inhibitory effects in several cancers; however, its inhibitory effect on osteosarcoma has not been reported. This study aimed to evaluate the inhibitory effect of PSO on osteosarcoma and elucidate the underlying molecular mechanisms.MethodsCrystal violet, cell counting kit-8 (CCK8), and 5-Ethynyl-2′-deoxyuridine (EdU) staining assays were used to assess the inhibitory effect of PSO on the proliferation of 143B and MG63 osteosarcoma cells. Wound healing and Transwell assays were conducted to evaluate the effects of PSO on osteosarcoma cell migration and invasion. The cell cycle and apoptosis were analyzed using flow cytometry. To determine the possible molecular mechanisms, RNA-sequencing was performed and protein expression was analyzed by western blotting. The inhibitory effect of PSO on osteosarcoma in vivo was analyzed using a mouse model of orthotopic osteosarcoma and immunohistochemistry.ResultsPSO inhibited osteosarcoma cell proliferation in a concentration-dependent manner, inhibited cell migration and invasion, and induced cell-cycle arrest and apoptosis. Mechanistically, PSO treatment significantly inhibited the focal adhesion kinase (FAK) and phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathways by downregulating ITGB1 expression in both MG63 and 143B cells. Furthermore, we demonstrated that PSO restrained osteosarcoma growth in vivo.ConclusionPSO may suppress osteosarcoma via the FAK and PI3K/Akt signaling pathways by downregulating ITGB1 expression.

Mammalian Ste20-like kinase 1 inhibition as a cellular mediator of anoikis in mouse bone marrow mesenchymal stem cells

The low survival rate of mesenchymal stem cells (MSCs) caused by anoikis, a form of apoptosis, limits the therapeutic efficacy of MSCs. As a proapoptotic molecule, mammalian Ste20-like kinase 1 (Mst1) can increase the production of reactive oxygen species (ROS), thereby promoting anoikis. Recently, we found that Mst1 inhibition could protect mouse bone marrow MSCs (mBMSCs) from H2O2-induced cell apoptosis by inducing autophagy and reducing ROS production. However, the influence of Mst1 inhibition on anoikis in mBMSCs remains unclear. To investigate the mechanisms by which Mst1 inhibition acts on anoikis in isolated mBMSCs. Poly-2-hydroxyethyl methacrylate-induced anoikis was used following the silencing of Mst1 expression by short hairpin RNA (shRNA) adenovirus transfection. Integrin (ITGs) were tested by flow cytometry. Autophagy and ITGα5β1 were inhibited using 3-methyladenine and small interfering RNA, respectively. The alterations in anoikis were measured by Terminal-deoxynucleoitidyl Transferase Mediated Nick End Labeling and anoikis assays. The levels of the anoikis-related proteins ITGα5, ITGβ1, and phospho-focal adhesion kinase and the activation of caspase 3 and the autophagy-related proteins microtubules associated protein 1 light chain 3 II/I, Beclin1 and p62 were detected by Western blotting. In isolated mBMSCs, Mst1 expression was upregulated, and Mst1 inhibition significantly reduced cell apoptosis, induced autophagy and decreased ROS levels. Mechanistically, we found that Mst1 inhibition could upregulate ITGα5 and ITGβ1 expression but not ITGα4, ITGαv, or ITGβ3 expression. Moreover, autophagy induced by upregulated ITGα5β1 expression following Mst1 inhibition played an essential role in the protective efficacy of Mst1 inhibition in averting anoikis. Mst1 inhibition ameliorated autophagy formation, increased ITGα5β1 expression, and decreased the excessive production of ROS, thereby reducing cell apoptosis in isolated mBMSCs. Based on these results, Mst1 inhibition may provide a promising strategy to overcome anoikis of implanted MSCs.

Integrin β3 regulates apical dendritic morphology of pyramidal neurons throughout hippocampal CA3.

In excitatory hippocampal pyramidal neurons, integrin β3 is critical for synaptic maturation and plasticity in vitro. Itgb3 is a potential autism susceptibility gene that regulates dendritic morphology in the cerebral cortex in a cell-specific manner. However, it is unknown what role Itgb3 could have in regulating hippocampal pyramidal dendritic morphology in vivo, a key feature that is aberrant in many forms of autism and intellectual disability. We found that Itgb3 mRNA is expressed in the stratum pyramidale of CA3. We examined the apical dendritic morphology of CA3 hippocampal pyramidal neurons in conditional Itgb3 knockouts and controls, utilizing the Thy1-GFP-M line. We fully reconstructed the apical dendrite of each neuron and determined each neuron's precise location along the dorsoventral, proximodistal, and radial axes of the stratum pyramidale. We found a very strong effect for Itgb3 expression on CA3 apical dendritic morphology: neurons from conditional Itgb3 knockouts had longer and thinner apical dendrites than controls, particularly in higher branch orders. We also assessed potential relationships between pairs of topographic or morphological variables, finding that most variable pairs were free from any linear relationships to each other. We also found that some neurons from controls, but not conditional Itgb3 knockouts, had a graded pattern of overall diameter along the dorsoventral and proximodistal axes of the stratum pyramidale of CA3. Taken together, Itgb3 is essential for constructing normal dendritic morphology in pyramidal neurons throughout CA3.

Inhibition of Integrin αvβ3-FAK-MAPK signaling constrains the invasion of T-ALL cells

ABSTRACT The role of adhesion receptor integrin αvβ3 in T-ALL was unclear. Firstly, we performed quantitative real-time PCR to assess medullary expression of integrin β3(ITGB3) in T-ALL patients and high ITGB3 expression was relevant with the central nervous system leukemia(CNSL) incidence. Decreasing of cell invasion was observed in Jurkat and Molt4 treated with integrin αvβ3 specific antibody and inhibitor as well as cells with ITGB3 interference. Further, phosphorylation of FAK, cRAF, MEK and ERK decreased in cells with integrin αvβ3 inhibition or interference. Invasion decreased in T-ALL cells treated with FAK and ERK inhibitors. In conclusion, inhibition of integrin αvβ3 signals significantly limits the cell invasion of T-ALL cells.

Tumor cell integrin β4 and tumor stroma E-/P-selectin cooperatively regulate tumor growth in vivo

BackgroundThe immunological composition of the tumor microenvironment has a decisive influence on the biological course of cancer and is therefore of profound clinical relevance. In this study, we analyzed the cooperative effects of integrin β4 (ITGB4) on tumor cells and E-/P-selectin on endothelial cells within the tumor stroma for regulating tumor growth by shaping the local and systemic immune environment.MethodsWe used several preclinical mouse models for different solid human cancer types (xenograft and syngeneic) to explore the role of ITGB4 (shRNA-mediated knockdown in tumor cells) and E-/P-selectins (knockout in mice) for tumor growth; effects on apoptosis, proliferation and intratumoral signaling pathways were determined by histological and biochemical methods and 3D in vitro experiments; changes in the intratumoral and systemic immune cell composition were determined by flow cytometry and immunohistochemistry; chemokine levels and their attracting potential were measured by ELISA and 3D invasion assays.ResultsWe observed a very robust synergism between ITGB4 and E-/P-selectin for the regulation of tumor growth, accompanied by an increased recruitment of CD11b+ Gr-1Hi cells with low granularity (i.e., myeloid-derived suppressor cells, MDSCs) specifically into ITGB4-depleted tumors. ITGB4-depleted tumors undergo apoptosis and actively attract MDSCs, well-known to promote tumor growth in several cancers, via increased secretion of different chemokines. MDSC trafficking into tumors crucially depends on E-/P-selectin expression. Analyses of clinical samples confirmed an inverse relationship between ITGB4 expression in tumors and number of tumor-infiltrating leukocytes.ConclusionsThese findings suggest a distinct vulnerability of ITGB4Lo tumors for MDSC-directed immunotherapies.Graphical

Expression profile of genes related to pregnancy maintenance in Dromedary Camel during the first trimester

So far, few signals involved in embryo-maternal dialogue have been identified in pregnant she-camel. Our objective was to investigate expression profiles of genes relevant to uterine extracellular matrix remodeling (ITGB4, SLCO2A1, FOS, and JUN), uterine tissue vascularization, and placental formation (VEGFA, PGF, and PDGFA), embryonic growth and development (IGF1 and PTEN), plus cell death of uterine tissue (BCL2) in early pregnant versus non-pregnant she-camels. Forty genital tracts (20 pregnant and 20 non-pregnant) and blood samples were collected from abattoirs. Total RNA was extracted from uterine tissues and qRT-PCR was conducted for candidate genes. Serum concentrations of progesterone (P4) and estradiol17-β (E2) were measured. Expression of ITGB4, FOS, and PGF genes increased (P < 0.001) in the right uterine horn of pregnant versus non-pregnant she-camels. Moreover, JUN, SLCO2A1, VEGFA, and PTEN mRNAs were up-regulated (P < 0.001) in various segments of uterine tissues in pregnant groups. The PDGFA transcript was over-expressed (P < 0.001) in both uterine horns of pregnant groups. Additionally, IGF1 was higher (P < 0.001) in the right horn and the uterine body of pregnant groups, and expression of BCL2 was increased (P < 0.001) in the pregnant uterine body. Moreover, serum concentrations of P4 were higher (P < 0.001) and E2 lower (P < 0.05) in pregnant she-camels. Taken together, the fine-tuning of genes related to implantation, matrix formation, vascularization, and placental formation is highly required for successful pregnancy in she-camels.

ITGB1-mediated molecular landscape and cuproptosis phenotype induced the worse prognosis in diffuse gastric cancer.

Diffuse type gastric cancer was identified with relatively worse prognosis than other Lauren's histological classification. Integrin β1 (ITGB1) was a member of integrin family which played a markedly important role in tumorigenesis and progression. However, the influence of ITGB1 in diffuse gastric cancer (DGC) remains uncertain. Here, we leveraged the transcriptomic and proteomic data to explore the association between ITGB1 expression and clinicopathologic information and biological process in DGC. Cell phenotype experiments combined with quantitative-PCR (q-PCR) and western blotting were utilized to identify the potential molecular mechanism underling ITGB1.Transcriptomics and proteomics both revealed that the higher ITGB1 expression was significantly associated with worse prognosis in DGC, but not in intestinal GC. Genomic analysis indicated that the mutation frequency of significantly mutated genes of ARID1A and COL11A1, and mutational signatures of SBS6 and SBS15 were markedly increased in the ITGB1 low expression subgroup. The enrichment analysis revealed diverse pathways related to dysregulation of ITGB1 in DGC, especially in cell adhesion, proliferation, metabolism reprogramming, and immune regulation alterations. Elevated activities of kinase-ROCK1, PKACA/PRKACA and AKT1 were observed in the ITGB1 high-expression subgroup. The ssGSEA analysis also found that ITGB1 low-expression had a higher cuproptosis score and was negatively correlated with key regulators of cuproptosis, including FDX1, DLAT, and DLST. We further observed that the upregulated expression of mitochondrial tricarboxylic acid (TCA) cycle in the ITGB1 low-expression group. Reduced expression of ITGB1 inhibited the ability of cell proliferation and motility and also potentiated the cell sensitive to copper ionophores via western blotting assay. Overall, this study revealed that ITGB1 was a protumorigenic gene and regulated tumor metabolism and cuproptosis in DGC.