Integrin Αvβ3 Expression Research Articles

Single-Cell Proteomic Profiling Identifies Nanoparticle Enhanced Therapy for Triple Negative Breast Cancer Stem Cells.

Breast cancer remains a major cause of cancer-related deaths in women worldwide. Chemotherapy-promoted stemness and enhanced stem cell plasticity in breast cancer is a cause for great concern. The discovery of drugs targeting BCSCs was suggested to be an important advancement in the establishment of therapy that improves the efficacy of chemotherapy. In this work, by using single-cell mass cytometry, we observed that stemness in spheroid-forming cells derived from MDA-MB-231 cells was significantly increased after doxorubicin administration and up-regulated integrin αvβ3 expression was also observed. An RGD-included nanoparticle (CS-V) was designed, and it was found that it could promote doxorubicin’s efficacy against MDA-MB-231 spheroid cells. The above observations suggested that the combination of RGD-included nanoparticles (CS-V) with the chemo-drug doxorubicin could be developed as a potential therapy for breast cancer.

68Ga]Ga-NODAGA-E[(cRGDyK)]2 Angiogenesis PET/MR in a Porcine Model of Chronic Myocardial Infarction.

Angiogenesis is crucial in tissue repair and prevents scar tissue formation following an ischemic event such as myocardial infarction. The ischemia induces formation of new capillaries, which have high expression of integrin αvβ3. [68Ga]Ga-NODAGA-E[(cRGDyK)]2 ([68Ga]Ga-RGD) is a promising PET-radiotracer reflecting angiogenesis by binding to integrin αvβ3. A Göttingen mini-pig underwent transient catheter-induced left anterior descending artery (LAD) occlusion for 120 min, and after 8 weeks was imaged on a Siemens mMR 3T PET/MR system. A large antero-septal infarction was evident by late gadolinium enhancement (LGE) on the short axis and 2–4 chamber views. The infarcted area corresponded to the area with high [68Ga]Ga-RGD uptake on the fused PET/MR images, with no uptake in the healthy myocardium. To support the hypothesis that [68Ga]Ga-RGD uptake reflects angiogenesis, biopsies were sampled from the infarct border and healthy myocardium. Expression of αvβ3 was evaluated using immunohistochemistry. The staining showed higher αvβ3 expression in the capillaries of the infarct border compared to those in the healthy myocardium. These initial data confirm in vivo detection of angiogenesis using [68Ga]Ga-RGD PET in a translational model, which overall support the method applicability when evaluating novel cardio-protective therapies.

Nuclear αvβ3 integrin expression, post translational modifications and regulation in hematological malignancies.

αvβ3 integrin, a plasma membrane protein, is amply expressed on an array of tumors. We identified nuclear αvβ3 pool in ovarian cancer cells and were interested to explore this phenomenon in two rare and aggressive types of leukemia, T-cell acute lymphoblastic leukemia (T-ALL) and Mast cell leukemia (MCL) using Jurkat and HMC-1 cell lines, respectively. Moreover, we collected primary cells from patients with chronic lymphocytic leukemia (CLL, n=11), the most common chronic adult leukemia and used human lymphoblastoid cell lines (LCL) generated from normal B cells. Nuclear αvβ3 integrin was assessed by Western blots, confocal microscopy, and the ImageStream technology which combines flow-cytometry with microscopy. We further examined post translational modifications (phosphorylation/glycosylation), nuclear trafficking regulation using inhibitors for MAPK (U0126) and PI3K (LY294002), as well as nuclear interactions by performing Co-immunoprecipitation (Co-IP). αvβ3 integrin was identified in all cell models within the nucleus and is N-glycosylated. In primary CLL cells the β3 integrin monomer is tyrosine Y759 phosphorylated, suggesting an active receptor conformation. MAPK and PI3K inhibition in Jurkat and CLL cells led to αvβ3 enhancement in the nucleus and a reduction in the membrane. The nuclear αvβ3 integrin interacts with ERK, Histone H3 and Lamin B1 in Jurkat, Histone H3 in CLL cells, but not in control LCL cells. To conclude, this observational study provides the identification of nuclear αvβ3 in hematological malignancies and lays the basis for novel cancer-relevant actions, which may be independent from the membrane functions.

Abstract P103: Molecular Imaging Analysis Of Integrin αvβ3 Expression With Gallium-68-labeled Arg-gly-asp In A Murine Model Of Thoracic Aortic Aneurysms

Background/Objective: Angiogenesis is a complex process. Studies in aneurysmal aortic wall in human tissue and animal models have demonstrated an increased mural neovascularization, and Integrin αVβ3 is a molecular indicator of angiogenesis. We aim to study and characterize Integrin αVβ3 expression with Gallium-68(68Ga)-labeled Arg-Gly-Asp (RGD) as a potential radiotracer using micro Positron Emission Tomography (PET) in a Murine model of Thoracic Aortic Aneurysms (TAA). Methods: Eight specimens of Wistar rat with Calcium Chloride (CaCl) TAA-induced were evaluated. The equipment utilized was an Albira microPET/CT unit (Bruker, Spain), access was obtained in one of the caudal veins for the radiolabeled 68Ga-RGD infusion. The PET uptake and activity were expressed as median value with interquartile range and the Mann-Whitney U test was employed to analyze and compare the chemically induced-site (Aneurysm) and intact aortic tissue (Control) in the same specimens. Statistical analysis was performed with GraphPad Prism 8.0 software. Results: All specimens tolerated well the procedures through a left thoracotomy (Upper panel figure A, B), and underwent imaging studies. A focal increased uptake was observed in the thoracic aortic lesions compared to control regions. The median standard uptake value (SUV) of 68Ga-RGD in the chemically induced-injury site (aneurysm) was 0.0125, while the uptake in the intact site (control) was 0.0003 (P value=0.0117) during a mean follow up period of 32 days (Lower panel figure A-C). Conclusion: We found significant differences of Integrin αVβ3 expression uptake with microPET imaging in chemically induced aneurysms and the intact aortic tissue within 5 weeks of vessel injury. Longitudinal studies with this imaging modality may assist in the molecular characterization of angiogenesis and aneurysmal progression in animal models, and its potential translation in clinical research studies in human patients with aortic aneurysm.

Expression of SGLT1 in the Mouse Endometrial Epithelium and its Role in Early Embryonic Development and Implantation.

Many functional activities of endometrium epithelium are energy consuming which are very important for maintaining intrauterine environment needed by early embryonic development and establishment of implantation window. Glucose is a main energy supplier and one of the main components of intrauterine fluid. Obviously, glucose transports in endometrium epithelium involve in for these activities but their functions have not been elucidated. In this research, we observed a spatiotemporal pattern of sodium glucose transporter 1 (SGLT1) expression in the mouse endometrium. We also determined that progesterone can promote the expression of SGLT1 in the mouse endometrial epithelium in response to the action of oestrogen. Treatment with the SGLT1 inhibitor phlorizin or small interfering RNA specific for SGLT1 (SGLT1-siRNA) altered glucose uptake in primary cultured endometrial epithelial cells, which exhibited reduced ATP levels and AMPK activation. The injection of phlorizin or SGLT1-siRNA into one uterine horn of each mouse on day 2 of pregnancy led to an increased glucose concentration in the uterine fluid and decreased number of harvested normal blastocysts and decreased expression of integrin αVβ3 in endometrial epithelium and increased expression of mucin 1 and lactoferrin in endometrial epithelium and the uterine homogenates exhibited activated AMPK, a decreased ATP level on day 4, and a decreased number of implantation sites on day 5. In embryo transfer experiments, pre-treatment of the uterine horn with phlorizin or SGLT1-siRNA during the implantation window led to a decreased embryo implantation rate on day 5 of pregnancy, even when embryos from normal donor mice were used. In conclusion, SGLT1, which participates in glucose transport in the mouse endometrial epithelium, inhibition and/or reduced expression of SGLT1 affects early embryo development by altering the glucose concentration in the uterine fluid. Inhibition and/or reduced expression of SGLT1 also affects embryo implantation by influencing energy metabolism in epithelial cells, which consequently influences implantation-related functional activities.

Dicalcin suppresses in vitro trophoblast attachment in human cell lines

Implantation is a highly organized process that involves an interaction between a competent blastocyst and a receptive uterus. Despite significant research efforts, the molecular mechanisms governing this complex process remain elusive. Here, we investigated the effect of dicalcin, an S100-like Ca2+-binding protein, on the attachment of choriocarcinoma cells (BeWo cells) onto a monolayer of endometrial carcinoma cells (Ishikawa cells). Extracellularly administered dicalcin bound to both BeWo and Ishikawa cells. Pretreatment of BeWo spheroids with dicalcin reduced the attachment ratio of the spheroids onto the monolayer, whereas that of Ishikawa cells showed no apparent change. We identified the partial amino acid sequence of human dicalcin that exhibited maximum suppression for BeWo spheroid attachment. Transmission electron microscopy analysis revealed that the dicalcin-derived peptide caused a dilation of the intercellular junction between BeWo and ISK cells. Peptide treatment of BeWo spheroids downregulated the expression of integrinαvβ3 in BeWo cells, and induced alterations in their phalloidin-staining pattern, as measured by the length of each F-actin fiber and the thickness of the cortical stress fiber. Thus, dicalcin affects reorganization of the intracellular actin meshwork and subsequently the intensity of attachment, functioning as a novel suppressor of implantation.

Evaluation of Integrin αvβ3 Expression in Murine Xenograft Models: [68Ga]Ga-DOTA-C(RGDfK) PET Study with Immunohistochemical Confirmation.

Tumor blood flow (TBF) is related to drug delivery and hypoxia, both of which can impact the efficacy of anti-cancer therapies. Although integrin αvβ3 expression is related to tumor angiogenesis, it remains unclear whether the degree of angiogenesis affects TBF. This study aimed to evaluate the expression of integrin αvβ3 in mouse tumor models using [68Ga]Ga-DOTA-c(RGDfK) peptide positron emission tomography (PET) and immunohistochemical staining. PET studies were conducted using mouse C6 glioma models and MIA PaCa-2 (n = 6 each). The [68Ga]Ga-DOTA-c(RGDfK) peptide was injected via the tail vein (2.17 ± 0.28 MBq), and 10 min static PET scans were performed. Immunohistochemical analysis was conducted using an integrin αVβ3 antibody. [68Ga]Ga-DOTA-c(RGDfK) peptide PET revealed higher uptake of the radiotracer in C6 gliomas than in MIA PaCa-2 tumors. The mean standardized uptake value was significantly higher in C6 gliomas (0.35 ± 0.058) than in MIA PaCa-2 tumors (0.17 ± 0.045). Histological analysis revealed intense integrin αVβ3 expression in the C6 gliomas, whereas the MIA PaCa-2 tumors had low expression levels. This study showed that the expression of integrin αvβ3 can be differentiated by the [68Ga]Ga-DOTA-c(RGDfK) peptide, suggesting the potential applicability of this peptide in the evaluation of the relationship between angiogenesis and TBF.

SPECT/CT Imaging, Biodistribution and Radiation Dosimetry of a 177Lu-DOTA-Integrin αvβ6 Cystine Knot Peptide in a Pancreatic Cancer Xenograft Model.

IntroductionPancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive malignant neoplasms, as many cases go undetected until they reach an advanced stage. Integrin αvβ6 is a cell surface receptor overexpressed in PDAC. Consequently, it may serve as a target for the development of probes for imaging diagnosis and radioligand therapy. Engineered cystine knottin peptides specific for integrin αvβ6 have recently been developed showing high affinity and stability. This study aimed to evaluate an integrin αvβ6-specific knottin molecular probe containing the therapeutic radionuclide 177Lu for targeting of PDAC.MethodsThe expression of integrin αvβ6 in PDAC cell lines BxPC-3 and Capan-2 was analyzed using RT-qPCR and immunofluorescence. In vitro competition and saturation radioligand binding assays were performed to calculate the binding affinity of the DOTA-coupled tracer loaded with and without lutetium to BxPC-3 and Capan-2 cell lines as well as the maximum number of binding sites in these cell lines. To evaluate tracer accumulation in the tumor and organs, SPECT/CT, biodistribution and dosimetry projections were carried out using a Capan-2 xenograft tumor mouse model.ResultsRT-qPCR and immunofluorescence results showed high expression of integrin αvβ6 in BxPC-3 and Capan-2 cells. A competition binding assay revealed high affinity of the tracer with IC50 values of 1.69 nM and 9.46 nM for BxPC-3 and Capan-2, respectively. SPECT/CT and biodistribution analysis of the conjugate 177Lu-DOTA-integrin αvβ6 knottin demonstrated accumulation in Capan-2 xenograft tumors (3.13 ± 0.63%IA/g at day 1 post injection) with kidney uptake at 19.2 ± 2.5 %IA/g, declining much more rapidly than in tumors.Conclusion 177Lu-DOTA-integrin αvβ6 knottin was found to be a high-affinity tracer for PDAC tumors with considerable tumor accumulation and moderate, rapidly declining kidney uptake. These promising results warrant a preclinical treatment study to establish therapeutic efficacy.

Molecular magnetic resonance imaging of Alpha-v-Beta-3 integrin expression in tumors with ultrasound microbubbles

Microbubbles (MB) are used as ultrasound (US) contrast agents and can be efficiently targeted against markers of angiogenesis and inflammation. Due to their gas core, MB locally alter susceptibilities in magnetic resonance imaging (MRI), but unfortunately, the resulting contrast is low and not sufficient to generate powerful molecular MRI probes. Therefore, we investigated whether a potent molecular MR agent can be generated by encapsulating superparamagnetic iron oxide nanoparticles (SPION) in the polymeric shell of poly (n-butylcyanoacrylate) (PBCA) MB and targeted them against αvβ3 integrins on the angiogenic vasculature of 4T1 murine breast carcinomas. SPION-MB consist of an air core and a multi-layered polymeric shell enabling efficient entrapment of SPION. The mean size of SPION-MB was 1.61 ± 0.32 μm. Biotin-streptavidin coupling was employed to functionalize the SPION-MB with cyclic RGDfK (Arg-Gly-Asp) and RADfK (Arg-Ala-Asp) peptides. Cells incubated with RGD-SPION-MB showed enhanced transverse relaxation rates compared with SPION-MB and blocking αvβ3 integrin receptors with excess free cRGDfK significantly reduced RGD-SPION-MB binding. Due to the fast binding of RGD-SPION-MB in vivo, dynamic susceptibility contrast MRI was employed to track their retention in tumors in real-time. Higher retention of RGD-SPION-MB was observed compared with SPION-MB and RAD-SPION-MB. To corroborate our MRI results, molecular US was performed the following day using the destruction-replenishment method. Both imaging modalities consistently indicated higher retention of RGD-SPION-MB in angiogenic vessels compared with SPION-MB and RAD-SPION-MB. Competitive blocking experiments in mice further confirmed that the binding of RGD-SPION-MB to αvβ3 integrin receptors is specific. Overall, this study demonstrates that RGD-SPION-MB can be employed as molecular MR/US contrast agents and are capable of assessing the αvβ3 integrin expression in the neovasculature of malignant tumors.

Bone Morphogenetic Protein 2 Increases Human Trophoblast Invasion by Up-Regulating Integrin Beta3

The adequate invasion of extravillous trophoblast cells (EVTs) is indispensable for the implantation of embryos and subsequent remodeling of uterine spiral arteries in early human gestation. Bone morphogenetic protein 2 (BMP2), which is abundantly expressed at the maternal-fetal interface, has been shown to promote the human EVT invasion process (1). Integrin switching (i.e., a switch from α6β4 to αvβ3) plays essential roles in cell-extracellular matrix adhesion and has been reported to influence EVT migration and invasion (2). Moreover, integrin β3 has been found to promote the adhesion, invasion, and migration abilities of embryonic trophoblasts (3). However, whether integrin β3 participates in BMP2 signaling and mediates BMP2-increased-human trophoblast invasion remains unknown. The purpose of our study was to explore the effects of BMP2 on integrin αvβ3 expression and the possible mediation role of integrin β3 in BMP2-regulated human trophoblast invasion. We used immortalized human trophoblast cell line (HTR8/SVneo) and primary human extravillous trophoblast cells (EVTs) isolated from first-trimester villi as study models. RT-qPCR and Western blot assay were respectively utilized to detect the messenger RNA and protein levels of intergrin αv and β3. The function of the target protein was studied by siRNA knockdown, and the trophoblast invasion ability was checked by Matrigel-coated transwell invasion assays. Our results demonstrated that the mRNA and protein levels of integrin β3, rather than integrin αv, were up-regulated after BMP2 treatment in HTR8/SVneo and primary EVT cells. Importantly, siRNA-mediated down-regulation of integrin β3 significantly inhibited basal and BMP2-induced HTR8/SVneo cell invasionas measured by transwell invasion assay. In conclusion, we findings support that BMP2 promotes human trophoblast cell invasion by up-regulating integrin β3 expression, benefiting the in-depth understanding of the pro-invasive effect of BMP2 on human trophoblasts during early pregnancy. Reference: (1) Hong-Jin Zhao et al., Cell Death Dis 2018;9:174. (2) Damsky, C.H. et al, Development 1994; 120, 3657-3666. (3) Dong-Mei He et al., Reproduction 2019;157:423-430.

αvβ3 Integrin Expression and Mitogenic Effects by Thyroid Hormones in Chronic Lymphocytic Leukemia.

Background: Chronic lymphocytic leukemia (CLL) is the most common adult leukemia. The thyroid hormones, T3 and T4, bind the αvβ3 integrin and activate phosphorylates ERK (pERK). These tumor-promoting actions were reported in a number of malignancies, but not in CLL. Methods: Primary cells from 22 CLL patients were verified for disease markers (CD5/CD19/CD23) and analyzed for αvβ3 by flow cytometry (FC), ImageStream, Western blots (WB), and immunohistochemistry (IHC) in archival bone marrow (BM, n = 6) and lymph node (LN, n = 5) tissues. Selected samples (n = 8) were incubated with T3 (1–100 nM) or T4 (0.1–10 µM) for 30 min, and the expression levels of αvβ3, pERK and PCNA (cell proliferation marker) were determined (WB). Results: αvβ3 was detected on the membrane of circulating CLL cells and in the BM but not in the LN. T3 and T4 enhanced αvβ3 protein levels in primary CLL cells. Similarly, pERK and PCNA were rapidly induced in response to T3 and T4 exposure. Conclusions: αvβ3 integrin is expressed on primary CLL cells and is induced by thyroid hormones. We further suggest that the hormones are mitogenic in these cells, presumably via αvβ3-mediated signaling.

Inhibition by Thyroid Hormones of Cell Migration Activated by IGF-1 and MCP-1 in THP-1 Monocytes: Focus on Signal Transduction Events Proximal to Integrin αvβ3.

Interaction between thyroid hormones and the immune system is reported in the literature. Thyroid hormones, thyroxine, T4, but also T3, act non-genomically through mechanisms that involve a plasma membrane receptor αvβ3 integrin, a co-receptor for insulin-like growth factor-1 (IGF-1). Previous data from our laboratory show a crosstalk between thyroid hormones and IGF-1 because thyroid hormones inhibit the IGF-1-stimulated glucose uptake and cell proliferation in L-6 myoblasts, and the effects are mediated by integrin αvβ3. IGF-1 also behaves as a chemokine, being an important factor for tissue regeneration after damage. In the present study, using THP-1 human leukemic monocytes, expressing αvβ3 integrin in their cell membrane, we focused on the crosstalk between thyroid hormones and either IGF-1 or monocyte chemoattractant protein-1 (MCP-1), studying cell migration and proliferation stimulated by the two chemokines, and the role of αvβ3 integrin, using inhibitors of αvβ3 integrin and downstream pathways. Our results show that IGF-1 is a potent chemoattractant in THP-1 monocytes, stimulating cell migration, and thyroid hormone inhibits the effect through αvβ3 integrin. Thyroid hormone also inhibits IGF-1-stimulated cell proliferation through αvβ3 integrin, an example of a crosstalk between genomic and non-genomic effects. We also studied the effects of thyroid hormone on cell migration and proliferation induced by MCP-1, together with the pathways involved, by a pharmacological approach and docking simulation. Our findings show a different downstream signaling for IGF-1 and MCP-1 in THP-1 monocytes mediated by the plasma membrane receptor of thyroid hormones, integrin αvβ3.

Talin1 regulates endometrial adhesive capacity through the Ras signaling pathway

AimsBlastocyst implantation is mainly depended on the adhesion between cells and cell matrix. Endometrial adhesion plays an important role in establishing embryo implantation, but the underlying mechanisms are remains unclear. Talin1 is a local adhesion complex protein that is necessary for cell adhesion and movement. However, the role and mechanisms of Talin1 in embryo implantation are still unclear. Main methodsThe expression of Talin1 and Integrin αvβ3 was measured in the receptive endometrium from the RIF (Recurrent implantation failure) cohort and NC (normal fertile control group) cohort. A JEG-3 trophoblast and endometrial epithelial cell adhesion model and pregnant mouse model were established. The molecular mechanism of Talin1-mediated cell adhesion was explored by RNA sequencing, RT-qPCR, as well as western blotting assays. Key findingsTalin1 enhances endometrial cell adhesion by regulating the Ras signaling pathway, and ultimately facilitates embryo implantation. SignificanceThis study revealed the molecular mechanisms of regarding the pathogenesis of RIF caused by endometrial receptivity insufficiency. Further pharmacological research on the Ras signaling pathway would be valuable and might provide new therapeutic targets for RIF patients.

Laminin alpha 4 promotes bone regeneration by facilitating cell adhesion and vascularization

Selective cell retention (SCR) has been widely used as a bone tissue engineering technique for the real-time fabrication of bone grafts. The greater the number of mesenchymal stem cells (MSCs) and endothelial progenitor cells (EPCs) retained in the scaffold, the better the osteoinductive and angiogenic properties of the scaffold's microenvironment. Improved bioscaffold properties in turn lead to improved bone graft survival, bone regeneration, and angiogenesis. Laminin plays a key role in cell-matrix adhesion, cell proliferation, and differentiation. We designed a collagen-binding domain (CBD) containing the core functional amino acid sequences of laminin α4 (CBD-LN peptide) to supplement the functional surface of a collagen-based decalcified bone matrix (DBM) scaffold. This scaffold promoted MSCs and EPCs early cell adhesion through up-regulating the expression of integrin α5β1 and integrin αvβ3 respectively, thus accelerated the following cell spreading, proliferation, and differentiation. Interestingly, it promoted the retention of MSCs (CD90+/CD105+ cells) and EPCs (CD31+ cells) in the scaffold following the use of clinical SCR technology. Furthermore, the DBM/CBD-LN scaffold induced the formation of type H vessels through the activation of the HIF-1α signaling pathway. The DBM/CBD-LN scaffold displayed rapid bone formation and angiogenesis in vivo, suggesting that it might be used as a new biomaterial in bone tissue engineering. Statement of significanceSelective cell retention technology (SCR) has been utilized in clinical settings to manufacture bioactive bone grafts. Specifically, demineralized bone matrix (DBM) is a widely-used SCR clinical biomaterial but it displays poor adhesion performance and angiogenic activity. In this work, we designed a collagen-binding domain (CBD) containing the core functional amino acid sequences of laminin α4 to supplement the functional surface of a collagen-based DBM scaffold. This bioscaffold promoted SCR-mediated MSCs and EPCs early cell adhesion, thus accelerated the following cell spreading, proliferation, and differentiation. Our results indicate this bioscaffold greatly induced osteogenesis and angiogenesis in vivo. In general, this bioscaffold has a good prospect for SCR application and may provide highly bioactive bone implant in clinical environment.

Genetic and epigenetic changes in the eutopic endometrium of women with endometriosis: association with decreased endometrial αvβ3 integrin expression

About 40% of women with infertility and 70% of women with pelvic pain suffer from endometriosis. The pregnancy rate in women undergoing IVF with low endometrial integrin αvβ3 (LEI) expression is significantly lower compared to the women with high endometrial integrin αvβ3 (HEI). Mid-secretory eutopic endometrial biopsies were obtained from healthy controls (C; n=3), and women with HEI (n=4) and LEI (n=4) and endometriosis. Changes in gene expression were assessed using human gene arrays and DNA methylation data were derived using 385 K Two-Array Promoter Arrays. Transcriptional analysis revealed that LEI and C groups clustered separately with 396 differentially expressed genes (DEGs) (P<0.01: 275 up and 121 down) demonstrating that transcriptional and epigenetic changes are distinct in the LEI eutopic endometrium compared to the C and HEI group. In contrast, HEI vs C and HEI vs LEI comparisons only identified 83 and 45 DEGs, respectively. The methylation promoter array identified 1304 differentially methylated regions in the LEI vs C comparison. The overlap of gene and methylation array data identified 14 epigenetically dysregulated genes and quantitative RT-PCR analysis validated the transcriptomic findings. The analysis also revealed that aryl hydrocarbon receptor (AHR) was hypomethylated and significantly overexpressed in LEI samples compared to C. Further analysis validated that AHR transcript and protein expression are significantly (P<0.05) increased in LEI women compared to C. The increase in AHR, together with the altered methylation status of the 14 additional genes, may provide a diagnostic tool to identify the subset of women who have endometriosis-associated infertility.

Assessing Doxorubicin-Induced Cardiomyopathy by 99mTc-3PRGD2 Scintigraphy Targeting Integrin αvβ3 in a Rat Model.

The present study evaluated interstitial alterations in doxorubicin-induced cardiomyopathy using a radiolabeled RGD peptide 99mTc-3PRGD2 specific for integrin αvβ3 that targets myofibroblasts.Cardiomyopathy was induced in 20 Sprague-Dawley rats by intraperitoneal doxorubicin injections (2.5 mg/kg/week) for up to six weeks. 99mTc-3PRGD2 scintigraphy was performed in control rats (n = 6) at baseline and three, six, and nine weeks after first doxorubicin administration (n = 6, 6, and 5 for each time point). For another three rats of 6-week modeling, cold c(RGDyK) was co-injected with 99mTc-3PRGD2 to evaluate specific radiotracer binding. Semi-quantitative parameters were acquired to compare radiotracer uptake among all groups. The biodistribution of 99mTc-3PRGD2 was evaluated by a γ-counter after scintigraphy. Haematoxylin and eosin, and Masson's staining were used to evaluate myocardial injury and fibrosis, while western blotting and immunofluorescence co-localization were used to analyze integrin αvβ3 expression in the myocardium.The 99mTc-3PRGD2 half-life in the cardiac region (Heartt 1/2) of the 9-week model and heart radioactivity percentage (%Heart20 min, %Heart40 min and %Heart60 min) of the 6 and 9-week models were significantly increased compared to the control. Heart-to-background ratio (HBR20 min, HBR40 min and HBR60 min) increase began in the third week, continued until the sixth week, and was reversed in the ninth week, which paralleled the changing trend of cardiac integrin αvβ3 expression. The myocardial biodistribution of 99mTc-3PRGD2 was significantly correlated with integrin β3 expression.The 99mTc-3PRGD2 scintigraphy allows for non-invasive visualization of interstitial alterations during doxorubicin-induced cardiomyopathy.

Targeting Interstitial Myofibroblast-Expressed Integrin αvβ3 Alleviates Renal Fibrosis.

Renal fibrosis is the final manifestation of various chronic kidney diseases. Interstitial myofibroblasts, which are reported to highly express integrin αvβ3, are the effector cells in renal fibrogenesis. Since current therapies do not efficiently target these cells, there is no effective therapeutic method for preventing or mitigating the disease. Here, we modified sterically stable PEGylated liposomes with the pentapeptide cRGDfC (RGD-Lip), which has a high affinity for αvβ3, to specifically deliver drug to renal interstitial myofibroblasts. Our results showed that attaching cRGDfC to liposomes significantly increased their uptake by activated renal fibroblasts NRK-49F cells, and this effect was greatly abolished by adding excess-free cRGDfC or a knockdown of αvβ3. Systemic administration of RGD-Lip gave rise to significant accumulation in a fibrotic kidney, which is ascribed to the specific recognition with integrin αvβ3 on interstitial myofibroblasts. When loaded with celastrol, RGD-guided liposomes dramatically depressed the proliferation and activation of NRK-49F cells in vitro. Additionally, celastrol-loaded RGD-Lip markedly attenuated renal fibrosis, injury, and inflammation induced by unilateral ureteral obstruction (UUO) in mice, without inducing significant systemic toxicity. Thus, this liposomal system shows great promise for delivering therapeutic agents to interstitial myofibroblasts for renal fibrosis treatment with minimal side effects.

Differential expression of αVβ3 and αVβ6 integrins in prostate cancer progression.

Neuroendocrine prostate cancer (NEPrCa) arises de novo or after accumulation of genomic alterations in pre-existing adenocarcinoma tumors in response to androgen deprivation therapies. We have provided evidence that small extracellular vesicles released by PrCa cells and containing the αVβ3 integrin promote neuroendocrine differentiation of PrCa in vivo and in vitro. Here, we examined αVβ3 integrin expression in three murine models carrying a deletion of PTEN (SKO), PTEN and RB1 (DKO), or PTEN, RB1 and TRP53 (TKO) genes in the prostatic epithelium; of these three models, the DKO and TKO tumors develop NEPrCa with a gene signature comparable to those of human NEPrCa. Immunostaining analysis of SKO, DKO and TKO tumors shows that αVβ3 integrin expression is increased in DKO and TKO primary tumors and metastatic lesions, but absent in SKO primary tumors. On the other hand, SKO tumors show higher levels of a different αV integrin, αVβ6, as compared to DKO and TKO tumors. These results are confirmed by RNA-sequencing analysis. Moreover, TRAMP mice, which carry NEPrCa and adenocarcinoma of the prostate, also have increased levels of αVβ3 in their NEPrCa primary tumors. In contrast, the αVβ6 integrin is only detectable in the adenocarcinoma areas. Finally, analysis of 42 LuCaP patient-derived xenografts and primary adenocarcinoma samples shows a positive correlation between αVβ3, but not αVβ6, and the neuronal marker synaptophysin; it also demonstrates that αVβ3 is absent in prostatic adenocarcinomas. In summary, we demonstrate that αVβ3 integrin is upregulated in NEPrCa primary and metastatic lesions; in contrast, the αVβ6 integrin is confined to adenocarcinoma of the prostate. Our findings suggest that the αVβ3 integrin, but not αVβ6, may promote a shift in lineage plasticity towards a NE phenotype and might serve as an informative biomarker for the early detection of NE differentiation in prostate cancer.

Nano Diaminopropane tetrac and integrin αvβ3 expression in different cancer types: Anti-cancer efficacy and Safety

Integrins are a family of heterodimeric plasma membrane glycoproteins, which regulate tumor growth, angiogenesis, migration, and metastasis. Integrin αvβ3 has been recognized as a putative target for the treatment of several cancers. Thus, the characterization of αvβ3 distribution in different human tumors is of substantial interest in tumor targeting and its suppression. In this study we evaluated the expression of integrin αvβ3 in different cancer types to define the expression pattern in cancer model. Furthermore, we investigated the effect of novel αvβ3 antagonist Diaminopropane Tetraiodothyroacetic acid conjugated to poly (lactic-co-glycolic acid) polymer and its nanoformulated form (NDAT), on different cancer cell lines both in vitro and in xenografts. In vitro, NDAT downregulated αv and β3 monomer expression. In vivo in tumor xenografts, similarly, NDAT downregulated αv and β3. Distinct reduction in tumor weight and viability was observed in glioblastoma xenografts treated with NDAT. Furthermore, NDAT was safe and tolerable in mice treated with high doses. In conclusion, NDAT is an effective and safe inhibitor of integrin αvβ3 expression in various cancer types, which indicates its impact on the targetability and suppression of αvβ3-associated tumor functions.

SPARC Aggravates Blood-Brain Barrier Disruption via Integrin αVβ3/MAPKs/MMP-9 Signaling Pathway after Subarachnoid Hemorrhage.

Blood-brain barrier (BBB) disruption is a common and critical pathology following subarachnoid hemorrhage (SAH). We investigated the BBB disruption property of secreted protein acidic and rich in cysteine (SPARC) after SAH. A total of 197 rats underwent endovascular perforation to induce SAH or sham operation. Small interfering ribonucleic acid (siRNA) for SPARC or scrambled siRNA was administered intracerebroventricularly to rats 48 h before SAH. Anti-SPARC monoclonal antibody (mAb) 236 for functional blocking or normal mouse immunoglobulin G (IgG) was administered intracerebroventricularly 1 h after SAH. Selective integrin αVβ3 inhibitor cyclo(-RGDfK) or phosphate-buffered saline was administered intranasally 1 h before SAH, along with recombinant SPARC treatment. Neurobehavior, SAH severity, brain edema, immunohistochemical staining, and Western blot were evaluated. The expression of SPARC and integrin αVβ3 was upregulated after SAH in the endothelial cells. SPARC siRNA and anti-SPARC mAb 236 prevented neuroimpairments and brain edema through protection of BBB as measured by IgG extravasation 24 and 72 h after SAH. Recombinant SPARC aggravated neuroimpairments and cyclo(-RGDfK) suppressed the harmful neurological effects via inhibition of activated c-Jun N-terminal kinase, p38, and matrix metalloproteinase-9 followed by retention of endothelial junction proteins. SPARC may induce post-SAH BBB disruption via integrin αVβ3 signaling pathway.