Sexual dimorphism in reactive oxygen species production and a role for integrin α1ß1 in estrogen receptor α and ß expression in articular cartilage.

BACKGROUND: Osteoarthritis (OA) is a debilitating disease involving cartilage degradation. A need remains for the discovery of new molecular targets in cartilage for pharmaceutical intervention of OA. One potential target is integrin α1ß1 that protects against OA when it is upregulated by chondrocytes early in the disease process. Integrin α1ß1 offers this protection by dampening epidermal growth factor receptor (EGFR) signaling, and its effects are more robust in females compared to males. The aim of this study, therefore, was to measure the impact of itga1 on chondrocyte EGFR activity and downstream reactive oxygen species (ROS) production in male and female mice. Furthermore, chondrocyte expression of estrogen receptor (ER) α and ERß was measured to investigate the mechanism for sexual dimorphism in the EGFR/integrin α1ß1 signaling axis. We hypothesized that integrin α1ß1 would decrease ROS production and pEGFR and 3-nitrotyrosine expression, with this effect being greater in females. We further hypothesized that chondrocyte expression of ERα and ERß would be greater in females compared to males, with a greater effect seen in itga1-null compared to wild-type mice. MATERIALS AND METHODS: Femoral and tibial cartilage of male and female, wild-type and itga1-null mice were processed for ex vivo confocal imaging of ROS, immunohistochemical analysis of 3-nitrotyrosine, or immunofluorescence of pEGFR and ERα and ERß. RESULTS: We show that ROS-producing chondrocytes are more abundant in female itga1-null compared to wild-type mice ex vivo; however, itga1 had limited influence on the percent of chondrocytes stained positively for 3-nitrotyrosine or pEGFR in situ. In addition, we found that itga1 influenced ERα and ERß expression in femoral cartilage from female mice, and that ERα and ERß were coexpressed as well as colocalized in chondrocytes. Finally, we show sexual dimorphism in ROS and 3-nitrotyrosine production, but surprisingly not in pEGFR expression. CONCLUSIONS: Together these data highlight sexual dimorphism in the EGFR/integrin α1ß1 signaling axis and underline the need for further investigation into the role of ERs in this biological paradigm. Understanding the molecular mechanisms underlying the development of OA is essential for the development of individualized, sex-specific treatments in this age of personalized medicine.

Integrin Targeting Enhances the Antimelanoma Effect of Annexin V in Mice.

Malignant melanoma, an increasingly common form of skin cancer, is a major threat to public health, especially when the disease progresses past skin lesions to the stage of advanced metastasis. Targeted drug development is an effective strategy for the treatment of malignant melanoma. In this work, a new antimelanoma tumor peptide, the lebestatin-annexin V (designated LbtA5) fusion protein, was developed and synthesized by recombinant DNA techniques. As a control, annexin V (designated ANV) was also synthesized by the same method. The fusion protein combines annexin V, which specifically recognizes and binds phosphatidylserine, with the disintegrin lebestatin (lbt), a polypeptide that specifically recognizes and binds integrin α1ß1. LbtA5 was successfully prepared with good stability and high purity while retaining the dual biological activity of ANV and lbt. MTT assays demonstrated that both ANV and LbtA5 could reduce the viability of melanoma B16F10 cells, but the activity of the fusion protein LbtA5 was superior to that of ANV. The tumor volume growth was slowed in a mouse xenograft model treated with ANV and LbtA5, and the inhibitory effect of high concentrations of LbtA5 was significantly better than that of the same dose of ANV and was comparable to that of DTIC, a drug used clinically for melanoma treatment. The hematoxylin and eosin (H&E) staining test showed that ANV and LbtA5 had antitumor effects, but LbtA5 showed a stronger ability to induce melanoma necrosis in mice. Immunohistochemical experiments further showed that ANV and LbtA5 may inhibit tumor growth by inhibiting angiogenesis in tumor tissue. Fluorescence labeling experiments showed that the fusion of ANV with lbt enhanced the targeting of LbtA5 to mouse melanoma tumor tissue, and the amount of target protein in tumor tissue was significantly increased. In conclusion, effective coupling of the integrin α1ß1-specific recognition molecule lbt confers stronger biological antimelanoma effects of ANV, which may be achieved by the dual effects of effective inhibition of B16F10 melanoma cell viability and inhibition of tumor tissue angiogenesis. The present study describes a new potential strategy for the application of the promising recombinant fusion protein LbtA5 in the treatment of various cancers, including malignant melanoma.

The effects of inhibition and siRNA knockdown of collagen-binding integrins on human umbilical vein endothelial cell migration and tube formation.

Blood vessels in the body are lined with endothelial cells which have vital roles in numerous physiological and pathological processes. Collagens are major constituents of the extracellular matrix, and many adherent cells express several collagen-binding adhesion receptors. Here, we study the endothelium-collagen interactions mediated by the collagen-binding integrins, α1ß1, α2ß1, α10ß1 and α11ß1 expressed in human umbilical vein endothelial cells (HUVECs). Using qPCR, we found expression of the α10 transcript of the chondrocyte integrin, α10ß1, along with the more abundant α2, and low-level expression of α1. The α11 transcript was not detected. Inhibition or siRNA knockdown of the α2-subunit resulted in impaired HUVEC adhesion, spreading and migration on collagen-coated surfaces, whereas inhibition or siRNA knockdown of α1 had no effect on these processes. In tube formation assays, inhibition of either α1 or α2 subunits impaired the network complexity, whereas siRNA knockdown of these integrins had no such effect. Knockdown of α10 had no effect on cell spreading, migration or tube formation in these conditions. Overall, our results indicate that the collagen-binding integrins, α1ß1 and α2ß1 play a central role in endothelial cell motility and self-organisation.

Assessment of functional roles and therapeutic potential of integrin receptors in osteoarthritis: A systematic review and meta-analysis of preclinical studies.

BACKGROUND: Integrins are heterodimeric transmembrane receptors that mediate a variety of biological function and plays a critical role in osteoarthritis (OA) pathogenesis, which may provide new targets for the development of OA therapies. However, the roles of integrins in different stages of OA remain elusive. OBJECTIVES: This study aimed to synthesize all published preclinical evidence on the roles of integrin receptors in different stages of OA to identify the potential target for drug development in alleviating OA pathogenesis. METHODS: Major electronic databases were used to identify related original articles. The methodological quality of all included studies was appraised using the SYRCLE risk of bias tool. We used the generic inverse variance with random effects model to calculate standardized mean differences (SMDs) and 95% confidence interval (CI). RESULTS: Seventeen studies were included in this systematic review. Integrin α5ß1 activation increases the histopathological score both in early [SMD, 6.39; 95%CI (2.90, 9.87); p = 0.0003] and late [SMD, 3.41; 95%CI (2.44, 4.38); p < 0.00001] stage of OA. Integrin α5ß1 also increased the core catabolic factors like MMP-3, IL-1ß, and TNF-α. Interestingly, the inactivation of α5ß1 integrin did not change the histopathological score (p = 0.84). Similarly, ß1 integrin notably increased histopathological score at both stages of OA [early; SMD, 7.13; 95%CI (2.01, 12.24); p = 0.006]; [late; SMD, 10.25; 95%CI (5.11, 15.39); p < 0.0001], and increased the MMP-13 levels. However, integrin ß1 was upregulated at the early stage and downregulated at the late stage of OA. Furthermore, α2ß1 integrin significantly increased histopathological score [SMD, 3.14; 95%CI (2.18, 4.10); p < 0.00001] and MMP-13 [SMD, 2.24; 95%CI (0.07, 4.41); p = 0.04]. Deactivating integrin α1ß1 increased histopathological score in late [SMD, 1.53; 95%CI (0.80, 2.26); p < 0.0001], but not in early [SMD, 0.90; 95%CI (-1.65, 3.45); p = 0.49] stage of OA. CONCLUSION: This study provides evidence that α5ß1, α2ß1, and α1ß1 integrin might be the potential target for future drug development in alleviating OA pathogenesis. Further work is required to establish our findings through activating/deactivating these receptors in different stages of OA.

Chemotherapy treatment induces pro-invasive changes in liver ECM composition.

Metastasis accounts for 90% of cancer-related deaths, yet the mechanisms by which cancer cells colonize secondary organs remain poorly understood. For breast cancer patients, metastasis to the liver is associated with poor prognosis and a median survival of 6 months. Standard of care is chemotherapy, but recurrence occurs in 30% of patients. Systemic chemotherapy has been shown to induce hepatotoxicity and fibrosis, but how chemotherapy impacts the composition of the liver extracellular matrix (ECM) remains unknown. Individual ECM proteins drive tumor cell proliferation and invasion, features that are essential for metastatic outgrowth in the liver. First, we find that the ECM of livers isolated from chemotherapy-treated MMTV-PyMT mice increases the invasion, but not proliferation, of metastatic breast cancer cells. Proteomic analysis of the liver ECM identified Collagen V to be more abundant in paclitaxel-treated livers. We show that Collagen V increases cancer cell invasion via α1ß1 integrins and MAPK signaling, while also increasing the alignment of Collagen I, which has been associated with increased invasion. Treatment with obtustatin, an inhibitor specific to α1ß1 integrins, inhibits tumor cell invasion in decellularized ECM from paclitaxel-treated livers. Overall, we show chemotherapy treatment alters the liver microenvironment, priming it as a pro-metastatic niche for cancer metastasis.

Molecular dynamics simulations, docking and MMGBSA studies of newly designed peptide-conjugated glucosyloxy stilbene derivatives with tumor cell receptors.

In this work, for the first time, we designed derivatives of beta-D-glucosyloxy-3-hydroxy-trans-stiblene-2-carboxylic acid (GHS), by conjugating GHS with tumor targeting peptides RPARPAR and GGKRPAR to target over-expressed receptors in tumor cells. The sequences RPARPAR and GGKRPAR are known to target the neuropilin1 (NRP1) receptor due to the C-terminal Arg domain; however, their effectiveness has never been examined with other commonly over-expressed receptors in tumor cells, particularly of chronic lymphocytic leukemia that include integrin α1ß1 and CD22. By conjugating these peptides with GHS, which is known for its inherent anti-cancer properties, the goal is to further enhance tumor cell targeting by developing compounds that can target multiple receptors. The physicochemical properties of the conjugates and individual peptides were analyzed using Turbomole and COSMOthermX20 in order to determine their hydrogen bond accepting and donating capabilities. The web server POCASA was used in order to determine the surface cavities and binding pockets of the three receptors. To explore the binding affinities, we conducted molecular docking studies with the peptides and the conjugates with each of the receptors. After molecular docking, the complexes were analyzed using Protein-Ligand Interaction Profiler to determine the types of interactions involved. Molecular dynamics simulation studies were conducted to explore the stability of the receptor-ligand complexes. Our results indicated that in most cases the conjugates showed higher binding and stability with the receptors. Additionally, highly stable complexes of conjugates were obtained with CD22, NRP1 and in most cases with the integrin α1ß1 receptor as well. The binding energies were calculated for each of the receptor ligand complexes through trajectory analysis using MMGBSA studies. SwissADME studies revealed that the compounds showed low GI absorption and were not found to be CYP inhibitors and had bioavailability score that would allow them to be considered as potential drug candidates. Overall, our results for the first time show that the designed conjugates can target multiple over-expressed receptors in tumor cells and may be potentially developed as future therapeutics for targeting tumor cells.

Distinctive roles of fibrillar collagen I and collagen III in mediating fibroblast-matrix interaction: A nanoscopic study.

One major goal in tissue engineering is to create functional materials, mimicking scaffolds in native tissues, to modulate cell function for tissue repair. Collagen is the most abundant structural protein in human body. Though collagen I (COLI) and collagen III (COLIII) are the predominant collagen types in connective tissues and they form stable hybrid fibrils at varied ratios, cell responses to the hybrid matrices are underinvestigated. In this work, we aim to explicate the distinctive roles of COLI and COLIII in fibroblast activation. Unidirectionally aligned COLI, COLIII and COLI-COLIII hybrid nanofibrils were generated via epitaxial growth of collagen on mica. AFM analyses revealed that, with the increase of COLI/COLIII ratio, the fibril width and stiffness increased and the binding affinity of cells to the matrix decreased. A hybrid matrix was found to activate fibroblasts the most effectively, characterized by extensive cell polarization with rigid stress fiber bundles and high α-SMA expression, and by the highest-level of collagen synthesis. It is ascribed to the fine balance between biochemical and biophysical cues achieved on the hybrid matrix. Thus, matrices of aligned COLI-COLIII hybrid fibrils and their derived multifunctional composites can be good candidates of implantation scaffolds for tissue regeneration.

Identification of a novel collagen-like peptide by high-throughput screening for effective wound-healing therapy.

Tissue regeneration and wound healing are still serious clinical complications globally and lack satisfactory cures. Inspired by the impressive regeneration ability of the post-injury earthworms and their widely accepted medicinal properties, we screened and identified a novel collagen-like peptide from the amputated earthworms using high-throughput techniques, including transcriptomics, proteomics, and mass spectrum. The identified collagen-like peptide col4a1 was cloned and expressed to comprehensively investigate the wound healing effect and underlying mechanism. It exerted significant effects on wound healing both in vitro and in vivo, including enhanced viability, proliferation, migration of fibroblasts, granulation, and collagen deposition. Moreover, the col4a1 functioned via binding with integrin α2ß1 and upregulating the RAS/MAPK signaling pathway. This work demonstrates that the novel collagen-like peptide col4a1 obtained from the amputated earthworms enables enhanced wound healing and provides new opportunities for wound care.

EGF receptor-mediated FUS phosphorylation promotes its nuclear translocation and fibrotic signaling.

Excessive accumulation of collagen leads to fibrosis. Integrin α1ß1 (Itgα1ß1) prevents kidney fibrosis by reducing collagen production through inhibition of the EGF receptor (EGFR) that phosphorylates cytoplasmic and nuclear proteins. To elucidate how the Itgα1ß1/EGFR axis controls collagen synthesis, we analyzed the levels of nuclear tyrosine phosphorylated proteins in WT and Itgα1-null kidney cells. We show that the phosphorylation of the RNA-DNA binding protein fused in sarcoma (FUS) is higher in Itgα1-null cells. FUS contains EGFR-targeted phosphorylation sites and, in Itgα1-null cells, activated EGFR promotes FUS phosphorylation and nuclear translocation. Nuclear FUS binds to the collagen IV promoter, commencing gene transcription that is reduced by inhibiting EGFR, down-regulating FUS, or expressing FUS mutated in the EGFR-targeted phosphorylation sites. Finally, a cell-penetrating peptide that inhibits FUS nuclear translocation reduces FUS nuclear content and collagen IV transcription. Thus, EGFR-mediated FUS phosphorylation regulates FUS nuclear translocation and transcription of a major profibrotic collagen gene. Targeting FUS nuclear translocation offers a new antifibrotic therapy.

Outer membrane protein OmpV mediates Salmonella enterica serovar typhimurium adhesion to intestinal epithelial cells via fibronectin and α1ß1 integrin.

Salmonella typhimurium is an invasive Gram-negative enteric bacterium, which causes salmonellosis, a type of gastroenteritis in humans and typhoid-like symptoms in mice. Upon entering through the contaminated food and water, S. typhimurium adheres, colonises, and invades intestinal epithelial cells (IECs) of the small intestine. In this study, we have shown that upon deletion of the outer membrane protein OmpV, there is a significant decrease in adherence of S. typhimurium to the IECs, indicating that OmpV is an important adhesin of S. typhimurium. Further, our study showed that OmpV binds to the extracellular matrix component fibronectin and signals through α1ß1 integrin receptor on the IECs and OmpV-mediated activation of α1ß1, resulting in the activation of focal adhesion kinase and F-actin modulation. Actin modulation is crucial for bacterial invasion. To the best of our knowledge, this is the first report of an adhesin mediated its effect through integrin in S. typhimurium. Further, we have observed a decrease in pathogenicity in terms of increased LD50 dose, lesser bacterial numbers in stool, and less colonisation of bacteria in different organs of mice infected with Δompv mutant compared with the wild-type bacteria, thus confirming the crucial role of OmpV in the pathogenesis of S. typhimurium.

VLA-1 Binding to Collagen IV Controls Effector T Cell Suppression by Myeloid-Derived Suppressor Cells in the Splenic Red Pulp.

Myeloid-derived suppressor cells (MDSCs) represent a major population controlling T cell immune responses. However, little is known about their molecular requirements for homing and T cell interaction to mediate suppression. Here, we investigated the functional role of the homing and collagen IV receptor VLA-1 (α1ß1-integrin) on in vitro GM-CSF generated murine MDSCs from wild-type (WT) and CD49a/α1-integrin (Itga1-/-) gene-deficient mice. Here, we found that effector (Teff) but not naive (Tn) CD4+ T cells express VLA-1 and monocytes further up-regulated their expression after culture in GM-CSF when they differentiated into the monocytic subset of resting MDSCs (R-MDSCs). Subsequent activation of R-MDSCs by LPS+IFN-γ (A-MDSCs) showed increased in vitro suppressor potential, which was independent of VLA-1. Surprisingly, VLA-1 deficiency did not influence A-MDSC motility or migration on collagen IV in vitro. However, interaction times of Itga1-/- A-MDSCs with Teff were shorter than with WT A-MDSCs on collagen IV but not on fibronectin substrate in vitro. After injection, A-MDSCs homed to the splenic red pulp where they co-localized with Teff and showed immediate suppression already after 6 h as shown by inhibition of T cell proliferation and induction of apoptosis. Injection of A-MDSCs from Itga1-/- mice showed equivalent homing into the spleen but a reduced suppressive effect. Interaction studies of A-MDSCs with Teff in the subcapsular red pulp with intravital two-photon microscopy revealed also here that MDSC motility and migration parameters were not altered by VLA-1 deficiency, but the interaction times with Teff were reduced. Together, our data point to a new role of VLA-1 adhesion to collagen IV as a prerequisite for extended contact times with Teff required for suppression.

The antitumor efficacy of monomeric disintegrin obtustatin in S-180 sarcoma mouse model.

Obtustatin, isolated from the Levantine Viper snake venom (Macrovipera lebetina obtusa -MLO), is the shortest known monomeric disintegrin shown to specifically inhibit the binding of the α1ß1 integrin to collagen IV. Its oncostatic effect is due to the inhibition of angiogenesis, likely through α1ß1 integrin inhibition in endothelial cells. To explore the therapeutic potential of obtustatin, we studied its effect in S-180 sarcoma-bearing mice model in vivo as well as in human dermal microvascular endothelial cells (HMVEC-D) in vitro, and tested anti-angiogenic activity in vivo using the chick embryo chorioallantoic membrane assay (CAM assay). Our in vivo results show that obtustatin inhibits tumour growth by 33%. The expression of vascular endothelial growth factor (VEGF) increased after treatment with obtustatin, but the level of expression of caspase 8 did not change. In addition, our results demonstrate that obtustatin inhibits FGF2-induced angiogenesis in the CAM assay. Our in vitro results show that obtustatin does not exhibit cytotoxic activity in HMVEC-D cells in comparison to in vivo results. Thus, our findings disclose that obtustatin might be a potential candidate for the treatment of sarcoma in vivo with low toxicity.

RK, the first scorpion peptide with dual disintegrin activity on α1ß1 and αvß3 integrins.

Scorpion peptides are well known for their pharmaceutical potential on different targets. These include mainly the ion channels which were found to be highly expressed in many diseases, including cancer, auto-immune pathologies and Alzheimer. So far, however, the disintegrin activity had only been characterized for snake venom molecules. Herein, we present the first short peptide, purified from the venom of Buthus occitanus tunetanus, (termed RK) able to inhibit the cell adhesion of Glioblastoma, Melanoma and Rat pheochromocytoma to different extracellular matrix (ECM) receptors. Anti-integrin antibody assay suggests that RK interacts with both α1ß1 and αvß3 with a more pronounced effect for the former. The examination of the primary structure of RK suggests the involvement of two motifs: KSS, analogue to KTS which was characterized for α1ß1 Snake venom disintegrins, and ECD, analogue to RGD which was found to be active on αvß3. To assess their roles in the disintegrin activity of RK, we conducted a computational analysis. The molecular docking study shows that RK involves mainly two segments to interact with the α1ß1 integrin, but the peptide does not implicate the KSS motif in the interaction. The molecular modeling study, suggests the key contribution of the ECD segment in the interaction with αvß3 integrin.

Magnesium Activates Microsecond Dynamics to Regulate Integrin-Collagen Recognition.

Integrin receptors bind collagen via metal-mediated interactions that are modulated by magnesium (Mg2+) levels in the extracellular matrix. Nuclear magnetic resonance-based relaxation experiments, isothermal titration calorimetry, and adhesion assays reveal that Mg2+ functions as both a structural anchor and dynamic switch of the α1ß1 integrin I domain (α1I). Specifically, Mg2+ binding activates micro- to millisecond timescale motions of residues distal to the binding site, particularly those surrounding the salt bridge at helix 7 and near the metal ion-dependent adhesion site. Mutagenesis of these residues impacts α1I functional activity, thereby suggesting that Mg-bound α1I dynamics are important for collagen binding and consequent allosteric rearrangement of the low-affinity closed to high-affinity open conformation. We propose a multistep recognition mechanism for α1I-Mg-collagen interactions involving both conformational selection and induced-fit processes. Our findings unravel the multifaceted role of Mg2+ in integrin-collagen recognition and assist in elucidating the molecular mechanisms by which metals regulate protein-protein interactions.

Targeting α1 inserted domain (I) of α1ß1 integrin by Lebetin 2 from M. lebetina transmediterranea venom decreased tumorigenesis and angiogenesis.

Through the recent development of knowledge in biotechnology and bioinformatics, snake venoms are widely used to develop new drugs to treat diseases such as hypertension and cancer. We have previously reported that Lebetin 2 isolated from Macrovipera lebetina transmediterranea venom displays a potent anti-platelet activity and exerts a cardioprotective effect in ischemia-reperfusion (IR) injury model. Here, we report that Lebetin 2 possess an anti-tumor effect by targeting the integrin receptor function. It was thus able to inhibit both adhesion and migration of pheochromocytoma cells (PC12) and α1ß1 integrin-expressing CHO cells (CHO-α1) to type I and IV collagens. Moreover, this peptide affects proliferation of PC12 cells by modulating AKT phosphorylation. Furthermore, Lebetin 2 exhibits a potent anti-angiogenic effect as assessed in vitro and ex vivo, using both the embryo chick chorioallantoic membrane model (CAM) and rat aortic ring assay. Interestingly, the interaction mode of Lebetin 2 with the integrin α1ß1, assessed in silico, showed that the peptide represents a steric obstruction preventing the collagen from enforcing the interactions with the integrin.

Pulmonary antigen encounter regulates the establishment of tissue-resident CD8 memory T cells in the lung airways and parenchyma.

Resident memory CD8 T (TRM) cells in the lung parenchyma (LP) and airways provide heterologous protection against influenza virus challenge. However, scant knowledge exists regarding factors necessary to establish and maintain lung CD8 TRM. Here we demonstrate that, in contrast to mechanisms described for other tissues, airway, and LP CD8 TRM establishment requires cognate antigen recognition in the lung. Systemic effector CD8 T cells could be transiently pulled into the lung in response to localized inflammation, however these effector cells failed to establish tissue residency unless antigen was present in the pulmonary environment. The interaction of effector CD8 T cells with cognate antigen in the lung resulted in increased and prolonged expression of the tissue-retention markers CD69 and CD103, and increased expression of the adhesion molecule VLA-1. The inability of localized inflammation alone to establish lung TRM resulted in decreased viral clearance and increased mortality following heterosubtypic influenza challenge, despite equal numbers of circulating memory CD8 T cells. These findings demonstrate that pulmonary antigen encounter is required for the establishment of lung CD8 TRM and may inform future vaccine strategies to generate robust cellular immunity against respiratory pathogens.

Semaphorin7A aggravates coxsackievirusB3-induced viral myocarditis by increasing α1ß1-integrin macrophages and subsequent enhanced inflammatory response.

Semaphorin7A (Sema7A) has been reported to play various roles in nerve axon growth, tumor suppression, and tissue remodeling, as well as regulation of intestinal inflammation diseases. Viral myocarditis (VMC) characterized by viral-myocardial-cell necrosis and inflammatory cell infiltration is a common clinical disease of the cardiovascular system. However, the role of Sema7A in coxsackievirus B3 (CVB3)-induced VMC has not been reported. In this study, we generated an acute VMC mouse model by CVB3 infection, and manipulated Sema7A expression by in vivo polyethyleneimine-mediated Sema7A down-regulation. Our results indicated that Sema7A was up-regulated in cardiomyocytes during VMC, and that Sema7A down-regulation following short hairpin RNA interference or mAb neutralization effectively protected mice from VMC. Additionally, reduced inflammatory responses were observed along with Sema7A down-regulation. Furthermore, adoptive transfer of α1ß1-integrin macrophages exacerbated CVB3-induced myocarditis, suggesting the significance of α1ß1-integrin macrophages in response to VMC. We observed that co-culture of neonatal myocardiocytes with macrophages increased the percentage of α1ß1-integrin macrophages, while Sema7A neutralization reduced α1ß1-integrin macrophages in heart tissue of VMC mice. These results demonstrated that Sema7A, as an inflammation regulator in CVB3-induced VMC, might interact with α1ß1-integrin in macrophages to enhance the inflammatory response and aggravate disease severity. Our findings provided insight into the potential role of Sema7A as a therapeutic treatment for VMC.

ANGPTL1 Interacts with Integrin α1ß1 to Suppress HCC Angiogenesis and Metastasis by Inhibiting JAK2/STAT3 Signaling.

Downregulation of tumor suppressor signaling plays an important role in the pathogenesis of hepatocellular carcinoma (HCC). Here, we report that downregulation of the angiopoietin-like protein ANGPTL1 is associated with vascular invasion, tumor thrombus, metastasis, and poor prognosis in HCC. Ectopic expression of ANGPTL1 in HCC cells effectively decreased their in vitro and in vivo tumorigenicity, cell motility, and angiogenesis. shRNA-mediated depletion of ANGPTL1 exerted opposing effects. ANGPTL1 promoted apoptosis via inhibition of the STAT3/Bcl-2-mediated antiapoptotic pathway and decreased cell migration and invasion via downregulation of transcription factors SNAIL and SLUG. Furthermore, ANGPTL1 inhibited angiogenesis by attenuating ERK and AKT signaling and interacted with integrin α1ß1 receptor to suppress the downstream FAK/Src-JAK-STAT3 signaling pathway. Taken together, these results suggest ANGPTL1 as a prognostic biomarker and novel therapeutic agent in HCC. Cancer Res; 77(21); 5831-45. ©2017 AACR.

Expression and role of VLA-1 in resident memory CD8 T cell responses to respiratory mucosal viral-vectored immunization against tuberculosis.

Lung resident memory T cells (TRM) characterized by selective expression of mucosal integrins VLA-1 (α1ß1) and CD103 (αEß7) are generated following primary respiratory viral infections. Despite recent progress, the generation of lung TRM and the role of mucosal integrins following viral vector respiratory mucosal immunization still remains poorly understood. Here by using a replication-defective viral vector tuberculosis vaccine, we show that lung Ag-specific CD8 T cells express both VLA-1 and CD103 following respiratory mucosal immunization. However, VLA-1 and CD103 are acquired in differential tissue sites with the former acquired during T cell priming in the draining lymph nodes and the latter acquired after T cells entered the lung. Once in the lung, Ag-specific CD8 T cells continue to express VLA-1 at high levels through the effector/expansion, contraction, and memory phases of T cell responses. Using a functional VLA-1 blocking mAb, we show that VLA-1 is not required for trafficking of these cells to the lung, but it negatively regulates them in the contraction phase. Furthermore, VLA-1 plays a negligible role in the maintenance of these cells in the lung. Our study provides new information on vaccine-inducible lung TRM and shall help develop effective viral vector respiratory mucosal tuberculosis vaccination strategies.

The binding capacity of α1ß1-, α2ß1- and α10ß1-integrins depends on non-collagenous surface macromolecules rather than the collagens in cartilage fibrils.

Interactions of cells with supramolecular aggregates of the extracellular matrix (ECM) are mediated, in part, by cell surface receptors of the integrin family. These are important molecular components of cell surface-suprastructures regulating cellular activities in general. A subfamily of ß1-integrins with von Willebrand-factor A-like domains (I-domains) in their α-chains can bind to collagen molecules and, therefore, are considered as important cellular mechano-receptors. Here we show that chondrocytes strongly bind to cartilage collagens in the form of individual triple helical molecules but very weakly to fibrils formed by the same molecules. We also find that chondrocyte integrins α1ß1-, α2ß1- and α10ß1-integrins and their I-domains have the same characteristics. Nevertheless we find integrin binding to mechanically generated cartilage fibril fragments, which also comprise peripheral non-collagenous material. We conclude that cell adhesion results from binding of integrin-containing adhesion suprastructures to the non-collagenous fibril periphery but not to the collagenous fibril cores. The biological importance of the well-investigated recognition of collagen molecules by integrins is unknown. Possible scenarios may include fibrillogenesis, fibril degradation and/or phagocytosis, recruitment of cells to remodeling sites, or molecular signaling across cytoplasmic membranes. In these circumstances, collagen molecules may lack a fibrillar organization. However, other processes requiring robust biomechanical functions, such as fibril organization in tissues, cell division, adhesion, or migration, do not involve direct integrin-collagen interactions.