[Effect of epidermal growth factor signal pathway on integrin alpha5 beta1 in prostate cancer cell line DU145].

OBJECTIVE: To investigate the molecular mechanism of epidermal growth factor (EGF) signal pathway on the expression of integrin alpha5 beta1 in prostate cancer cell line DU145. METHODS: Using flow-cytometry, the effects of EGF and the mitogen-activated protein kinase (MAPK) signal pathway inhibitor PD98059 on the expression of integrin alpha5 and beta1 subunits on DU145 cell surface were analyzed. RT-PCR and Western blot methods were used to examined the expression of mRNA and cell total protein of integrin alpha5 and beta1 subunits. And the metastatic phenotypes in DU145 cell were investigated. RESULTS: The expression levels of integrin alpha5 beta1, which was the receptor for fibronectin, were changed. EGF up-regulated the protein and mRNA expression of beta1 subunit on DU145 cell surface, 231% and 248% (P < 0.01) compared to the control respectively, and it could significantly promote the ability of DU145 cell adhesion to fibronectin and migration. However PD98058, which was the inhibitor of MAPK signal pathway, down-regulated the protein and mRNA expression of beta1 subunit, 60% and 63% (P < 0.01) compared to the control respectively, and it had the contrary function on the adhesion and migration ability of DU145 cell. But both had no effect the expression of alpha5 subunit. CONCLUSIONS: EGF might promote the metastatic ability mainly by up-regulating the expression of beta1 subunit by activating MAPK signal pathway in DU145 cells. Their regulation effects are on the mRNA transcriptional level.

Characterization of integrin and anchorage dependence in mammary epithelial cells following c-erbB2-induced epithelial-mesenchymal transition.

Signalling from the proto-oncogene c-erbB2 in mammary epithelial cells has earlier been shown to result in epithelial-mesenchymal transition (EMT) giving rise to fibroblast-like cells, and acquisition of anchorage-independent growth (AIG) usually determined by growth capacity in soft agar. In this study, we have analysed AIG associated with c-erbB2-induced EMT in a human mammary epithelial cell line. Intriguingly, cells capable of growth in soft agar were shown to be dependent on the function of beta(1) integrin extracellular matrix receptors for growth in collagen. We therefore tested the hypothesis that apparent AIG was due to deposition of extracellular matrix in the agar. Although the fibroblastic cells had strongly upregulated expression of the fibronectin receptor subunit integrin alpha(5) andabundant fibronectin fibrils, these properties did not have a positive correlation with AIG. Furthermore, antibody blocking of integrin alpha(5) and beta(1) failed to inhibit AIG. These results indicate that the anchorage-independent cells are not dependent on connection to extracellular matrix, but instead may be subject to a growth-inhibitory effect from the collagen in the absence of integrin signalling. This notion was supported by the finding that integrin blocking of the fibroblastic cells in fibrin was without effect on proliferation.

Sensory neuron subtypes have unique substratum preference and receptor expression before target innervation.

The factors controlling the specification and subsequent differentiation of sensory neurons are poorly understood. Data from embryological manipulations suggest that either sensory neuron fates are specified by the targets they encounter or sensory neurons are considerably more "plastic" with respect to specification than are neurons of the CNS. The prevailing view that sensory neurons are specified late in development is not consistent, however, with the directed outgrowth of sensory neurons to their targets and the characteristic spatial distribution of sensory neuron fates within the peripheral ganglia. To address when in development different classes of sensory neurons can first be distinguished, we investigated the interactions of early dorsal root ganglia neurons with the extracellular matrix before neurite outgrowth to targets. We found that subclasses of sensory neurons in early dorsal root ganglia show different patterns of neurite outgrowth and integrin expression that are predictive of their fates. In the absence of neurotrophins, presumptive proprioceptive neurons extend neurites robustly on both laminin and fibronectin, whereas presumptive cutaneous neurons show a strong preference for laminin. Cutaneous afferents that have innervated targets show a similar strong preference for laminin and show higher levels of integrin alpha7beta1 than do proprioceptive neurons. Finally, presumptive proprioceptive neurons express fibronectin receptors, integrin alpha3beta1, alpha4beta1, and alpha5beta1, at higher levels than do presumptive cutaneous neurons. Our results indicate that subtypes of sensory neurons have unique patterns of neurite outgrowth and receptor expression before target innervation.

The alpha5beta1 integrin selectively enhances epidermal growth factor signaling to the phosphatidylinositol-3-kinase/Akt pathway in intestinal epithelial cells.

We have investigated EGF-driven signaling processes in rat intestinal epithelial cell lines that overexpress either the alpha5beta1 integrin or the alpha2beta1 integrin. Both cell types display efficient activation of Erk/MAP kinase, but only the alpha5beta1 expressing cells display a strong activation of Akt. A complex is formed between activated EGFR and alpha5beta1, but not with alpha2beta1; this complex also contains ErbB3 and p85. Thus alpha5beta1 can support efficient activation of both the Erk and the phosphatidylinositol-3-kinase/Akt branches of the EGFR signaling cascade, whereas alpha2beta1 can support only the Erk branch.

Stromal-derived factor-1 in human tumors recruits and alters the function of plasmacytoid precursor dendritic cells.

Dendritic-cell (DC) trafficking and function in tumors is poorly characterized, with studies confined to myeloid DCs (DC1s). Tumors inhibit DC1 migration and function, likely hindering specific immunity. The role of plasmacytoid DCs (DC2s) in tumor immunity is unknown. We show here that malignant human ovarian epithelial tumor cells express very high levels of stromal-derived factor-1, which induces DC2 precursor (preDC2) chemotaxis and adhesion/transmigration, upregulates preDC2 very late antigen (VLA)-5, and protects preDC2s from tumor macrophage interleukin-10-induced apoptosis, all through CXC chemokine receptor-4. The VLA-5 ligand vascular-cell adhesion molecule-1 mediated preDC2 adhesion/transmigration. Tumor preDC2s induced significant T-cell interleukin-10 unrelated to preDC2 differentiation or activation state, and this contributed to poor T-cell activation. Myeloid precursor DCs (preDC1s) were not detected. Tumors may weaken immunity by attracting preDC2s and protecting them from the harsh microenvironment, and by altering preDC1 distribution.

Interference of tenascin-C with syndecan-4 binding to fibronectin blocks cell adhesion and stimulates tumor cell proliferation.

Tenascin-C is an adhesion-modulatory extracellular matrix molecule that is highly expressed in tumors. To investigate the effect of tenascin-C on tumor cells, we analyzed its antiadhesive nature and effect on tumor cell proliferation in a fibronectin context. Glioblastoma and breast carcinoma cell adhesion was compromised by a mixed fibronectin/tenascin-C substratum, which concomitantly caused increased tumor-cell proliferation. We identified the antiadhesive mechanism as a specific interference of tenascin-C with cell binding to the HepII/syndecan-4 site in fibronectin through direct binding of tenascin-C to the 13th fibronectin type III repeat (FNIII13). Cell adhesion and proliferation levels were restored by the addition of FNIII13. Overexpression of syndecan-4, but not syndecan-1 or -2, reverted the cell adhesion defect of tenascin-C. We characterized FNIII13 as the binding site for syndecan-4. Thus we describe a novel mechanism by which tenascin-C impairs the adhesive function of fibronectin through binding to FNIII13, thereby inhibiting the coreceptor function of syndecan-4 in fibronectin-induced integrin signaling.

Impairment of gingival fibroblast adherence by IL-6/sIL-6R.

Interleukin-6 (IL-6) binds to human gingival fibroblasts (HGF) in the presence of a soluble form of IL-6 receptor (sIL-6R). We investigated the effects of IL-6 on the functions of HGF in the presence of sIL-6R. HGF changed their morphology from spindle-shaped to round, and detached from the culture dish by stimulation with IL-6/sIL-6R. In this condition, a signal transducer gp130 and a transcription factor Stat3 were phosphorylated, resulting in activation of transcription factors Stat3 and C/EBPbeta. Cytoskeletal beta-actin and adhesion molecule integrin-alpha5, a subunit of alpha5beta1 integrin (VLA-5), were found to possess potential binding domains for these transcription factors in their promoters. Accumulation of beta-actin and integrin-alpha5 mRNA decreased, contrary to the expectation of the induction of gene transcription. Furthermore, the decrease in their mRNAs was associated with reduced expression of both actin and VLA-5 proteins. These results suggest that the expression of VLA-5 and actin was down-regulated in HGF through an IL-6 signaling pathway, resulting in impairment of HGF adherence.

Adult neuronal regeneration induced by transgenic integrin expression.

In a variety of adult CNS injury models, embryonic neurons exhibit superior regenerative performance when compared with adult neurons. It is unknown how young neurons extend axons in the injured adult brain, in which adult neurons fail to regenerate. This study shows that cultured adult neurons do not adapt to conditions that are characteristic of the injured adult CNS: low levels of growth-promoting molecules and the presence of inhibitory proteoglycans. In contrast, young neurons readily adapt to these same conditions, and adaptation is accompanied by an increase in the expression of receptors for growth-promoting molecules (receptors of the integrin family). Surprisingly, the regenerative performance of adult neurons can be restored to that of young neurons by gene transfer-mediated expression of a single alpha-integrin.

Tumour necrosis factor-alpha provokes upregulation of alpha2beta1 and alpha5beta1 integrins, and cell migration in OST osteosarcoma cells.

OST cells enhance the induction of matrix metalloproteinase (MMP)-9 by tumour necrosis factor (TNF)-alpha and the corresponding metastasis to lungs in vivo (Kawashima et al., 1994). We focused on the adhesive and migratory properties of OST cells, and investigated the expression of integrins in OST cells stimulated by TNFalpha in vitro. OST cells potentiated not only adhesion to the extracellular matrix (ECM) but also the migration on ECM. On competitive reverse transcription-polymerase chain reaction (RT-PCR) analyses, the amounts of alpha2 (4.9-fold), alpha5 (1.2-fold) and alpha(v) (4.9-fold) were upregulated by TNFalpha at the transcriptional level. Alpha-5 showed a slight increase by flow cytometry; however, alpha2 and alphav integrins remained unchanged at the protein level. Immunofluorescence study disclosed integrins of alpha2beta1 and alpha5beta1 were much clustered at cell processes by TNFalpha stimulation, probably related to increased cell adhesion and migration. Therefore, the upregulation of alpha2beta1 and alpha5beta1 integrins seems to contribute to tumour invasion and metastatic potential.

[Quantitative expression of the adhesion receptors VLA-4, VLA-5, L-selectin, MAC-1, and ICAM-1 on the surface of CD34+ cells]. / Expression quantitative des récepteurs d'adhérence VLA-4, VLA-5, L-sélectine, MAC-1, et ICAM-1 à la surface des cellules CD34+.

The aim of this work was to quantify by flow cytometry the main adhesion receptors on CD34+ cells. These cells were isolated from bone marrow (BM) or mobilized peripheral blood (PB). The proportions of CD34+/CD49d+ and CD34+/CD49e+ are weaker on PB cells, without quantitative expression variation. This phenotypic variation may induce CD34+ cells exist from BM into circulation, promoting the mobilization. The homing to the BM implicate the CD62L receptor, which expression was found more frequently and stronger on PB cells than on BM. The CD11b, CD18 and CD54 receptors are implicated in CD34+ cells adhesion to BM micro-environment. No significant variation in CD34+/CD11b+ and CD34+/CD18+ cells frequency was noted. Moreover, CD54 receptor was more frequently expressed on PB cells. Quantitative analysis revealed that CD18 was more strongly expressed on BM than on PB cells. This quantitative variation could promote progenitor adhesion by interacting with stromal cells. Finally, quantitative expression of the main receptors on CD34+ cells provides an original option for studying CD34+ cells during the mobilization, the homing or the adhesion to BM micro-environment.

[The adhesive and migrating function of human epithelial cells opsonized by fibronectin].

OBJECTIVE: To investigate the adhesive and migrating function of human epithelial keratiocytes during in vitro culture. METHODS: The adhesive and migrating function was determined in freshly isolated EKCs and those cultured for 7 similar 10 days after being opsonized by fibronectin (FN). The expressions of alpha(5)beta(1) receptors in EKC were examined with indirect immunofluorescence staining methods before and after culture. RESULTS: (1) The adhesive and migrating indices after FN opsonization of EKCs freshly isolated EKCs were obviously lower than those cultured for 7 similar 10 days (P < 0.01). (2) There exhibited positive staining of the expression of alpha(5)beta(1) receptors in the EKC after 1 day culture and in the proliferative epithelia after in vitro culturing of a tissue mass. While there exhibited strongly positive staining of the peripheral proliferative epithelia of the EKCs and tissue mass after 7 days of culture, negative or weakly positive stainings were found in freshly isolated EKCs. CONCLUSION: (1) There existed significant difference of biological function between the freshly isolated EKCs and those cultured for 7 similar 10 days. (2) The strongest expression of alpha(5)beta(1) receptors was observed at the active proliferative site (peripheral proliferative epithelia of a tissue mass) and in the biological active period (cultured for 7 days) of EKCs. (3) The adhesive and migrating function of EKCs could be effectively induced and activated by in vitro culture, which enabled the EKC to alter from a relatively biologically functional static state to an actively functional state.

Integrin modulating factor: a 30-kD protein that modulates the expression and function of alpha5beta1 integrin receptor.

The integrin family of cell surface receptors consists of transmembrane glycoproteins involved in cellular morphology, cytoarchitecture, cell-cell, and cell-extracellular matrix interaction. Changes in integrin receptor expression are associated with malignant transformation. The adhesion promoting activity of several members of the integrin receptors may be modulated. Integrin Associated Proteins and integrin modulating factor that may modulate integrin receptors expression and function have been reported. In this article, we report the identification of a 30-kD protein produced in SiHa cell culture medium that can modulate the expression and function of alpha5beta1 integrin receptor in HeLaS3 cells. The cell adhesion assay clearly demonstrated that HeLaS3 cells grown in a serum-free culture medium of SiHa cells (fresh medium: culture medium = 3:1) stimulated the ligand binding activity of alpha5beta1 receptor to fibronectin in a time-dependent manner, having a peak activity at 72 hours of culture. Immunocytochemical localization showed a very high expression of alpha5beta1 receptor in HeLaS3 cells grown in a SiHa culture medium for 72 hours. The (NH4)2SO4 fractionation demonstrated that proteins present in 80-100% (NH4)2SO4 saturated fraction of serum-free SiHa culture medium have a significant stimulatory effect on the binding of HeLaS3 cells to fibronectin ligand via the alpha5beta1 integrin receptor. High pressure liquid chromatography (HPLC) separation of 80-100% (NH4)2SO4 saturated fraction showed a 30- kD protein in polyacrylamide gel electrophoresis (PAGE) analysis that has a maximum stimulatory effect on the binding of HeLaS3 cells to fibronectin ligand via the alpha5beta1 integrin receptor. In conclusion, our observations indicated that human cervical tumor cells SiHa produce a 30-kD protein that can modulate the expression and function of alpha5beta1 fibronectin integrin receptor of HeLaS3 cells. These findings strengthen the concept that some cellular proteins, also called Integrin Associated Protein, may regulate the integrin receptor expression and function. Studies are in progress to characterize this 30-kD integrin modulating factor and its role in the regulation of integrin receptor function.

Human hematopoietic stem cells stimulated to proliferate in vitro lose engraftment potential during their S/G(2)/M transit and do not reenter G(0).

An understanding of mechanisms regulating hematopoietic stem cell engraftment is of pivotal importance to the clinical use of cultured and genetically modified transplants. Human cord blood (CB) cells with lymphomyeloid repopulating activity in NOD/SCID mice were recently shown to undergo multiple self-renewal divisions within 6 days in serum-free cultures containing Flt3-ligand, Steel factor, interleukin 3 (IL-3), IL-6, and granulocyte colony-stimulating factor. The present study shows that, on the fifth day, the transplantable stem cell activity is restricted to the G(1) fraction, even though both colony-forming cells (CFCs) and long-term culture-initiating cells (LTC-ICs) in the same cultures are approximately equally distributed between G(0)/G(1) and S/G(2)/M. Interestingly, the G(0) cells defined by their low levels of Hoechst 33342 and Pyronin Y staining, and reduced Ki67 and cyclin D expression (representing 21% of the cultured CB population) include some mature erythroid CFCs but very few primitive CFCs, LTC-ICs, or repopulating cells. Although these findings suggest a cell cycle-associated change in in vivo stem cell homing, the cultured G(0)/G(1) and S/G(2)/M CD34(+) CB cells exhibited no differences in levels of expression of VLA-4, VLA-5, or CXCR-4. Moreover, further incubation of these cells for 1 day in the presence of a concentration of transforming growth factor beta(1) that increased the G(0)/G(1) fraction did not enhance detection of repopulating cells. The demonstration of a cell cycle-associated mechanism that selectively silences the transplantability of proliferating human hematopoietic stem cells poses both challenges and opportunities for the future improvement of ex vivo-manipulated grafts. (Blood. 2000;96:4185-4193)

Regulation of fibronectin matrix assembly by activated Ras in transformed cells.

Fibronectin extracellular matrix plays a critical role in the microenvironment of cells. Loss of this matrix frequently accompanies oncogenic transformation, allowing changes in cell growth, morphology, and tissue organization. The HT1080 human fibrosarcoma cell line is deficient in formation of fibronectin matrix fibrils but assembly can be induced by the glucocorticoid dexamethasone. Here we show that fibronectin assembly can also be restored by stimulation of alpha5beta1 integrin with activating antibody or with Mn2+ suggesting that integrin activity is reduced in these cells. While dexamethasone promoted actin stress fiber formation, actin filaments remained cortical following Mn2+ treatment showing that the dexamethasone effect is not due solely to cytoskeletal changes. HT1080 cells have one activated allele of N-ras and PD98059 inhibition of signaling from Ras through ERK increased fibronectin matrix accumulation. Conversely, the p38 MAP kinase inhibitor SB203580 blocked induction of matrix and increased ERK phosphorylation. Thus, two MAP kinase pathways contribute to the control of integrin-mediated fibronectin assembly. ERK activity and fibronectin assembly were linked in three different ras-transformed cell lines but not in SV40- or RSV-transformed cells indicating that oncogenic Ras uses a distinct mechanism to down-regulate cell-fibronectin interactions.

Expression of integrin alpha5beta1 and MMPs associated with epithelioid morphology and malignancy of uveal melanoma.

PURPOSE: Altered expression of the alpha5beta1 integrin and matrix metalloproteinases (MMPs) is recognized as a hallmark of invasive tumor cells. The purpose of the present study was to investigate the expression of integrin subunit alpha5, its corresponding ligand fibronectin (FN), and the expression pattern for MMPs in four highly proliferative human choroidal melanomas (TP17, TP31, SP8.0, and SP6.5) to evaluate whether any correlation can be established between these markers and cell tumorigenicity. METHODS: Cell tumorigenicity was evaluated by subcutaneous injection of uveal melanoma cell lines in immunodeficient nude mice. Anchorage dependency was evaluated by growth assays in soft agar. The invasive ability of each cell type was also determined using a modified Boyden chamber. Expression of both the alpha5 integrin subunit and FN was determined at the mRNA level by RT-PCR. The protein level (for alpha5) was determined by flow cytometry and inhibition of adhesion assays by using an antibody directed against the alpha5 subunit. Expression of MMPs was determined by standard gelatin zymography. RESULTS: Assays in nude mice provided evidence that the cell lines possess a range of tumorigenic ability of TP17>TP31>SP8.0>SP6.5. Antibody inhibition of cell adhesion and flow cytometry demonstrated that TP17 cells have no detectable membrane-bound alpha5beta1, whereas low levels are found in primary cultured melanocytes, as well as in SP6.5, SP8.0, and TP31 cells. RT-PCR analyses provided evidence that both FN and alpha5 expression may be regulated at the transcriptional level. Gelatin zymography revealed that all cell lines, as well as normal melanocytes, express MMP-2 at varying levels but that only the highly invasive TP17 cell line secretes a distinctive MMP with a high molecular weight of 117 kDa. CONCLUSIONS: Among the four melanoma cell lines selected for the completion of this study, TP17 exhibited the most aggressive phenotype, which also correlated with the mostly epithelioid morphology of these cells. The cell morphology of the TP17 cell line could be related to the loss of alpha5beta1, whereas its invasive properties are more likely related to the expression of the 117-kDa MMP.

Over expression of integrin alpha 5 beta 1 in human hepatocellular carcinoma cell line suppresses cell proliferation in vitro and tumorigenicity in nude mice.

Integrin alpha 5 beta 1 and alpha 2 beta 1 are the major integrin receptors in human hepatocytes. However, in human hepatocellular carcinoma cells it was found that the expression of integrin alpha 5 beta 1 was decreased and another integrin alpha 6 beta 1 increased. In this study, the SMMC7721 human hepatocellular carcinoma cells cotransfected or singly transfected with integrin alpha 5 and/or beta 1 cDNAs were established, and designated alpha 5 beta 1.6-7721, alpha 5.3-7721, and beta 1.6-7721 cell lines, respectively. Transfection with cDNAs of integrin alpha 5 and beta 1 subunits resulted in the over expression of each integrin and modified biological properties, including a slowed growth rate, changes in the cell cycle from 15.5% of control cells in the G2/M phase to 12.1%, 9.6% and 9.4% in alpha 5.3-7721, beta 1.6-7721, alpha 5 beta 1.6-7721, respectively, and a decrease in the Cell Mitosis Index from 1.6 in controls to 0.96, 0.95, and 0.72, and 34%, 28% and 52% derived from colony forming ability, respectively. Tumorigenicity was also tested in nude mice with inoculation of cells subcutaneously. Tumor masses growing in nude mice following inoculation with beta 1.6-7721, and alpha 5 beta 1.6-7721 cells weighed only 52% or 31% those of control cells. These results indicated that deletion or low expression of integrin alpha 5 beta 1 may play an important role in the development of hepatocellular carcinoma. Therefore, induction of expression of the integrin alpha 5 beta 1 in malignant cells could be a potential means of treating hepatocellular carcinoma.

TGF-beta 1 modulated the expression of alpha 5 beta 1 integrin and integrin-mediated signaling in human hepatocarcinoma cells.

Integrins are a family of cell surface adhesion molecules which mediate cell adhesion and initiate signaling pathways that regulate cell spreading, migration, differentiation, and proliferation. TGF-beta is a multifunctional factor that induces a wide variety of cellular processes. In this study, we show that, TGF-beta 1 treatment enhanced the amount of alpha 5 beta 1 integrin on cell surface, the mRNA level of alpha 5 subunit, and subsequently stimulated cell adhesion onto a fibronectin (Fn) and laminin (Ln) matrix in SMMC-7721 cells. TGF-beta 1 could also promote cell migration. Furthermore, our results showed that TGF-beta1 treatment stimulated the tyrosine phosphorylation level of FAK, which can be activated by the ligation and clustering of integrins. PTEN can directly dephosphorylate FAK, and the results that TGF-beta 1 could down-regulate PTEN at protein level suggested that TGF-beta 1 might stimulate FAK phosphorylation through increasing integrin signaling and reducing dephosphorylation of FAK. These studies indicated that TGF-beta 1 and integrin-mediated signaling act synergistically to enhance cell adhesion and migration and affect downstream signaling molecules of hepatocarcinoma cells.

alpha5beta1 integrin protects intestinal epithelial cells from apoptosis through a phosphatidylinositol 3-kinase and protein kinase B-dependent pathway.

Renewal of the gastrointestinal epithelium involves a coordinated process of terminal differentiation and programmed cell death. Integrins have been implicated in the control of apoptotic processes in various cell types. Here we examine the role of integrins in the regulation of apoptosis in gastrointestinal epithelial cells with the use of a rat small intestinal epithelial cell line (RIE1) as a model. Overexpression of the integrin alpha5 subunit in RIE1 cells conferred protection against several proapoptotic stimuli. In contrast, overexpression of the integrin alpha2 subunit had no effect on cell survival. The antiapoptotic effect of the alpha5 subunit was partially retained by a mutated version that had a truncation of the cytoplasmic domain. The antiapoptotic effects of the full-length or truncated alpha5 subunit were reversed upon treatment with inhibitors of phosphatidylinositol 3-kinase (PI-3-kinase), suggesting that the alpha5beta1 integrin might interact with the PI-3-kinase/Akt survival pathway. When cells overexpressing alpha5 were allowed to adhere to fibronectin, there was a moderate activation of protein kinase B (PKB)/Akt, whereas no such effect was seen in alpha2-overexpressing cells adhering to collagen. Furthermore, in cells overexpressing alpha5 and adhering to fibronectin, there was a dramatic enhancement of the ability of growth factors to stimulate PKB/Akt; again, this was not seen in cells overexpressing alpha2 subunit and adhering to collagen or fibronectin. Expression of a dominant negative version of PKB/Akt in RIE cells blocked to ability of alpha5 to enhance cell survival. Thus, the alpha5beta1 integrin seems to protect intestinal epithelial cells against proapoptotic stimuli by selectively enhancing the activity of the PI-3-kinase/Akt survival pathway.

Expression of beta1-integrins and N-cadherin in bladder cancer and melanoma cell lines.

Changes in the expression of integrins and cadherins might contribute to the progression, invasion and metastasis of transitional cell cancer of the bladder and of melanomas. The expression of alpha5 (P < 0.001), alpha2 and beta1 (P < 0.05 - P < 0.001) integrin subunits in melanoma cells from noncutaneous metastatic sites (WM9, A375) were significantly increased as compared to cutaneous primary tumor (WM35) and metastatic (WM239) cell lines. These differences might be ascribed to the invasive character of melanoma cells and their metastasis to the noncutaneous locations. The significantly heterogeneous expression of beta1 integrin subunit in two malignant bladder cancer cell lines (T24 and Hu456) and nonsignificant differences in the expression of alpha2, alpha3, and alpha5 subunits between malignant and non-malignant human bladder cell lines do not allow an unanimous conclusion on the role of these intergrin subunits in the progression of transitional cancer of bladder. The adhesion molecule, expressed in all studied melanoma and bladder cell lines, that reacted with anti-Pan cadherin monoclonal antibodies was identified as N-cadherin except in the HCV29 non-malignant ureter cell line. However, neither this nor any other bladder or melanoma cell line expressed E-cadherin. The obtained results imply that the replacement of E-cadherin by N-cadherin accompanied by a simultaneous increase in expression of alpha2, alpha3 and alpha5 integrin subunits clearly indicates an increase of invasiveness of melanoma and, to a lesser extent, of transitional cell cancer of bladder. High expression of N-cadherin and alpha5 integrin subunit seems to be associated with the most invasive melanoma phenotype.

Autocrine IL-6 production by human transitional carcinoma cells upregulates expression of the alpha5beta1 firbonectin receptor.

PURPOSE: Studies have demonstrated elevated expression and secretion of IL-6 by transitional cell carcinomas (TCC) following bacillus Calmette-Guerin (BCG) therapy. At present, the role of IL-6 on the biology of TCC is poorly understood. This study evaluated the influence of IL-6 expression on a critical variable regulating BCG-tumor interaction, the tumor expression of alpha5beta1 integrin. MATERIALS AND METHODS: A human TCC cell line (253J) was transfected with an expression vector containing the full-length IL-6 cDNA sequence. Overexpression of IL-6 mRNA and protein was confirmed by Northern analysis and ELISA, respectively. Clones found to overexpress IL-6 were then assayed for alpha5beta1 expression using Northern analysis and flow cytometry. The effect of alterations in alpha5beta1 expression on tumor adherence to fibronectin (FN), and BCG adherence to tumor cells was determined using specific adherence assays. RESULTS: mRNAs for both the alpha5 and beta1 subunits of the FN receptor were increased an average of 9.4 fold and 125.7 fold respectively in the IL-6 overexpressing transfectants relative to the parental 253J cells. Increased mRNA of alpha5 and beta1 was associated with increased cell surface expression of both proteins. Increased protein expression resulted in greater FN substrate binding affinity and increased adherence of BCG to tumor cells. CONCLUSIONS: Autocrine expression of IL-6 upregulates the expression of FN receptor subunits in TCC. Increased alpha5beta1 expression increases cellular adherence to FN, and BCG adherence to tumor cells. These results suggest a role for IL-6 in mediating the antitumor activity of BCG by influencing BCG's adherence to TCC.