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PubMed1997

[Expression of the integrin alpha 5 subunit in hepatocellular carcinoma].

OBJECTIVE: To elucidate the roles of integrins in tumorigenesis, progression, differentiation, invasiveness and metastasis, and the role of the integrin alpha 5 subunit in hepatocellular carcinoma (HCC). METHODS: 79 formalin-fixed sections of hepatocellular carcinoma (HCC) were obtained. The integrin alpha 5 subunit was measured by immunohistochemistry assay (ABC method). RESULTS: In 79 cases of HCC, cancerous tissues had lower expression than their adjacent non-tumor tissues (32.9% vs 81.0%, P < 0.01). The alpha 5 positive rate in small HCC (= < 5 cm in diameter) was higher than large HCC > 10 cm in diameter), being 55.6% and 10.0% respectively. Well-differentiated HCC expressed higher alpha 5 than poorly differentiated ones. The alpha 5 positive rates were lower in highly invasive HCC than those to low invasive ones. CONCLUSION: Our study indicated that expression of the integrin alpha 5 subunit is correlated with growth, differentiation, invasiveness and metastasis of HCC. It is possible that alpha 5 subunit is a negative regulator to these biological parameters of HCC.

Adult↗
PubMed2003

Binding of alpha5 monoclonal antibody to cell surface alpha5beta1 integrin modulates MMP-2 and MMP-7 activity in B16F10 melanoma cells.

UNLABELLED: The integrin multigene family, comprising at least 21 distinct heterodimeric complexes, includes receptors for all major extracellular proteins such as fibronectin, laminin, vitronectin, and collagens. These receptors play important roles in various physiological processes. Matrixmetallo-proteinases (MMPs) are a family of zinc-dependent endopeptidases that degrade the extracellular matrix (ECM) components in physiological conditions. MMPs can degrade all structural components of the ECM, which is one of the critical aspects of tumor cell invasion. The role of matrilysin (MMP-7) in the progression of various carcinomas was recently addressed. Evidence of interplay between integrin function and ECM matrix integrity has come from experiments showing that integrin-ECM contacts can regulate expression and function of MMPs. In this communication, we report on an interesting interrelationship between cell surface alpha5beta1 integrin (fibronectin receptor) and MMPs (MMP-2 and MMP-7). RESULTS: The zymographic analysis showed that ligation of cell surface alpha5beta1 integrin by alpha5 monoclonal antibody leads to the expression and activation of MMP-2 and MMP-7 in B16F10 melanoma cells. The immunoblot confirmed the tyrosine phosphorylation of FAK in B16F10 cells grown in presence of alpha5 monoclonal antibody, indicating the transduction of signal via FAK. The immunocytochemical study demonstrated that alpha5beta1 integrin stimulation causes rearrangement of actin fibers and its accumulation in focal adhesion sites. CONCLUSION: alpha5beta1 integrin-induced expression and activation of MMP-2 and -7 indicates the role of tumor cell surface integrin receptor in the modulation of MMPs and, thereby, the invasive property of tumor cells.

Animals↗
PubMed2004

Expression of integrin subunits alpha2, alpha3, alpha5, alphav, beta1, beta3 and beta4 in different histological types of ameloblastoma compared with dental germ, dental lamina and adult lining epithelium.

OBJECTIVE: To analyze integrin expression and distribution in different histological types of ameloblastoma, compared with dental germ, dental lamina and adult lining epithelium. MATERIALS AND METHODS: Three-micrometer sections from paraffin-embedded specimens were evaluated employing a streptavidin-biotin immunohistochemical method and anti-integrin alpha2, alpha3, alpha5, alphav, beta1, beta3 and beta4 antibodies. RESULTS: All integrins were present in all specimens, exhibiting different patterns. In follicular ameloblastoma, the integrin staining was stronger in the periphery while integrin alpha2 was not present in the central cells. Acanthomatous ameloblastoma showed a similar pattern, with positive staining for integrins alpha3, alpha5, alphav, beta1 and beta4 in the metaplastic cells. In the unicystic, integrin staining was uniform except for integrins alpha5 and beta3 which showed weaker staining in the upper layers. In the plexiform ameloblastoma, dental germ and lamina integrin staining was uniform. In the adult lining epithelium, staining for integrins alpha2, alpha5 and beta4 was confined to the basal layer, while integrins alphav and beta3 were present in the basal and parabasal, with integrins alpha3 and beta1 in the upper layers. CONCLUSION: Acanthomatous, follicular and unicystic ameloblastomas showed integrin staining patterns similar to the adult lining epithelium while the plexiform ameloblastoma was similar to the dental germ and lamina.

Adult↗
PubMed2005

Prostaglandin E2 stimulates fibronectin expression through EP1 receptor, phospholipase C, protein kinase Calpha, and c-Src pathway in primary cultured rat osteoblasts.

Fibronectin (Fn) is involved in the early stages of bone formation, and prostaglandin E (PGE) is an important factor regulating osteogenesis. Here we found that PGE(2) enhanced extracellular Fn assembly in rat primary osteoblasts, as shown by immunofluorescence staining and enzyme-linked immunosorbent assay. PGE(2) also increased the protein levels of Fn by using Western blotting analysis. By using pharmacological inhibitors or activators or genetic inhibition by the EP receptor, antisense oligonucleotides revealed that the EP(1) receptor but not other PGE receptors is involved in PGE(2)-mediated up-regulation of Fn. At the mechanistic level, Ca(2+) chelator (1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester)), phosphatidylinositol-phospholipase C inhibitor (U73122), or Src inhibitor (PP2) attenuated the PGE(2)-induced Fn expression. Protein kinase C (PKC) inhibitor (GF109203X) also inhibited the potentiating action of PGE(2). Furthermore, treatment with antisense oligonucleotides of various PKC isoforms, including alpha, beta, epsilon, and delta, demonstrated that alpha isozyme plays an important role in the enhancement action of PGE(2) on Fn assembly. Flow cytometry and reverse transcription-PCR showed that PGE(2) and 17-phenyl trinor PGE(2) (EP(1)/EP(3) agonist) increased the surface expression and mRNA level of alpha5 or beta1 integrins. Fn promoter activity was enhanced by PGE(2) and 17-phenyl trinor PGE(2) in cells transfected with pGL2F1900-Luc. Cotransfection with dominant negative mutants of PKCalpha or c-Src inhibited the potentiating action of PGE(2) on Fn promoter activity. Local administration of PGE(2) or 17-phenyl trinor PGE(2) into the metaphysis of the tibia via the implantation of a needle cannula significantly increased the Fn and alpha5beta1 integrin immunostaining and bone volume of secondary spongiosa in tibia. Taken together, our results provided evidence that PGE(2) increased Fn and promoted bone formation in rat osteoblasts via the EP(1)/phospholipase C/PKCalpha/c-Src signaling pathway.

Animals↗
PubMed1999

Transcriptional profiling of human microvascular endothelial cells in the proliferative and quiescent state using cDNA arrays.

Endothelial cells are known to be a rich source of transcriptional gene expression. Recent technological advances now permit the detailed profiling of mRNA expression using arrays of known cDNAs on blots. We have used such arrays to examine expression of mRNA by primary isolates of human foreskin microvascular endothelial cells in the proliferative and quiescent state. Cells were stimulated by a combination of known growth factors for these cells including epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), and 'endothelial cell growth supplement (ECGS)' either alone or in combination. Analysis showed the expression of many mRNAs but of the 588 examined, only one, namely monocyte chemotactic protein-1 (MCP-1), showed a decrease on treatment with EGF. A combination of image grabbing followed by subtractive densitometry enabled identification of the mRNAs upregulated in proliferating endothelium. In consideration of the possibility of selective vascular targeting, of particular interest was the increase in expression of the mRNA for the cell surface proteins vascular endothelial (VE-) and neural (N-) cadherin and alpha5, alphav, beta1 and beta3 integrins. The alpha5 integrin offers a previously unrecognized opportunity for vascular targeting.

Journal Article↗
PubMed1999

CD9 is involved in invasion of human trophoblast-like choriocarcinoma cell line, BeWo cells.

The CD9 molecule is expressed on human extravillous trophoblasts, which invade the endometrium during implantation and placentation. To elucidate the role of CD9 in trophoblastic function, we investigated the expression of CD9 protein and mRNA in BeWo cells, a human trophoblast-like choriocarcinoma cell line, using immunohistochemistry, Western blotting and reverse transcription-polymerase chain reaction (RT-PCR). When BeWo cells were cultured with anti-CD9 monoclonal antibodies (mAb), their invasion through the extracellular matrices was significantly enhanced in a dose-dependent manner. Cell proliferation and human chorionic gonadotrophin production were unaffected. On the other hand, culture in the presence of mAb against integrins alpha3, alpha5 and beta1, which partially block the interaction with the extracellular matrices, inhibited BeWo cell invasion. Anti-CD9 monoclonal antibody had a stimulatory effect on BeWo cell invasion in the presence of anti-integrin alpha3 antibody. In contrast, it had no effect in the presence of mAb against integrins alpha5 and beta1, which were also highly expressed on BeWo cells. These findings suggest that CD9 has a function connected with the invasive properties of BeWo cells, which is partially mediated by integrin alpha5beta1. This may relate to the involvement of CD9 in trophoblastic invasion.

Antibodies, Monoclonal↗
PubMed2005

Effect of phase limited inhibition of MyoD expression on the terminal differentiation of bovine myoblasts: no alteration of Myf5 or myogenin expression.

To investigate the roles played by MyoD in the terminal differentiation of satellite cell-derived myoblasts, the effect of antisense inhibition of MyoD expression was examined in bovine adult myoblast culture, in which inhibition treatment was limited to the terminal differentiation phase. MyoD antisense oligonucleotide DNA (AS-mD) suppressed the formation of multinucleated myotubes in the cell culture. Myotube formation was suppressed even when AS-mD treatment was limited to the period preceding the onset of myotube formation. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis revealed that treatment with AS-mD suppressed the expression of myosin heavy chain embryonic isoform and troponin T isoforms at 4 days after the induction of differentiation. AS-mD also suppressed the expression of MRF4, but did not alter the expression of either Myf5 or myogenin, in contrast to previous results using mouse cells possessing MyoD(-/-) genetic background. These findings suggest that MyoD controls myogenesis but not Myf5 or myogenin mRNA expression during the terminal differentiation phase. Furthermore, among the alpha4, alpha5, alpha6, and alpha7 integrins, alpha4, alpha5, and alpha7 integrin expression was suppressed by AS-mD treatment, in parallel with the suppression of myotube formation, which suggests that MyoD controls myotube formation by regulating the expression of alpha4, alpha5, and alpha7 integrins.

Animals↗
PubMed1999

Phenotypical and functional properties of human bone marrow mesenchymal progenitor cells.

Bone marrow stroma provides the microenvironment for hematopoiesis and is also the source of mesenchymal progenitors (mesenchymal or marrow stromal cells [MSC]) that may serve as long-lasting precursors for bone, cartilage, lung, and muscle. While several studies have indicated the differentiation potential of MSC, few studies have been performed on the cells themselves. In an attempt to further expand our knowledge on these cells, we have performed studies on their cell cycle, immuno- and adhesive-phenotype, ex vivo expansion, and differentiation properties. MSC cultures have been initiated from human bone marrow low-density mononuclear cells and maintained in the absence of differentiation stimuli and hematopoietic cells. The homogenous layer of adherent cells thus formed exhibits a typical fibroblastlike morphology, a population doubling time of 33 h, a large expansive potential, and cell cycle characteristics including a subset (20%) of quiescent cells. The antigenic phenotype of MSC is not unique, borrowing features of mesenchymal, endothelial, and epithelial cells. Together, MSC express several adhesion-related antigens, like the integrin subunits alpha4, alpha5, beta1, integrins alphavbeta3 and alphavbeta5, ICAM-1, and CD44H. MSC produce and functionally adhere to extracellular matrix molecules. When incubated under proper stimuli, MSC differentiate into osteoblasts or adipocytes. Taken together, these results demonstrate that adherent marrow-derived cells cultured in the absence of hematopoietic cells and differentiation stimulus give rise to a population of cells with phenotypical and functional features of mesenchymal progenitors. The existence of a subset of quiescent cells in MSC cultures seems to be extremely significant, since their number and properties should be enough to sustain a steady supply of cells that upon proliferation and commitment may serve as precursors for a number of nonhematopoietic tissues.

Adipocytes↗
PubMed2002

Expression of integrins by human periodontal ligament and gingival fibroblasts and their involvement in fibroblast adhesion to enamel matrix-derived proteins.

We showed recently that human periodontal ligament (PDL) and gingival fibroblasts adhere and spread on enamel matrix protein (EMP) coatings. In the present study, we investigated whether this interaction can be attributed to integrin expression. Human PDL and gingival fibroblasts were cultured for periods up to 24 h on EMP coatings, in the presence of synthetic RGD-containing peptide or an antibody against the beta1 integrin subunit. The cells were first cultured for 24 h under serum-free conditions and then cultured on EMP coatings for 48 h. Integrin expression levels were assessed by flow cytometry analysis. It was found that attachment and spreading on EMP was inhibited by the synthetic RGD-containing peptide, but not by a synthetic RGE-peptide. Both PDL and gingival fibroblasts showed expression of the integrin subunits, alpha2, alpha5, beta1, and the integrin, alphavbeta3. Incubation with an antibody against the beta1 subunit significantly inhibited the attachment and spreading of PDL and gingival fibroblasts on EMP coatings. We conclude that integrins are involved in the interaction of PDL and gingival fibroblasts with EMP.

Adolescent↗
PubMed2003

The function of Xenopus germ cell nuclear factor (xGCNF) in morphogenetic movements during neurulation.

The germ cell nuclear factor (GCNF, NR6A1) is a nuclear orphan receptor first described in the mouse testis and subsequently identified as an essential transcription factor in vertebrate embryogenesis. Here, we analyze the phenotype of Xenopus embryos after depletion of embryonic GCNF (xEmGCNF) protein using a specific morpholino antisense oligonucleotide. Morphological defects after xEmGCNF knockdown became obvious from neurulation onward. Among the abnormalities observed, defective formation of the neural tube and a short and curved main body axis were the most remarkable traits. Histological analysis, lineage tracing of injected blastomeres, and Keller sandwich explants revealed that xEmGCNF function is required for different patterns of cell intercalation during neurulation and consequently for the sequence of morphogenetic movements leading to formation of the neural tube. Further characterization of the phenotype at the molecular level showed an abnormal distribution of the extracellular matrix protein fibronectin and a reduction in the expression level of the integrin subunits alpha5 and alpha6, the limiting components of the laminin and fibronectin receptors, respectively. We propose integrin-mediated cell-matrix interaction as a process that requires xEmGCNF function and provides, in concert with cadherins-mediated cell-cell interactions, a molecular basis for morphogenetic cell movements during neurulation.

Animals↗
PubMed2004

Differential expression of integrin alpha5beta1 in human abdominal aortic aneurysm and healthy aortic tissues and its significance in pathogenesis.

BACKGROUND: Abdominal aortic aneurysm (AAA) is a common aged disease of human aorta with increasing incidence. It is characterized by dramatic vascular remodeling via proteolysis and degradation of matrix proteins. Integrins are important cellular receptors for matrix proteins, which may have an association with pathological remodeling. The present study was undertaken to analyze the expression of integrin subunits in human aneurysmal aortas and with healthy aortic tissues as controls. MATERIALS AND METHODS: The expression of integrin genes in AAA specimens and healthy human aortic tissues was detected by RT-PCR technique. The correlation of variation and distribution of smooth muscle cells (SMCs) and integrin protein expression in the corresponding tissues were studied immunohistochemically. RESULTS: The gene transcripts coding for integrin alpha4, alpha5, alphaV, beta1, beta3, beta5, and beta6 subunits were constitutively expressed in the normal aortas. Only gene expressions of integrin alpha5 and beta1 were significantly decreased by 81% and 85%, respectively, in AAA specimens (P < 0.005) when compared with healthy aortic specimens. No age dependence of the expression of integrin alpha5beta1 genes was found. Significant reduction of medial SMC density was confirmed in corresponding AAA compared with control aortas. Immunoreactivity of integrin alpha5beta1 receptor was found to be exclusively localized within the medial layer of the parallel normal aortic sections, whereas this protein was absent in the destructive media of aneurysmal aortic sections. CONCLUSIONS: The marked decrease in integrin alpha5beta1 expressions was unique to aneurysmal aortic tissues and correlated to a decrease in density of SMCs, which are the major cells in maintaining the structure stability of normal aortas. As integrin alpha5beta1 specifically binds fibronectin and collagen, those results may suggest that the absence of integrin alpha5beta1 activity impair matrix protein attachment and alter the architecture in aortic media thereby lead to the deformity of aorta and aneurysm formation.

Adult↗
PubMed2005

Role for beta1 integrin and its associated alpha3, alpha5, and alpha6 subunits in development of the human fetal pancreas.

The integrin receptors play a major role in tissue morphogenesis and homeostasis by regulating cell interactions with extracellular matrix proteins. We have examined the expression pattern of integrin subunits in the human fetal pancreas (8-20 weeks fetal age) and the relevance of beta1 integrin function for insulin gene expression and islet cell survival. Its subunits alpha3, alpha5, and alpha6 beta1 integrins are expressed in ductal cells at 8 weeks, before glucagon- and insulin-immunoreactive cells bud off; their levels gradually increase in both ductal cells and islet clusters up to 20 weeks. Colocalization of alpha3, alpha5 and alpha6 beta1 integrins with endocrine cell markers was frequently observed in 8- to 20-week fetal pancreatic cells. When the beta1 integrin receptor was functionally blocked in cultured islet-epithelial clusters with a beta1 immunoneutralizing antibody or following transient beta1 integrin small interfering RNA treatment, there was inhibition of cell adhesion to extracellular matrices, decreased expression of insulin, and increased cell apoptosis. These data offer evidence for dynamic and cell-specific changes in integrin expression during human pancreatic islet neogenesis. They also provide an initial insight into a molecular basis for cell-matrix interactions during islet development and suggest that beta1 integrin plays a vital role in regulating islet cell adhesion, gene expression, and survival.

Apoptosis↗
PubMed1999

Development of cytotrophoblast columns from explanted first-trimester human placental villi: role of fibronectin and integrin alpha5beta1.

Human first-trimester floating mesenchymal villi explanted onto gels of collagen I or Matrigel were observed to undergo de novo development of anchoring sites. These consisted of cytotrophoblast columns that formed by proliferation of stem villous cytotrophoblast cells, as revealed by whole-mount and thin-section microscopy and incorporation of bromodeoxyuridine into DNA. Column formation occurred exclusively at the distal tips of the villi. No column formation was observed in tissue explanted onto agarose. On Matrigel, the developing columns penetrated downwards into the matrix, whereas on collagen I, cytotrophoblast sheets spread across the surface of the gel and merged to form a shell. The developing columnar cytotrophoblast up-regulated integrins alpha1beta1 and alpha5beta1 and produced an extracellular matrix containing oncofetal fibronectin, as in vivo. Function-blocking antibodies were used to investigate the role of the integrin-fibronectin interaction in anchoring villus development on collagen I. Antibodies to fibronectin and the integrin subunits alpha5 and beta1, added at 24 h, all changed the pattern of cytotrophoblast outgrowth. Anti-fibronectin caused cell rounding within the cytotrophoblast sheet and increased the population of single cells at its periphery. Anti-integrin alpha5 caused rounding and redistribution of cells within the outgrowth. In the presence of anti-integrin beta1, cell-collagen interactions within the sheet were destabilized, often leading to the appearance of an annulus of aggregated cells at the periphery. These results show that 1) mesenchymal villi retain the potential to form anchoring sites until at least the end of the first trimester, 2) adhesion to a permissive extracellular matrix stimulates cytotrophoblast proliferation and differentiation along the extravillous lineage, 3) integrin alpha5beta1-fibronectin interactions contribute significantly to anchorage of the placenta to uterine extracellular matrix. We suggest that as the developing placenta ramifies, new sites of anchorage form whenever peripheral villi contact decidua. This process is predicted to contribute to the stability of the placental-decidual interface.

Animals↗
PubMed1997

Fibronectin provides a conduit for fibroblast transmigration from collagenous stroma into fibrin clot provisional matrix.

After injury, the wound space is filled with a fibrin/fibronectin clot containing growth factors released by platelets and monocytes. In response to these factors, fibroblasts migrate into the fibrin clot and contribute to the formation of granulation tissue. The functional mechanisms allowing fibroblasts to leave the collagenous matrix of normal connective tissue and invade the provisional matrix of the fibrin clot have not been fully defined. To investigate these mechanisms we established a new in vitro model which simulates specific aspects of early wound healing, that is, the migration of fibroblasts from a three-dimensional collagen matrix into a fibrin clot. This transmigration could be induced by physiological concentrations of platelet releasate or platelet-derived growth factor BB (PDGF-BB) in a concentration-dependent manner. At 24 hours irradiated fibroblasts invaded the fibrin gel almost as well as non-irradiated cells, indicating that transmigration was independent of proliferation. Plasminogen and its activators appear to be necessary for invasion of the fibrin clot since protease inhibitors decreased the amount of migration. These serine proteases, however, were not necessary for exit from the collagen gel as fibroblasts migrated out of the collagen gel onto a surface coated with fibrin fibrils even in the presence of inhibitors. Removal of fibronectin (FN) from either the collagen gel or the fibrin gel markedly decreased the number of migrating cells, suggesting that FN provides a conduit for transmigration. Cell movement in the in vitro model was inhibited by RGD peptide, and by monoclonal antibodies against the subunits of the alpha5 beta1 and alpha v beta3 integrin receptor. Thus, the functional requirements for fibroblast transmigration from collagen-rich to fibrin-rich matrices, such as occurs in early wound healing, have been partially defined using an in vitro paradigm of this important biologic process.

Antigens, CD↗
PubMed2006

Interleukin-1alpha enhances the aggressive behavior of pancreatic cancer cells by regulating the alpha6beta1-integrin and urokinase plasminogen activator receptor expression.

BACKGROUND: In human pancreatic cancer progression, the alpha6beta1-integrin is expressed on cancer cell surface during invasion and metastasis formation. In this study, we investigated whether interleukin (IL)-1alpha induces the alterations of integrin subunits and urokinase plasminogen activator/urokinase plasminogen activator receptor (uPA/uPAR) expression in pancreatic cancer cells. We hypothesize that the alterations of integrin subunits and uPA/uPAR expression make an important role in signaling pathways responsible for biological behavior of pancreatic cancer cells. RESULTS: IL-1alpha upregulated the expression of alpha6 and beta1 integrins without any alterations of alpha5 and alphav integrins expression. IL-1alpha also induced enhancement in the expression of uPA/uPAR in pancreatic cancer cells. IL-1alpha enhanced the proliferation, adhesion, and migration in pancreatic cancer cells, and IL-1alpha-induced alterations of uPA/uPAR expression correlated with the increased the migration of pancreatic cancer cells. Upregulation of alpha6 integrin subunit and uPA/uPAR correlated with the activation of Ras and downstream extracellular signal-regulated kinase (ERK) pathways. IL-1alpha-induced activation of Ras and downstream ERK can be inhibited by using inhibitory antibodies against alpha6 and beta1 integrin and uPAR, consistent with the inhibition of proliferation, adhesion and migration of pancreatic cancer cells. Immunohistochemical analysis demonstrated a significant association between strong expressions of alpha6 integrin with uPAR in pancreatic cancer specimens. Furthermore, the strong expression of alpha6 integrin and uPAR was found to be independent prognosticator in pancreatic cancer patients. CONCLUSION: Based on these findings, we conclude that IL-1alpha can induce selective upregulation of alpha6beta1-integrin and uPA/uPAR in pancreatic cancer cells and these changes may modulate the aggressive functions of pancreatic cancer.

Aged↗
PubMed2002

Retroviral gene transfer to human epidermal keratinocytes correlates with integrin expression and is significantly enhanced on fibronectin.

Human epidermal keratinocytes are an important target for gene therapy because they can be easily expanded in culture and used to generate skin substitutes for the treatment of wounds, genetic diseases of the skin, and for delivery of proteins to the systemic circulation. Although retroviral transduction results in permanent genetic modification, differentiation and loss of transduced cells from the epidermis results in temporary transgene expression. To ensure permanent genetic modification, epidermal stem cells must be transduced with high efficiency. We evaluated gene transfer on two different substrates and found that the efficiency of gene transfer is substantially higher on a substrate of recombinant fibronectin (FN), when compared to tissue culture plastic (TCP). The rate of retroviral transduction on FN is four times faster than transduction on tissue culture plates and is independent of polybrene (PB). The transduction efficiency correlates with the levels of expression of integrin subunits alpha5, alpha2, and beta1, which have been shown to correlate with stem cell phenotype. Notably, cells that adhere rapidly to FN are transduced more efficiently than slowly adherent cells. In addition, integrin-blocking antibodies decrease the efficiency of gene transfer in a dose-dependent manner. Our results suggest that FN may enhance retroviral gene transfer to the least differentiated cells, thereby increasing the potential of genetically modified keratinocytes to treat short- and long-term disease states.

Antibodies, Monoclonal↗
PubMed1998

Fibronectin-mediated cell adhesion is required for induction of 92-kDa type IV collagenase/gelatinase (MMP-9) gene expression during macrophage differentiation. The signaling role of protein kinase C-beta.

Induction of the 92-kDa gelatinase (MMP-9) gene expression is associated with macrophage differentiation. In this study, we explored the regulatory mechanisms underlying this differentiation-associated MMP-9 gene expression in human HL-60 myeloid leukemia cells and human peripheral blood monocytes. Phorbol 12-myristate 13-acetate (PMA) markedly induced MMP-9 gene expression in HL-60 cells; the induction closely paralleled the timing and extent of PMA-induced cell adhesion and spreading, a hallmark of macrophage differentiation. Similarly, treatment with PMA or macrophage-colony stimulating factor stimulated adherence and spreading of blood monocytes with a concurrent 7- or 5-fold increase in MMP-9 production, respectively. In protein kinase C (PKC)-beta-deficient HL-60 variant cells (HL-525), PMA failed to induce cell adhesion and MMP-9 gene expression. Transfecting HL-525 cells with a PKC-beta expression plasmid restored PKC-beta levels and PMA inducibility of cell adhesion and spreading as well as MMP-9 gene expression. Induction of cell adhesion and MMP-9 gene expression in HL-60 cells and blood monocytes was strongly inhibited by neutralizing monoclonal antibodies to fibronectin (FN) and its receptor alpha5 beta1 integrin. HL-525 cells, which constitutively display high levels of surface alpha5 beta1 integrin, adhered and spread on immobilized FN with concomitant induction of MMP-9 gene expression. Cytochalasins B and D were each a potent inhibitor of MMP-9 production. Our results suggest that alpha5 beta1 integrin-mediated interaction of immature hematopoietic cells with FN plays a critical role in modulating matrix-degrading activities during macrophage differentiation.

Cell Adhesion↗
PubMed1998

Expression of alpha5 (CD49e) and alpha6 (CD49f) integrin subunits on T cells in the circulation and the lamina propria of normal and inflammatory bowel disease colonic mucosa.

Intestinal lamina propria T cells are believed to be derived, via the systemic circulation, from gut-associated lymphoid tissue. After migration into the lamina propria, T cells are capable of luminally directed migration following the loss of surface epithelial cells. For adhesion and migration within the extracellular matrix, T cells are likely to utilize the integrin family of adhesion molecules. The aim of this study was to quantitatively and qualitatively investigate the expression of alpha5 and alpha6 integrin subunits on the surface of human T cells that: (a) migrated out of the lamina propria, (b) remained resident within the matrix and (c) were present in the circulation. In both subpopulations of CD4 and CD8-positive T cells, from both normal and inflamed (inflammatory bowel disease) colonic mucosa, there were significantly fewer alpha5 and alpha6-positive cells than in the peripheral blood. In addition, there were significantly fewer alpha6 integrin molecules on the surface of CD4 and CD8-positive lamina propria T-cell subpopulations, compared with those in the circulation. Our studies suggest that, following migration into the lamina propria, there is down-regulation of alpha5 and alpha6 integrin-subunit expression on the surface of T cells. Molecules other than members of very late activation antigen-5 (VLA-5) (alpha5beta1) and VLA-6 (alpha6beta1) families of adhesion molecules are likely to be important in interactions with extracellular components in the lamina propria of normal and inflamed human colonic mucosa.

Antigens, CD↗