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PubMed2003

SPARC/osteonectin is a frequent target for aberrant methylation in pancreatic adenocarcinoma and a mediator of tumor-stromal interactions.

Deregulated expression of SPARC/osteonectin, a secreted glycoprotein with multiple biological functions, has been associated with the progression of various cancers. Using microarrays, we previously identified SPARC as one of the genes induced by treatment with a DNA methylation inhibitor in pancreatic cancer cells. We therefore analysed the expression pattern and methylation status of the SPARC gene in pancreatic cancer. Gene expression profiling by oligonucleotide microarray and reverse transcription-PCR analyses demonstrated that SPARC mRNA was expressed in non-neoplastic pancreatic ductal epithelial cells, but was not expressed in a majority of pancreatic cancer cell lines. The loss of SPARC expression was associated with aberrant hypermethylation of its CpG island. Immunohistochemical labeling revealed that the SPARC protein was overexpressed in the stromal fibroblasts immediately adjacent to the neoplastic epithelium in primary pancreatic cancers, but rarely expressed in the cancers themselves. Primary fibroblasts derived from pancreatic cancer strongly expressed SPARC mRNA and secreted SPARC protein into the conditioned media, and treatment of pancreatic cancer cells with exogenous SPARC resulted in growth suppression. SPARC expression in fibroblasts from noncancerous pancreatic tissue was augmented by coculture with pancreatic cancer cells. These findings suggest that SPARC is a frequent target for aberrant methylation in pancreatic cancer and that SPARC expression in fibroblasts adjacent to pancreatic cancer cells is regulated through tumor-stromal interactions.

Adenocarcinoma↗
PubMed2003

Mammalian fibroblasts lacking mitochondrial NAD+-dependent methylenetetrahydrofolate dehydrogenase-cyclohydrolase are glycine auxotrophs.

Primary fibroblasts established from embryos of NAD-dependent mitochondrial methylenetetrahydrofolate dehydrogenase-cyclohydrolase (NMDMC) knockout mice were spontaneously immortalized or transformed with SV40 Large T antigen. Mitotracker Red CMXRos staining of the cells indicates the presence of intact mitochondria with a membrane potential. The nmdmc(-/-) cells are auxotrophic for glycine, demonstrating that NMDMC is the only methylenetetrahydrofolate dehydrogenase normally expressed in the mitochondria of these cell lines. Growth of null mutant but not wild type cells on complete medium with dialyzed serum is stimulated about 2-fold by added formate or hypoxanthine. Radiolabeling experiments demonstrated a 3-10 x enhanced incorporation of radioactivity into DNA from formate relative to serine by nmdmc(-/-) cells. The generation of one-carbon units by mitochondria in nmdmc(-/-) cells is completely blocked, and the cytoplasmic folate pathways alone are insufficient for optimal purine synthesis. The results demonstrate a metabolic role for NMDMC in supporting purine biosynthesis. Despite the recognition of these metabolic defects in the mutant cell lines, the phenotype of nmdmc(-/-) embryos that begin to die at E13.5 is not improved when pregnant dams are given a glycine-rich diet or daily injections of sodium formate.

Aminohydrolases↗
PubMed1997

A benzo[a]pyrene-induced cell cycle checkpoint resulting in p53-independent G1 arrest in 3T3 fibroblasts.

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor of the basic helix-loop-helix family. Although physiological ligands for the AhR have not been identified, carcinogenic polycyclic aromatic hydrocarbons such as Benzo[a]pyrene (B[a]P) are high affinity AhR ligands that induce nuclear translocation and sequence-specific DNA binding of the AhR. AhR-regulated genes include members of the cytochrome P-450 family that are known to oxidize B[a]P to form genotoxic (DNA-damaging) metabolites. Murine Swiss 3T3 cells express high levels of AhR. Treatment of Swiss 3T3 cells with B[a]P during the G1 phase of the cell cycle resulted in growth arrest, as shown by inhibition of growth factor-stimulated DNA synthesis. By contrast, other murine 3T3 fibroblasts not expressing detectable levels of AhR did not undergo growth arrest in response to B[a]P. The AhR antagonist alpha-naphthoflavone prevented B[a]P-induced growth arrest, further demonstrating that cessation of cell growth was mediated by the activated AhR. A nongenotoxic AhR ligand (2,3,7,8-tetrachlorodibenzo-p-dioxin) did not elicit growth arrest, showing that ligand activation of the AhR alone was insufficient to block cell cycle progression. However, genomic DNA from B[a]P-treated Swiss 3T3 cells contained covalent adducts, whereas that from 2,3,7,8-tetrachlorodibenzo-p-dioxin-treated cells did not, showing that G1 arrest correlated with DNA damage resulting from genotoxic B[a]P metabolites. B[a]P-induced DNA damage and growth arrest was coincident with elevated levels of nuclear p53 protein and induction of the p53-regulated mdm-2 proto-oncogene. However, Swiss 3T3 fibroblasts expressing "dominant negative" mutant p53, as well as primary fibroblasts from p53-/- "knockout" mice, also underwent growth arrest in response to B[a]P. Therefore, B[a]P-induced growth arrest occurs via p53-independent mechanisms.

3T3 Cells↗
PubMed2005

Hypertrophic scar fibroblasts have increased connective tissue growth factor expression after transforming growth factor-beta stimulation.

BACKGROUND: Hypertrophic scars and keloids respond to dermal disruption with excessive collagen deposition and increased transforming growth factor (TFG)-beta expression. Connective tissue growth factor (CTGF) is a downstream mediator of TGF-beta activity that is associated with scar and fibrosis. The authors hypothesize that there is increased expression of CTGF by hypertrophic scar and keloid fibroblasts in response to TGF-beta stimulation. METHODS: Primary fibroblasts were isolated in culture from human hypertrophic scar (n = 2), keloid (n = 2), and normal skin (n = 2). After 18 hours of serum starvation, the cells were stimulated with 10 ng/ml of TGF-beta1, TGF-beta2, and TGF-beta3 for 24 hours. Quantitative real-time polymerase chain reaction was performed on extracted RNA samples to assay for CTGF mRNA expression. RESULTS: Baseline CTGF expression was increased 20-fold in unstimulated hypertrophic scar fibroblasts and 15-fold in keloid fibroblasts compared with normal fibroblasts. CTGF expression increased greater than 150-fold when stimulated with TGF-beta1 (p < 0.002) and greater than 100-fold when stimulated by TGF-beta2 or TGF-beta3 compared with normal fibroblasts (p < 0.02 and p < 0.002, respectively). CTGF expression was greatest after TGF-beta1 stimulation in hypertrophic scar fibroblasts compared with TGF-beta2 (p < 0.04) and TGF-beta3 (p < 0.02). Keloid fibroblast CTGF expression also increased greater than 100-fold after stimulation with TGF-beta1 (p = 0.16) and greater than 75-fold after addition of TGF-beta2 and TGF-beta3 (p = 0.06 and p = 0.22, respectively). CONCLUSIONS: Hypertrophic scar fibroblasts have both intrinsic up-regulation of CTGF transcription and an exaggerated capacity for CTGF transcription in response to TGF-beta stimulation. These data suggest that blockage of CTGF activity may reduce pathologic scar formation.

Adult↗
PubMed2002

Transformed and tumor-derived human cells exhibit preferential sensitivity to the thiol antioxidants, N-acetyl cysteine and penicillamine.

Thiol antioxidants, typified by N-acetyl cysteine, are known to induce p53-dependent apoptosis in transformed mouse embryo fibroblasts but not in normal mouse embryo fibroblasts. We now report that this is also the case for human cells. First, we used an isogenic fibroblast cell lineage exhibiting progressive stages of transformation, from primary derived cells to v-MYC immortalized to tumorigenic. At the immortalization stage, cells became 12- and 480-fold more sensitive to the thiol antioxidants N-acetyl cysteine (NAC) and penicillamine (PEN), respectively. Although immortalization of these cells was associated with v-MYC expression, overexpression of MYC was not sufficient for sensitizing these cells to antioxidants. To test whether sensitivity to antioxidants is a general property of immortalized human cells, including fully transformed cells, 12 tumor-derived cell lines were treated with PEN, the more potent of the two antioxidants. Ten of 11 caspase-proficient tumor cell lines underwent apoptosis after treatment, whereas primary fibroblasts and keratinocytes were resistant. The difference between normal and transformed cells was apparent whether the assay used measured caspase 3 activation, Annexin V binding, or cell viability. Tumor cell lines containing wild-type p53 were more sensitive than p53-null cell lines. The requirement for p53 was tested using the p53 inhibitor, pifithrin-alpha, or using stable transfectants of a v-MYC-immortalized, telomerase-positive cell line that expresses HPV16 E6 to bind and degrade p53. In the latter case, > or = 80% of the PEN-induced apoptosis was dependent on the presence of wild-type p53. These studies suggest that treatment with thiol-containing antioxidants, such as PEN, may offer a useful approach for preferential induction of apoptosis in preneoplastic and neoplastic cells.

Acetylcysteine↗
PubMed1989

Differential activation of the hprt gene on the inactive X chromosome in primary and transformed Chinese hamster cells.

We have investigated the genetic activation of the hprt (hypoxanthine-guanine phosphoribosyltransferase) gene located on the inactive X chromosome in primary and transformed female diploid Chinese hamster cells after treatment with the DNA methylation inhibitor 5-azacytidine (5azaCR). Mutants deficient in HPRT were first selected by growth in 6-thioguanine from two primary fibroblast cell lines and from transformed lines derived from them. These HPRT- mutants were then treated with 5azaCR and plated in HAT (hypoxanthine-methotrexate-thymidine) medium to select for cells that had reexpressed the hprt gene on the inactive X chromosome. Contrary to previous results with primary human cells, 5azaCR was effective in activating the hprt gene in primary Chinese hamster fibroblasts at a low but reproducible frequency of 2 x 10(-6) to 7 x 10(-6). In comparison, the frequency in independently derived transformed lines varied from 1 x 10(-5) to 5 x 10(-3), consistently higher than in the nontransformed cells. This increase remained significant when the difference in growth rates between the primary and transformed lines was taken into account. Treatment with 5azaCR was also found to induce transformation in the primary cell lines but at a low frequency of 4 x 10(-7) to 8 x 10(-7), inconsistent with a two-step model of transformation followed by gene activation to explain the derepression of hprt in primary cells. Thus, these results indicate that upon transformation, the hprt gene on the inactive Chinese hamster X chromosome is rendered more susceptible to action by 5azaCR, consistent with a generalized DNA demethylation associated with the transformation event or with an increase in the instability of an underlying primary mechanism of X inactivation.

Animals↗
PubMed1992

Primary chimpanzee skin fibroblast cells are fully permissive for human cytomegalovirus replication.

Cytomegaloviruses generally display a host range restricted to differentiated cell types from the species they infect. For human cytomegalovirus (HCMV) this has meant that with few exceptions tissue culture systems have relied on the use of primary foreskin fibroblast (HF) cells or primary human embryonic lung cells to study gene expression and virus replication functions. We have observed that primary skin fibroblast (CF) cells derived from the chimpanzee (Pan troglodytes) support the replication of a laboratory strain (Towne) of HCMV. The kinetics of gene expression of the Towne strain grown in CF or HF cells appeared to be equivalent. Titres of progeny virions grown in CF cells appeared to be reduced 10-fold relative to those of virus grown in HF cells. In contrast, replication of the Towne virus was not supported by growth in WES cells (ATCC no. CRL 1609), a chimpanzee skin fibroblast cell line transformed by an adenovirus 12-simian virus 40 hybrid. This study shows that HCMV is less parochial in its host range than previously thought.

Animals↗
PubMed2002

Identification of novel candidates for replicative senescence by functional proteomics.

To identify the underlying mechanisms that limit the mitotic potential of normal somatic cells, we have undertaken a high resolution differential proteomic analysis aimed at identifying proteins that were differentially expressed upon replicative senescence. Since replicative senescence in heterogeneous primary fibroblast cultures is asynchronous, we analysed a group of conditionally immortalized rat embryo fibroblast cell lines that have previously been shown to undergo synchronous senescence upon inactivation of SV40 tsA58 T antigen. This identified 43 spots that were differentially expressed in these cell lines. Comparison of the identity of these features with those identified in a complimentary independent differential proteomic analysis of replicative senescence, directly in primary rat embryo fibroblasts upon serial passaging, identified nine features that were in common between the two studies even though they had been conducted entirely separately. None of these proteins have previously been recognized to be involved with replicative senescence. Thus, they represent novel starting points for elucidating the underlying mechanism that regulates the finite mitotic life span of somatic cells and how it can be overcome in cancer cells.

Animals↗
PubMed1995

DNA repair synthesis following irradiation with 254-nm and 312-nm ultraviolet light is not diminished in fibroblasts from patients with dysplastic nevus syndrome.

The DNA excision repair capacity of 23 primary fibroblast lines from patients with dysplastic nevus syndrome was investigated and DNA repair synthesis ("unscheduled DNA synthesis") was determined after UV exposure. Seventeen fibroblast lines from normal donors served as controls. The dose/response experiments included up to ten dose levels and two wavelength ranges: UV-C (using a low-pressure mercury lamp emitting predominantly 254-nm light) and UV-B (artificial "sunlamp" radiation centering around 312-nm light). For each dose level, silver grains over fibroblast nuclei were counted by visual inspection. Twelve cell lines were also evaluated for both UV wavelength ranges using a new semi-automatic image analyzing system. This system included components for rapid sequential identification of both fibroblast nuclei and silver grains sited above them. Silver grains over 100 nuclei were determined for each UV dose level. Dose/response curves were established and analyzed by linear regression. As a quantitative term for assessing DNA excision repair capacity of a cell line we calculated the linear increase (G0) in the number of grains per nucleus, when the UV dose was multiplied by the factor e (i.e. 2.72). The sensitivity of grain detection and resolution of overlapping grains was approximately threefold better in visual than in automatic counting, especially when there were more than 70 grains over nuclei. The time required for visual counting, however, was tenfold that of automatic counting. The variance-weighted mean G0v.w of all fibroblast lines from patients with dysplastic nevus syndrome was found to be 79.1 (+/- 1.8- grains/nucleus, that of fibroblast lines from normal donors was 74.2 (+/- 1.7) grains/nucleus. This difference revealed a slightly better repair capability for cell lines from patients but was at the borderline of detection and, therefore, should not be overinterpreted. From the experimental accuracy achieved by determination of the variance-weighted means of the two groups, we would have been able to detect a difference of 7 and more grains [> 2 x (sigma normal+sigma patients)]. The variance-weighted mean G0v.w of all fibroblast lines from patients with dysplastic nevus syndrome was found to be 76.4 (+/- 1.4) grains/nucleus, whereas that of fibroblast lines from normal donors was only 66.6 (+/- 1.8) grains/nucleus. This difference was statistically significant and, contrary to expectation, revealed better, not worse post-UV DNA repair capability in cell lines from patients that in those from normal donors.(ABSTRACT TRUNCATED AT 400 WORDS)

Autoradiography↗
PubMed1980

Cytolysis of fibroblasts by C3a.

C3a was found to be more cytolytic to transformed than to primary fibroblast cultures from mouse and man. Fibroblasts made quiescent with caffeine were lysed by the same concentrations of C3a as untreated cells. These findings may have implications in tumour immunity.

Animals↗
PubMed2005

Oxidative and heat stress gene changes in hypertrophic scar fibroblasts stimulated with interleukin-1beta.

INTRODUCTION: Cutaneous wounds that involve loss of tissue heal through a complex process of generating granulation tissue to initially cover the wound, followed by epithelialization, and contraction. Normal healing requires a delicate balance between cellular, matrix, and vascularity build up and breakdown. Defects in the regulation of this balance can alter normal scar formation through fibroblastic hyperproliferation, which is characteristic of hypertrophic scar formation. METHODS: Primary fibroblasts cultures from hypertrophic scar or adjacent normal skin were seeded in growth medium. Half of each was stimulated with interleukin (IL)-1beta for 6 h and half served as control nonstimulated fibroblasts. Supernatants were tested for expressed proteins and the fibroblasts for changes in gene expression. RESULTS: Comparison between normal skin and hypertrophic scar fibroblasts stimulated with IL-1beta indicate that 15 genes increased and 8 genes decreased expression. When normal skin was stimulated with IL-1beta, there were increases in the expression of heat shock transcription factor-1, hsp70, and IL-6 When hypertrophic scar was stimulated with IL-1beta, there was a decrease in nuclear factor-kappaB, GADD45-alpha, p53, p53 binding protein, and Cox-2 genes. These genes may play specific but different roles in controlling the cellular response to cell stress and apoptosis. CONCLUSION: Data presented suggest that oxidative and heat stress proteins, stimulated through IL-1beta, may be important mediators of abnormal scarring.

Adolescent↗
PubMed2003

Behavioral correction of Parkinsonian rats following the transplantation of immortalized fibroblasts genetically modified with TH and GCH genes.

Eukaryotic plasmid vectors encoding the tyrosine hydroxylase (TH) gene and GTP cyclohydrolase-1 (GCH) gene were constructed and introduced into immortalized fibroblasts obtained from SV40 large antigen (LT(AG)) transformed rat primary fibroblasts. TH and GCH positive clones were selected and identified by immunohistochemistry and RT-PCR, respectively. Hemi-parkinsonian rats created using 6-hydroxydopamine (6-OHDA) were used to assess the therapeutic effect created by the co-implantation of immortalized fibroblasts genetically modified by TH or GCH genes. Animal behavior was significantly improved two weeks following implantation and behavioral correction was maintained for over 14 weeks. Behavioral improvement was paralleled by exogenous TH gene expression, identified by TH immunohistochemistry and RT-PCR analyses. The transplanted cells survived for at least 38 weeks as demonstrated by fibronectin immunohistochemical staining. Tumor formation or host reaction was not seen, although TH expression was negative for 20 weeks after the implantation. This work demonstrates that the co-transplantation of immortalized fibroblasts genetically modified by TH and GCH genes may be developed as a valuable approach to the treatment of Parkinson's disease.

Animals↗
PubMed2005

Human skin and gingival keratinocytes show differential regulation of matrix metalloproteinases when combined with fibroblasts in 3-dimensional cultures.

BACKGROUND: Matrix metalloproteinases (MMP) and their inhibitors are expressed in tissues during interactions between keratinocytes and fibroblasts. Maintaining the balance between MMPs and their inhibitors is critical; failure to do so can lead to severe tissue damage or complete destruction, as seen in periodontal disease. Previously we showed that 3-dimensional (3-D) cultures of homotypically-combined skin and gingival cells mimicked the tissues in protein and lipid production, but heterotypic cultures did not. METHODS: We examined the production and activation of MMPs in these homotypic and heterotypic combinations of skin and gingival keratinocytes and fibroblasts during the critical time that they reformed the tissues. Primary fibroblasts and keratinocytes were isolated from normal human gingiva and skin and grown in 3-D cultures for up to 42 days. MMP-1, MMP-2, and MMP-9 in the media and inhibition of MMPs from these cultures were analyzed. RESULTS: These experiments determined that skin fibroblasts grown with skin or gingival keratinocytes secrete increased amounts of MMP-1 compared to gingival fibroblasts; that the interaction of keratinocytes with fibroblasts decreases the amount of MMP-2 produced by the fibroblasts in 3-D cultures; that skin keratinocytes, but not gingival keratinocytes, interact with fibroblasts to upregulate expression of the active form of MMP-9; and that medium conditioned by gingival 3-D cultures does not contain an inhibitor of MMP-9. CONCLUSION: Varying the type of fibroblast beneath the keratinocytes allowed us to determine that skin and gingival keratinocytes differentially regulate the production and activation of MMP-9, but not MMP-2, a finding that could influence the success of tissue grafting after periodontal surgery.

Animals↗
PubMed1998

Defective growth correlates with reduced accumulation of a viral DNA replication protein after low-multiplicity infection by a human cytomegalovirus ie1 mutant.

To investigate the importance of the IE1 p72 regulatory protein during human cytomegalovirus replication, a recombinant virus unable to synthesize IE1 p72 was constructed. The Towne strain mutant CR208 lacked exon 4 of the major immediate-early gene and was isolated and complemented in an IE1-expressing immortalized human fibroblast line (ihfie1.3). Replication of CR208 in primary human fibroblasts was completed after an input multiplicity of 10 PFU/cell but was severely-impaired at 0.1 PFU/cell. CR208 formed plaques with lower efficiency on primary fibroblasts than on ihfiel.3 cells, and the relationship between the CR208 inoculum size and the resulting number of undersized plaques was nonlinear, indicating that multiple particles of CR208 were required to initiate lytic replication in a single primary fibroblast. After infection of primary fibroblasts with CR208 at 5 PFU/cell, a normal pattern of viral antigens was detected, although IE1 p72 was absent. During lower-multiplicity infections, IE2 protein was consistently detected at similar levels in a similar proportion of CR208-infected cells relative to the case for a Towne infection, but many fewer CR208-infected cells contained the ppUL44 polymerase accessory protein when evaluated at 24 or 48 h after infection. Furthermore, fibroblasts infected with CR208 at a low multiplicity failed to form viral DNA replication compartments, despite having expressed IE2 p86. These low-multiplicity growth and expression defects were corrected in two rescued derivatives of CR208 able to synthesize IE1 p72. One rescued virus (CR249) carried a deletion removing the large intron between exons 1 and 2 of the ie1-ie2 locus, revealing that this intron was dispensable for growth in cell culture.

Antigens, Viral↗
PubMed1983

Induction of DNA repair synthesis by ultraviolet radiation and methylmethanesulphonate in cultured mouse lymphocytes.

The induction of DNA repair synthesis by UV-radiation and methyl-methanesulphonate (MMS) was studied in mouse lymphocytes and leukemic cells by means of autoradiography and scintillation counting, after labelling in vitro with tritiated thymidine ([3H]dThd). Repair stimulation was detected by both procedures in LSTRA and YC8 leukemic cell lines as well as in primary fibroblasts of BALB/c and BALB/Mo mice. No stimulation was observed in primary cultures of lymphocytes from the spleen, thymus and lymph-nodes of the same mice. In primary lymphocytes neither stimulation with concanavalin A (Con A) nor pre-incubation with 5-bromodeoxyuridine (BUdR) were effective in making evident DNA repair. The data put into question the reliability of the repair test for the prediction of carcinogenic potential of chemicals.

Animals↗
PubMed2006

Primary mouse dermal fibroblast cell cultures as an in vitro model system for the differential pathogenicity of cross-species herpesvirus papio 2 infections.

Infection of mice with herpesvirus papio 2 (HVP2) parallels zoonotic monkey B virus infections. A major benefit of the HVP2/mouse model is the existence of two HVP2 subtypes: HVP2nv rapidly invades and destroys the CNS while HVP2ap produces no clinical signs and mild histopathological lesions. However, in the natural baboon host, no difference in pathogenicity of HVP2 subtypes is evident. Primary dermal fibroblast cells were evaluated as a model system for defining virus-host interactions that influence the outcome of a cross-species infection. No differences in plaque formation or virus replication were observed between HVP2 subtypes in primary baboon dermal fibroblast cultures. In contrast, when primary mouse dermal fibroblasts (PMDF) were infected, HVP2nv replicated to higher titers and was more efficient at shutting down host-cell protein synthesis compared to HVP2ap. HVP2ap-infected PMDF cells produced more IFN-beta compared to HVP2nv, and IFN-beta pretreatment of PMDF cultures inhibited HVP2ap replication but did not affect HVP2nv. The differential pathogenicity of HVP2 subtypes in mice and the lack of such differences in the natural baboon host are recapitulated in the primary dermal fibroblast cell culture system. This model may prove useful in examining early, local, host responses that influence the outcome of cross-species infections.

Animals↗
PubMed2000

The angiogenic factors Cyr61 and connective tissue growth factor induce adhesive signaling in primary human skin fibroblasts.

The angiogenic inducers cysteine-rich angiogenic protein 61 (Cyr61) and connective tissue growth factor (CTGF) are structurally related, extracellular matrix-associated heparin-binding proteins. Both can stimulate chemotaxis and promote proliferation in endothelial cells and fibroblasts in culture and induce neovascularization in vivo. Encoded by inducible immediate early genes, Cyr61 and CTGF are synthesized upon growth factor stimulation in cultured fibroblasts and during cutaneous wound healing in dermal fibroblasts. Recently, we have shown that adhesion of primary human fibroblasts to immobilized Cyr61 is mediated through integrin alpha(6)beta(1) and cell surface heparan sulfate proteoglycans (HSPGs) (Chen, N., Chen, C.-C., and Lau, L.F. (2000) J. Biol. Chem. 275, 24953-24961), providing the first demonstration of an absolute requirement for HSPGs in integrin-mediated cell attachment. We show in this study that CTGF also mediates fibroblast adhesion through the same mechanism and demonstrate that fibroblasts adhesion to immobilized Cyr61 or CTGF induces distinct adhesive signaling responses consistent with their biological activities. Compared with fibroblast adhesion to fibronectin, laminin, or type I collagen, cell adhesion to Cyr61 or CTGF induces 1) more extensive and prolonged formation of filopodia and lamellipodia, concomitant with formation of integrin alpha(6)beta(1)-containing focal complexes localized at leading edges of pseudopods; 2) activation of intracellular signaling molecules including focal adhesion kinase, paxillin, and Rac with similar rapid kinetics; 3) sustained activation of p42/p44 MAPKs lasting for at least 9 h; and 4) prolonged gene expression changes including up-regulation of MMP-1 (collagenase-1) and MMP-3 (stromelysin-1) mRNAs and proteins sustained for at least 24 h. Together, these results establish Cyr61 and CTGF as bona fide adhesive substrates with specific signaling capabilities, provide a molecular basis for their activities in fibroblasts through integrin alpha(6)beta(1) and HSPG-mediated signaling during attachment and indicate that these proteins may function in matrix remodeling through the activation of metalloproteinases during angiogenesis and wound healing.

Cell Adhesion↗
PubMedDate not supplied

Adenosine deaminase complexing protein in cancer studies.

ADCP is a dimeric glycoprotein of about 200KD, for which the physiological role is still obscure. This protein occurs mainly in a membrane bound form in various human tissues. In this paper we review the current literature on ADCP in cancer studies. Soluble ADCP was described to be consistently decreased or absent in cancers of lung, liver, kidney and colon. These findings could not be confirmed by immunohistochemical and quantitative biochemical studies in a series of colorectal-, prostatic-, and renal carcinomas. Only in a third of these tumors a decrease could be demonstrated, whereas in the other cases unaltered or even increased amounts were observed. However, in virally transformed human fibroblasts a consistent decrease or complete absence of ADCP was seen, while primary fibroblasts were found to contain high amounts of this protein. Recently, the use of ADCP as a differentiation marker in colonic cancer has been advocated. Furthermore the presence of ADCP in the serum of renal adenocarcinoma patients was found to be indicative of a better chance of five year survival. These studies suggest that ADCP may be a differentiation marker useful for immunohistochemical characterization of colonic and renal carcinomas as well as a serum marker useful for follow-up studies of these types of cancer, analogous to CEA. Finally, ADCP has been found to be selectively expressed by certain T-cell subsets and henceforth may be useful in the studies on leukemias.

Carrier Proteins↗