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PubMed2001

Perineuronal nets show intrinsic patterns of extracellular matrix differentiation in organotypic slice cultures.

Perineuronal nets (PNs), consisting of extracellular matrix proteoglycans, complexed with hyaluronan and colocalized with tenascins, are associated with distinct neuronal populations in mature mammalian brain. PNs have been shown to appear postnatally during the period of synaptic refinement and myelination, indicating the commencement of mature physiological properties of neurons. Here we show that the developmental patterns of formation of PNs are well preserved in organotypic slice cultures prepared from rats on postnatal day 3-5 and maintained in vitro for 3-10 weeks. Staining of cultures with Wisteria floribunda agglutinin and immunocytochemical detection of chondroitin sulfate proteoglycans revealed developing PNs in the basal forebrain, mesencephalic regions, and the cerebellum after 2 weeks in vitro, and later in the neocortical areas and hippocampus. In contrast, neurons known to be devoid of PNs in the adult rat brain such as cholinergic basal forebrain neurons and catecholaminergic tegmental neurons differentiate without any formation of PNs in slice cultures. We show further that environmental factors influence the development of PNs around the net-associated types of neurons. Notably, chronic depolarization of the cultures, imposed by an elevated concentration of external potassium ions, enhanced the development of PNs. Blocking of calcium channels with magnesium chloride or with the L-type calcium channel blocker nifedipine, suppressed the development of PNs, while a block of voltage-gated sodium channels by tetrodotoxin had no obvious effects. The results show that extracellular matrix components specifically contribute to the organotypic patterns that develop in brain slice cultures. Evidence is provided that the differentiation of PNs is regulated by calcium-dependent signaling.

Acetylcholine↗

PubMed1996

Characterization of gelatinases linked to extracellular matrix invasion in ovarian adenocarcinoma: purification of matrix metalloproteinase 2.

Substantial evidence indicates that proteolytic degradation of the extracellular matrix is necessary for invasion and metastasis by cancer cells. Our previous work has demonstrated elevated secretion by cultured ovarian adenocarcinoma cells of two gelatinolytic metalloproteinases, a 72-kDa enzyme resembling matrix metalloproteinase 2 (MMP-2) and a 92-kDa enzyme resembling MMP-9 (Moser et al, Int. J. Cancer 56, 552-559, 1994). To assess the potential in vivo relevance of these enzymes, we have examined ovarian carcinoma ascites using gelatin substrate zymography. MMP species identical to those secreted from several well-characterized ovarian adenocarcinoma cell lines were found in the majority of ascites: MMP-2-like gelatinase (23 of 23 cases) and MMP-9-like gelatinase (18 of 23 cases), suggesting a prevalence of these species in the ovarian carcinoma microenvironment and their availability for tumor-associated proteolysis. The contribution of these proteinases to ovarian cancer invasion was further demonstrated by experiments measuring tumor cell-mediated proteolysis of native endothelial cell extracellular matrix (ECM) and tumor cell invasion of reconstituted basement membrane (Matrigel). These data showed that secretion of type IV collagenase activity by a series of independently isolated ovarian adenocarcinoma cell lines correlated well with the ability of these cells to proteolyze the ECM and invade the basement membrane. Furthermore, we have identified and characterized an ovarian carcinoma-associated gelatinase, the 72-kDa MMP found in conditioned media of the DOV 13 cell line, as MMP-2. This enzyme was identical to the previously described MMP-2 from other sources by Western blot, amino terminal sequence, and substrate specificity. Additionally, a large portion of the MMP-2 activity found in DOV 13 conditioned media is active without organomercurial treatment, suggesting that ovarian cancer cells have an endogenous activator of the zymogen. Together, these data suggest that ECM proteolysis mediated by tumor-associated proteinases plays an important role in the invasion and/or metastasis of ovarian carcinoma.

Adenocarcinoma↗

PubMed1984

The effect of the extracellular matrix on the detachment of human endothelial cells.

Human umbilical vein endothelial cells can be serially passaged by supplementing medium with a partially purified growth factor. Cell-substratum detachment of early and late passage endothelial cells was examined using trypsin, collagenase, or homocysteine. Late-passage cells detached more rapidly than early passage cells under all conditions tested. The rate of detachment was dependent upon the specific agent used. Protease-mediated detachment was most rapid, occurring over minutes, in contrast to homocysteine-induced detachment, which occurred over hours. When detached cells were collected and replated in the absence of the detaching agent, these cells reattached, spread, and continued to proliferate. No significant difference was observed in the rate of adhesion of either early or late passage cells to a gelatin matrix. When early or late-passage endothelial cells were plated and grown to confluence on a matrix synthesized by the opposite cell type, the rate of protease-mediated cell detachment resembled the cell type from which the matrix was derived. The ease of endothelial cell detachment was determined by the origin of the extracellular matrix. Examination of the extracellular matrices from early and late passage cells revealed significant differences in the amounts of glycosaminoglycans and sulfated proteins present. These studies demonstrate the importance of the endothelial cell extracellular matrix in protease-mediated cell detachment. The rate of cell detachment was controlled by the extracellular matrices are not altered by the endothelial cells.

Cell Adhesion↗

PubMed1996

Differential expression of extracellular matrix components during the morphogenesis of human gastric mucosa.

BACKGROUND: Components of extracellular matrix play a crucial role in morphogenesis and epithelial cell differentiation. In this study, we examined the spatiotemporal expression and distribution of several major extracellular macromolecules of developing and adult human gastric mucosa, with particular emphasis on the pit-gland axis. METHODS: Indirect immunofluorescence was performed on cryosections of developing and adult gastric tissues by using specific antibodies. RESULTS: From 8 weeks of gestation onward, including adults, heparan sulfate proteoglycan, type IV collagen, and laminin alpha 1, beta 1, and gamma 1 chains were systematically and uniformly located at the basement membrane of the stratified epithelium (8-11 weeks) and of surface and pit-gland epithelia. Between 8 and 20 weeks of gestation, fibronectin and tenascin were colocalized throughout the entire mesenchyme. In adult mucosa, fibronectin was distributed at all levels of surface and glandular epithelium, whereas tenascin expression was restricted to surface and pit epithelial cells. The expression of the alpha 2 laminin chain was first detected at 12 weeks at the base of forming gastric glands. In the adult, the alpha 2 as opposed to the alpha 1 chain of laminin was confined to the basement membrane of the glandular and lower part of gastric pit epithelia. CONCLUSIONS: This analysis of the composition of the extracellular matrix in human gastric mucosa suggests an important role for some of its components in morphogenesis and maintenance of gastric glands.

Collagen↗

PubMed2002

Regulation of endothelin synthesis by extracellular matrix in human endothelial cells.

BACKGROUND: Vascular diseases are characterized by the presence of structural changes and the progressive loss of endothelial function. Although the biochemical basis of these structural changes have started to be outlined, it seems that accumulation of normal extracellular matrix proteins as well as the appearance of interstitial collagens, mainly collagen type I, characterize this process. On the other hand, a role for endothelial vasoactive factors has been proposed in the genesis of endothelial dysfunction, and it is generally accepted that changes in extracellular matrix composition may modify cell behavior. METHODS: Experiments were designed to test the influence of the supporting matrix on endothelin-1 (ET-1) synthesis by endothelial cells. Northern blot experiments were performed to analyze the prepro-endothelin-1 (prepro-ET-1) mRNA expression. ET-1 production was measured by ELISA. RESULTS: Cells grown on collagen type I (Col I) showed an increase of prepro-ET-1 mRNA level when compared with cells cultured on collagen type IV (Col IV). According to these results, the release of ET-1 to culture medium was also higher in Col I-grown cells than in those cultured on Col IV. Treatment of cells with a peptide that interferes with Col I integrins (D6Y), or with protein tyrosine kinase inhibitors such as genistein and herbimycin, completely abolished the effect of Col I. Moreover, experiments with antibodies against integrins suggest that these cell surface receptors could be involved in the modulation of ET-1 system by extracellular matrix. CONCLUSIONS: These results suggest that the presence of an abnormal extracellular matrix could stimulate endothelin synthesis by human endothelial cells, through integrin activation.

Cells, Cultured↗

PubMed1993

The effect of extracellular matrix molecules on the in vitro behavior of bovine endothelial cells.

Extracellular matrix (ECM) is an important mediator of endothelial functions such as adhesion, spreading, migration, proliferation, and maintenance of differentiated functions. Attachment of cultured cells to tissue culture polystyrene (TCPS) is dependent on vitronectin which adsorbs onto the surface from the serum in the culture medium. Vitronectin (VN) will adsorb efficiently to TCPS even if the latter has been coated with another matrix molecule and blocked with albumin. This means that studies of the interactions of cells with individual coated ECM molecules will be confounded by the presence of adsorbed VN if serum is present in the culture medium. In this study, the adhesion, spreading, growth, and output of endogenous matrix molecules by bovine corneal endothelial (BCE) cells were measured on five different matrix substrates using medium which had been depleted of vitronectin to avoid such confounding effects. The same cell adhesion and spreading maxima were achieved on vitronectin, fibronectin (FN), laminin (LM), and types I and IV collagen (col I, col IV). The coating concentrations required to achieve these maxima, however, differed among the substrates, LM needing considerably higher concentrations than the other substrates for both maximal adhesion and spreading and FN needing higher concentrations for cell spreading. When cells were continuously passaged on each of the five substrates coated at concentrations optimal for cell spreading, no differences in cell proliferation rates or cell morphology were observed. Significant differences, however, were observed in the subcellular output of endogenous matrix molecules (FN, LM, col IV, and thrombospondin) between the different substrates. Col I was a poor substrate for the production of all ECM molecules tested over the 10 passages of the experiment, whereas col IV was a consistently good substrate. LM and FN substrates displayed differential effects on the output of different ECM molecules. VN was unique in that BCE cells at early passage on this substrate produced high levels of endogenous matrix molecules, whereas with continued passage on this substrate, a progressive decline in ECM secretion was observed. These results show that incorporation of individual molecules into the ECM by BCE cells in culture is significantly affected by the nature of the substratum. They further suggest that passage of endothelial cells in media containing serum (which results in coating of VN onto the substrate) may result in a progressive reduction of ECM output.

Animals↗

PubMed2006

Dermatopontin, a novel player in the biology of the extracellular matrix.

Dermatopontin is a widely distributed small molecular weight protein in the extracellular matrix (ECM) and today its homologues are known in five mammals and several invertebrates. The structures of these homologues are relatively well conserved among the species. In the skin, dermatopontin is located mainly on the surface of the collagen fibers. It is found in the conditioned medium and also in the cytoplasm of cultured fibroblasts. Early studies focused on ECM assembly (collagen fibrillogenesis) and interactions (with the proteoglycan decorin). Subsequently, a targeted disruption of dermatopontin resulted in a phenotype similar to Ehlers-Danlos syndrome. In addition, a cell adhesion activity of this protein for dermal fibroblasts and several other cells was found, and this activity might suggest this protein's involvement in wound healing. The expression of dermatopontin around an infarct zone of experimental myocardial infarction may support this possibility. In invertebrates, dermatopontin homologues act mainly as adhesion/agglutination molecules. In addition, we found that transforming growth factor-beta1 interacts with dermatopontin and the function of this cytokine is modified by dermatopontin. Recently, the involvement of this protein in cell proliferation has been indicated. In this review we describe the reported functions of this protein and speculate on the multiple roles of this largely uncharacterized matrix molecule.

Amino Acid Sequence↗

PubMed1993

Interaction of osteoblasts with extracellular matrix: effect of mast cell chymase.

We studied the effect of mast cell chymase on the interaction between osteoblasts and extracellular matrix. Chymase was purified from mast cell lysate by anion exchange chromatography. Osteoblasts were isolated from rat calvarias by collagenase digestion. Incubation of osteoblasts with mast cell lysate (40-170 micrograms/ml) or purified chymase (8-32 micrograms/ml) resulted in changes in cell-matrix interaction and cell morphology. Osteoblasts treated with chymase also showed a gradual detachment from the artificial substrata and from the biomatrix (collagen-digested rib fragment). A similar effect of mast cell chymase on the osteoblasts was found in vitro on endosteum of an intact parietal bone. Neutral protease inhibitors abolished the effect of both crude and purified enzyme preparations on the cell-matrix interaction. Mast cell chymase had no effect on osteoblast viability. The effect of enzyme on osteoblast proliferation was studied with lower concentrations of enzyme (0.2 micrograms/ml) in order to avoid cell detachment; there was no effect on either the metaphase index or on the number of cells after 5 days of incubation with chymase. Osteoblast attachment and cell spreading on different matrix proteins (fibronectin, vitronectin, extract of noncollagenous matrix proteins from rat bone) were significantly altered by their pretreatment with chymase. Matrix fibronectin of osteoblasts in culture as well as soluble vitronectin and fibronectin were digested by rat mast cell chymase. Our data suggest that mast cells through action of neutral protease chymase may alter molecules in extracellular matrix that are important in osteoblast adhesion, cell spreading, maintenance of cell morphology, and, most likely, cell function.

Animals↗

PubMed1987

Distribution of extracellular matrix proteins in primary human brain tumours: an immunohistochemical analysis.

Using immunohistochemical techniques, we localized several glycoproteins of the extracellular matrix in paraffin-embedded sections of 4 normal brain and 38 primary intracranial tumour specimens. All specimens were positively immunostained to various degrees by monoclonal antibodies to type IV collagen and procollagen III and by antisera to laminin and fibronectin. Staining was consistently most intense at sites of contact between neuroepithelial and mesenchymal or leptomeningeal elements; there was no demonstrable staining within or between neuroepithelial elements in the neuropil. Tumour cells from meningiomas and from the sarcomatous portion of a gliosarcoma were positively immunostained for fibronectin and laminin. The integrity of the glial limitans externa was demonstrated by the positive linear reaction product produced by immunostains for type IV collagen and laminin, even in the most malignant gliomas. The deposition of extracellular matrix glycoproteins at the glial-mesenchymal interface observed in this study of primary human brain tumours is a manifestation of one of the interactions between tumour and stromal cells in the central nervous system. A loss of coordination and an alteration in the interactions between epithelial cells and stromal cells across extracellular matrices such as basement membranes are thought to be fundamental steps in the development and progression of cancer. Further characterization studies focusing on other markers of the extracellular matrix are needed to elucidate completely the function of this structure in the central nervous system.

Antibodies, Monoclonal↗

PubMed1988

Immunohistochemical studies on the tissue localization of collagen types I, III, IV, V and VI in schwannomas. Correlation with ultrastructural features of the extracellular matrix.

The distinctive tissue localization of collagen types in typical schwannomas with Antoni type A and B areas was demonstrated immunohistochemically using affinity-purified antibodies against types I, III, IV, V and VI collagen and comparative ultrastructural studies were made on the extracellular matrix components. Antoni type A tissue, which was composed of tightly packed spindle cells with long cytoplasmic processes surrounded by a continuous basement membrane and a few fibrillar components of the extracellular matrix, was almost exclusively immunoreactive for type IV collagen, presumably representing the basement membrane. Verocay bodies, which are organoid structures of Antoni type A tissue, had a variety of more abundant extracellular fibrous components, such as banded collagen fibrils, fibrous long-spacing fibrils and microfibrils. These were positive for type I and III, as well as type IV collagen. In Antoni type B areas, where two types to tumor cells designated Schwann cell-like and fibroblast-like were scattered in large amounts of amorphous extracellular matrix containing microfibrils and thick banded collagen fibrils, type VI collagen as well as types I, III and IV collagen were consistently detected. Type V collagen was localized in dense fibrous tissue areas and around blood vessels. These findings indicate that the differently organized cellular patterns of schwannomas, identified as Antoni types A and B, are characterized not only by the ultrastructural features of the extracellular matrix, but also by the distinctive collagen types produced by neoplastic Schwann cells.

Adult↗

PubMed1999

Cell-extracellular matrix interactions and EGF are important regulators of the basal mammary epithelial cell phenotype.

The mammary epithelium is composed of a luminal epithelium and a basal layer containing myoepithelial cells and undifferentiated precursors. Basal cells express specific protein markers, such as keratin 14 (K14) and P-cadherin. To study the factors that regulate the basal mammary epithelial cell phenotype, we have established two clonal derivatives of the mouse HC11 cell line, BC20 and BC44, expressing high levels of K14 and P-cadherin. Unlike the parental HC11 cells, these basal cells did not produce beta-casein in response to lactogenic hormone treatment; however their phenotype appeared to be plastic. Cultured in EGF-free medium, they exhibited enhanced cell-extracellular matrix adhesions and deficient cell-cell junctions, whereas long-term treatment with EGF induced a decrease of focal contact number and establishment of cell-cell junctions, resulting in downregulation of K14 and P-cadherin expression at the protein and mRNA levels. To determine whether cell-extracellular matrix interactions mediated by integrins have a role in the regulation of the expression of K14 and P-cadherin, the amounts of transcripts for the two proteins were analysed in the basal cells, which were plated on the function-blocking antibodies against beta1 and alpha6 integrin chains, on fibronectin and on laminin 5. The amount of P-cadherin transcript was 2- to 4-fold higher in cells plated on the function-blocking anti-integrin antibodies and on the extracellular matrix proteins, as compared to cells plated on poly-L-lysine, whereas the K14 transcript levels were not significantly modified in response to adhesion. The data demonstrate that integrin-mediated cell interaction with extracellular matrix is directly implicated in the control of P-cadherin expression, and that EGF and cell-extracellular matrix adhesion events are important regulators of the basal mammary epithelial cell phenotype.

Animals↗

PubMed2005

Mechanical, biochemical, and extracellular matrix effects on vascular smooth muscle cell phenotype.

The vascular smooth muscle cell (VSMC) is surrounded by a complex extracellular matrix that provides and modulates a variety of biochemical and mechanical cues that guide cell function. Conventional two-dimensional monolayer culture systems recreate only a portion of the cellular environment, and therefore there is increasing interest in developing more physiologically relevant three-dimensional culture systems. This review brings together recent studies on how mechanical, biochemical, and extracellular matrix stimulation can be applied to study VSMC function and how the combination of these factors leads to changes in phenotype. Particular emphasis is placed on in vitro experimental studies in which multiple stimuli are combined, especially in three-dimensional culture systems and in vascular tissue engineering applications. These studies have provided new insight into how VSMC phenotype is controlled, and they have underscored the interdependence of biochemical and mechanical signaling. Future improvements in creating more complex in vitro culture environments will lead to a better understanding of VSMC biology, new treatments for vascular disease, as well as improved blood vessel substitutes.

Adaptation, Physiological↗

PubMed1990

Extracellular matrix from normal but not Steel mutant mice enhances melanogenesis in cultured mouse neural crest cells.

The Steel mutation is a non-cell-autonomous defect in mice that affects the development of several stem cell populations, including germ cells, hematopoietic cells, and neural crest-derived pigment cells. To characterize the environmental lesion caused by the Steel mutation, we have compared the ability of normal and mutant extracellular matrix material to support the differentiation of normal mouse neural crest cells in vitro. Extracellular matrix deposited by cultured skin cells isolated from normal fetuses enhanced melanogenesis by crest cells over that observed on plastic substrata. In contrast, matrix material produced by Steel-Dickie (Sld) fetal skin cells failed to enhance melanogenesis. Adrenergic differentiation by neural crest-derived cells was promoted equally by both normal and mutant extracellular matrix compared to control substrata. We conclude that the environmental defect in mutant embryos selectively affects a melanogenic subpopulation of neural crest cells and resides, at least in part, in the extracellular matrix.

Animals↗

PubMed1989

The extracellular matrix of Volvox carteri: molecular structure of the cellular compartment.

The extracellular matrix (ECM) of Volvox contains insoluble fibrous layers that surround individual cells at a distance to form contiguous cellular compartments. Using immunological techniques, we identified a sulfated surface glycoprotein (SSG 185) as the monomeric precursor of this substructure within the ECM. The primary structure of the SSG 185 poly-peptide chain has been derived from cDNA and genomic DNA. A central domain of the protein, 80 amino acid residues long, consists almost exclusively of hydroxyproline residues. The chemical structure of the highly sulfated polysaccharide covalently attached to SSG 185 has been determined by permethylation analysis. As revealed by EM, SSG 185 is a rod-shaped molecule with a 21-nm-long polysaccharide strand protruding from its central region. The chemical nature of the cross-links between SSG 185 monomers is discussed.

Amino Acid Sequence↗

PubMed2002

Expression of adhesion and extracellular matrix molecules in the developing human brain.

Cell adhesion molecules and extracellular matrix molecules have important roles in cell migration and connection. Their developmental expression has not been fully described in humans. In this report, these molecules were examined by immunohistochemistry in frontal tissue samples from 14- to 28-week-old fetuses aborted for obstetric reasons (n = 20) and four fetuses with nervous system abnormalities. Neural cell adhesion molecule (NCAM), tenascin, and laminin were expressed after 17 weeks. Neural cell adhesion molecule was observed in the neuropil, whereas tenascin and laminin also had cellular and vascular expression. Thrombospondin and fibronectin, apparent after 14 weeks, showed a redistribution from periventricular to outer cortical layers after midgestation. N-cadherin and integrin were observed in mid- and late gestation. Maternal or environmental conditions seemed to influence the pattern of expression. Fetuses with nervous system abnormalities had altered expression of several molecules. The descriptive data obtained in this study might constitute a basis for further studies investigating the role of these molecules in developmental abnormalities of the brain.

Brain↗

PubMedDate not supplied

Channel catfish, Ictalurus punctatus rafinesque, neutrophil adhesion to selected extracellular matrix proteins, lipopolysaccharide, and catfish serum.

Adhesion of leukocytes to endothelium and extracellular matrix proteins is an important step in the inflammatory process. Therefore, the adhesion of channel catfish neutrophils to a surface coated with extracellular matrix proteins, LPS, and non-immune catfish serum was evaluated. Stimulation of neutrophils with phorbol dibutyrate (PDBU) resulted in at least two-fold increases in cellular adhesion to all substrates tested except laminin. When EDTA was included during or after PDBU stimulation, neutrophil adhesion to fibrinogen and LPS coated surfaces was reduced to the level of unstimulated neutrophils or to 50-60% of that for stimulated neutrophils. Similarly, EDTA and Ca2+/Mg2+ deficient medium reduced homotypic aggregation of PDBU stimulated neutrophils to background levels. Adhesion of stimulated neutrophils to fibrinogen coated surfaces was inhibited 44, 33, and 50% when soluble fibrinogen, fibronectin, and serum, respectively, were used to block the adhesion assay. The tripeptide integrin adhesion recognition sequence, Arg-Gly-Asp (RGD), caused 83% reduction and the fibrinogen-binding inhibitor protein caused 10% reduction in binding of stimulated neutrophils to fibrinogen coated surfaces. Two hexapeptides tested did not reduce neutrophil adhesion to fibrinogen. The binding of channel catfish neutrophils to the matrices used in the present study is suggestive that integrin mediated adhesion occurs during biological and pathological processes of teleosts.

Animals↗

PubMed1993

Tenascin is a cytoadhesive extracellular matrix component of the human hematopoietic microenvironment.

Tenascin is a large extracellular matrix (ECM) glycoprotein found in restricted tissue locations in the adult organism. It is copiously synthesized in regenerative organs or regenerating tissues and by certain tumors. We have analyzed the expression of tenascin in human long term bone marrow cultures as well as in cryostat sections of native bone marrow and found it strongly expressed by the stromal cells of the microenvironment. Two different protein subunits of 280 and 220 kD were detected by immunoblotting. These two forms are derived most likely from two different mRNA splice variants of 6 and 8 kb detected by Northern blotting. The in vivo analysis of cryostat sections showed a codistribution with other ECM molecules such as fibronectin and collagen type III in the microenvironment surrounding the maturing hematopoietic cells. Using two independent cell adhesion assays tenascin could be shown to function as a cytoadhesive molecule for hematopoietic cells. These data suggest a direct involvement of tenascin in the retention of hematopoietic progenitor cells in the stroma.

Blotting, Northern↗

PubMed2005

Developmental regulation and coordinate reexpression of FKBP65 with extracellular matrix proteins after lung injury suggest a specialized function for this endoplasmic reticulum immunophilin.

AFKBP65 (65-kDa FK506-binding protein) is an endoplasmic reticulum (ER)-localized peptidyl-prolyl cis-trans isomerase predicted to play a role in the folding and trafficking of secretory proteins. In previous studies, we have shown that FKBP65 is developmentally regulated and associates with the extracellular matrix protein, tropoelastin, during its maturation and transport through the ER. In this study, we show that FKBP65 is expressed in the lung with the same developmental pattern as tropoelastin and other matrix proteins. To test the hypothesis that FKBP65 is upregulated at times when extracellular matrix proteins are being actively synthesized and assembled, adult mice were treated with bleomycin to cause reinitiation of matrix protein production during the ensuing development of pulmonary fibrosis. After bleomycin instillation, FKBP65 expression was reactivated in the lung with a pattern similar to that observed for tropoelastin and type I collagen. Using human lung fibroblast cultures, we showed that FKBP65 does not undergo the unfolded protein response, a response associated with an upregulation of resident ER proteins that occurs after increased ER stress. When fibroblasts were treated with transforming growth factor (TGF)-beta1, which is upregulated during the development of pulmonary fibrosis and known to induce matrix production, FKBP65 expression and synthesis was also increased. Similar to type I collagen and tropoelastin, this response was completely inhibited in a dose-dependent manner by GGTI-298, a geranylgeranyl transferase I inhibitor. Treatment of fibroblasts with an inhibitor of ribonucleic acid (RNA) polymerase II after TGF-beta1 treatment showed that the effect of TGF-beta1 was not because of increased stabilization of the FKBP65 messenger RNA. In summary, we have shown that FKBP65 is highly expressed in lung development, downregulated in the adult, and can be reactivated in a coordinated manner with extracellular matrix proteins after lung injury. The expression pattern of FKBP65, which is atypical for general ER foldases, suggests that FKBP65 has a distinct set of developmentally regulated protein ligands. The response to injury, which may be in part a direct response to TGF-beta1, assures the presence of FKBP65 in the ER of cells actively producing components of the extracellular matrix.

Animals↗