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M Kan

Publications and source records attributed to M Kan.

At least 19 recordsLinked to original sources
PubMed1992

Expression and immunochemical analysis of rat and human fibroblast growth factor receptor (flg) isoforms.

Potentially 96 splice variants among four genes that code for the human heparin-binding fibroblast growth factor receptor family complicate study of structure, metabolism, and function of single isoforms in mammalian cells. As an alternative, we expressed structural subdomains and isoforms of the flg receptor gene in bacteria and baculoviral-infected insect cells. We developed and characterized a panel of 16 isoform and domain-specific polyclonal and monoclonal antibodies. The panel of antibodies was used to distinguish mature glycosylated ligand-binding and kinase-active and -inactive recombinant isoforms in baculoviral insect cells and transfected mammalian cells and natural isoforms in rat prostate and human liver cells. The results revealed a cell type-specific expression of the flg gene and isoforms that result from combinations of splice variations. Reactive epitopes of monoclonal antibodies against both the three (alpha) and two (beta) immunoglobulin-like disulfide loop extracellular domain isoforms were mapped by cross-reactivity with synthetic polypeptide sequences and deletion mutants expressed in bacteria. The native alpha and beta receptor isoforms differed in display of shared epitopes and suggested that the NH2-terminal Loop I and COOH-terminal Loops II and III of the alpha isoform are interactive. Although the common Loops II and III appear qualitatively sufficient for ligand binding, the results suggest that tertiary relationships among loops in the three and two loop isoforms are distinct and, therefore, the two isoforms may have distinct activities. Spatial models for arrangement of immunoglobulin-like loops in the extracellular domain of the two isoforms are presented.

Amino Acid Sequence↗
PubMed1992

Substitution of putative half-cystine residues in heparin-binding fibroblast growth factor receptors. Loss of binding activity in both two and three loop isoforms.

Alternate use of an exon coding for an 89-residue NH2 terminal immunoglobulin-like disulfide loop results in isoforms of the heparin-binding fibroblast growth factor receptor (FGF-R) with three (FGF-R alpha) and two (FGF-R beta) Ig-like loops in the extracellular domain. Both FGF-R alpha and FGF-R beta isoforms exhibit qualitatively similar ligand-binding activities. In this report, we show by site-directed mutagenesis and analysis of ligand-binding activity in transfected cells that substitution of a cysteine that potentially forms an intra-loop disulfide in either juxtamembrane Loop II or III disrupted maturation and formation of the ligand-binding site in both FGF-R alpha and FGF-R beta isoforms. Neither three loop FGF-R alpha constructions coding for intact Loops I and II adjacent to defective Loop III nor intact Loops I and III separated by defective Loop II exhibited ligand-binding activity. In addition, a two-loop molecule of tandem Loops I and III was inactive. The results suggest that single Loops I, II, or III of FGF-R are insufficient to form a ligand-binding site. Loop I does not form an independent ligand-binding site with either Loop II or III, but interacts with a common ligand-binding site formed by Loops II and III (Xu, J., Nakahara, M., Crabb, J. W., Shi, E., Matuo, Y., Fraser, M., Kan, M., Hou, J., and McKeehan, W. L. (1992) J. Biol. Chem. 267, 17792-17803, 1992).

Amino Acid Sequence↗
PubMed1992

Identification of two enhancer elements in the gene encoding the type 1 glucose transporter from the mouse which are responsive to serum, growth factor, and oncogenes.

The type 1 glucose transporter (GLUT1) gene encodes an integral membrane glycoprotein responsible for facilitating transfer of glucose across plasma membrane and is rapidly activated by serum, growth factors, and by oncogenic transformation. To elucidate the molecular mechanisms of regulation of GLUT1 gene expression, we isolated and characterized the mouse GLUT1 gene. DNA elements regulating transcription of the gene were analyzed in transient expression assays after transfection of NIH/3T3 cells with a low background chloramphenicol acetyltransferase (CAT) vector system pSVOOCAT. We identified two enhancer elements; the first one is located 2.7 kilobases upstream of the cap site of the gene which contains the homologous sequences with two 12-O-tetradecanoylphorbol-13-acetate-responsive elements (TREs), a serum response element, a cyclic AMP-responsive element (CRE) and three GC boxes, and the second one is located in the second intron of the gene which contains the homologous sequences with two TREs and one CRE. With the promoter alone the transcription of the gene is activated by src, only slightly activated by ras and is not activated by serum and platelet-derived growth factor. When the gene is accompanied by one of these enhancers, the transcription is activated by all these stimuli.

3T3 Cells↗
PubMed1992

cis-4-Hydroxy-L-proline and ethyl-3,4-dihydroxybenzoate prevent myogenesis of C2C12 muscle cells and block MyoD1 and myogenin expression.

cis-4-Hydroxy-L-proline (cis-OH-Pro) and ethyl-3,4-dihydroxybenzoate (EDHB), two distinct inhibitors of collagen synthesis, prevented myogenesis in C2C12 mouse skeletal muscle cells. Both inhibitors blocked myotube formation and the expression of sarcomeric myosin heavy chain. Northern blot analysis showed that cis-OH-Pro- and EDHB-treated C2C12 muscle cells did not express the myogenic regulatory genes, MyoD1 and myogenin, but continued to express non-muscle isoforms of actin (beta and gamma) and alpha-tropomyosin. 10TFL2-3B cells, a C3H10T1/2 cell line permanently transfected with myogenin cDNA, constitutively expressed exogenous myogenin in the presence of cis-OH-Pro but failed to activate endogenous myogenin and to undergo myogenesis. These results demonstrate that commitment to terminal differentiation and activation of myogenic regulatory genes requires active synthesis of the extracellular matrix component collagen.

Animals↗
PubMed1992

The effect of Ninjinyoeito on Werner's syndrome skin fibroblasts.

The effect of Ninjinyoeito on three cases of Werner's syndrome fibroblasts was investigated. In all three cases, groups which were treated with Ninjinyoeito showed improved life-span doubling levels compared to groups which were not treated with Ninjinyoeito. Also, all of the treated group in all three cases showed significantly higher values in the rates of DNA synthesis including two cases which showed significantly higher rates of protein synthesis.

Adult↗
PubMed1992

Small renal cell carcinoma: pathologic and radiologic correlation.

The authors correlated the radiologic and pathologic findings of 36 patients with small renal cell carcinoma (less than or equal to 3 cm in diameter). Tumors were discovered incidentally or by means of mass surveys with ultrasound (US). Computed tomography (CT) and angiography were performed before surgery. Of the 36 tumors, 24 were of solid (alveolar) architecture, five were of papillary architecture, three were of tubular architecture, and four were of multilocular cystic architecture. Cell arrangement was closely correlated with radiologic appearance, especially in regard to tumor vascularity and echogenicity. Histologically homogeneous tumors of solid architecture were hypoechoic at US and hypervascular at angiography and contrast medium-enhanced CT. Tumors of papillary, tubular, and multilocular cystic architecture were hyperechoic at US and hypovascular at angiography. There was no correlation of cell differentiation and cell architecture with echogenicity or tumor vascularity. Tumors with hemorrhage showed marked hyperattenuation at CT. A tumor capsule was observed in 25 patients (69%); this was demonstrated as a rim at US or angiography.

Adult↗
PubMed1992

Heparin-binding (fibroblast) growth factors are potential autocrine regulators of esophageal epithelial cell proliferation.

A serum-free culture system supplemented with neural tissue extract for normal human esophagus was applied to the culture of mouse esophageal epithelium. Similar to mouse mesenchyme and skin epithelium, esophageal epithelial lines (MEE) emerged after serial culture. The cells had an apparent unlimited life-span, but retained morphology and other characteristics of normal epithelial cells. A screen for growth factors that stimulated growth of MEE cells in the absence of neural extract revealed that epidermal growth factor (EGF) and heparin-binding (fibroblast) growth factors (HBGF) were most effective. An HBGF-like activity was apparent in extracts of rapidly proliferating, but not quiescent MEE cells at low or confluent densities. A cloned cell line (MEE/C8) was selected from MEE cell cultures in the absence of neural extract. MEE/C8 cells proliferated independent of either EGF or HBGF at rates equal to MEE cells, cell extracts exhibited HBGF-like activity at all stages of proliferation, and the cells formed invasive tumors in syngenic hosts. The HBGF-like activity present in extracts of tumorigenic MEE/C8 cells had properties similar to HBGF-1 (acidic fibroblast growth factor).

Animals↗
PubMed1992

[Cerebral salt wasting syndrome secondary to head injury: a case report].

A case of cerebral salt wasting syndrome secondary to head injury is reported here. A 4-year-old boy was admitted to our hospital with head injury. Neurological examination revealed no abnormal findings other than consciousness disturbance. Plain skull X-ray demonstrated a linear fracture of the bilateral parietal bones, and CT scan demonstrated subarachnoid hemorrhage of the tentorium of the cerebellum. He gradually improved, but on the 6th day deterioration of consciousness developed. At that time CT scan demonstrated no abnormal findings. Biochemical analysis showed hyponatremia (116mEq/L) with increased natriuresis. Although a high dose of NaCl was supplied, serum sodium levels did not normalize. So we suspected that SIADH might be causing the hyponatremia, and water restriction was started. He lost 1 kg in body weight over 3 days, but serum sodium levels remained low (118mEq/L) with increased natriuresis. We found that the hyponatremia was caused by cerebral salt wasting syndrome, so we treated the patient with fludrocortisone acetate. Consciousness disturbance improved two days after the medication with fludrocortisone acetate, and serum sodium levels became normal (137mEq/L) on the 27th day. The administration of fludrocortisone acetate was able to be stopped two months after admission, and then the patient was discharged without any neurological deficits. We discussed in detail the diagnosis and the treatment of cerebral salt wasting syndrome.

Child, Preschool↗
PubMed1991

Vascular endothelial cell growth factor (VEGF) produced by A-431 human epidermoid carcinoma cells and identification of VEGF membrane binding sites.

A distinct family of endothelial cell mitogens that are homologous to platelet-derived growth factor has recently been identified. Unlike other known endothelial cell mitogens, these vascular endothelial cell growth factors (VEGFs) are secreted and appear to act specifically on endothelial cells. We have purified VEGF 2083-fold to apparent homogeneity from protein-free culture medium conditioned by A-431 human epidermoid carcinoma cells. This A-431-derived VEGF was characterized as a homodimer composed of 22-kDa subunits with an N-terminal sequence that was similar to VEGFs produced by human HL-60 leukemic and U-937 histiocytic lymphoma cells. A-431 VEGF was used to identify specific and saturable binding sites for VEGF on human umbilical vein endothelial cells (HUVEC). By affinity cross-linking, VEGF-binding site complexes of 230, 170, and 125 kDa were detected on HUVEC. VEGF specifically induced the tyrosine phosphorylation of a 190-kDa polypeptide, which was similar in mass to the largest binding site detected by affinity cross-linking.

Amino Acid Sequence↗
PubMed1991

16-kilodalton heparin binding (fibroblast) growth factor type one appears in a stable 40-kilodalton complex after receptor-dependent internalization.

Incubation of 16-kDa 125I-labeled heparin binding (acidic fibroblast) growth factor type one (HBGF-1) with human hepatoma cells and normal rat hepatocytes resulted in the appearance of a stable 125I-labeled complex with an apparent molecular mass of 40 kDa. The complex could be isolated with specific antibodies against HBGF-1. In contrast to membrane receptor-bound 125I-HBGF-1, the complex was resistant to dissociation by detergents, acid, heat, and reducing or denaturing agents. Formation of a stable complex did not require treatment with cross-linking agents. Appearance of the 40-kDa complex was dependent on time, temperature, and enriched culture medium. Conditions that enhanced or reduced display of specific HBGF-1 membrane receptor sites enhanced or reduced the appearance of the 40-kDa complex. Dansylcadaverine, chloroquine, and staurosporine blocked the appearance of the 40-kDa complex concurrent with the blockage of internalization of the receptor-bound HBGF-1. Two-dimensional gel electrophoretic analysis, metabolic labeling with L-[35S]cysteine, and recovery of 16-kDa HBGF-1 from the 40-kDa complex after base treatment suggest involvement of a 24-kDa cellular protein in the complex formation. These results suggest a potentially novel receptor-dependent pathway for metabolism of HBGF-1.

Alkaloids↗
PubMed1991

Hepatocyte growth factor/hepatopoietin A stimulates the growth of rat kidney proximal tubule epithelial cells (RPTE), rat nonparenchymal liver cells, human melanoma cells, mouse keratinocytes and stimulates anchorage-independent growth of SV-40 transformed RPTE.

Hepatocyte growth factor/hepatopoietin A is a mitogen for primary hepatocytes and may mediate regeneration after liver damage. To date, the activity of this novel factor has been restricted to hepatocytes. We now show that the factor is also a mitogen for a number of primary epithelial cells but is inactive with human foreskin fibroblasts, human endothelial cells and HEP3B cells. The factor also substitutes for HBGF-2 (basic FGF) in stimulating the anchorage-independent growth of SV-40 transformed rat kidney epithelial cells. Therefore, hepatocyte growth factor/hepatopoietin A appears to act on a variety of epithelial, but not mesenchymal, cells which respond to HBGFs.

Animals↗
PubMed1991

Regulation of rat proximal tubule epithelial cell growth by fibroblast growth factors, insulin-like growth factor-1 and transforming growth factor-beta, and analysis of fibroblast growth factors in rat kidney.

Growth factors may play an important role in regulating the growth of the proximal tubule epithelium. To determine which growth factors could be involved, we have investigated the mitogenicity of various purified factors in rat kidney proximal tubule epithelial (RPTE) cells cultured in defined medium. Fibroblast growth factors, aFGF (acidic FGF) and bFGF (basic FGF), stimulate DNA synthesis in a dose-dependent manner, with ED50 values of 4.5 and 3.2 ng/ml, respectively; their effects are not additive. With cholera toxin in the medium, both aFGF and bFGF can replace insulin or epidermal growth factor (EGF) to attain the maximum level of cell growth, but they cannot replace cholera toxin. Cholera toxin specifically potentiates the effects of FGFs on DNA synthesis. At high cell density, both insulin and insulin-like growth factor 1 (IGF-1) induce DNA synthesis more effectively than EGF, FGFs and cholera toxin. The high concentration (0.2-1.0 microgram/ml) of insulin required for cell growth can be replaced by a low concentration of IGF-1 (10-20 ng/ml), indicating that insulin probably acts through a low affinity interaction with the IGF-1 receptor. Transforming growth factor-beta 1 (TGF-beta 1) inhibits DNA synthesis induced by individual factors and combinations of factors in a concentration-dependent manner. Northern blot analysis shows that mRNA for TGF-beta 1, IGF-1, and aFGF, but not bFGF are present in rat kidney. Western blot analysis and bioassay data confirmed that the majority of FGF-like protein in rat kidney is aFGF. The data suggest that in addition to EGF, IGFs, and TGF-beta, FGFs may also be important kidney-derived regulators of proximal tubule epithelial cell growth in vivo and in vitro.

Animals↗
PubMed1991

Heparin-binding (fibroblast) growth factors are potential autocrine regulators of esophageal epithelial cell proliferation.

A serum-free culture system supplemented with neural tissue extract for normal and tumor human esophagi was applied to the culture of mouse esophageal epithelium. Similar to mouse mesenchyme and skin epithelium, esophageal epithelial lines (MEE) emerged after serial culture. The cells had an apparent unlimited life span but retained morphology and other characteristics of normal epithelial cells. The cells formed a small cyst consisting of keratinized squamous epithelium in syngenic hosts. A screen for growth factors that stimulated growth of the nonmalignant MEE cells in the absence of neural extract revealed that epidermal growth factor (EGF) and heparin-binding (fibroblast) growth factors (HBGF) were most effective. An HBGF-like activity was apparent in extracts of rapidly proliferating but not quiescent MEE cells at low or confluent densities. A cloned cell line (MEE/C8) was selected from MEE cell cultures in the absence of neural extract. MEE/C8 cells proliferated independent of either EGF or HBGF at rates equal to MEE cells, cell extracts exhibited HBGF-like activity at all stages of proliferation, and the cells formed large invasive tumors in syngenic hosts. The HBGF-like activity present in extracts of tumorigenic MEE/C8 and proliferating nonmalignant MEE cells had properties similar to HBGF-1 (acidic fibroblast growth factor). These results constitute a cultured mouse esophageal epithelial cell model for study of conversion of immortalized premalignant cells to malignant cells, and suggest that conversion from a state of cell cycle-dependent autocrine expression of one or more members of the HBGF family to a state of constitutive expression correlates with and may contribute to malignancy.

Animals↗
PubMed1991

Primary culture of human gallbladder epithelial cells.

Growth requirements of epithelial cells isolated from the human gallbladder were examined. The growth of gallbladder epithelial cells (GBEC) was stimulated by medium conditioned with gallbladder fibroblasts (CM-GBF) in a dose-dependent manner. The conditioned medium derived from human embryo lung fibroblasts also showed similar growth stimulating activity for GBEC, suggesting that fibroblasts secrete a factor or factors which induce GBEC growth in vitro. CM-GBF in the presence of 10% fetal bovine serum increased GBEC growth up to 10 times the growth in the absence of CM-GBF supplement. GBEC cultured with CM-GBF showed hexagonal shape under a phase-contrast microscope, and also expressed cytokeratin and mucopolysaccharide in the cytoplasm, both of which are specific for GBEC. Electron microscopy revealed desmosomes and tight junctions between the cells and microvilli on the apical plasma membrane, suggesting that they regained morphological polarity in the medium containing CM-GBF. These results shows that CM-GBF is essential for the growth and the differentiation of GBEC in culture.

Animals↗
PubMed1990

Fibronectin, not laminin, mediates heparin-dependent heparin-binding growth factor type I binding to substrata and stimulation of endothelial cell growth.

Fibronectin and heparin-binding growth factors (HBGF) are essential for growth of cultured endothelial cells. The stimulation of endothelial cell growth by HBGF type one (HBGF-1) in particular requires heparin or a similar glycosaminoglycan. The requirement for fibronectin and heparin for HBGF-1-stimulated endothelial cell growth may be related. HBGF-1 absorbed to the natural subcellular matrix of endothelial cells supports cell growth. [125I]HBGF-1 specifically associates with a sequentially reconstituted matrix of collagen-fibronectin-heparin, and HBGF-1 absorbed to the reconstituted matrix supports growth of the endothelial cells. A reconstituted matrix of collagen-laminin-heparin neither supported binding of [125I]HBGF-1 nor HBGF-1-stimulated endothelial cell growth. Association kinetics of [125I]HBGF-1 to heparinlike sites and membrane receptor sites on endothelial cell monolayers suggest that fibronectin-heparinlike binding sites in the subcellular matrix may be an obligatory reservoir of active HBGF-1 that binds to specific cell membrane receptors.

Cell Division↗
PubMed1990

Heparin-binding (fibroblast) growth factors type one and two genes are co-expressed in proliferating normal human vascular endothelial and smooth muscle cells in culture.

Complimentary ribonucleic acid (cRNA) probes were used to detect expression of the genes for heparin binding (fibroblast) growth factor type one (HBGF-1) and two (HBGF-2) in cultured endothelial and smooth muscle cells from normal human blood vessels. Hybridization in situ revealed that transcripts for both HBGF-1 and HBGF-2 are expressed in endothelial cells from both umbilical vein and aorta. Relative intensity of radioactive grains suggest that HBGF-1 gene expression may exceed HBGF-2 expression in aortic smooth muscle cells. Collective expression of both HBGF-1 and HBGF-2 in smooth muscle cells may exceed that in endothelial cells.

Cell Division↗
PubMed1990

Opposite effects of monokines (interleukin-1 and tumor necrosis factor) on proliferation and heparin-binding (fibroblast) growth factor binding to human aortic endothelial and smooth muscle cells.

Heparin-binding (fibroblast) growth factors (HBGF) are mitogens for both human aortic endothelial and smooth muscle cells. Under similar conditions, both vascular cells display similar numbers of specific HBGF binding sites with similar apparent affinity for HBGF. The monokines, interleukin-1 and tumor necrosis factor, inhibit endothelial cell growth and stimulate smooth muscle cell growth. The opposite mitogenic effects correlate with reduction and increase in HBGF receptor number displayed by endothelial and smooth muscle cells, respectively. These results suggest that the two monokines may depress endothelial cell regeneration and augment smooth muscle cell hyperplasia by differential modulation of the HBGF receptor in the two vascular cell types.

Aorta↗