Joachim Sasse - Academia.edu (original) (raw)
Papers by Joachim Sasse
Journal of Cell Biology, 1991
Compared to basic fibroblast growth factor (bFGF), a widely distributed, broad spectrum mitogen a... more Compared to basic fibroblast growth factor (bFGF), a widely distributed, broad spectrum mitogen and mesoderm inducer, acidic fibroblast growth factor (aFGF) is reported to have an essentially neural distribution and to be undetectable in the early embryo. In the present investigation, we used immunoblotting and immunochemistry to assess the cellular and tissue distributions of aFGF and bFGF in 11-20-d rat embryos. Immunoblotting of crude and heparin-bound embryo extracts revealed faint bands at the expected 17-18-kD and predominant bands at an apparent molecular mass of 26 to 28-kD (despite reducing conditions) using multiple specific antibodies for aFGF and bFGF. Pretreatment with 8 M urea yielded 18-20-kD aFGF and bFGF and some 24-26-kD bFGF. Immunoreactivity for both aFGF and bFGF was positive and similar in the cytoplasm, nuclei, and extracellular matrix of cells of neuroectodermal and mesodermal origin, while it was negative in endoderm-derived cells. The distribution of immuno...
The American journal of pathology, 1988
The basement membranes of bovine cornea are found to contain an angiogenic endothelial cell mitog... more The basement membranes of bovine cornea are found to contain an angiogenic endothelial cell mitogen, basic fibroblast growth factor (FGF), as determined by heparin-affinity chromatography, immunoblotting, immunofluorescence, and stimulation of capillary endothelial cell proliferation. The growth factor appears to be bound to heparan sulfate and is released from the cornea by treatment with heparin, a hexasaccharide heparin fragment, heparan sulfate, or heparanase, but not by chondroitin sulfate or chondroitinase. These findings indicate that basement membranes of the cornea may serve as physiologic storage depots for an angiogenic molecule. Abnormal release of this growth factor could be responsible for corneal neovascularization in a variety of ocular diseases. Physiologic and pathologic neovascularization in other tissues may also be initiated by release of stored angiogenic factors from the basement membrane. The sequestration of angiogenic endothelial mitogens in the basement me...
Journal of Biological Chemistry, 1992
The small, leucine-rich proteoglycans, decorin and biglycan, are prominent components of many ext... more The small, leucine-rich proteoglycans, decorin and biglycan, are prominent components of many extracellular matrices and are differentially regulated in various tissues. We have examined the effects of retinoic acid (RA) on the expression of biglycan and decorin at the protein and mRNA levels in cultured bovine articular chondrocytes. Biglycan protein expression is rapidly turned off after 1-2 days of treatment with RA. In contrast, decorin protein expression is increased 12-18-fold following 3 days of RA treatment. The level of biglycan mRNA was also rapidly reduced upon RA treatment, mirroring the protein expression. The reduction was apparent by 6 h, and, by 4 days, the levels were nearly undetectable. In contrast, decorin mRNA was induced upon treatment with RA. The increase in decorin message levels was first apparent by 24 h, reaching maximum by 2 days, and remained constant through 4 days. The repression of biglycan mRNA displayed equal sensitivity to RA concentrations from 10(-5) to 10(-9) M. Decorin mRNA was induced in a dose-dependent fashion by RA. Retinoic acid at a concentration of 10(-5) M, the highest dose examined, resulted in maximal induction of the message, and control levels were obtained with 10(-8) M. The protein synthesis inhibitor cycloheximide inhibited the induction of decorin mRNA, indicating that the induction by RA was a secondary event. In contrast, the repression of biglycan by RA was not significantly altered by cycloheximide, showing that the repression was a direct effect. Actinomycin D inhibited the induction of decorin mRNA, indicating that transcription was required for the induction. Nuclear run-on assays confirmed that RA was regulating biglycan mRNA expression at the transcription level. A 24-h RA treatment decreased the level of transcription of the biglycan gene 5-fold. In contrast, no increase in transcription from the decorin gene could be detected by nuclear run-on assays. Therefore, the elevation in decorin mRNA levels observed after RA treatment was the result of a post-transcriptional event, most likely the consequence of stabilization of the message. This study demonstrates that the genes for these two similar proteoglycans are under very different forms of regulation by RA in chondrocytes. The pattern of differential expression of biglycan and decorin could serve as an additional marker for indicating changes of the cartilage phenotype.
The Journal of Urology, 1987
Human male urine contains large amounts of prostatic acid phosphatase (PAP). Although pros ta tic... more Human male urine contains large amounts of prostatic acid phosphatase (PAP). Although pros ta tic fluid was thought to be the major source of urinary PAP, results of our earlier studies indicate that female urine also contains significant amounts of PAP, suggesting the possibility of an extraprostatic source of PAP in male urine as well. The present study was conducted in an att0111µt to ascertain the presence and source of extra-prostatic urinary PAP. Urine samples were obtained from patients before and after total prostatectomy for prostate carcinoma. PAP levels in these samples were analyzed by radioimmunoassay (RIA) and expressed as ng. PAP per mg. urinary creatinine. Urinary PAP levels in preoperative patients and in healthy males were 258±136 (N-3) and 180±39 (N-5) respectively,
Journal of Cell Biology, 1984
We have addressed the problem of the segregation of cell lineages during the development of carti... more We have addressed the problem of the segregation of cell lineages during the development of cartilage and muscle in the chick limb bud. The following experiments demonstrate that early limb buds consist of at least two independent subpopulations of committed precursor cells--those in (a) the myogenic and (b) the chondrogenic lineage--which can be physically separated. Cells obtained from stage 20, 21, and 22 limb buds were cultured for 5 h in the presence of a monoclonal antibody that was originally isolated for its ability to detach preferentially myogenic cells from extracellular matrices. The detached limb bud cells were collected and replated in normal medium. Within 2 d nearly all of the replated cells had differentiated into myoblasts and myotubes; no chondroblasts differentiated in these cultures. In contrast, the original adherent population that remained after the antibody-induced detachment of the myogenic cells differentiated largely into cartilage and was devoid of muscl...
Journal of Visualized Experiments, 2008
Proceedings of the National Academy of Sciences, 1986
A human hepatoma cell line synthesizes, as evidenced by metabolic labeling, an endothelial cell m... more A human hepatoma cell line synthesizes, as evidenced by metabolic labeling, an endothelial cell mitogen that is found to be mostly cell associated. The hepatoma-derived growth factor (HDGF) has been purified to homogeneity by a combination of Bio-Rex 70, heparin-Sepharose, and reverse-phase chromatography; it is a cationic polypeptide with a molecular weight of about 18,500-19,000. HDGF is structurally related to basic fibroblast growth factor (FGF). Immunological analysis demonstrates that antiserum prepared against a synthetic peptide corresponding to the amino-terminal sequence of basic FGF cross-reacts with HDGF when analyzed by electrophoretic blotting and by immunoprecipitation. Sequence analysis of tryptic fragments demonstrates that HDGF contains sequences that are homologous to both amino-terminal and carboxyl-terminal sequences of basic FGF.
Clinical Orthopaedics and Related Research, 1994
A canine model of distraction osteogenesis has recently been developed that permitted the evaluat... more A canine model of distraction osteogenesis has recently been developed that permitted the evaluation of bone formation and its vascularization during bifocal callotasis. In this model, the authors examined the composition of the blood vessels during distraction osteogenesis of the mandible for laminin and for Type IV collagen, both constituents of the vascular basement membrane. At the fibrous distraction site, at the juncture of the free cortical surface and the regenerated bone, and at the abutting cortical surfaces at the distal margin of the defect, laminin and Type IV collagen were present in all vessels.
Proceedings of the National Academy of Sciences, 1987
Bovine aortic and corneal endothelial cells synthesize a growth factor that remains mostly cell-a... more Bovine aortic and corneal endothelial cells synthesize a growth factor that remains mostly cell-associated but can also be extracted from the subendothelial extracellular matrix (ECM) deposited by these cells. The endothelial cell-derived growth factors extracted from cell lysates and from the extracellular matrix appear to be structurally related to basic fibroblast growth factor by the criteria that they bind to heparin-Sepharose and are eluted at 1.4-1.6 M NaCl, have a molecular weight of about 18,400, cross-react with anti-basic fibroblast growth factor antibodies when analyzed by electrophoretic blotting and immunoprecipitation, and are potent mitogens for bovine aortic and capillary endothelial cells. It is suggested that endothelium can store growth factors capable of autocrine growth promotion in two ways: by sequestering growth factor within the cell and by incorporating it into the underlying extracellular matrix.
Migration of endothelial cells is involved in normal and pathological angiogenesis and in reendot... more Migration of endothelial cells is involved in normal and pathological angiogenesis and in reendothelialization after vascular injury or rupture of atherosclerotic plaques. Several types of endothelial cells are known to synthesize basic fibroblast growth factor (bFGF); in some of these, migration is increased by exogenous bFGF and inhibited by anti-bFGF antibodies. Using immunocytochemical techniques and RNase protection analysis, we studied endothelial cells from bovine coronary arteries and veins as well as from adrenal microvessels. We found that bFGF mRNA and peptide were present in confluent endothelial cells and were upregulated during migration stimulated by removal of some cells from the monolayer. During migration, extracellular matrix stores of bFGF were depleted, and bFGF immunoreactivity began to accumulate in the cytoplasm of endothelial cells between 2 and 6 E a ndothelial cell migration is a feature of angiogenesis during embryogenesis, wound repair, and tumor growth&...
Cancer Research, Apr 15, 1988
Growth factor activity was partially purified from human renal tumors and a human bladder cancer ... more Growth factor activity was partially purified from human renal tumors and a human bladder cancer cell line by heparin-Sepharose chromatography. This activity stimulated bovine capillary endothelial cell prolifer ation and DNA synthesis in BALB/c 3T3 cells. Partially purified growth factor preparations from these tumors contained a protein with an ap proximate molecular weight of 17,000 which was recognized by a polyclonal antiserum raised against a peptide fragment of basic fibroblast growth factor (FGF). This growth factor activity appears to be related to basic fibroblast growth factor. Measurement of FGF-like activity in 50 urine samples from 32 adult males showed that 55% (6 of 11) of the urine samples from patients with bladder cancer and 100% (7 of 7) of the urine samples from patients with kidney cancer contained activity equivalent to more than 20 ng of basic FGF/h of urine production. In contrast, only 6% (2 of 32) of the urine samples from controls, patients with a benign disease, or patients with a history of bladder or kidney cancer contained this level of growth factor activity. These results suggest that patients with bladder or kidney cancer release an FGF-like factor into urine which may be used as a marker for these tumors.
Chemie in unserer Zeit, 1971
Otolaryngology -- Head and Neck Surgery, 1994
A model of bifocal distraction osteogenesis in the canine model was used to assess and quantitate... more A model of bifocal distraction osteogenesis in the canine model was used to assess and quantitate the mineral content of the newly forming bone within the canine mandible. A 2-cm defect was created in the body of the mandible, and after a posterior osteotomy, the transport disk was advanced at 0.25 mm per 8 hours for 21 days and then held in rigid fixation for an additional week. As a control for this study, three additional dogs underwent the same procedure with the exception that the transport disk was not advanced. Electron dispersive spectroscopy analysis was performed on the newly formed regenerate bone and compared with areas of existing cortical bone of both the transport disk and the mandible. In the control model, special note was made of the pericortical callus at the osteotomy site as well as of the regenerative bone that filled the 2-cm defect in the body of the mandible. Calcium/phosphorous ratios were used to assess the composition of the mineralized regions of the man...
Methods in Enzymology, 1987
The Journal of urology, 1988
Homogenates of human testes, epididymides and prostate, and calf testes and epididymides are mito... more Homogenates of human testes, epididymides and prostate, and calf testes and epididymides are mitogenic for cultured human foreskin fibroblasts. The growth factors appear similar in that they are inactivated by boiling and acid, but not by treatment with reducing agent. The growth factor in human and bovine testes was partially purified from tissue homogenates, prepared in high ionic strength buffer (pH 7.6) containing protease inhibitors, by ammonium sulfate precipitation and two cycles of heparin-Sepharose chromatography. The growth factor in calf testes was also partially purified from tissue extracted in ammonium sulfate without protease inhibitors, acidified to pH 4.5, and precipitated by ammonium sulfate followed by two cycles of heparin-affinity chromatography. A predominant 17,500 molecular weight (MW) growth factor was identified from alkaline homogenates of human and calf testes by its reactivity with antisera prepared against synthetic peptides whose sequences corresponded...
Cancer research, Jan 15, 1988
Growth factor activity was partially purified from human renal tumors and a human bladder cancer ... more Growth factor activity was partially purified from human renal tumors and a human bladder cancer cell line by heparin-Sepharose chromatography. This activity stimulated bovine capillary endothelial cell proliferation and DNA synthesis in BALB/c 3T3 cells. Partially purified growth factor preparations from these tumors contained a protein with an approximate molecular weight of 17,000 which was recognized by a polyclonal antiserum raised against a peptide fragment of basic fibroblast growth factor (FGF). This growth factor activity appears to be related to basic fibroblast growth factor. Measurement of FGF-like activity in 50 urine samples from 32 adult males showed that 55% (6 of 11) of the urine samples from patients with bladder cancer and 100% (7 of 7) of the urine samples from patients with kidney cancer contained activity equivalent to more than 20 ng of basic FGF/h of urine production. In contrast, only 6% (2 of 32) of the urine samples from controls, patients with a benign di...
The Journal of urology, 1988
Tissue homogenates of rat ventral prostate were examined for growth factor activity using a fibro... more Tissue homogenates of rat ventral prostate were examined for growth factor activity using a fibroblast mitogenesis assay. G-75 Sephadex gel filtration separated the growth factor activity into two peaks. A broad first peak contained 98% of the protein and several growth factor moieties. A smaller second peak (MW 6,000) contained epidermal growth factor (EGF) as determined by binding in both a competitive receptor binding assay and a radioimmunoassay using anti-mouse epidermal growth factor (anti-mEGF). The broad first peak also contained substantial amounts of EGF-like activity as higher MW forms of EGF. The broad first peak was further fractionated by heparin-Sepharose affinity chromatography. A major fraction with growth factor activity eluted at 1.5 M NaCl and this fraction was shown to contain bFGF by immunostaining with antisera prepared against synthetic peptides corresponding to amino acid sequences 1-12 (amino terminal), 33-43 (internal), and 136-145 (carboxy terminal) of ba...
The Anatomical Record, 1995
Cartilage canals are perichondral invaginations of blood vessels and connective tissue that are f... more Cartilage canals are perichondral invaginations of blood vessels and connective tissue that are found within the epiphyses of most mammalian long bones. Functionally, they provide a means of transport of nutrients to the hyaline cartilage, a mechanism for removal of metabolic wastes, and a conduit for stem cells that are capable of initiating and sustaining ossification of the chondroepiphysis. Morphological and biomolecular changes of the chondroepiphyses appear to potentiate ossification within the chondroepiphyses of developing bones. As both cell migration and vascular invasion are anchorage dependent processes, antibodies to laminin and Type IV collagen were used to assess compositional changes in the basement membrane of cartilage canals accompanying epiphyseal ossification. Differences in chronological appearance, as well as, in distribution between the two components were noted in the chondroepiphysis. Laminin was distributed throughout the connective tissue of cartilage canal at all stages of development, and not limited to an association with the vascular lumen. Type IV collagen was not present during the initial perichondral invagination. Although staining for Type IV collagen was later acquired, its distribution was restricted to a discontinuous rimming of the periphery of the canal, and a diffuse presence within the intra-canalicular mesenchyme. Concurrent with chondrocyte hypertrophy and mineralization of the hyaline matrix, rapid changes in both the morphology of the vessel and distribution of the antibodies were detected. In addition to the presence of laminin at the interface of the endothelium and the hyaline matrix, a wide distribution within the connective tissue components of the newly ossifying matrix of epiphyseal bone could be detected. Type IV collagen remained closely associated with the lumens of the intra-canalicular vessels throughout the transition. Following ossification of the secondary center, staining for Type IV collagen could then be detected in the bone-forming regions of transforming matrix as well, clearly delineating the individual vessels within the newly formed marrow spaces. This suggests that bone formation is intimately related to vessel staining for collagen type IV, and that acquired vessel competence is a facet of endochondral bone formation that results from provisional matrix changes. Furthermore, the data suggests that during bone formation under tension, basement membrane deposition can be demonstrated without an intermediary hyaline matrix hypertrophic chondrocyte phase. This data was interpreted to suggest that chondrocyte hypertrophy at the growth plate may be a reaction to vascular invasion, that in turn, stimulates adjacent chondrocyte proliferation.
Fibronectin, the major cell surface glycoprotein of fibroblasts, is absent from differentiated ca... more Fibronectin, the major cell surface glycoprotein of fibroblasts, is absent from differentiated cartilage matrix and chondrocytes in situ. However, dissociation of embryonic chick sternal cartilage with collagenase and trypsin, followed by inoculation in vitro reinitiates fibronectin synthesis by chondrocytes. Immunoflu- orescence microscopy with antibodies prepared against plasma fibronectin (cold insoluble globulin (CIG)) reveals fibronectin associated with the chondrocyte surface. Synthesis and secretion of fibronectin into the medium are shown by anabolic labeling with (3~S)methionine or (3H)glycine, and identification of the secreted proteins by immunoprecipitation and sodium dodecyl sulfate (SDS)-disc gel electrophoresis. When chondrocytes are plated onto tissue culture dishes, the pattern of surface- associated fibronectin changes from a patchy into a strandlike appearance. Where epithelioid clones of polygonal chondrocytes develop, only short strands of fibronectin appear pre...
The Journal of Cell Biology, 1988
Extracellular matrix proteins and their proteolytic products have been shown to modulate cell mot... more Extracellular matrix proteins and their proteolytic products have been shown to modulate cell motility. We have found that certain tumor cells display a chemotactic response to degradation products of the matrix protein elastin, and to an elastin-derived peptide, VGVAPG. The hexapeptide VGVAPG is a particularly potent chemotaxin for lung-colonizing Lewis lung carcinoma cells (line M27), with 5 nM VGVAPG eliciting maximal chemotactic response when assayed in 48-microwell chemotaxis chambers. Binding of the elastin-derived peptide to M27 cells was studied using a tyrosinated analog (Y-VGVAPG) to allow iodination. Scatchard analysis of [125I]Y-VGVAPG binding to viable M27 tumor cells at both 37 and 4 degrees C indicates the presence of a single class of high affinity binding sites. The dissociation constant obtained from these studies (2.7 X 10(-9) M) is equivalent to the concentration of VGVAPG required for chemotactic activity. The receptor molecule was identified as an Mr 59,000 spe...
Journal of Cell Biology, 1991
Compared to basic fibroblast growth factor (bFGF), a widely distributed, broad spectrum mitogen a... more Compared to basic fibroblast growth factor (bFGF), a widely distributed, broad spectrum mitogen and mesoderm inducer, acidic fibroblast growth factor (aFGF) is reported to have an essentially neural distribution and to be undetectable in the early embryo. In the present investigation, we used immunoblotting and immunochemistry to assess the cellular and tissue distributions of aFGF and bFGF in 11-20-d rat embryos. Immunoblotting of crude and heparin-bound embryo extracts revealed faint bands at the expected 17-18-kD and predominant bands at an apparent molecular mass of 26 to 28-kD (despite reducing conditions) using multiple specific antibodies for aFGF and bFGF. Pretreatment with 8 M urea yielded 18-20-kD aFGF and bFGF and some 24-26-kD bFGF. Immunoreactivity for both aFGF and bFGF was positive and similar in the cytoplasm, nuclei, and extracellular matrix of cells of neuroectodermal and mesodermal origin, while it was negative in endoderm-derived cells. The distribution of immuno...
The American journal of pathology, 1988
The basement membranes of bovine cornea are found to contain an angiogenic endothelial cell mitog... more The basement membranes of bovine cornea are found to contain an angiogenic endothelial cell mitogen, basic fibroblast growth factor (FGF), as determined by heparin-affinity chromatography, immunoblotting, immunofluorescence, and stimulation of capillary endothelial cell proliferation. The growth factor appears to be bound to heparan sulfate and is released from the cornea by treatment with heparin, a hexasaccharide heparin fragment, heparan sulfate, or heparanase, but not by chondroitin sulfate or chondroitinase. These findings indicate that basement membranes of the cornea may serve as physiologic storage depots for an angiogenic molecule. Abnormal release of this growth factor could be responsible for corneal neovascularization in a variety of ocular diseases. Physiologic and pathologic neovascularization in other tissues may also be initiated by release of stored angiogenic factors from the basement membrane. The sequestration of angiogenic endothelial mitogens in the basement me...
Journal of Biological Chemistry, 1992
The small, leucine-rich proteoglycans, decorin and biglycan, are prominent components of many ext... more The small, leucine-rich proteoglycans, decorin and biglycan, are prominent components of many extracellular matrices and are differentially regulated in various tissues. We have examined the effects of retinoic acid (RA) on the expression of biglycan and decorin at the protein and mRNA levels in cultured bovine articular chondrocytes. Biglycan protein expression is rapidly turned off after 1-2 days of treatment with RA. In contrast, decorin protein expression is increased 12-18-fold following 3 days of RA treatment. The level of biglycan mRNA was also rapidly reduced upon RA treatment, mirroring the protein expression. The reduction was apparent by 6 h, and, by 4 days, the levels were nearly undetectable. In contrast, decorin mRNA was induced upon treatment with RA. The increase in decorin message levels was first apparent by 24 h, reaching maximum by 2 days, and remained constant through 4 days. The repression of biglycan mRNA displayed equal sensitivity to RA concentrations from 10(-5) to 10(-9) M. Decorin mRNA was induced in a dose-dependent fashion by RA. Retinoic acid at a concentration of 10(-5) M, the highest dose examined, resulted in maximal induction of the message, and control levels were obtained with 10(-8) M. The protein synthesis inhibitor cycloheximide inhibited the induction of decorin mRNA, indicating that the induction by RA was a secondary event. In contrast, the repression of biglycan by RA was not significantly altered by cycloheximide, showing that the repression was a direct effect. Actinomycin D inhibited the induction of decorin mRNA, indicating that transcription was required for the induction. Nuclear run-on assays confirmed that RA was regulating biglycan mRNA expression at the transcription level. A 24-h RA treatment decreased the level of transcription of the biglycan gene 5-fold. In contrast, no increase in transcription from the decorin gene could be detected by nuclear run-on assays. Therefore, the elevation in decorin mRNA levels observed after RA treatment was the result of a post-transcriptional event, most likely the consequence of stabilization of the message. This study demonstrates that the genes for these two similar proteoglycans are under very different forms of regulation by RA in chondrocytes. The pattern of differential expression of biglycan and decorin could serve as an additional marker for indicating changes of the cartilage phenotype.
The Journal of Urology, 1987
Human male urine contains large amounts of prostatic acid phosphatase (PAP). Although pros ta tic... more Human male urine contains large amounts of prostatic acid phosphatase (PAP). Although pros ta tic fluid was thought to be the major source of urinary PAP, results of our earlier studies indicate that female urine also contains significant amounts of PAP, suggesting the possibility of an extraprostatic source of PAP in male urine as well. The present study was conducted in an att0111µt to ascertain the presence and source of extra-prostatic urinary PAP. Urine samples were obtained from patients before and after total prostatectomy for prostate carcinoma. PAP levels in these samples were analyzed by radioimmunoassay (RIA) and expressed as ng. PAP per mg. urinary creatinine. Urinary PAP levels in preoperative patients and in healthy males were 258±136 (N-3) and 180±39 (N-5) respectively,
Journal of Cell Biology, 1984
We have addressed the problem of the segregation of cell lineages during the development of carti... more We have addressed the problem of the segregation of cell lineages during the development of cartilage and muscle in the chick limb bud. The following experiments demonstrate that early limb buds consist of at least two independent subpopulations of committed precursor cells--those in (a) the myogenic and (b) the chondrogenic lineage--which can be physically separated. Cells obtained from stage 20, 21, and 22 limb buds were cultured for 5 h in the presence of a monoclonal antibody that was originally isolated for its ability to detach preferentially myogenic cells from extracellular matrices. The detached limb bud cells were collected and replated in normal medium. Within 2 d nearly all of the replated cells had differentiated into myoblasts and myotubes; no chondroblasts differentiated in these cultures. In contrast, the original adherent population that remained after the antibody-induced detachment of the myogenic cells differentiated largely into cartilage and was devoid of muscl...
Journal of Visualized Experiments, 2008
Proceedings of the National Academy of Sciences, 1986
A human hepatoma cell line synthesizes, as evidenced by metabolic labeling, an endothelial cell m... more A human hepatoma cell line synthesizes, as evidenced by metabolic labeling, an endothelial cell mitogen that is found to be mostly cell associated. The hepatoma-derived growth factor (HDGF) has been purified to homogeneity by a combination of Bio-Rex 70, heparin-Sepharose, and reverse-phase chromatography; it is a cationic polypeptide with a molecular weight of about 18,500-19,000. HDGF is structurally related to basic fibroblast growth factor (FGF). Immunological analysis demonstrates that antiserum prepared against a synthetic peptide corresponding to the amino-terminal sequence of basic FGF cross-reacts with HDGF when analyzed by electrophoretic blotting and by immunoprecipitation. Sequence analysis of tryptic fragments demonstrates that HDGF contains sequences that are homologous to both amino-terminal and carboxyl-terminal sequences of basic FGF.
Clinical Orthopaedics and Related Research, 1994
A canine model of distraction osteogenesis has recently been developed that permitted the evaluat... more A canine model of distraction osteogenesis has recently been developed that permitted the evaluation of bone formation and its vascularization during bifocal callotasis. In this model, the authors examined the composition of the blood vessels during distraction osteogenesis of the mandible for laminin and for Type IV collagen, both constituents of the vascular basement membrane. At the fibrous distraction site, at the juncture of the free cortical surface and the regenerated bone, and at the abutting cortical surfaces at the distal margin of the defect, laminin and Type IV collagen were present in all vessels.
Proceedings of the National Academy of Sciences, 1987
Bovine aortic and corneal endothelial cells synthesize a growth factor that remains mostly cell-a... more Bovine aortic and corneal endothelial cells synthesize a growth factor that remains mostly cell-associated but can also be extracted from the subendothelial extracellular matrix (ECM) deposited by these cells. The endothelial cell-derived growth factors extracted from cell lysates and from the extracellular matrix appear to be structurally related to basic fibroblast growth factor by the criteria that they bind to heparin-Sepharose and are eluted at 1.4-1.6 M NaCl, have a molecular weight of about 18,400, cross-react with anti-basic fibroblast growth factor antibodies when analyzed by electrophoretic blotting and immunoprecipitation, and are potent mitogens for bovine aortic and capillary endothelial cells. It is suggested that endothelium can store growth factors capable of autocrine growth promotion in two ways: by sequestering growth factor within the cell and by incorporating it into the underlying extracellular matrix.
Migration of endothelial cells is involved in normal and pathological angiogenesis and in reendot... more Migration of endothelial cells is involved in normal and pathological angiogenesis and in reendothelialization after vascular injury or rupture of atherosclerotic plaques. Several types of endothelial cells are known to synthesize basic fibroblast growth factor (bFGF); in some of these, migration is increased by exogenous bFGF and inhibited by anti-bFGF antibodies. Using immunocytochemical techniques and RNase protection analysis, we studied endothelial cells from bovine coronary arteries and veins as well as from adrenal microvessels. We found that bFGF mRNA and peptide were present in confluent endothelial cells and were upregulated during migration stimulated by removal of some cells from the monolayer. During migration, extracellular matrix stores of bFGF were depleted, and bFGF immunoreactivity began to accumulate in the cytoplasm of endothelial cells between 2 and 6 E a ndothelial cell migration is a feature of angiogenesis during embryogenesis, wound repair, and tumor growth&...
Cancer Research, Apr 15, 1988
Growth factor activity was partially purified from human renal tumors and a human bladder cancer ... more Growth factor activity was partially purified from human renal tumors and a human bladder cancer cell line by heparin-Sepharose chromatography. This activity stimulated bovine capillary endothelial cell prolifer ation and DNA synthesis in BALB/c 3T3 cells. Partially purified growth factor preparations from these tumors contained a protein with an ap proximate molecular weight of 17,000 which was recognized by a polyclonal antiserum raised against a peptide fragment of basic fibroblast growth factor (FGF). This growth factor activity appears to be related to basic fibroblast growth factor. Measurement of FGF-like activity in 50 urine samples from 32 adult males showed that 55% (6 of 11) of the urine samples from patients with bladder cancer and 100% (7 of 7) of the urine samples from patients with kidney cancer contained activity equivalent to more than 20 ng of basic FGF/h of urine production. In contrast, only 6% (2 of 32) of the urine samples from controls, patients with a benign disease, or patients with a history of bladder or kidney cancer contained this level of growth factor activity. These results suggest that patients with bladder or kidney cancer release an FGF-like factor into urine which may be used as a marker for these tumors.
Chemie in unserer Zeit, 1971
Otolaryngology -- Head and Neck Surgery, 1994
A model of bifocal distraction osteogenesis in the canine model was used to assess and quantitate... more A model of bifocal distraction osteogenesis in the canine model was used to assess and quantitate the mineral content of the newly forming bone within the canine mandible. A 2-cm defect was created in the body of the mandible, and after a posterior osteotomy, the transport disk was advanced at 0.25 mm per 8 hours for 21 days and then held in rigid fixation for an additional week. As a control for this study, three additional dogs underwent the same procedure with the exception that the transport disk was not advanced. Electron dispersive spectroscopy analysis was performed on the newly formed regenerate bone and compared with areas of existing cortical bone of both the transport disk and the mandible. In the control model, special note was made of the pericortical callus at the osteotomy site as well as of the regenerative bone that filled the 2-cm defect in the body of the mandible. Calcium/phosphorous ratios were used to assess the composition of the mineralized regions of the man...
Methods in Enzymology, 1987
The Journal of urology, 1988
Homogenates of human testes, epididymides and prostate, and calf testes and epididymides are mito... more Homogenates of human testes, epididymides and prostate, and calf testes and epididymides are mitogenic for cultured human foreskin fibroblasts. The growth factors appear similar in that they are inactivated by boiling and acid, but not by treatment with reducing agent. The growth factor in human and bovine testes was partially purified from tissue homogenates, prepared in high ionic strength buffer (pH 7.6) containing protease inhibitors, by ammonium sulfate precipitation and two cycles of heparin-Sepharose chromatography. The growth factor in calf testes was also partially purified from tissue extracted in ammonium sulfate without protease inhibitors, acidified to pH 4.5, and precipitated by ammonium sulfate followed by two cycles of heparin-affinity chromatography. A predominant 17,500 molecular weight (MW) growth factor was identified from alkaline homogenates of human and calf testes by its reactivity with antisera prepared against synthetic peptides whose sequences corresponded...
Cancer research, Jan 15, 1988
Growth factor activity was partially purified from human renal tumors and a human bladder cancer ... more Growth factor activity was partially purified from human renal tumors and a human bladder cancer cell line by heparin-Sepharose chromatography. This activity stimulated bovine capillary endothelial cell proliferation and DNA synthesis in BALB/c 3T3 cells. Partially purified growth factor preparations from these tumors contained a protein with an approximate molecular weight of 17,000 which was recognized by a polyclonal antiserum raised against a peptide fragment of basic fibroblast growth factor (FGF). This growth factor activity appears to be related to basic fibroblast growth factor. Measurement of FGF-like activity in 50 urine samples from 32 adult males showed that 55% (6 of 11) of the urine samples from patients with bladder cancer and 100% (7 of 7) of the urine samples from patients with kidney cancer contained activity equivalent to more than 20 ng of basic FGF/h of urine production. In contrast, only 6% (2 of 32) of the urine samples from controls, patients with a benign di...
The Journal of urology, 1988
Tissue homogenates of rat ventral prostate were examined for growth factor activity using a fibro... more Tissue homogenates of rat ventral prostate were examined for growth factor activity using a fibroblast mitogenesis assay. G-75 Sephadex gel filtration separated the growth factor activity into two peaks. A broad first peak contained 98% of the protein and several growth factor moieties. A smaller second peak (MW 6,000) contained epidermal growth factor (EGF) as determined by binding in both a competitive receptor binding assay and a radioimmunoassay using anti-mouse epidermal growth factor (anti-mEGF). The broad first peak also contained substantial amounts of EGF-like activity as higher MW forms of EGF. The broad first peak was further fractionated by heparin-Sepharose affinity chromatography. A major fraction with growth factor activity eluted at 1.5 M NaCl and this fraction was shown to contain bFGF by immunostaining with antisera prepared against synthetic peptides corresponding to amino acid sequences 1-12 (amino terminal), 33-43 (internal), and 136-145 (carboxy terminal) of ba...
The Anatomical Record, 1995
Cartilage canals are perichondral invaginations of blood vessels and connective tissue that are f... more Cartilage canals are perichondral invaginations of blood vessels and connective tissue that are found within the epiphyses of most mammalian long bones. Functionally, they provide a means of transport of nutrients to the hyaline cartilage, a mechanism for removal of metabolic wastes, and a conduit for stem cells that are capable of initiating and sustaining ossification of the chondroepiphysis. Morphological and biomolecular changes of the chondroepiphyses appear to potentiate ossification within the chondroepiphyses of developing bones. As both cell migration and vascular invasion are anchorage dependent processes, antibodies to laminin and Type IV collagen were used to assess compositional changes in the basement membrane of cartilage canals accompanying epiphyseal ossification. Differences in chronological appearance, as well as, in distribution between the two components were noted in the chondroepiphysis. Laminin was distributed throughout the connective tissue of cartilage canal at all stages of development, and not limited to an association with the vascular lumen. Type IV collagen was not present during the initial perichondral invagination. Although staining for Type IV collagen was later acquired, its distribution was restricted to a discontinuous rimming of the periphery of the canal, and a diffuse presence within the intra-canalicular mesenchyme. Concurrent with chondrocyte hypertrophy and mineralization of the hyaline matrix, rapid changes in both the morphology of the vessel and distribution of the antibodies were detected. In addition to the presence of laminin at the interface of the endothelium and the hyaline matrix, a wide distribution within the connective tissue components of the newly ossifying matrix of epiphyseal bone could be detected. Type IV collagen remained closely associated with the lumens of the intra-canalicular vessels throughout the transition. Following ossification of the secondary center, staining for Type IV collagen could then be detected in the bone-forming regions of transforming matrix as well, clearly delineating the individual vessels within the newly formed marrow spaces. This suggests that bone formation is intimately related to vessel staining for collagen type IV, and that acquired vessel competence is a facet of endochondral bone formation that results from provisional matrix changes. Furthermore, the data suggests that during bone formation under tension, basement membrane deposition can be demonstrated without an intermediary hyaline matrix hypertrophic chondrocyte phase. This data was interpreted to suggest that chondrocyte hypertrophy at the growth plate may be a reaction to vascular invasion, that in turn, stimulates adjacent chondrocyte proliferation.
Fibronectin, the major cell surface glycoprotein of fibroblasts, is absent from differentiated ca... more Fibronectin, the major cell surface glycoprotein of fibroblasts, is absent from differentiated cartilage matrix and chondrocytes in situ. However, dissociation of embryonic chick sternal cartilage with collagenase and trypsin, followed by inoculation in vitro reinitiates fibronectin synthesis by chondrocytes. Immunoflu- orescence microscopy with antibodies prepared against plasma fibronectin (cold insoluble globulin (CIG)) reveals fibronectin associated with the chondrocyte surface. Synthesis and secretion of fibronectin into the medium are shown by anabolic labeling with (3~S)methionine or (3H)glycine, and identification of the secreted proteins by immunoprecipitation and sodium dodecyl sulfate (SDS)-disc gel electrophoresis. When chondrocytes are plated onto tissue culture dishes, the pattern of surface- associated fibronectin changes from a patchy into a strandlike appearance. Where epithelioid clones of polygonal chondrocytes develop, only short strands of fibronectin appear pre...
The Journal of Cell Biology, 1988
Extracellular matrix proteins and their proteolytic products have been shown to modulate cell mot... more Extracellular matrix proteins and their proteolytic products have been shown to modulate cell motility. We have found that certain tumor cells display a chemotactic response to degradation products of the matrix protein elastin, and to an elastin-derived peptide, VGVAPG. The hexapeptide VGVAPG is a particularly potent chemotaxin for lung-colonizing Lewis lung carcinoma cells (line M27), with 5 nM VGVAPG eliciting maximal chemotactic response when assayed in 48-microwell chemotaxis chambers. Binding of the elastin-derived peptide to M27 cells was studied using a tyrosinated analog (Y-VGVAPG) to allow iodination. Scatchard analysis of [125I]Y-VGVAPG binding to viable M27 tumor cells at both 37 and 4 degrees C indicates the presence of a single class of high affinity binding sites. The dissociation constant obtained from these studies (2.7 X 10(-9) M) is equivalent to the concentration of VGVAPG required for chemotactic activity. The receptor molecule was identified as an Mr 59,000 spe...