Biomedical subjects

H C Bauer

Publications and source records attributed to H C Bauer.

At least 19 recordsLinked to original sources

PubMed1992

Comparison of gene expression of extracellular matrix molecules in brain microvascular endothelial cells and astrocytes.

By use of random-primed cDNA probes the expression of extracellular matrix molecules in cerebral microvascular endothelial cells (cEC) and in astrocytes from mouse brain was examined. Two phenotypically different batches of cloned cEC were used. Expression of major adhesive ECM molecules, constituting the endothelial basement membrane (i.e., fibronectin, laminin A, B and collagen IV) and of other attachment factors, such as SPARC (osteonectin), tenascin and thrombospondin 1, was examined. We have demonstrated that cEC of different morphology display variations in the expression of fibronectin (FN), thrombospondin 1 (TSP1) and collagen IV (C IV). Astrocytes were shown to contain FN, TSP1, TN and SPARC mRNA. Unexpectedly, SPARC mRNA could not be detected in any of the capillary endothelial cells examined. Therefore, we suggest that astrocytes are likely to be involved in endothelial differentiation and function in the central nervous system via ECM molecule secretion.

Animals↗

PubMed1992

Expression and functional activity of P-glycoprotein in cultured cerebral capillary endothelial cells.

Analysis of a panel of endothelial cells passaged between 5 and 25 times and derived from various organs and species demonstrated that murine and porcine cerebral capillary endothelial cells actively excluded the fluorescent dye rhodamine 123, a substrate of P-glycoprotein. In addition, rhodamine 123 accumulation could be enhanced by the multidrug resistance chemosensitizer verapamil, known to reduce P-glycoprotein-mediated drug efflux. Cloned murine and porcine cerebral capillary endothelial cells were immunoreactive with the C219 monoclonal antibody to P-glycoprotein, and a C219 epitope-specific blocking peptide could abolish staining. The antiproliferative and cytotoxic effects of vincristine, but not cis-platinum(II) diamminedichloride, were increased by the addition of either verapamil or cyclosporin A to brain endothelial cell cultures in a 72-h assay, as determined by [3H]thymidine incorporation and total protein measurement. Cyclosporin A was a more effective reversal agent than verapamil. Thus, a P-glycoprotein isoform may be constitutively expressed in brain endothelial cells in vitro and supports the available data on in situ immunohistochemical staining of P-glycoprotein at the blood-brain barrier. In addition, these findings may indicate that one function of P-glycoprotein in vivo at the blood-brain barrier is the exclusion of xenobiotics from central nervous system tissues.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1992

Embryonic development of the CNS microvasculature in the mouse: new insights into the structural mechanisms of early angiogenesis.

Vascularization and expression of blood-brain barrier (bbb) associated morphological characteristics were studied by ultrastructural analysis during the embryonic development of the mouse CNS. At day 9 (E9) capillaries were only found in the perineural mesenchymal tissue exhibiting fenestrations and loosely attached pericytes. At E10 endothelial cells (EC) together with pericytes and other mesenchymal cells invaded the intraneural domain. The immigrating EC showed numerous protrusions and lost their fenestrations as soon as the new intraneural capillaries were formed. Intraneural EC showed prominent nuclei, many pinocytotic vesicles and junctional complexes which appeared tight in some places. These results indicate that from the first day of intraneural vascularization onwards, the morphological properties of the bbb are present in the early embryonic mouse cerebral cortex.

Animals↗

PubMed1992

Two cloned cerebral endothelial cell phenotypes: an in vitro model for angiogenesis?

Expression of smooth muscle alpha-actin and migratory behaviour of cloned cerebral endothelial cells (cEC) which exhibited two distinct phenotypes (type I, type II) were studied. Removal of mitogenic factors (alpha ECGF, ECGS) and heparin from the culture medium resulted in a smooth muscle-like appearance (type II) of the cells, expression of smooth muscle alpha-actin protein and smooth muscle actin mRNA and in an increased migratory activity. In contrast, addition of growth factors and heparin led to a cobblestone-like phenotype (type I) which lacked the expression of smooth muscle alpha-actin but expressed other proteins as determined by 2-D-gel electrophoresis.

Actins↗

PubMed1991

Reversible expression of sm alpha-actin protein and sm alpha-actin mRNA in cloned cerebral endothelial cells.

The expression of smooth muscle (sm) alpha-actin was studied in cloned capillary cerebral endothelial cells of two phenotypes. Type I cells were cultured in medium containing 10% FCS, heparin and ECGS (or alpha-ECGF) and stained positive for a specific endothelial cell marker (Bandeiraea simplicifolia). Depletion of heparin and ECGS resulted in a smooth muscle-like appearance after 2-3 days. Cells of this phenotype, (type II) stained positive for the endothelial cell marker and for sm alpha-actin. In contrast to type I cells, type II cells expressed sm alpha-actin protein and mRNA as evidenced by Immunoblots and Northern blots. This phenotypic switch was shown to be reversible and so was the expression of sm alpha-actin.

Actins↗

PubMed1991

Glial cells and neurons induce blood-brain barrier related enzymes in cultured cerebral endothelial cells.

The blood-brain barrier (BBB) in mammals is created and maintained by cerebral endothelial cells (cEC) that express specialized functional properties, including intercellular tight junctions, absence of fenestrae and specific membrane transport systems. It has been proposed that the differentiation of these characteristics, acquired during brain development, is controlled by the neural environment. Co-culture experiments of cloned cEC with astroglial cells, C6 glioma cells and cortical neurons, with plasma membranes or conditioned media of these cells, were used to study induction of some BBB characteristics in vitro. Activities of Na+,K(+)-ATPase and gamma-glutamyl transpeptidase (GGTP), an enzyme responsible for amino acid transport across the BBB, were taken as parameters for BBB function. Co-culture of cEC with C6 glioma cells caused a two-fold increase in GGTP activity and this activity was likewise amplified by incubation with plasma membrane fractions derived from C6 glioma cells, embryonic brain cells and cortical neurons; conditioned media (soluble factors) had no effect. Na+,K(+)-ATPase activity, estimated from the ouabain inhibitable fraction of 86Rb uptake, was increased by about 90% in cEC incubated with C6 glioma plasma membranes. We propose from these data that both neurons and glial cells confer BBB characteristics on cEC via cell-cell contact.

Animals↗

PubMed1991

DNA content prognostic in soft tissue sarcoma. 102 patients followed for 1-10 years.

In a prospective study of 102 patients with soft tissue sarcoma, the prognostic importance of DNA content and clinicopathologic features was analyzed. Based on DNA flow cytometry, 37 lesions were diploid (normal DNA content) and 65 were nondiploid (abnormal DNA content). The 5-year metastasis-free survival rate of the whole series was 0.59. The survival rate was 0.77 for patients with diploid tumors and 0.48 for those with nondiploid tumors (P = 0.01). Multivariate analysis identified two independent metastatic risk factors: increasing tumor size and nondiploidy. Unexpectedly, high malignancy grade (III-IV) was not found to be an independent risk factor for metastasis. The risk of metastasis was strongly related to the number of risk factors present. Thus, the 5-year survival for the 76 patients with no or one risk factor was 0.69, as compared with 0.30 for the 26 patients with two risk factors (P less than 0.0001). Our study shows that metastatic disease in soft tissue sarcoma is closely related to nondiploidy. A prognostication model based on DNA content and tumor size was found to discriminate between patients with a good and a poor prognosis after surgical treatment. The model can be used to identify patients who should be excluded from trials with adjuvant chemotherapy.

Adult↗

PubMed1990

Effects of progesterone, epipregnanolone and RU 38486 on potassium uptake in cultured cortical neurons.

It was previously reported that progesterone and its metabolites influence electrical properties of the CNS in many different ways. In the present study we elicited the effects of progesterone, its 5 beta reduced metabolite epipregnanolone and the anti-progestin compound RU 38486 on potassium uptake in cultured cortical neurons. K+ was substituted by the tracer substance 86Rb. When hormone treatment (10(-9)-10(-7) M/l) was performed for 3 days, addition of progesterone and epipregnanolone led to a significant decrease of 86Rb uptake whereas treatment with RU 38486 markedly increased 86Rb uptake. The effect of the anti-progestin could be reversed by the addition of increasing amounts of progesterone. Hormone actions were dose-dependent and most distinct when performed from the very first day of culture. Short-term (15 min) hormone treatment of neurons did not significantly alter 86Rb uptake. These findings suggest a specific receptor mediated progestin action which, in a long-term course, controls potassium uptake across excitable membranes.

Animals↗

PubMed1990

Gamma-glutamyl-transpeptidase (GGTP) and NA+K(+)-ATPase activities in different subpopulations of cloned cerebral endothelial cells: responses to glial stimulation.

Glial stimulation of Na+K(+)-ATPase and gamma-glutamyl-transpeptidase was taken as parameter for blood brain barrier function in cloned cerebral endothelial cells of different phenotypes. In type I cells ("cobblestone" phenotypus) gamma-glutamyl-transpeptidase activity increased 10-12 fold and Na+K(+)-ATPase activity was 2-fold increased after glial stimulation. In type II cells ("spindle-form" phenotype) gamma-glutamyl-transpeptidase was only 2-fold increased, whereas Na+K(+)-ATPase was even depressed. K(+)-(86Rb) uptake was twice as high in type I cells. These data indicate that type I cells are involved in blood brain barrier function.

Animals↗

PubMed1990

Influence of angiotensin converting enzyme inhibitor (captopril) on kidney epithelial cells in vitro: studies on potassium (86Rb) influx and cellular proliferation.

The effects of captopril on potassium influx and cellular proliferation in a dog kidney epithelial cell line (Madin-Darby canine kidney cells, MDCK) were studied. Na+K(+)-ATPase activity and the loop diuretic sensitive Na/K/2Cl- cotransport were measured using 86Rb as tracer substance. Cells were incubated with various concentrations of captopril (1-10 mmol/l). The furosemide sensitive Na/K/2Cl- cotransport was significantly decreased from 1 mmol/l onwards. Na+/K(+)-ATPase activity was lowered only when high amounts (10 mmol/l) of the drug were used. Cell proliferation was measured via [3H]thymidine incorporation. After incubation with 1 mmol/l captopril proliferation was strongly decreased (greater than 50%). Higher amounts (5-10 mmol/l) did not further suppress cell proliferation. The data suggest that natriuresis following ACE inhibition in vivo does not involve a direct effect of captopril on Na+K(+)-ATPase. However, the effect on cell proliferation may be of clinical relevance in respect to a possible mitogenic effect of angiotensin II.

Animals↗

PubMed1990

Comparison of the effects of adriamycin and methotrexate on orthotopic and induced heterotopic bone in rats.

The effect of the two antineoplastic drugs, Adriamycin and methotrexate, on orthotopic bone, and on the induction of experimental heterotopic bone in rats was analyzed. The drugs were administered as single injections: Adriamycin in s.c. doses of 0.5 and 2 mg/kg body weight and methotrexate i.v. 100 and 250 mg/kg body weight followed by leucovorin rescue after 2 h. A passing, but significant, decrease in body weights occurred in the methotrexate-treated animals, but not in those given Adriamycin. Analysis of the amount of heterotopic bone formed 4 weeks after induction by demineralized bone matrix revealed a 30-40% decrease in the groups treated with either of the antineoplastic agents, whereas orthotopic bone was unaffected. Six weeks after the treatment the net effect on the induced bone had decreased. The present study shows that the two antineoplastic drugs Adriamycin and methotrexate inhibit heterotopic new bone formation induced by demineralized bone matrix in rats to an equal extent, although their mode of action on the cellular level is entirely different, and that the inhibitory effect of a single treatment diminishes in the presence of a continuous inductive process.

Animals↗

PubMed1990

Bone resorption in orthotopic and heterotopic bone of dichloromethylene bisphosphonate-treated rats.

The effect of the bisphosphonate dichloromethylene bisphosphonate (Cl2MBP) on orthotopic and heterotopic bone, induced by implants of demineralized bone matrix (DBM) in rats, was analyzed, with special reference to bone resorption. The heterotopic bone was formed by induction for 3 weeks; at this time, the rats were given daily subcutaneous injections of 3 mg/kg of body weight of Cl2MBP or saline, until sacrifice. Prior to the start of treatment, the animals were given 45Ca and [3H]proline to label the inorganic and organic components of bone, respectively. Groups of rats were sacrificed at intervals from 1 to 31 days after isotope injection, and the net formation of bone and the elimination rates of the two isotopes were studied in the heterotopic bone, in diaphyseal and metaphyseal bone, and in teeth. The treatment with Cl2MBP caused a doubling of the daily net increase in mineral of the induced heterotopic bone, and a less pronounced increase in the ash content of tibiae. The treatment decreased the elimination rates of both isotopes in the orthotopic and heterotopic bone, showing that decreased bone resorption is the cause of the increased net bone formation.

Animals↗

PubMed1990

Glial-conditioned medium and attachment to ConA are essential for long-term culture of cortical neurons.

Cortical brain cells from 14-day-old mouse embryos were seeded on various substrates and cultivated in serum-free medium with or without conditioned medium from astrocytes or C6 glioma cells. Poly-L-lysine was shown to be the best substrate for cell attachment followed by Concanavalin A (ConA) and adhesion particles derived from glia cells. Cells grown on ConA sprouted rapidly and formed large networks. Survival of neurons was greatly prolonged when glia-conditioned medium (GCM) was present in the culture medium. Cells grown on ConA were then viable for more than 4 weeks. Without GCM, neurons survived in culture for about 2 weeks, regardless of the substrate. Endothelial cell growth supplement or acidic fibroblast growth factor increased survival of neurons but also stimulated proliferation of astrocytes.

Animals↗

PubMed1989

Cytogenetic and quantitative DNA analysis of primary and xenografted human osteosarcomas.

We have analyzed, cytogenetically and by flow DNA cytometry, three human osteosarcomas. From one case, both the primary tumor (T9) and three of its serial passages in nude mice were analyzed. From the two other tumors (T1, T4), nude mouse xenografts were examined. Complex karyotypic rearrangements were invariably found in the short-term culture preparations. The major clones were hypodiploid in T1 but near-triploid in all other tumor samples. No rearrangement common to all tumors could be identified. The DNA indexes were 1.0/2.0 (T1), 1.4 (T4), 0.9/1.8 (T9 primary), and 1.7 (all T9 xenografts). Thus, there was good correlation between the DNA indexes and the chromosome numbers. Chromosomal evolution could be studied in one case, in which both the primary tumor (T9) and its xenograft passages (1, 4, and 7), obtained 1, 7, and 11 months after the first transplantation of T9 cells to nude mice, were analyzed. All but one of the clonal marker chromosomes found in the primary tumor were retained in all passages. On the other hand, several new clonal markers developed.

Animals↗

PubMed1989

Tears of the medial meniscus associated with increased radionuclide activity of the proximal tibia. Report of three cases.

Three patients who presented with longstanding medial knee pain and normal radiographs had an isotope bone scan in an attempt to arrive at a diagnosis. In each case there was a well-defined area of increased uptake over the medial tibial condyle and arthroscopy subsequently revealed meniscal damage. After arthroscopic menisectomy each patient became asymptomatic and the bone scan returned to normal. We suggest that patients with incapacitating knee pain, normal radiographs and a positive bone scan undergo an arthroscopy even in the absence of clinical signs of meniscal damage.

Adult↗

PubMed1989

Micromethod for the determination of 3-beta-HSD activity in cultured cells.

A modified radioassay for the determination of the 3 beta-hydroxy-delta 5-steroid dehydrogenase (3-beta-HSD) is described. The assay is based on the conversion of [3H]pregnenolone to [3H]progesterone followed by a digitonin precipitation step. The method was applied to neurons, glial cells, C6 glioma cells and adrenal tumor cells in culture. Adrenal tumor cells and C6 glioma cells showed higher enzyme activity than primary cultures of astrocytes and neurons. Dependence of enzyme activity on pH, protein concentration and reaction time was demonstrated for C6 cells. A pH optimum was shown between 7.5 and 8.1, and the reaction was linear up to 2 h. beta-oestradiol inhibited 3-beta-HSD activity completely. The assay presented is fast, highly reproducible, and offers the possibility of studying 3-beta-HSD activity in differentiating cells in culture without preparation of microsomes or extraction of reaction products.

3-Hydroxysteroid Dehydrogenases↗

PubMed1989

Isolation, characterization, and long-term cultivation of porcine and murine cerebral capillary endothelial cells.

We present a simple method for isolation and long-term cultivation of porcine and murine cerebral capillary endothelial cells (cEC). Two major points are made. First, that the "characteristic" morphology of the endothelial cells depends mainly on the presence of endothelial cell growth factors in the culture medium and second, that the identification of the cells as endothelial cells requires a special lectin instead of criteria used for large vessel endothelial cells, such as factor VIII staining or LDL uptake. Pure cerebral capillaries were isolated by means of a series of centrifugation steps; endothelial cells were released by collagenase treatment and cultivated on plastic petri dishes, which proved to be better for cell attachment than collagen or gelatin coating. The microvascular cells were cultivated in either the presence or absence of growth factors. Medium 199 + 10% FCS produced mainly spindle-shaped cells, growing in the "hills and valleys" pattern, which, if not passaged for weeks, showed three dimensional tubular structures. Cells of the "cobblestone" phenotype were promoted in medium 199 + 10% FCS, enriched with endothelial cell growth supplement (ECGS) and heparin (referred to as complete medium). These cells retained their phenotype for months and could be passaged up to 35 times till now. If ECGS and heparin were omitted from these cultures, the cells became elongated and resembled smooth muscle cells. This effect was reversible when the cells were transferred to complete medium. With cEC, cloned by limiting dilution, we noticed this reversal phenomenon as well. We used several markers to characterize the microvascular cells and could show that the lectin of Bandeiraea simplicifolia is a highly reliable marker for endothelial cells and that the monoclonal antibody alpha-sm-1 (anti-smooth muscle cell actin) is excellent for determining smooth muscle cells.

Animals↗

PubMed1989

Effect of human interferon-alpha and interferon-gamma on growth, histology, and DNA content of human osteosarcomas in nude mice.

The antitumor effect of human natural and recombinant interferon-gamma (IFN-gamma) was evaluated in human osteosarcomas grown as xenografts in nude mice. IFN-gamma was given as daily subcutaneous injections, alone or in combination with IFN-alpha. The growth of two out of three tested osteosarcomas was inhibited by 2 x 10(5) IU of natural IFN-gamma. A five times higher dose of recombinant IFN-gamma, as compared with natural (n) IFN-gamma, was needed to obtain growth inhibition of one osteosarcoma. This difference in dose-response could be explained by differences in pharmacokinetics. Hence, subcutaneously administered natural IFN-gamma gave 10 times higher serum levels than obtained with the recombinant type. Combination treatment with IFN-alpha and IFN-gamma induced a potentiation of the antitumor effect in one osteosarcoma. In another osteosarcoma, 2-4 x 10(5) IU of nIFN-gamma did not effect tumor growth and could not potentiate the antitumor effect of 2-4 x 10(5) IU of nIFN-alpha. By using DNA analysis in cell suspension and tissue section, the proportion of aneuploid tumor cells within the xenograft could be estimated. This analysis showed that the antitumor effects of IFN were more pronounced than mere measurement of tumor volume suggested. IFN-inhibited tumors were partly replaced by fibroblasts or bone tissue. In conclusion, at the doses given nIFN-gamma appeared to have similar antitumor effects as IFN-alpha in two osteosarcomas, whereas one was sensitive to only IFN-alpha. Combination IFN treatment induced a potentiation of the antitumor effect in one osteosarcoma but not in another. The differences between the osteosarcomas in obtained antitumor effect of IFN treatment probably reflects individual IFN sensitivity and demonstrates the importance of assessing several tumors of the same neoplastic entity.

Animals↗