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Biomedical subjects

K Keller

Publications and source records attributed to K Keller.

At least 91 records · Page 5Linked to original sources
PubMed1990

Effects of isolated rheumatoid synovial cells on cartilage degradation in vitro.

Rheumatoid synovium in coculture with cartilage has been shown to release a factor(s) that stimulates the depletion of glycosaminoglycans (GAG) from cartilage matrix. Human rheumatoid synovium was enzymatically disaggregated and the isolated cells were subjected to a variety of mechanical and immunological treatments. Synovial cell conditioned media (SCCM) were prepared and analyzed for their ability to stimulate GAG depletion. SCCM prepared from increasing concentrations of isolated synovial cells demonstrated cartilage degradative activity in a dose-dependent manner. This activity was characterized as interleukin-1 like and was found mostly within the adherent cell population where the synovial macrophages retained significant degradative ability. T cells alone were found to have no direct degradative effect on cartilage, but their presence appeared to augment the response of the adherent cells. The techniques described here provide a quantitative model for examining the degradative factors from synovium as well as the cellular interactions that promote their release.

Animals↗
PubMed1990

Binding of proteolytically-degraded human colonic mucin glycoproteins to the Gal/GalNAc adherence lectin of Entamoeba histolytica.

Rat and human colonic mucin glycoproteins bind to the Gal/GalNAc adherence lectin on the surface of Entamoeba histolytica in vitro, thus inhibiting the organism from adhering to and lysing the target cells. Human colonic mucin glycoproteins were isolated by Sepharose 4B gel filtration chromatography, they were proteolytically degraded with trypsin, pronase, and papain, and the glycoprotein fractions were reisolated by Sephacryl S-200 gel filtration chromatography. Binding of the mucin glycoprotein fractions to amoebae was quantitated by the inhibition of adherence of Chinese hamster ovary cells to the surface of the amoebae. Trypsin and papain digests caused 40 and 20% reductions, respectively, in the excluded fractions (void volume) that contained all the carbohydrates; pronase digests resulted in extensive degradation of the mucin glycoprotein with the carbohydrate fractions eluting over 40% of the gel bed volume. 3H-labelled mucin glycoprotein and sodium dodecylsulfate-polyacrylamide gel electrophoresis confirmed the presence of the high molecular weight carbohydrate-rich glycoproteins with no subunits in the excluded fractions and the absence of sugars in the included peptides. Only the high molecular weight carbohydrate-containing fractions bind amoebae and inhibit amoebic adherence to Chinese hamster ovary cells. The trypsin digested mucins in the excluded volume were more efficient than the native undigested mucins in binding amoebae. The carbohydrate-containing fractions of the pronase digests were the least effective in binding amoebae and inhibiting adherence of Chinese hamster ovary cells. This suggests that proteolytically-degraded colonic mucins that are glycosylated, as well as the undegraded native mucin glycoproteins of the gut, may play a protective role in binding to amoebae, thus preventing contact of amoebae with mucosal epithelial cells and potential invasion.

Acetylgalactosamine↗
PubMed1990

Antibody to interleukin 1 inhibits the cartilage degradative and thymocyte proliferative actions of rheumatoid synovial culture medium.

Cartilage breakdown in rheumatoid arthritis results from (a) lytic action by synovial enzymes, and (b) release of synovial catabolin, now believed to be a form of interleukin 1 (IL-1), causing chondrocytes to degrade their matrix. Rheumatoid synovial culture media were tested for their ability to stimulate cartilage degradation (proteoglycan release from bovine nasal cartilage discs) and thymocyte proliferation (3H-thymidine incorporation) in the absence or presence of anti-IL-1. Degradation of living cartilage, stimulated 2-fold by synovial culture media, was inhibited up to 80% by anti-IL-1. Residual breakdown in living cartilage and synovial culture media induced breakdown in dead cultures were of similar magnitude, and both were unaffected by antibody treatment. Proteoglycan products released from synovial culture media treated cartilage were of smaller average molecular weight (Sepharose CL-2B), and such size reduction was inhibited by anti-IL-1 treatment. Synovial culture media that stimulated cartilage degradation also stimulated thymocyte proliferation; the latter was fully suppressible by anti-IL-1. One of 8 synovial culture media contained an inhibitor(s) of thymocyte proliferation, removable by dialysis. We conclude (1) rheumatoid synovial catabolin activity is due to a form of IL-1. (2) A minor nonsuppressible component of synovial culture media stimulated breakdown, identical in living and killed cartilage, is due to passive transfer of enzymic activity. (3) Cultured rheumatoid synovium releases both IL-1 and an inhibitor(s) of IL-1 action.

Antibodies↗
PubMed1990

[Quantitative autoradiography of the glucose transport protein with (3H)cytochalasin-B in the mammalian eye].

In vitro autoradiography with [3H]cytochalasin for the anatomic localization of the glucose transport protein in eyes of rats, rabbits, baboons, marmosets and humans reveals high glucose carrier densities in the ciliary body, especially ciliary processes, iris, retina and, in some species, in the trabecular meshwork and lens. In the lens, specific [3H]cytochalasin B binding sites are mainly concentrated in the lens nucleus. Lower concentrations are found in the cortex. During again, the glucose transporter concentration increases up to the age of 8 years in the marmoset lens nucleus, but decreases in the cortex and retina. Moderate amounts of carrier are located in the corneal endo- and epithelium. These findings suggest a possible involvement of glucose transporter-related mechanisms in cataractogenesis, e.g. in that resulting from glycosylation of lens proteins. The enrichment of glucose carrier protein in the trabecular meshwork suggests a high metabolic activity and a possible involvement in the regulation of intraocular pressure. The relationship between glucose transport and utilization is also briefly discussed.

Animals↗
PubMed1990

Different mammalian facilitative glucose transporters expressed in Xenopus oocytes.

Xenopus oocytes exhibit an extremely low basal glucose transport and are thus ideally suited for the expression of heterologous glucose transporters. They have, therefore, proven to be a very valuable expression system to functionally express the erythrocyte/brain (GLUT1), the liver (GLUT2), and the adipocyte/muscle (GLUT4) glucose transporters. Characterization of their functional properties indicates that they fulfill the criteria of a glucose transport protein by demonstrating saturation kinetics, stereospecificity, and inhibition by cytochalasin B. Although mammalian facilitative glucose transporters have been expressed in bacteria and eukaryotic cell lines, the present data emphasize the advantage of using Xenopus oocytes for comparative kinetic analysis of the various members of the facilitated diffusion glucose transporter gene family.

Animals↗
PubMed1990

Biochemical and histological evaluation of the synovial-like tissue around failed (loose) total joint replacement prostheses in human subjects and a canine model.

The tissue around loose total joint replacement prostheses displays a synovial-like lining comprised of cells that produce IL-1 and PGE2, mediators of inflammation that stimulate bone resorption. Particles of titanium alloy, as well as cobalt-chromium alloy and polyethylene, were found to aggravate the histiocytic response and production of IL-1 and PGE2. Tissue with similar histological and biochemical features was produced in a canine model of the aseptic loose cemented femoral stem.

Animals↗
PubMed1989

Functional expression of the human HepG2 and rat adipocyte glucose transporters in Xenopus oocytes. Comparison of kinetic parameters.

Facilitated glucose transport is a ubiquitous characteristic of animal cells carried out by a family of membrane glycoproteins. Two members of this gene family are the well characterized human erythrocyte protein that has been cloned from the HepG2 cell line and the insulin-sensitive transporter that has been cloned from adipocytes and muscle tissue. In the present study the HepG2 and adipocyte glucose transporters were functionally expressed in Xenopus oocytes after injection of synthetic mRNAs produced by transcription in vitro from cloned cDNAs. Both 2-deoxyglucose uptake and 3-O-methylglucose transport were increased several-fold over basal levels in mRNA-injected oocytes. Increased uptake of 2-deoxyglucose was inhibited completely in the presence of cytochalasin B, and 3-O-methylglucose transport was blocked by D-glucose but not by L-glucose. The half-saturation constant and turnover number for 3-O-methylglucose transport at 22 degrees C via the HepG2 transporter were estimated to be 21 mM and 2.2 x 10(3) s-1 under equilibrium exchange conditions. The half-saturation constant for 3-O-methylglucose transport via the adipocyte transporter under the same conditions was estimated to be 1.8 mM. These data prove the functional identity of the cloned HepG2 and adipocyte cDNAs and indicate that the HepG2 and adipocyte transporters display similar kinetic behavior when expressed in the frog oocyte membrane as compared with their native membrane environments. Thus, the difference in the equilibrium exchange kinetic parameters for glucose transport in the erythrocyte and the adipocyte is a result of the expression of two distinct glucose transporter proteins.

Adipose Tissue↗
PubMed1989

Endogenous regulation of 2-deoxyglucose uptake in C6 glioma cells correlates with cytoskeleton-mediated changes of surface morphology.

The cellular basis of the membrane-limited state of glucose utilization and the mechanism of the endogenous regulation of hexose uptake in dense monolayers of C6 glioma cells were investigated. In an earlier study, it was shown that at high rates of glucose transport and phosphorylation combined with the inhibition of glycolytic adenosine triphosphate (ATP) production by iodoacetate, an endogenous regulatory response occurred that resulted in rapid, periodic variations of the glucose uptake rates (Lange et al., 1982). Similar time-dependent periodic changes of uptake rates also occurred during incubation of C6 glioma cells with 2 mM 2-deoxyglucose (2-DG) without pretreatment of the cells with iodoacetate. These changes were accompanied by variations of the intracellular ATP content, by distinct alterations of the shape and arrangement of microvilli and lamellae (lamellipodia) on the cell surface, and by changes of the cytoskeletal F-actin content. Because the changes of 2-DG uptake rates occurred independent of the intracellular 2-DG concentration, the bulk of this 2-DG pool was assumed to be localized apart from the membranal transport sites. Downregulation of 2-DG uptake appeared to be triggered by a rapid decrease of a small pool of the cellular ATP involved in the phosphorylation of transported hexose. Scanning and transmission electron microscopic observations of cells fixed in different states of the endogenous uptake regulation supported the assumption that the interior of lamellae and microvilli may represent a small entrance compartment for transported hexoses in which occurred the observed close coupling between hexose transport and phosphorylation as well as the rapid variations of ATP content. Hexose uptake is supposed to be regulated by cytoskeleton-mediated changes of volume and diffusional accessibility of this compartment, modulating the degree of its metabolic coupling with the cytoplasmic main compartment.

Actins↗
PubMed1989

A prospective randomized trial of low molecular weight heparin-DHE and conventional heparin-DHE (with acenocoumarol) in patients undergoing gynaecological surgery.

The antithromboembolic efficacy of once a day low molecular weight heparin in fixed combination with dihydroergotamine (LMWH-DHE) was compared with conventional heparin-DHE in combination with Acenocoumarol (heparin-DHE/A) in 191 patients undergoing gynaecological surgery. LMWH-DHE proved equally effective in preventing thromboembolic complications, with a similar incidence of postoperative bleeding and side effects. Deep vein thrombosis occurred once in each group and one non-fatal pulmonary embolism occurred in the LMWH-DHE group. The main advantage of LMWH-DHE was significantly better patient acceptance of the single daily subcutaneous injection as compared with the two injections of conventional heparin-DHE (P = 0.02). On the other hand, LMWH-DHE was associated with significantly increased incidence of intraoperative bleeding (P less than 0.02). The bleeding did not, however, cause any clinical problems. Discontinuation of therapy due to bleeding or pain at the site of injection occurred three times in each group. We consider the use of LMWH-DHE to be an attractive, economic and safe method of thromboembolic prophylaxis.

Acenocoumarol↗
PubMed1989

Cloning and functional expression of a human pancreatic islet glucose-transporter cDNA.

Previous studies have suggested that pancreatic islet glucose transport is mediated by a high-Km, low-affinity facilitated transporter similar to that expressed in liver. To determine the relationship between islet and liver glucose transporters, liver-type glucose-transporter cDNA clones were isolated from a human liver cDNA library. The liver-type glucose-transporter cDNA clone hybridized to mRNA transcripts of the same size in human liver and pancreatic islet RNA. A cDNA library was prepared from purified human pancreatic islet tissue and screened with human liver-type glucose-transporter cDNA. We isolated two overlapping cDNA clones encompassing 2600 base pairs, which encode a pancreatic islet protein identical in sequence to that of the putative liver-type glucose-transporter protein. Xenopus oocytes injected with synthetic mRNA transcribed from a full-length cDNA construct exhibited increased uptake of 2-deoxyglucose, confirming the functional identity of the clone. These cDNA clones can now be used to study regulation of expression of the gene and to assess the role of inherited defects in this gene as a candidate for inherited susceptibility to non-insulin-dependent diabetes mellitus.

Animals↗
PubMed1988

Association of glycolytic enzymes with the cytoplasmic side of the plasma membrane of glioma cells.

A latex phagocytosis technique was used to prepare relatively pure plasma membranes with inside-out orientation. This method was adapted through a number of modifications in order to evaluate the association of glycolytic enzymes with the cytoplasmic side of the plasma membrane of C6 glial cells. As phosphorylation is strictly coupled with transport in these cells, glycolytic enzymes, especially hexokinase, could metabolize glucose in close vicinity to its transporter. Of the enzymes tested, hexokinase is present in considerable quantities on these membranes (nearly 40% of homogenate specific activity), followed by D-glyceraldehyde-3-phosphate dehydrogenase (10%), pyruvate kinase (8%), and 3-phosphoglycerate kinase (1%). Except for hexokinase, the enzyme pattern presented here is different from that published for other membrane preparations.

Cell Membrane↗
PubMed1986

Viral and bacterial organisms associated with acute pharyngitis in a school-aged population.

To investigate the causes and clinical characteristics of acute pharyngitis among school-aged children (4 to 18 years), we obtained throat cultures for respiratory viruses, Mycoplasma pneumoniae, group A streptococcus, and Chlamydia trachomatis from 320 patients with sore throat and 308 controls without respiratory complaints. The study was conducted from January to April 1985 in a private pediatric practice in central New York State. Sixty percent of the patients and 26% of the control subjects had positive cultures for at least one organism. Forty percent of patients had positive cultures for group A streptococcus, compared with 11.9% of the controls. Fifty (16%) patients had positive viral cultures, compared with eight (2.6%) controls; the predominant viral isolate was influenza A Philippines. Patients infected with influenza A were significantly more likely to complain of cough and hoarseness, and were less likely to have pharyngeal exudate or tender cervical adenopathy, than were patients who had positive cultures for group A streptococcus. Although 49 (15.8%) patients with acute pharyngitis had cultures positive for M. pneumoniae, 53 (17.6%) asymptomatic controls were also had M. pneumoniae-positive cultures. Thus detection of M. pneumoniae in the throat of school-aged children with pharyngitis may not be sufficient to establish a diagnosis of disease caused by this organism. C. trachomatis was not isolated from any patient or control.

Acute Disease↗
PubMed1986

[Obstructive endometriosis of the efferent urinary tract].

Even though, in every 7th to 8th patient with endometriosis, it occurs in the efferent urinary tract, little attention has been paid this fact, generally speaking. This may be due in part to the atypical symptomatology of ureteral or respectively vesical endometriosis. However, its consequences can be serious in both younger patients as well as postmenopausal women. On the basis of two of the authors' own cases and the literature, the clinical symptoms and treatment of vesical endometriosis and ureteral obstruction associated with pelvic endometriosis are discussed.

Adult↗
PubMed1986

Glucose transporter in plasma membranes of cultured neural cells, as characterized by cytochalasin B binding.

Identification of hexose transporter sites by cytochalasin B binding was conducted with a centrifugation assay. The determination of KD and Bmax values by LIGAND computer analysis provided binding data that are similar in primary astrocytes (238 nM and 14 pmol/mg protein) and neuroblastoma cells (179 nM and 13.6 pmol/mg protein). In contrast, only an insignificant number of transporter sites was detectable in C6 glioma cells, irrespective of whether membrane fractions were obtained by a two-phase polymer system or by a latex phagocytosis technique yielding inside-out plasma membranes. The latter membrane preparation was utilized to identify and quantitate the transporter molecules at the inner membrane surface of primary astrocytes, i.e., 160 nM (KD) and 5.8 pmol/mg protein (Bmax), respectively.

Animals↗