Search PubMed⌕ Search

Biomedical subjects

R H Kelly

Publications and source records attributed to R H Kelly.

At least 37 records · Page 2Linked to original sources
PubMed1993

Complement C3 fragments in urine: detection in systemic lupus erythematosus patients by western blotting.

We characterized urinary excretion of C3 fragments among patients with systemic lupus erythematosus (SLE) as a possible indicator of renal involvement. 28 patients, representing a broad range of disease activity were admitted to our study. Urinary proteins were separated on 4-20% gradient SDS-PAGE gels, under reducing conditions, and transblotted to nitrocellulose. Western blots were developed with a polyvalent goat-anti-human C3d antiserum, and an alkaline phosphatase-conjugated rabbit anti-goat IgG. Three patterns were obtained: 1) no bands detected; 2) bands suggesting the presence of intact C3; and 3) samples with additional low molecular (< 4 x 10(4)) bands. The 12 patients with no C3 bands had minimal disease activity (e.g. fatigue, arthralgia, arthritis, rash, oral ulcers). The seven patients with intact C3 patterns also had minimally active disease. Their primary clinical findings included fatigue, pleurisy, renal disease which had been treated, hemolytic anemia, and arthritis. Patients with low molecular weight C3 fragments in their urine formed two sub-sets, based upon their presenting features. The first group had severe disease and contained all patients with active lupus nephritis (n = 4), while the second consisted of non-renal patients with primary clinical findings of moderate disease activity (e.g. thrombocytopenia, pneumonitis, arthritis). Our results suggest urinary excretion of low molecular weight C3 fragments correlates with active renal disease, but is a variable finding among SLE patients with non-renal manifestations of disease activity.

Blotting, Western↗
PubMed1993

Beta-2 transferrin assay in clinical management of cerebral spinal fluid and perilymphatic fluid leaks.

Beta-2 transferrin is a protein found in cerebral spinal fluid (CSF) and inner ear perilymph, but not in blood, nasal or ear secretions. The purpose of this paper is to evaluate the clinical usefulness of the current assay for beta-2 transferrin for detecting CSF and perilymphatic leaks. We reviewed the hospital records of the first 88 patients having specimens submitted for beta-2 transferrin analysis at our institution. Both CSF and perilymph leaks were identified. However, confirmation of the absence or presence of beta-2 transferrin was directly used in the clinical management of only 55% of the patients. This was largely secondary to the time delay in test processing and initial lack of physician confidence with the test. However, our review of the clinical outcomes relating to the use of the beta-2 transferrin analysis suggests high sensitivity and specificity for the test. Analysis of beta-2 transferrin appears to be a valuable test for detecting CSF leakage and a promising test for confirming perilymphatic leaks. However, to achieve greater clinical usefulness a rapid clinical assay needs to be developed and further information gained regarding the sensitivity and specificity of the beta-2 transferrin assay for detecting CSF and perilymphatic fluid leakage.

Adult↗
PubMed1993

Transferrin allelic variants may cause false positives in the detection of cerebrospinal fluid fistulae.

Variations in either the polypeptide sequence or the carbohydrate moieties of transferrin may result in altered electrophoretic mobility of this molecule. We report a case of an allelic (polypeptide) variant of transferrin with mobility similar to that of the beta 2 (sialic acid-depleted) transferrin found in cerebrospinal fluid (CSF) and a few other body fluids. Allelic variation and other transferrin anomalies may be mistaken for the CSF isoform, resulting in false diagnoses of CSF fistulae.

Alleles↗
PubMed1992

Lack of increase in heterozygous alpha 1-antitrypsin deficiency phenotypes among patients with hepatocellular and bile duct carcinoma.

Homozygous alpha 1-antitrypsin deficiency (PiZZ phenotype) is known to be associated with increased risk of cirrhosis and primary liver cancer. Although a relationship between heterozygous alpha 1-antitrypsin deficiency and chronic liver disease was suggested recently, it is still a matter of controversy whether such patients are at increased risk of liver cancer. The goal of this study was to determine the prevalence of heterozygous alpha 1-antitrypsin deficiency of different phenotypes among patients with primary hepatobiliary cancers. We studied 82 patients with primary hepatobiliary cancer; 59 had hepatocellular carcinoma and 23 had bile duct carcinoma. alpha 1-Antitrypsin quantitation and phenotyping were performed in each patient using standard methods. The distribution of the various Pi phenotypes was compared with that found in a normal population and reported elsewhere. Odds-ratio and chi 2 tests were used to measure the relative risk and the significance of association, respectively, between primary hepatobiliary cancers and heterozygous alpha 1-antitrypsin deficiency. Four patients in each of the cancer groups were heterozygous. Among the hepatocellular carcinoma patients, three had the PiMS phenotype and one had the PiMZ phenotype. Of these four heterozygous patients, only two had cirrhosis; one had cryptogenic cirrhosis and the other had hepatitis B virus-related cirrhosis. One noncirrhotic patient with a PiMZ phenotype had a fibrolamellar carcinoma. Of the four patients with bile duct carcinoma, three had the PiMS phenotype and one had the PiMZ phenotype. Of the four heterozygous patients, two had primary sclerosing cholangitis without associated inflammatory bowel disease and one patient had had previous biliary operations.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenoma, Bile Duct↗
PubMed1992

Association between heterozygous alpha 1-antitrypsin deficiency and genetic hemochromatosis.

Primary hemochromatosis is a genetically determined autosomal recessive disorder characterized by the excessive accumulation of body iron, most of which is deposited in the parenchymal cells of various organs. alpha 1-Antitrypsin deficiency is characterized among others by defective secretion of alpha 1-antitrypsin from liver cells. Whereas the risk of cirrhosis is increased in homozygous patients (PI ZZ) and possible in heterozygous patients (non-PI MM) as well, a greater risk for hepatocellular carcinoma has been suggested only in homozygous patients. Because these two metabolic disorders are relatively common, it has been difficult to determine whether they are associated with each other. In this study, we tried to determine the relationship between these two disorders using the case material seen at the University of Pittsburgh during a 7-yr period. We studied 15 patients with genetic hemochromatosis. alpha 1-Antitrypsin quantitation and phenotyping were performed in each case using standard methods. The distribution of the various Pi phenotypes was compared with that found in a normal population and reported elsewhere. Odds ratio and chi 2 tests were used to measure the relative risk and significance of association, respectively. Eleven patients (73%) were found to be PI M and four (27%) were identified as being heterozygotes: three (20%) were PI MZ, and one (7%) was PI MS. The prevalence of the PI MS phenotype was similar to that in the general population (7% vs. 6.4%; NS). The PI MZ phenotype, however, was statistically more common in patients with hemochromatosis than in the general population (20% vs. 2.2%; p less than 0.004).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1992

Beta 2 transferrin application in otology.

The diagnosis and management of perilymphatic fistula has received considerable attention in recent years. Despite the use of sophisticated technology, the diagnosis of perilymphatic fistula continues to rest primarily upon clinical suspicion and the exclusion of other disorders. In addition, the confirmation of a perilymphatic fistula during surgical exploration is usually based upon the subjective observation of fluid pooling in niches of the middle ear. A sensitive and objective laboratory test for identifying perilymph in the middle ear would be a useful adjunct for the diagnosis and management of perilymphatic fistula. The objective of this paper is to demonstrate the potential utility of beta 2 (beta 2) transferrin assay in the diagnosis of perilymphatic fistula. To accomplish this objective, we confirmed that beta 2 transferrin is present in living human perilymph and is absent in the normal or inflamed middle ear. In addition, the utility of beta 2 transferrin assay in the diagnosis of cerebrospinal fluid otorrhea is presented.

Aged↗
PubMed1991

The immune system as a neural network: a multi-epitope approach.

The term "neural network" has been applied to arrays of simple activation units linked by weighted connections. If the connections are modified according to a defined learning algorithm, such networks can be trained to store and retrieve patterned information. Memories are distributed throughout the network, allowing the network to recall complete patterns from incomplete input (pattern completion). The major biological application of neural network theory to date has been in the neurosciences, but the immune system may represent an alternative organ system in which to search for neural network architecture. Previous applications of parallel distributed processing to idiotype network theory have focused upon the recognition of individual epitopes. We argue here that this approach may be too restrictive, underestimating the power of neural network architecture. We propose that the network stores and retrieves large, complex patterns consisting of multiple epitopes separated in time and space. Such a network would be capable of perceiving an entire bacterium, and of storing the time course of a viral infection. While recognition of solitary epitopes occurs at the cellular level in this model, recognition of structures larger than the width of an antibody binding site takes place at the organ level, via network architecture integration of, i.e. individual epitope responses. The Oudin-Cazenave enigma, the sharing of idiotypic determinants by antibodies directed against distinct regions of the same antigen, suggests that some network level of integration of the individual clonal responses to large antigens does occur. The role of cytokines in prior neural network models of the immune system is unclear. We speculate that cytokines may influence the temperature of the network, such that changes in the cytokine milieu serve to "anneal" the network, allowing it to achieve the optimum steady-state in the shortest period of time.

Artificial Intelligence↗
PubMed1991

Anti-estrogen antibodies in systemic lupus erythematosus: a quantitative evaluation of serum levels.

We measured the beta-estradiol binding capacity of serum gamma-globulins in four subject groups; 1) normal men, 2) normal women who had never taken oral contraceptives, 3) normal women who had a history of oral contraceptive use and, 4) patients with systemic lupus erythematosus (SLE). We used dextran-coated charcoal to strip endogenous estradiol from serum proteins, added 3H-estradiol, and measured its association with proteins in various electrophoretic fractions following zone separation on agarose gels. Most of the bound radioactivity was present in the albumin, beta and gamma-globulin fractions. Binding to gamma-globulins was elevated in SLE patients, and normal controls who had taken oral contraceptives, as opposed to other controls (p less than 0.005). Gamma-region radioactivity could be removed by protein-G adsorption prior to zone electrophoresis. Isoelectric focusing revealed a pattern of tritiated-E2 binding consistent with polyclonal B-cell activation in all groups. There was no correlation between the extent of gamma-region binding and the total serum immunoglobulin level for any of the groups studied, nor was there a correlation between E2 binding and anti-DNA titers in the SLE group. The average anti-estradiol antibody concentrations in SLE sera (assuming equimolar binding) was 105 ng/ml (95% CL = 92-118), whereas their average anti-DNA antibody concentration was in the microgram/ml range. Thus, quantitatively, the level of anti-estradiol antibodies is at least an order of magnitude lower than the anti-DNA antibodies characteristic of this disease.

Adolescent↗
PubMed1991

Response of low-passage human malignant gliomas in vitro to stimulation and selective inhibition of growth factor-mediated pathways.

The proliferation of many nonglial tumors in vitro depends on the presence of nanomolar concentrations of one or more growth factors. To define the growth factor requirements of malignant glial tumors, the authors examined the response properties of four low-passage human malignant glioma lines to the following mitogens: epidermal growth factor (EGF), acidic and basic fibroblast growth factors (FGF's), insulin-like growth factor I (IGF-I), nerve growth factor (NGF), platelet-derived growth factor (PDGF), 12-O-tetradecanoyl-13-phorbol acetate (TPA), and serum. Each of the tumors showed increased deoxyribonucleic acid (DNA) synthesis (assessed by acid-precipitable [3H]-thymidine incorporation) in response to PDGF with a maximum effect at 50 ng/ml. Three tumors responded to EGF, three to IGF-I, two to acidic FGF, two to basic FGF, and two to TPA with maximum effects at 10, 50, 1, 1, and 10 ng/ml, respectively. None of the tumors responded to NGF. In the responsive tumors, optimum concentrations of EGF, IGF, TPA, acidic FGF, and basic FGF induced, at most, a two- to fourfold increase in [3H]-thymidine incorporation, which was only 30% to 50% of the response seen in 10% serum. In contrast, PDGF increased DNA synthesis eight- to 10-fold, equaling the effect of 10% serum. Measurements of cell proliferation also demonstrated a significant response to PDGF in each of the tumors. Appropriate concentrations of an anti-PDGF neutralizing antibody inhibited baseline DNA synthesis and proliferation in the absence of added growth factors, suggesting the possible role of PDGF in autocrine stimulation of these cells. However, this antibody produced only slight inhibition of serum-induced mitogenesis. Trapidil, an agent reported to inhibit the effects of PDGF, and polymyxin B, an inhibitor of protein kinase C, strongly inhibited baseline as well as PDGF- and serum-induced mitogenesis. It is concluded that, in the malignant gliomas studied, PDGF may be acting as a dominant mitogen to enhance DNA synthesis, and may function in autocrine stimulation. However, other factors contained in serum can also contribute to cell division.

Antibodies↗
PubMed1990

Evidence for locally synthesized and clonally restricted immunoglobulin in the synovial fluid from rheumatoid arthritis patients.

In this study, we examined the immunoglobulin (Ig) present in synovial fluid (SF) from patients with rheumatoid arthritis (RA) to determine if it was locally produced and to assess the presence of clonally restricted (oligoclonal) immunoglobulin. We studied SF/serum pairs from 55 RA patients and 23 patients with degenerative joint disease (DJD). We found increases in total protein, IgG, IgA, and IgM in RA vs DJD SF (P less than 0.01). The immunoglobulin present in RA appeared to be locally produced as evidenced by significant increases (P less than 0.01) in the immunoglobulin indices. Regression analysis among the levels of IgG, IgA, and IgM RF and the Ig indices suggested that only a minority of the locally synthesized Ig present was specific for RF. To provide evidence of clonal restriction, we further analyzed the SF specimens by isoelectric focusing and assessed the presence of oligoclonal bands present only in RA SF. In 7/55 RA specimens (13%) we found unique SF IgG bands. All bands were of similar isoelectric point (pI), being quite cathodic with pI greater than 7.5. Our evidence supports synthesis of Ig within RA synovium, with a minority of patients showing prominent and unique SF Ig bands. This suggests an oligoclonal response in SF of some patients, but polyclonal Ig synthesis in most.

Arthritis, Rheumatoid↗
PubMed1990

Effect of tamoxifen on DNA synthesis and proliferation of human malignant glioma lines in vitro.

Previous studies in our laboratory have shown that proliferation of human malignant gliomas in vitro depends in part upon the activation of protein kinase C (PKC) and, conversely, can be blocked by inhibitors of PKC. Here, we examined the effect of tamoxifen, a known PKC inhibitor, on DNA synthesis and proliferation of an established human glioma line (U138) and two low passage cultures of explanted human glioblastomas. Tamoxifen produced a profound, dose-dependent inhibition of both [3H] thymidine incorporation and cell proliferation, with a 50% effective dose of 20 ng/ml under serum-free conditions and 50 to 200 ng/ml in the presence of 10% serum. These tumors were estrogen receptor negative and showed no mitogenic response to estradiol. Furthermore, concentrations of estradiol as high as 10 micrograms/ml had no effect on the tamoxifen-induced inhibition. This suggests that the mechanism of growth inhibition by tamoxifen in these gliomas did not involve an estrogen receptor-mediated process but may instead result from its inhibition of PKC. In view of the profound effect of tamoxifen on cultured gliomas at concentrations that can safely be achieved therapeutically, further in vitro and in vivo studies of this agent are warranted.

Cell Division↗
PubMed1990

Response of malignant glioma cell lines to activation and inhibition of protein kinase C-mediated pathways.

To evaluate the role of protein kinase C-mediated pathways in the proliferation of malignant gliomas, this study examined the effect of a protein kinase C (PKC)-activating phorbol ester (12-O-tetradecanoyl-13-phorbol acetate or TPA) and a protein kinase C inhibitor (polymyxin B) on deoxyribonucleic acid (DNA) synthesis of malignant glioma cells in vitro. A serum-free chemically defined medium, MCDB 105, was employed for all studies. Two established human malignant glioma cell lines (T98G and U138), two rat glioma lines (9L and C6), and two low-passage human glioma lines (obtained from surgical specimens) were studied. With the exception of the C6 line, all tumors responded in a dose-dependent fashion to nanomolar concentrations of TPA with a median effective dose that varied from 0.5 ng/ml for the U138 glioma to 1 ng/ml for the T98G glioma. At optimal concentrations (5 to 10 ng/ml), TPA produced a two- to five-fold increase in the rate of DNA synthesis (p less than 0.05) as assessed by incorporation of 3H-thymidine. However, TPA had no additive effect on the mitogenic response produced by epidermal growth factor (EGF) or platelet-derived growth factor (PDGF). Inhibition of PKC using the antibiotic polymyxin B (20 micrograms/ml) abolished the TPA-induced mitogenic response in the five responsive lines tested. In two tumors (U138 and 9L), polymyxin B also eliminated EGF-, PDGF-, and serum-induced DNA synthesis as well as abolishing baseline DNA synthesis. These cells remained viable, however, as assessed by trypan blue exclusion; after removal of polymyxin B from the medium, they were able to resume DNA synthesis in response to TPA and serum. In the three other tumors (T98G and the two low-passage human glioma lines), growth factor-induced and serum-induced DNA synthesis were inhibited by approximately 25% to 85%. It is concluded that PKC-mediated pathways affect DNA synthesis in the human malignant glial tumors studied. The response of the glioma cells to TPA is similar to the responses seen in fetal astrocytes, but differs significantly from those reported for normal adult glial cultures. Because the response of the 9L glioma to TPA is similar to the responses seen in the human tumors, the 9L rat glioma model may prove useful for examining the role of PKC-mediated pathways in controlling glioma growth in vivo.

Animals↗
PubMed1990

Response of malignant glioma cell lines to epidermal growth factor and platelet-derived growth factor in a serum-free medium.

The use of a serum-free culture system for assessing the growth factor responsiveness of malignant glial cells is described. The mitogenic properties of epidermal growth factor (EGF) and platelet-derived growth factor (PDGF) were examined in three human malignant glioma cell lines (T98G, U87, and U138). Each of the three had high-affinity EGF receptors and all responded in a dose-dependent fashion to physiological concentrations of EGF. These cell lines also showed a pronounced mitogenic response to PDGF which equaled or exceeded that achieved with EGF. Simultaneous stimulation with both factors produced an additive response, which approximated that obtained in medium supplemented with 10% fetal calf serum. The authors conclude that functional EGF and PDGF receptors were present in the human malignant glial tumors studied. The response of the human glioma lines to these growth factors in many respects parallels the response seen in fetal astrocytes tested under similar conditions. In contrast, the behavior of two chemically induced rat gliomas (9L and C6) differed significantly from that seen in the human lines, suggesting that the rat lines may not be entirely acceptable as models for studying the growth characteristics of human malignant glial tumors.

Brain Neoplasms↗
PubMed1989

A method for recovery of native, clonally-restricted immunoglobulins from agarose gels.

Multiple low level, clonally-restricted, immunoglobulins (Ig) are commonly encountered on routine serum protein electrophoresis by clinical laboratories using high resolution zone electrophoresis on agarose. We sought a method for recovering the clonally-restricted Ig, in native configuration, from clinical laboratory gels as a first step in the investigation of its clinical significance. We found that a two-stage electrophoretic procedure gave consistently good recoveries. After routine agarose gel electrophoresis, portions of the electropherogram, containing clonally-restricted Ig, were excised and subjected to flatbed isoelectric focusing in agarose to enhance separation of the individual antibody clonotypes. Multiple slabs, containing the same clonally-restricted Ig, could be cut from adjacent tracks (i.e., tracks loaded with the same specimen) on the zone electropherogram and applied to a single track on the focusing gel to improve separation and increase yields. The focused gels were cut to isolate slabs containing individual clonotypes. These slabs were washed to remove carrier ampholytes and held at -20 degrees C overnight. Ig was extracted from the thawed gels, with 61-68% recovery, by ultracentrifugation following physical disruption of the gel. Antigen binding activity of the recovered Ig was verified by rate nephelometry. Clonally-restricted antibodies were successfully isolated from an immune animal serum by this procedure and biotinylated for use as probes on Western blots.

Electrophoresis↗
PubMed1988

Biclonal IgM gammopathy in chronic lymphocytic leukemia.

The association of chronic lymphocytic leukemia (CLL) with serum paraproteinemia (ie, monoclonal immunoglobulin production and secretion) is well known. We, however, could find only three previous reports of CLL where multiple serum paraproteins were encountered. We describe a case of biclonal gammopathy in CLL, involving IgM/kappa and IgM/lambda, with each paraprotein reaching serum levels of approximately 10 g/L (1 g/dL). Using immunohistochemical techniques, we identified two morphologically similar lymphocyte populations, which could be stained for either mu and kappa or mu and lambda. The peripheral blood contained a majority of mu/kappa-containing cells (kappa/lambda = 17.5:1), while the bone marrow only contained a modest excess of cells staining for mu/kappa (kappa/lambda = 2.4:1). The clinical significance and prognosis of biclonal IgM gammopathies is uncertain, since so few cases have been reported. Our patient has now been followed up for more than four years.

Aged↗
PubMed1987

Antibodies to native and denatured DNA. Quantitation using an immuno-slot-blot technique.

The attachment of DNA to nitrocellulose by suction filtration has been used as a prelude to DNA hybridization experiments, and several filter manifolds for this purpose are available. One suction manifold, the 'Minifold II slot-blotter', creates small, rectangular slots of bound DNA which can be probed using immunoenzymatic techniques and then evaluated with quantitative scanning densitometry. We applied this technique to the detection and quantitation of naturally occurring anti-DNA antibodies. After probing slot-blots of native or denatured DNA with human sera, the slot-blots were stained using an alkaline phosphatase second antibody system. The serum concentration of anti-DNA antibodies was determined by comparing the peak areas of the positive blots to a standard IgG curve. When native DNA was used as a substrate, these serum concentrations correlated well with the anti-DNA titer as measured by the Crithidia luciliae indirect immunofluorescence assay.

Antibodies↗