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Biomedical subjects

C H Campbell

Publications and source records attributed to C H Campbell.

At least 19 recordsLinked to original sources
PubMed2001

A comparison of membrane vs. intracellular estrogen receptor-alpha in GH(3)/B6 pituitary tumor cells using a quantitative plate immunoassay.

The plasma membrane form of the estrogen receptor-alpha (mER-alpha) is involved in rapid estrogen-induced prolactin release from GH(3)/B6 rat pituitary tumor cells and can be detected immunocytochemically using several estrogen receptor-alpha (ER-alpha) antibodies. We recently described staining of fixed cells via a biotin-avidin-alkaline phosphatase sandwich assay. From this protocol, we have developed a rapid, quantifiable 96-well plate immunoassay for mER-alpha, using a different alkaline phosphatase substrate, para-nitrophenylphosphate, which generates a soluble yellow product, para-nitrophenol. We also permeabilized cells with detergent during fixation to measure intracellular ER-alpha (iER-alpha) with the same assay and then compared intracellular versus membrane ER-alpha levels in two GH(3)/B6 cell subclones originally selected for high and absent mER-alpha expression by immunocytochemistry. While the F10 subclone expresses plentiful amounts of the mER-alpha, the D9 subclone has undetectable levels of mER-alpha using this assay. In addition, there is a seven-fold difference in iER-alpha expression between the high (F10) and no (D9) mER-alpha expressing subclones. In the high mER-alpha expressing cell line, the mER-alpha totals approximately one third of total cellular ER-alpha. Neither membrane or intracellular forms of ER-beta were detected with this assay. The pNp assay allows convenient and quantitative comparison of multiple parameters of mER-alpha and iER-alpha regulation and should be applicable to other antigens that are expressed on the cell surface as well as intracellularly.

Analysis of Variance↗
PubMed1999

Membrane oestrogen receptors on rat pituitary tumour cells: immuno-identification and responses to oestradiol and xenoestrogens.

Our laboratory has identified plasma membrane oestrogen receptors on a GH3/B6 rat pituitary tumour cell line and several sublines which produce rapid (within minutes), non-genomic responses to oestrogens. Oestrogen receptors have been identified by their binding to nine different antibodies (Abs) which together recognize at least seven epitopes on the oestrogen receptor-alpha. GH3/B6/F10 cells, a membrane oestrogen receptor-enriched subline, elevate intracellular calcium levels in response to 10 nM oestradiol. Prolactin release in these cells is triggered by both 1 pM and 1 nM oestradiol and diethylstilbestrol (DES). A membrane oestrogen receptor-alpha immunocytochemical signal rapidly disappears (at 3 min) and reappears (at 12-15 min) when 1 nM oestradiol, 10 nM diethylstilbestrol, or 10 nM nonylphenol is applied to the cells. This suggests that both oestrogens and xenoestrogens can utilize this alternative pathway for oestrogenic action. Xenoestrogens, which have so far shown weak effects in genomic assay systems, should now be retested for activity in eliciting membrane-initiated oestrogenic responses.

Animals↗
PubMed1997

The role of HIV counseling and testing in the developing world.

The role of voluntary HIV counseling and testing is still under debate, especially in the developing world. HIV counseling-and-testing (HIV CT) services are a major component of HIV and AIDS control programs in the industrialized world and are increasingly being advocated in the developing world. In the United States, voluntary HIV CT has been a major component of HIV prevention efforts since the HIV antibody test became available in 1985. Yet even in the United States, questions about the management, cost, and effectiveness of voluntary HIV CT services continue to be raised. Because HIV CT has multiple goals, the evaluation of its effectiveness is a complicated task. Worldwide, a broad range of ethical, social, policy, technical, and economic issues encompass this HIV prevention activity. This article identifies the substantial barriers and serious concerns that are raised about HIV CT services and attempts to highlight the potential advantages of providing HIV CT as part of a developing country's comprehensive HIV prevention strategy.

AIDS Serodiagnosis↗
PubMed1995

Annexins I, II and III are specific choline binding proteins.

We have isolated choline binding proteins from the plasma membrane fraction fraction of human lung epithelium-derived cell line (A549) by means of detergent solubilization, anion exchange and affinity chromatography. One of the affinity purified proteins had a specific choline binding activity of 44-57 pmol/mg, representing a two to three hundredfold enrichment relative to the specific activity of freshly prepared plasma membranes. The purified protein has a molecular mass of 38 kDa by SDS PAGE analysis and was identified as annexin II by N-terminal microsequencing. Annexin II, however, had not previously been known for choline binding activity. We therefore prepared a mixture of authentic annexins (I-V) from A549 cells. The mixture had a choline binding activity of 15 to 18 pmol/mg. The annexin mixture was subsequently affinity chromatographed on the choline-conjugated Sepharose 6B column. Analyses by SDS PAGE and immunoblot revealed that annexins I, II, and III are bound to the choline column while annexins IV and V did not. These results indicate that some of the annexins have specific choline binding activities.

Amino Acid Sequence↗
PubMed1993

Purification of a novel glycosylated ferritin from horse heart.

We have previously shown that mRNA coding for ferritin L subunit is present on both cytosolic ribosomes and endoplasmic reticulum-bound ribosomes in rat heart tissue [Campbell et al. (1989) Arch Biochem Biophys 273:89-98]; from this we infer that heart tissue is capable of making a secreted ferritin. We now report the purification from horse heart, of a ferritin that specifically binds to Concanavalin A-Sepharose and is immunologically cross-reactive with antibodies raised against both horse cellular ferritin and horse serum ferritin. Where cellular ferritin is 10 nm in diameter and contains primarily 21-kDa subunits (as determined by gel exclusion chromatography and electron microscopy), the glycosylated heart ferritin is smaller with diameters of 3-5 nm. Antisera raised against serum ferritin cross-reacted with the glycosylated heart ferritin did but did not show significant cross-reactivity with cellular ferritin thus indicating that serum ferritin and glycosylated heart ferritin have antigenic determinants which may not be present on cellular ferritin. The glycosylated ferritin also differs from cellular ferritin in subunit composition, with subunits of 66, 60.5, 53.5, 43.5, and 29.5 kDa, as shown by SDS-PAGE and Western blot analysis. Interestingly, ferritin purified from horse serum contains subunits of similar size.

Animals↗
PubMed1992

Herpes simplex virus infection as a risk factor for human immunodeficiency virus infection in heterosexuals.

To determine if infection with herpes simplex virus (HSV) type 2 is associated with human immunodeficiency virus (HIV) type 1 infection among patients attending sexually transmitted diseases clinics, a case-control study was done on coded sera from 179 HIV-1-infected patients and 367 age-, race-, and gender-matched HIV-1-seronegative patients. Although only 13 (2.3%) of 546 patients had a history of genital herpes treatment, 72% and 56.6%, respectively, had serologic evidence of prior infection with HSV-1 and -2. HSV-1 antibody prevalence was similar among both patient groups; however, HSV-2 antibodies were more common among those infected with HIV-1. Among heterosexual men, 62.7% of those infected with HIV-1 had HSV-2 antibodies compared with 46.7% of those not infected (P less than .01). The HSV-2 seroprevalence among women with or without HIV infection was 78.1% and 57.7%, respectively (P less than .02). A history of intravenous drug use and a reactive serologic test for syphilis were each independently associated with HIV-1 infection in heterosexuals. These data suggest that the two most common causes of genital ulcerative disease in the United States, genital herpes and syphilis, may contribute to increased risk for HIV-1 infection among heterosexuals.

Antibodies, Viral↗
PubMed1989

Ferritin mRNA is found on bound as well as on free polyribosomes in rat heart.

Free and endoplasmic reticulum-bound polyribosomes from rat heart were examined for their ferritin mRNA content. A procedure for separation and purification of the two ribosome populations that produced good yields of homogeneous mono- and polyribosomes with no contaminating ultrastructures and gave distinctive sedimentation profiles in 15-50% sucrose gradients was developed. 14C-labeled free and bound polyribosomes added to heart preparations indicated that only 3% of free and 5.5% of bound polyribosomes cross-contaminated the bound and free fractions, respectively. RNA from both polyribosome populations hybridized with [32P]cDNA for rat ferritin. The extent of hybridization with mRNA from endoplasmic reticulum (ER)-derived polyribosomes was much greater than what could be accounted for by cross-contamination with free polyribosomes. This indicates that heart ferritin is synthesized not only on free polyribosomes for internal use in iron storage but also on ER-bound polyribosomes, where it may be destined for secretion into the plasma.

Animals↗
PubMed1989

Translational regulation of ferritin synthesis in rat spleen: effects of iron and inflammation.

Translational control of ferritin synthesis was studied in rat spleen, and compared with that for liver, heart and brain, in response to iron and inflammation. Spleen concentrations of total RNA in the ribonucleoprotein (mRNP) fraction was comparable to that for liver, while polyribosomal RNA was less. Both fractions were ten-fold lower in heart and brain. In untreated animals, the mRNP fraction of all tissues had the largest portion of the ferritin mRNA, as determined by slot blot hybridization with 32P-labeled cDNA for the L subunit. Acute treatment with ferric ammonium citrate shifted the spleen ferritin mRNA to the polyribosome fraction. This was also so in liver but not in the heart and brain which took up much less iron. The findings were confirmed by hybridization studies of mRNPs and polyribosomes separated in sucrose gradients. Turpentine-induced inflammation also caused a shift in ferritin mRNA from the mRNP to the polyribosome fraction of spleen and liver, over 12 h. We conclude that as in liver, spleen ferritin synthesis is under translational control by iron, and that both tissues also respond to inflammation by shifting of ferritin mRNA to the polyribosomes.

Animals↗
PubMed1988

Human immunodeficiency virus infection among patients attending clinics for sexually transmitted diseases.

To assess the prevalence and associated risk factors for human immunodeficiency virus (HIV) infection in patients attending inner-city clinics for sexually transmitted diseases in Baltimore, we screened 4028 patients anonymously, of whom 209 (5.2 percent) were seropositive for HIV. HIV-seropositivity rates were higher among men (6.3 percent) than women (3.0 percent) (P less than 0.001) and among blacks (5.0 percent) than whites (1.2 percent) (P less than 0.02). Among men, but not women, HIV seroprevalence increased markedly and steadily up to the age of 40. In men, HIV seropositivity was independently associated with increased age, black race, a history of homosexual contact, and the use of parenteral drugs. In women, a history of parenteral drug use or of being a sexual partner of a bisexual man or parenteral drug user were independently predictive of HIV seropositivity. In men, HIV seropositivity was also associated with a history of syphilis or a reactive serologic test for syphilis, and in women, with a history of genital warts. Since these associations were independent of the type and number of reported sexual partners, they raise the possibility that sexually transmitted diseases that disrupt epithelial surfaces may be important in the transmissibility of HIV. In addition, on a self-administered questionnaire, one third of HIV-infected men and one half of infected women did not acknowledge previous high-risk behavior for HIV exposure. These data suggest that patients at clinics for sexually transmitted diseases represent a group at high risk for HIV infection, and that screening, counseling, and intensive education should be offered to all patients attending such clinics.

Acquired Immunodeficiency Syndrome↗
PubMed1987

Binding sites for elastase on cultured human fibroblasts that do not mediate internalization.

The proteolytic actions of elastases have been implicated in extracellular matrix damage, which is characteristic of a variety of pathological conditions including emphysema and rheumatoid arthritis. In order to elucidate the molecular events involved in elastase interaction with connective tissue cells, the present study was designed to investigate the association of elastase with human fibroblasts at 4 degrees C. Elastase bound saturably to binding sites that were present on the surface of these cells. Analysis of cell-bound elastase by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed the presence of a high molecular weight complex (Mr 54,000) that was not formed with elastase whose catalytic site serine was derivatized with a diisopropylphosphate group. The complex did not represent elastase bound to either protease nexin or contaminating serum. The cellular component with which elastase formed a complex could not be detected in the cell culture medium. Unexpectedly, elastase that had been pre-bound at 4 degrees C was not internalized after cells were warmed to 37 degrees C. The elastase binding site described in this report is therefore distinct from high affinity binding sites involved in receptor-mediated endocytosis and intracellular degradation.

Binding Sites↗
PubMed1986

Interactions of serine proteases with cultured fibroblasts.

This review summarizes the mechanisms by which several serine proteases, particularly urokinase, thrombin, and elastase, interact with cultured fibroblasts. Many of these studies were prompted by findings that interactions of these proteases with cells and the extracellular matrix are important in a number of physiologic and pathologic processes. Two main pathways have been identified for specific interactions of these proteases with fibroblasts. One involves surface binding sites for the free protease that appear to bind only one particular protease. An unusual feature collectively shared by the binding sites for urokinase, thrombin, and elastase is that the bound protease is not detectably internalized by the fibroblasts. The other pathway by which serine proteases interact with fibroblasts involves proteins named protease nexins (PNs). Three PNs have been identified. They are secreted by fibroblasts and inhibit certain serine proteases by forming a covalent complex with the protease catalytic site serine. The complexes then bind back to the fibroblasts via the PN portion of the complex and are internalized and degraded. Recent studies showing that the fibroblast surface and extracellular matrix accelerate the inactivation of thrombin by PN-1 support the hypothesis that the PNs control protease activity at and near the cell surface. The PNs differ from plasma protease inhibitors in their molecular properties, absence in plasma, site of synthesis, and site of clearance of the inhibitor:protease complexes.

Amyloid beta-Protein Precursor↗
PubMed1985

Immunogenicity of bluetongue virus inactivated by gamma irradiation.

The immunogenicity of vaccines prepared from bluetongue viruses inactivated by gamma irradiation was tested by their ability to stimulate the production of neutralizing antibody in mice. Antibody titres induced by monovalent and polyvalent vaccines, vaccines prepared from virus produced in mice and cell cultures and vaccines containing virus exposed to six, eight and ten megarads of irradiation indicated that immunogenicity was not adversely affected by the inactivation procedure. The results suggest that gamma irradiation would be an effective means of inactivating bluetongue virus without affecting antigenic determinants for neutralizing antibody.

Animals↗