Search PubMed⌕ Search

Biomedical subjects

R Ross

Publications and source records attributed to R Ross.

At least 73 records · Page 4Linked to original sources
PubMed1998

Interactions between synthetic vanilloids and the endogenous cannabinoid system.

The chemical similarity between some synthetic agonists of vanilloid receptors, such as olvanil (N-vanillyl-cis-9-octadecenoamide), and the 'endocannabinoid' anandamide (arachidonoyl-ethanolamide, AEA), suggests possible interactions between the cannabinoid and vanilloid signalling systems. Here we report that olvanil is a stable and potent inhibitor of AEA facilitated transport into rat basophilic leukemia (RBL-2H3) cells. Olvanil blocked both the uptake and the hydrolysis of [14C]AEA by intact RBL-2H3 cells (IC50 = 9 microM), while capsaicin and pseudocapsaicin (N-vanillyl-nonanamide) were much less active. Olvanil was more potent than previously reported inhibitors of AEA facilitated transport, i.e. phloretin (IC50 = 80 microM), AM404 (12.9% inhibition at 10 microM) or oleoylethanolamide (27.5% inhibition at 10 microM). Olvanil was a poor inhibitor of [14C]AEA hydrolysis by RBL-2H3 and N18TG2 cell membranes, suggesting that the inhibitory effect on [14C]AEA breakdown observed in intact cells was due to inhibition of [14C]AEA uptake. Olvanil was stable to enzymatic hydrolysis, and (i) displaced the binding of high affinity cannabinoid receptor ligands to membrane preparations from N18TG2 cells and guinea pig forebrain (Ki = 1.64-7.08 microM), but not from cells expressing the CB2 cannabinoid receptor subtype; (ii) inhibited forskolin-induced cAMP formation in intact N18TG2 cells (IC50 = 1.60 microM), this effect being reversed by the selective CB1 antagonist SR141716A. Pseudocapsaicin, but not capsaicin, also selectively bound to CB1 receptor-containing membranes. These data suggest that some of the analgesic actions of olvanil may be due to its interactions with the endogenous cannabinoid system, and may lead to the design of a novel class of cannabimimetics with potential therapeutic applications as analgesics.

Animals↗
PubMed1998

The actin-bundling protein fascin is involved in the formation of dendritic processes in maturing epidermal Langerhans cells.

Dendritic cells (DC) are characterized by their unique potential to prime naive T cells. Epidermal Langerhans cells (LC), the DC resident in the epidermis, gain this immunostimulatory capacity following Ag contact in vivo or during in vitro culture of epidermal cell suspensions. To analyze differential gene expression in maturing LC, we constructed a highly representative cDNA library of cultivated LC (cLC) in lambda ZAP II containing 18 x 10(6) independent clones. This library was screened with freshly isolated Langerhans cell (fLC)- and cLC-derived probes for cLC-specific cDNAs. The cDNAs identified were sequenced and analyzed by database searches. Two cDNA fragments were identified as fragments of fascin, indicating that fascin is differentially expressed in LC. By competitive RT-PCR, we confirmed that fascin is highly expressed in cLC cultivated for 1, 2, and 3 days, while no signals were obtained with fLC. Western blot and immunofluorescence analysis revealed cLC-specific expression of fascin on the protein level as well. Fascin is known to be involved in the organization of the actin cytoskeleton in cytoplasmatic extensions of nerve growth cones. Its differential expression in maturing LC coincides with the formation of numerous dendritic projections in LC. Their formation was inhibited by incubation of LC with fascin antisense oligonucleotides during cultivation. Therefore, we conclude that fascin is necessary for the formation of the dendritic processes of maturing Langerhans cells and may thus influence T cell-LC interaction.

Actins↗
PubMed1998

Caspase-mediated cleavage of focal adhesion kinase pp125FAK and disassembly of focal adhesions in human endothelial cell apoptosis.

Normal endothelial and epithelial cells undergo apoptosis when cell adhesion and spreading are prevented, implying a requirement for antiapoptotic signals from the extracellular matrix for cell survival. We investigated some of the molecular changes occurring in focal adhesions during growth factor deprivation-induced apoptosis in confluent monolayers of human umbilical vein endothelial cells. Among the first morphologic changes after initiation of the apoptotic process are membrane blebbing, loss of focal adhesion sites, and retraction from the matrix followed by detachment. We observe a specific proteolytic cleavage of focal adhesion kinase (pp125FAK), an important component of the focal adhesion complex, and identify pp125FAK as a novel substrate for caspase-3 and caspase-3-like apoptotic caspases. The initial cleavage precedes detachment, and coincides with loss of pp125FAK and paxillin from focal adhesion sites and their redistribution into the characteristic membrane blebs of apoptotically dying cells. Cleavage of pp125FAK differentially affects its association with signaling and cytoskeletal components of the focal adhesion complex; binding of paxillin, but not pp130(Cas) (Cas, Crk-associated substrate) and vinculin, to the COOH terminally truncated pp125FAK is abolished. Therefore, caspase-mediated cleavage of pp125FAK may be participating in the disassembly of the focal adhesion complex and actively interrupting survival signals from the extracellular matrix, thus propagating the cell death program.

Apoptosis↗
PubMed1998

Cleavage of p21Cip1/Waf1 and p27Kip1 mediates apoptosis in endothelial cells through activation of Cdk2: role of a caspase cascade.

Apoptosis of human endothelial cells after growth factor deprivation is associated with rapid and dramatic up-regulation of cyclin A-associated cyclin-dependent kinase 2(cdk2) activity. In apoptotic cells, the C termini of the cdk inhibitors p21Cip1/Waf1 and p27Kip1 are truncated by specific cleavage. The enzyme involved in this cleavage is CPP32 and/or a CPP32-like caspase. After cleavage, p21Cip1/Waf1 loses its nuclear localization sequence and exits the nucleus. Cleavage of p21Cip1/Waf1 and p27Kip1 results in a substantial reduction in their association with nuclear cyclin-cdk2 complexes, leading to a dramatic induction of cdk2 activity. Dominant-negative cdk2, as well as a mutant of p21Cip1/Waf1 resistant to caspase cleavage, partially suppress apoptosis. These data suggest that cdk2 activation, through caspase-mediated cleavage of cdk inhibitors, may be instrumental in the execution of apoptosis following caspase activation.

Apoptosis↗
PubMed1998

Maturation of epidermal Langerhans cells in vitro is accompanied by downregulation of 4F2 (CD98) as determined by differential display.

Following short-term culture, Langerhans cells mature morphologically and functionally into potent immunostimulatory cells. As regulation of gene expression accompanies this maturation process, it is likely that differentially expressed genes are involved in the maturation events. Using the recently described method of differential display, we generated cDNA expression patterns starting with mRNA of murine epidermal Langerhans cells isolated either directly (fLC) or following 3 d cultivation (cLC). Five hundred putative differentially expressed cDNA fragments were recovered from the gel. For a part of the fragments differential expression was confirmed by dot blot and Southern hybridization procedures. These cDNA fragments were subcloned and sequenced following the verification step. Database searches revealed that unknown genes as well as already characterized genes were identified. A cDNA fragment preferentially hybridizing with fLC was identified as the murine surface marker 4F2 (CD98). Downregulation of the activation marker 4F2/CD98 was confirmed by additional analysis at the mRNA and protein level. The downregulation of 4F2 surface expression on cLC is compatible with the notion that the committed, terminally differentiated cLC downregulate proteins involved in proliferation and cell survival.

Animals↗
PubMed1998

Cleavage of beta-catenin and plakoglobin and shedding of VE-cadherin during endothelial apoptosis: evidence for a role for caspases and metalloproteinases.

Growth factor deprivation of endothelial cells induces apoptosis, which is characterized by membrane blebbing, cell rounding, and subsequent loss of cell-matrix and cell-cell contacts. In this study, we show that initiation of endothelial apoptosis correlates with cleavage and disassembly of intracellular and extracellular components of adherens junctions. beta-Catenin and plakoglobin, which form intracellular links between vascular endothelial cadherin (VE-cadherin) and actin-binding alpha-catenin in adherens junctions, are cleaved in apoptotic cells. In vitro incubations of cell lysates and immunoprecipitates with recombinant caspases indicate that CPP32 and Mch2 are involved, possibly by initiating proteolytic processing. Cleaved beta-catenin from lysates of apoptotic cells does not bind to endogenous alpha-catenin, whereas plakoglobin retains its binding capacity. The extracellular portion of the adherens junctions is also altered during apoptosis because VE-cadherin, which mediates endothelial cell-cell interactions, dramatically decreases on the surface of cells. An extracellular fragment of VE-cadherin can be detected in the conditioned medium, and this "shedding" of VE-cadherin can be blocked by an inhibitor of metalloproteinases. Thus, cleavage of beta-catenin and plakoglobin and shedding of VE-cadherin may act in concert to disrupt structural and signaling properties of adherens junctions and may actively interrupt extracellular signals required for endothelial cell survival.

Antigens, CD↗
PubMed1998

Involvement of NO in contact hypersensitivity.

The NO synthases (NOS) generate NO from L-arginine. High concentrations of NO have been shown to be responsible for tissue injury and cell death, while low concentrations of NO induce vasodilatation and other signaling effects. We have investigated the involvement of NO in contact hypersensitivity (CHS) reactions. CHS induced by treatment of BALB/c mice with the contact allergen 2,4-dinitrofluorobenzene (DNFB) was significantly reduced by the NOS inhibitor N-methyl-L-arginine (L-NMA), but not by the stereoisomer D-NMA, as shown by reduced ear swelling responses and evaluation of ear tissue sections. The CHS response was also reduced by aminoguanidine, which is known to preferentially inhibit the inducible isoform of the enzyme (iNOS), suggesting that iNOS contributed to the inflammatory response. We therefore investigated whether iNOS was expressed by epidermal cells. Epidermal Langerhans cells produced low but significant amounts of iNOS mRNA at the single-cell level as indicated by RT-PCR. Likewise, keratinocytes expressed basic iNOS mRNA levels. Elicitation of a CHS response by DNFB in vivo resulted in enhanced iNOS mRNA expression in Langerhans cells and keratinocytes, with higher levels of expression in Langerhans cells. The enhanced mRNA expression in Langerhans cells correlated with iNOS protein production as shown by immunofluorescence staining of epidermal sheets performing double staining with anti-iNOS and anti-MHC class II antibodies. Our data suggest that epidermal cell-derived NO contributes to the ear swelling reaction in CHS.

Allergens↗
PubMed1998

Does adipose tissue influence bioelectric impedance in obese men and women?

Bioelectric-impedance analysis overestimates fat-free mass in obese people. No clear hypotheses have been presented or tested that explain this effect. This study tested the hypothesis that adipose tissue affects measurements of resistance by using data for whole body and body segment resistance and by using muscle, adipose tissue, and bone volumes from magnetic resonance imaging for 86 overweight and obese men and women (body mass index > 27 kg/m2; age 38.5 +/- 10.2 yr). In multiple-regression analysis, muscle volumes had strong associations with resistance, confirming that the electric currents are conducted primarily in the lean soft tissues. Subcutaneous adipose tissue had a slight but statistically significant effect in women, primarily for the leg, suggesting that adipose tissue can affect measured resistance when the volume of adipose tissue is greater than muscle volume, as may occur in obese women in particular. This resulted in a slight overestimation of fat-free mass (e.g., +3 kg) when a bioelectric-impedance-analysis equation calibrated for nonobese female subjects was applied.

Adipose Tissue↗
PubMed1998

Cadaver validation of skeletal muscle measurement by magnetic resonance imaging and computerized tomography.

Magnetic resonance imaging (MRI) and computerized tomography (CT) are promising reference methods for quantifying whole body and regional skeletal muscle mass. Earlier MRI and CT validation studies used data-acquisition techniques and data-analysis procedures now outdated, evaluated anatomic rather than adipose tissue-free skeletal muscle (ATFSM), studied only the relatively large thigh, or found unduly large estimation errors. The aim of the present study was to compare arm and leg ATFSM cross-sectional area estimates (cm2) by using standard MRI and CT acquisition and image-analysis methods with corresponding cadaver estimates. A second objective was to validate MRI and CT measurements of adipose tissue embedded within muscle (interstitial adipose tissue) and surrounding muscle (subcutaneous adipose tissue). ATFSM area (n = 119) by MRI [38.9 +/- 22.3 (SD) cm2], CT (39.7 +/- 22.8 cm2), and cadaver (39.5 +/- 23.0 cm2) were not different (P > 0.001), and both MRI and CT estimates of ATFSM were highly correlated with corresponding cadaver values [MRI: r = 0.99, SE of estimate (SEE) 3.9 cm2, P < 0.001; and CT: r = 0.99, SEE = 3.8 cm2, P < 0.001]. Similarly good results were observed between MRI- and CT-measured vs. cadaver-measured interstitial and subcutaneous adipose tissue. For MRI-ATFSM the intraobserver correlation for duplicate measurements in vivo was 0. 99 [SEE = 8.7 cm2 (2.9%), P < 0.001]. These findings strongly support the use of MRI and CT as reference methods for appendicular skeletal muscle, interstitial and subcutaneous adipose tissue measurement in vivo.

Adipose Tissue↗
PubMed1998

Upregulation of VCAM-1 and ICAM-1 at atherosclerosis-prone sites on the endothelium in the ApoE-deficient mouse.

Focal recruitment of monocytes and lymphocytes is one of the earliest detectable cellular responses in the formation of lesions of atherosclerosis. This localized accumulation of leukocytes is a multistep process in which the endothelium remains intact and may regulate leukocyte recruitment by expressing specific adhesion molecules. To examine the relationship of adhesion molecule expression to initiation factors and the sites of lesion formation, we analyzed the expression of vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), and platelet-endothelial cell adhesion molecule-1 (PECAM-1) en face on the aortic endothelium of control mice and homozygous apolipoprotein E-deficient (ApoE -/-) mice that develop complex lesions of atherosclerosis similar to those in humans. In control mice, VCAM-1 staining was weak and limited to sites of altered blood flow. In contrast, in the ApoE -/- mice, VCAM-1 appeared to be localized over the surface of groups of endothelial cells in lesion-prone sites. Expression of VCAM-1 preceded lesion formation, and increased expression above control levels appeared to be correlated with the extent of exposure to plasma cholesterol. Although ICAM-1 was the most prominent adhesion molecule in lesion-prone sites, its expression appeared to be independent of plasma cholesterol levels and was upregulated in both ApoE -/- and control mice. At lesion-prone sites associated with altered blood flow, ICAM-1 was located over the surface of each endothelial cell and on microvilli, whereas VCAM-1 was confined to the cell periphery in non-lesion-prone sites. PECAM-1 was localized at the cell periphery throughout the aorta, and its expression did not appear to be regulated. Thus, the levels, localization, and characteristics of expression of VCAM-1, ICAM-1, and PECAM-1 appear to be differentially regulated. Upregulation of VCAM-1 and ICAM-1 is associated with sites of lesion formation.

Animals↗
PubMed1998

Age, ethnicity, and comorbidity in a national sample of hospitalized alcohol-dependent women veterans.

OBJECTIVE: Treatment patterns in a national sample of hospitalized women veterans diagnosed with alcohol dependence were identified with the goal of improving health services to women veterans with alcohol-related disorders. METHODS: Information from VA's patient treatment file for fiscal year 1993 was used to identify 854 women veterans diagnosed with alcohol dependence. Of that group, 546 received a primary diagnosis of alcohol dependence, and 308 received a secondary diagnosis of alcohol dependence after they sought treatment for other health problems. Chi square tests and multivariate logistic regression analyses were used to examine relationships between the sociodemographic profiles of these women and the types of services they received. RESULTS: The study population's largest age group (49 percent) was 30 to 39 years old. Fifty-two percent of the women were divorced or separated, and 62 percent were Caucasian. The overwhelming majority of comorbid diagnoses were of psychiatric disorders. Overall, only 47 percent of the 854 patients received formal treatment for their alcohol disorder, and only 34 percent completed alcohol treatment. Women over age 60 were significantly less likely than women in other age groups to enter or complete formal treatment. Native-American women were significantly more likely than Caucasians or African Americans to receive formal alcohol treatment services. CONCLUSIONS: The results indicate a need for targeting interventions more effectively in certain groups of women veterans diagnosed with alcoholism. Low completion rates also suggest a need for greater incentives for patients to complete treatment programs.

Adult↗
PubMed1998

Proteoglycan distribution in lesions of atherosclerosis depends on lesion severity, structural characteristics, and the proximity of platelet-derived growth factor and transforming growth factor-beta.

The accumulation of proteoglycans (PGs) in atherosclerosis contributes to disease progression and stenosis and may partly depend on local regulation by growth factors such as platelet-derived growth factor (PDGF) and transforming growth factor (TGF)-beta. In this study, the distribution of the major extracellular PGs is compared with that of PDGF and TGF-beta isoforms in developing lesions of atherosclerosis from hypercholesterolemic nonhuman primates. Strong immunostaining for decorin, biglycan, versican, and hyaluronan is observed in both intermediate and advanced lesions. Perlecan staining is weak in intermediate lesions but strong in advanced lesions in areas bordering the plaque core. Immunostaining for PDGF-B and TGF-beta1 is particularly prominent in macrophages in intermediate and advanced lesions. In contrast, TGF-beta2 and TGF-beta3 and PDGF-A are present in both macrophages and smooth muscle cells. Overall, PG deposits parallel areas of intense growth factor immunostaining, with trends in relative localization that suggest interrelationships among certain PGs and growth factors. Notably, decorin and TGF-beta1 are distributed similarly, predominantly in the macrophage-rich core, whereas biglycan is prominent in the smooth muscle cell matrix adjoining TGF-beta1-positive macrophages. Versican and hyaluronan are enriched in the extracellular matrix adjacent to both PDGF- and TGF-beta1-positive cells. These data demonstrate that PG accumulation varies with lesion severity, structural characteristics, and the proximity of PDGF and TGF-beta.

Animals↗
PubMed1997

The mitogen-activated protein kinase pathway can mediate growth inhibition and proliferation in smooth muscle cells. Dependence on the availability of downstream targets.

Activation of the classical mitogen-activated protein kinase (MAPK) pathway leads to proliferation of many cell types. Accordingly, an inhibitor of MAPK kinase, PD 098059, inhibits PDGF-induced proliferation of human arterial smooth muscle cells (SMCs) that do not secrete growth-inhibitory PGs such as PGE2. In striking contrast, in SMCs that express the inducible form of cyclooxygenase (COX-2), activation of MAPK serves as a negative regulator of proliferation. In these cells, PDGF-induced MAPK activation leads to cytosolic phospholipase A2 activation, PGE2 release, and subsequent activation of the cAMP-dependent protein kinase (PKA), which acts as a strong inhibitor of SMC proliferation. Inhibition of either MAPK kinase signaling or of COX-2 in these cells releases them from the influence of the growth-inhibitory PGs and results in the subsequent cell cycle traverse and proliferation. Thus, the MAPK pathway mediates either proliferation or growth inhibition in human arterial SMCs depending on the availability of specific downstream enzyme targets.

Cell Division↗