Biomedical subjects

J North

Publications and source records attributed to J North.

At least 19 recordsLinked to original sources

PubMed2003

Measurement and interpretation of pneumococcal IgG levels for clinical management.

The detection of pneumococcal IgG antibodies is helpful for the evaluation of response to pneumococcal vaccination and need for revaccination. Results generated by the clinical assay which is currently used, in which the 23 valent polysaccharide vaccine is the antigen, were compared to those obtained by a capsular polysaccharide serotype-specific assay that measures IgG antibodies to 9 common serotypes causing invasive disease. Discrepancies in 21/47 (45%) of the results were observed in a direct comparison between the two assays. In each case a positive titre was obtained on the clinical assay but IgG levels on the serotype-specific assay were below the putative protective level of 0.2 micro g/ml for at least one of the 9 serotypes assayed. The generation of false positives by the current clinical assay is due to its lack of specificity. Antibodies to C-polysaccharide and all of the 23 serotypes included in the pneumococcal polysaccharide vaccine are incorporated into the final titre whereas the serotype-specific assay adsorbs out noncapsular polysaccharide antibodies. The discrepancies between the two assays highlight the importance of standardized assays that measure putative correlates of protection and demonstrate the need to re-evaluate the current clinical assay. A tool that allows the interpretation of the results of the serotype-specific assay is provided and its potential for assessing individual susceptibility levels to vaccine preventable pneumococcal infection is discussed.

Adolescent↗

PubMed2002

The pre-operative identification of low-risk endometrialcancer: an audit of women treated in the South Island of New Zealand 1998-2000.

OBJECTIVE: To determine whether pre-operative investigations identify a group of patients with low-risk endometrial cancer, who do not require tertiary referral for surgical staging or pelvic radiotherapy. DESIGN: Retrospective chart review. SETTING: South Island of New Zealand gynaecological oncology services. SAMPLE: One hundred and forty consecutive patients with newly diagnosed endometrial cancer from 1988 to 2000. METHODS: The results of preoperative investigations were compared with the final pathology. MAIN OUTCOME MEASURES: Correlation of preoperative investigations with low risk disease. For the purpose of the study, women with grade 1 or 2 endometrioid tumours confined to the uterine body and less than 50% myometrial invasion were considered to have low risk disease. RESULTS: In total, 50 women had low risk disease. Only 53% of patients with grade 1 tumours on initial biopsy had low risk disease. Women who had a grade 1 tumour at biopsy and, an ultrasound report with an endometrial thickness of less than 20 mm, and no evidence of myometrial invasion, cervical involvement or adnexal metastasis had a 76% chance of having low risk disease. CONCLUSION: We were unable to accurately define the low risk group from pre-operative assessment.

Curettage↗

PubMed2001

Use of Leu3a/3b for the accurate determination of CD4 subsets for measurement of intracellular cytokines.

BACKGROUND: Identification of human T-helper cell subsets is possible by measurement of intracellular cytokines after coincubation of lymphocytes with phorbol myristate acetate (PMA), calcium ionophore, and brefeldin A for up to 20 h. However, exposure to PMA leads to internalization of membrane CD4 and to loss of resolution of the CD4+ cells. Detection of CD3+CD8- cells or preselection of CD4+ cells prior to stimulation is more cumbersome than direct measurement of CD4+ cells. We report the use of the Leu3a/Leu3b multiclone for the accurate determination of CD4 cells after PMA stimulation. METHODS: Peripheral blood lymphocytes were isolated from healthy normal donors and the proportion of CD3+ / CD4+ T cells was determined by flow cytometry before and after incubation with PMA, calcium ionophore, and brefeldin A for 20 h using a variety of anti-CD4 monoclonal antibodies. RESULTS: The Leu3a/3b multiclone reagent was the only anti-CD4 monoclonal antibody capable of resolving more than 98% of the initial CD4+ events after incubation with PMA. CONCLUSIONS: The higher signal-to-noise ratio associated with Leu3a3b reagent, compared with other CD4-specific antibodies available, allows the direct and accurate identification of the CD4 subset even after PMA treatment of cells.

CD4 Antigens↗

PubMed2001

Subfamilies of CR1 non-LTR retrotransposons have different 5'UTR sequences but are otherwise conserved.

CR1 elements and CR1-related (CR1-like) elements are a novel family of non-LTR retrotransposons that are found in all vertebrates (reptilia, amphibia, fish, and mammals), whereas more distantly related elements are found in several invertebrate species. CR1 elements have several features that distinguish them from other non-LTR retrotransposons. Most notably, their 3' termini lack a polyadenylic acid (poly A) tail and instead contain 2-4 copies of a unique 8 bp repeat. CR1 elements are present at approximately 100,000 copies in the chicken genome. The vast majority of these elements are severely 5' truncated and mutated; however, six subfamilies (CR1-A through CR1-F) are resolved by sequence comparisons. One of these subfamilies (i.e. CR1-B) previously was analyzed in detail. In the present study, we identified several full-length elements from the CR1-F subfamily. Although regions within the open reading frames and 3' untranslated regions of CR1-F and CR1-B elements are well conserved, their respective 5' untranslated regions are unrelated. Thus, our results suggest that new CR1 subfamilies form when elements with intact open reading frames acquire new 5' UTRs, which could, in principle, function as promoters.

Amino Acid Sequence↗

PubMed2001

Detection of different strains of Potato virus Y and their mixed infections using competitive fluorescent RT-PCR.

A competitive fluorescent RT-PCR assay (CF RT-PCR) was developed for the rapid and reliable detection and discrimination of the two most common strains of Potato virus Y (PVY) found in potato (necrotic and ordinary). The assay incorporates two strain specific primers labelled with fluorescent labels, used in conjunction with a universal PVY primer. The strain specific primers compete for the same annealing site which further increases specificity. Discrimination is conferred by the fluorescent labels; green PCR products for PVY(O) and red for PVY(N), whilst mixed infections are detected as orange PCR products without the need for staining agarose gels. The assay can be scaled up for the processing of 96 samples simultaneously, with the detection of PCR products directly using a fluorescent microtitre plate reader. The assay successfully discriminated between 20 isolates of PVY tested, and could be used for the direct detection of PVY in potato tubers.

Antibodies, Viral↗

PubMed2001

Grit-blasted titanium femoral stem in cementless primary total hip arthroplasty: a 5- to 10-year multicenter study.

A total of 133 cementless primary total hip arthroplasties using the Zweymuller-Alloclassic grit-blasted titanium tapered stem were performed in 3 institutions. The patient cohort was divided into 2 subgroups, nonselected and selected, on the basis of excellent bone stock and age (<65 years old at surgery). Acetabular components were all cementless, and bearing surfaces were all alumina-ceramic on polyethylene. After a 7.3-year average follow-up period (range, 5-10 years), 118 primary femoral replacements in 109 patients could be reviewed fully. Mean age at surgery was 55.7 years (range, 27-84 years). According to the Merle d'Aubigne and Charnley rating system, clinical results were graded excellent and good in 89% of hips and fair in 11%. Radiologically, early subsidence >2 mm could be detected in 4 hips (3.4%). Calcar atrophy and spot welds were noted in 77% and 82% of hips. Femoral osteolysis granuloma was noted in 4 hips (3.4%). There has been no stem fracture and no ceramic head breakage. The survivorship at 10 years with definite femoral aseptic loosening (radiographic failure) as the endpoint was 100% (95% Wilson confidence interval, 78.4%-100%; worst scenario, 95.4%). A significant difference between the nonselected and selected patient subgroups was observed only for early reoperation (P =.03) and proximal stress shielding (P =.01). Press-fitting but not filling the femoral canal with a rough titanium, straight, tapered femoral component represents, at intermediate follow-up, a promising cementless option in primary total hip arthroplasty.

Adult↗

PubMed2000

Interleukin-9 enhances interleukin-5 receptor expression, differentiation, and survival of human eosinophils.

Interleukin-9 (IL-9) has been implicated in the pathogenesis of allergic disorders. To examine the interaction between IL-9 and eosinophils, we evaluated mature peripheral blood eosinophils for their expression of the specific alpha-subunit of the IL-9 receptor (IL-9R-alpha). The expression of IL-9R-alpha by human eosinophils was detected at the messenger RNA (mRNA) and protein levels by reverse transcriptase-polymerase chain reaction (RT-PCR), flow cytometry, and immunocytochemical analysis, respectively. Functional analyses demonstrated that recombinant human (rh)IL-9 inhibited in vitro peripheral blood human eosinophil apoptosis in a concentration-dependent manner. We then examined the role of IL-9 in eosinophil differentiation using the human cord blood CD34(+) cells and human promyelocytic leukemia cells (HL-60). The addition of IL-9 to CD34(+) cells cultured in IL-3 and IL-5 enhanced eosinophil development, and IL-9 alone induced the expression of IL-5R-alpha. IL-9 also up-regulated the IL-5R-alpha chain cell surface expression during terminal eosinophil differentiation of the HL-60 cell line. Our findings suggest that IL-9 may potentiate in vivo eosinophil function by increasing their survival and IL-5-mediated differentiation and maturation. Taken together, these results suggest a mechanism by which IL-9 potentiates airway and tissue eosinophilia.

Antigens, CD34↗

PubMed2000

Interleukin 5 regulates the isoform expression of its own receptor alpha-subunit.

The receptor for interleukin 5 (IL-5) consists of a cytokine-specific alpha chain (IL-5Ralpha) and a signaling beta chain, which is shared with interleukin 3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF). These 3 cytokines can act in eosinophil development and activation in vitro, but gene deletion or antibody blocking of IL-5 largely ablates eosinophilic responses in models of allergic disease or helminth infection. We investigated factors acting in differential IL-5Ralpha gene splicing to generate either the membrane-anchored isoform (TM-IL-5Ralpha) which associates with the common beta chain to allow IL-5 responsiveness, or a secreted, antagonist variant (SOL-IL-5Ralpha). In a murine myeloid cell line (FDC-P1), transfected with minigenes allowing expression of either IL-5Ralpha variant, IL-5 itself, but not IL-3 or GM-CSF, stimulated a reversible switch toward expression of TM-IL-5Ralpha. A switch from predominantly soluble isoform to TM-IL-5Ralpha messenger RNA (mRNA) expression was also seen during IL-5-driven eosinophil development from human umbilical cord blood-derived CD34(+) cells; this was accompanied by surface expression of IL-5Ralpha and acquisition of functional responses to IL-5. IL-3 and GM-CSF also supported eosinophil development and up-regulation of TM-IL-5Ralpha mRNA in this system, but this was preceded by expression of IL-5 mRNA and was inhibited by monoclonal antibody to IL-5. These data suggest IL-5-specific signaling, not shared by IL-3 and GM-CSF, leading to a switch toward up-regulation of functional IL-5Ralpha and, furthermore, that IL-3 and GM-CSF-driven eosinophil development is dependent on IL-5, providing an explanation for the selective requirement of IL-5 for expansion of the eosinophil lineage. (Blood. 2000;95:1600-1607)

Animals↗

PubMed2000

A quality assurance audit: phase III trial of maximal androgen deprivation in prostate cancer (TROG 96.01).

In 1997 the Trans-Tasman Radiation Oncology Group (TROG) performed a quality assurance (QA) audit of its phase III randomized clinical trial investigating the effectiveness of different durations of maximal androgen deprivation prior to and during definitive radiation therapy for locally advanced carcinoma of the prostate (TROG 96.01). The audit reviewed a total of 60 cases from 15 centres across Australia and New Zealand. In addition to verification of technical adherence to the protocol, the audit also incorporated a survey of centre planning techniques and a QA time/cost analysis. The present report builds on TROG's first technical audit conducted in 1996 for the phase III accelerated head and neck trial (TROG 91.01) and highlights the significant progress TROG has made in the interim period. The audit provides a strong validation of the results of the 96.01 trial, as well as valuable budgeting and treatment planning information for future trials. Overall improvements were detected in data quality and quantity, and in protocol compliance, with a reduction in the rate of unacceptable protocol violations from 10 to 4%. Audit design, staff education and increased data management resources were identified as the main contributing factors to these improvements. In addition, a budget estimate of $100 per patient has been proposed for conducting similar technical audits. The next major QA project to be undertaken by TROG during the period 1998-1999 is an intercentre dosimetry study. Trial funding and staff education have been targeted as the key major issues essential to the continued success and expansion of TROG's QA programme.

Androgen Antagonists↗

PubMed1999

Selective removal of plutonium 238 from a canal sediment using a carbonate-chelant soil washing technology (ACTDECON).

The Mound laboratory site in Miamisburg, OH, a former plutonium processing facility, contains approximately 40000 yd(3) (30,580 m3) of plutonium- and thorium-contaminated soils and sediments at levels that require remediation. Existing applicable remediation technologies are unsatisfactory, because they are expensive and do not provide volume reduction. ACT*DE*CON is a chemical soil leaching technology for the treatment of soils that utilizes contaminant dissolution via dilute selective solutions to remove radionuclides. In bench-scale tests, process parameters were developed for the optimal treatment of the Miami Erie Canal soil at the Mound site, combining the maximum plutonium removal with an acceptable amount of soil dissolution and minimizing the costs of reagents. Parameters evaluated included soil to extractant mass ratio, temperature, rinse solution composition, kinetics, and the application of several dewatering aids. Plutonium removal rates of >95% were achieved, and the residual plutonium in the treated soil proved to be very immobile-confirming that the process had removed the most accessible species of the radionuclide. Currently being tested at Mound is an engineering scale-up that includes an attrition scrubber, a counter-current extractor, and a reverse osmosis system. Economic evaluations based on bench-scale results put the treatment cost at US$278/yd(3) (US$364/m3), compared to US$350/yd(3) (US$458/m3) for the 'box-and-bury' baseline alternative treatment system.

Carbonates↗

PubMed1999

Increased mature and immature CCR3 messenger RNA+ eosinophils in bone marrow from patients with atopic asthma compared with atopic and nonatopic control subjects.

BACKGROUND: Eosinophil infiltration of the bronchial mucosa is characteristic of asthma. Eosinophils differentiate from CD34(+) progenitors. Animal models suggest cooperation between IL-5 and eotaxin to allow rapid mobilization of a pool of bone marrow eosinophils followed by recruitment to the airway mucosa. OBJECTIVE: The purpose of this study was to enumerate CD34(+) cell numbers in blood and bone marrow from atopic asthmatics and control subjects and to test the hypothesis that there is an increased bone marrow pool of CCR3(+) eosinophils in patients with atopic asthma, as compared with control subjects. METHODS: Bone marrow aspirates and peripheral blood were obtained from volunteers with asthma and control volunteers. CD34(+) cell numbers were evaluated by flow cytometry, and eosinophil colony-forming activity was evaluated by methylcellulose cultures. Mature eosinophils, eosinophil myelocytes, metamyelocytes, and band forms (immature eosinophils) were enumerated by morphologic findings and immunocytochemistry for eosinophil cationic protein. CCR3 and eotaxin mRNA expression was examined by in situ hybridization, and protein expression was examined by immunocytochemistry. CCR3(+) cells were further identified with Chromotrope 2R staining. RESULTS: CD34(+) cell numbers in bone marrow were increased in atopic subjects. Numbers of eosinophil colony-forming units in blood and bone marrow did not differ between groups. Percentages of both mature and immature eosinophils were increased in bone marrow from patients with atopic asthma, but not atopic patients with no asthma or normal control subjects. CCR3 was expressed by immature and mature bone marrow eosinophils. Eotaxin was expressed by bone marrow cells from all 3 groups, but there was no increase in subjects with asthma. CONCLUSION: These findings suggest that in humans there is an increased bone marrow pool of CCR3(+) mature and immature eosinophils available for rapid mobilization in subjects with asthma but not in atopic subjects with no asthma.

Adult↗

PubMed1999

CD34(+)/interleukin-5Ralpha messenger RNA+ cells in the bronchial mucosa in asthma: potential airway eosinophil progenitors.

Eosinophil differentiation is thought to occur by the action of interleukin (IL)-5 on CD34(+) progenitor cells. The allergen-induced increase in eosinophil numbers in isolated airway preparations in vitro, and detection of increased numbers of circulating CD34(+) cells in atopic subjects, led us to the hypothesis that the eosinophil infiltration of the airway in asthma may result from local mucosal differentiation, in addition to recruitment from the bone marrow. We examined CD34(+) cell numbers by immunohistochemistry and IL-5 receptor alpha (IL-5Ralpha) messenger RNA (mRNA) expression by in situ hybridization in bronchial biopsies from atopic asthmatic patients, and from atopic and nonatopic control subjects. CD34(+) cell numbers were increased in the airway in atopic asthmatic and atopic nonasthmatic subjects. In contrast, CD34(+)/ IL-5Ralpha mRNA+ cells were increased in asthmatic subjects when compared with both atopic and nonatopic control subjects. Airway numbers of CD34(+)/IL-5Ralpha mRNA+ cells were correlated to airway caliber in asthmatic subjects and to eosinophil numbers. These findings support the concept that eosinophils may differentiate locally in the airway in asthma.

Adult↗

PubMed1999

The killing effect of photodynamic therapy using benzoporphyrin derivative on retinoblastoma cell line in vitro.

OBJECTIVE: To evaluate the killing effect of photodynamic therapy (PDT) using new photosensitizer benzoporphyrin derivative complexed with human low density lipoprotein on retinoblastoma (RB) cell line in vitro. METHODS: The experiment of photodynamic killing effect on RB cell line in vitro was performed by using benzoporphyrin derivative (BPD or Verteporfin) and monochromatic light at the wavelength around 690 nm. Seven BPD concentrations (2,500 ng/ml, 1,250 ng/ml, 625 ng/ml, 312.5 ng/ml, 156.25 ng/ml, 78.125 ng/ml, 39.0625 ng/ml) and three energy densities (1.2 J/cm2, 2.4 J/cm2 and 3.6 J/cm2) were applied. The damage of the tumor cells was evaluated by MTT assay 24 hours after PDT. The changes of the ultrastructure of RB cells were observed under eleceronic microscope 4 hours after PDT. RESULTS: There was a significant dose-response relationship between tumor cell damage and BPD concentration in the medium under light irradiation at the energy density of 1.2 J/cm2. At each BPD concentration, the inhibition rate increased with the rise of energy density. RB tumor cell necrosis was found widely under electronic microscope. CONCLUSIONS: This study suggested that RB cells are very sensitive to the PDT induced by BPD in vitro. RB tumor cell were directly killed by photodynamic effect induced by BPD.

Cytotoxicity, Immunologic↗

PubMed1998

Costimulation through CD86 is involved in airway antigen-presenting cell and T cell responses to allergen in atopic asthmatics.

Atopic allergic asthma is characterized by activation of Th2-type T cells in the bronchial mucosa. Previous reports have suggested an important role for costimulation through the CD28/CTLA4-CD80/CD86 pathway in allergen activation of T cells in animal models of inhaled allergen challenge. However, human allergen-specific lines and clones were reported to be costimulation independent. We therefore examined CD80 and CD86 dependence of allergen-induced T cell proliferation and cytokine production in peripheral blood and bronchoalveolar lavage from atopic asthmatic subjects and controls. Both allergen-induced proliferation and IL-5 production from PBMC were inhibited by CTLA4-Ig fusion protein and anti-CD86, but not anti-CD80 mAbs. When allergen-specific CD4+ T cell lines from peripheral blood were examined, proliferation and cytokine production were found to be independent of CD80 or CD86 costimulation. However, when cells obtained directly from the airways were examined, allergen-induced proliferation of bronchoalveolar lavage T cells from atopic asthmatic subjects was inhibited by anti-CD86 but not anti-CD80. In addition, bronchoalveolar lavage-adherent cells from asthmatic, but not control subjects showed APC activity to autologous T cells. This was also inhibited by anti-CD86 but not anti-CD80. Thus allergen-induced T cell activation and IL-5 production in the airway in asthmatic subjects is susceptible to blockade by agents interfering with costimulation via CD86, and this may hold therapeutic potential in asthma.

Abatacept↗

PubMed1998

IL-3, IL-5, granulocyte-macrophage colony-stimulating factor receptor alpha-subunit, and common beta-subunit expression by peripheral leukocytes and blood dendritic cells.

BACKGROUND: IL-3, IL-5, and granulocyte-macrophage colony-stimulating factor (GM-CSF) receptors consist of cytokine-specific alpha-subunits, which associate with a shared signalling common beta-subunit (beta(c)) to form a high-affinity complex. The expression of IL-3, IL-5, and GM-CSF is upregulated in atopic inflammation, and these cytokines are thought to contribute to pathology through mechanisms that include eosinophil activation. OBJECTIVE: We sought to examine the distribution of receptor expression between cells relevant to allergic inflammation from individual subjects and to compare atopic and nonatopic individuals. METHODS: Peripheral blood was obtained from atopic and nonatopic volunteers. Cytokine-receptor expression was examined by flow cytometry with monoclonal antibodies specific for alpha-subunits and beta(c) in combination with phenotypic markers for eosinophils, basophils, neutrophils, dendritic cells, monocytes, and T cells. RESULTS: Using a ligand-independent system, we confirmed the cellular distribution of IL-5Ralpha, IL-3Ralpha, and GM-CSFRalpha. IL-3Ralpha and GM-CSFRalpha were detected on high-affinity IgE receptor blood dendritic cells. Beta(c) expression was detected on basophils, eosinophils, neutrophils, and, at low levels, on monocytes and dendritic cells. There was intense staining of basophils for IL-3Ralpha relative to IL-5Ralpha, GM-CSFRalpha, and beta(c), whereas eosinophil-staining intensity was similar for IL-3Ralpha, IL-5Ralpha, GM-CSFRalpha, and beta(c). There were no significant differences between atopic and nonatopic subjects in cytokine-receptor staining. CONCLUSION: IL-3Ralpha and GM-CSRalpha are shown on a newly defined population of Fc(epsilon)RI-high dendritic cells. The intense staining of basophils for IL-3Ralpha, relative to that of IL-5Ralpha and GM-CSFRalpha, is in contrast to eosinophils from the same subjects and may explain the higher sensitivity of basophils to IL-3 compared with IL-5 and GM-CSF. We found no evidence for downregulation of receptor expression in atopic compared with nonatopic subjects, suggesting that these receptors remain accessible as potential targets for therapeutic intervention in atopic allergic disease.

Adolescent↗

PubMed1998

Na-dependent transport of S-(1,2-dichlorovinyl)-L-cysteine by renal brush-border membrane vesicles.

Cytotoxicity after exposure to the nephrotoxicant S-(1, 2-dichloro-vinyl)-L-cysteine (DCVC) requires transport of this cysteine conjugate across the cell membrane. Although several basolateral transport pathways have been implicated in the uptake of this compound into renal proximal cells, the identity of the process or processes associated with transport across the luminal membrane is unclear. We used a preparation of luminal brush-border membrane vesicles to characterize the transport of [35S]DCVC in rabbit kidney. An inwardly directed Na-gradient stimulated the initial rate of DCVC uptake by 16-fold compared to uptake measured in the absence of Na+. The Na-dependent component of DCVC uptake was stimulated by imposition of an inside-negative electrical potential difference and was blocked by the presence of 5 mM unlabeled DCVC in the extravesicular solution. Transport of DCVC was adequately described by Michaelis-Menten kinetics with an apparent Kt of 0.5 mM. DCVC uptake was blocked by the presence in the extravesicular solution of 10 mM concentrations of phenylalanine, leucine and cysteine, but not by glycine, proline, lysine, taurine, N-acetyl DCVC, p-aminohippurate, lactate or succinate. Unlabeled DCVC inhibited uptake of [14C]phenylalanine by a mechanism that exerted a greater effect on the apparent Kt than on the Jmax of phenylalanine, implicating a possible competitive interaction between these compounds. The carrier-mediated permeability of DCVC (defined as the ratio of Jmax/Kt) in luminal brush border membranes was as large as or larger than that reported for a battery of other organic electrolytes, including several amino acids and organic anions. We conclude that luminal transport of DCVC in rabbit proximal cells is limited to a single Na-cotransport process that also handles phenylalanine.

Animals↗

PubMed1998

Relation of cysteine conjugate nephrotoxicity to transport by the basolateral organic anion transport system in isolated S2 segments of rabbit proximal renal tubules.

We examined basolateral transport of the radiolabeled zwitterionic nephrotoxic cysteine S-conjugate, S-(1,2-dichlorovinyl)-L-cysteine (DCVC), inhibition of such transport and the effects of inhibition of transport on the toxicity produced by DCVC in isolated S2 segments of rabbit proximal tubules. High concentrations of unlabeled DCVC itself and an unlabeled nontoxic cysteine S-conjugate, S-(2-benzothiazole)-L-cysteine cis-inhibited the basolateral uptake of radiolabeled DCVC by approximately 80 to 85%. High concentrations of para-aminohippurate, the prototype substrate for the basolateral organic anion transport system, and probenecid, a well-known inhibitor of basolateral organic anion transport, cis-inhibited the basolateral uptake of radiolabeled DCVC by approximately 70%, whereas a high concentration of L-phenylalanine had little effect. High concentrations of S-(2-benzothiazole)-L-cysteine and para-aminohippurate in the bathing medium with DCVC inhibited the loss of 86Rb (used as a K+ surrogate to measure toxicity) from S2 segments produced by DCVC alone to approximately the same extent as they inhibited uptake of DCVC. Under the same circumstances, probenecid completely inhibited 86Rb loss. These data indicate that in rabbit proximal renal S2 tubules basolateral entry of DCVC can occur to a major extent via the organic anion transport pathway and that inhibition of such entry can reduce toxicity to approximately the same extent that entry is reduced. They also suggest that probenecid provides additional protection from DCVC toxicity.

Animals↗

PubMed1997

Reduction of trans-4,5-dihydroxy-1,2-dithiane by cellular oxidoreductases activates gadd153/chop and grp78 transcription and induces cellular tolerance in kidney epithelial cells.

trans-4,5-Dihydroxy-1,2-dithiane, the intramolecular disulfide form of dithiothreitol (DTTox) transcriptionally activates the stress-responsive genes gadd153(chop) and grp78. Herein, we used a renal epithelial cell line, LLC-PK1, to investigate the mechanism(s) whereby DTTox activates a molecular stress response. DTTox activated both grp78 and gadd153 transcriptionally, but gadd153 mRNA stability also increased suggesting that both transcriptional and posttranscriptional mechanisms are involved. DTTox did not activate hsp70 transcription indicating that a heat shock response was not induced. Structure-activity studies showed that DTTox analogues lacking the intramolecular disulfide were inactive. Furthermore, the ring-open intermolecular disulfide form of DTTox, 2-hydroxyethyl disulfide, was only a weak inducer of grp78 and gadd153 but was a strong inducer of hsp70 mRNA and a potent oxidant that lowered the NADPH/NADP+ ratio and depleted reduced glutathione (GSH). DTTox had little effect on the overall GSH and NADPH levels; thus cells were not undergoing oxidative stress; however, the NADPH/NADP+ ratio decreased slightly indicating that reducing equivalents were consumed. LLC-PK1 cells reduced DTTox to DTT, and the kinetics as well as the concentration dependence for reduction correlated with induction of both grp78 and gadd153 mRNA. Prior treatment with DTTox rendered cells tolerant to the potent nephrotoxicant S-(1,1,2, 2-tetrafluoroethyl)-L-cysteine. Bacitracin, an inhibitor of plasma membrane oxidoreductases, blocked DTTox reduction and gene activation as well as DTTox-induced tolerance. Thus, activation of stress genes and induction of cellular tolerance by DTTox is mediated by a novel mechanism involving cellular oxidoreductases.

Animals↗