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Biomedical subjects

R J Albertini

Publications and source records attributed to R J Albertini.

At least 91 records · Page 5Linked to original sources
PubMed1989

Molecular analyses of in vivo hprt mutations in human T-lymphocytes. III. Longitudinal study of hprt gene structural alterations and T-cell clonal origins.

The hprt clonal assay detects mutations occurring in vivo in the hypoxanthine-guanine phosphoribosyltransferase (hprt) gene of human T-lymphocytes. Analysis of 94 wild-type and 326 hprt mutant clones from 3 normal males was performed using Southern blotting with hprt and T-cell receptor (TCR) gene probes. Gross structural alterations of the hprt gene occurred in approximately 14% of the in vivo derived mutants. Breakpoints were randomly distributed across the gene with one possible mutational "hot spot" observed. Most hprt mutants were independent as judged by TCR gene rearrangement patterns indicating that the measured hprt mutant frequency is a good measure of the actual hprt mutation frequency. However, sibling mutants (generally doublets and triplets except for one nonamer) were detected. Information on the timing in vivo of the hprt mutational events and the persistence in vivo of sibling mutants was also obtained.

Adult↗
PubMed1988

Molecular analyses of in vivo hypoxanthine-guanine phosphoribosyltransferase mutations in human T-lymphocytes: II. Demonstration of a clonal amplification of hprt mutant T-lymphocytes in vivo.

Recent molecular analysis of in vivo-derived hprt mutant T-lymphocytes cloned from human blood show that mutants occurring at the normal frequency (approximately 5 X 10(-6) in healthy young individuals generally represent independent hprt mutations. Here we report that in an individual with a high mutant frequency (86-620 X 10(-6],92% (61/66) of the mutant clones are descendents of an original mature T-cell precursor that has undergone in vivo clonal expansion. Therefore, these mutants could represent as few as one original hprt mutation. If so, correcting for the clonal expansion yields a revised calculated mutant frequency (Mf) value for this individual that is near the normal range. These hprt mutant clones all showed identically rearranged T-cell receptor (TCR) beta and gamma gene patterns by Southern blot analysis. All the clones were surface marker CD4+, showed no obvious chromosomal aberration, and had no detectable hprt gene structural alteration. This TCR-defined T-cell clone appears to have expanded in the blood of the individual over a 6-month period and persists at high levels after nearly 4 years. This finding illustrates the need to analyze mutants from individuals with high mutant frequencies at the molecular level in order to estimate hprt mutation frequency from the calculated hprt mutant frequency. The possibility that spontaneous hprt mutants might arise in vivo preferentially in dividing cells, and implications of this, are discussed.

Adult↗
PubMed1988

An immunogenetic study of suppressor cell activity in autoimmune chronic active hepatitis.

Some patients with autoimmune chronic active hepatitis as well as their disease-free first degree relatives show decreased suppressor cell activity of peripheral blood T lymphocytes. Studies were therefore undertaken in families ascertained by the presence of a single chronic active hepatitis patient to determine if this abnormality of immune regulation represents a genetic phenotype simply controlled by a gene or genes at a putative disease susceptibility locus and, further, if this locus showed linkage to either the HLA or the immunoglobulin constant region loci. In addition to determining circulating autoantibody status and genotyping for HLA and immunoglobulin allotypes, suppressor T cells were evaluated by surface markers and by determining their ability to suppress IgG secretion in vitro. The results suggest that immunoregulatory dysfunction in autoimmune chronic active hepatitis is a familial abnormality, but that this abnormality occurs independent of circulating autoantibody status and of the segregation of genes for HLA or immunoglobulin allotypes.

Antigens, Differentiation↗
PubMed1988

Mutagenicity monitoring in humans by autoradiographic assay for mutant T lymphocytes.

Somatic cell mutation which occurs in vivo in humans can be determined by measurement of the frequency of the 6-thioguanine-resistant (TGr) T lymphocytes in samples of peripheral blood. This frequency can be determined by either a short-term autoradiographic methodology or a longer cell-cloning methodology. The advantage of the former is the relative simplicity of the assay, while the latter allows recovery of mutant clones for further characterization. This report presents results of a longitudinal study of cancer chemotherapy nurses and other health care personnel by use of the autoradiography assay. The use of this assay in human mutagenicity monitoring and the analysis of the TGr cell frequencies are discussed in terms of age effects and validation of 'elevated' frequencies by use of the clonal assay. This report then presents evidence that both assays yield similar TGr cell frequencies in two groups of 'normal adults'. The mean variant frequency (+/- S.D.) for 82 autoradiographic assays was 8.7 (+/- 6.1) X 10(-6), while the mean mutant frequency (+/- S.D.) for 115 clonal assays was 6.5 (+/- 4.8) X 10(-6). In addition, concurrent autoradiographic and clonal assays on 33 individuals yielded mean values (+/- S.D.) of 8.4 (+/- 8.5) X 10(-6) and 10.5 (+/- 6.3) X 10(-6), respectively.

Age Factors↗
PubMed1988

Association and linkage studies of immunoglobulin heavy chain allotypes in inflammatory bowel disease.

Inflammatory bowel disease is a familial disorder of unknown etiology, characterized by a variety of immunological abnormalities for which there are no proven genetic markers. Studies of immunoglobulin heavy chain and light chain allotypes were undertaken in 89 patients with Crohn's disease and in 18 families with ulcerative colitis or Crohn's disease in at least two first-degree relatives, to search for association and/or linkage between a postulated disease susceptibility locus and an immunoglobulin allotype locus. No evidence of association or linkage with a specific immunoglobulin allotype was found.

Adolescent↗
PubMed1988

Effect of commercial peritoneal dialysis fluids on the lytic function of lymphokine-activated killer cells.

Intraperitoneal (ip) immunotherapies are often administered in saline or peritoneal dialysis solutions. In preparation for a Phase I i.p. interleukin-2/lymphokine-activated killer cell (IL-2/LAK) trial in patients with recurrent ovarian cancer, we evaluated the effect of six solutions, including two 1.5% dextrose peritoneal dialysis solutions (PDS), on the lytic function of LAK. In vitro IL-2-activated LAK cells were exposed to solutions, washed with Hanks' balanced salt solution, and lytic activity was measured in a standard 4-h chromium release assay using Daudi as a tumor target. LAK function was abrogated after 2 h of exposure to Inpersol and after only 20 min of exposure to Dianeal PD-2. Five percent dextrose also significantly decreased lytic capabilities of LAK, whereas 0.9% sodium chloride or lactated Ringers had no deleterious effect on function. Adjustment of PDS to pH greater than 6.5 decreased the damaging effect on LAK function, suggesting that the low pH of PDS and 5% dextrose results in a loss of LAK cell viability and therefore lytic function. Based on these data, we have chosen to administer IL-2/LAK i.p. in lactated Ringers.

Cytotoxicity Tests, Immunologic↗
PubMed1987

Refinement of a T-lymphocyte cloning assay to quantify the in vivo thioguanine-resistant mutant frequency in humans.

Cell cloning by limiting dilution in 96-well microtiter plates has been employed to isolate colonies of human T-lymphocytes resistant to the purine analogue, 6-thioguanine (TG). These colonies show stability of the TG-resistant (TG1) phenotype, lack hypoxanthine guanine phosphoribosyl transferase (HPRT) activity and thus appear to be the result of in vivo somatic cell mutation events. In order to employ this T-lymphocyte cloning assay for quantitative determination of the in vivo TGr mutant frequency in humans, we have defined the optimal conditions for T-cell colony formation with both nonselected and TG-selected cells. The parameters investigated include medium, serum, amount of the mitogen phytohaemagglutinin, amount of T-cell growth factor (TCGF) and the number of irradiated feeder or accessory cells. Under the optimal conditions, the fraction of positive wells is proportional to the number of cells plated per well with both nonselection and TG selection conditions. T-cell cloning efficiencies therefore are independent of inoculum size. There was some evidence for a decline in TGr mutant cell cloning at densities greater than 2 x 10(4) cells per round-bottom well, possibly due to metabolic cooperation between wild-type and mutant cells. The conditions defined in this study appear to provide a quantitative measurement of the in vivo TGr mutant frequency in human T-lymphocytes.

Antigen-Presenting Cells↗
PubMed1987

Molecular analyses of in vivo hprt mutations in human T-lymphocytes. I. Studies of low frequency 'spontaneous' mutants by Southern blots.

Fifty wild-type and 164 in vivo-derived hprt mutant T-cell clones obtained from eight non-mutagen-exposed adult males with mutant frequency values in the normal range (usually less than 10 X 10(-6) were studied by Southern blot analyses to determine frequency and extent of gross structural alterations in the hprt gene. Sixteen (9.8%) of the mutant clones showed hprt changes. No site or type of lesion predominated. Relative frequencies of gross structural alterations in the recovered hprt mutants did not differ among the eight individuals, within limits detectable by the study. DNA from 201 of these 214 clones was also studied with a T-cell receptor (TCR) beta gene probe as a marker for independence of in vivo-derived clones. Some clones were also studied with a TCR gamma gene probe. Ninety-four percent of wild-type and 89% of the hprt mutants were found to originate from independent in vivo precursors. Therefore, most of the recovered hprt mutants in the study were presumably derived from separate in vivo mutations. For non-mutagenized adults with normal mutant frequencies, in vivo mutant frequencies are thus reasonable approximations of in vivo mutation frequencies, although elsewhere we show that this is not necessarily true for individuals with grossly elevated mutant frequencies.

Adult↗
PubMed1986

Use of T-cell receptor gene probes to quantify the in vivo hprt mutations in human T-lymphocytes.

T-cell receptor (Ti) gene restriction fragment patterns (RFPs) were determined by Southern blots of genomic DNA obtained from T-lymphocyte colonies isolated from a single normal individual. 4 wild-type colonies and 11 in vivo derived 6-thioguanine-resistant mutant colonies with previously characterized hprt gene structural alterations were studied. Among the hprt mutants, 10 of the 11 showed unique Ti RFPs indicating their origins in different in vivo progenitors. Unique Ti RFPs were also seen among the wild-type T-cell colonies. One, however, shared its Ti RFP with a mutant. These results suggest that mutation in vivo of the hprt gene in human T-lymphocytes occurred after thymic maturation and that the 11 recovered hprt mutants probably resulted from 11 independent mutational events.

DNA↗
PubMed1986

HLA typing and autoantibodies in hepatitis B surface antigen-negative chronic active hepatitis.

HLA types, serum autoantibodies and serum globulin levels were surveyed in 46 patients with HBsAg-negative chronic active hepatitis. Patients with chronic active hepatitis with viral risk factors were less likely than those without viral risk factors to be HLA type B8 (10% vs 44%) or to have autoantibodies (antinuclear and/or anti-smooth muscle antibodies) (38% vs 84%). Thirty patients (10 with and 20 without viral risk factors) were treated with glucocorticosteroids. Of the 11 patients who were HLA-B8, 100% responded to treatment. Of the 20 patients who were ANA positive, 80% responded. The data suggest that the presence of HLA-B8 may be a useful predictor of response to anti-inflammatory treatment and may define a genetic subset of HBsAg-negative chronic active hepatitis that will benefit from glucocorticosteroid therapy.

Adult↗
PubMed1986

The HPRT mutant T-cell assays for human mutagenicity monitoring.

Both T-cell assays are useful for human mutagenicity monitoring. The autoradiographic assay is inexpensive, relatively short term and easily applicable to population studies. The clonal assay will provide information on the nature of spontaneous mutation in vivo and, by analysis of mutational spectra, may yield insights into induced mutational events. In addition to the characterizations noted above, we have recently shown that analysis of in vivo derived hprt mutant T-cell colonies with cDNA probes for the and T-cell receptor genes allows quantification of the number of in vivo hprt mutations that give rise to an observed collection of mutants. (Nicklas, et. al, 1985, Albertini et. al 1985b) Refinements of this assay should increase its applicability for population studies. The utility of both assays should be better defined by studying the relationship between their respective end-points. Both assays yield similar frequencies of TGr cells in individuals (Albertini, 1985), and may thus be equivalent. Future studies should answer this question.

Autoradiography↗
PubMed1985

Confidence intervals and sample size calculations to compare variant frequencies.

Direct mutagenicity tests offer the opportunity of monitoring human populations to detect evidence of genetic damage that occurs in vivo. As such these tests offer the potential of linking earlier exposures to mutagenic agents to subsequent health effects. One such test detects mutant T-lymphocytes that arise in vivo in human peripheral blood. Statistical analysis of the value observed, the variant frequency (Vf), is the subject of this paper. We present and illustrate procedures for finding confidence intervals for a single variant frequency and for the ratio of two independent variant frequencies. We also derive formulae for required sample size to ensure adequate power to detect a specified change in a single variant frequency or a specified difference between two variant frequencies. Our approach is to exploit the approximate normality of the natural logarithm of a Poisson distributed variable. The procedures developed appear to be quite accurate even for the small (5-15) values often observed in the variant frequency assay. Moreover, the procedures are very easy to use and should prove valuable to investigators involved in direct mutagenicity testing.

Drug Resistance↗
PubMed1983

Pure RBC aplasia and myasthenia gravis. Coexistence of two diseases associated with thymoma.

Pure RBC aplasia (PRCA) and myasthenia gravis (MG) are occasionally associated with thymoma, although the occurrence of all three in a single patient is rare. We describe a patient in whom PRCA and MG occurred in conjunction with the presence of serum autoantibodies, evidence for a serum inhibitor of erythropoiesis, and elevation of circulating thymosin alpha 1. Although a thymoma could not be detected before death, postmortem examination of the mediastinum found scattered cells within a nodule, suggestive of a lymphoepithelial thymic neoplasm.

Aged↗
PubMed1982

T-cell cloning to detect the mutant 6-thioguanine-resistant lymphocytes present in human peripheral blood.

Rare thioguanine-resistant T lymphocytes, present in vivo in human peripheral blood, were isolated and grown in vitro as thioguanine-resistant cultured T cells. The conditions for their selection in vitro were such that thioguanine resistance had to have arisen in vivo. The mutant cells bore T-cell surface markers, maintained their thioguanine resistance in vitro in the presence or absence of selection, and were deficient in hypoxanthine-guanine phosphoribosyltransferase activity.

Antigens, Surface↗
PubMed1980

Suppression by progesterone of nonspecific in vitro lymphocyte stimulation in mice as a mechanism for the enhancement of herpes simplex virus type 2 vaginal infection.

The role of antibody, interferon, and cell-mediated immunity (CMI) were studied to determine the mechanisms for progesterone enhancement of vaginal herpes simplex virus type 2 (HSV 2) infection in mice. Three groups of mice were studied: nonpregnant control, pregnant, and nonpregnant progesterone-treated mice. Vaginal infection with HSV 2 did not elicit a neutralizing antibody or a systemic interferon response in any of the groups tested. Splenic lymphocytes from noninfected and infected mice were stimulated in vitro with a nonspecific T-cell mitogen concanavalin (Con A) to measure the proliferative phase of CMI in these groups of mice. No suppression of (3H) thymidine (3HTdR) uptake was found in the pregnant or nonpregnant, progesterone-treated animals as compared to nonpregnant control mice. When progesterone was added directly to the splenic lymphocytes and continuously present during Con A stimulation a statistically significant depression of 3HTdR incorporation was found. We concluded that progesterone depresses Con A stimulation of murine lymphocytes, but progesterone must be continuously present to do so.

Animals↗