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Biomedical subjects

P W Mueller

Publications and source records attributed to P W Mueller.

At least 19 recordsLinked to original sources
PubMed2005

High-resolution genotyping of HLA-DQA1 in the GoKinD study and identification of novel alleles HLA-DQA1*040102, HLA-DQA1*0402 and HLA-DQA1*0404.

In order to achieve high-resolution HLA-DQA1 genotyping, it is necessary to identify polymorphisms in exons 1, 2 and 3. We present a high-resolution sequence-based typing (SBT) strategy for genotyping exons 1, 2 and 3 of the polymorphic HLA-DQA1 locus. This method is an improvement upon previously presented methods, because it utilizes the minimum number of SSP-PCR assays to obtain clear DNA sequence in both the forward and reverse directions of all three exons. All known HLA-DQA1 alleles are resolved with the exception of HLA-DQA1*010101 and HLA-DQA1*010102 for which the distinguishing polymorphism is located in exon 4 and does not result in an amino acid change. This method has enabled our laboratory to identify three new HLA-DQA1 alleles - HLA-DQA1*040102, HLA- DQA1*0402 and HLA-DQA1*0404 - in the Genetics of Kidneys in Diabetes (GoKinD) study population. Additionally, we present single-allele amplification methods, which identify the coding sequences of HLA-DQA1 exons 1, 2, 3, intron 2 and 300 bp of the HLA-DQA1 promoter (QAP). This study, also describes the QAP for most of the known HLA-DQA1 alleles, three HLA-DQA2 promoter sequences and the intron 2 sequences for HLA-DQA1*040101, HLA-DQA1*040102, HLA-DQA1*0402 and HLA-DQA1*0404.

Alleles↗
PubMed2001

High-resolution sequence-based typing strategy for HLA-DQA1 using SSP-PCR and subsequent genotyping analysis with novel spreadsheet program.

We present a new sequence-based typing (SBT) strategy for the polymorphic HLA-DQA1 locus that is based on sequence-specific primer - polymerase chain reaction (SSP-PCR) amplification from genomic DNA. This method allows high-resolution genotyping in the second exon of the DQA1 gene. This gene presents a unique situation in which half of the known alleles contain an inframe three base pair deletion of codon 56. This deletion confounds direct SBT methodologies of heterozygous individuals containing both a deletion and nondeletion allele. The primary HLA haplotype associated with type 1 diabetes susceptibility is DR3/DR4. The DQA1 genotype for these two haplotypes are DQA1 *0501, a non-deletion allele and *0301, a deletion allele, thus creating a situation that cannot be resolved using a direct sequencing approach. Our group-specific SBT strategy isolates the deletion alleles from the nondeletion alleles, allowing them to be resolved by direct sequencing. Additionally, we present a novel spreadsheet program that accurately assigns the genotype of both homozygous and heterozygous persons.

Alleles↗
PubMed1999

Current activities at the Centers for Disease Control and Prevention's National Diabetes Laboratory.

In 1997, the Centers for Disease Control and Prevention established the National Diabetes Laboratory in order to help prevent and treat type 1 diabetes. This state-of-the-art laboratory collaborates with research scientists and key national and international organizations throughout the world to identify and study risk factors for type 1 diabetes by developing measurements for glycosylated proteins, developing and evaluating technology for measuring genetic risk factors for the disease, and working to standardize autoantibody measurements. Developing improved technologies for diagnosing and managing diabetes and developing reference materials for properly calibrating and standardizing blood glucose meters are also critical aspects of the laboratory's work. In addition, the laboratory provides quality storage for valuable collections of biologics and other materials and facilitates sharing of specimens, associated epidemiologic data, and test results. Working with our partners in diabetes research, we are improving the diagnosis, treatment, and prevention of type 1 diabetes.

Autoantibodies↗
PubMed1998

New approaches for detecting thresholds of human nephrotoxicity using cadmium as an example.

Damage to the kidneys is one of the primary toxic actions of metals. Nephrotoxic substances not only cause renal disease directly, but they can also destroy renal reserve capacity, potentially placing those people with additional risk factors, such as diabetes, hypertension, cardiovascular disease, and genetic predispositions, at greater risk. To detect nephrotoxicity in people at a stage where intervention can be effective, sensitive methods are needed. One of the major advantages of using sensitive biomarkers of renal damage is that people who may be particularly susceptible to renal damage can be identified early, at a reversible stage of damage, and the progression to end-stage renal disease may be halted or delayed. Various categories of tests can be used to detect effects of nephrotoxic substances on the kidney. Through the use of biomarkers of damage to various parts of the nephron, U.S. and European studies have both shown a similar pattern of damage among men occupationally exposed to cadmium. These studies indicate various thresholds of renal effects, which researchers suggest represent a cascade of progressively severe damage to the kidney. Research into new biomarkers of damage caused by exposure to nephrotoxic substances centers around mechanisms of cell death, including necrosis and apoptosis; mechanisms of cell growth, regeneration, and proliferation, including factors that control cell cycle, influence gene expression, and modulate nucleic acid synthesis; and genetic factors that increase susceptibility to renal disease. Examples of types of candidate biomarkers include cytokines, lipid mediators, growth factors, transcription factors and protooncogenes, extracellular matrix components (collagen, glycoproteins, and proteoglycans), and cell adhesion molecules. Research into new categories of biomarkers may provide additional insights into the mechanisms of damage caused by nephrotoxins.

Biomarkers↗
PubMed1995

An in-depth examination of the excretion of albumin and other sensitive markers of renal damage in mild hypertension.

In an in-depth examination to better define the renal effects of mild hypertension, we used urinary proteins to indicate damage to the glomerulus (albumin), tubular reabsorption capability (retinol-binding protein), and turnover of tubular tissue (alanine aminopeptidase and N-acetyl-beta-D-glucosaminidase) in a group of 18 people with mild hypertension not associated with diabetes and a control group (n = 12). The participants' activity was controlled on a high normal salt diet for 3 days followed by a low salt diet for 4 days. Two distinct patterns of albumin excretion were evident in the hypertensive group: 22% had elevated, highly variable excretion patterns, and the rest had tightly grouped values below 16 mg/g creatinine, 16 micrograms/min, or 16 mg/L, with the lowest within-person biological variability given by albumin calculated as a ratio to creatinine. Albumin and NAG excretion primarily correlated with systolic blood pressure and the best correlations were given by ratios to creatinine. A marked decrease in salt excretion of 71% (to 50.8 mEq/day) resulted in significant (P < .0005) decreases in systolic (13.9 mm Hg), diastolic (6.4 mm Hg), and mean arterial pressures (8.9 mm Hg) only in the group with mild hypertension. However, albumin excretion did not decrease when dietary salt content was lowered. The group with hypertension also had higher urinary excretion of lysosomal N-acetyl-beta-D-glucosaminidase (P < .01), and whites in the group had a higher excretion of retinol-binding protein than did whites in the control group (P < .02). Retinol-binding protein values, however, were within the normal range, indicating that the elevated albumin values were the result of changes in selectivity of the glomerulus.

Acetylglucosaminidase↗
PubMed1992

Chronic renal effects in three studies of men and women occupationally exposed to cadmium.

We measured sensitive indicators of renal damage in three different populations occupationally exposed to cadmium, and examined the degree of variation in damage and the relative sensitivity of different types of indicators. The three studies included (1) men exposed in a cadmium recovery plant, (2) men exposed in a nickel/cadmium battery plant, and (3) women exposed in the latter plant. The indicators of renal damage were urinary proteins in three categories: (1) the high molecular weight enzymes alanine aminopeptidase (AAP) and N-acetyl-beta-D-glucosaminidase (NAG), (2) the intermediate molecular weight protein albumin (ALB), and (3) the low molecular weight proteins retinol-binding protein (RBP) and beta 2-microglobulin (B2M). These tests indicate that exposed groups with higher urine cadmium levels had varying degrees of renal damage. All exposed groups showed evidence of renal damage when compared with their respective control groups. A higher percentage of elevated protein levels was noted in the exposed group of Study 1 than in the exposed groups of Studies 2 and 3. In Study 1, the means of all five protein levels and ALB, RBP, and B2M fractional clearances were significantly elevated in the group with higher urine cadmium concentrations when compared with the groups with lower urine cadmium concentrations. Highly significant dose-response relationships for all of the urinary protein tests, including fractional clearances, were found. All of the tests were more sensitive in detecting evidence of subclinical renal damage than serum creatinine, a commonly used indicator of renal function. The order of test sensitivity in men was determined by considering three factors: (1) the magnitude of the correlation coefficient between the test and the urine cadmium concentration in the study with the most advanced damage, (2) the relative cadmium level predicted by the dose-response model at which there is a 10% chance of observing an elevated test value, and (3) the ability of the tests to detect renal effects in the population with less advanced damage. The tests in order of decreasing sensitivity in men are ALB, AAP, NAG, RBP approximately B2M. The women with higher urine cadmium levels in Study 3 had a higher percentage of elevated AAP and NAG values when compared with the control group.

Adult↗
PubMed1991

Considerations when measuring urinary albumin: precision, substances that may interfere, and conditions for sample storage.

The measurement of small but abnormal amounts of albumin in urine is important in evaluating kidney disease in people with diabetes mellitus, hypertension, or possible adverse health effects from exposure to nephrotoxins. Routine laboratory methods for measuring albumin are not sensitive enough to measure the amounts that are significant in urine (less than 30 mg/L). In our laboratory we used three immunoassays for measuring urinary albumin: enzyme-linked immunosorbent assay (EIA), radioimmunoassay (RIA), and immunoturbidimetric assay (IT). We calculated the CVs of the three methods, investigated potential interfering substances at three times their normal concentrations, and stored urine under different conditions to find the best way to protect the sample until assay. The potential interferents we checked were transferrin, urea, beta 2-microglobulin, retinol-binding protein, creatinine, kappa and lambda light chains, IgG, hemoglobin, ketone, and glucose. The stability study involved two study temperatures (-20 and -70 degrees C) and four treatments (centrifuging or filtering, before or after storage). We found the following: the RIA had the lowest CV; the results from the interference study showed no interference from normal physiological concentrations of the substances investigated; storage at -70 degrees C regardless of the treatment should be adequate to prevent loss of albumin immunoreactivity.

Albuminuria↗
PubMed1991

Interlaboratory comparison of the measurement of albumin in urine.

Because of increased interest in the assay of albumin in urine and the sensitivity required to quantify concentrations associated with (a) increased risk of developing end-stage renal disease and cardiovascular disease among people with diabetes and (b) renal damage caused by exposure to nephrotoxic substances, we conducted a pilot study of the variation of these measurements within and among five laboratories that use various immunoassays. These assays included two different enzyme immunoassays, two different immunoturbidimetric assays, a fluorescent immunoassay, and a zone immunoelectrophoresis assay. The results indicate considerable variation both within and among laboratories for measurements at or near the normal range. Variability is equally attributable to the precision of individual immunoassays and to the variation of the mean values obtained by each laboratory. Individual laboratory CVs ranged from 5.8% to 18.2% for mid- and high-concentration samples treated with preservative and from 8.4% to 23.6% for mid- and high-concentration samples containing no preservative. The relative bias of individual laboratory means ranged from -56.4% to 20.5% for the two preserved materials and from -32.6% to 0.8% for the two materials containing no preservative. To reduce the chance of misdiagnosing the risk associated with above-normal albumin concentrations in urine, we need to address the problems contributing to imprecision and inaccuracy, particularly laboratory-to-laboratory variability.

Albuminuria↗
PubMed1990

Indicators of acute renal-transplant rejection in patients treated with cyclosporine.

We evaluated the ability of three enzymes--N-acetyl-beta-D-glucosaminidase (NAG; EC 3.2.1.30), alanine aminopeptidase (AAP; microsomal aminopeptidase, EC 3.4.11.2), and gamma-glutamyltransferase (GGT; EC 2.3.2.2)--and adenosine deaminase binding protein (ABP) in urine to predict or confirm renal-transplant rejection in patients treated with cyclosporine. We measured the enzymes daily during the early post-transplant hospital stay of 104 renal-transplant recipients (72 men and 32 women). We also measured ABP in 32 of these patients. We analyzed the data by calculating the activity ratio of each day's test value to the previous day's result and optimized the sensitivity (SN) and specificity (SP) to determine the optimal ratio for each test. The results indicate that cyclosporine treatment reduces the optimal sensitivity and specificity of these tests. Three comparable tests (ABP, GGT, and AAP) yield the best optimal values (SN = 0.77, 0.69, 0.77; and SP = 0.71, 0.74, 0.63, respectively), and the NAG test yields the lowest combination of sensitivity and specificity (SN = 0.62, SP = 0.66). All four tests were less sensitive and specific than the plasma creatinine test (optimal day-to-day difference = 5 mg/L). However, the ABP and AAP tests gave indications of rejection at least 24 h before clinical diagnosis for 50% of the patients experiencing rejection, while early plasma creatinine increases of 5 mg/L occurred in only 19% of this group.

Adolescent↗
PubMed1989

Chronic renal tubular effects in relation to urine cadmium levels.

In this study the urine activities of two brush border membrane enzymes, alanine aminopeptidase (AAP) and gamma-glutamyltranspeptidase (GGT), and the lysosomal enzyme N-acetyl-beta-D-glucosaminidase (NAG) were measured in men exposed to cadmium to investigate chronic renal toxicity. The subjects consist of a group with urine cadmium levels less than 2.0 micrograms/l and a group with higher cadmium levels (urine cadmium greater than or equal to 2.0 micrograms/l) with past or present occupational exposure to cadmium. The mean NAG value corrected for creatinine in the higher cadmium group (2.95 U/g creatinine) is significantly different from the mean low cadmium group value (0.92 U/g creatinine, p = 0.0083). Likewise, AAP in the higher group (13.83 U/g creatinine) is significantly different from that of the low group (6.58 U/g creatinine; p = 0.0018). NAG and AAP also give significant correlations with cadmium levels in urine (NAG: r = 0.51; p = 0.0001; AAP: r = 0.56; p = 0.0001). There is no similar statistically significant difference in GGT means in the two groups. In contrast to cadmium correlations, blood lead does not correlate with NAG, AAP, or GGT. A dose-response relationship was found between NAG and cadmium and between AAP and cadmium. The analysis of this relationship gives estimates of a 10% chance of observing an elevated NAG value at a cadmium level of 6.3 micrograms/g creatinine (8.0 micrograms/l) and a 10% chance of observing an elevated AAP at a cadmium level of 5.0 micrograms/g creatinine (3.4 micrograms/l). These data indicate elevations of NAG and AAP at urine cadmium levels below the level of 10 micrograms/g creatinine recommended as an upper limit by the 1980 World Health Organization Study Group. Since elevated levels of NAG and AAP in cadmium-exposed workers may reflect chronic renal tubular nephrotoxicity, these findings indicate that cadmium levels below 10 micrograms/g creatinine may be accompanied by subclinical tubular dysfunction, and that WHO guidelines should be interpreted cautiously, particularly with reference to workers who are no longer exposed and may have had higher cadmium body burdens in the past.

Acetylglucosaminidase↗
PubMed1987

Alanine aminopeptidase in serum: automated optimized assay, and effects of age, sex, smoking, and alcohol consumption in a selected population.

In the diagnosis of chronic (as opposed to acute) liver diseases, combinations of indicators are needed to improve specificity. Alanine aminopeptidase (AAP; microsomal aminopeptidase, EC 3.4.11.2) activity in serum reportedly is a very sensitive indicator of intrahepatic cholestasis and biliary obstruction; it is also particularly useful in diagnosing chronic liver disease when combined with an indicator of hepatocyte damage such as aspartate aminotransferase or alanine aminotransferase. We optimized the assay of AAP in serum, automated the assay by using a centrifugal analyzer, then used this automated assay to determine activity in 202 individuals, ages one to 73 years. The preliminary results were analyzed in terms of the effects of age, sex, smoking, and alcohol consumption on AAP activity in serum. Striking sex-related differences were observed: AAP activity in males declined 2.5 times more rapidly with age than did that in females; indeed, activity in adult females remained essentially constant. Moreover, AAP values were higher in men who smoked than in those who did not, the difference being of borderline significance by analysis of covariance (p = 0.0865) but significant by partial correlations (p = 0.02). No similar differences were seen for women smokers and non-smokers. When the effects of other variables were held constant, alcohol consumption alone did not significantly correlate with AAP activity in men or women.

Adolescent↗
PubMed1986

Effects of polychlorinated biphenyls and lipemia on serum analytes.

Twelve serum analytes [triglycerides, cholesterol, total and conjugated bilirubin, high-density-lipoprotein cholesterol (HDL-C), alkaline phosphatase (AP), gammaglutamyl transferase (GGT), aspartate aminotransferase (AST), alanine aminotransferase (ALT), beta-glucuronidase (beta-glu), alanine aminopeptidase (AAP), and 5'-nucleotidase (5'nuc)] were measured to investigate their correlation with exposure to polychlorinated biphenyls (PCBs) and 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (DDT). The relationship between serum lipids, lipophilic toxicants, and the analytes was also evaluated. The beta-glu, 5'nuc, triglycerides, cholesterol, and total bilirubin correlated positively and significantly with log concentrations of serum total PCBs and 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE), a metabolite of DDT. The more highly chlorinated PCBs (Aroclor 1260) had significant, positive correlations with several serum analytes, but the less chlorinated PCBs (Aroclor 1242) correlated significantly and negatively only with HDL-cholesterol. Triglyceride- and cholesterol-rich lipoproteins were added to serum to determine the effects of lipids on these assays. Several were spuriously elevated. AP and beta-glu were not affected by lipoprotein addition with the methods used in this study. AAP was increased significantly only at triglyceride concentrations exceeding 400 mg/dl. Lipoproteins may be elevated because of deranged lipid metabolism in response to PCBs, or PCBs may be elevated because elevated lipoproteins are present, as in familial triglyceridemia, a relatively common dyslipoproteinemia. Because this relationship is not well understood with respect to cause and effect, we propose the further use in epidemiological investigations of assay methods that are little affected by blood lipids yet are correlated with PCB concentrations. Congener-specific quantification of PCBs would help elucidate the effects of PCBs on assays used to monitor health effects.

5'-Nucleotidase↗
PubMed1983

Dopamine-B-hydroxylase: suicide inhibition by the novel olefinic substrate, 1-phenyl-1-aminomethylethene.

Dopamine-B-hydroxylase [E.C.1.14.17.1] plays a key role in the biosynthetic interconversion of neurotransmitters. It is now demonstrated for the first time that dopamine-B-hydroxylase also catalyzes the oxygenation of an olefinic substrate, 1-phenyl-1-aminomethylethene, producing 2,3-dihydroxy-2-phenylpropylamine after acid workup. This reaction gives the normal oxygenase stoichiometry of electrons to O2 to product of 2:1:1, and is kinetically comparable to other oxygenase activities of dopamine-B-hydroxylase, with a kcat value of 10 sec-1 and a KM of 8.3 mM. 1-Phenyl-1-aminomethylethene is also a time-dependent, first-order inactivator of dopamine-B-hydroxylase. The inactivation process exhibits the characteristics of mechanism-based, irreversible inactivation, giving a KI value of 13 mM and a kinac of 0.04 min-1. The central role of dopamine-B-hydroxylase in catecholamine metabolism suggests possible pharmacological uses for olefinic inhibitors of this enzyme.

Adrenal Glands↗