Plasma acidic glycohydrolases in insulin-dependent diabetes mellitus

Summary We assayed plasma activities of -galactosidase, -hexosaminidase, -mannosidase, -fucosidase and -galactosidase involved in degradation of the glycoprotein molecule in 110 insulin-dependent diabetics aged 3-1/2 to 19 years and compared them to a group of normal youngsters. We correlated the plasma enzyme activities with the duration, control and sequelae of insulin-dependent diabetes. Insulin-dependent diabetics had a significantly higher plasma activity of -hexosaminidase and -mannosidase (p<0.01) and a significantly lower plasma activity of -fucosidase and -galactosidase (p<0.01). Of the 5 enzymes studied, only plasma -hexosaminidase correlated with fasting and postprandial blood sugar (p<0.01), cholesterol and triglycerides (p<0.05). Additionally, poor control of diabetes was also associated with a significantly higher plasma -hexosaminidase activity (p<0.01). Proteinuria or an abnormal Addis count suggestive of renal involvement was associated with various changes in plasma acidic hydrolases. These changes may be related to insulin deficiency rather than hyperglycemia and may be genetically determined.Deceased on August 2, 1981.     

Prevention of diabetic glomerulopathy in streptozotocin diabetic rats by insulin treatment

Summary Glomerular basement membrane thickness (GBMT) has been measured in streptozotocin diabetic rats treated with insulin. The study included 3 groups of 8 rats each: 1) a well-controlled group of diabetic rats under insulin treatment with a plasma glucose level reasonable close to normal values, 2) a poorly-controlled group also under insulin treatment with constant high plasma glucose values, and 3) an age and weight matched non-diabetic control group. After 6 months of diabetes, GBMT was measured applying an intercept method on 3 glomerular cross sections from each of the 24 animals. The measurements showed that mean GBMT was 132.2. nm in the non-diabetic control rats and 131.6 nm in the well-controlled diabetic rats. In the poorly-control-led group the mean GBMT was 140.4 nm, i.e. statistically significantly increased when compared to each of the two other groups, 2p=0.022 and 0.012 respectively.The results demonstrate that good blood glucose control in rats preserves normal GBMT.     

Mutation of E1α gene in a female patient with pyruvate dehydrogenase deficiency due to rapid degradation of E1 protein

Summary A mutation of an insertion of 4 bp in the gene for the subunit of pyruvate dehydrogenase (E1) was found in a female with pyruvate dehydrogenase deficiency due to the rapid degradation of and subunit proteins of pyruvate dehydrogenase. This mutation caused a frameshift that altered the amino acid sequence and created a premature stop codon. This 4-bp insertion has been found in an unrelated female patient with E1 deficiency. It is rare that the same mutation is found in unrelated patients with this rare inborn error of metabolism. Furthermore, short deletions or duplications in the E1 gene of patients with E1 deficiency have been found only in exons 10 and 11. These exons may be hot spots for the mutations by the recombinational processes. This patient was heterozygous for the normal and a mutant allele. However, in most of the cultured skin fibroblasts from this patient, the mutant allele was expressed. These observations suggest that the X chromosome containing the normal allele was predominantly inactivated so that she developed lactic acidaemia and neurological abnormalities despite being heterozygous. The mutant subunit protein failed to form a stable structure of pyruvate dehydrogenase, so that both and subunit proteins were degraded rapidly.     

Elliptocytosis associated with an abnormal α glycophorin

Summary A case of elliptocytosis associated with an undescribed abnormal glycophorin (GP) is reported. Using immunoblotting techniques, a clear-cut minor band 6 was detected emerging just behind the monomer of GP (band 6) when probed with anti- GP antiserum. It also reacted with anti-peptide C antiserum, suggesting that this new band with a molecular weight of 24 K is related to the structural alteration of GP and not GP. The erythrocyte membrane proteins of the patient exhibited a quite normal pattern, with a normal spectrin/ spectrin ratio, but the reaction with anti-protein 4·1 serum confirmed the increase in proteolytic susceptibility of her protein 4·1. The results of DNA mapping implied that the abnormality may be due to a short deletion of the heterozygote. The significance of deviation involving the GP and protein 4·1 to the elliptocytic change of erythrocyte shape is briefly discussed.This work was supported by grant no. HL-21016 from the National Institutes of Health, Bethesda, MD     

Studies on the oxidation of phytanic acid and pristanic acid in human fibroblasts by acylcarnitine analysis

The -oxidation of phytanic acid and the -oxidation of pristanitc acid were investigated in cultured fibroblasts from controls and patients affected with different peroxisomal disorders using deuterated substrates. Formation of [-2H6]4,8-dimethylnonanoylcarnitine ([-2H6]C11-carnitine) from [-2 H6]phytanic acid and [-2H6]pristanic acid was used as marker for these processes. Analysis was performed by tandem mass spectrometry.In normal cells, formation of [-2H6]C11-carnitine from both [-2H6]phytanic acid and [-2H 6]pristanic acid was observed. When peroxisome-deficient fibroblasts were incubated with these substrates, [-2H6]C11-carnitine was not detectable or, in two cases, very low, which results from deficiencies in both peroxisomal - and -oxidation. In cells with an isolated -oxidation defect at the level of the peroxisomal bifunctional protein, formation of [-2H6]C11-carnitine could also not be detected.Cells with an isolated defect in the -oxidation of phytanic acid, obtained from patients affected with Refsum disease (McKusick 266500) or rhizomelic chondrodysplasia punctata (McKusick 215100), did not form [-2H6]C11 -carnitine from [-2H6]phytanic acid. The observed formation of [-2H6]C11-carnitine from [-2H6]pristanic acid in these cells is in accordance with a normal peroxisomal -oxidation in these disorders.This study shows that separate incubation of fibroblasts with [-2H 6]phytanic acid and [-2H6]pristanic acid, followed by acylcarnitine analysis in the medium by tandem mass spectrometry, can be used for screening cell lines for deficiencies in the peroxisomal - and -oxidation pathways.     

Schindler disease: An inherited neuroaxonal dystrophy due to α-N-acetylgalactosaminidase deficiency

Summary The clinical, pathological and biochemical features of a neuroaxonal dystrophy resulting from the deficient activity of lysosomal -N-acetylgalactosaminidase are described. This neurodegenerative disorder was recognized in two brothers who had the typical clinical manifestations and neuropathological lesions observed in patients with Seitelberger disease, the infantile form of neuroaxonal dystrophy. Axonal spheroids were observed histologically in the grey matter, and ultrastructural examination revealed the characteristic formations in dystrophic axons in the myenteric plexus and neocortex. Using a newly synthesized fluorogenic substrate, 4-methylumbelliferyl--N-acetylgalactosaminide, the markedly deficient activity of-N-acetylgalactosaminidase was demonstrated in the affected brothers while their consanguineous parents had intermediate activities, consistent with the autosomal recessive transmission of this disease. No detectable-N-acetylgalactosaminidase was seen in immunoblots using monospecific rabbit antihuman-N-acetylgalactosaminidase antibodies. Abnormally increased amounts of urinary glycopeptides were observed by high resolution thin layer chromatography. Analytical studies identified four of the accumulating urinary compounds, the blood group A trisaccharide GalNAc1 3(Fuc1 2)Gal and threeO-linked glycopeptides, GalNAc1 O-serine and -threonine, NeuNAc2 3Gal1 3(NeuNAc2 6)GalNAc1 O-serine and -threonine, and NeuNAc2 3Gal1 4GlcNAc1 6(NeuNAc2 3Gal1 3)GalNAc1 O-serine and -threonine. Of eight unrelated patients diagnosed as having infantile neuraxonal dystrophy by pathological studies, none had deficient-N-acetylgalactosaminidase activity, emphasizing the biochemical heterogeneity underlying this diagnostic entity. These findings document the first delineation of a metabolic defect in an inherited neuroaxonal dystrophy and suggest that the axonal pathology in this disorder, and perhaps in the other neuroaxonal dystrophies, results from abnormal glycoprotein metabolism involvingO-linked glycopeptides.     

Rapid detection of −α 4.2 deletion of α-thalassemia-2 by polymerase chain reaction

Summary We sequenced part of the X boxes of-thalassemia-1 of Southeast Asia type (- -SEA) with– ^4.2,– ^3.7,– ^G-Taichung, and ^CS. We found the X box of– ^3.7 belonged to the X box of 2 globin gene and the X box of ^cs contained X boxes of both al and2 globin gene, whereas the X box of– ^4.2 and– ^G-Taichung was a hybrid of X boxes of 2 and 1 globin gene. We also found there are two types of– ^4.2 deletion; type 1 is a common type of– ^4.2 deletion and type 2 is linkage to– ^G-Taichung. We used a combination of two methods, the amplification refractory mutation system (ARMS) and the amplified created restriction sites (ACRS), to amplify the hybrids of X boxes specifically. The upstream primer for X box of2 globin gene was designed following the standard ARMS procedure to amplify the X segment of the-globin gene. The downstream primer was designed according to the ACRS method to check the specificity of PCR products. Using this approach, we can diagnose the different types of– ^4.2 deletion. This kind of approach can also be used to amplify the specific region from the cluster of highly homologous genes.     

Thalassemia intermedia: compound heterozygous β∘/β+-thalassemia and co-inherited heterozygous α+-thalassemia

Summary The relative excess of - over -globin chains in the erythroid precursors is the chief pathophysiological factor of homozygous -thalassemia. The clinical picture is usually characterized by a transfusion-dependent dyserythropoietic anemia (thalassemia major). However, some patients present with moderate anemia that does not require regular blood transfusions (thalassemia intermedia). The molecular heterogeneity of -thalassemia mutations and changes of - and -globin gene expression play an important role in modifying the clinical phenotype. We report here on a female Greek patient with homozygous -thalassemia but normal growth and development, excellent exercise tolerance, and no need of blood transfusions. She is thus mildly affected clinically, although there is marked pallor, jaundice, and hepatosplenomegaly. These signs correspond to her marked hypochromic, microcytic anemia with erythroid hyperplasia of the bone marrow. -Globin genotyping shows her to be compound heterozygous for the codon 39 C T -nonsense mutation and for the T C ⁺-mutation at position 6 of the splice consensus at the exon 1/intron 1 junction (CD39 C T/IVS 1–6 T C). -Globin gene mapping demonstrates the presence of a 3.7-kb ⁺-thalassemia deletion on one allele (–^3.7/). Taken together, this study identifies a complex interaction of genetic factors that do not significantly alter the clinical phenotype when present alone but ameliorate the course of homozygous -thalassemia when inherited in combination.Abbreviations Hb hemoglobin - Hct hematocrit - HPFH hereditary persistence of fetal hemoglobin - IVS intervening sequence - MCH mean corpuscular hemoglobin - MCV mean corpuscular volume - PCR polymerase chain reaction     

Isolierung von freien Nucleotiden aus verschiedenen Geweben

Zusammenfassung Es wurden die freien Nucleotide aus dem säurelöslichen Extrakt des Sarkom 37 (Ascitesform) durch Anionenaustauschchromatographie isoliert. Die Befunde werden mit den Ergebnissen an normalen und anderen Tumorgeweben verglichen.In der Arbeit werden folgende Abkürzungen verwendet Ad Adenosin - A2MP Adenosin-2-monophosphat - A3MP Adenosin-3-monophosphat - AMP Adenosin-5-monophosphat - ADP Adenosin-5-monophosphat - ATP Adenosin-5-triphosphat - G2MP Guasonin-2-monophosphat - G3MP Guanosin-3-monophosphat - GMP Guanosin-5-monophosphat - GDP Guanosin-5-diphosphat - GTP Guanosin-5-triphosphat - C2MP Cytidin-2-monophosphat - C3MP Cytidin-3-monophosphat - CMP Cytidin-5-monophosphat - CDP Cytidin-5-diphosphat - CTP Cytidin-5-triphosphat - U2MP Uridin-2-monophosphat - U3MP Uridin-3-monophosphat - UMP Uridin-5-monophosphat - UDP Uridin-5-diphosphat - UTP Uridin-5-triphosphat - UDPA Uridin-5-diphosphat-N-acetylglucosamin - UDPG Uridin-5-diphosphat-glucose - UDPGa Uridin-5-diphosphatgalaktose - UDPGl Uridin-5-diphosphatglucuronsäure - TPN Triphosphopyridinnucleotid - DPN Diphosphopyridinnucleotid - FAD Flavin-adenin-dinucleotid - HS Harnsäure - RNS Ribonucleinsäure - IMP Inosin-5-monophosphat Mit 6 Textabbildungen.Den soliden Tumor verdanken wir Herrn Dr. Dr.Ch. Hackmann, Farbenfabriken Bayer, Wuppertal-Elberfeld. Dieser Tumor wurde von Herrn Dr.H. Wrba im hiesigen Institut in die Ascitesform übergeführt.     

Inhibition of Parietal Cell Acid Secretion Is Mediated by the Classical Epidermal Growth Factor Receptor

Epidermal growth factor (EGF) and transforminggrowth factor- (TGF-) inhibit gastric acidsecretion both in vivo and in vitro. Previous studieshave indicated that EGF and TGF- bind to the same EGF/TGF- receptor. Nevertheless, weand others have previously demonstrated that inhibitionof acid secretion by these growth factors requiresconcentrations of the peptides that are 10-fold higher than those necessary for induction ofmitogenesis. Therefore, we have sought to investigatewhether gastric parietal cells may possess a secondEGF/TGF- receptor class. Two systems werestudied: First, [¹²⁵I]TGF- was cross-linkedto the receptor in isolated rabbit parietal cellmembranes, and labeled species were resolved onSDS-PAGE. Second, acid secretion was evaluated inpylorus-ligated waved-2 mutant mice, which carry a disabling pointmutation in their classical EGF/TGF- receptor. Inisolated parietal cells, [¹²⁵I]TGF-was cross-linked into a single species of 170 kDa.Cross-linking was inhibited in the presence of unlabeledTGF- with an IC of 80 nM. In the pylorus-ligatedmice, control littermate mice demonstrated adose-dependent inhibition of acid secretion by EGF withan IC of 20 g/kg. In contrast, EGF had noinhibitory effect on acid secretion in 50 waved-2 miceat concentrations up to 100 g/kg. No alterations inparietal cell or gastrin cell numbers were observed.These results in both isolated rabbit parietal cellsand waved-2 mice support the existence of only a singleclass of EGF/TGF- receptors in parietal cells.Differences in growth factor affinity are likely due to the modification of the receptor or oneof its coordinate regulators.     

Diabetic retinopathy and visual disability

Summary Data from blind and partial sight registers and from special surveys have been analysed to derive estimates of the number of adults in England and Wales visually disabled by diabetic retinopathy. If visual disability is defined as less than 6/18 Snellen, approximately 30 per 100.000 adults living at home are disabled by diabetic retinopathy; if defined around traditional concepts of blindness (approximately 3/60 vision or less) the estimated figure is about 20 per 100.000 total population. Annual additions to the blind register for diabetic retinopathy are about two per 100.000 total population. Weaknesses in data gathering are discussed.     

Mutations in the R-type pyruvate kinase gene and altered enzyme kinetic properties in patients with hemolytic anemia due to pyruvate kinase deficiency

Summary The biochemical properties of erythrocyte pyruvate kinase (PK) together with mutations found in the coding sequence of the R-PK gene in five patients with severe hemolytic anemia due to PK deficiency are described. The enzyme variants were designated PK Mosul (homozygote), PK Bukarest_1,2, PK Hamburg₁, PK Köln₁, and PK Essen (compound heterozygote). PK Mosul showed normal positive cooperative substrate binding, PK Bukarest_1,2 exhibited noncooperative behavior, and PK Hamburg₁ and PK Köln₁ displayed mixed cooperativity, whereas PK Essen was negative cooperative. PK Mosul was found to be homozygous for the mutation 1151 ACG to ATG, resulting in an amino acid substitution 384 Thr to Met. In one allele of PK Bukarest_1,2 a single nucleotide substitution GAG-TAG was found at nucleotide 721, causing a change of 241 Glu to a chain termination codon (PK Bukarest₁). Additionally, in the second allele of this patient a point mutation at position 1594 (CGG-TGG) occurs, changing 532 Arg to Trp (PK Bukarest₂). Direct sequencing showed the heterozygosity of the patient's mother (PK Bukarest₁/normal) at position 721 and of the patient's father (PK Bukarest₂ /normal) at position 1594. A point mutation at position 1529 (CGA-CAA), causing an amino acid substitution 510 Arg-Gln, was identified in PK Hamburg₁ and PK Köln₁. The second mutation in these variants was not detected. In PK Essen no mutation in the coding sequence was found at all. Screening for the mutation at position 1529 in further compound heterozygote patients and in normal subjects of Western European origin showed that this exchange is a common mutation responsible for PK deficiency in this population.Supported by theDeutsche Forschungsgemeinschaft, Grants no. La 527/1 and Ne 416/1.     

Enzymatic determination of serum 3α-sulfated bile acids concentration with bile acid 3α-sulfate sulfohydrolase

A newly developed enzymatic method for determining urinary 3-sulfated bile acids was used to measure serum 3-sulfated bile acid levels in 114 patients with hepatobiliary diseases and 56 healthy subjects. The lowest measurable amount of the 3-sulfated bile acids was 0.5 µmol/liter. The standard curves for glycolithocholic acid 3-sulfate, glycoursodeoxycholic acid 3-sulfate, and lithocholic acid 3-sulfate were linear from 0.5 to 250 µmol/liter. Specificity of the assay was satisfactory and intra- and interassay variations ranged from 0.8 to 4.4% and from 1.2 to 7.9%, respectively. Analytical recovery was more than 91%. The values obtained by this assay were well correlated with those by gas-liquid chromatography measurement (r=0.91,P<0.01). The fasting serum 3-sulfated bile acids level in healthy subjects ranged from undetectable to 1.9 µmol/liter (mean±se; 0.9±0.1 µmol/liter). The percentage of 3-sulfated bile acids in total bile acids (sum of 3-sulfated and 3-hydroxy bile acids) in serum was 16.8±1.5%. In subjects with hepatobiliary diseases, serum 3-sulfated bile acids levels were elevated; however, the percentage of 3-sulfated bile acids in total bile acids was decreased and correlated with the severity of hepatocellular insufficiency. This enzymatic assay is simple, rapid, and accurate for the determination of serum 3-sulfated bile acids.     

Effect of Interferon-α Treatment of Chronic Hepatitis C on Health-Related Quality of Life

Studies of interferon- (IFN-)therapy for chronic hepatitis C have focused on viralclearance; however, few have evaluated patient'shealth-related quality of life during therapy. Thisstudy evaluates health-related quality of life and theprevalence of anxiety and depression in patients withchronic hepatitis C before, during, and followingIFN- therapy. Patients undergoing IFN-therapy for chronic hepatitis C were asked to completehealth status measures as well as anxiety and depressioninventories before, during, and following IFN-therapy. These measures were compared to the results of healthy adults in the general US population.Thirty-eight of forty-eight eligible patients (79%) withchronic hepatitis C completed the questionnaires.Respondents demonstrated a significant increase in depression during the sixth month ofinterferon therapy in comparison to pretreatmentresults. Anxiety scores improved significantly after onemonth of IFN- in comparison to pretreatmentresults. Scores on the health status measures did notvary with IFN- therapy. Patient responses wereanalyzed with respect to biochemical response(normalized transaminases) to IFN-. IFN-responders, who were aware of their transaminase results,exhibited lower scores on anxiety subscales during andafter therapy (P = 0.02-0.04). Scores on the healthstatus subscale, role emotional, improved in IFN- responders compared to nonresponders during thesixth month of therapy (P = 0.02). Response toIFN- therapy was not associated with any otherdifferences on subscale analysis. Patients with chronichepatitis C exhibited health perceptions similar to thegeneral US population, and these were unchanged duringIFN- therapy. However, the incidence ofdepression significantly increased during the sixthmonth of IFN- therapy. IFN- respondersexhibited fewer emotional problems as well as a lowerincidence of anxiety during and followingtherapy.     

Uptake and metabolism of radiolabelled GM1-ganglioside in skin fibroblasts from controls and patients with GM1-gangliosidosis

Summary The uptake and metabolism of [3-³H-sphingosine]GM1-ganglioside was measured in cultured skin fibroblasts from controls and patients with infantile, juvenile and adult GM1-gangliosidosis. When dissolved in medium with phosphatidylserine, GM1-ganglioside was efficiently taken up by cultured skin fibroblasts and transferred into lysosomes. A linear increase in GM1-ganglioside endocytosis was shown with phosphatidylserine concentrations of up to 40m/ml. A pulse-chase study revealed that [³H]GM1-ganglioside was metabolized to GM2-ganglioside, GM3-ganglioside, ceramide dihexoside, ceramide monohexoside, ceramide and sphingosine. Sphingosine was recycled to sphingomyelin. In a 20-h pulse study, cell lines from patients with GM1-gangliosidosis of infantile, juvenile and adult types hydrolysed 2–5%, 20–44% and 54–58% of the total endocytosed GM1-ganglioside respectively. These values were lower than in control cells (64.17 ± 5.43% (n=10)). The hydrolysis rates of exogenous [³H]GM1-ganglioside in cultured fibroblasts from patients with various types of GM1-gangliosidosis closely reflected the clinical severity.Abbreviations GM1 Gal13GalNAc14(NeuAc23)Gal14Galc-1-ceramide - GM2 GalNAc14(NeuAc23)Gal14Glc-1-ceramide - GM3 NeuAc23Gal14Glc-1-ceramide - CDH Gal14Glc-1-ceramide     

Effects of short-term high dose intake of evening primrose oil on plasma and cellular fatty acid compositions, α-tocopherol levels,and erythropoiesis in normal and Type 1 (insulin-dependent) diabetic men

Summary In addition to their usual diet, nine Type 1 (insulin-dependent) diabetic men and ten male control subjects took 20 g d,ga-tocopheryl acetate enriched evening primrose oil (14.45 g 182c,6, 1.73g 183c,6, 400 mg d,-tocopheryl acetate) daily for one week. At start, diabetic patients had more 140, 150 and 18 2c,6, and less 160, 161c,7, 181c,7, 183c,6, 203c,9, 203c,6, 204c,6 and 226c,3 in plasma, erythrocytes and/or platelets. Furthermore, they had lower 161c,7/160, 181c,7/160, and 204c,6/203c,6 ratios and a higher 203c,6/183c,6 ratio. In diabetic patients, -tocopherol levels in erythrocytes were lower, whereas those in plasma were normal. In both groups, oil intake changed fatty acid profiles. Most markedly, 203c,6 increased, whereas the ratios 203c,6/ 183c,6 and 204c,6/203c,6 decreased. 204c,6 increased in control subjects, but not in diabetic patients. Erythrocytes and platelets responded differently in their fatty acid profiles, -tocopherol rose in plasma and, although less for diabetic patients, in erythrocytes. In diabetic patients as well as in control subjects, erythrocyte count, haemoglobin level, mean corpuscular haemoglobin content and concentration increased and glycosylated haemoglobin percentage decreased without an apparent decline in blood glucose levels. Plasma -thromboglobulin and platelet factor 4 decreased, especially in diabetic patients. In conclusion, diabetic patients had abnormal fatty acid patterns, suggesting an impaired 9, 6 and 5 desaturation and an enhanced chainelongation, and had lower erythrocyte a-tocopherol levels; and short-term high dose intake of evening primrose oil increased 203c,6 in both groups, but 204c,6 only in control subjects, gave fatty acid responses which were different for erythrocytes and platelets, enhanced erythropoiesis, and lowered indices of in vivo platelet activation.     

Inactivation of HIV in plasma derivatives by β-propiolactone and UV irradiation

Summary The inactivation of HIV in human plasma and plasma derivatives by combined treatment with -propiolactone and UV-irradiation was investigated. -propiolactone inactivated 3.5 log₁₀ and UV 2.8 log₁₀ HIV in plasma and -propiolactone 3.5 log₁₀ in cryoprecipitate and UV irradiation 4.5 log₁₀ in factor VIII concentrate.

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Inaktivierung von HIV in Plasmaderivaten durch -Propiolacton in Kombination mit UV-Bestrahlung

Zusammenfassung Untersucht wurde die Inaktivierung von HIV in humanem Plasma und Plasmaderivaten durch die kombinierte Behandlung mit -Propiolacton und UV-Bestrahlung. HIV wurde durch -Propiolacton um 3,5 log₁₀ und UV um 2,8 log₁₀ in Plasma und durch -Propiolacton um 3,5 log₁₀ in Kryopräzipitat bzw. durch UV um 4,5 log₁₀ in Faktor VIII-Konzentrat inaktiviert.

     

The effects of parabiosis on serum and kidney glycosidase activities in spontaneously diabetic mice

Summary Spontaneously diabetic non-obese mice of the ICR strain were newly inbred in Shionogi laboratory, Japan. Animals became diabetic suddenly, more frequently and severely in females. Blood glucose levels were 452±73 mg/100 ml with serum insulin levels of < 1.0 U/ml in the fed state. Parabiosis with normal control ICR mice for 2 weeks decreased the blood glucose level to 260±51 mg/ 100ml (P<0.01) and resulted in serum insulin levels of 46.0±18.0 U/ml (P<0.01). Kidney homogenate -N-acetylglucosaminidase and -galactosidase activities were reduced in diabetic mice (42% and 44% decrease respectively) (P<0.025 and P<0.001), and restored almost to normal after 2 weeks of parabiosis. Renal -mannosidase activity was decreased 43% (P<0.001) in the diabetic mice but unaffected by parabiosis. Serum -N-acetylglucosaminidase, -galactosidase and -glucosidase activities were significantly increased in diabetic mice (179%; 233% and 58% increase respectively) (P<0.005, P<0.001 and P<0.001), and returned to normal with parabiosis.     

Analytical Performance of the New Coagulation Monitoring System INRatio™ for the Determination of INR Compared with the Coagulation Monitor Coaguchek® S and an Established Laboratory Method

An evaluation of the INRatio Prothrombin Time Monitoring system for determination of INR was done in two centers with a total of 5 healthy subjects and 77 subjects on oral anticoagulation. The INRatio and the Coaguchek® S were compared with an established laboratory method. The correlation coefficient of the comparison with the laboratory was r = 0.954 for INRatio and r = 0.937 for Coaguchek® S. The mean relative deviation from the lab method calculated according to Hill was 6.87% for INRatio, which is rated very goo, and 9.72% for Coaguchek® S (goo). The imprecision in the normal range (INR = 1.1) showed a coefficient of variation (CV) of 7.8% and a standard deviation (SD) of 0.09. In the therapeutic range (INR 3.9) the CV was 5.4%, the SD 0.21 and above therapeutic range (INR 5.3), the CV was 8.4% (SD 0.44), rated satisfactory. The concordances of the compared methods with the routine were 81% for INRatio and 79% for Coaguchek® S, which is considered state-of-the-art. Most of the patients perceptions of the INRatio were very positive.In the hands of professionals the INRatio demonstrated very good accuracy and precision and an excellent technical reliability. Further studies using INRatio for self testing by patients are warranted.     

Detection of clonality by polymerase chain reaction in childhood B-lineage acute lymphoblastic leukemia

Summary DNA-based PCR with various sets of primers for TCR /, and Ig heavy chain (IgH) genes were used to study clonality in childhood B-lineage acute lymphoblastic leukemia. Amplification of the IgH CDR-III was observed in 75 of 120 analyzed cases (62.5%). From all analyzed groups, the IgH gene rearrangement was most often observed in pre-B ALL (85.7%) and was rather rare in null-ALL (34.5%). TCR delta gene rearrangement was the most common, and was observed in 77 patients (64.2%). The typical pattern of rearrangements was defined as anincomplete V2 to D3, V2 to D2, or D3 to D3 to D2 recombination product. Rearrangements of TCR gamma gene we observed in 61 cases (50.8%). TCR gamma gene rearrangements were detected predominantly in null-ALL and early B-ALL (55.2% and 60%, respectively) and were rather rare in other groups. Of all eight V segments of VI group, the most frequent gene usage concerns regions V2, V4, and V7. We have confirmed that IgH gene amplification, together with TCR gamma and delta gene amplification, provides a rapid, sensitive approach to assessing clonality in ALL almost in 100% of cases.This work was financed by KBN grants 4.0551.91.01 and 6.6346.92.03