Hypomagnesaemia in Type 2 (non-insulin-dependent) diabetes mellitus is not corrected by improvement of long-term metabolic control

Summary Low levels of magnesium have frequently been reported in diabetes mellitus especially in poorly controlled Type 1 (insulin-dependent) diabetic patients. Furthermore hypomagnesaemia might contribute to insulin resistance in Type 2 (non-insulin-dependent) diabetes. As the influence of improved metabolic control on plasma magnesium levels is unknown in Type 2 diabetic patients we studied magnesium plasma levels in 50 patients 1) before, 2) one and 3) three months after the initiation of insulin therapy or intensified treatment with oral hypoglycaemic agents. Magnesium plasma levels were measured by a colorimetric method and were significantly reduced in diabetic patients compared to healthy control subjects (0.79±0.01 mmol/l vs 0.88±0.01 mmol/l; p<0.0001). Metabolic control was significantly improved as documented by reduced HbA_1C levels in both insulin-treated patients or the patients on oral hypoglycaemic agents (p<0.003). However, plasma magnesium levels remained unchanged during the follow-up in the insulin-treated group (10.79±0.02 mmol/l; 20.81±0.02 mmol/l; 30.79±0.01 mmol/l) as well as in the patients on oral hypoglycaemic agents (10.79±0.03 mmol/l; 20.78±0.02 mmol/ l; 30.84±0.04 mmol/l). This study shows that even marked improvement of glycaemic control does not correct hypomagnesaemia in Type 2 diabetes. We conclude that hypomagnesaemia might be related to the insulin-resistant state and that possible beneficial effect of chronic magnesium administration should be evaluated in these patients.     

Pharmacokinetics of intravenous glibenclamide investigated by a high performance liquid Chromatographic assay

Summary The relationship between blood glucose and glycosylated haemoglobin (HbA_1c) has been investigated during an 8 week period in 53 Type 1 (insulin-dependent) diabetic women studied during the third trimester of pregnancy. Blood glucose estimations (fasting and 2 h post-prandially) were made an average of 41 times in each patient during this period and HbA_1c was determined once at the end of the study. There was a significant correlation between both the mean blood glucose over the preceding 8 weeks and the standard deviation of the fasting blood glucose with HbA_1c (r=0.69, p<0.001; r=0.46, p<0.001, respectively). A glycosylation index was calculated for each patient (HbA_1c divided by the mean blood glucose value). There was a significant correlation between the glycosylation index and duration of diabetes (r=0.68, p<0.001). In contrast, there was no correlation between red cell 2,3-diphosphoglycerate and HbA_1c or glycosylation index. These findings suggest that increasing duration of diabetes influences the post-translational formation of HbA_1c and that isolated HbA_1c values need to be interpreted with caution in the pregnant diabetic.     

Gewinnung von Granulozyten zur Transfusion mit dem IBM-Blutzellseparator von gesunden Spendern

Zusammenfassung Durch Variation der mit dem IBM-Zellseparator zur Gewinnung von Leukozyten gebräuchlichen Methoden konnte gezeigt werden, daß die Antikoagulierung mit einer Kombination von Heparin und ACD, die Verwendung von Neoplasmagel^® sowie die Reduzierung des extrakorporalen Blutvolumens die besten Voraussetzungen zur Gewinnung von Granulozyten sind. So konnten von einem gesunden Spender in 6 h im Mittel (n=5) 1,6x10¹⁰ Granulozyten isoliert werden bei einem Reinheitsgrad von Granulozyten: Lymphozyten: Thrombozyten: Erythrozyten=11,211151. Die Ergebnisse konnten darüber hinaus durch Erhöhung der Granulozytenausgangszahl beim Spender mit Prednisolon gesteigert werden auf eine mittlere Ausbeute (n=5) von 5,6x10¹⁰ bei einem Reinheitsgrad von Granulozyten: Lymphozyten: Thrombozyten: Erythrozyten=10,23,240. Die Viabilität der isolierten Granulozyten wurde geprüft durch Untersuchung ihres Verhaltens nach Markierung mit DF³²P und autologer Transfusion. Gegenüber nicht isolierten, im Vollblut markierten Granulozyten zeigte sich beiobne Prednisolon isolierten Granulozyten eine Verminderung, beimit Prednisolon isolierten Granulozyten eine Erhöhung der Transfusionseffektivität.

---

Summary By varying the common methods for separating leukocytes with the IBM cell separator the anticoagulation with a combination of Heparin and ACD, the use of Neoplasmagel^® as well as the reduction of the extracorporal blood volume have shown to be the best conditions to separate granulocytes. From 5 healthy donors a mean yield of 1.6x10¹⁰ granulocytes could be obtained during six h with a degree of purity of granulocytes: lymphocytes: thrombocytes: erythrocytes=11.211151. These results could be improved by increasing the count of granulocytes in the donor's blood with prednisolone in another 5 donors to a mean yield of 5.6x10¹⁰ with a degree of purity of granulocytes: lymphocytes: thrombocytes: erythrocytes=10.23.240. The viability of separated granulocytes was proved by investigating their circulation after labeling with³²P and autologous transfusion. Comparing with granulocytes which were not separated but labeled in fresh whole blood the effectivity of transfusion was diminished when separated granulocytes were transfused without stimulation with prednisolone and was increased when separated granulocytes were transfused after stimulation with prednisolone.

---

---

Studie im Rahmen des Assoziationsvertrages EURATOM-GSF No. 031-64-I BIAD und des Sonderforschungsbereiches 37 der Universität München.     

Weakened cellular scavenging activity against oxidative stress in diabetes mellitus: regulation of glutathione synthesis and efflux

Summary Glutathione functions to scavenge oxidants or xenobiotics by covalently binding them and transporting the resulting metabolites through an adenosine 5-triphosphate-dependent transport system. It has been reported that the intracellular concentration of glutathione decreases in diabetes mellitus. In order to elucidate the physiological significance and the regulation of anti-oxidants in diabetic patients, changes in the activity of the glutathione-synthesizing enzyme, -glutamylcysteine synthetase, and transport of thiol [S-(2,4-dinitrophenyl)glutathione] were studied in erythrocytes from patients with non-insulin-dependent diabetes and K562 cells cultured with 27 mmol/l glucose for 7 days. The activity of -glutamylcysteine synthetase, the concentration of glutathione, and the thiol transport were 77%, 77% and 69%, respectively in erythrocytes from diabetic patients compared to normal control subjects. Treatment of patients with an antidiabetic agent for 6 months resulted in the restoration of -glutamylcysteine synthetase activity, the concentration of glutathione, and the thiol transport. A similar impairment of glutathione metabolism was observed in K562 cells with high glucose levels. The cytotoxicity by a xenobiotic (1-chloro-2,4-dinitrobenzene) was higher in K562 cells with high glucose than in control subjects (50% of inhibitory concentration. 300±24 mol/l vs 840±29 mol/l, p<0.01). Expression of -glutamylcysteine synthetase protein was augmented in K562 cells with high glucose, while enzymatic activity and expression of mRNA were lower than those in the control subjects. These results suggest that inactivation of glutathione synthesis and thiol transport in diabetic patients increases the sensitivity of the cells to oxidative stresses, and these changes may lead to the development of some complications in diabetes mellitus.Abbreviations ATP Adenosine 5-triphosphate - NIDDM non-insulin-dependent diabetes mellitus - GSH -glutamylcysteinyl glycine - GSSG glutathione disulphide - -GCS -glutamylcysteine synthetase - mRNA messenger ribonucleic acid - DNA deoxyribonucleic acid - C₅₀ 50% inhibitory concentration - CDNB 1-chloro-2,4-dinitrobenzene - GS-DNP S-(2,4-dinitrophenyl)glutathione - PSL photostimulated luminescence     

Plasma and urine free L-Carnitine in human diabetes mellitus

Summary L-carnitine is essential for the transport of long-chain fatty acids into mitochondria and their oxidation. Recently, a relationship between plasma free fatty acids (FFA) and L-carnitine metabolism has been observed. Plasma free L-carnitine (FC), FFA, triglycerides, cholesterol, blood glucose concentration and daily excretion of FC were determined in 20 diabetic patients as well as in 18 control subjects. Both in male diabetics and in male controls, plasma FC was significantly higher than in females. Mean plasma FC was found to be significantly reduced in diabetics (21 ± 2vs 35 ± 2 mol/1 in non-diabetic subjects; p<0.005). Daily urinary excretion of FC was clearly lower in diabetic patients than in controls (172 ± 34vs 403 ± 38 mol/24 h; p<0.001). The reduced plasma FC in diabetes mellitus may be due to redistribution between circulating free and esterified carnitine and to increased utilization of FC for synthesis of acylcarnitine in tissues.     

Glucose intolerance and impairment of insulin secretion in relation to vitamin D deficiency in East London Asians

Summary Vitamin D deficiency reduces insulin secretion and still occurs in East London Asians in whom the prevalence of diabetes mellitus is at least four times that of Caucasians. Vitamin D status was assessed in 44 of 65 non-diabetic subjects at risk of diabetes (spot blood glucose level >6.0 mmol/l <2 h post cibum, or >4.6 mmol/l >2 h post cibum on two separate occasions) and in 15 of 60 age and sex-matched low-risk control subjects who attended for oral glucose tolerance test (OGTT) after screening of 877 omnivorous subjects not known to have diabetes. It was found that 95% of at-risk and 80% of low-risk subjects were vitamin D deficient (serum 25-hydroxy-vitamin D <11 ng/ml). Diabetes was present in 16, impaired glucose tolerance in 12 and normoglycaemia in 19 at-risk subjects, impaired glucose tolerance in 2, and normoglycaemia in 13 low-risk subjects. Correlations of 30-min OGTT blood glucose, specific insulin and C-peptide levels with 25-hydroxy-vitamin D concentrations in 44 at-risk subjects were –0.31 (p=0.04), 0.59 (p=0.0001) and 0.44 (p=0.006). In 15 not-at-risk subjects 30-min OGTT specific insulin and C-peptide levels correlated with 25-hydroxy-vitamin D, r=0.39 (p=0.04) and 0.16 (p=0.43), respectively. Serum alkaline phosphatase concentration was higher in at-risk than not-at-risk subjects (59.6 vs 46.5 IU/l, p=0.012); corrected calcium concentrations were comparable (2.38 vs 2.39 mmol/l, p=0.7). Following treatment with 100,000 IU vitamin D by i.m. injection, specific insulin, C-peptide [30 min on OGTT] and 25-hydroxy-vitamin D concentrations had risen 8–12 weeks later [means±SD] from 57±62 to 96.2±82.4 mU/l [p=0.0017], 1.0±0.4 to 1.7±0.8 pmol/ml [p=0.0001] and 3.6±1.8 to 13.5±7.4 ng/ml [p=0.0001], (but not to low-risk group values of 179±89 mU/l, 2.7±1.14 pmol/ml and 8.16±6.4 ng/ml), respectively. Both total serum alkaline phosphatase and corrected calcium concentrations rose following vitamin D treatment in the at-risk subjects by 11.1±8.22 (from 44 to 55 IU/l) and 0.15±0.18, (2.43 to 2.57 mmol/l), respectively (p=0.004). Abnormal glucose tolerance was unchanged by vitamin D treatment. The value of early and sustained repletion with vitamin D in diabetes prophylaxis should be examined in communities where vitamin D depletion is common.Abbreviations OGTT Oral glucose tolerance test - IGT impaired glucose tolerance - p.c. post cibum - CV coefficient of variation - NEFA non-esterified fatty acids     

Colonic mucosal T lymphocytes in ulcerative colitis: Expression of CD7 antigen in relation to MHC class II (HLA-D) antigens

T-cell subsets and their activation state were examined by double-label immunofluorescence of cryostat tissue sections of the colon from 21 patients with ulcerative colitis (UC) and 30 histologically normal controls. Expression of MHC class I (HLA-A, B, C) and class II (HLA-D) antigens was studied in parallel. In the normal colonic mucosa, the CD4CD8 ratio in the epithelial compartment approximated 11, and in the lamina propria, 2.551. Of the CD8⁺ (cytotoxic/suppressor) subset, approximately half did not express the CD5 pan-T marker in either compartment. Virtually no Leu 8⁺ cells were observed, implying that the CD4⁺ subset consisted of helper, rather than suppressor-inducer cells. Classical markers of T-cell activation (CD25, HLA-D) and proliferation were absent, and strong expression of the CD7 immunostimulation marker was approximately equal in both CD4 and CD8 subsets. The epithelium was uniformly negative for class II antigens, but positive for class I. In UC, there were no significant alterations in CD4CD8 ratios in either compartment, and there were no changes with respect to phenotype of the subsets. In 11 of 19 patients (mainly with total colitis), enterocytes were HLA-D⁺. In this HLA-D⁺ group, there was an increase in the percentage of CD4⁺ cells coexpressing CD7; this difference was significant (P<0.02) in the lamina propria. Increased expression of CD7 was also found by the CD6⁺ T cell subset (P<0.05). These results suggest that class II expression is mediated by immunostimulated T helper cells in UC, with consequences for antigen presentation and maintenance of the chronic inflammatory state.HLA-D is used as a generic term for class II major histocompatibility complex (MHC) gene products (HLA-DR, DP, DQ) unless specified otherwise.     

Effect of β-adrenergic stimulation on free fatty acid content in subepicardial and subendocardial layers of the dog heart in situ. Separation and assay by capillary gas chromatography

Summary The effects of -adrenergic stimulation produced by an infusion of isoproterenol (1 g·kg^–1 min^–1, 30 min) were studiedin situ in the anaesthetized dog placed under a total cardiopulmonary bypass. Samples of the subepicardial and the subendocardial layers were homogenized separately prior to the extraction and methylation of free fatty acids (FFA). Gas chromatography on Carbowax 20 M capillary columns was used for the quantitation of myristic (C 140), palmitic (C 160), palmitoleic (C 161), stearic (C 180), oleic (C 181), linoleic (C 182), and arachidonic (C 204) acids.Within 5 min, isoproterenol decreased the tissue content of FFA significantly. The decrease was more pronounced in the endocardial layer where the FFA concentration reached its minimum at the 5th or the 15th min. In the epicardial layer, all the FFA reached their minimal concentration at the 30th min of the isoproterenol infusion. In both layers, lactate content remained unchanged at 5 and 15 min and rose at the 30th min only and content in phosphorylated compounds (ATP, creatine-phosphate—CP) did not show any significant variation during the -stimulation period. A significant correlation was found between the chronotropic effect of isoproterenol and the reduction of FFA concentration.With the technical assistance of Agnès Bacconin.     

Hypoadiponectinaemia and high risk of type 2 diabetes are associated with adiponectin-encoding (ACDC) gene promoter variants in morbid obesity: evidence for a role of ACDC in diabesity

Aims/hypothesis Morbid obesity (BMI>40 kg/m²) affecting 0.5–5% of the adult population worldwide is a major risk factor for type 2 diabetes. We aimed to elucidate the genetic bases of diabetes associated with obesity (diabesity), and to analyse the impact of corpulence on the effects of diabetes susceptibility genes.Methods We genotyped known single nucleotide polymorphisms (SNPs) in the adiponectin-encoding adipocyte C1q and collagen-domain-containing (ACDC) gene (–11,391G>A, –11,377C>G, +45T>G and +276G>T), the peroxisome proliferator-activated receptor gamma (PPARG) Pro12Ala SNP and ACDC exon 3 variants in 703 French morbidly obese subjects (BMI 47.6±7.4 kg/m²), 808 non-obese subjects (BMI<30 kg/m²) and 493 obese subjects (30BMI<40 kg/m²).Results Two 5-ACDC SNPs –11,391G>A, –11,377C>G were associated with adiponectin levels (p=0.0003, p=0.008) and defined a low-level haplotype associated with decreased adiponectin levels (p=0.0002) and insulin sensitivity (p=0.01) and with a risk of type 2 diabetes that was twice as high (p=0.002). In contrast, the prevalence of the PPARG Pro12Ala was identical in diabetic and normoglycaemic morbidly obese subjects. The PPARG Pro12 allele only displayed a trend of association with type 2 diabetes in the non-obese group. ACDC exon 3 variants were associated with type 2 diabetes in the non-obese group only (odds ratio 7.85, p<0.0001). In contrast, the 5-ACDC low-level haplotype was associated with type 2 diabetes in obese and morbidly obese subjects (odds ratio 1.73 and 1.92) but not in non-obese individuals.Conclusions/interpretation These data clarify the contribution of the 5-ACDC SNPs to the risk of diabesity. Their interaction with corpulence suggests for the first time a different genetic profile of type 2 diabetes in morbidly obese patients compared with in less obese individuals.     

Effect of 14 days' subcutaneous administration of the human amylin analogue,pramlintide (AC137), on an intravenous insulin challenge and response to a standard liquid meal in patients with IDDM

Summary Individuals with insulin-dependent diabetes mellitus (IDDM or type 1 diabetes) are deficient in both insulin and amylin, peptides secreted by the beta cell. We have investigated the effects of amylin replacement therapy employing the human amylin analogue, pramlintide (25, 28, 29-pro-human amylin, previously referred to as AC137), upon the responses to a standardized insulin infusion (40 mU · kg^–1 · h^–1) for 100 min and a liquid Sustacal meal (360 kcal) in 84 healthy IDDM patients. Following baseline evaluations, patients were randomly assigned to receive subcutaneous injections of placebo, 30, 100 or 300 g pramlintide 30 min before meals for 14 days. There was no meaningful difference between adverse events reported by the 30-g pramlintide and the placebo groups, but ten subjects withdrew due to nausea, eight of these in the 300-g dose group. Peak plasma pramlintide concentrations for the 30-g group were 21±3 and 29±5 pmol/l on Days 1 and 14, respectively. These values are similar to postprandial plasma amylin concentrations in normal volunteers. The plasma glucose, free insulin, glucagon, epinephrine and norepinephrine concentrations during the insulin infusion test before and after therapy were identical in each of the groups. Prior to pramlintide therapy, Sustacal ingestion produced a 4.0–4.8 mmol/l rise in plasma glucose concentrations in each of the groups. Pramlintide therapy reduced postprandial hyperglycaemia as reflected by the 3-h incremental AUC_glucose (AUC_glucose above or below fasting glucose concentration) Day 1 vs Day 14: 30 g, 322±92 vs –38±161 mmol/l · min, p=0.010; 100 g, 317±92 vs –39±76 mmol/l · min, p=0.001; and 300 g, 268±96 vs –245±189 mmol/l · min, p=0.077. Thus, pramlintide therapy with these regimens did not appear to impair either in vivo insulin action or the counter-regulatory response to hypoglycaemia but did show a clear effect of blunting postprandial hyperglycaemia following a standardized meal.Abbreviations IDDM Insulin-dependent diabetes mellitus - AUC_glucose area under the plasma glucose concentration curve - ANOVA analysis of variance     

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.     

Comparison of mRNA contents of interleukin-1Β and nitric oxide synthase in pancreatic islets isolated from female and male nonobese diabetic mice

Summary Interleukin-1 (IL-1) has been suggested to mediate beta-cell destruction in insulin-dependent diabetes mellitus (IDDM) by inducing nitric oxide production. In this study, we assessed the levels of IL-1 and the inducible form of nitric oxide synthase (iNOS), using a semi-quantitative polymerase chain reaction assay, and performed determinations of nitrite accumulation and IL-1 bioactivity, on pancreatic islets isolated from 5- and 16-week-old female and male nonobese diabetic (NOD) mice and from nondiabetes prone NMRI mice. NOD mouse islets contained notable amounts of IL-1 mRNA. At 5 weeks of age, but not at 16 weeks, the values were higher in islets isolated from NOD females compared to males. The IL-1 bioactivity showed differences roughly reflecting the mRNA levels in the NOD mouse islets. In the NMRI mouse islets the IL-1 bioactivity was very low. The expression of iNOS mRNA increased in both male and female islets between 5 and 16 weeks of age. Immunocytochemistry of pancreatic sections indicated the presence of macrophages especially in the peri-insular area of the NOD mice which suggests that IL-1 was produced by macrophages. The levels of IL-1 activity and mRNA in freshly isolated islets from NOD 5-week-old females did not correlate to the iNOS mRNA content or to the nitrite production. However, after incubation with IL-1 in vitro, both NOD and NMRI islets responded with a marked increase in nitric oxide production. It is concluded that the presence of IL-1 in isolated NOD mouse islets, via an induction of iNOS expression and nitric oxide production, cannot explain the gender difference in diabetes incidence in NOD mice.Abbreviations BB BioBreeding - GAPDH glyceraldehyde-3-phosphate dehydrogenase - IDDM insulin-dependent diabetes mellitus - iNOS inducible form of nitric oxide synthase - IL-1 interleukin-1 - IL-1ra interleukin-1 receptor antagonist protein - IL-6 interleukin-6 - NMRI Naval Marine Research Institute - NO nitric oxide - NOD nonobese diabetic - PAP peroxidase-antiperoxidase - PCR polymerase chain reaction - SDS sodium dodecyl sulphate - TNF tumour necrosis factor - OD optical density     

Expression of glomerular extracellular matrix components in human diabetic nephropathy: decrease of heparan sulphate in the glomerular basement membrane

Summary Diabetic nephropathy is characterized by albuminuria which proceeds to overt proteinuria. The highly negatively stained HS side chain of heparan sulphate proteoglycan (HSPG) is a major determinant of the charge-dependent permeability of the GBM. We set out to study the presence of HS and HSPG in the GBM of patients with diabetic nephropathy using newly developed monoclonal antibodies, and to compare HSPG expression to the expression of other previously investigated glomerular extracellular matrix compounds. Immunohistochemically, glomerular extracellular matrix components were analysed in 14 renal biopsies of patients with diabetic nephropathy and compared with those of normal control subjects. Monoclonal antibodies used were: JM403 against the HS side chain of GBM HSPG and JM72 against the HSPG-core protein. Also, a polyclonal antiserum (B31) against human GBM-HSPG-core protein was used. Additionally, antibodies were used against collagen types I, III, IV and against 1(IV)NC, 3(IV)NC and fibronectin. Staining was scored for intensity and for staining pattern by four independent observers who had no previous knowledge of the sample origin. No glomerular staining was seen for collagen type I. Collagen type III was present in some diabetic nodules. Anti-collagen type IV showed a decreased GBM staining in patients with diabetic nephropathy (p = 0.04). With anti-1(IV)NC no changes in GBM staining intensity were observed; with anti-3(IV)NC brilliant GBM staining was seen in both groups. Increased mesangial staining (p = 0.003) was seen with anti-collagen type IV in biopsies with nodular lesions. No differences were observed for fibronectin although it was abundantly present in the mesangial area of biopsies from patients with diabetic nephropathy. In biopsies with mesangial expansion and in biopsies with diabetic nodules, we observed a decreased GBM (p = 0.001) HS side chain staining (JM403) without changes in HSPG-core protein staining (JM72,B31). The HS staining pattern regularly changed from a linear to a more granular and irregular pattern. In patients with a creatinine clearance of more than 15 ml/min, the intensity of GBM HS staining showed an inverse correlation with the rate of proteinuria (r = -0.85, p = 0.004), suggesting a functional relationship. The decreased HS staining in the GBM may reflect the potentially disrupted charge barrier in diabetic nephropathy.Abbreviations HS Heparan sulphate - GBM glomerular basement membrane - HSPG heparan sulphate proteoglycan - NC noncollagenous globular domain - IDDM insulin-dependent diabetes mellitus - NIDDM non-insulin-dependent diabetes mellitus     

K-value and low insulin secretion in a non-obese white population: predicted glucose tolerance after 25 years

Aims/hypothesis Insulin resistance and insulin deficiency are proposed as risk factors for IGT and type 2 diabetes. We assessed the predictive value of initial parameters for the outcome of an OGTT performed 24.3±2.9 years later in an unselected healthy non-obese population.Methods The K-value of an IVGTT was determined in 267 healthy subjects (mean±SD: age 31.0±12.0 years, BMI 21.8±2.8 kg/m²). First-phase insulin response to a glucose infusion test was estimated as an incremental 5- or 10-min (I5 or I10) value, and as insulinogenic indices (I5/G5 or I10/G10) adjusted for insulin sensitivity determined by homeostasis model assessment for insulin resistance ([I5/G5]/HOMA-IR).Results At follow-up, six subjects had type 2 diabetes and 47 had IGT; 214 retained normal glucose tolerance. Insulin sensitivity and early (30 min) insulin response decreased with decreasing outcome OGTT. Blood glucose (2 h) at OGTT correlated positively with initial age and BMI, and negatively with I5/G5, (I5/G5)/HOMA-IR and K-value. In multiple linear regression analysis, (I5/G5)/HOMA-IR, I10, K-value, age, HOMA estimate of insulin secretion, and fasting plasma glucose were significantly associated with 2-h OGTT blood glucose. Similar results were obtained on comparing differences between subjects with normal and decreased (IGT+diabetes) glucose tolerance.Conclusions/interpretation In 267 non-obese healthy subjects, initial K-value and first-phase insulin response to glucose adjusted for insulin sensitivity, but not insulin sensitivity itself, were strong predictors of the outcome of an OGTT performed 25 years later. Thus, in contrast to obese or other high-risk populations, in lean subjects, decreased beta cell function, but not insulin resistance itself, determines future glucose tolerance.This paper is dedicated to Rolf Luft, our mentor and collaborator over several decades, on the occasion of his 90th birthday.     

Association of Trp64Arg mutation of the β3-adrenergic-receptor with NIDDM and body weight gain

Summary A possible pathogenic mutation in the 3-adrenergic-receptor gene (Trp64Arg) has been reported to be associated with an earlier age of onset of non-insulin-dependent diabetes mellitus (NIDDM) and clinical features of the insulin resistance syndrome in Pima Indian, Finnish and French subjects. Since marked heterogeneity has been reported in the association of mutations of candidate genes with NIDDM between Japanese and other ethnic groups, we investigated the association of Trp64Arg with NIDDM in Japanese subjects. The allele frequency of the mutation (Arg) was slightly, but not significantly, higher in NIDDM than in control subjects (70 out of 342 alleles [20.5%] vs 40 out of 248 [16.1%], respectively, p>0.2). When our data were combined with those of Pima Indian and Finnish subjects, however, the Arg/Arg genotype was significantly associated with NIDDM as compared with the other two genotypes (p<0.005, relative risk [RR] 2.13, 95% confidence interval [CI] 1.28–3.55). The Arg allele was also associated with NIDDM (p<0.05, RR 1.27, 95% CI 1.06–1.52). Japanese subjects homozygous for the mutation had a significantly higher body mass index (mean ± SD25.5±3.9 kg/ m²) than heterozygotes (22.6±4.1, p<0.05) and normal homozygotes (22.8±3.8, p<0.05). NIDDM patients homozygous for the mutation tended to have an earlier age of onset of NIDDM than those with other genotypes. These data suggest that the Trp64Arg mutation not only contributes to weight gain and age-at-onset of NIDDM but is also associated with susceptibility to NIDDM.Abbreviations NIDDM Non-insulin-dependent diabetes mellitus - 3-AR 3-adrenergic-receptor - Trp64Arg a mutation in the 3-adrenergic-receptor gene causing a Trp to Arg change at codon 64 - BMI body mass index - ADRB3 3-adrenergic receptor gene - PCR polymerase chain reaction - RFLP restriction fragment length polymorphism - RR relative risk - CI confidence interval     

Large-bowel cancer in the young: A national survival study

Large-bowel cancer in young patients is reported to be a more aggressive and advanced disease at presentation and is believed to be associated with a relatively poor prognosis. Of 2420 patients registered in New Zealand (1968 to 1970), 131 were under 40 years of age and 2289 were over 40 years of age. The annual average incidence of treatable colorectal cancer in patients under 40 years of age was 2.36 per 100,000 and 82.93 in patients over 40 year of age. There were predominantly more females in both age groups with colonic tumors, 5044 (femalemale), and 759652 (femalemale). The rectal tumor male-to-female ratio of 10.68 in those over 40 years of age was reversed in those under 40 years of age (12.08). There was no significant difference in the subsite distribution of colorectal cancers between the two groups. There was a higher proportion of Stage 1 tumors in those under 40 years of age and a correspondingly higher proportion of Stage 2 tumors in those over 40 years of age. The overall crude and relative five-year survival rates for patients under 40 years of age were both 60 percent, whereas the crude rate for older patients was 42 percent, with a corresponding relative rate of 53 percent. Ten-year survival rates were generally higher in younger patients. From this study, there was no evidence to suggest that younger patients (less than 40 years old) with colorectal cancer had worse prognoses and did not survive as long as older patients (40 years and over).Poster presentation at the meeting of The American Society of Colon and Rectal Surgeons, Anaheim, California, June 12 to 17, 1988.     

The effect of hand osteoarthritis on grip and pinch strength and hand function in postmenopausal women

The aim of the study was to investigate the effect of osteoarthritis on hand function in postmenopausal women. One hundred patients with hand OA and 70 healthy volunteers as controls were evaluated. Grip and pinch strength measurements and Dreisers functional index were used for hand function. Pain was assessed by a visual analog scale, and tenderness was assessed by palpation and scored, depending on the severity of tenderness, as 0, 1 or 2. Heberdens and Bouchards nodules and joint involvement were also recorded. The number of patients with only distal interphalangeal joint involvement was 50 (50%), those with distal interphalangeal joint plus proximal interphalangeal joint involvement was 49 (49%), and those with carpometacarpal joint involvement numbered 18 (18%). The incidence of Heberdens and Bouchards nodules was 85% and 36%, respectively. Eighty-six (86%) patients were suffering from pain and 57 were found to have tenderness. Grip and pinch strength was significantly lower (p<0.05) and Dreisers functional index score was significantly higher (p<0.001) in the study group (particularly in grade 4 OA). Grip strength was lower in hand OA patients with distal interphalangeal joint plus proximal interphalangeal joint involvement than in those with only distal interphalangeal joint and carpometacarpal joint involvement. Pinch strength was also lower in patients with distal interphalangeal joint plus proximal interphalangeal joint plus carpometacarpal joint involvement. The patients with Heberdens and Bouchards nodules had lower grip and pinch strength than controls. Also, pain and tenderness had significant (p<0.05) effects on hand function. Dreisers total score ranged from 0 to 10 in 80 (80%) patients and from 11 to 20 in 20 patients. In conclusion, hand osteoarthritis contributes to hand dysfunction, mainly related to the severity of osteoarthritis, pain, joint involvement and the presence of nodules.Abbreviations CMC Carpometacarpal - DIP Distal interphalangeal - OA Osteoarthritis - PIP Proximal interphalangeal     

Marine lipids normalize cholesteryl ester transfer in IDDM

Summary Patients with insulin-dependent diabetes mellitus (IDDM) have a pathological increase in cholesteryl ester transfer (CET) that enriches the apolipoprotein B-containing lipoproteins with cholesteryl ester and increases their atherogenicity. Since we have shown earlier that omega-3 (n-3) fatty acids present in marine lipids normalize both CET and lipoprotein composition in non-diabetic patients with hypercholesterolaemia, we sought to determine whether the same beneficial effects could be achieved in nine normolipidaemic (triglycerides 1.10; cholesterol 4.94, high density lipoprotein 1.10 mmol/l) IDDM patients (fructosamine 424±156; normal 174–286 mol/l) treated for 2 months with n-3 fatty acids (4.6 g/day). Before treatment, CET measured by both mass and isotopic assays was abnormally accelerated (p<0.001). While marine lipids modestly decreased triglyceride levels (–14%; p<0.05), CET fell dramatically in all subjects (mass assay: –97% at 1 h; isotopic assay: –58%; p<0.001) to below control levels with no change in glycaemic control (fructosamine 408±103 mol/l). The mass of cholesteryl ester transfer protein paradoxically increased significantly (pre-treatment: 2.04±0.86 vs post-treatment 2.48±0.97 g/ml; p<0.05). Since it is believed that accelerated CET promotes the formation of atherogenic cholesteryl ester-enriched apo B-containing lipoproteins, the capacity of marine lipids to reverse this functional abnormality without altering glycaemic control suggests that these agents may have an adjunctive role to play in the nutritional therapy of IDDM.Abbreviations IDDM Insulin-dependent diabetes mellitus - NIDDM non-insulin-dependent diabetes mellitus - CET cholesteryl ester transfer - CETP cholesteryl ester transfer protein - HDL high density lipoprotein - LpL lipoprotein lipase - apo B apolipoprotein B - n-3 fatty acids omega-3 fatty acids     

Peripheral blood lymphocyte apoptosis and circulating dendritic cells in patients with systemic lupus erythematosus: correlation with immunological status and disease-related symptoms

We investigated in vivo the relationship between the degree of peripheral blood lymphocyte apoptosis and circulating dendritic cells (DC) as well as the immunological status in 45 patients with systemic lupus erythematous (SLE). Apoptosis was detected by phosphatidylserine externalization and assays to detect caspase activation. The total DC count (tDC) and their myeloid, mDC1 (BDCA1+) and mDC2 (BDCA3+), and plasmacytoid, pDC (BDCA3+), subtypes were assessed. Moreover, several immunological parameters, such as complement proteins, interferons (IFN), tumor necrosis factor (TNF)-, interleukin (IL)-4, and IL-6 levels were assessed. There were no significant differences in both the intensity of apoptosis and DC counts between active and inactive SLE as well as between untreated patients and those treated with steroids. The incidence of lymphocyte apoptosis correlated positively with T-cell count, both T-helper (p=0.004) and cytotoxic T cells (p=0.001), but not with B or natural killer (NK) cells. The intensity of apoptosis enhanced along with the increase in complement C3 (p=0.016) and decrease in IFN- (p=0.040) levels. The apoptotic cell count correlated with tDC (p=0.031), mDC1 (p=0.007), and pDC (p=0.039) counts. Both tDC and mDC1 counts correlated positively with antinuclear antibody (ANA) titers (p=0.017 and 0.032, respectively). Moreover, tDC correlated with C4 (p=0.039) and pDC with both C3 (p=0.032) and C4 (p=0.007) levels. The DC counts correlated inversely with IFN- (tDC, p=0.038; mDC1, p=0.009), IL-6 (mDC2, p=0.031), or serum IgG levels (tDC, p=0.006; mDC1, p<0.001; mDC2, p=0.001). We found a positive correlation between lymphocyte apoptosis and peripheral blood DC count as well as the level of complement proteins in patients with SLE. The enhanced lymphocyte apoptosis was accompanied by the decrease in concentration of some cytokines, such as IFN- or IL-6, as well as serum IgG level. This finding may reflect pathogenetic events during development of the disease, which include a persistent signal derived from circulating apoptotic lymphocytes, mobilizing the complement system, and attracting peripheral blood DC.     

Clinical usefulness of free light chain concentration as a tumor marker in multiple myeloma

Monoclonal immunoglobulin, as a marker for monoclonal gammopathy, is evaluated by protein electrophoresis (PEP) and immunofixation electrophoresis (IFE). However, PEP and IFE are not satisfactory in sensitivity, objectivity, and facility. Recently, a highly sensitive, automated immunoassay for measurement of free light chain (FLC) concentrations in serum and urine has been developed for the identification and monitoring of patients with monoclonal gammopathy. To explore the clinical usefulness of measurement of FLC concentrations, we measured the and FLC concentrations and calculated the / FLC ratios for three groups [multiple myeloma (MM), other diseases, and control] and compared the results of the FLC assay with the results of PEP or IFE. The concentrations of serum and FLCs and the / FLC ratios for the MM group and non-MM groups were distinct. In the MM group, some sera and urine samples had no evidence of M protein on PEP and IFE, but FLC assay showed abnormal concentrations of FLCs and abnormal / FLC ratios in most cases. As compared with the PEP, the / FLC ratio revealed higher sensitivity in all diagnostic ranges with different cutoff values. Particularly, when the cutoff value 2.0 for / FLC ratio was used, specificity and positive predictive value were largely improved than when the cutoff values 1.2 and 1.5 were used. These findings indicated that FLC assay enables to detect myeloma patients with very low M protein due to early stage or after therapy and to distinguish patients with monoclonal increase of FLC from patients with polyclonal increase of FLC due to other conditions, particularly using / FLC ratio 0.3–2.0 as a diagnostic range. Despite some technical limitations of the assay, the incorporation of / FLC ratios with FLC concentrations is useful in the detection of M protein, particularly with negative serum or urine IFE results, and differentiation of monoclonal gammopathies from patients with polyclonal increase in FLC due to other conditions.