High glucose and hydrogen peroxide increase c-Myc and haeme-oxygenase 1 mRNA levels in rat pancreatic islets without activating NFκB

Aims/hypothesis Hyperglycaemia and the pro-inflammatory cytokine IL-1 induce similar alterations of beta cell gene expression, including up-regulation of c-Myc and haeme-oxygenase 1. These effects of hyperglycaemia may result from nuclear factor-kappa B (NFB) activation by oxidative stress. To test this hypothesis, we compared the effects of IL-1, high glucose, and hydrogen peroxide, on NFB DNA binding activity and target gene mRNA levels in cultured rat islets.Methods Rat islets were pre-cultured for 1 week in serum-free RPMI medium containing 10 mmol/l glucose, and further cultured in glucose concentrations of 5–30 mmol/l plus various test substances. Islet NFB activity was measured by ELISA and gene mRNA expression was measured by RT-PCR.Results IL-1 consistently increased islet NFB activity and c-Myc, haeme-oxygenase 1, inducible nitric oxide synthase (iNOS), Fas, and inhibitor of NFB alpha (IB) mRNA levels. In comparison, 1- to 7-day culture in 30 mmol/l instead of 10 mmol/l glucose stimulated islet c-Myc and haeme-oxygenase 1 expression without affecting NFB activity or iNOS and IB mRNA levels. Fas mRNA levels only increased after 1 week in 30 mmol/l glucose. Overnight exposure to hydrogen peroxide mimicked the effects of 30 mmol/l glucose on haeme-oxygenase 1 and c-Myc mRNA levels without activating NFB. On the other hand, the antioxidant N-acetyl-l-cysteine inhibited the stimulation of haeme-oxygenase 1 and c-Myc expression by 30 mmol/l glucose and/or hydrogen peroxide.Conclusions/interpretation In contrast to IL-1, high glucose and hydrogen peroxide do not activate NFB in cultured rat islets. It is suggested that the stimulation of islet c-Myc and haeme-oxygenase 1 expression by 30 mmol/l glucose results from activation of a distinct, probably oxidative-stress-dependent signalling pathway.     

Diagnosis of the mucopolysaccharidoses using cultured skin fibroblasts and amniotic fluid cells

Assay of-L-iduronidase, heparin sulphamidase,N-acetyl--D-glucosaminidase, arylsulphatase B,-L-fucosidase,-glucuronidase,-galactosidase and-D-mannosidase in cultured cells is described. Activities in deficient fibroblast strains are compared to control fibroblast strains. The first case of Sanfilippo B in the United Kingdom is reported. A comparison of enzyme activities in cultured fibroblasts and amniotic fluid cells is made.     

GlutathioneS-transferase expression in malignant mesothelioma and non-neoplastic mesothelium: an immunohistochemical study

Expression of the glutathioneS-transferase (GST) subclasses , and was investigated immunohistochemically in 20 normal or hyperplastic mesothelium and in 57 malignant mesothelioma cases. These results were correlated with survival and also with P-170 glycoprotein expression. Nearly all the non-neoplastic mesothelium cases were positive for GST and . About half of the non-neoplastic cases were positive for . Twenty-nine (51%) malignant mesotheliomas were positive for at least one of the GST species; 21 (37%) showed immunoreactivity for , 18 (31.5%) for and 21 (37%) for . A total of 54 mesothelioma cases displayed immunoreactivity for the P-170 glycoprotein. For GST and GST, a statistical significance between expression and increased survival was found (respectivelyP=0.012 and 0.024) while for GST no significance was found. The results of this study demonstrate that expression of GST correlates positively with increased survival in malignant mesothelioma. It is also concluded that, in mesothelioma, GST and P-170 glycoprotein may contribute to the resistance to cytotoxic drugs frequently observed in these tumours. No correlation between GST and P-170 expression was demonstrated.Abbreviation GST glutathioneS-transferase     

Islet B-cell dysfunction and the time course of recovery following chronic overinsulinisation of normal rats

Summary Appropriate insulin therapy may preserve or improve islet B-cell function whereas the effects of overinsulinisation are unclear. Pancreatic islet B-cell function was therefore studied after overinsulinisation of normal rats for 4 weeks (fed blood glucose 2.2–4.5 mmol/l, controls 4.1–7.0 mmol/l). Insulin secretion was assessed by a 3-h hyperglycaemic clamp (10.0 mmol/l) performed 1, 48, and 120 h after insulin withdrawal (n=6 in each group). When the clamp was performed 1 h after insulin withdrawal, clamp insulin concentration was 1.6±0.1 g/l, compared to 9.3±1.0 g/l in control rats. The integrated area under the plasma insulin concentration curve was also significantly decreased (4.8±0.4 vs 20.3±2.2 g·l^–1·h^–1, p<0.001), but recovered to 9.4±1.0 g·l^–1·h^–1 after 48 h, and to 17.5±1.4 g·l^–1·h^–1 after 120 h. Pancreatic insulin contents were decreased at 1 h (6±1 g/g wet wt) and 48 h (54±12 g/g wet wt) but not at 120 h (221±30 g/g wet wt) after withdrawal (controls, 303±29 /g wet wt) and there was a strong relationship with pancreatic preproinsulin mRNA and the clamp insulin response. Thus, overinsulinisation with prolonged periods of low blood glucose concentrations impairs islet B-cell function, but is reversible over 5 days.     

Interactive influences of peroxisome proliferator-activated receptor α activation and glucocorticoids on pancreatic beta cell compensation in insulin resistance induced by dietary saturated fat in the rat

Aims/hypothesis We sought to elucidate whether excess glucocorticoids and increased dietary lipids act synergistically to impair glucose tolerance and, if so, whether activation of peroxisome proliferator-activated receptor (PPAR) has an adverse or beneficial effect on glucose tolerance.Methods Dexamethasone (100 g kg^–1 body weight day^–1; 5 days) was administered to insulin-resistant rats fed a high-saturated-fat (HF) diet for 4weeks. The PPAR agonist WY14643 was administered (50 mg kg^–1 body weight intraperitoneally) 24 h before sampling. Glucose-stimulated insulin secretion (GSIS) was assessed in vivo after an acute glucose bolus injection, and in vitro using step-up and step-down islet perifusions.Results Although neither PPAR activation nor dexamethasone alone affected fasting glycaemia in the HF group, dexamethasone in combination with PPAR activation elicited marked postabsorptive hyperglycaemia. Dexamethasone treatment of HF rats had little effect on GSIS after an acute glucose challenge in vivo, but induced glucose intolerance. PPAR activation augmented GSIS in dexamethasone-treated HF rats in vivo, restoring glucose tolerance. Contrasting with data obtained in vivo, greatly enhanced peak rates of GSIS were observed ex vivo in perifusions of islets from dexamethasone-treated HF rats compared with those from untreated HF rats, an effect attenuated by antecedent PPAR activation.Conclusions/interpretation The study demonstrates that glucocorticoid excess precipitates the development of glucose intolerance in rats maintained on a high-saturated-fat diet. It does this by interrupting the negative feedback loop between insulin sensitivity and secretion in vivo, such that further enhancement of compensatory insulin secretion is not possible. PPAR activation restores the coupling between insulin secretion and action.     

Isolation and characterization of the normal canine β-galactosidase gene and its mutation in a dog model of GM1-gangliosidosis

The acid -galactosidase cDNA of Portuguese Water dogs was isolated and sequenced. The entire coding region of the gene consists of 2004 nucleotides encoding a protein of 668 amino acids. Its encoding sequence indicates approximately 86.5% identity at the nucleotide level and about 81% identity at the amino acid level with the encoding region of the human acid -galactosidase gene. The deduced amino acid sequence contains a 24-amino-acid putative signal sequence, six possible glycosylation sites, and seven cysteine residues. A homozygous recessive mutation, causing canine GM1-gangliosidosis, was identified at nucleotide G²⁰⁰A in exon 2 resulting in an Arg⁶⁰His (mutation R60H) amino acid substitution. The mutation creates a new restriction enzyme site for Pml1. Genotyping 115 dog samples for this acid -galactosidase gene alteration readily distinguished affected homozygous recessives (n=5), heterozygous carriers (n=50) and normal homozygotes (n=60). DNA mutation analysis provided a method more specific than enzyme assay of -galactosidase for determination of carriers.     

Contributions of nuclear medicine to the therapy of malignant tumors

Radionuclides are applied in oncology for diagnosis and therapy. The former demands gamma — emitting radionuclides for labeling specific substrates for localizing malignant tissue and for analyzing tumor metabolism in vivo. Here, positron emission tomography (PET) may register in vivo the metabolism, for example, of glucose, amino acids, and receptors and of potentially useful cytotoxic agents. The advantage of the positron emitting radionuclides of carbon, nitrogen and fluorine is the labeling of substrates without changing substrate specificity within the metabolic reaction chain; also, substrate concentration in situ may be quantified. With regard to therapy radionuclides that emit -and -particles or decay by electron capture with the Auger effect, are administered in ionic form or with tumor seeking substrates. Examples are radioiodine for treating thyroid malignancy and radiophosphorus for myelopoliferative diseases. Organically bound radionuclides are given as labeled ligands for specific receptors, such as meta-iodo-benzylguanidine (MIBG) for treating the catecholamine producing tumors phaeochromocytoma and neuroblastoma and labeled monoclonal antibodies for tumors specific receptors. Highly localized energy depositions come from Auger emitters such as¹²⁵I and by the neutron capture therapy, where boron-10 in the tumor cell is exposed to thermal neutrons for initiating the B¹⁰ (n; ) Li⁷ reaction, especially for treating neuro- and glioblastoma and melanoma. Endogenous radiotherapy with radionuclides rely on the success of delivering a proper amount of energy into individual tumor cells with optimal protection of normal tissue. The inevitable heterogeneity of energy deposition events from such approaches demands careful dosimetric assessment for which the classical methods of dosimetry for percutaneous radiotherapy are not applicable.The Journal of Cancer Research and Clinical Oncology publishes in loose succession Editorials and Guest editorials on current and/or controversial problems in experimental and clinical oncology. These contributions represent exclusively the personal opinion of the author The Editors     

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.     

Demonstration of a relatively hepatoselective effect of covalent insulin dimers on glucose metabolism in dogs

Summary Insulin analogues with relatively greater effect on hepatic glucose production than peripheral glucose disposal could offer a more physiological approach to the treatment of diabetes mellitus. The fact that proinsulin exhibits this property to a minor degree may suggest that analogues with increased molecular size may be less able than insulin to obtain access to peripheral receptor sites. Covalent insulin dimers have previously been shown to possess lower hypoglycaemic potencies than predicted by their in vivo receptor binding affinities. Reduced rates of diffusion to peripheral target tissues-might be an explanation for the lower in vivo potency compared to insulin. To test the relative hepatic and peripheral effects of covalent insulin dimers, glucose clamp procedures with D-[3-³H] glucose tracer infusions were used in anaesthetised greyhounds to establish dose-response curves for rates of hepatic glucose production and glucose disposal with insulin, N^B1, N^{B 1},-suberoyl-insulin dimer, and N^B29, N^{B 29},-suberoyl-insulin dimer. With N^B1, N^{B 1},-suberoyl-insulin dimer molar potencies relative to insulin were 68%, (34–133) (mean and 95% fiducial limits), for inhibition of hepatic glucose production and 14.7%, (10.3–20.9) for glucose disposal. With N^B29,N^{B 29},-suberoyl-insulin dimer potencies were 75%, (31–184) and 2.5%, (1.5–4.3), for inhibition of hepatic glucose production and for glucose disposal, respectively. The demonstration that both dimers exhibit a significantly greater effect on glucose production than on glucose disposal supports the suggestion that analogues with increased molecular size may exhibit reduced ability to gain access to peripheral target cells.Abbreviations B1-B 1D N^B1,N^{B 1},-suberoyl-insulin dimer - B29-B 29D N^B29,N^{B 29},-suberoyl-insulin dimer - Ra hepatic glucose production rate - Rd peripheral glucose disposal rate - M_r relative molecular weight - MCR metabolic clearance rate - ANOVA analysis of variance     

Sulphonylurea stimulates glucose uptake in rats through an ATP-sensitive K+ channel dependent mechanism

Summary We studied the effect of gliclazide, a second-generation sulphonylurea, on rat skeletal muscle glucose uptake using perfused hindquarter muscle preparations. Gliclazide at concentrations of 10 to 1000 g/ml increased (p<0.05) the basal glucose uptake. The effect of gliclazide on glucose uptake was immediate and dose-dependent, reaching a plateau at a concentration of 300 g/ml; the half-maximal effect was obtained between 25 and 50 g/ml. The glucose uptake stimulated by gliclazide (300–1000 g/ ml) did not differ from that achieved by 10^–9 mol/l insulin, and was lower (p<0.05) than that obtained with 10^–7 mol/l insulin. The combination of gliclazide (300 g/ml) and 10^–9 mol/l insulin produced an increase in glucose uptake (7.7±0.6 mol · g^–1 · h^–1, n=8, mean±SEM) which was higher (p<0.05) than that achieved with 10^–9 mol/l insulin (5.6±0.7 mol · g^–1 · h^–1, n=11) and not different from that obtained with 10^–7 mol/l insulin (9.8±1.0 mol · g^–1 · h^–1, n=11). Diazoxide (100 mol/l), an ATP-sensitive K⁺ channel opener, reversed the stimulatory effect of gliclazide (100 g/ml) on muscle glucose uptake from 3.1±0.4 to 0.5±0.2 mol · g^–1 · h^–1, (n=7, p<0.001). The addition of diazoxide prior to gliclazide into the perfusion medium blocked the gliclazide-induced glucose uptake by the hindquarter muscle preparations. In conclusion, gliclazide alone has an immediate stimulatory effect on glucose uptake by skeletal muscle and together with insulin has an additive effect on muscle glucose uptake. The effect of gliclazide on muscle glucose uptake seems to be due to the inhibition of ATP-sensitive K⁺ channels.Abbreviations NIDDM Non-insulin-dependent diabetes mellitus - GLUT glucose transporter     

Twenty-four hour profiles of plasma C-peptide in Type 1 (insulin-dependent) diabetic children

Summary Twenty-four hour profiles of plasma C-peptide an index of endogenous insulin secretion, were performed in 15 Type 1 (insulin-dependent) diabetic children. Plasma C-peptide was detectable in six children, of whom four (C-peptide producers) had peak values above normal fasting levels. In each of the six children with residual B cell function, there was a close correlation between plasma C-peptide and simultaneous blood glucose (r> 0.50, p< 0.05). Post-breakfast peak blood glucose was 10.2 ± 1.7 mmol/l (mean ±SEM) in the C-peptide producers and 18.7 ± 1.7 mmol/l in the 11 children with low or no detectable C-peptide. Mean M-value, an index of deviation from an ideal blood glucose, was lower in the C-peptide producers (p<0.05). It is concluded that residual functioning B cells in diabetic children behave physiologically in that insulin secretion fluctuates in accordance with the prevailing blood glucose; and that the pattern of action of injected insulin is more critical in non-C-peptide producers who lack the post-prandial dampening effect provided by residual endogenous insulin secretion.     

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.     

Different effects of glucose and glyburide on insulin secretion in rat pancreatic islets pre-exposed to interleukin-1β. Possible involvement of K+ and Ca2+ channels

Summary In vitro islet exposure to interleukin 1 inhibits the beta-cell response to glucose. We have studied whether a similar inhibition also occurs in response to the sulphonylurea glyburide. Rat pancreatic islets were cultured for 24 h in the presence or absence of 50 U/ml interleukin 1 and then stimulated with either glucose or glyburide for 1 h at 37 °C. In control islets basal insulin secretion was 117±32 pg · islet^–1 · h^–1 (mean ± SEM, n=7) and greatly increased in response to 16.7 mmol/l glucose (2140±293) or 10 mol/l glyburide (1464±234). When islets were pre-exposed to interleukin 1, insulin release was significantly reduced in response to glucose (323±80, p<0.001) but not in response to glyburide (1316±185). Since both glucose and glyburide influence beta-cell K⁺ and Ca²⁺ efflux, to further investigate this different response in islets exposed to interleukin 1 we measured both Rb⁺ efflux (as index of the ATP-sensitive K⁺ channel activity) and Ca²⁺ uptake. In control islets, the increased insulin secretion in response to 16.7 mmol/l glucose or 10 mol/l glyburide was associated with a reduction of ⁸⁶Rb efflux (decrement of –50±1.2 % and –49±2.3 %, respectively, mean ± SEM, n=5). In contrast, in interleukin 1pre-exposed islets both glucose and glyburide stimulation only slightly modified ⁸⁶Rb efflux (decrement of –19±1.9% and –5.3±3.1 %, respectively, n=5, p<0.001). ⁴⁵Ca²⁺ uptake in control islets was 2.6±0.4 pmol · islet^–1 · 20 min^–1 under basal conditions (at 2.8 mmol/l glucose), and increased to 16.8±3.2 and 10.7±2.1 pmol · islet^–1 · 20 min^–1 in islets stimulated with 16.7 mmol/l glucose or 10 mol/l glyburide, respectively (mean ± SEM, n=6). ⁴⁵Ca²⁺ uptake in interleukin 1 treated islets was higher than in control islets under basal conditions (4.6±0.6 pmol · islet^–1 · 20 min^–1 at 2.8 mmol/l glucose, p<0.05), but was significantly reduced in response to glucose 16.7 mmol/l (7.1±1.1, p<0.01 with respect to control islets). In contrast to glucose, 10 mol/l glyburide was able to stimulate calcium uptake in interleukin 1 treated islets in a similar way to control islets (12.8±2.5). The present data demonstrate that rat pancreatic islets treated with interleukin 1 for 24 h lose their responsivity to glucose, but not to glyburide. The difference between the two secretagogues is associated with the persistent ability of glyburide to influence Ca²⁺ uptake even in islets with impaired K⁺-channel function.     

Lack of α2-Antiplasmin Enhances ADP Induced Platelet Micro-Aggregation through the Presence of Excess Active Plasmin in Mice

Background: The role of 2-antiplasmin (2-AP) on platelet aggregation was investigated using mice deficient in 2-AP (2-AP^–/–) or using wild type mice (2-AP^+/+). Methods: Blood samples were taken from each mouse under anesthesia with ether and platelet rich plasma (PRP) was prepared. Platelet aggregation induced by various doses of ADP (0.3–30 M) was detected using a laser-light scattering (LS) system. Aggregated forms were observed using a scanning electron microscopy (SEM). Results: Dose-dependent platelet aggregation was not different in both types of mice. However, platelet micro-aggregate formation in 2-AP^–/– mice induced by low dose of ADP (1.0 M) markedly increased compared to the situation in wild type mice. Aggregated form detected by SEM showed supported data from LS analysis. When washed platelets of 2-AP^+/+ mice were resuspended in plasma of 2-AP^–/– mice, platelet micro-aggregation was also increased. On the contrary, when washed platelets of 2-AP^–/– mice were suspended in plasma of 2-AP^+/+ mice, platelet micro-aggregation did not change. In separate experiments, tPA (1.0 g/ml) was added to PRP before the stimulation of ADP. tPA had no effect on platelet aggregation in 2-AP^+/+ mice, however platelet micro-aggregation in 2-AP^–/– mice was markedly increased by the treatment with tPA. Moreover, the amount of released ATP from stimulated platelets was increased in 2-AP^–/– mice treated with tPA. Conclusion: Lack of 2-AP increased platelet micro-aggregation, and plasmin plays an important role in the formation of platelet aggregation when 2-AP knockout mice are used. Consequently, the reduction of 2-AP could be a risk factor for the activation of platelets resulting in thrombus formation.     

Influence of anti-ulcer drugs used in Japan on the result of <Superscript>13</Superscript>C-urea breath test for the diagnosis of <Emphasis Type="Italic">Helicobacter pylori</Emphasis> infection

Background The ¹³C-urea breath test (UBT) is a simple test for the diagnosis of Helicobacter pylori infection, but several factors have been reported to affect the results of this test. In this study, the effects of the anti-ulcer drugs used in Japan on the results of the UBT were determined.Methods The subjects of the study were 64 adult volunteers who tested positive for H. pylori infection by the serum antibody method. Eight classes of anti-ulcer drugs used in Japan were administered at their usual doses to these subjects: lansoprazole, a proton pump inhibitor (PPI); nizatidine, an H₂-receptor antagonist (H₂RA); and polaprezinc, ecabet sodium, rebamipide, teprenone, cetraxate hydrochloride, and sucralfate, all mucoprotective agents. The study drugs were randomized for administration to the subjects, and each of the drugs was administered for 14 consecutive days. The UBT was performed on days 0, 14, and 21.Results The mean ¹³C in the lansoprazole group was significantly decreased on day 14, to below 10, in 4 of 16 subjects, and in 1 of the 4 subjects, the test result was negative, with the ¹³C falling to 1.7. The value returned to baseline 1 week after the discontinuation of lansoprazole. The other drugs administered had no significant effect on the result of the UBT, except that the mean ¹³C showed a tendency to decrease after the administration of ecabet sodium and rebamipide.Conclusions Administration of a PPI may produce a false-negative UBT result, while other anti-ulcer drugs, for the most part, have little effect on the result of the UBT when used alone.     

Gene expression of cytokines suppressing hematopoietic progenitor cells in lymphoid malignancies

Summary The expression of cytokine genes for tumor necrosis factor (TNF), lymphotoxin and transforming growth factor (TGF), all of which are known to suppress normal hematopoiesis, was investigated in 32 patients with lymphoid malignancies using Northern blot analysis. Messenger RNA (mRNA) for TNF, lymphotoxin and TGF was detected in 9 cases, 2 cases and 7 cases, respectively. When the relationship between cytokine gene expression and surface phenotype was analyzed, the expression of CD19 correlated significantly with expression of the TNF gene (P<0.05). This suggests that B cell malignancies are likely to produce TNF. When the hematological parameters of patients expressing and not expressing the gene were compared, the expression of TNF mRNA was found to correlate with more profound anemia in acute lymphoblastic leukemia (P<0.05). Both granulocyte and platelet counts were lower in patients expressing TNF mRNA; however, the decreases were not significant. Neither lymphotoxin nor TGF gene expression correlated significantly with any hematological parameter.Abbreviations TNF tumor necrosis factor - TGF transforming growth factor - IL interleukin - ALL acute lymphoblastic leukemia - CLL chronic lymphocytic leukemia - ATLL adult T-cell leukemia/lymphoma - NHL non-Hodgkin's lymphoma - MM multiple myeloma Partly supported by grants-in-aid from the Ministry of Education, Science and Culture of Japan (60770949, 63015063, 02256102, 03670325) and from the Fukuoka Anti-Cancer-Society.     

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.     

The circulating molecular weight forms of infused recombinant insulin-like growth factor-I and effects on glucose and fat metabolism in lambs

Summary We have investigated the relationship between the plasma distribution of infused recominant insulin-like growth factor-I across the insulin-like growth factor binding proteins and the resultant effects on glucose and fat metabolism. The studies were performed in 24-h fasted ram lambs which received primed constant infusions of ³H labelled glucose tracer. When isotopic equilibrium had been reached, the animals received 90-min infusions of human insulin-like growth factor-I at various doses (2.5, 20, 40 and 120 g· kg^–1·h^–1, n=3 for each dose). Total plasma insulin-like growth factor-I was significantly elevated by infusion at a rate of 40 g·kg^–1·h^–1 (from 185±14 g/l to 442±41 g/l, p<0.05) and 120g·kg^–1h^–1 (from 181±2 g/l to 953±39 g/1, p<0.005). The plasma concentrations of insulin-like growth factor-I not associated with binding proteins remained undetectable (<15 g/l) at the end of the 2.5 and 20 g·kg^–1·h^–1 doses, but were significantly elevated at the end of the 40 and 120 g·kg^–1·h^–1 infusions (to 71±14 g/l, p<0.05 and 176±55 g/l, p<0.01 respectively). The infused insulin-like growth factor-I associated primarily with 35–60 kilodalton binding proteins. Glucose kinetics were significantly altered only by the highest dose infusion, during which there was a fall in plasma glucose concentration from 3.5±0.2 mmol/l to 1.9±0.2 mmol/l (p<0.05). This was due to a 51% increase in the rate of glucose clearance. There was no significant change in the rate of glucose production. The plasma concentrations of glycerol and non-esterified fatty acid were not changed by any of the doses infused. We conclude that the hypoglycaemic action of infused recombinant insulin-like growth factor-I relates to a marked elevation of free insulin-like growth factor-I in the plasma, but that a threshold concentration of free insulin-like growth factor-I must be exceeded before this action is observed. The hypoglycaemic action of recominant insulin-like growth factor-I results primarily from an increase in glucose clearance while glucose metabolism was more sensitive than fat metabolism to infused recominant insulin-like growth factor-I. Both these actions contrast with those of insulin, and suggest that the acute metabolic effects of recombinant insulin-like growth factor-I are not mediated simply by cross-reaction with insulin receptors.     

Palmitate dependence of insulin secretion, “de novo” phospholipid synthesis and 45Ca2+-turnover in glucose stimulated rat islets

Summary Palmitate ability to modify D-[U-¹⁴C]glucose incorporation into different lipids (de novo synthesis), as well as sugar-stimulation of insulin release and ⁴⁵Ca²⁺-fluxes, was investigated in islets of fed and 48-h starved rats. The fatty-acid induced dose-dependent, correlative increments of insulin secretion, ⁴⁵Ca²⁺-influx and the de novo synthesis of each phospholipid fraction analysed at 20 mmol/l (but not 3 mmol/l) glucose. Omission of calcium reduced drastically (p<0.001) insulin release and the de novo synthesis of neutral glycerolipids, leaving unaltered that of acidic phospholipids (phosphatidate and phosphoinositides). The increased synthesis of the latter is therefore not the consequence of stimulated secretion. It could initiate or contribute to maintain an increased turnover of islet phosphoinositides, thus generating some mediators of the calcium signalling system (inositol phosphates). Starvation led to a drastic reduction (p<0.001) of both insulin secretion, de novo synthesis of each lipid fraction, and ⁴⁵Ca²⁺-influx in response to glucose and palmitate. The presence of a fatty-acid oxidation inhibitor (2-bromostearate or 2-tetradecylglycidate) prevented the effect of starvation on ⁴⁵Ca²⁺-influx, as it has been shown to do on insulin secretion and palmitate incorporation into islet lipids. It is finally suggested that palmitate might amplify the insulin secretory response of islets to glucose, through the stimulation of the de novo synthesis of phosphoinositides and the subsequent generation of inositol phosphates, which would contribute to accelerated calcium turnover.     

Production of interleukin-1β and tumour necrosis factor-α in patients with benign or malignant ovarian tumours

Summary To assess the role of interleukin-1 (IL-1) and tumour necrosis factor (TNF) in the physiological host defence mechanisms against malignancies, the production of these cytokines in sera, ascitic and cyst fluids and in the tumour tissues of patients with benign or malignant ovarian tumours was studied. IL-1 was found neither in the sera nor in the ascitic fluids of these patients. It was also virtually absent from the cyst fluid samples. However, a mean value of 790 pg IL-1/g tumour was found. Like IL-1, TNF was virtually absent in the serum samples. It was, however, detectable in the ascitic and cyst fluids and tumour tissues. The TNF concentrations were highest in the tumour tissues, with a mean level of 328 pg/g tumour. When comparing the level of IL-1 and TNF in patients with benign tumours to that seen in patients with malignant tumours, no differences in production were observed, regardless of the origin of the test samples. Our results indicate the production of IL-1 and TNF in patients with ovarian tumours. More importantly, the finding that the production of these cytokines in patients with benign tumours is similar to that in patients with malignant tumours supports the conclusion that the production of these cytokines is more a nonspecific indicator of an inflammatory process than a specific response to a malignant process.Abbreviations IL interleukin - TNF tumour necrosis factor