bluuebunny/crossref_metadata_2025 huggingface dataset:
Large rearrangements of the LDL receptor gene and lipid profile in a FH Spanish population
Authors: Chaves F. J. , Real J. T. , García‐García A. B. , Puig O. , Ordovas J. M. , Ascaso J. F. , Carmena R. , Armengod M. E.
Published: March 2003

Background Familial hypercholesterolemia (FH) is an autosomal dominant disease caused by mutations in the low‐density lipoprotein receptor (LDLR) gene. To date, there has not been a systematic survey of the frequency of gross mutations in the LDLR gene in the Spanish population. The objective of our study was to investigate large rearrangements in the Spanish FH population and the relation between the kind of large rearrangement and the phenotype in carrier families.

Material and methods The LDLR gene was screened to detect major rearrangements in a sample of 89 probands. Southern blot, long polymerase chain reaction (PCR), reverse transcription (RT) ‐PCR and DNA sequencing were used to detect and characterize the mutations.

Results Five large rearrangements were found in six probands. Two mutations were due to duplications of internal regions of the gene, whereas the rest were caused by partial deletions, which eliminated the promoter region in two cases. The internal rearrangements, two duplications and one deletion, were apparently caused by recombination between ALU sequences and the study of their mRNA indicated that the reading frame was maintained. The analysis of the lipid profile between patients with similar characteristics (age, sex, body mass index, etc.) but carrying mutations that either eliminated the promoter region or produced internal rearrangements showed significant differences (total cholesterol: 366·6 ± 81·8 vs. 304·6 ± 25·1 P = 0·023, and LDL cholesterol: 317·7 ± 65·1 vs. 249·2 ± 27·4 P = 0·003).

Conclusions The frequency of large mutations in a Spanish FH sample was close to 7% and at least four of the mutations found had not been described in other populations. Mutations that eliminate the promoter region originate more severe hypercholesterolemia than defective mutations, which suggests that the absence of the promoter region and transcription of the LDLR gene is worse compensated than the synthesis of a defective LDL receptor.
Effect of nitric oxide inhibition on nasal airway resistance after nasal allergen challenge in allergic rhinitis
Authors: Maniscalco M. , Sofia M. , Carratù L. , Higenbottam T.
Published: March 2003

Background Nitric oxide has been detected by chemiluminescence in the lumen of nasal airway, which is increased in nasal breathing in patients with seasonal rhinitis during a chronic exposure. The purpose of this study was to determinate the effect of a NO‐synthase inhibitor NGL‐arginine methyl ester (L‐NAME) on nasal airway resistance (NAR) in patients with seasonal allergic rhinitis after an acute challenge to the allergen.

Methods Nitric oxide levels in the nose were measured by the chemiluminescence method in nine non‐atopic volunteers and in seven patients with seasonal rhinitis at rest and after an acute challenge with the allergen. NAR were measured by active anterior rhinomanometry.

Results Basal nasal NO concentration in allergic rhinitis was 496·5 ± 151·4 parts per billion (ppb). (n = 7) and it was not significantly different from levels found in the control group: 458·4 ± 105·9 ppb (n = 9). The topical administration of L‐NAME in allergic rhinitis reduced the NO concentration (338·6 ± 99·3 ppb, P < 0·001; n = 7). In the rhinitic patients the challenge with the allergen did not modify the nasal NO levels (504·5 ± 138·5 ppb). The application of the allergen after the pretreatment with placebo caused a significant increase in NAR (from 0·32 ± 0·11 Pa s cm−3 to 1·01 ± 0·12 Pa s cm−3, P < 0·001; n = 7). Pre‐treatment with L‐NAME did not prevent the increase in NAR induced by allergen challenge (from 0·36 ± 0·15 Pa s cm−3 to 1·06 ± 0·26 Pa s cm−3).

Conclusions The results indicate that nasal administration of a NOS inhibitor L‐NAME, at doses capable of decreasing nasal NO levels, has no effect on NAR and it does not prevent the NAR increase induced by an acute challenge with allergen in subjects with seasonal rhinitis.
Gene therapy for therapeutic angiogenesis in critically ischaemic lower limb – on the way to the clinic
Authors: Rissanen T. T. , Vajanto I. , Ylä‐Herttuala S.
Published: March 2003

Currently, no effective pharmacological treatment is available for vascularisation defects in lower limbs. Many patients presenting with persistent pain and ischaemic ulcers are not suitable candidates for surgical or endovascular approaches. Further refinement of the available methods will undoubtedly lead to a more active approach towards treatment of peripheral arterial occlusive disease (PAOD). Recently, therapeutic angiogenesis, in the form of recombinant growth factor administration or gene therapy, has emerged as a novel tool to treat these patients. However, improved gene transfer methods and better understanding of blood vessel formation are required to bring therapeutic angiogenesis to clinical practice. Here we review the clinical problem (PAOD), mechanisms of blood vessel formation (angiogenesis, vasculogenesis and arteriogenesis), experimental evidence and clinical trials for therapeutic angiogenesis in critically ischaemic lower limbs. Also, angiogenic growth factors, including vascular endothelial growth factors (VEGFs) and fibroblast growth factors (FGFs), delivery methods, and vectors for gene transfer in skeletal muscle, are discussed. In addition to vascular growth, gene transfer of growth factors may enhance regeneration, survival, and innervation of ischaemic skeletal muscle. Nitric oxide (NO) appears to be a key mediator in vascular homeostasis and growth, and a reduction in its production by age, hypercholesterolemia or diabetes leads to the impairment of ischaemic disorders.
Gemfibrozil treatment potentiates oxidative resistance of high‐density lipoprotein in hypertriglyceridemic patients
Authors: Chen M.‐F. , Wang T.‐D. , Yeh H.‐T. , Hsu H.‐C. , Lee Y.‐T.
Published: March 2003

Background Studies suggest that both oxidized low and high density lipoprotein (LDL and HDL) play a role in the pathogenesis of atherosclerosis. Gemfibrozil is widely used and is reported to increase cholesterol of LDL and HDL in hypertriglyceridemic patients. The aim of this study was to investigate the effect of gemfibrozil treatment on the oxidative status of lipoprotein particles in Fredrickson phenotype IV hypertriglyceridemic patients.

Methods Twenty‐two patients, aged 38–64 years, with fasting plasma triglyceride concentrations between 2·90 and 8·97 mmol L−1, were recruited and were given gemfibrozil 300 mg three times daily for 12 weeks. Venous blood samples were collected before gemfibrozil treatment, after 4, 8, or 12 weeks of treatment, and 4 weeks after termination of treatment, and used to analyse the plasma lipid profile, isolate lipoproteins, and analyse the chemical composition and in vitro oxidation of lipoprotein particles.

Results Gemfibrozil treatment resulted in a decrease in plasma total triglyceride levels and the triglyceride content of all lipoproteins. Plasma total cholesterol levels were decreased as a result of a decrease in very low density lipoprotein (VLDL) cholesterol levels. A slight increase in LDL cholesterol levels was observed, whereas the thiobarbituric acid‐reactive substances (TBARS) of LDL were decreased and the lag and peak time of LDL to oxidation were unchanged and maximal diene production was decreased. Plasma HDL cholesterol levels, the surface‐to‐core ratio of HDL particles, and the resistance of HDL to oxidation were increased.

Conclusion The decreased TBARS and diene production of LDL, increased HDL cholesterol levels, and increased resistance of HDL to oxidation may, in part, explain why gemfibrozil treatment was found to be generally beneficial in terms of protection against coronary heart disease.
CCK‐B/gastrin receptors in human colorectal cancer
Authors: Schmitz F. , Otte J.‐M. , Stechele H. U. , Reimann B. , Banasiewicz T. , Fölsch U. R. , Schmidt W. E. , Herzig K.‐H.
Published: March 2003

Background Mature amidated gastrin (G17 amide) mediates its effects in the gastrointestinal tract by activating G protein‐coupled CCK‐B/gastrin receptors. Although trophic actions of gastrin on the gastric mucosa have been well‐established, the effect of G17 amide, progastrin and intermediates to colon neoplasia in humans is controversial. While epidemiological evidence from patients with elevated serum gastrin levels related to pernicious anaemia does not support an increased risk for colon cancer [1], a recent study suggests that prolonged hypergastrinaemia is associated with an increased risk for colon cancer [2]. The extent to which trophic actions of gastrin in colorectal cancer are mediated by functional gastrin receptors remains to be defined. The aim of the present study was to determine CCK‐B/gastrin receptor expression, structure, and function in 79 patients with colon cancer.

Materials and methods CCK‐B/gastrin receptor cDNAs were isolated from 79 human colorectal cancer specimens and 15 control tissues, subcloned into the eukaryotic expression vector pCR3.1 and subjected to DNA sequence analysis. Wild‐type and mutant cDNAs were transiently expressed in COS‐7 cells to determine ligand affinities by 125I‐labelled CCK‐8S competition binding. Activation of the MAP kinase signalling cascade by G17 amide was determined in transfected Colo 320 cells expressing the wild‐type or mutant CCK‐B/gastrin receptors. Clonal expansion of single cells was quantified in transfected Colo 320 cells.

Results Gastrin mRNA is expressed in 44% of colorectal cancers and in 13% of control tissues. CCK‐B/gastrin receptor mRNA is expressed in 38% of colorectal cancers and 13% of normal colonic tissue. Co‐expression of gastrin and CCK‐B/gastrin receptor message is significantly increased in colorectal cancer specimens (32% vs. 0%). There is no correlation between CCK‐B/gastrin receptor expression and disease stage or histological grading. DNA sequence analysis revealed one spontaneous CCK‐B/gastrin receptor mutation within the third intracellular loop with an exchange of valine‐287 for phenylalanine. Pharmacological characterisation of the 287V → F CCK‐B/gastrin receptor reveals wild‐type affinities for G17 amide, glycine‐extended gastrin, CCK‐8S and L‐365,260. Mutation 287V → F is associated with a loss of gastrin‐induced MAPK p44/p42 signalling in Colo 320 cells while clonal expansion from single cells is increased by 53·1 ± 15·9% when compared to Colo 320 cells expressing wild‐type CCK‐B/gastrin receptors.

Conclusions Structural alterations of CCK‐B/gastrin receptors may account for increased growth‐promoting effects of amidated gastrins in colorectal cancer.
Heterogeneity of smooth muscle cells in advanced human atherosclerotic plaques: intimal smooth muscle cells expressing a fibroblast surface protein are highly activated by platelet‐released products
Authors: Martínez‐González J. , Berrozpe M. , Varela O. , Badimon L.
Published: March 2003

In vascular disease, smooth muscle cells (SMC) undergo phenotypic modulation and may acquire properties resembling those of fibroblasts in tissue wound healing.

We aimed to show the differential expression of a fibroblast surface protein (FSP) by SMC in atherosclerotic lesions.

In early human coronary atherosclerotic lesions the expression of FSP in the intima was absent. In contrast, 29 of 29 middle/advanced lesions contained intimal SMC expressing high levels of FSP. Fibroblast surface protein positive SMC were negative for desmin but expressed variable levels of α‐SM actin, SM caldesmon, SM myosin heavy chain and vimentin. Explants from advanced atherosclerotic lesions yielded two main SMC subpopulations. SMC over‐expressing FSP exhibited higher in vitro mitogenic response (premitotic DNA synthesis) to sera (2‐ to 8‐fold) and platelet‐released products (8‐ to 26‐fold), especially from thrombin‐activated platelets, than FSP‐negative SMC.

Our results suggest that the expression of FSP in SMC could indicate an activated phenotype, and the presence of highly positive FSP cells in the atherosclerotic lesions might be indicative of an increased SMC responsiveness to processes that locally generate thrombin and activate platelets.
Mixed cryoglobulinemia is associated with increased risk for death, or neoplasia in HIV‐1 infection
Authors: Kordossis T. , Sipsas N. V. , Kontos A. , Dafni U. , Moutsopoulos H. M.
Published: March 2003

Backround Cryoglobulinemia has been reported in several chronic infectious and autoimmune diseases, and in patients with HIV‐1 infection. Cryoglobulinemia associated with hepatitis C virus infection is considered a risk factor for the development of neoplasia, especially B‐cell non‐Hodgkin lymphoma. This study was undertaken to investigate whether the presence of circulating cryoglobulins is associated with survival or development of neoplastic disease in HIV‐1 infection.

Design We evaluated 87 unselected consecutive HIV‐1 infected patients for the presence of cryoglobulinemia and they were prospectively followed up for a median of 34 months, with clinic visits at 4‐month intervals. None of the patients had neoplasia at study entry. Time‐to‐event analysis for death, neoplasm and B‐cell lymphoproliferative disorder were performed with Cox proportional hazards models.

Results Mixed cryoglobulinemia (types II and III) was detected in 24 (28%) of the 87 patients. During the follow up, 12 patients died and 8 developed neoplastic disease. Multivariate analysis showed that circulating cryoglobulins were an independent predictor of death [relative risk (RR), 4·97; 95% confidence intervals (CI), 1·26–19·63] and development of neoplasia (RR, 5·18; 95% CI, 1·23–21·83). In addition, cryoglobulinemia reached borderline significance as a predictor of lymphoproliferative disorder of B‐cell origin (P = 0·08; RR, 4·53; 95% CI, 0·83–24·75).

Conclusions Our results suggest that cryoglobulinemia is associated with an increased risk for death, neoplasia or development of lymphoproliferative disorder of B‐cell origin, in HIV‐1 infected patients.
Procion orange tracer dye technique vs. identification of intrafibrillar fibronectin in the assessment of sarcolemmal damage
Authors: Palacio J. , Gāldiz J. B. , Alvarez F. J. , Orozco‐Levi M. , Lloreta J. , Gea J.
Published: March 2003

Background The use of Procion orange dye (POD) is one of the most widely accepted techniques to assess sarcolemmal damage. This phenomenon has been related to functional adaptation in skeletal muscles. The POD method includes intravenous injection of this colorant in vivo, enabling its identification inside those fibres with membrane leaks (fluorescence). However, the safety of the use of POD has not been proven.

Aim This study was designed to compare POD with a safer alternative, involving the identification of intracellular fibronectin using specific antibodies.

Method Eight Swiss mice were submitted to electrical stimulation of the lower limbs at different frequencies (10–80 Hz). Subsequently, the POD solution was infused, and samples from the vastus medialis muscle were obtained 24 h later. Samples were processed and serial sections were analysed using immunohistochemistry (monoclonal antibodies against fibronectin) and epifluorescence microscopy.

Results Ninety‐eight per cent of the fibres were equally classified by both techniques, which in addition showed good correlation (percentages of damaged fibres, r = 0·998, P < 0·001) and concordance (R1 = 0·82) in quantitative terms.

Conclusions Although the two techniques compared here are based on different principles, both are comparable in assessing sarcolemmal damage. This would facilitate comparisons between human and experimental studies. In addition, the fibronectin technique appears to be a suitable alternative for long‐term studies including repeated biopsies.
Growth hormone replacement therapy induces codeine clearance
Authors: Gil Berglund E. , Johannsson G. , Beck O. , Bengtsson B‐Å. , Rane A.
Published: March 2003

Background The increasing clinical use of growth hormone (GH) has raised questions about other than growth‐related metabolic effects of this treatment. GH regulates the expression of several hepatic drug metabolising enzymes in the rat, but it is not known whether GH treatment alters the expression of such liver enzymes in man. We have investigated the effects of GH on codeine clearance and two enzymes of the cytochrome P450 (CYP) family, CYP3A and CYP2D6, and UDP‐glucuronosyl transferase (UDPGT). These enzymes have a superior importance in hepatic biotransformation of numerous drugs. In addition, CYP3A and UDPGT are catalysts of many reactions with endobiotics such as steroid hormones.

Methods We used codeine as a probe drug for assessment of the enzyme activities. Codeine was administered as a single‐dose prior to, and after 3 months of GH substitution in GH‐deficient patients. Total clearance, and clearance along each of the three primary metabolic pathways of codeine, was assessed.

Results Three months of GH substitution increased the total clearance of codeine (21%, P < 0·01) and clearance catalysed by UDPGT significantly (31%, P < 0·05). The treatment tended to increase the clearance via the CYP3A pathway (83%, P = 0·05).

Conclusions The effects of GH replacement therapy on drug metabolism may have clinical implications when combined with drugs that are substrates of UDPGT and CYP3A. Effects on steroid hormone metabolism with endocrine consequences can not be ruled out.
Nitric oxide attenuates platelet‐activating factor‐induced nasal airway plasma extravasation in healthy subjects
Authors: Maniscalco M. , Ferrara G. , Carratù L. , Sofia M.
Published: March 2003

Background Paranasal sinuses and the nose are important sources of nitric oxide (NO) in humans but the relevance of NO production to the control of nasal airway plasma exudation and its response to inflammatory mediators such as platelet‐activating factor (PAF) in healthy subjects is not well known.

Design In this study we aimed to evaluate the effect of the nitric oxide synthase (NOS) inhibitor NGl‐arginine methyl ester (l‐NAME) on nasal airway plasma extravasation at baseline and after an acute challenge with PAF that induces most symptoms of rhinitis. Eleven healthy subjects were enrolled in the study. Plasma extravasation in the nasal airway was assessed by measuring the albumin content of nasal lavage.

Results PAF challenge caused a significant increase in concentrations of albumin in the nasal lavage fluid (from 0·59 ± 0·13 mg dL−1 to 2·46 ± 0·45 mg dL−1) after placebo. Pretreatment with l‐NAME significantly prevented the increase of albumin in the nasal lavage fluid induced by PAF as compared to placebo (from 0·53 ± 0·11 mg dL−1 to 1·70 ± 0·28 mg dL−1; P < 0·005).

Conclusion Topical administration of a NO inhibitor is able to attenuate the nasal airway plasma extravasation induced by PAF, suggesting that NO release in vivo is involved in the nasal response to PAF.
Methylenetetrahydrofolate reductase gene C677T mutation is related to the defects in the internal elastic lamina of the artery wall
Authors: Hämelahti P. , Järvinen O. , Sisto T. , Wirta V. , Ilveskoski E. , Koivula T. , Lehtimäki T.
Published: March 2003

Background The C677T mutation of the methylenetetrahydrofolate reductase (MTHFR) gene leads to C/C, C/T and T/T genotypes, which affect the plasma homocysteine concentration in humans. In mini‐pigs, high serum homocysteine levels are associated with defects in the internal elastic lamina (IEL) of the artery wall, which are apparently related to the migration of smooth muscle cells into the intima during atherogenesis. We studied the association between the MTHFR genotypes and the number of gaps in the IEL in the wall of the five major abdominal arteries.

Materials and methods The autopsy study included 123 subjects (90 males and 33 females) aged 18–93. For the light microscopy, a 0·5 cm circular segment of the coeliac, the superior mesenteric, the inferior mesenteric and the renal arteries were cut and embedded in paraffin blocks. The circumference of the IEL, the thickness of the intima and the number of the gaps per millimetre in the IEL were measured by MOP 3 image analysis.

Results The T‐allele carriers (C/T and T/T) of the MTHFR gene had significantly less gaps in the IEL than the subjects with the C/C genotype in the superior mesenteric and in the left renal arteries (2·02 ± 2·25 vs. 2·53 ± 1·89, P < 0·04 and 0·56 ± 1·09 vs. 1·82 ± 2·66, P < 0·02, respectively). The trend was similar for the coeliac and the right renal arteries.

Conclusions Our result suggests that MTHFR polymorphism may be involved in the fragmentation of the IEL.
G‐455A polymorphism of the fibrinogen beta gene and deep vein thrombosis
Authors: Renner W. , Cichocki L. , Forjanics A. , Köppel H. , Gasser R. , Pilger E.
Published: March 2003

Background Elevated fibrinogen levels have been linked to increased risk for deep venous thrombosis, although it is not clear whether fibrinogen is causal or rather a marker for the presence of other risk factors. A common G/A polymorphism in the gene for the fibrinogen beta‐chain (FGB G‐455A) is associated with elevated fibrinogen levels. The present study was designed to analyze the role of this genetic marker for deep venous thrombosis.

Materials and Methods We performed a case–control study including 307 patients with documented deep venous thrombosis and 316 control subjects. β‐fibrinogen genotypes were determined by allele‐specific polymerase chain reaction.

Results GG, GA and AA genotype frequencies were similar among the patients (53·1%, 41·0, 5·9) and controls (51·6%, 42·1, 6·3; P = 0·92). Fibrinogen levels of the patients (median 3·72 g l−1; range 1·93–11·6) did not differ significantly from those of the controls (3·76; 2·17–9·99). Carriers of the homozygous AA genotype had significantly higher fibrinogen levels than noncarriers (patients: 5·32 vs. 3·59; P = 0·024; controls: 6·29 vs. 3·72; P = 0·048).

Conclusion Our data suggest that the fibrinogen‐elevating FGB G‐455A gene polymorphism is not linked to an increased risk for deep venous thrombosis.
Is the gender difference in LDL size explained by the metabolic complications of visceral obesity?
Authors: Lemieux I. , Pascot A. , Lamarche B. , Prud’homme D. , Nadeau A. , Bergeron J. , Després J.‐P.
Published: March 2003

Background Several studies have reported a significant gender difference in low‐denstiy lipoprotein (LDL) size, with men being characterized by smaller, denser LDL particles than women, and it has been suggested that the contribution of the greater accumulation of visceral adipose tissue in men compared with women may be a factor potentially contributing to the gender difference in LDL heterogeneity.

Materials and methods We measured LDL particle size by 2–16% nondenaturing polyacrylamide gradient gel electrophoresis in 299 men and 231 women in whom visceral adipose tissue accumulation was measured by computed tomography. A fasting plasma lipoprotein‐lipid profile was also obtained in all subjects.

Results Overall, the men were characterized by a more deteriorated metabolic risk factor profile, which included higher plasma insulin and triglyceride levels, a greater visceral adipose tissue accumulation (P < 0·001) and smaller LDL particles (251·7 ± 5·2 vs. 254·4 ± 4·2 Å, P < 0·0001). This gender difference in LDL peak particle diameter remained significant (252·4 ± 4·3 vs. 253·5 ± 4·3 Å, P < 0·01) after adjustment for sex‐specific differences in plasma triglyceride levels by covariance analysis. Significant negative correlations were noted between the LDL particle diameter and the triglyceride concentrations in both genders (r = −0·52 and r = −0·36, P < 0·0001 for the men and women, respectively), with no gender difference in this relationship being found. However, viscerally obese women (visceral adipose tissue levels > 100 cm2) with increased plasma triglyceride concentrations (> 2·0 mmol L−1) still had larger LDL particles than viscerally obese men with a similar elevation in their triglyceride levels (251·6 ± 4·9 vs. 248·7 ± 4·5 Å, P < 0·01).

Conclusions Results of the present study suggest that the reduced LDL particle size observed in men compared with women cannot be entirely explained by their higher visceral adipose tissue accumulation and increased plasma triglyceride levels. Moreover, the gender difference in LDL size could be influenced, at least in part, by the severity of the hypertriglyceridaemic state.
Differential expression of α‐ and β‐defensins in human peripheral blood
Authors: Fang X.‐M. , Shu Q. , Chen Q.‐X. , Book M. , Sahl H.‐G. , Hoeft A. , Stuber F.
Published: March 2003

Background  Human defensin peptides with broad‐spectrum antimicrobial activity have been implicated in the human defence response towards microbial invasion. Two families of defensins designated α‐ and β‐defensins, respectively, have been identified. Little is known about the expression of both defensin families in human peripheral blood. The purpose of this study was to examine the expression of α‐ and β‐defensin genes in human peripheral blood.

Material and methods  Fifty‐one healthy blood donors were screened for defensin expression. Blood from defensin responders was stimulated by lipopolysaccharide or heat‐inactivated Pseudomonas aeruginosa ex vivo. Levels of mRNA were assessed by semiquantitative RT‐PCR. Southern blot analysis and sequencing were used to confirm the identity of defensin gene transcripts. Western blotting analysis was used to detect the expression of defensin peptides.

Results  β‐defensin was undetected in human peripheral blood without stimulation. Following stimulation by lipopolysaccharide or heat‐inactivated bacterial cells, the majority (88·2%) of healthy individuals had a detectable expression for β‐defensin‐1 gene and 39·2% for β‐defensin‐2 gene, compared with none for β‐defensin‐3. β‐defensin‐1 and ‐2 mRNAs in the stimulated human peripheral blood of responders became detectable at 3 h and showed a maximum at 6 h following induction by 100 ng mL−1 of lipopolysaccharide or bacterial cells. In contrast, human α‐defensins 1–3 mRNA are constitutively expressed in peripheral leukocytes but not up‐regulated by lipopolysaccharide or bacterial cells.

Conclusions  In human peripheral blood, β‐defensin‐1 and ‐2 genes were transiently transcribed and translated following the induction of lipopolysaccharide or heat‐inactivated bacterial cells, whereas α‐defensins 1–3 genes were constitutively transcribed, and β‐defensin‐3 gene was not expressed. The inducible expression of β‐defensin‐1 and ‐2 genes showed interindividual variability.
Lipopolysaccharide represses cholesterol 7‐alpha hydroxylase and induces binding activity to the bile acid response element II
Authors: Dikopoulos N. , Weidenbach H. , Adler G. , Schmid R. M.
Published: March 2003

Background Inflammatory states such as hepatitis and sepsis are frequently associated with alterations of bile acid synthesis. These conditions are mediated by bacterial wall products like lipopolysaccharide (LPS). Cholesterol 7‐alpha hydroxylase is the rate‐limiting enzyme of bile acid synthesis. Hydrophobic bile acids repress cholesterol 7‐alpha hydroxylase transcription via binding to the farnesoid X receptor and interaction with the bile acid response element II in the cholesterol 7‐alpha hydroxylase promoter.

Methods We tested the effect of LPS on hepatic expression of cholesterol 7‐alpha hydroxylase in C57BL/6 mice and Wistar rats. Further, we analyzed the binding activity of hepatic nuclear extracts to the bile acid response element II and the binding site for farnesoid X receptor heterodimers (ecdysone response element).

Result Lipopolysaccharide caused a 100‐fold reduction of cholesterol 7‐alpha hydroxylase mRNA levels in mice and a 10‐fold reduction in rats. Protein levels of cholesterol 7‐alpha hydroxylase also decreased in both species. These changes inversely correlated with the increased binding activity of nuclear proteins to the bile acid response element II and the ecdysone response element.

Conclusion Lipopolysaccharide‐induced repression of cholesterol 7‐alpha hydroxylase occurs at the transcriptional level. The underlying mechanism involves an increased binding activity of nuclear proteins to the bile acid response element II and the ecdysone response element. In conclusion, the farnesoid and retinoid X receptors participate in LPS‐induced cholesterol 7‐alpha hydroxylase repression.
No acute impact of lipid apheresis treatment on free radical scavenging enzyme gene expression in white blood cells
Authors: Schettler V. , Krontal J. , Scheel A. , Wieland E.
Published: March 2003

Background Lipid apheresis (LA) treatment has been suggested to cause oxidative stress. Defense against oxygen‐radical‐mediated damage is provided by nonenzymatic and enzymatic antioxidants. In the present investigation we have investigated whether gene expression of free radical scavenging enzymes (FRSE) is affected in leukocytes of patients undergoing LDL‐apheresis.

Materials and methods For this purpose cellular glutathione peroxidase (GPx‐1), phospholipid glutathione peroxidase (GPx‐4), glutathione reductase (GSSG‐R), glutathione synthetase (GSH‐S), Cu/Zn‐superoxide dismutase (SOD‐1) and catalase (CAT) mRNA expression were followed at the start (SA) and immediately after (EA) LA treatment (n = 25). Gene expression was determined by quantitative RT‐PCR with the LightCycler® instrument (Roche Diagnostics, Mannheim, Germany) and transcription elongation factor‐2 as reference gene.

Results The expression of GPx‐1, GPx‐4, GSSG‐R, GSH‐S, SOD‐1, CAT mRNA was not affected by a single LA treatment. Free radical scavenging enzymes mRNAs were significantly (P < 0·05) increased in the LA patients (GPx‐1: 2·00 ± 1·37; GPx‐4: 0·52 ± 0·46; GSSG‐R: 0·07 ± 0·03; GSH‐S: 0·04 ± 0·03; SOD‐1: 1·12 ± 0·74; CAT: 0·15 ± 0·07) when compared with 26 healthy blood donors (GPx‐1: 1·1 ± 0·6; GPx‐4: 0·35 ± 0·19; GSSG‐R: 0·02 ± 0·01; GSH‐S: 0·03 ± 0·01; SOD‐1: 0·16 ± 0·08; CAT: 0·09 ± 0·05; mean ± SD).

Conclusions These results show that the LA procedure does not acutely affect the antioxidant defense system on the gene level but suggests that the chronic stress resulting from hyperlipidaemia and/or LA may cause FRSE gene induction.
Molecular insights into insulin action and secretion
Authors: Rhodes C. J. , White M. F.
Published: March 2003

Tightly co‐ordinated control of both insulin action and secretion is required in order to maintain glucose homeostasis. Gene knockout experiments have helped to define key signalling molecules that affect insulin action, including insulin and insulin‐like growth factor‐1 (IGF‐1) receptors, insulin receptor substrate (IRS) proteins and various downstream effector proteins. β‐cell function is also a tightly regulated process, with numerous factors (including certain signalling molecules) having an impact on insulin production, insulin secretion and β‐cell mass. While signalling molecules play important roles in insulin action and secretion under normal circumstances, abnormal insulin signalling in muscle, adipose tissue, liver and pancreas leads to insulin resistance and β‐cell dysfunction. In particular, the signalling protein IRS‐2 may have a central role in linking these abnormalities, although other factors are likely to be involved.
Tumour necrosis factor β gene polymorphisms in myasthenia gravis
Authors: Zelano G. , Lino M. M. , Evoli A. , Settesoldi D. , Batocchi A. P. , Torrente I. , Tonali P. A.
Published: March 2003

Genetic analyses indicate that genes within the major histocompatibility complex (MHC) can be involved in susceptibility to autoimmune disease. To investigate the role of the tumour necrosis factor beta (TNFB) gene in myasthenia gravis (MG) susceptibility, we analysed an NcoI polymorphism within the TNFB gene in 63 MG patients and 93 healthy individuals. When patients were subdivided according to thymic pathology, we found differences between MG patients with thymic hyperplasia and thymoma versus controls. In MG patients with thymic hyperplasia we found a positive association with the TNFB*1 allele [Relative risk (RR): 2.6; P<0.001] and phenotype (RR: 1.8; P<0.005) and a negative association with the TNFB*2/2 genotype (RR: 0.2; P<0.001) when compared to the controls. On the other hand, in MG patients with thymoma we found a positive association with the TNFB*2/2 genotype (RR: 5.6; P<0.01) and a negative association with the TNFB*1 allele (RR: 0.3; P<0.05) and *1/2 genotype (RR: 0.2; P<0.01). These data suggest that the two different forms of MG can have different pathogenesis and that the TNFB gene could influence susceptibility to MG.
Two novel biallelic polymorphisms in the IL‐2 gene
Authors: John S. , Turner D. , Donn R. , Sinnott P. , Worthington J. , Ollier W. E. R. , Hutchinson I. V. , Hajeer A. H.
Published: March 2003

We have detected two novel single nucleotide polymorphisms in the IL‐2 gene, at positions –330 and +166 relative to the transcription start site. The +166 change occurs within the leader peptide and does not affect amino acid sequence. The –330 polymorphism has two common alleles, making it an ideal marker for genetic association studies.
Analysis of the polymorphism of the tumour necrosis factor (TNF) gene and promoter and of circulating TNF‐alpha levels in heart‐transplant patients suffering or not suffering from severe rejection
Authors: Abdallah A. N. , Cucchi‐Mouillot P. , Biteau N. , Cassaigne A. , Haras D. , Iron A.
Published: March 2003

Plasma TNF‐α levels are generally higher in heart‐graft patients who experience a rejection episode than in those who do not. Because the TNF gene and its promoter are polymorphic, we studied the relationships between genetic variability at the TNF locus, the occurrence of graft rejection and TNF‐α plasma levels in 62 heart‐transplant patients in order to investigate inter‐individual differences in plasma TNF‐α levels after allogeneic stimulation. TNF‐α was immunoenzymatically measured in blood specimens collected on the same day as endomyocardial biopsy. After PCR amplification of DNA, NcoI and AspHI polymorphisms were characterized by their restriction profiles, TNFa microsatellites by electrophoretic separation on acrylamide and the promoter region by sequencing. Plasma levels and molecular genetic results were compared to the grade of heart graft rejection established according to pathological criteria. In our study, allograft rejection was associated neither with NcoI or AspHI polymorphism nor with nucleotide changes in the TNF‐Α promoter. We observed low TNF‐α levels in n1/n1 homozygous patients and in subjects with G→A at position – 308 of the promoter sequence. Concerning the polymorphism of the TNFa microsatellite, our results might suggest an association with graft rejection but we have to be very careful in drawing conclusions because of the small size of the sample.
Susceptibility to coeliac disease in Tunisian children and GM immunoglobulin allotypes
Authors: Bouguerra F. , Dugoujon J. M. , Babron M. C. , Greco L. , Khaldi F. , Debbabi A. , Bennaceur * B. , Clerget‐Darpoux F.
Published: March 2003

Coeliac disease is a malabsorption disorder of the small intestine resulting from ingestion of gluten. The immunogenetic component is clearly demonstrated by the association of the disease with human leukocyte antigens (HLA). Among other candidate genes are the GM allotypes, which are the markers of the constant parts of heavy chains of the subclasses IgG1, IgG2 and IgG3. GM immunoglobulin allotypes have been analysed in 131 unrelated Tunisian children with coeliac disease. All patients and their parents were tested for G1M(1, 2, 3, 17), G2M(23) and G3M(5, 6, 10, 11, 13, 14, 15, 16, 21, 24, 28) by the classical haemagglutination method. Genotypes and haplotypes were deduced from phenotypes in patients and their parents. Transmission disequilibrium tests were performed in 79 informative families. The GM*3;..;5* haplotype was transmitted more often (23) than not (8) by heterozygous parents (χ2 = 7.26; P = 0.007). This difference remained significant after correction for multiple testing. This study provides evidence for association and linkage between GM and coeliac disease. It suggests that GM or genes close to GM play a role in the development of the disease.
Complex interaction between HLA DR and DQ in conferring risk for childhood type 1 diabetes
Authors: Kockum I. , Sanjeevi C. B. , Eastman S. , Landin‐Olsson M. , Dahlquist G. , Lernmark Å.
Published: March 2003

Type 1 (insulin‐dependent) diabetes mellitus is associated with HLA DR and DQ factors, but the primary risk alleles are difficult to identify because recombination events are rare in the DQ–DR region. The risk of HLA genotypes for type 1 diabetes was therefore studied in more than 420 incident new onset, population‐based type 1 diabetes children and 340 age, sex and geographically matched controls from Sweden. A stepwise approach was used to analyse risk by relative and absolute risks, stratification analysis and the predispositional allele test. The strongest relative and absolute risks were observed for DQB1*02‐DQA1*0501/DQB1*0302‐DQA1*0301 heterozygotes (AR 1/46, P < 0.001) or the simultaneous presence of both DRB1*03 and DQB1*0302 (AR 1/52, P < 0.001). Stratification analysis showed that DQB1*0302 was more frequent among DRB1*04 patients than DRB1*04 controls (P < 0.001), while DRB1*03 was more frequent among both DQA1*0501 (P < 0.001) and DQB1*02 (P < 0.001) patients than respective controls. The predispositional allele test indicated that DRB1*03 (P < 0.001) would be the predominant risk factor on the DRB1*03‐DQA1*0501‐DQB1*02 haplotype. In contrast, although DQB1*0302 (P < 0.001) would be the predominant risk factor on the DRB1*04‐DQA1*0301‐DQB1*0302 haplotype, the predispositional allele test also showed that DRB1*0401, but no other DRB1*04 subtype, had an additive risk to that of DQB1*0302 (P < 0.002). It is concluded that the association between type 1 diabetes and HLA is due to a complex interaction between DR and DQ since (1) DRB1*03 was more strongly associated with the disease than DQA1*0501‐DQB1*02 and (2) DRB1*0401 had an additive effect to DQB1*0302. The data from this population‐based investigation suggest an independent role of DR in the risk of developing type 1 diabetes, perhaps by providing diseases‐promoting transcomplementation molecules.
HLA class II polymorphism in Thai patients with non‐Hodgkin’s lymphoma
Authors: Nathalang O. , Tatsumi N. , Hino M. , Prayoonwiwat W. , Yamane T. , Suwanasophon C. , Sriphaisal T.
Published: March 2003

The distribution of HLA‐DRB1 alleles and DQB1 alleles in 100 Thai patients with non‐Hodgkin’s lymphoma (NHL) was analysed using the polymerase chain reaction with sequence‐specific primer (PCR–SSP) method, and the association between the disease and the presence of certain HLA class II alleles was investigated. The frequencies of HLA‐DRB1*1502 and DRB1*09012 were increased while those of DRB1*0404, DRB1*0803 and DRB1*1106 were decreased. On the other hand, the incidence of HLA‐DQB1 alleles was similar to that in the normal population. Interestingly, only HLA‐DRB1*1502 showed a significant positive association with NHL, especially in patients ≤ 45 years and in male patients. It is concluded that the DRB1*1502 allele may contribute to NHL susceptibility in the Thai population. However, further studies on the functional roles of the HLA class II alleles are necessary to elucidate NHL susceptibility.
Further characterization of MHC haplotypes demonstrates conservation telomeric of HLA‐A: update of the 4AOH and 10IHW cell panels
Authors: Cattley S. K. , Williamson J. F. , Tay G. K. , Martinez O. P. , Gaudieri S. , Dawkins R. L.
Published: March 2003

Cell panels have been used extensively in studies of polymorphism and disease associations within the major histocompatibility complex (MHC), but the results from these panels require continuous updates with the increasing availability of novel data. We present here an updated table of the typings of the 10IHW and 4AOH panels. Local data included are HFE, HERV‐K(C4) and six microsatellites telomeric of HLA‐A. Typings for class I, MICA (PERB11.1), MICB (PERB11.2), XA, XB, LMP2 and 10 microsatellites reported by others have also been consolidated in this table. The tabulation shows that the length of conservation in the human MHC is even more extensive than previously thought. Human MHC ancestral haplotypes are inherited as a conserved region of genomic sequence spanning some 6–8 megabases from the HLA class II region and beyond the HLA class I region up to and including the HFE gene. Numerous examples of historical recombinations were also observed.
Molecular variation of HLA class I genes in the Corsican population: approach to its origin
Authors: Grimaldi M.‐C. , Crouau‐Roy B. , Contu L. , Amoros J.‐P.
Published: March 2003

Three human leucocyte antigen (HLA) class I loci (HLA‐A, ‐B and ‐Cw) were typed for the first time at the DNA level in the Corsican population. This analysis was performed on 100 individuals of Corsican origin living in central Corsica (46 individuals) and in south‐western Corsica (54 individuals). The genetic structure of these two subpopulations was analysed on the basis of the molecular polymorphisms of the HLA‐A, ‐B and ‐Cw genes. A phylogenetic and a haplotypic analysis were performed. The genotypic analysis did not detect genetic differentiation. Examination of the allelic and haplotypic frequencies did, however, reveal some significant differences between the two zones. Similarities with the Sardinian population were found, and were particularly evident in the south‐western sample. However, Corsica has probably been subject to greater West European influence, which can be seen in the Corte sample (Haute Corse).
Asthma and IL‐4 receptor alpha gene variants
Authors: Wjst M. , Kruse S. , Illig T. , Deichmann K.
Published: March 2003

Linkage of allergy to chromosome 16 has been described in several studies, together with a positive association with interleukin 4 receptor α gene variants. Our aim was to replicate these findings in a sample of German and Swedish families recruited through sib‐pairs affected by bronchial asthma. None of the markers showed linkage with the main phenotype of asthma or with total serum IgE. Seropositivity to D. pteronyssinus showed borderline significance in a region flanking the IL4Rα location. Single nucleotide polymorphisms (SNPs) leading to the protein exchanges I50V, E375A, C406R, S478P and Q551R in the IL‐4 receptor α were examined for allele sharing in sibs with asthma. Multiple regression analysis was performed for association with total serum IgE and specific IgE. Allele sharing of IL4Rα SNPs in asthmatic children was not significantly increased for any of the examined SNPs except for the intracytoplasmatic polymorphism 551R (0.79 vs. 0.84 expected, P = 0.044). The variants 50V, 478P and 551R were associated with slightly increased, and 375A and 406R with decreased total IgE levels, all at a non‐significant level. None of the examined IL4Rα variants were correlated to asthma severity. In summary, a single gene effect of IL4Rα variants or any other gene on chromosome 16 could not be shown in this selected population of children with asthma. As there could be interactions with multiple genetic and environmental factors, IL4Rα could still be involved in asthma pathogenesis.
Frequencies of the ‐330 (T → G) IL‐2 and ‐590 (T → C) IL‐4 gene polymorphisms in a population from south‐eastern Brazil
Authors: Scarel‐Caminaga R. M. , Trevilatto P. C. , Souza A. P. , Brito R. B. , Line S. R. P.
Published: March 2003

Polymorphisms in the promoter regions of cytokine genes may affect their transcription. A T/G substitution at position −330 of the interleukin‐2 (IL‐2) gene and a T/C substitution at position −590 of the interleukin‐4 (IL‐4) gene have been described previously. The −590 (T → C) IL‐4 gene polymorphism was associated with asthma and atopy in US and Japanese populations. Population genetics is a useful tool for determination of the biological significance of genetic polymorphisms. The aim of this study was to investigate the frequencies of polymorphisms in the promoter regions of the IL‐2 and IL‐4 genes in a population from south‐eastern Brazil and to compare them with those published for other populations. Allele frequencies were estimated in 114 unrelated individuals from São Paulo State. These subjects had an average age of 41.2 years (± 12.4 years) and the ethnic composition of the sample was: 78.07% Caucasian, 11.4% Black and 10.53% Mulatto. DNA from subjects was extracted from epithelial buccal cells, and the PCR‐RFLP technique was employed to investigate the −330 (T → G) IL‐2 and −590 (T → C) IL‐4 gene polymorphisms. The allele frequency of the IL‐2 gene polymorphism obtained in our study was similar to that found in UK Caucasoid groups. The T allele frequency of the IL‐4 gene polymorphism observed in the Caucasian Brazilian group was similar to that found in UK and Australian populations, while the frequency observed for the Black Brazilian group was similar to that found in Japanese and Kuwaiti Arab populations. The results for the −330 (T → G) IL‐2 and −590 (T → C) IL‐4 polymorphisms are consistent with the high contribution of European lineages to the population in south‐eastern Brazil.
Soil organic matter pools in a volcanic‐ash soil under fallow or cultivation with applied chicken manure
Authors: GIJSMAN A. J. , SANZ J. I.
Published: March 2003

Frequent applications of chicken manure to a volcanic‐ash soil resulted in an increased crop yield several years after the applications ceased. Improved nutrient cycling through the soil organic matter (SOM) was thought be the cause of this. An area that for several years had been under either unimproved fallow or continuous bean‐maize intercropping (with chicken manure application) was planted with maize intercropped with the legume Arachis pintoi. Soil samples were fractionated into several sizes of aggregates and incubated for 6 weeks in intact form and after crushing the aggregates. Between 1.2 and 3.1% of the C in any aggregate size class exited in easily decomposable (i.e. not physically or chemically protected) form. Less than 0.8% of the macroaggregate C was physically protected by the aggregate structure. More than 97% of the C in any aggregate size class can be considered resistant. The large‐or small‐macroaggregate‐protected SOM (i.e. in aggregates > 2000 μm and 250–2000 μm, respectively). once made available, was more easily decomposed than the unprotected SOM. The large SOM content (45.8–57.3 g C kg−1) but yet limited mineralizability indicates that mechanisms other than soil macroaggregation are important for protecting SOM in this soil. Binding of organic molecules onto allophane minerals is likely to be such a mechanism.

The larger yield and nutrient uptake by the maize in the former bean‐maize plots compared with the former fallow plots could not be explained by differences in SOM decomposition. We think that the frequent application of chicken manure to the former bean‐maize plots increased the available phosphorus in these strongly P‐sorbing soils by increasing the cycling of organic‐P or by blocking some of the P‐reactive sites. This, however, needs to be further investigated.