Stromelysin-1 polymorphism as a new potential risk factor in progression of chronic obstructive pulmonary disease.

BACKGROUND: Chronic obstructive lung disease (COPD) is characterised by partially reversible usually progressive airflow limitation caused by inflammation and remodelling. Stromelysin-1 (MMP-3) has regulatory activity on other matrix-metalloproteinases. Altered MMP-3 activity has been described in different diseases. We investigated the role of a promoter MMP-3 polymorphism in determining susceptibility and severity of COPD. METHODS: We studied 147 patients with COPD in stable conditions and distinguished two groups based on FEV1 values. In 100 patients functional modifications across a two-year period were noted. 133 healthy subjects were used as controls. Genotyping for the -1171 5A/6A MMP-3 polymorphism was performed using nucleotide sequencing. RESULTS: No difference was noted in the genotype distribution between COPD patients and controls. However, among patients with severe disease 6A/6A genotype and 6A allelic frequency were significantly more represented than among mild-moderate patients (p < 0.05). The 6A/6A genotype was also associated with a higher FEV1 decline over time. CONCLUSIONS: Our data suggests that -1171 6A allele does not represent a risk factor for the development of COPD while it is associated with more severe disease and different functional decline. We hypothesis that a disregulation of MMP-3, possibly caused by the -1171 5A/6A polymorphism or other linked variants, may lead to different progression in COPD.

Ser128Arg gene polymorphism for E-selectin and severity of atherosclerotic arterial disease.

AIM: Genetic factors appear to be important in the pathogenesis of cardiovascular and cerebrovascular disease. Adhesion molecules like the members of the selectin family participate in the interaction between leukocytes and the endothelium. They are also involved in the pathogenesis of the atherosclerotic process. In E-selectin, exchange from serine to arginine (position 128, S128R) is correlated with early atherosclerosis. The aim of this study was to assess E-selectin Ser128Arg polymorphism in subjects with clinical and instrumental evidence of atherosclerosis and to analyze the correlations with clinical severity. METHODS: A total of 144 subjects (100 men and 44 women, mean age 72 years, range 48-78) with atherosclerotic disease in different vascular sites documented by angiography were studied; 138 volunteers were recruited as a control group. Whole blood was collected; DNA was extracted with a commercial kit and amplified with 2 primers. The PCR was performed by standard procedure. To assess the disease severity all patients were classified by an arbitrary clinical and angiographic score scale. RESULTS: The genotype distribution between patients and controls was different, although statistical significance was not achieved (p=0.06). In patients a significant difference in Arg allele frequency was observed between mild and severe atherosclerotic disease (OR 2.28; 95% CI 1.15-4.52; p=0.017). Four ho-mozygous cases for S128R were found in patients, none in controls. All these 4 patients had the highest severity score, that means a more severe atherosclerotic disease. CONCLUSION: Our study suggests that the E-selectin polymorphism may be associated with severity of atherosclerotic disease, but does not allow us to conclude that it is actually a risk factor for atherosclerosis.

Phenylalanine hydroxylase mutations and phenylalanine-tyrosine metabolism in heterozygotes for phenylalanine hydroxylase deficiency.

UNLABELLED: The aim of this study was to determine whether any relationship exists between the severity of mutation of the phenylalanine hydroxylase (PAH) gene and the plasma concentrations of phenylalanine (Phe) and tyrosine (Tyr) under fasting and semifasting conditions among heterozygotes in a matched case-control study. Parents of patients affected by PAH deficiency (n = 25) detected through the Italian Neonatal Screening Program and referred from January 1994 to June 2000, and parents of healthy children were investigated. In total, 68 subjects without any disease, 34 hyperphenylalaninaemia (HPA) heterozygous parents and 34 age- and gender-matched controls, were recruited. Plasma concentrations of Phe and Tyr in fasting and semifasting (1600 mg Phe oral load) conditions were the main outcome measures. DNA analysis for PAH mutations was performed in all 68 subjects. Compared with controls, heterozygotes showed higher fasting and semifasting Phe concentrations (p < 0.0001), lower semifasting Tyr concentrations (p = 0.015), lower Tyr variations (p = 0.003) and a higher Phe/Tyr ratio (p < 0.0001) in switching from fasting to semifasting conditions. Heterozygotes carrying a severe mutation showed semifasting plasma Tyr concentrations lower than controls (p = 0.019) but not significantly different from Tyr levels found in non-severe carriers (p = 0.197). The Tyr variations were minor in severe carriers than controls (p < 0.001) and non-severe carriers too, although with lower significance (p = 0.089). In six carriers of A403V mutation, parents of mild hyperphenylalaninaemics on an unrestricted diet, significant differences in variations from fasting to semifasting conditions were found compared with parents of patients on a diet. CONCLUSION: Although the great heterogeneity of PAH mutations limits any general conclusion, the results suggest that monitoring plasma Tyr variations may be more sensitive than plasma Phe in assessing the severity of PAH mutations in HPA heterozygotes.

Matrix metalloproteinase-1 promoter polymorphism 1G/2G is correlated with colorectal cancer invasiveness.

PURPOSE: Matrix metalloproteinase-1 (MMP-1) is likely to be involved in invasion and metastasis of several tumors by degrading the extracellular matrix. A single guanine insertion polymorphism (2G) in the MMP-1 promoter region creates an Ets binding site causing the elevation of transcriptional level and local expression of MMP-1. The aim of this study was to evaluate the impact of this 2G insertion type polymorphism on invasion and metastasis of colorectal cancer (CRC). EXPERIMENTAL DESIGN: We genotyped for this 1G/2G polymorphism 60 patients, who were operated on for CRC and followed for 6-30 months (median: 21). A control population of 164 age- and sex-matched tumor-free subjects was also genotyped for the same polymorphism. RESULTS: The proportion of 2G homozygotes was higher in the CRC group than in the controls (P = 0.014; odds ratio, 2.21; 95% confidence interval, 1.17-4.16). The CRC group was divided in a group without metastasis (M-) and a group that had developed metastasis (M+). At the time of diagnosis, 2G homozygotes were more represented in the M+ group than in M- (P = 0.0082; odds ratio, 4.73; 95% confidence interval, 1.46-15.26). The difference between M- patients and controls did not achieve statistical significance (P = 0.52). CONCLUSIONS: Our results suggest that the presence of 2G polymorphism at the MMP-1 promoter region may favor the growth and the metastatic process in CRC patients and could be looked at as a risk factor for a worse prognosis.

Homocysteine concentration in primary and systemic sclerosis associated Raynaud's phenomenon.

OBJECTIVE: To investigate whether patients with systemic sclerosis (SSc) have raised homocysteine (Hcy) plasma levels, thought to be an independent risk factor for vascular disease, and to study the relationship between Hcy and endothelial damage, and between Hcy and methylene-tetrahydrofolate reductase (MTHFR) genotypes, and patients' vitamin nutritional status, which are among the more frequent causes of hyperhomocysteinemia. METHODS: We measured Hcy, von Willebrand factor (vWF), folic acid, and vitamin B12 plasma levels and analyzed the frequencies of MTHFR mutations in 30 patients with SSc and 12 patients with primary Raynaud's phenomenon (RP); 29 healthy subjects served as controls. RESULTS: Patients with SSc had higher Hcy and vWF concentrations than those with RP (p < 0.01 and p < 0.02, respectively) or controls (p < 0.02 and p < 0.0001, respectively). Folic acid and vitamin B12 were lower in SSc than in RP (p < 0.01 and p < 0.02, respectively) or controls (p < 0.05). MTHFR genotype did not influence Hcy, folate, or vitamin B12 concentrations, but patients homozygous for the mutant gene had higher vWF levels. CONCLUSION: Patients with SSc, but not those with RP, had significantly higher Hcy and vWF plasma levels. Nutritional rather than inherited factors seem to have a pathogenic role in SSc hyperhomocysteinemia.

Should genetic analysis in newborn screening and a heterozygote test for hyperphenylalaninaemia be recommended? An Italian study.

OBJECTIVE: To determine whether the introduction of genetic analysis for phenylalanine hydroxylase (PAH) deficiency into regional screening programmes can be supported by the benefit-cost ratio. METHOD: Tests for the genetic PAH locus were carried out in 151 patients with hyperphenylalaninaemia originally from all of the Italian regions. PAH mutations were identified by extraction of genomic DNA from leucocytes (whole blood in EDTA), PAH exon amplification was determined by polymerase chain reaction, restriction enzyme analysis was carried out for some recognised mutations, and DNA sequence analysis for the other mutations. RESULTS: It was found that the eight most common mutations in the population accounted for 49% of the mutant alleles, which is well below the required standard for effective population screening (90%). CONCLUSIONS: Genetic screening for PAH deficiency in Italy does not increase the sensitivity of the methodology and the benefit-cost ratio, and thus provides no advantage, particularly as the correlation between genotype and the metabolic phenotype needed to optimise dietary intervention is still being studied.

Exogenous cGMP prevents decrease in diuresis and natriuresis induced by inhibition of NO synthesis.

We previously demonstrated that the intravenous infusion of the specific inhibitor of nitric oxide (NO) synthesis, NG-nitro-L-arginine methyl ester (L-NAME), over a period of 60 min elevates mean arterial pressure (MAP) and reduces renal hemodynamics and excretory function. The objective of the present study was to determine the ability of a guanosine 3',5'-cyclic monophosphate (cGMP) analogue, 8-bromo-cGMP (8-BrcGMP), in preventing the increase in MAP and the reductions in renal plasma flow (RPF), glomerular filtration rate (GFR), urine flow (UV), and sodium excretion rate (UNaV) induced by intravenous infusion of L-NAME in rats. As expected, the infusion of L-NAME (50 micrograms.kg-1.min-1) increased (P < 0.05) MAP and reduced (P < 0.05) RPF, GFR, UV, and UNaV. The administration of 8-BrcGMP (100 micrograms.kg-1.min-1) and L-NAME resulted in no change in MAP, RPF, and GFR. However, decreased (P < 0.05) UV and UNaV were still observed. When 8-BrcGMP (200 micrograms.kg-1.min-1) and L-NAME were infused together, no significant changes in MAP or in renal function were observed. To prove the specificity of the 8-BrcGMP preventive effects, dibutyryl cAMP (200 micrograms.kg-1.min-1) and L-NAME (50 micrograms.kg-1.min-1) were infused together. Under these conditions, MAP, RPF, GFR, UV, and UNaV were modified in a manner similar to that observed during the infusion of L-NAME.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of the endothelium-dependent relaxing factor nitric oxide on renal function.

The role of nitric oxide in renal function has been assessed with pharmacologic and physiologic interventions. Pharmacologically, the renal vasodilation and, to some extent, the natriuresis produced by endothelium-dependent vasodilators such as acetylcholine and bradykinin are mediated by nitric oxide and also by prostaglandins. However, prostaglandins and nitric oxide do not participate in the renal effects produced by endothelium-independent vasodilators such as atrial natriuretic peptide, prostaglandin I2, and nitroprusside. Physiologically, nitric oxide and prostaglandins exert a strong regulation on the effects produced by changes in renal perfusion pressure. Increments in renal perfusion pressure within the range of RBF autoregulation appear to inhibit prostaglandin synthesis while simultaneously enhancing the formation of nitric oxide. Nitric oxide modulates autoregulatory vasoconstriction and at the same time inhibits renin release. Conversely, a decrease of renal perfusion pressure to the limit of or below RBF autoregulation may inhibit the synthesis of nitric oxide but may trigger the release of prostaglandins, whose vasodilator action ameliorates the fall in RBF and stimulates renin release. Nitric oxide and prostaglandins are also largely responsible for mediating pressure-induced natriuresis. However, unlike prostaglandins, mild impairment of the synthesis of nitric oxide in systemic circulation produces a sustained decrease in sodium excretion, which renders blood pressure susceptible to be increased during high-sodium intake. This effect suggests that a deficiency in the synthesis of nitric oxide could constitute the most effective single disturbance to foster the development of a syndrome similar to that seen in salt-sensitive hypertension.

Zonal changes of guanidine 3', 5'-cyclic monophosphate related to endothelium-derived relaxing factor in dog renal medulla.

This study was undertaken to determine the basal and the stimulated profiles of endothelium-derived relaxing factor (EDRF) production along the cortical medullary axis of the dog kidney. To this end, slices (0.5 mm thick) were obtained from six zones equally spaced along the cortical medullary axis. Zone 1 included the medullary crest, while zones 2 and 3 included the inner medulla, zone 4 the outer medulla, zones 5 and 6 the the middle and superficial cortex, respectively. The guanidine 3', 5'-cyclic monophosphate (cGMP) content (an index of EDRF production) was determined by radioimmunoassay under basal conditions and after acetylcholine (10(-5) M), bradykinin (10(-5) M) and SIN-1 (10(-4) M) stimulation. Under basal and stimulated conditions, the cGMP concentrations were highest in the midinner medulla and decreased progressively to lowest concentration in the cortex. These responses were inhibited by NG-monomethyl-L-arginine (LNMMA), a specific antagonist of EDRF synthesis. In contrast, LNMMA did not alter the stimulation of cGMP produced by SIN-1 (10(-4) M) an endothelium-independent vasodilator. This particular localization of EDRF-mediated stimulation on the midinner medulla may have a specific role in the regulation of sodium tubular reabsorption.

The calcium channel antagonist S 11568 causes endothelium-dependent relaxation in canine arteries.

The new dihydropyridine calcium channel antagonist S 11568 and its optical isomers, S 12967 and S 12968 (3 x 10(-6) to 10(-4) M), caused, unlike nifedipine (10(-4) M), equipotent and rapid endothelium-dependent relaxations and increased the content of cyclic GMP in rings of canine femoral arteries. These effects were observed in the presence of indomethacin and were prevented by methylene blue, hemoglobin and NG-monomethyl-L-arginine. Thus these effects must involve endothelium-derived relaxing factor (EDRF) and be distinct from the calcium channel antagonistic effect, which is stereoselective and of slow onset. The compounds did not potentiate relaxations of rings without endothelium to nitric oxide. In bioassay experiments, the compounds produced endothelium-dependent relaxation only when applied to endothelial donors. These results are compatible with an increased release of EDRF induced by the dihydropyridine compounds.

Cold-induced changes in plasma norepinephrine, epinephrine and dopamine concentrations in patients with Raynaud's phenomenon.

To investigate the role of the sympathetic system in Raynaud's phenomenon, which has not yet been elucidated, we measured the levels of norepinephrine, epinephrine and dopamine before, immediately and 30 min after the cold pressor test in plasma from 17 patients with primary, 6 with secondary Raynaud's phenomenon and 19 volunteers, matched for age and sex. Patients had significantly low baseline epinephrine (0.13 +/- 0.02 vs 0.37 +/- 0.04, nmol/l, p less than 0.001, mean +/- S.E.), but normal norepinephrine and dopamine (norepinephrine: 1.77 +/- 0.16 and 2.06 +/- 0.18; dopamine: 0.10 +/- 0.01 and 0.11 +/- 0.02, patients and controls). Immediately after the cold test norepinephrine significantly increased (p less than 0.001) in patients (2.42 +/- 0.22) and controls (3.24 +/- 0.28); epinephrine increased in patients (0.18 +/- 0.02, p less than 0.02); dopamine did not show any significant change (0.13 +/- 0.01 and 0.13 +/- 0.02, patients and controls). In the recovery period, while norepinephrine and epinephrine returned to baseline in both groups, dopamine increased in controls (0.21 +/- 0.04, p less than 0.005) but remained unchanged in patients (0.11 +/- 0.01). We conclude that there is no sympathetic overactivity in Raynaud's phenomenon and propose a role for circulating dopamine in post-ischaemic vasodilatation as an explanation for the particular behaviour of dopamine.

L-arginine therapy in Raynaud's phenomenon?

Since L-Arginine is the substrate for nitric oxide synthesis by vascular endothelial cells the effects of L-arginine treatment on the digital vascular response to local stimuli were investigated in patients with primary or secondary Raynaud's phenomenon. After therapy, patients with Raynaud's phenomenon secondary to systemic sclerosis showed: (1) higher digital vasodilation after local warming, (2) cold-induced digital vasodilation, and (3) increase of plasma levels of tissue-type plasminogen activator.

Evidences for the existence of endothelium-derived relaxing factor in the renal medulla.

The basal levels of cGMP in renal medulla slices were enhanced when the slices were stimulated with both endothelium-dependent (acetylcholine) and endothelium-independent (molsidomine) vasodilators. When preincubated with NG-mono-methyl-L-arginine, a specific inhibitor of endothelium-derived relaxing factor, only the acetylcholine-stimulated increase was completely abolished. Furthermore, a preincubation with L-arginine, a selective precursor of endothelium-derived relaxing factor, enhanced the cGMP levels. The results indicate that the renal medulla, presumably the endothelial cells of the vasa recta, is able to produce endothelium-derived relaxing factor.

Effect of chronic ketanserin treatment on serotonin-induced platelet aggregation in patients with Raynaud's phenomenon.

To investigate whether the variable efficacy in Raynaud's phenomenon of long-term oral ketanserin treatment might be related to variation in the sensitivity of 5-HT2 receptors to ketanserin, serotonin-induced platelet aggregation was measured in ten patients with Raynaud's phenomenon at various times after treatment with ketanserin. Platelet aggregation was completely inhibited 90 min after 40 mg ketanserin, but not 12-14 h after the last dose of 40 mg on the 31st day of continuous twice daily administration. However, 90 min after an additional dose of 40 mg, platelet aggregation was again completely inhibited. The present results indicate that ketanserin 40 mg b.d. does not continuously inhibit platelet 5-HT2 receptors in patients with Raynaud's phenomenon and suggest that more frequent intake might be more effective.

Platelet and plasma serotonin in patients with liver cirrhosis.

To evaluate the role of serotonin in liver cirrhosis, serotonin was determined by high-performance liquid chromatography in plasma, platelets and ascitic fluids from 14 cirrhotic patients. Plasma-free serotonin was within the normal range, but intraplatelet serotonin was significantly low in cirrhosis (p less than 0.001) and this decrease paralleled the severity of the disease. The concentration of serotonin in ascitic fluids was 12% of the corresponding plasma concentrations. Our data indicate that serotonin levels are influenced by hepatic injury, but the reasons for these changes are still unclear.