Do the COL1A1 and Taq 1 vitamin D receptor polymorphisms have a role in identifying individuals at risk of developing osteoporosis?

The distribution of the Taq 1 polymorphism in the vitamin D receptor (VDR) gene and the MSc 1 polymorphism in the collagen 1 alpha 1 (COL1A1) gene were studied in 266 female and 55 male patients attending an osteoporosis clinic. Allele frequency in control (T- or Z-score >-1.0) and osteoporotic (T- or Z-scores <-2.5) groups were compared using Chi squared tests. No differences were found between the 2 groups with either of the polymorphisms. When allele frequency was compared in patients with and without history of fracture, no differences were found in the frequency of the COL1A1 alleles. However there were significantly more fracture patients, who had been previously treated with corticosteroids for other conditions, carrying the T allele of the VDR polymorphism (X2 = 5.65, p>0.01<0.02). In conclusion, neither of these polymorphisms aid in the prediction of osteoporosis but the VDRT allele may carry an increased fracture risk in patients who require corticosteroid treatment.

Plasma and urinary cytokine homeostasis and renal function during cardiac surgery without cardiopulmonary bypass.

Cardiopulmonary bypass (CPB) significantly contributes to the plasma pro-inflammatory cytokine response at cardiac surgery. Complementary plasma and urinary anti-inflammatory cytokine responses have been described. The pro-inflammatory cytokines interleukin 8 (IL-8), tumour necrosis factor alpha (TNF-alpha) and interleukin 1beta (IL-1beta) have lower molecular weights than the anti-inflammatory cytokines interleukin 10 (IL-10), interleukin 1 receptor antagonist (IL-1ra) and TNF soluble receptor 2 (TNFsr2) and thus undergo glomerular filtration more readily. In vitro work suggests that proximal tubular cells are vulnerable to pro-inflammatory cytokine mediated injury. Accordingly, this study investigated the hypothesis that cardiac surgery without CPB would not have significant changes in plasma and urinary cytokines and proximal renal dysfunction. Eight patients undergoing coronary artery bypass grafting (CABG) without CPB were studied. Blood and urine samples were analysed for pro- and anti-inflammatory cytokines. Proximal tubular dysfunction was measured using urinary Nu-acetyl-beta-D-glucosaminidase (NAG)/creatinine and alpha(1)-microglobulin/creatinine ratios. Plasma IL-8, IL-10, IL-1ra and TNFsr2 were significantly elevated compared with baseline. Urinary IL-1ra and TNFsr2 were significantly elevated, as were urinary NAG/creatinine and alpha(1)-microglobulin/creatinine ratios. Two hours following revascularization, urinary IL-1ra correlated with urinary alpha(1)-microglobulin/creatinine ratios (P<0.05). As previously reported in CABG surgery with CPB, we now report that non-CPB cardiac surgery also has significant changes in plasma and urinary cytokine homeostasis and early proximal tubular injury. The correlation between urinary IL-1ra and alpha(1)-microglobulin/creatinine ratios is consistent with earlier suggestions of a mechanistic link between cytokine changes and proximal tubular dysfunction. The relative roles of CPB and non-CPB processes in producing inflammation still require definition.

Plasma and urinary cytokine homeostasis and renal dysfunction during cardiac surgery.

BACKGROUND: Cardiac surgery induces changes in plasma cytokines. Proinflammatory cytokines have been associated with a number of renal diseases. The proinflammatory cytokines interleukin 8 (IL-8), tumor necrosis factor alpha (TNFalpha), and interleukin 1beta (IL-1beta) are smaller than the antiinflammatory cytokines interleukin 10 (IL-10), interleukin 1 receptor antagonist (IL-1ra), and TNF soluble receptor 2 (TNFsr2), and thus undergo glomerular filtration more readily. Accordingly, this study investigated the relation between plasma and urinary cytokines and proximal renal dysfunction during cardiac surgery. METHODS: Twenty patients undergoing coronary artery bypass grafting with cardiopulmonary bypass (CPB) were studied. Blood and urine samples were analyzed for proinflammatory and antiinflammatory cytokines. Proximal tubular dysfunction was measured using urinary N-acetyl-beta-d-glucosaminidase (NAG)/creatinine and alpha1-microglobulin/creatinine ratios. RESULTS: Plasma IL-8, IL-10, IL-1ra, and TNFsr2 values were significantly elevated compared with baseline. Urinary IL-1ra and TNFsr2 were significantly elevated. Urinary NAG/creatinine and alpha1-microglobulin/creatinine ratios were also elevated. Plasma TNFalpha at 2 h correlated with urinary NAG/creatinine ratio at 2 and 6 h (P < 0.05) and with urinary IL-1ra at 2 h (P < 0.05). Plasma IL-8 at 2 h correlated with NAG/creatinine at 6 h (P < 0.05). Urinary IL-1ra correlated with urinary NAG/creatinine ratio after cross-clamp release and 2 and 6 h after CPB (P < 0.05). CONCLUSIONS: Cardiac surgery using CPB leads to changes in plasma and urinary cytokine homeostasis that correlate with renal proximal tubular dysfunction. This dysfunction may be related to the renal filtration of proinflammatory mediators. Renal autoprotective mechanisms may involve the intrarenal generation of antiinflammatory cytokines.

Mutation screening and genotype:phenotype correlation in familial hypercholesterolaemia.

The aim of this study was to develop a mutation screening protocol for familial hypercholesterolaemia (FH) patients and to assess genotype/phenotype effects in terms of pre-treatment lipid profiles and presentation of tendon xanthomata (TX). A total of 158 families with clinical definitions of possible (120) or definite (38) FH were studied using a tiered screening protocol. Mutations were identified in 52 families, 44 families showing 23 different LDLR gene defects and eight families showing the common Apo B100 gene defect R3500Q. LDLR defects were detected in various regions of the gene with 56% in the LDL binding domain (exons 2-6) and 37% in the EGF precursor homology domain (exons 7-14). The most common mutations were D461N(7), C210X(5), 932delA(5), and C163Y(4). Frameshift mutations accounted for 20% with nonsense 13%, mis-sense 35%, splice 3%, Apo B 13% and 2% large deletion, 13% of clinically definite FH remained undefined. In conclusion, DNA based diagnosis is possible in 79% (30/38) of clinically definite FH families and of the 120 possible FH families at the start of the screening program, 18% (22/120) now have defined mutations. Overall 60 families from the original 158 meet the clinical and/or genetic criteria for definite FH. Tendon xanthomata were present in only 58% (30/52) of genetically defined FH families, thus limiting its use as a strict diagnostic criteria. Families with low density lipoprotein receptor (LDLR) defects present with higher total and LDL cholesterol levels and a higher incidence of TX than do those with the common Apo B variant, and frameshift mutations appear to have the most severe presentation.

Oxidative stress during acute respiratory exacerbations in cystic fibrosis.

BACKGROUND: Patients with cystic fibrosis experience chronic systemic oxidative stress. This is coupled with chronic inflammation of the lung involving bronchial polymorphonuclear neutrophil accumulation and activation. We hypothesised that, during periods of acute respiratory exacerbation, free radical activity and consequent damage would be most marked and that intensive treatment of the infection would result in improvement towards values found during stable periods. METHODS: Plasma and red blood cells were collected from 12 healthy normal volunteers and from 12 patients with cystic fibrosis with an acute respiratory exacerbation (increased respiratory symptoms, reduction in forced expiratory volume in one second (FEV1) of more than 10%, and a decision to treat with intravenous antibiotics). Further samples were collected from patients following two weeks of treatment. Samples were analysed for inflammatory markers, markers of free radical damage, and aqueous and lipid phase scavengers. RESULTS: During respiratory exacerbations FEV1 and forced vital capacity (FVC) were lower than in controls (mean differences -2.82 (95% CI -2.12 to -3.52) and -3. 79 (-3.03 to -4.55) l, respectively) but improved following treatment (mean change 0.29 (95% CI 0.18 to 0.40) and 0.33 (0.23 to 0.43) l, respectively). Inflammatory markers during exacerbations were significantly higher in patients than in controls with the following mean (95% CI) differences: C reactive protein (CRP), 46 (17 to 75) g/l; neutrophil elastase alpha1-antiprotease complexes (NEAPC), 4.4 (1.77 to 7.07) mg/l; white cell count (WCC), 5.3 (4.7 to 5.9) x 10(9)/l. These markers decreased significantly following treatment with the following mean (95% CI) changes: CRP -26 (-10 to -42) g/l; NEAPC -3.1 (-1.3 to -4.9) mg/l; WCC -1.5 (-1.3 to -1.7) x 10(9)/l. Malondialdehyde (MDA) as a marker of free radical activity was significantly higher in patients during exacerbations than in controls with a mean (95% CI) difference of 193 (107 to 279) which improved with treatment (mean change -56 (95% CI -28 to -84) nmol/mmol cholesterol). Red blood cell polyunsaturated fatty acids were significantly lower in patients than in controls with a mean difference of -4.4(95% CI -2.6 to -6.2) moles percent, but did not improve significantly after treatment. Protein carbonyls during exacerbations were not different from controls but did increase with treatment compared with levels during the exacerbation (mean change 0.39 (95% CI 0.11 to 0.67) micromol/g protein). Aqueous and lipid phase scavengers in patients during exacerbations were significantly lower than in controls with the following mean (95% CI) differences: ascorbate, -19.0 (-2.7 to -35.3) micromol/l; sulphydryls, -122 (-77 to -167) micromol/l; retinol, -237 (-47 to -427) nmol/mmol cholesterol; beta-carotene, -52.8 (-11.8 to -93.8) nmol/mmol cholesterol; luteine, -50.4 (-10.4 to -90.4) nmol/mmol cholesterol; lycopene, -90.1 (-30.1 to -150.1) nmol/mmol cholesterol. Treatment resulted in improvement with the following mean (95% CI) changes: sulphydryls, 50 (32 to 68) micromol/l; retinol, 152 (47 to 257) nmol/mmol cholesterol; alpha- and beta-carotene, 0.6 (0.0 to 1.2) and 7.6 (0.0 to 15.2) nmol/mmol cholesterol, respectively; alpha-tocopherol, 839 (283 to 1405) nmol/mmol cholesterol; and lycopene, 8.2 (0.0 to 16.2) nmol/mmol cholesterol. CONCLUSIONS: Abnormalities of markers of inflammation, free radical activity, and radical scavengers were significantly more extreme during acute respiratory exacerbations and showed improvement with treatment. The need to provide protection from inflammation and free radical damage should therefore be dynamic and related to the inflammatory and oxidative processes.

Familial defective apolipoprotein B-100 (R3500Q) in Northern Ireland.

Familial defective apolipoprotein B-100 (FDB) R3500Q is an autosomal co-dominant disorder caused by the substitution of glutamine for arginine at amino acid residue 3500 of the apolipoprotein B-100 gene. It is associated with hypercholesterolaemia of varying severity, and with coronary artery disease. Hypercholesterolaemic patients (n = 158) from Northern Ireland were screened for the defect by polymerase chain reaction-mediated, site-directed mutagenesis. Clinical presentation ranged from moderate hypercholesterolaemia with a family history of hypercholesterolaemia or heart disease (n = 104) to those classified as definitely having familial hypercholesterolaemia (FH) (n = 54). Eight (5.1%) unrelated individuals were found to be heterozygous for the FDB R3500Q mutation, including two (3.7%) of those 54 classified clinically as having FH. Treatment with HMG-CoA-reductase-inhibiting drugs (statins) decreased total cholesterol by 22-44% and low-density lipoprotein cholesterol by 34-46% in all but one FDB heterozygote.

Oxidative stress in cyclosporin and azathioprine treated renal transplant patients.

The major cause of death following transplantation is cardiovascular disease. Among the many processes involved in atherogenesis, oxidative stress and modification of low density lipoprotein has been assigned a major role. This in turn may be affected by the immunosuppressive regime used. We studied oxidative stress in 40 renal transplant patients receiving two different immunosuppressive regimens (20 on cyclosporin, 20 on azathioprine/prednisolone), and 19 normal controls. Changes in lipid peroxidation (assessed as thiobarbituric acid reacting substances, TBARS), antioxidant enzyme activities (glutathione reductase GSHPx, glutathione peroxidase GSHPx and superoxide dismutase SOD) vitamin E and antioxidant associated trace metals (selenium, copper, zinc) were studied. Alteration of erythrocyte membrane fluidity was examined using the fluorescent probe 1,6 diphenyl-1,3,5-hexatriene (DPH). Both transplant groups showed no difference in TBARS, lipid standardised vitamin E, copper or selenium compared to controls. Zinc was significantly increased in both the cyclosporin and azathioprine groups compared to controls (P < 0.05). SOD was reduced in both transplant groups compared to controls (P < 0.001). GSHPx was elevated in both groups compared to controls but only reached significance in the azathioprine treated group (P < 0.005). GSHRx was slightly elevated in both transplant groups but did not reach significance. Erythrocyte membrane anisotropy was decreased in the cyclosporin treated group (P < 0.05). There was no difference in the azathioprine group compared to controls. The present results suggest an adaptive response to increased oxidative stress in both transplant groups sufficient to minimise markers of oxidative stress (TBARS and anisotropy). The results also suggest no significant difference between the two immunosuppressive regimes with regard to oxidative stress.

The peri-operative use of the oesophageal Doppler monitor in patients undergoing coronary artery revascularisation. A comparison with the continuous cardiac output monitor.

This study was carried out to evaluate the ease of use and reliability of cardiac output estimations performed by an oesophageal Doppler monitor and to compare its use with that of a continuous cardiac output pulmonary flotation catheter. Measurements were made during and after surgery in 16 patients scheduled to undergo coronary revascularisation. Both devices suffered significant intra-operative problems which led us to question their suitability as operating theatre monitors. After surgery the continuous cardiac output monitor provided stable results while the oesophageal Doppler monitor required the continuous presence of an experienced anaesthetist to ensure comparable cardiac output estimations.

Comparison of two fluorescent probes for the measurement of erythrocyte membrane fluidity in renal dialysis patients.

Two fluorescent probes were used for the measurement of membrane fluidity in patients on haemodialysis and continuous ambulatory peritoneal dialysis. 1,6-Diphenyl-1,3,5-hexatriene (DPH) anisotropy gives an indication of lipid order and pyrene measures lateral diffusion through the membrane. Pyrene dimer/monomer ratio was significantly lower than controls in both pre-dialysis and post-dialysis samples but DPH anisotropy was unchanged. Both methods showed an increase in membrane fluidity across a 4 hour haemodialysis session. There was an increase in membrane fluidity in CAPD patient samples which was more marked using DPH than pyrene. These results suggest that the two probes give different but complementary information about changes in membrane fluidity and may be more informative when used together rather than singly.

Organophosphates and monocyte esterase deficiency.

AIMS: To examine the possibility that monocyte esterase deficiency (MED) could be caused by exposure to organophosphates. METHODS: Pseudocholinesterase, paraoxonase and arylesterase activities were measured in the serum and acetylcholinesterase activity was measured in the red cells of a group of monocyte esterase deficient subjects and compared with the enzyme activities of a control group of monocyte esterase positive subjects. RESULTS: No significant difference was found between the enzyme activities of the monocyte esterase deficient group and the control group for any of the esterases investigated. CONCLUSION: Current or recent exposure to organophosphorus is not the cause of MED.

Oxidative stress and erythrocyte membrane fluidity in patients undergoing regular dialysis.

Oxidative damage due to free radical production is increased in uraemic patients and has been suggested as a possible factor contributing to the anaemia of chronic renal failure (CRF) and the pathogenesis of atherosclerosis. Oxidative stress was assessed in 40 patients with CRF maintained by either haemodialysis (HD) or continuous ambulatory peritoneal dialysis (CAPD) and in 18 healthy controls. Lipid peroxidation (assessed as malondialdehyde, MDA), total glutathione (TG), antioxidant enzyme (glutathione reductase (GSHRx), glutathione peroxidase (GSHPx) and superoxide dismutase (SOD)) activity and antioxidant associated trace metal (selenium, copper, zinc) levels were studied. Erythrocyte membrane fluidity was examined using the fluorescent probe 1,6 diphenyl-1,3,5-hexatriene (DPH). The results indicate increased levels of oxidative stress and altered erythrocyte membrane fluidity in patients treated with CAPD compared with controls and patients treated with HD. Only minor changes were observed in patients treated with HD. Altered free radical activity, oxidative stress and altered erythrocyte membrane fluidity observed in patients with CRF may contribute to the increase in vascular disease in such patients and to the anaemia of CRF.

Effect of red wine consumption on lipoprotein (a) and other risk factors for atherosclerosis.

Epidemiological studies have pointed to the role of alcohol, and red wine in particular, in reducing the incidence of coronary heart disease. This study attempted to distinguish, in vivo, the effects of components specific to red wine and those of alcohol on lipoproteins, antioxidant status and membrane fluidity. Volunteers (n = 20) were given 200 ml of red wine per day for 10 days. Following a 6-week washout, this was repeated with white wine. Changes within treatment groups were analysed by paired t tests and repeated measures analysis of variance was used to distinguish effects of red wine components and alcohol. LDL was prepared by ultracentrifugation and all other assays were by conventional laboratory techniques. No effect with either treatment was detected on total cholesterol, triglycerides, HDL or measures of antioxidant status, including the susceptibility of LDL to oxidation. Red wine reduced LDL cholesterol (p < 0.01), and both treatments reduced LDL apo B (p < 0.01) and increased LDL chol:apo B ratio (p < 0.01), implying an increase in LDL size. Potential anti-atherogenic changes specific to red wine were reduction in lipoprotein (a) (p < 0.001) and increased membrane fluidity (p < 0.01). These results are not in keeping with the proposed role of red wine components in free-radical protection, but the reduction in lipoprotein (a) merits further investigation.

Does pretreatment with cimetidine and ranitidine affect the disposition of bupivacaine?

Thirty patients who chose extradural analgesia for elective Caesarean section were pretreated, by random selection, with cimetidine 400 mg, ranitidine 150 mg or no H2-blocker. Following the administration of 0.5% bupivacaine, no significant difference was found between peak plasma bupivacaine concentrations or area under the plasma bupivacaine concentration-time curve (AUC) in these three groups. This study shows that a single dose of cimetidine or ranitidine does not affect significantly the disposition of bupivacaine in the obstetric patient.

Postoperative gastric emptying in outpatient anesthesia: the effect of opioid supplementation.

Acetaminophen (paracetamol) 20 mg/kg was administered orally to 45 gynecological outpatients who had received either alfentanil 5 micrograms/kg, fentanyl 1 microgram/kg, or no analgesic supplement immediately prior to the induction of general anesthesia. Postoperative gastric emptying, assessed by acetaminophen absorption, was significantly inhibited in those given alfentanil. This inhibition is unlikely to be of great clinical importance and was much less than that found in previous studies using longer-acting opioids such as morphine.

Plasma concentrations of bupivacaine during extradural anaesthesia for caesarean section. The effect of adrenaline.

The clinical effects and plasma levels associated with the use of 0.5% bupivacaine with and without the addition of 1:200,000 adrenaline (5 micrograms/ml) were studied in 30 patients who underwent extradural anaesthesia for elective Caesarean section. The addition of adrenaline to bupivacaine prolongs analgesia, reduces the degree of hypotension and delays its onset. Plasma bupivacaine levels were consistently lower when adrenaline was added, but this difference was significant only at 10 minutes after the initial dose. Prolonging the interval between increments seems to be a more reliable way to reduce plasma concentration than the addition of the catecholamine.

I.v. temazepam: theoretical and clinical considerations.

Two injectable forms of temazepam, in 90% propylene glycol or 40% salicylic acid, were studied in volunteers, and before surgery in healthy patients. The volunteers also received two forms (capsule and elixir) by mouth. The salicylate preparation was painful on injection and both i.v. formulations caused an unacceptably high incidence of venous thrombosis. Temazepam was detected in plasma earlier following the elixir preparation than the capsule. Plasma concentrations were similar following both injectable preparations. The potency of i.v. temazepam in inducing drowsiness in patients was much less than expected and doses greater than 0.6 mg kg-1 were required to produce adequate sedation. There was a significant reduction in thiopentone induction dose in patients receiving temazepam i.v.

A double-blind comparison of intramuscular pethidine and nalbuphine in labour.

A double-blind, between-patient comparison of intramuscular pethidine 100 mg and nalbuphine 20 mg for the relief of pain during labour in 80 patients is described. Severity of pain was assessed before and after treatment by subjective pain scores and visual analogue scales. Neither of these methods showed a significant difference between the treatments. Nalbuphine was associated with less maternal nausea and vomiting than pethidine, but this possible advantage was somewhat offset by a tendency of the drug to produce more maternal sedation and dizziness. The mean umbilical vein/maternal vein ratio was significantly higher for nalbuphine (0.78, SEM 0.03) than for pethidine (0.61, SEM 0.02), which suggests easier placental transfer of the former. This finding was reflected in significantly lower 2-4 hour neurobehavioural scores for the infants of mothers given nalbuphine, but there was no significant difference between these scores at 24 hours. On the basis of this study, nalbuphine does not offer a substantial improvement over pethidine for pain relief in labour.

A method for producing constant plasma concentrations of drugs. Application to methohexitone.

A delivery system in which a dilute infusion of methohexitone is continuously added to a smaller volume of more concentrated solution has been used to infuse exponentially decreasing drug concentrations at a constant rate of infusion. The constants for calculation of concentrations and infusion rate are described. It was possible to achieve and maintain the target plasma concentration required to produce anaesthesia in conjunction with nitrous oxide in oxygen. Methohexitone is the only available drug suitable for this technique, but it requires frequent opioid supplementation and is not the ideal drug for such a technique.