Management of venous thromboembolism.

INTRODUCTION: Venous Thromboembolism is an important healthcare problem the world over, resulting in significant morbidity, mortality and resource expenditure. The rationale for use of thromboprophylaxis is based on solid principles and scientific evidence. Indian perspective on this topic is lacking due to the non-availability of published Indian data. This document reviews the available International and Indian data and discusses the relevance of recommendations for prevention and management of Venous Thromboembolism (VTE) in the Indian context. MATERIALS AND METHODS: Meetings of various specialists from different Indian hospitals in the field of Gastrointestinal Surgery, General and Vascular Surgery, Hematology, Intensive Care, Obstetrics and Gynecology, Oncology and Orthopedics were held in the months of August 2005 to January 2006. The guidelines published by American College of Chest Physicians (ACCP), the International Union of Angiology (IUA), and the Royal College of Obstetricians and Gynecologists (RCOG), were discussed during these meetings. The relevance of these guidelines and the practical implications of following these in a developing country like India were also discussed. Any published data from India was collected from data base searches and the results, along with personal experiences of the participating specialists were discussed. The experiences and impressions of the experts during these meetings have been included in this document. Data from recent sources (International Union of Angiology and the National Comprehensive Cancer Network (NCCN) Practice guidelines in Oncology on Venous thromboembolic disease) was subsequently also included in this document. RESULTS: The suggestions formulated in this document are practical, and would intend to serve as a useful practical reference. CONCLUSIONS: A number of unanswered questions remain in the field of thromboprophylaxis, and carefully designed research protocols may help answer some of these. Implementation of the suggestions outlined in the document remains to be studied in the Indian context.

Inhibition of human alpha-thrombin by a phosphonate tripeptide proceeds via a metastable pentacoordinated phosphorus intermediate.

X-ray crystallographic studies of human alpha-thrombin with a novel synthetic inhibitor, an acyl (alpha-aminoalkyl)phosphonate, reveal the existence of a pentacovalent phosphorus intermediate state. Crystal structures of the complex of alpha-thrombin with the phosphonate compound were determined independently using crystals of different ages. The first structure, solved from a crystal less than seven days old, showed a pentacoordinated phosphorus moiety. The second structure, determined from a crystal that was 12 weeks old, showed a tetracoordinated phosphorus moiety. In the first structure, a water molecule, made nucleophilic by coordination to His57 of alpha-thrombin, is bonded to the pentacoordinated phosphorus atom. Its position is approximately equivalent to that occupied by the water molecule responsible for hydrolytic deacylation during normal hydrolysis. The pentacoordinated phosphorus adduct collapses to give the expected pseudo tetrahedral complex, where the phosphorus atom is covalently bonded to Ser195 O(gamma). The crystallographic data presented here therefore suggest that the covalent bond formed between the inhibitor's phosphorus atom and O(gamma) of Ser195 proceeds via an addition-elimination mechanism, which involves the formation of a pentacoordinate intermediate.

Bemiparin and fluid flow modulate the expression, activity and release of tissue factor pathway inhibitor in human endothelial cells in vitro.

We investigated the localisation, gene expression, and activity of tissue factor pathway inhibitor (TFPI) in endothelial cells (EC) grown in static conditions or under shear stress, in the presence of unfractionated heparin (UFH) and two low-molecular-weight heparins (LMWHs). dalteparin and bemiparin (a second generation of LMWHs). All three preparations induced increased release, cellular redistribution, and enhanced activity of TFPI on the cell surface in static EC. In EC grown under shear stress (0.27, 4.1 and 19 dyne/cm2) and incubated with each heparin for 24 h, the release of TFPI was significantly correlated with the level of flow for bemiparin and dalteparin, but not for UFH. For all three levels of flow tested, bemiparin induced the highest secretion and increase of both cellular TFPI and cell surface activity of the inhibitor. The expression of TFPI mRNA, determined by Northern blotting, was specifically modulated by heparins. All three preparations increased the expression of TFPI by 60 to 120% in EC under minimal flow, but only bemiparin enhanced TFPI mRNA in EC under the arterial flow. Immunogold electron microscopy revealed that EC exhibited strong cellular labelling for TFPI when grown under arterial flow in the presence of bemiparin. We conclude that in EC subjected to shear stress in vitro bemiparin is more efficient than UFH or dalteparin in modulating the expression. release and activity of TFPI. We therefore suggest that bemiparin may be superior over the conventional heparins in maintaining the anticoagulant properties of the endothelium.

Phosphate diabetes in patients with chronic fatigue syndrome.

Phosphate depletion is associated with neuromuscular dysfunction due to changes in mitochondrial respiration that result in a defect of intracellular oxidative metabolism. Phosphate diabetes causes phosphate depletion due to abnormal renal re-absorption of phosphate be the proximal renal tubule. Most of the symptoms presented by patients with phosphate diabetes such as myalgia, fatigue and mild depression, are also common in patients with chronic fatigue syndrome, but this differential diagnosis has not been considered. We investigated the possible association between chronic fatigue syndrome and phosphate diabetes in 87 patients who fulfilled the criteria for chronic fatigue syndrome. Control subjects were 37 volunteers, who explicitly denied fatigue and chronic illness on a screening questionnaire. Re-absorption of phosphate by the proximal renal tubule, phosphate clearance and renal threshold phosphate concentration were the main outcome measures in both groups. Of the 87 patients with chronic fatigue syndrome, nine also fulfilled the diagnostic criteria for phosphate diabetes. In conclusion, we report a previously undefined relationship between chronic fatigue syndrome and phosphate diabetes. Phosphate diabetes should be considered in differential diagnosis with chronic fatigue syndrome; further studies are needed to investigate the incidence of phosphate diabetes in patients with chronic fatigue syndrome and the possible beneficial effect of vitamin D and oral phosphate supplements.

Central cooling effects in patients with hypercholesterolaemia.

1. A prospective study has been carried out, and 68 patients with hypercholesterolaemia have been investigated to study the effects of central cooling on serum lipid levels. 2. Central cooling was obtained by the exposure of the whole body to cold water. All patients were trained to gradually reduce the water temperature from 22 to 14 degrees C and to increase the time of exposure from 5 to 20 min over a period of 90 days. The 33 male and 35 female patients were aged between 40 and 60 years at entry with total cholesterol of 6.0 mmol/l or greater and low-density lipoprotein (LDL)-cholesterol of 4. 0 mmol/l or greater. Thyroid-stimulating hormone, free thyroxine (FT4), total T3, total cholesterol, LDL-cholesterol, high-density lipoprotein (HDL)-cholesterol, triacylglycerols and total fat mass (determined by dual-energy X-ray absorptiometry scan) were obtained at baseline and after 3 months treatment with hydrotherapy. 3. Central cooling obtained by hydrotherapy results in a median fall in tympanic temperature from 0.2 degrees C (P<0.001) to 0.8 degrees C (P<0.001). We have observed in these patients a significant reduction in total cholesterol (-0.2 mmol/l, P=0.006) and LDL-cholesterol (-0.2 mmol/l, P=0.004). Serum FT4 level was higher than baseline results in 30 of these hypercholesterolaemic patients (15.5 pmol/l to 17.3 pmol/l) and there was no significant change in serum thyroid-stimulating hormone and total T3. 4. In conclusion, in our patients with hypercholesterolaemia we have observed a significant reduction of total cholesterol and LDL-cholesterol after body temperature regulation.

Detection and characterization of seven novel protein S (PROS) gene lesions: evaluation of reverse transcript-polymerase chain reaction as a mutation screening strategy.

The molecular genetic analysis of protein S deficiency has been hampered by the complexity of the protein S (PROS) gene and by the existence of a homologous pseudogene. In an attempt to overcome these problems, a reverse transcript-polymerase chain reaction (RT-PCR) mutation screening procedure was developed. However, the application of this mRNA-based strategy to the detection of gene lesions causing heterozygous type I protein S deficiency appears limited owing to the high proportion of patients exhibiting absence of mRNA derived from the mutation-bearing allele ("allelic exclusion"). Nevertheless, this strategy remains extremely effective for rapid mutation detection in type II/III protein S deficiency. Using the RT-PCR technique, a G-to-A transition was detected at position +1 of the exon IV donor splice site, which was associated with type I deficiency and resulted in both exon skipping and cryptic splice site utilization. No abnormal protein S was detected in plasma from this patient. A missense mutation (Asn 217 to Ser), which may interfere with calcium binding, was also detected in exon VIII in a patient with type III protein S deficiency. A further three PROS gene lesions were detected in three patients with type I deficiency by direct sequencing of exon-containing genomic PCR fragments: a single base-pair (bp) deletion in exon XIV, a 2-bp deletion in exon VIII, and a G0to-A transition at position -1 of the exon X donor splice site all resulted in the absence of mRNA expressed from the disease allele. Thus, the RT-PCR methodology proved effective for further analysis of the resulting protein S-deficient phenotypes. A missense mutation (Met570 to Thr) in exon XIV of a further type III-deficient proband was subsequently detected in this patient's cDNA. No PROS gene abnormalities were found in the remaining four subjects, three of whom exhibited allelic exclusion. However, the father of one such patient exhibiting allelic exclusion was subsequently shown to carry a nonsense mutation (Gly448 to Term) within exon XII.

Selenium enhances prostacyclin production by cultured endothelial cells: possible explanation for increased bleeding times in volunteers taking selenium as a dietary supplement.

Selenium added to the culture medium of confluent pig aortic endothelial cells caused a time-related elevation in the activity of the hydroperoxide scavenging enzyme: glutathione peroxidase. This increased activity was associated with an enhanced ability to produce prostacyclin irregardless of whether the agonist was arachidonic acid or thrombin. Since prostacyclin synthetase is believed to be irreversibly inhibited by alkyl hydroperoxides, we feel that the greater production of prostacyclin by selenium-treated cells as compared with control cells may reflect a protective effect of GSH.Px towards the synthetase enzyme. The results from this study may explain the observations made on a group of human volunteers ingesting selenium as a dietary supplement. After six weeks treatment with selenium, bleeding time in this group was prolonged suggesting an improved ability to synthesize prostacyclin as a result of selenium-dependent glutathione peroxidase activation in the vessel wall.

Prostaglandins and acute pancreatitis--experimental and clinical studies.

Acute pancreatitis has a mortality of 10-20 per cent, and in cases of acute haemorrhagic pancreatitis this rises to 80-90 per cent. At present there is no reliable treatment for this condition. Based on the hypothesis that the release locally and systemically of the intracellularly activated pancreatic digestive enzymes is due to cell membrane instability, we have studied the cytoprotective (cell membrane stabilizing) role of prostaglandins in this condition. In the first part of this study, an animal model of acute pancreatitis with a mortality of 100 per cent by 14 days has been established by feeding mice a choline-deficient ethionine supplemented diet. Using this model we have demonstrated improved survival (16 out of 50 survived 14 days or longer) by administering prostaglandin E2 subcutaneously (P less than 0 X 02). We have demonstrated that the improved survival is dose-dependent, in the range 2 X 5-5 X 0 mg/kg body weight 8 hourly (P less than 0 X 02) and time-dependent, still being effective if treatment is delayed for 24 h (P less than 0 X 02). Great emphasis has been placed on not commencing treatment until after the induction of acute pancreatitis. In the second part we have used cell membrane marker enzymes to study the cell membrane stabilizing effect of prostaglandin E2 in the human. It has been demonstrated that cell membrane instability occurs. In 12 of 50 episodes treated by prostaglandin E2 infusion, the lysosomal, mitochondrial and cell membranes are stabilized, thus decreasing the release of intracellular enzymes.