Depsides as non-redox inhibitors of leukotriene B(4) biosynthesis and HaCaT cell growth, 2. Novel analogues of obtusatic acid

Aseries of obtusatic acid analogues has been synthesized and evaluated as inhibitors of leukotriene B(4) (LTB(4)) biosynthesis and as antiproliferative agents. The 4-O-benzylated and the 4-O-demethylated congeners were the most potent inhibitors of LTB(4) production of the depside class of compounds, with IC(50) values in the submicromolar range. Furthermore, these compounds do not function as redox-based inhibitors because they were not reactive against a stable free radical, 2,2-diphenyl-1-picrylhydrazyl, and did not produce appreciable amounts of deoxyribose degradation as a measure of their potency to generate hydroxyl radicals. Some obtusatic acid congeners were also potent inhibitors of keratinocyte growth. Growth inhibition was not mediated by damage to the cell membrane, as the activity of lactate dehydrogenase released from the cytoplasm was in the control range.     

The effect of hormone replacement therapy and route of administration on selected cardiovascular risk factors in post-menopausal women

BACKGROUND: There is increasing use of hormone replacement therapy (HRT) by post-menopausal women. Observational epidemiological studies have shown reductions in cardiovascular risk factors in HRT users in the USA, but no randomized controlled trials of HRT have been carried out in the primary practice setting. Previous studies of cardiovascular risk factors have shown a variety of responses according to type of progestagen and oral or topical administration. None has examined the effect of route using an identical progestagen. OBJECTIVES: Our aim was to establish differences, if any, in alteration in cardiovascular risk factors with HRT in post-menopausal women according to route of administration of HRT, oral, transdermal and implant, using first oestrogen alone then oestrogen plus norethisterone, or testosterone for implant. METHODS: Subjects were recruited by letter of invitation to women aged 50-65 years from lists in general practices local to the Charing Cross Hospital Lipid Clinic in West London. Their menopausal status was confirmed and they were randomized to one of three treatment groups or acted as controls. They attended for three visits; at baseline, HRT was initiated as oestrogen alone, oral or transdermal. At the 3-month visit, HRT with the progestagen, norethisterone, was given cyclically, continuously or transdermally until the final visit at 6 months. A separate group of women from the menopause clinic at Chelsea and Westminster Hospital were studied on oestrogen implant then on implanted oestrogen and testosterone. The outcome measures studied were the separate effects of the four regimes as compared with controls on lipoproteins, glucose, insulin, fibrinogen, factor VII and E-selectin, together with weight, waist:hip ratio and blood pressure. RESULTS: The continuous combined oestrogen-progestagen therapy had similar effects on cardiovascular risk factors as oestrogen with cyclical progestagen. All regimes lowered low-density lipoprotein cholesterol, the oral route being more potent than the parenteral; the effect of transdermal HRT was similar to the implant. Lp(a) was reduced only with the oral route. Reductions in factor VII and E-selectin were observed in both the oral and transdermal routes. There was no increase in body mass index, waist:hip ratio, blood pressure or glucose and insulin levels with any of the HRT regimes used. Systolic blood pressure was reduced with the transdermal route. CONCLUSIONS: This study supports the evidence that oestrogen-progestagen HRT, both oral and transdermal, although attenuating some of the benefit of oestrogen alone on fibrinogen and high-density lipoprotein, significantly reduces cardiovascular risk factors, which should diminish post-menopausal risk of coronary disease.     

Down-regulation of urea transporters in the renal inner medulla of lithium-fed rats

BACKGROUND: Lithium is commonly used to treat bipolar psychiatric disorders but can cause reduced urine concentrating ability. METHODS: To test whether lithium alters UT-A1 or UT-B urea transporter protein abundance or UT-A1 phosphorylation, rats were fed a standard diet supplemented with LiCl for 10 or 25 days, and then compared to pair-fed control rats. To investigate another potential mechanism for decreased urea transport, inner medullary collecting duct (IMCD) suspensions from lithium-fed or control rats were incubated with 32P-orthophosphate to measure the phosphorylation of UT-A1. RESULTS: In lithium-fed rats (25 days), UT-A1 abundance was reduced to 50% of control rats in IM tip and to 25% in IM base, and UT-B abundance was reduced to 40% in IM base. Aquaporin-2 (AQP2) protein abundance was reduced in both IM regions. Vasopressin (100 pmol/L) increased UT-A1 phosphorylation in IMCD suspensions from control but not from lithium-fed rats; a higher vasopressin concentration (100 nmol/L) increased UT-A1 phosphorylation in control and lithium-fed rats. CONCLUSIONS: Decreases in UT-A1, UT-B, and AQP2 protein abundance, and/or vasopressin-stimulated phosphorylation of UT-A1, can contribute to the reduced urine concentrating ability that occurs in lithium-treated rats.     

Molecular approaches to urea transporters

Urea plays a critical role in the urine-concentrating mechanism in the inner medulla. Physiologic data provided evidence that urea transport in red blood cells and kidney inner medulla was mediated by specific urea transporter proteins. Molecular approaches during the past decade resulted in the cloning of two gene families for facilitated urea transporters, UT-A and UT-B, encoding several urea transporter cDNA isoforms in humans, rodents, and several nonmammalian species. Polyclonal antibodies have been generated to the cloned urea transporter proteins, and the use of these antibodies in integrative animal studies has resulted in several novel findings, including: (1) the surprising finding that UT-A1 protein abundance and urea transport are increased in the inner medulla during conditions in which urine concentrating ability is reduced; (2) vasopressin increases UT-A1 phosphorylation in rat inner medullary collecting duct; (3) UT-A protein abundance is upregulated in uremia in both liver and heart; and (4) UT-B is expressed in many nonrenal tissues and endothelial cells. This review will summarize the knowledge gained from using molecular approaches to perform integrative studies into urea transporter protein regulation, both in normal animals and in animal models of human diseases, including studies of uremic rats in which urea transporter protein is upregulated in liver and heart.     

Evaluation of PCR-ELISA for determination of telomerase activity in prostate needle biopsy and prostatic fluid specimens

The conventional TRAP assay will determine telomerase activity in tissue or other specimens. However, methodological disadvantages limit its clinical use. We evaluated a modified TRAP assay, the telomerase PCR-ELISA, as a practical clinical system for measuring its activity in conjunction with prostate cancer (PCa). We examined telomerase activity by both TRAP and PCR-ELISA assays in 48 sextant needle biopsy (SNB) specimens from dye-marked areas of the prostate glands of 7 PCa patients. Each specimen was histologically confirmed as cancerous or cancer-free by examining a paired specimen taken from the same marked area. In addition, prostatic fluid (PF) specimens were analyzed from 18 patients, 9 of whom were diagnosed with PCa while 9 were diagnosed as cancer-free but mostly with BPH. The results on individual SNB specimens matched well for the two methods. The sensitivity (91%) and specificity (69%) for the PCR-ELISA measurements were consistent with those for the conventional TRAP assay, 88% and 81%, respectively. Quantitatively, with the PCR-ELISA assay, the mean telomerase activity (24.5+/-28.4 units) per needle core with PCa cells was significantly higher than that in needle cores without PCa cells (7.2+/-2.2 unit), as it was with the conventional TRAP assay, namely 25.6+/-27.8 units and 7.3+/-1.8 units, respectively. In PF specimens from PCa patients, which had a lower mean telomerase than was found in needle cores containing PCa cells (7.1+/-1.5 units in the PCR-ELISA, 7.2+/-1.8 units in the conventional TRAP assay), statistical analysis showed good matching between the results from the two assays, overall. In conclusion, the PCR-ELISA can be considered a reliable method to determine telomerase activity as an adjunct in the diagnosis and treatment of prostate cancer.     

A collaborative study to establish the 2nd International Standard for Fibrinogen, Plasma

An International Collaborative Study involving 12 laboratories in 7 different countries was undertaken in order to replace the 1st International Standard (IS) for Fibrinogen, Plasma (89/644). The candidate replacement standard was the ampouled and freeze-dried residue of solvent/detergent treated plasma and was calibrated as coded duplicates (A and B) versus the 1st IS Fibrinogen, Plasma by automated Clauss assay and by a recommended clot collection (gravimetric) assay. This latter method had been used to calibrate the 1st IS Fibrinogen, Plasma. Comparing the ratios of the potency estimates of sample A to sample B (the coded duplicates), all of the laboratories obtained a ratio within 5% of the expected value of 1.0 by automated Clauss assay, which suggests that the laboratories were able to perform this assay well. Scrutiny of the data obtained from the gravimetric assays revealed that in almost all cases the results were invalid. The results of these assays are included in this report but clearly should be treated with caution and indeed produced significantly lower mean estimates of potency than the other assay methods. The overall geometric mean of all estimates of potency of the proposed 2nd IS Fibrinogen, Plasma (98/612) is 2.19 mg/ampoule by the automated Clauss assay. These data have been presented to the Fibrinogen Sub-Committee of the Standardisation and Scientific Committee (SSC) of the International Society on Thrombosis and Haemostasis (ISTH) (Washington, DC, August 1999), which recommended the establishment of 98/612 as the 2nd IS Fibrinogen, Plasma. This report has been presented to the Expert Committee on Biological Standardisation of the World Health Organisation (ECBS-WHO) at their 1999 session and 98/612 was established as the 2nd IS Fibrinogen, Plasma with a potency of 2.2 mg/ampoule.     

Prescreen evaluation of situs inversus patients

Situs inversus is a congenital visceral malrotation anomaly that occurs in approximately 2 per 10,000 live births, but it may go unrecognized until discovered during emergency surgery. The differential diagnosis in situs inversus patients may not be readily seen in the emergency setting. Historical symptoms include reversed locations for common physical complaints, whereas physical signs can be used to diagnose and treat these patients. Laboratory data may also be used to diagnose and treat patients with this anomaly. This study was prompted by a postmortem investigation of a patient with situs inversus and dextrocardia. Several other cases of congenital malrotation, with attention to anatomical variants coupled with medical data, provide guidelines in prescreen evaluation and medical/surgical treatment of similar patients. Careful attention to laboratory and radiologic findings are paramount to quality patient care and prevention of complications. Educating these patients about their malrotation would also aid in future treatment.