Early renal post-ischaemic tissue damage and dysfunction with contribution of A1-adenosine receptor activation in rat.

AIM: This study investigated the effect of a selective A(1)-adenosine receptor (A(1)-AR) antagonist, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), on the renal dysfunction and histological damage induced by ischaemia/reperfusion at an early stage. METHODS: Pentobarbital anaesthetised rats were prepared for measuring renal functional variables. Ischaemia was induced by bilateral renal artery clamping for 30 min followed by a 4 h reperfusion period. In DPCPX-treated rats, it was infused (i.v.) at 10 microg/kg per min before and after renal ischaemia. Both kidneys were examined using light and electron microscopy. RESULTS: The renal ischaemic challenge resulted in major histological and ultrastructural damages, which were associated with decreased creatinine clearance, absolute potassium-excretion and effective free-water reabsorption, but increased fractional sodium-excretion and urine flow during reperfusion period. In DPCPX-treated rats, the histological and ultrastructural damage to the kidneys was improved along with the decrease in creatinine clearance and increase in fractional sodium-excretion being smaller, but the increase in urine flow being larger than those of the non-treated rats, while absolute potassium-excretion and effective free-water reabsorption were equal to those of the sham-operated rats. CONCLUSION: These findings suggest that endogenous activation of A(1)-AR contributes to the early development of renal ischaemia/reperfusion injury.

Expression of spinal cord Fos protein in response to intrathecal adrenomedullin and CGRP in conscious rats.

Adrenomedullin (AM) immunoreactivity and mRNA, in addition to a large number of specific AM-binding sites, exist in the rat spinal cord. However, no phenotype has been reported for AM in the spinal cord. Here, expression of c-fos in response to intrathecal (i.t.) administration of AM, proadrenomedullin N-terminal 20 peptide (PAMP) and calcitonin gene-related peptide (CGRP) was examined in the thoracic, lumbar and sacral regions of spinal cord in conscious rats. Two hours after i.t. administration of either CGRP (2.5 and 10 microg) or AM (10 microg), the number of c-Fos immunoreactive nuclei was increased in all the spinal regions examined in this study, with the highest increase observed in the superficial dorsal horn. Few cells with c-fos immunoreactivity were found in the spinal cord of rats 2 h after i.t. injection of either saline or PAMP. Effects of AM (10 microg) and CGRP (2.5 microg) on c-fos expression were blocked when rats were pretreated with 40 microg of intrathecal CGRP8-37 (CGRP1 receptor antagonist). Fos-like immunoreactivity induced by i.t. CGRP and/or AM were also significantly abolished by i.t. administration of the nitric oxide (NO) inhibitor, l-NAME, indicating that endogenous NO is a necessary intermediary in CGRP and AM induced c-fos expression in the rat spinal cord. In conclusion, AM induces c-fos expression in rat spinal cord when administered intrathecally, with the pattern being similar to those produced by i.t. CGRP. Effects of the two peptides are sensitive to CGRP8-37 and l-NAME.

Antihypertriglyceridemic effect of walnut oil.

Hyperlipidemia is an important modifiable risk factor of coronary heart diseases. So far, several studies, have indicated the beneficial effects of nuts on plasma lipid profile. Previously, in a pilot study the authors have shown that administration of 20 g/day of Persian walnut (Juglans regia L.) for 8 weeks could decrease plasma triglyceride (TG) concentration by 17% (p value < 0.05). Walnut also increased the plasma HDL-cholesterol level markedly (p value < 0.05). To make the measurements more reliable and to avoid the unwanted walnut side effects (eg, rash, pruritus), this randomized, double blind case-control study was conducted to evaluate the lipid-lowering effect of Persian walnut oil in the population of southern Iran. Sixty hyperlipidemic subjects were randomized into 2 groups; group A patients (n = 29) received walnut oil encapsulated in 500 mg capsules, 3 g/day, for 45 days. Group B patients (n = 31) received placebo and served as the control group. Lipid profiles of both groups were checked before; on days 15, 30, and 45 after the beginning; and 15 days after termination of the study. Plasma TG concentrations decreased by 19% to 33% of baseline in group A patients (p value < 0.05). No statistically significant change was observed in other measured parameters. It was concluded that walnut oil is a good antihypertriglyceridemic natural remedy and should be further explored in more detail.

Effects of intracerebroventricular injection of glucagon like peptide-1 and its related peptides on serotonin metabolism and on levels of amino acids in the rat hypothalamus.

High concentrations of glucagon-like peptide-1 (7-36) amide (GLP-1) and its specific receptor (GLP-1R) have been found in the rat hypothalamus. In this study the actions of GLP-1 and its related peptides, exendin-4 (GLP-1R agonist), exendin (9-39) (GLP-1R antagonist) and GLP-1 (9-36) amide (the major GLP-1 metabolite) on levels of serotonin (5-HT), 5-hydroxyindolacetic acid (5-HIAA) and amino acids (Glu, Asp, Gln, Gly, Tyr, Trp, GABA) in the hypothalamus were investigated. Intracerebroventricular (ICV) injection of GLP-1 (4 nmol) produced a significant reduction in levels of 5-HT (54%) and all measured amino acids (34 to 56%) compared with saline injected controls, whereas exendin (9-39) (4 nmol) was ineffective. ICV injection of exendin-4 produced a significant reduction in the levels of 5-HT, 5-HIAA, Trp, Glu, and Tyr. ICV injection of GLP-1(9-36) amide showed a statistically significant increase in the level of 5-HT, 5-HIAA and all the amino acids tested in this study. Prior administration of exendin (9-39) or GLP-1 (9-36) amide blocked the effects of GLP-1 on the levels of 5-HT and the amino acids. These data are consistent with exendin-4 being a GLP-1R agonist and exendin (9-39) being a specific GLP-1R antagonist. GLP-1 (9-36) amide, a primary metabolite of GLP-1, appears to act as an endogenous antagonist at the GLP-1R.

Erysipeloid leishmaniasis: an unusual clinical presentation.

Old World cutaneous leishmaniasis has many different clinical presentations. A rare and unusual presentation of cutaneous leishmaniasis is the erysipeloid type. This clinical form is not only unusual in its clinical features but also in the specific category of patients it seems to afflict. In this report 5 Iranian patients, predominantly females, between 50 and 70 years of age, presented with infiltrative erythematous lesions covering the center of the face and resembling erysipelas. Skin smears and/or skin biopsies revealed the diagnosis of cutaneous leishmaniasis. The reason for this type of presentation is unclear, although factors such as the specific species involved, the host's immune response, the hormonal changes encountered with increasing age, and the changes in skin barrier with ageing can be speculated as being important points in causing such an unusual presentation.

A comparison of drug binding sites on mammalian albumins.

The fluorescent probes warfarin and dansylsarcosine are known to selectively interact with binding sites I and II, respectively, on human albumin. This paper investigates whether similar binding sites exist on bovine, dog, horse, sheep and rat albumins. Binding sites on albumins were studied by: (1) displacement of warfarin and dansylsarcosine by site I (phenylbutazone) and site II (diazepam) selective ligands; (2) the effects of non-esterified fatty acids (carbon chain lengths: C5-C20) and changes in pH (6-9) on the fluorescence of warfarin and dansylsarcosine; and (3) the ability of site selective ligands to inhibit hydrolysis of 4-nitrophenyl acetate. For bovine, dog, horse, human and sheep albumins the fluorescence of bound warfarin and dansylsarcosine was selectively decreased by phenylbutazone and diazepam, respectively. For these albumins medium chain fatty acids (C1-C12) reduced the fluorescence of dansylsarcosine (maximum inhibition with C9) whereas long chain acids (C12-C20) enhanced the fluorescence of warfarin (maximum increases with C12). In addition, changes in pH from 6 to 9 increased the fluorescence of warfarin and although site I ligands (warfarin/phenylbutazone) had no pronounced effects on 4-nitrophenyl acetate hydrolysis, site II ligands (dansylsarcosine/diazepam) significantly inhibited this reaction. Rat albumin behaved differently from the other albumins studied in that the C12-C20 fatty acids and changes in pH did not enhance the fluorescence of warfarin. Moreover, the differential effects of site I and site II ligands on the fluorescence of warfarin/dansylsarcosine and hydrolysis of 4-nitrophenyl acetate were less apparent with rat albumin. The results suggest bovine, dog, horse and sheep albumins have binding sites for warfarin and dansylsarcosine with similar properties to sites I and II on human albumin. By contrast, the warfarin binding site and to a lesser degree the dansylsarcosine site, of rat albumin have different characteristics from these sites on the other albumins studied.

Further characterization of the protective effect of 8-cyclopentyl-1,3-dipropylxanthine on glycerol-induced acute renal failure in the rat.

In the rat, treatment with the alkylxanthine 8-cyclopentyl-1,3-dipropylxanthine (CPX) at a dose of 0.1 mg kg-1 antagonizes adenosine-induced falls in renal blood flow and reduces the severity of glycerol-induced acute renal failure. Treatment of glycerol-injected rats with 0.03 mg kg-1 of CPX resulted in no significant improvements in a range of indices of renal function. However, treatment with 0.1 or 0.3 mg kg-1 doses of CPX did significantly ameliorate acute renal failure although there were no significant differences in the degree of protection of renal function afforded by these two doses. In glycerol-injected rats, 0.1 or 0.3 mg kg-1 CPX administered either as a single dose or repeated doses every 12 h for two days did not inhibit renal phosphodiesterase. Thus the beneficial effects of CPX can be produced by doses that have no effect on renal phosphodiesterase activity whereas 0.1 mg kg-1 of CPX has been shown previously to antagonize the actions of adenosine. The findings provide further evidence that the beneficial effect of CPX in glycerol-induced acute renal failure is a consequence of adenosine antagonism and not phosphodiesterase inhibition.

The effect of 8-cyclopentyl-1,3-dipropylxanthine on the development of cyclosporin-induced acute renal failure.

The effect of the selective A1-adenosine receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (CPX, 0.1 mg kg-1 i.v.) administered twice daily to rats has been assessed on the development of renal dysfunction induced by four daily injections of cyclosporin (60 mg kg-1 i.p.). The series of cyclosporin injections resulted in a polyuria accompanied by a 64-70% increase in plasma urea and creatinine concentrations and a 50% reduction in inulin clearance. However, cyclosporin administration resulted in no change in p-aminohippurate clearance nor was there any evidence of tubular necrosis or vascular damage. CPX treatment did not improve any index of renal function perturbed by cyclosporin. The findings provide evidence that adenosine does not play a role in the pathophysiology of cyclosporin nephrotoxicity.

Analysis of the binding of fluorescent ligands to soluble proteins. Use of simultaneous non-linear least squares regression to obtain estimates of binding parameters.

The binding of three fluorescent ligands (warfarin, dansylsarcosine and 1-anilino-8-naphthalene sulphonate) to human albumin was analysed using simultaneous non-linear least squares regression analysis. Both mock and actual fluorescence data were examined and the results indicated that reliable estimates of the binding parameters as well as the molar fluorescence of bound ligand could be obtained. The advantage of this method of analysis is that it makes full use of all the experimental data and it eliminates the need for the graphical procedures usually employed to estimate the molar fluorescence of bound ligand and its binding constants. This type of analysis can be extended to other systems where some physical property of the bound ligand varies with increasing protein concentration.

Effect of valproic acid, its unsaturated metabolites and some structurally related fatty acids on the binding of warfarin and dansylsarcosine to human albumin.

The sites to which valproic acid and its main unsaturated metabolites (2-en-2-propyl pentanoic acid and 4-en-2-propyl pentanoic acid) bind to on human albumin were investigated by (1) measuring their ability to displace the fluorescent probes warfarin and dansylsarcosine and (2) by assessing the extent to which they inhibited the hydrolysis of 4-nitrophenyl acetate. Valproate and its metabolites displaced both warfarin and dansylsarcosine, and they also inhibited the hydrolysis of 4-nitrophenyl acetate. The order of potency for inhibition of both binding and hydrolysis was: 2-en-2-propyl pentanoic acid greater than 4-en-2-propyl pentanoic acid greater than or equal to valproate. It is concluded that valproic acid and its unsaturated metabolites can displace ligands from the warfarin binding site (site I) and the benzodiazepine/indole binding site (site II), but the primary interaction is with site II. Furthermore, the introduction of a double bond into the carbon backbone of valproate increases affinity for albumin at both sites.

A pitfall in the use of double-reciprocal plots to estimate the intrinsic molar fluorescence of ligands bound to albumin.

Double-reciprocal plots of fluorescence intensity versus protein concentration are often used to obtain the intrinsic molar fluorescence (Fb) of ligands bound to acceptor molecules such as albumin. In this paper we show that these plots develop upward concave curvature as the concentration of albumin increases. Thus linear extrapolation of such plots cannot be employed to provide accurate values of Fb. It is suggested that a direct plot of fluorescence intensity versus log protein concentration should be employed to obtain Fb.