We investigated whether the glomerular synthesis of prostacyclin modulates the renal blood flow and glomerular filtration rate in chronic glomerular disease. The urinary excretion of 6-keto-prostaglandin F1 alpha, a stable breakdown product of prostacyclin, was significantly (P less than 0.01) reduced in 20 women with chronic glomerular disease, as compared with 19 controls, whereas excretion of urinary prostaglandin E2 was unchanged. In 10 patients randomly assigned to one week of treatment with ibuprofen, excretion of urinary 6-keto-prostaglandin F1 alpha and prostaglandin E2 was reduced by 80 per cent, the level of serum creatinine was increased by 40 per cent, and creatinine and para-aminohippurate clearances were reduced by 28 and 35 per cent, respectively. The reduction of both clearances was inversely related (P less than 0.01) to the basal urinary excretion of 6-keto-prostaglandin F1 alpha but not of prostaglandin E2. No functional changes were detected in five healthy women, despite a similar suppression of renal prostacyclin synthesis by ibuprofen. In contrast, one week of treatment with sulindac did not affect renal prostacyclin synthesis or renal function in the other 10 patients, despite a marked inhibition of extrarenal cyclooxygenase activity. We conclude that in patients with mild impairment of renal function, the renal blood flow and glomerular filtration rate are critically dependent on prostacyclin production. In such patients sulindac may be a safe substitute for other nonsteroidal antiinflammatory drugs.

We have examined the urinary excretion of stable immunoreactive eicosanoids in 23 female patients with systemic lupus erythematosus (SLE), 16 patients with chronic glomerular disease (CGD), and 20 healthy women. SLE patients had significantly higher urinary thromboxane B2 (TXB2) and prostaglandin (PG) E2 excretion and significantly lower 6-keto-PGFI. than did healthy women. In contrast, CGD patients only differed from controls for having reduced 6-keto-PGF1I excretion. The group of SLE patients with active renal lesions differed significantly from the group with inactive lesions for having a lower creatinine clearance and urinary 6-keto-PGFI. and higher urinary TXB2.Higher urinary TXB2 excretion was associated with comparable platelet TXB2 production in whole blood, undetectable TXB2 in peripheral venous blood, and unchanged urinary excretion of 2,3-dinor-TXB2. A significant inverse correlation was found between urinary TXB2 and creatinine clearance rate (Cc,). In contrast, the urinary excretion of 6-keto-PGFI. showed a significant linear correlation with both Cc, and para-aminohippurate clearance rate (CPAH). In four SLE and seven CGD patients, inhibition of renal cyclooxygenase activity by ibuprofen was associated with a significant reduction in urinary 6-keto-PGFIe, and TXB2 and in both Ccr and CPAH. However, the average decrease in both clearances was 50% lower in SLE patients than in CGD patients, when fractionated by the reduction in urinary 6-keto-PGFi, or PGE2 excretion.We conclude that the intrarenal synthesis of PGI2 and TXA2 is specifically altered in SLE. Such biochemical alterations are

A B ST R A C T The objectives of this investigation were: (a) to characterize the time and dose dependence of the effects of prostacyclin (PGI2) on renin release in healthy men; (b) to define whether PGI2-induced renin release is secondary to hemodynamic changes; (c) to determine the plasma and urine concentrations of 6-keto-PGFIa (the stable breakdown product of PGI2) associated with renin release induced by exogenous or pharmacologically enhanced endogenous PGI2. Intravenous PGI2 or 6-keto-PGFIa infusions at nominal rates of 2.5, 5.0, 10.0, and 20.0 ng/kg per min were performed in each of six normal human subjects; in three of them, PGI2 infusion was repeated after ,B-adrenergic blockade and cyclooxygenase inhibition.PGI2, but not 6-keto-PGFIa, caused a time-and dosedependent increase of plasma renin activity, which reached statistical significance at 5.0 ng/kg per min and was still significantly elevated 30 min after discontinuing the infusion. Although combined propranolol and indomethacin treatment significantly enhanced the hypotensive effects of infused PGI2, it did not modify the dose-related pattern of PGI2-induced renin release.Plasma 6-keto-PGFIa levels rose from undetectable levels (<7.5 pg/ml) in a stepwise fashion during increasingly higher infusion rates of PGI2 or 6-ketoPGFia. The threshold concentration of plasma 6-ketoReceived for publication 27 PGFIa associated with a statistically significant stimulation of renin release was -200 pg/ml. Upon discontinuing PGI2 or 6-keto-PGFIar infusion, the disappearance of 6-keto-PGFIa from blood showed an identical biphasic behavior, the initial phase having an apparent t1/2 of 3.2 min. The intravenous infusion of furosemide, which is known to stimulate renin release via a cyclooxygenase-dependent mechanism, caused a three-to fourfold increase of urinary 6-keto-PGF1, excretion rate, concomitant with the elevation of plasma renin activity levels, in six healthy women. 6-Keto-PGFIa remained undetectable in peripheral venous plasma throughout the study. We conclude that in human subjects: (a) PGI2-induced renin release occurs with a dose and time dependence similar to its reported platelet effects; (b) PGI2-induced renin release is not mediated by adrenergic stimuli or cyclooxygenase-dependent mechanisms secondary to hemodynamic changes; (c) furosemide-induced renin release is associated with increased renal PGI2 formation; and (d) PGI2 appears to act as a local modulator rather than a circulating hormone in controlling juxtaglomerular function.

Glomerular vasodilatation in the early stages of type I diabetes mellitus apparently results from arteriolar insensitivity to vasoconstrictors. Since cytosolic free calcium ([Ca2+]i) is a major signaling mechanism for smooth muscle contraction, we studied whether growth of smooth muscle-like rat glomerular mesangial cells in media with high glucose concentration affects [Ca2+]i responses to vasoconstrictors. In cells grown for five days in 22 mM glucose, we observed blunted responsiveness to three structurally unrelated vasoconstrictors that elevate [Ca2+]i via a phospholipase C-dependent mechanism, angiotensin II, prostaglandin F2 alpha, and arginine vasopressin. Inhibition of [Ca2+]i responses was not due to an osmotic effect of high glucose, since it was not mimicked by hypertonic mannitol. While the size of intracellular Ca2+ pools was unaffected by elevated glucose, Na+/Ca2+ exchange was markedly inhibited, thus ruling out both impaired filling of Ca2+ stores and enhanced counter-regulatory mechanisms. Impaired myoinositol transport or intracellular sorbitol accumulation were not responsible for the effects of high glucose, since supplementation of media with myo-inositol or with the aldose reductase inhibitor. Alcon 1576, failed to reverse insensitivity to vasoconstrictors. On the other hand, down-regulation or pharmacological inhibition of protein kinase C completely reversed the effects of high glucose, thus indicating involvement of this signal transduction pathway. These data suggest a possible intracellular mechanism for the impaired vascular sensitivity underlying early renal hemodynamic changes in diabetes mellitus.

We have studied, prospectively, 116 patients with motor deficits associated with herniation of a lumbar disc who underwent microdiscectomy. They were studied during the first six months and at a mean of 6.4 years after surgery. Before operation, muscle weakness was mild (grade 4) in 67% of patients, severe (grade 3) in 21% and very severe (grade 2 or 1) in 12%. The muscle which most frequently had severe or very severe weakness was extensor hallucis longus, followed in order by triceps surae, extensor digitorum communis, tibialis anterior, and others. At the latest follow-up examination, 76% of patients had complete recovery of strength. Persistent weakness was found in 16% of patients who had had a mild preoperative deficit and in 39% of those with severe or very severe weakness. Muscle strength was graded 4 in all patients with persistent weakness, except for four with a very severe preoperative deficit affecting the L5 or S1 nerve root. They showed no significant recovery. Excluding this last group, the degree of recovery of motor function was inversely related to the preoperative severity and duration of muscle weakness. The patients’ subjective functional capacity was not directly related to the degree of recovery except in those with persistent severe or very severe deficit.

Techniques are described in detail for the radioimmunological measurement of prostaglandin (PG) E2 and F2alpha in human urine. 50 ml urine samples are extracted with an organic solvent system and then purified by silicic acid column chromatography. The overall recovery after extraction and purification, calculated with labeled as well as unlabeled compounds, is in the order of 70%. The column eluates are assayed at 1:12-1:60 dilution in the "standard diluent" of the assay: 8 pg PGE2 or PGF2 alpha/ml of whole urine represents the lowest measurable concentration. A urine blank and a solvent blank were evaluated separately, subjecting 50 ml of urine obtained from an indomethacin treated subject ("PG-free" urine) or 50 ml of distilled water, respectively, to the extraction-purification procedures. Both were found not to interfere with the antigen-antibody reaction. Urinary PG-like immunoreactivity (LI) was characterized in terms of immunochemical and thin-layer chromatographic (TLC) behavior. Both urinary PGE2-LI and PGF2 alpha-LI behaved as authentic PGE2 and PGF2 alpha, upon dilution and on TLC. In 33 healthy female subjects (aged 19-58 yr), urinary excretion rates averaged 178 +/- 80 (mean +/- SD) ng/day for PGE2 and 498 +/- 181 ng/day for PGF2 alpha. In a group of 8 healthy men, both PGE2 and PGF2 alpha excretion rates were higher and more scattered than the female values. When two healthy women were given indomethacin (200 mg/day), urinary PGE2 and PGF2 alpha dropped to undetectable levels during the 4th day of drug therapy. Intravenous injection of furosemide (50 mg) in a female volunteer was followed by an immediate rise of urinary sodium, PGE2, PGF2 alpha and PGE2/PGF2 alpha ratio and plasma renin activity. In a 10 year old girl with Bartter's syndrome, urinary PG excretion rate was elevated with a 3 times higher than normal PGE2/PGF2 alpha ratio. Indomethacin therapy resulted in a prompt drop of PG excretion rate and of plasma renin activity. These results show that a combination of adequate purification steps and antisera with optimal characteristics provides a reliable method for measuring PGE2 and PGF2 alpha in urine, and therefore a valuable tool to further our knowledge of the physiology and physiopathology of the renal PG-system.

Diabetic glomerulopathy is a common long-term complication of insulin-dependent diabetes mellitus [1]. Hyperfiltration, resulting from a fall of afferent arteriolar resistances with increased glomerular capillary pressure, is an early sign of glomerular involvement [1,2]. Such haemodynamic changes are believed to result from reduced vascular sensitivity to vasoconstrictors [3]. Arteriolar relaxation could be explained by enhanced production of vasodilators, such as prostaglandins [4], or atrial natriuretic peptides [5]. Alternatively, reduced responsiveness of the contractile apparatus of the glomerulus should be considered [3,6]. Diabetologia (1997) Summary In cultured mesangial cells (MC), capacitative Ca 2 + influx via store-operated channels (SOC) is potentiated by agents that release Ca 2 + from intracellular stores, and inhibited by protein kinase C (PKC). Cells grown under high glucose conditions, as a model of the diabetic microenvironment, display reduced Ca 2 + signalling in response to vasoconstrictors, probably due to downregulation by elevated PKC activity. Since SOC might be relevant to this phenomenon, we assessed Ca 2 + influx by microfluorometry of fura-2-loaded rat MC cultured for 5 days in normal (5.5 mmol/l, NG) or high glucose (30 mmol/l, HG). The addition of 1-10 mmol/l Ca 2 + to NG cells equilibrated in Ca 2 + -free media induced an immediate Ca 2 + influx with a free cytosolic Ca 2 + ([Ca 2 + ] i ) plateau of 155 ± 50 and 318 ± 114 nmol/l, respectively. Basal influx was reduced to 88 ± 8 and 145 ± 17 nmol/l [Ca 2 + ]i (1-10 mmol/l Ca 2 + , p < 0.01) by 30 mmol/l d -glucose. This effect of HG was confirmed by Mn 2 + quenching of fura-2, indicating reduced entry of divalent cations via the capacitative pathway. Equimolar l -glucose had no effect on Ca 2 + influx, consistent with a non-osmotic mechanism.Arginine vasopressin (10 m mol/l) elicited weaker release of stored Ca 2 + and subsequent influx in HG cells (191 ± 33 vs 153 ± 24 nmol/l, 400 ± 76 vs 260 ± 33 nmol/l, 1-10 mmol/l Ca 2 + , NG/HG, p < 0.05). To examine the involvement of PKC in the effect of HG on capacitative Ca 2 + influx, the enzyme was activated or downregulated by treatment with 0.1 m mol/l phorbol myristate acetate (PMA) for 3 min or 24 h, respectively. PMA acutely inhibited Ca 2 + influx in NG cells, while PKC downregulation restored it in HG cells. Similarly, the PKC inhibitors staurosporin or H-7 normalized SOC activity in HG cells. In summary, impairment of Ca 2 + influx via SOC by HG is one mechanism of the reduced MC [Ca 2 + ] i responsiveness to vasoconstrictors. This event is mediated by PKC and may contribute to the glomerular haemodynamic changes in the initial stages of diabetes mellitus. [Diabetologia (1997) 40: 521-527]

Ca2+ influx is a major component of the response of cultured human mesangial cells (HMC) to vasoconstrictors. Activators of phospholipase C such as angiotensin II (Ang II) release Ca2+ from intracellular stores and enhance Ca2+ influx, which in turn is modulated by Na+/Ca2+ exchange. By microfluorometry we studied the mechanisms of Ca2+ entry in resting and stimulated fura-2-loaded monolayers or single HMC. Addition of 1 to 10 mM extracellular Ca2+ to cells equilibrated in Ca(2+)-free media resulted in a rapid, persistent elevation of free cytosolic Ca2+ ([Ca2+]i), from 52 +/- 5 to 113 +/- 18 and 226 +/- 37 nM, respectively. Ca2+ influx was blocked by lanthanum or chelation with EGTA, while it was only partially inhibited by voltage-operated Ca2+ channel (VOC) blockers, such as nifedipine or verapamil. The rise of [Ca2+]i at high external [Ca2+] was not due to a Ca(2+)-sensing mechanism with release of intracellular stored Ca2+, since it was prolonged, and it was not seen in cells maintained in normal 1.25 mM [Ca2+] media. Moreover, it was not abolished by prior depletion of Ca2+ stores with 0.5 microM thapsigargin or 5 microM ionomycin in Ca(2+)-free media, which transiently increased [Ca2+]i (to 281 +/- 39 and 380 +/- 51 nM, respectively). On the contrary, both agents markedly potentiated Ca2+ influx upon addition of 1 to 10 mM [Ca2+]e, (to a maximum of 686 +/- 111 and 633 +/- 150 nM, P < 0.05 vs. control).(ABSTRACT TRUNCATED AT 250 WORDS)