Pharmacokinetics, distribution, metabolism and excretion of.

PURPOSE: To evaluate the metabolic fate of UCN-01, a signal transduction inhibitor, blood and plasma concentrations, distribution, metabolism and excretion were investigated in rats and dogs after intravenous administration of [3H]UCN-01. METHODS: The radioactivity in plasma, blood and tissues was measured after intravenous administration of UCN-01. In addition, the radioactivity excreted in bile, urine and feces was also determined. RESULTS: The radioactivity in rat and dog plasma disappeared triphasically with terminal half-lives of 21.3 and 27.2 h, respectively. The ratios of the blood-to-plasma concentrations ranged from 0.82 to 1.13 in rats and 0.81 to 1.73 in dogs. From 0.5 to 4 h after giving [3H]UCN-01 to rats, the radioactivity in all tissues except the brain and testes was higher than in plasma. The highest concentration was observed in the lungs followed by the liver and kidneys. The radioactivity was mainly excreted in feces, reaching 96.0% of the radioactivity dose in rats and 78.4% in dogs up to 168 h after injection. Since the biliary excreted radioactivity was 67.2% over 48 h in bile duct-cannulated rats, most of the radioactivity excreted in feces was from biliary radioactivity. There were several metabolites in bile samples, but little UCN-01. CONCLUSIONS: UCN-01 is mainly eliminated by the liver, and there are high concentrations of radioactivity derived from [3H]UCN-01 in all tissues except the brain and testes.

Inflammatory malignant fibrous histiocytoma of the ovary producing interleukin-6: A case report.

OBJECTIVE: Inflammatory malignant fibrous histiocytoma (IMFH) is composed of fibroblastic and histiocytic elements with a marked inflammatory cell infiltrate. The authors examined the case of a 43-year-old woman with IMFH of the ovary. METHODS: The serum interleukin-6 (IL-6) levels were examined and immunohistochemical analysis of the tumor was performed using monoclonal antibodies against lysozyme, alpha 1-antitripsin, vimentin, and IL-6. RESULTS: The patient had marked general inflammatory reactions of fever (38.0 degrees C), leukocytosis with 78.3% neutrophils, elevated CRP (23.5 mg/dl), and accelerated ESR (166 mm/h). The serum IL-6 concentration was 499 pg/ml. The right ovary was occupied by the solid tumor, composed of both fibrous and histiocytic elements with a marked inflammatory cell infiltrate. These inflammatory reactions disappeared after surgical resection and recurred when the tumor reappeared. The immunohistochemical staining of lysozyme, alpha 1-antitrypsin, vimentin, and IL-6 was localized to the malignant cells, suggesting their histiocytic origin. CONCLUSIONS: This is the first case of IMFH of the ovary producing IL-6. Expression of IL-6 in the IMFH may account for both the local and the systemic inflammatory effects of tumors with these morphological features.

Novel missense mutation (Leu466Arg) of the DAX1 gene in a patient with X-linked congenital adrenal hypoplasia.

We identified a DAX1 missense mutation, a substitution of arginine for leucine at codon 466 (Leu466Arg), in an infant with X-linked congenital adrenal hypoplasia (AHC). A heterozygous substitution, Leu466Arg, was also identified in his mother and sister. Since leucine at position 466 is well conserved among other orphan nuclear hormone receptor superfamilies and Leu466Arg was not detected among 50 normal Japanese control individuals, the mutation is most likely responsible for X-linked AHC. It is interesting to note that Leu466Arg among all mutations ever reported is located at the most C-terminal region of the DAX-1 protein. Most mutations identified previously were located in the C-terminal presumptive ligand binding domain. Hence, the C-terminal end of the DAX-1 protein may play an important role in the biological function, such as in normal adrenal embryogenesis.

Characterization of a newly established human tumor cell line (TEN) from a patient with clear cell carcinoma of the uterine body and its sensitivity to anti-cancer agents.

A cell line derived from human endometrial clear cell adenocarcinoma was newly established and named TEN. The tumor cells were obtained from uterine body of a 74-year-old who had been undergone an abdominal simple hysterectomy. The histologic features of the tumor cells showed abundant clear cytoplasm with diastase digested glycogen granule growing in solid nest and tubular pattern. The TEN cells were continuously propagated in vitro during the past 45 months and they were at 75th passage. They grew in a monolayered sheet with a doubling time of about 53 hours. The TEN cells resembled the structure of the original tumor and had abundant glycogen granules, lipid droplets in the cytoplasm. The histopathology of the transplanted tumor in SCID mice resembled that of the original tumors. The TEN cells secreted a high content of CA125. Immunohistochemically, the TEN cells had c-erbB-2 and Cathepsin D immunoreactivity in some parts of the cell population. But they did not have estrogen, progesterone and EGF receptor. Sensitivities of the TEN cells to a variety of anti-cancer drugs were examined. In in-vitro tests, MTT assays employed. The results suggested that the TEN cells were not sensitive to any of 13 agents. On the other hand, in-vivo sensitivity test of transplanted tumor in SCID mice, the tumors were sensitive to CPT-11 and paclitaxel. We conclude that the TEN cell line will be effective material for chemosensitivities against the endometrial clear cell adenocarcinoma.

[Serum eosinophil cationic protein in infants during first wheezing episode].

We measured serum ECP levels in infants during first wheezing episode. Serum ECP in these infants are significantly higher than in control infants, although much higher in children with asthma. Serum ECP in these infants with high serum IgE and/or positive RAST score are higher than in infants with normal serum IgE and negative RAST score. In children with bronchial asthma serum ECP is correlated with peripheral eosinophil counts, but in infants during first wheezing episode serum ECP is often elevated not associated with increased peripheral eosinophil counts. These suggest that activated eosinophils could be responsible for bronchoconstriction in wheezing patients with atopic diathesis even in very early phase and that these eosinophilic inflammations could contribute to formation of increased airway reactivity and bronchial asthma.

H+ secretion in rabbit mesonephric collecting tubule.

The mesonephros, the precursor of the metanephros, the definitive kidney, is the functional excretory organ in the 12- to 20-day-old rabbit fetus. It is believed to acidify allantoic fluid. To determine whether H+ excretion occurs in the distal nephron, we examined isolated perfused mesonephric collecting tubules by microcalorimetry and pH-sensitive fluorescent dyes. Collecting tubules secreted H+ (absorbed HCO3-) at rates twice those observed in the mature outer medullary collecting duct (OMCD) of the metanephric kidney. H+ secretion was not inhibited by ouabain (18.5 +/- 2.2 vs. 16.7 +/- 4.0 pmol.min-1.mm-1; n = 7, P = NS) but was reversibly inhibited by removing Cl- from the bathing solution (15.1 +/- 2.3 to -0.6 +/- 3.7 to 15.5 +/- 1.1 pmol.min-1.mm-1; n = 5, P < 0.05); luminal application of N-ethylmaleimide (NEM), an inhibitor of the H(+)-ATPase, also inhibited H+ secretion (n = 2). These results suggested that H+ secretion in the mesonephric collecting tubule is mediated by transporters similar to those of the OMCD. To test this hypothesis, we stained collecting tubules from 15-20 day embryos with 6-carboxyfluorescein (6-CF) diacetate to identify intercalated-like cells or perfused them with 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein acetoxymethyl ester (BCECF-AM) to measure intracellular pH (pHi). We found that 139 +/- 15 cells/mm concentrated 6-CF or BCECF, consistent with the phenotype of metanephric intercalated cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of a novel diuretic, 7-chloro-2,3-dihydro-1-(2-methylbenzoyl)-4(IH)-quinolinone-4-oxime-o- sulfonic acid, potassium salt (M17055) on Na+ and K+ transport in the distal nephron segments.

By using an in vitro microperfusion technique, we examined whether a novel loop diuretic, 7-Chloro-2,3-dihydro-1-(2-methylbenzoyl)-4(IH)-quinolinone-4-oxime-o-sul fonic acid, potassium salt (M17055), a derivative of quinolinone oxime sulfonic acids, affects Na+ and K+ transport in the distal nephron segments, including the cortical collecting duct and connecting tubule (CNT) isolated from rabbit kidneys. M17055 added to the lumen at 1 mM caused a positive deflection of transepithelial voltage (VT) by 2.2 +/- 0.4 mV. The response was less than that evoked by 10 microM amiloride (8.9 +/- 0.1 mV). In the collecting duct cell of the cortical collecting duct from normal rabbits, M17055 depolarized the basolateral membrane by 9.2 +/- 1.3 mV, whereas amiloride hyperpolarized it by 7.6 +/- 2.4 mV. In the cortical collecting duct from deoxycorticosterone acetate-treated rabbits, despite the fact that both agents depolarized the basolateral membrane of the collecting duct cell, amiloride consistently hyperpolarized the apical membrane, whereas M17055 did not cause any significant changes in apical membrane voltage. In the presence of 2mM Ba++ in the lumen, the apical membrane voltage deflection by M17055 was abolished. In addition, the magnitude of the apical membrane voltage deflection caused by an abrupt increase in luminal K+ concentration from 5 to 50 mM was significantly reduced. In the CNT, both amiloride and M17055 caused a positive deflection of VT. However, M17055 depolarized the basolateral membrane by 6.6 +/- 1.6 mV, whereas amiloride hyperpolarized it by 4.4 +/- 1.1 mV.(ABSTRACT TRUNCATED AT 250 WORDS)

Adaptation of rabbit cortical collecting duct to in vitro acid incubation.

Cortical collecting ducts (CCDs) isolated from acid-loaded rabbits and perfused in vitro absorb HCO3-, whereas CCDs from normal animals secrete HCO3-. We have previously shown that CCDs incubated in vitro for 3 h at pH 6.9 show a reduction in net (baseline and stimulated) HCO3- secretion. In this study we ascertained the minimum duration of an acidic stimulus necessary to induce adaptive changes in stimulated HCO3- secretion (determined in the absence of basolateral Cl-) and the roles of protein synthesis and cytoskeletal function in this process. CCDs incubated in acid (pH 6.8, HCO3- 6 mM) for 1 h followed by incubation at pH 7.4 (HCO3- 25 mM) for 2 h showed a 41% reduction in stimulated HCO3- secretion (P < 0.001), similar to that observed after 3 h of incubation at pH 6.8. However, this incubation protocol failed to enhance stimulated HCO3- absorption (determined in the absence of luminal Cl-). Addition of 10 microM anisomycin, a reversible inhibitor of protein synthesis, throughout the entire period of incubation (1 h at pH 6.8 plus 2 h at pH 7.4) blocked adaptive reduction in HCO3- secretion, as did exposure to anisomycin only during the initial 1 h of acid incubation. In contrast, anisomycin application during the 2-h incubation at pH 7.4 failed to block this adaptation of HCO3- secretion. Application of 4 microM actinomycin D, an inhibitor of DNA transcription, during the acid incubation also prevented the adaptive response, as did application during the total or during the 2-h pH 7.4 incubation period of 0.2 microM cytochalasin D, an inhibitor of actin filament function.(ABSTRACT TRUNCATED AT 250 WORDS)

Electrophysiological identification of alpha- and beta-intercalated cells and their distribution along the rabbit distal nephron segments.

By cable analysis and intracellular microelectrode impalement in the in vitro perfused renal tubule, we identified alpha- and beta-intercalated (IC) cells along the rabbit distal nephron segments, including the connecting tubule (CNT), the cortical collecting duct (CCD), and the outer medullary collecting duct in the inner stripe (OMCDi). IC cells were distinguished from collecting duct (CD) cells by a relatively low basolateral membrane potential (VB), a higher fractional apical membrane resistance, and apparent high Cl- conductances of the basolateral membrane. Two functionally different subtypes of IC cells in the CCD were identified based on different responses of VB upon reduction of the perfusate Cl- from 120 to 12 mM: the basolateral membrane of beta-IC cells was hyperpolarized, whereas that of alpha-IC cells was unchanged. This is in accord with the hypothesis that the apical membrane of beta-IC cells contains some Cl(-)-dependent entry processes, possibly a Cl-/HCO3- exchanger. Further characterization of electrical properties of both subtypes of IC cells were performed upon lowering bath or perfusate Cl- from 120 to 12 mM, and raising bath or perfusate K+ from 5 to 50 mM. A 10-fold increase in the perfusate K+ had no effect on VB in both subtypes of IC cells. Upon abrupt changes in Cl- or K+ concentration in the bath, a large or a small depolarization of the basolateral membrane, respectively, was observed in both subtypes of IC cells. The electrical properties of alpha- and beta-IC cells were similar among the distal nephron segments, but their distribution was different: in the CNT, which consists of IC cells and CNT cells, 97.3% (36/37) of IC cells were of the beta type. In the CCD, which consists of IC cells and CD cells, 79.8% (79/99) of IC cells were of the beta-type, whereas in the OMCDi 100% (19/19) were of the alpha type, suggesting that the beta type predominates in the earlier and the alpha type in the later segment.

Mechanism of water transport across the upper portion of the descending thin limb of long-looped nephron of hamsters.

The mechanisms of water transport across the upper portion of the descending limb of long-looped nephron (LDLu) were examined by microperfusion of segments isolated from hamster kidneys. Because of cation permselectivity a streaming voltage was generated when a transmural osmotic gradient was imposed. When 100 mmol/l urea was added to the bath, the streaming voltage was -4.9 +/- 0.4 mV. Addition of 10(-4) mol/l parachloromercuribenzene sulfonate (PCMBS) decreased the voltage to -2.4 +/- 0.7 mV. This effect was associated with changes in osmotic water permeability (Pf, 10(-3) cm/s) from 243 +/- 42 to 47 +/- 15. PCMBS also decreased the transmural diffusional water permeability (Pdw, 10(-3) cm/s) from 9.4 +/- 0.6 to 7.2 +/- 0.6. The inhibitory effect of PCMBS was prevented by pretreatment with dithiothreitol. N-Butanol permeability was measured as an index of cellular resistance for diffusion. Large differences between Pf and Pdw can be explained both by cellular resistance to diffusion and by resistance through a water channel with a single file mechanism. The apparent activation energy for water transport, 13.3 x 10(3) joule/mol (3.16 kcal/mol), was low. These findings are compatible with the hypothesis that a water channel exists in this segment. PCMBS also inhibited the NaCl diffusion voltage, a parameter indicating cation permselectivity, in parallel with suppression of the streaming voltage, suggesting that the water channel is in part associated with cation permselectivity. The possibility that the PCMB-sensitive cation-permselective pathway exists in parallel cannot be ruled out.