Ischemia-induced enhancement of CFTR expression on the plasma membrane in neonatal rat ventricular myocytes

Pathophysiological functions of cardiac cystic fibrosis transmembrane conductance regulator (cCFTR) in ischemia are not well known. Using neonatal rat ventricular cardiomyocytes in primary culture in this study, we thus examined whether the CFTR protein is expressed and is functioning as a cAMP-activated anion channel on the plasma membrane under ischemic conditions. After the cells were subjected to simulated ischemia (O(2) and glucose deprivation), an up-regulation of the CFTR expression was transiently observed in the membrane fraction by Western blot. A peak expression of mature CFTR protein was found at 3 h of ischemia, and thereafter the signal diminished gradually. In contrast, the results of Northern blot indicated that the expression level of CFTR mRNA changed little until 3 h of ischemia, whereas the level slightly decreased after 8 h of ischemia. An immunohistochemical examination showed, in agreement with the results of Western blot analysis, that the expression of CFTR protein on the plasma membrane became most prominent at 3 h of ischemia, whereas the plasmalemmal CFTR signal was markedly reduced after 8 h of ischemia. Whole-cell recordings showed that the cardiomyocytes responded to cAMP with an activation of time- and voltage-independent currents that contained an anion-selective component sensitive to CFTR Cl(-) channel blockers (NPPB and glibenclamide) but not to a stilbene-derivative conventional Cl(-) channel blocker (SITS). This cAMP-activated Cl(-) channel current was found to be enhanced after an application of ischemic stress for 3 to 4 h. These findings indicate that a plasmalemmal expression of CFTR is transiently enhanced under glucose-free hypoxic conditions presumably because of a posttranslational control.     

Development of a contamination free 6 valve injector inline monitoring system for endotoxin measurement in dialysate

Use of dialysate as supplement fluid in hemodiafiltration requires controlling contamination by endotoxin of the dialysate. We thus aimed at developing an endotoxin monitoring system with complete exclusion of endotoxin contamination for simple, easy, and accurate measurement of endotoxin concentration in dialysate. In the present study, we used a 6 valve injector along with a high performance liquid chromatogram system. This new system showed a sensitivity of approximately 1 endotoxin units (EU)/L in the range of 0 to 30 EU/L endotoxin in dialysate and no trace of endotoxin contamination. In conclusion, the new endotoxin monitoring system showed high sensitivity and reproducibility, with easy operation.     

Two newly identified genomic mutations in a Japanese female patient with fructose-1,6-bisphosphatase (FBPase) deficiency

Fructose-1,6-bisphosphatase (FBPase) (EC 3.1.3.11) catalyzes the splitting of fructose-1,6-bisphosphate into fructose 6-phosphate and inorganic phosphate. FBPase deficiency is an autosomal recessive inherited disorder caused by distraction of the fructose-1,6-bisphosphatase 1 gene (FBP1) and features severely impaired gluconeogenesis. We studied a female patient with typical FBPase deficiency symptoms. The FBPase activity of her peripheral white blood cells was undetectable. Genetic analyses of FBP1 revealed her to be a compound-heterozygote of two new mutations F194S and P284R. Gene tracking in the family revealed the mother to be a heterozygote of F194S, and the father and a sister to be heterozygotes of P284R. As both Phe194 and Pro284 of FBPase are highly conserved in many species and close to crucial amino acid residues to FBPase functions, these mutations could be responsible for the loss of FBPase activities.     

Hypercalcemia in sarcoidosis

Although hypercalcemia has long been recognized as a complication of sarcoidosis, the incidence of hypercalcemia (> or = 11 mg/dl) in Japan is probably less than 5%. 1 alpha, 25(OH)2D3 is the main cause for hypercalcemia in sarcoidosis and overproduced by sarcoid granulomata. Gamma-interferon produced by activated lymphocytes and macrophages plays a major role in the synthesis of 1 alpha, 25(OH)2D3. PTH release is down regulated by high serum concentration of 1 alpha, 25(OH)2D3. Parathyroid hormone related protein may also contribute to the hypercalcemia of sarcoidosis. Treatment of hypercalcemia and hypercalciuria consists of a low calcium diet, adequate hydration, minimization of exposure to sunlight and reducing overproduction of 1 alpha, 25(OH)2D3. Prednisone, 15 to 25 mg/day, is the drug of choice to reduce the overproduction of 1 alpha, 25(OH)2D3.     

Neurosarcoidosis

Sarcoidosis is a chronic systemic disease of unknown causes characterized by noncauseating granuloma in various organs. Neurological involvement occurs in 5-6% of patients with sarcoidosis. Most patients with neurosarcoidosis have extraneurologic abnormalities and extraneurologic biopsies usually support the diagnosis, however, nearly half of the patients with neurosarcoidosis present with neurological manifestations without systemic symptoms. Several cases of isolated neurosarcoidosis have been reported, making the diagnosis difficult. Although several diagnostic criteria have been proposed, neuropathological examination is required for the definite diagnosis. Brain biopsy may still be required in selected patients. Since it is often difficult to perform, clinical assessment with various combinations of modern neuroimaging techniques is important. Lumbar puncture is useful to rule out other diseases but CSF changes are not specific. Elevated serum ACE level is also not a specific diagnosis test. Any part of the CNS can be involved, but there is predilection for hypothalamus, pituitary gland, leptomeninges, and cranial nerves particularly facial nerve. Although corticosteroids are the main stay of therapy, refractory cases can be treated by immunosuppressive and radiation therapy.     

No increase of blocking type anti-thyrotropin receptor antibodies during pregnancy in patients with Graves' disease

Serial changes in serum levels of anti-TSH receptor antibodies were examined during and after pregnancy in six patients with Graves' disease receiving no or minimal maintenance doses of antithyroid drugs. During pregnancy, serum levels of TSH-binding inhibitory Igs (P < 0.001) and thyroid-stimulating antibodies (TSAbs) (P < 0.01) decreased gradually but increased after delivery in all patients. Activities of thyroid-stimulation blocking antibodies (TSBAbs) were lower than the cut-off value in early pregnancy, and values significantly decreased in four patients during pregnancy. The other two patients showed no significant change during pregnancy. In contrast, TSBAb levels increased significantly (P < 0.01) after delivery in all patients. We found that activities of TSH-binding inhibitory Igs, TSAb, and TSBAb decrease during pregnancy and increase after delivery, suggesting that amelioration of Graves' disease during pregnancy is induced by decrease of TSAb but not by the appearance of TSBAb.     

Development of a new extracorporeal whole-liver perfusion system

We have developed a new extracorporeal whole-liver accommodation device in which a whole swine liver is placed in a physiological state by modeling the intraabdominal arrangement in the pig body, with the liver supported by a special inferior vena cava tube. Furthermore, we employed a diaphragm-type artificial heart in our system to produce pulsatile blood flow through the hepatic artery, which is considered to be indispensable to dilate peripheral vessels and supply oxygenated whole blood to the peripheral liver tissue. Beneficial effects were demonstrated in visual findings and bile juice secretion. The color of the liver surface in our system remained bright red, indicating that the liver vessels were well drained and free from congestion, and bile juice secretion was maintained at more than 10ml/h throughout the perfusion period. Our system exhibited excellent ammonia removal and urea nitrogen synthesis, and serum aspartate aminotransferase levels showed no increase, indicating the absence of hepatocyte destruction. Histological findings showed that the liver could expand appropriately and was free from compression caused by its own weight. In conclusion, our original liver accommodation device enabled appropriate expansion of the whole liver and supplied adequate oxygenated blood to peripheral areas by means of a pulsatile pump.