Improved specificity of a new homogeneous assay for LDL-cholesterol in serum with abnormal lipoproteins.

BACKGROUND: Although a homogeneous assay for serum LDL-cholesterol (LDL-C) has become a routine clinical procedure, problems remain in assay performance characteristics. METHODS: We examined the performance of a recently developed automated homogeneous assay (New-Daiichi assay) for serum LDL-C and compared the results with those obtained by the current homogeneous method (Denka-Seiken assay) or by ultracentrifugation as a control. RESULTS: The New-Daiichi assay showed satisfactory basic performance characteristics such as reproducibility, linearity, and stability. There was no interference in the assay by various substances examined. The LDL-C values obtained with this method correlated well with those obtained by ultracentrifugation. In samples from patients with obstructive jaundice, both methods detected cholesterol from abnormal lipoproteins (such as lipoprotein-X and -Y), but the New-Daiichi assay was less reactive and more specific for LDL-C. CONCLUSION: The new method has improved performance for the accurate measurement of LDL-C in clinical practice.

Nilvadipine inhibits nuclear factor-kappaB-dependent transcription in hepatic cells.

BACKGROUND: Recent findings suggest that some dihydropyridine-type calcium channel blockers, widely used as anti-hypertensive drugs, have direct anti-atherogenic action through their antioxidant properties. METHODS: We examined the effect of nilvadipine on the activity of a representative radical-sensitive transcription factor, nuclear factor kappa-B (NF-kappaB), in the human hepatocyte cell line HuH7 in vitro. RESULTS: Nilvadipine potently inhibited NF-kappaB-dependent transcription in a dose- and time-dependent manner, with a minimal effective concentration of 50 nmol/l. The effect was specific because no similar effects were found in the prototype dihydropyridine nifedipine. Electromobility shift assay showed reduced protein binding to the NF-kappaB-consensus sequence in nilvadipine-treated cells. Nilvadipine also reduced the expression of fibrinogen and plasminogen activator inhibitor-1 (PAI-1). CONCLUSIONS: Since NF-kappaB-mediated gene products, such as fibrinogen and PAI-1, are known to facilitate hypercoagulation, thrombosis and vascular events, we suggest that nilvadipine has a direct beneficial effect separate from its anti-hypertensive properties by inhibiting NF-kappaB-dependent gene expression and eventually inhibiting atherosclerosis.

Dehydroepiandrosterone-sulfate inhibits nuclear factor-kappaB-dependent transcription in hepatocytes, possibly through antioxidant effect.

Dehydroepiandrosterone (DHEA) and DHEA-sulfate (DHEAS), the representative sex steroid precursors, are postulated to have antiinflammatory effects, although the molecular background remains unknown. In this study, we examined the effects of these sex steroid precursors on cytokine-induced, nuclear factor-kappaB (NF-kappaB)-mediated transcription. The HuH7 human hepatocyte cell line was stably transfected with an NF-kappaB-luciferase reporter gene or transiently transfected with other representative response elements-luciferase fusion genes, and the effects of DHEA/DHEAS on proinflammatory cytokine-induced transcription were estimated by luciferase assay. The results showed that DHEA/DHEAS potently inhibited TNF-alpha-induced NF-kappaB-dependent transcription in a time- and dose-dependent manner. The effect was more obvious for DHEAS than for DHEA, and both steroids preferentially inhibited the cytokine-stimulated rather than basal NF-kappaB-mediated transcription. Similar effects were observed in activator protein-1-dependent but not constitutive Rous sarcoma virus promoter-dependent transcription. Two major downstream products of the sex steroid precursors, estradiol and testosterone, had no effect, indicating that the observed suppressive effect is not mediated by these metabolites. In contrast, glucocorticoids showed inhibitory effects on both basal and stimulated transcription and had an additive effect with DHEAS, suggesting the independent mechanisms of action of these steroid hormones. Finally, DHEAS eliminated hydroxyradical-induced activation of NF-kappaB-dependent transcription as well. Altogether, these results suggest that DHEA/DHEAS have an antiinflammatory effect in such a way that they inhibit proinflammatory cytokine-stimulated, NF-kappaB-mediated transcription, at least partly through their antioxidant properties.

Possible involvement of ryanodine receptor-mediated intracellular calcium release in the effect of corticotropin-releasing factor on adrenocorticotropin secretion.

We examined the role of intracellular calcium release in the regulation of CRH-induced ACTH secretion using the AtT20 corticotroph cell line. We found that ruthenium red, an inhibitor of ryanodine receptor, substantially diminished the secretory response, whereas Xestospongin C, an inositol 1,4,5-triphosphate receptor antagonist, had no effect. Expression of two ryanodine receptor subtypes (RyR1 and RyR3) was confirmed by RT-PCR. We also found that caffeine, a ryanodine receptor agonist, significantly stimulated, whereas thapsigargin, which causes depletion of intracellular calcium store, markedly diminished, the ACTH release. These results suggest that ryanodine receptor-mediated calcium-induced calcium release is involved in the regulation of CRH-induced ACTH release.

Sodium bicarbonate infusion test: a new method for evaluating parathyroid function.

We have developed a new test for estimating the secretory capacity of parathyroid hormone (PTH) from the parathyroid gland. Sodium bicarbonate solution [8.4% (w/v); 35 ml/m(2) body surface area] was infused for 2 min, and blood samples for the determination of plasma ionized calcium, plasma PTH (intact, midregion, carboxy-terminus) and related parameters were serially obtained. In 8 healthy volunteers, the mean (+/-SE) plasma ionized calcium fell promptly and significantly (from 1.21 +/- 0.01 to 1.11 +/- 0.01 mmol/L) after the sodium bicarbonate infusion. The mean (+/-SE) plasma intact PTH increased promptly and significantly, by more than four fold (42.3 +/- 4.2 to 182.4 +/- 34.7 pg/ml), and then gradually returned to basal levels. In patients with partial hypoparathyroidism who have detectable basal plasma levels of PTH, the absolute increment in PTH levels was much less, and in the plasma obtained from patients with complete hypoparathyroidism, absolutely no response was observed. Plasma obtained from patients diagnosed with primary hyperparathyroidism (parathyroid adenoma or hyperplasia) has high basal PTH levels. The response to the sodium bicarbonate infusion in these patients was markedly blunted (less than a two-fold increase in all cases examined). No significant adverse effects were observed during the procedure. Therefore, the sodium bicarbonate infusion test is a simple and sensitive method to stimulate PTH release, and is clinically useful for evaluating parathyroid gland function.

Composite tumor of mucinous cystadenoma and somatostatinoma of the kidney.

Approximately 30 cases of carcinoid tumor of the kidney have been reported in the English literature, including three cases found as components of teratomas. Renal composite tumors associated with somatostatinoma have not been described. A 53-year-old female presented with an incidentally found right renal cystic lesion. Computed tomography demonstrated a cystic lesion associated with a solid nodule in the right kidney and postcontrast dynamic MRI revealed enhancement of the solid nodule. The patient underwent radical nephrectomy for the kidney lesion and is now well without recurrence 21 months after the operation. From the histopathological findings we diagnosed the cystic lesion as a composite tumor composed of mucinous cystadenoma and carcinoid tumor. Immunohistochemistry demonstrated the majority of cells of in carcinoid portion to be positive for antisomatostatin staining. The present case is the first documented composite tumor of mucinous cystadenoma and somatostatinoma of the kidney.

Endometrial atypical hyperplasia with clear cell change spreading throughout the endometrium.

An unusually rare case of atypical endometrial hyperplasia with clear cell change (metaplasia) spreading through almost the whole endometrium of a 37-year-old woman with infertility and abnormal bleeding is reported.

Usefulness of p53 and Ki-67 immunohistochemical analysis for preoperative diagnosis of extremely well-differentiated gastric adenocarcinoma.

Of 987 cases of gastric adenocarcinoma seen at Nagoya University School of Medicine, we found 6 rare, extremely well-differentiated advanced gastric adenocarcinomas that could not be diagnosed as malignant tumors with only H&E staining, even with repeated biopsies under preoperative endoscopy. The aim of this study was to determine whether an immunohistochemical method using p53 and Ki-67 antibody would be helpfulfor preoperative pathologic diagnosis. The cancer control cases were 16 cases of ordinary well-differentiated advanced gastric adenocarcinoma, while the gastritis control cases were 22 cases of Helicobacter pylori-positive chronic gastritis. The p53 labeling index and the localization of Ki-67+ cells showed that the special adenocarcinomas in biopsy specimens were distinct from the surrounding normal mucosa and chronic gastritis, but not from the cancer control cases. These methods are useful markers for preoperative pathologic diagnosis of extremely well-differentiated gastric adenocarcinoma, which sometimes is confused with regenerative atypical glands before operation.

P53 labeling index in cholangioscopic biopsies is useful for determining spread of bile duct carcinomas.

BACKGROUND: Preoperative biopsy specimens obtained by means of percutaneous transhepatic cholangioscopy are useful for planning curative resection of bile duct carcinoma in an effort to improve survival. However, tissue diagnosis is sometimes difficult. This study evaluated the usefulness of p53 immunostaining of cholangioscopic specimens for determination of tumor spread. METHODS: A total of 107 biopsy specimens from 28 patients with bile duct carcinoma was selected. Before surgery, these specimens were diagnosed histopathologically (hematoxylin and eosin staining) as positive or negative for carcinoma. After definitive surgery, specimens were immunostained with anti-p53 antibody. RESULTS: Eighteen of 28 cases (64%) were positive for p53. Among these, 86% obtained from the main carcinomatous lesion or an area of superficial spread of the carcinoma exhibited a p53 labeling index (LI) over 25% as opposed to a p53 LI under 25% for all specimens obtained from noncarcinomatous lesions. Twelve specimens from 8 cases were classified before surgery as indeterminate (hematoxylin and eosin staining). The criterion of p53 LI over 25% was applicable in 11 of the 12 specimens. CONCLUSION: The p53 immunostaining of biopsy specimens obtained by means of percutaneous transhepatic cholangioscopy is helpful in determining tumor spread in bile duct carcinoma.

AT1 receptor blockade reduces cardiac calcineurin activity in hypertensive rats.

The possible role of calcineurin in the attenuation of cardiac hypertrophy and fibrosis by blockade of the angiotensin II type 1 (AT1) receptor was investigated in Dahl salt-sensitive (DS) rats. The effect of the calcineurin inhibitor FK506 was also studied. DS rats progressively developed severe hypertension when fed a diet containing 8% NaCl from 7 weeks of age. In addition, marked cardiac hypertrophy and fibrosis were apparent and the activity of calcineurin and its mRNA expression in the myocardium was increased in these animals at 12 weeks in comparison with age-matched Dahl salt-resistant rats. The abundance of angiotensin-converting enzyme (ACE) and transforming growth factor (TGF)-beta1 mRNAs was also increased in the hearts of DS rats at 12 weeks. Treatment of DS rats with a non-antihypertensive dose of the selective AT1 receptor blocker candesartan (1 mg/kg per day) or FK506 (0.1 mg/kg per day) from 7 to 12 weeks attenuated both calcineurin activity and its mRNA expression in the heart, as well as the development of cardiac hypertrophy and fibrosis, without affecting cardiac function. Treatment with candesartan, but not FK506, prevented the upregulation of ACE and TGF-beta1 gene expression. Both candesartan and FK506 prevented the load-induced induction of fetal-type cardiac genes. These results demonstrate that AT1 receptor blockade attenuates the development of cardiac hypertrophy and fibrosis as well as the activation of calcineurin, without an antihypertensive effect, in rats with salt-sensitive hypertension. Calcineurin may be downstream from TGF-beta1 in AT1 receptor-mediated angiotensin II signaling in vivo.