Pre-clinical evaluation of a new orally-active CCK-2R antagonist, Z-360, in gastrointestinal cancer models.

BACKGROUND: Gastrin has a role in gastrointestinal (GI) malignancy. This study provides pre-clinical evaluation of a novel, orally-active gastrin/cholecystokinin-2 receptor (CCK-2R) antagonist, Z-360. METHODS: (125)I gastrin-17 (G17) displacement and G17-stimulated calcium assays were used in classical CCK-2R-transfected cell lines. Akt phosphorylation was assessed by Western blotting. Z-360 efficacy in vivo was evaluated in three human xenograft models, and microvessel density and apoptosis in these models were investigated by immunohistochemistry. RESULTS: Z-360 inhibited (125)I G17 binding to cells expressing CCK-2R, and G17-stimulated signalling. Reduced Akt phosphorylation in an oesophageal cell-line treated with Z-360 was reversed by co-treatment with G17. Z-360 increased survival in a gastric ascites model (p=0.011) and decreased tumour growth in a hepatic metastasis model (81%, p=0.02). In an orthotopic pancreatic model, Z-360 combined with gemcitabine decreased final tumour weight compared to single agents (84%, p=0.002) and there was increased apoptosis and decreased microvessel density in ex vivo tumour tissue. CONCLUSIONS: These results show that the orally-active CCK-2R antagonist, Z-360 has high sub-nM affinity for classical CCK-2R, is well tolerated in vivo and exerts an anti-tumour effect.

Comparison of (18)F-FDG, (131)I-Na, and (201)Tl in diagnosis of recurrent or metastatic thyroid carcinoma.

UNLABELLED: There are several reports about the usefulness of (18)F-FDG PET in thyroid cancer. However, few studies have compared FDG PET with (131)I and (201)Tl scintigraphy. The aim of this study was to evaluate the clinical significance of whole-body FDG PET in differentiated thyroid cancer and to compare the results with those obtained from (131)I and (201)Tl scintigraphy. METHODS: Whole-body FDG PET was performed on 32 patients (10 men, 22 women; age range, 30-77 y; mean age, 54 y) with differentiated thyroid cancer (5 cases of follicular cancer and 27 of papillary cancer) after total thyroidectomy. An overall clinical evaluation was performed, including cytology, thyroglobulin level, sonography, MRI, and CT, to allow a comparison with functional imaging results for each patient. Metastatic regions were divided into five areas: neck, lung, mediastinum, bone, and other. Multiple lesions in one area were defined as one lesion. The tumor-to-background ratio (TBR) was measured for the lesions that were positive for both (201)Tl uptake and FDG PET uptake. RESULTS: The number of lesions totaled 47. Forty-one (87%) were detected by all scintigraphic methods. FDG uptake was concordant with (131)I uptake in only 18 lesions (38%). FDG uptake was concordant with (201)Tl uptake in 44 lesions (94%). Only one lesion was negative for FDG uptake and positive for (201)Tl uptake, and two lesions were positive for FDG uptake and negative for (201)Tl uptake. A significant correlation was seen between the TBR of (201)Tl and that of FDG (r = 0.69; P<0.05). CONCLUSION: These data indicate that for detecting metastatic lesions, FDG PET and (131)I scintigraphy may provide complementary information, whereas FDG PET may provide results similar to those of (201)Tl scintigraphy. Thus, the combination of (131)I scintigraphy and FDG PET (or (201)Tl scintigraphy) is the method of choice for detecting metastatic thyroid cancer after total thyroidectomy.

[The false positive reaction of the Triage panel drug-of-abuse by herbal drugs ma-huang (Ephedra sinica (Ephedraceae))].

We investigated false-positive reactions obtained from a drug screening test using a Triage panel. We detected 2 cases giving false-positive reaction for AMP (amphetamine, methamphetamine) during the screening of 187 normal subjects. Subsequent follow up testing by high-performance liquid chromatography (HPLC), showed both to be false-positive reactions. As both cases have a history of ingesting the herbal drug, Ma-huang (Ephedra sinica (Ephedraceae)), containing ephedrine, we examined the relationship between false-positive reactions on Triage and Ma-huang. All urine samples collected from 7 healthy volunteers following administration of Ma-huang indicated AMP positive on Triage. Also a high ratio of AMP positives was observed in the patients who were administered Ma-huang-containing drugs at the hospital. However, none of them were identified as true-positives by HPLC or gas chromatography mass spectrometry (GC/MS) analysis. The extract of Ma-huang contained in herbal drugs, which otherwise contain neither amphetamine nor its derivatives, gives (AMP) positive indications on Triage. We speculate that unidentified components of Ma-huang cause the false-positive reactions. We suggest that follow-up tests by GC/MS or HPLC are needed wherever a positive result is obtained from a screening test by Triage. Furthermore, it will be established to continue collecting information on prescribed and non-prescribed drugs.

Beta ray-induced scission of DNA in tritiated water and protection by a green tea percolate and (-)-epigallocatechin gallate.

The beta-ray induced scission of puC18 plasmid DNA from E. coli in tritiated water was examined in the presence or absence of a green tea percolate (TP) and the main constituent, (-)-epigallocatechin gallate (EGCg). An analysis of the ratio of the original closed-circular to the open-circular form of DNA, which was formed by the strand scission of DNA, revealed that TP and EGCg showed a protective effect on DNA scission depending on their concentrations. A new technique, named solid state spin trapping, was applied to examine this scavenging ability toward the hydroxyl (OH) radical generated in tritiated water. The result was kinetically analyzed to reveal that TP and EGCg showed the scavenging effect, suggesting that the protective effect on DNA scission was attributable to the scavenging effect on the OH radical.

Protecting effect of a green tea percolate and its main constituents against gamma ray-induced scission of DNA.

Gamma ray-induced scission of puC18 plasmid DNA prepared from E. coli was examined in the presence of a green tea percolate and its main constituents, L-ascorbic acid (used as the sodium salt) and (-)-epigallocatechin gallate. Each of these showed a protecting effect against DNA scission. The relationship between the protecting effect against DNA scission and the scavenging effect of the hydroxyl radical was examined, and is discussed from the viewpoint of interaction with DNA.

Effects of Chinese herbal drugs on serum lipids, lipoproteins and apolipoproteins in mild to moderate essential hypertensive patients.

We investigated the effects of the traditional Chinese herbal drugs, Dai-saiko-to (D) and Saiko-ka-ryukotsuboreito (S) on blood pressure, pulse rates, serum lipids, lipoproteins and apolipoproteins in 30 patients with mild to moderate hypertension in an open, randomised trial. After the drug treatment, BP remained unchanged, but pulse rates declined significantly after 3 months in the S treated group. Serum total cholesterol and triglyceride values did not change, but high density lipoprotein-cholesterol increased significantly (P < 0.05) in both groups. Apo-AI (P < 0.1 in S group) and apo-AII (P < 0.05 in D group, P < 0.1 in S group) tended to increase 3 months after treatment. These data indicate that both of these traditional Chinese medicines have a preferential effect on lipid metabolism with little antihypertensive action.

Reversion of thyrotoxic atrial fibrillation in hypothyroid state after radioiodine treatment.

Twenty patients with thyrotoxic Basedow's disease complicated by atrial fibrillation lasting more than one month despite treatment with antithyroidal drugs were treated with radioiodine supplemented with an antithyroidal drug or inorganic iodine. We classified the 20 patients on the basis of atrial fibrillation reversion into two groups, one with reversion (group I) and the other without reversion (group II). In all 12 patients in group I, T4 and T3 decreased to hypothyroid levels in 3.2 +/- 1.3 months, and one month later all patients had their sinus rhythm restored while T4 and T3 remained below normal (2.6 +/- 1.1 micrograms/dl and 77.9 +/- 34.4 ng/dl, respectively). Although T4 and T3 also decreased within 3.5 +/- 1.8 months in all 8 patients in group II, one month later, atrial fibrillation persisted while T4 and T3 (10.4 +/- 5.3 micrograms/dl and 157.7 +/- 67.5 ng/dl, respectively) rose significantly compared to those in group I (P less than 0.001 and P less than 0.01, respectively). For reversion of atrial fibrillation it is important that the onset of hypothyroidism is rapidly induced by radioiodine and that hypothyroidism continues for at least one month.

Plasma vasopressin and atrial natriuretic hormone levels in hypopituitarism with and without hydrocortisone treatments: responses to an acute water load.

To assess whether arginine vasopressin and atrial natriuretic hormone participate in impaired urinary dilution and excretion in glucocorticoid deficiency secondary to hypopituitarism, an acute oral water load of 20 ml.kg-1 BW was undertaken in the absence and presence of an oral hydrocortisone (60 mg) treatment in patients with ACTH deficiency (N = 7) and panhypopituitarism (N = 2). Plasma arginine vasopressin and atrial natriuretic hormone and renal water handling were simultaneously determined and compared with those in similarly water-loaded normal subjects. Plasma arginine vasopressin did not fall in response to decreased blood osmolality after an acute water load in the absence of hydrocortisone; plasma atrial natriuretic hormone did not change despite blood volume expansion; and impairment in urinary dilution and excretion remained. On the other hand, in the presence of hydrocortisone, plasma arginine vasopressin fell in response to a decrease in plasma osmolality and plasma atrial natriuretic hormone increased, thereby restoring urinary dilution and excretion. These results demonstrate that the impaired arginine vasopressin response to acute water loading play an essential role in deranged renal water and electrolyte handling in the state of glucocorticoid deficiency; the impaired release of atrial natriuretic hormone also may affect these disorders.

Effects of Chinese herbal drug, dai-saiko-to on plasma lipids, lipoproteins and liver lipid contents in guinea pig.

Effects of Chinese herbal drug, Dai-saiko-to, on plasma lipids, lipoproteins and liver lipid contents were investigated in guinea pig with/without cholesterol feeding. Guinea pigs were divided into 4 groups (Group 1: normal chow (N) diet, Group 2: N + 1% Dai-saiko-to (D) diet, Group 3: 1% cholesterol (C) diet, Group 4: 1% C+ 1% D diet), and were treated for 5 weeks. Significant body weight reductions were observed in Groups 3 and 4 compared to Groups 1 and 2. Significant reductions of plasma triglyceride (TG) and very low and low density lipoproteins [(V) LDL]-TG were found in Group 3 compared to Groups 1, 2, and 4. (HDL)-TG significantly decreased in Groups 3 and 4 compared to Groups 1 and 2, but no significant difference was noted between Groups 1 and 2, and Groups 3 and 4. Liver lipid contents analysis showed no significant changes between Groups 1 and 2, and Groups 3 and 4, respectively, but the distribution of lipid droplets as pathologically determined using a computer program showed Group 4 had less lipid deposition compared to Group 3. These data suggest that Dai-saiko-to acts on triglyceride metabolism in hypercholesterolemic guinea pigs.

Calcitonin gene-related peptide in the human hypothalamus.

Calcitonin gene-related peptide (CGRP)-like immunoreactivity (LI) in the human hypothalamus was investigated by radioimmunoassay and by immunocytochemistry. CGRP-LI was detected from two hypothalami obtained at autopsy (2.1 and 7.0 ng/g wet tissue) by radioimmunoassay. Reverse phase high performance liquid chromatography revealed that most of the CGRP-LI in the human hypothalamus was eluted in an identical position with synthetic human CGRP. For immunocytochemistry, human hypothalami obtained at autopsy were fixed and cryostat-sectioned at 40 microns. Free floating sections were immunostained with antibody to CGRP. CGRP-immunoreactive cell bodies were found in the supraoptic nucleus, paraventricular nucleus and infundibular nucleus. These findings indicate that CGRP exists in the cell bodies of the supraoptic nucleus, paraventricular nucleus and infundibular nucleus in the human hypothalamus and CGRP may play some roles in the endocrine and other functions of the human hypothalamus.

Immunoreactive somatostatin in male reproductive system in humans.

Somatostatin-like immunoreactivity is distributed widely in humans; the highest concentration is in the central nervous system, gastrointestinal tract, and pancreas. To determine if somatostatin is present in the male reproductive system, we analyzed human testis, epididymis, prostate, and semen. Somatostatin-like immunoreactivity was detectable in acid extracts of human testis, epididymis, and prostate (n = 6 each) in concentrations of 4.0 +/- 1.4 (+/- SD), 14.7 +/- 3.2, and 27.5 +/- 5.1 pmol/g wet wt, respectively. Considerable amounts of immunoreactive somatostatin also were detectable in semen; the mean value was 3.8 +/- 1.3 nmol/L (n = 6). This value was 200-fold higher than that in peripheral plasma. The somatostatin immunoreactivity in these tissues was characterized by gel filtration chromatography. Two peaks of somatostatin immunoreactivity, one coeluting with somatostatin-14 and the other with somatostatin-28, were found in the testis, epididymis, prostate, and hypothalamus. The amounts of the two sizes were nearly equal in the testis; somatostatin-14 predominated in the epididymis, prostate, and hypothalamus; whereas only somatostatin-28 was detected in semen. The presence of somatostatin in the male reproductive system suggests that somatostatin may play a role in the regulation of reproductive function in men.

Corticotropin-releasing hormone in the human hypothalamus. Free-floating immunostaining method.

We have clearly demonstrated corticotropin-releasing hormone (CRH) immunoreactive cell bodies and nerve fibers in the human hypothalamus by immunocytochemistry using free-floating sections instead of paraffin-embedded sections. Human hypothalami were obtained at autopsy, fixed and cryostat-sectioned at 40 microns. Free-floating sections were immunostained with antibody to CRH using the Vector ABC system. Most of CRH immunoreactive nerve fibers from the paraventricular nucleus pass under the fornix, while some CRH immunoreactive nerve fibers pass beyond the fornix and some through the fornix. Then the CRH immunoreactive nerve fibers run downward, medially to the supraoptic nucleus and toward the pituitary stalk. This method of immunocytochemistry is a very sensitive and suitable means for immunocytochemical studies of neuropeptides in the human brain.

Multiple forms of immunoreactive growth hormone-releasing hormone in human plasma, hypothalamus, and tumor tissues.

We examined the nature of GHRH in plasma of normal subjects and patients with acromegaly, hypothalamic tissue, pheochromocytoma, and GHRH-producing pancreatic tumor tissue using two RIAs of different specificity. One assay was a N-terminal assay that recognized GHRH-(1-44)-NH2, GHRH-(1-40)-OH, and GHRH-(1-37)-OH equally, and the other was a C-terminal assay that recognized only the COOH-terminal amidated sequence of GHRH-(1-44)-NH2. GHRH immunoreactivity was detectable in all samples in both assay systems, but the ratios of C- to N-terminal activity differed. The gel filtration profiles of plasma and tumor tissue revealed one peak in (or near) the position of synthetic GHRH-(1-44)-NH2. In contrast, two peaks were found in hypothalamic tissue; a major peak in the position of synthetic GHRH-(1-44)-NH2 and a higher mol wt peak. Ion exchange chromatography of the immunoreactive GHRH material from gel filtration of pooled plasma from normal subjects revealed three components of immunoreactive GHRH, one major peak in the position of GHRH-(1-40)-OH and two minor peaks in the positions of GHRH-(1-44)-NH2 and GHRH-(1-37)-OH. Two components of immunoreactive GHRH, a major peak in the position of GHRH-(1-44)-NH2 and a minor peak in the position of GHRH-(1-40)-OH, were found in hypothalamic tissue and pheochromocytomas. In the two ectopic GHRH-producing pancreatic tumors, three components of immunoreactive GHRH were detected: a major peak in the position of GHRH-(1-40)-OH, a smaller peak in the position of GHRH-(1-37)-OH, and a very small peak of GHRH-(1-44)-NH2. Synthetic GHRH-(1-44)-NH2 was not degraded by plasma during the extraction procedures. These results suggest that 1) the measured immunoreactive GHRH concentration differs when the same samples are measured by RIAs using antisera with different specificities; 2) such differences may be due to the presence of microheterogeneity of immunoreactive GHRH; 3) the microheterogeneity of immunoreactive GHRH in plasma is different from that in the hypothalamus; and 4) the posttranslational processing of GHRH in human hypothalamus is similar to that of pheochromocytomas but different from that of ectopic GHRH-producing pancreatic tumors.

Effects of active vitamin D3 and parathyroid hormone on the serum osteocalcin in idiopathic hypoparathyroidism and pseudohypoparathyroidism.

Serum osteocalcin was measured in patients with idiopathic hypoparathyroidism or pseudohypoparathyroidism, before or during the treatment with active vitamin D3 (1,25(OH)2D3 or 1 alpha OHD3). Serum osteocalcin and plasma 1,25(OH)2D were decreased in 11 patients with idiopathic hypoparathyroidism before treatment (2.8 +/- 1.27 ng/ml, P less than 0.001 and 14.3 +/- 4.27 pg/ml, P less than 0.001, respectively). In 24 patients with idiopathic hypoparathyroidism during the treatment, serum osteocalcin and plasma 1,25(OH)2D were within the normal range (4.5 +/- 0.74 ng/ml and 25.7 +/- 5.69 pg/ml, respectively). In five patients with pseudohypoparathyroidism before treatment, plasma 1,25(OH)2D was decreased (15.6 +/- 10.6 pg/ml, P less than 0.001) but serum osteocalcin was normal (7.8 +/- 1.66 ng/ml). In nine patients with pseudohypoparathyroidism during the treatment with active vitamin D3, serum osteocalcin and plasma 1,25(OH)2D were normal (6.8 +/- 1.47 ng/ml and 27.2 +/- 6.0 pg/ml, respectively). Serum PTH in pseudohypoparathyroidism was increased before treatment (0.70 +/- 0.34 ng/ml, P less than 0.05) and was normal during the treatment (0.50 +/- 0.13 ng/ml). In idiopathic hypoparathyroidism, the active vitamin D3 increased serum osteocalcin without PTH. In pseudohypoparathyroidism, PTH may increase serum osteocalcin or modulate the effect of active vitamin D3 on serum osteocalcin.

Potentiation of peptide histidine methionine (PHM) on corticotropin-releasing hormone (CRH) induced ACTH secretion in humans.

The effects of rat corticotropin-releasing hormone (CRH) and of synthetic peptide histidine methionine (PHM) on ACTH release were studied in vivo in humans. PHM alone did not alter the basal plasma ACTH or cortisol levels, but it strongly potentiated CRH-induced ACTH secretion. These results, together with our previous observation that PHM immunoreactivity is present in human hypothalamus and pituitary stalk, suggest that PHM may play a physiological role in the regulation of ACTH secretion in humans.

Hypotensive and natriuretic effects of nifedipine in essential hypertension. Role of renal kallikrein-kinin-prostaglandin and renin-angiotensin-aldosterone systems.

To assess the role of renal kallikrein-kinin-prostaglandin and renin-angiotensin-aldosterone systems in the diuretic and natriuretic actions of nifedipine, a calcium-channel blocker, 20 mg of nifedipine was administered orally to 15 patients with essential hypertension. Nifedipine promptly induced a hypotensive effect and an increase in pulse rate. Urine volume, urinary sodium excretion, and creatinine clearance were significantly increased after the administration of nifedipine by 63.5%, 48.5% and 12.4%, respectively. Urinary excretion of kallikrein and prostaglandin E were also significantly increased after the administration of nifedipine by 29.4% and 50.0%, respectively. The change in urinary kallikrein excretion was significantly correlated with that in urine volume (r = 0.70, p less than 0.01) or that in urinary sodium excretion (r = 0.86, p less than 0.01). In addition, the change in urinary prostaglandin E excretion was also significantly correlated with that in urine volume (r = 0.72, p less than 0.01) or that in urinary sodium excretion (r = 0.53, p less than 0.05). Plasma aldosterone concentration did not change despite of the marked increase in plasma renin activity, and plasma aldosterone concentration/plasma renin activity ratio decreased after the administration of nifedipine. These results suggest that the augmented renal kallikrein-kinin-prostaglandin system and the suppressed secretion of aldosterone may be associated with the diuretic and natiuretic action of nifedipine and may contribute to the reduction in blood pressure that is caused mainly by its vasodilatory action.

Immunocytochemical identification of CRF in the human hypothalamus.

Immunoreactive ovine corticotropin-releasing factor (CRF) was revealed in the cell bodies of parvocellular neurons in the paraventricular nucleus of the human hypothalamus by an immunocytochemical technique. The immunoreactivity was negative in magnocellular neurons. Immunoreactive nerve fibers were found in the subependymal layers of the third ventricle and in the perivascular space of the primary plexus in the lower hypothalamus. No CRF immunoreactive cell bodies were found in the supraoptic nucleus.

Carnitine depletion as a probable cause of hyperlipidemia in uremic patients on maintenance hemodialysis.

The mechanism of te development of hemodialysis hyperlipidemia was investigated in uremic patients on maintenance hemodialysis. Hemodialysis treatment lost large amounts of carnitine from blood into the dialysate fluid, resulting in the reduction in serum concentration of carnitine. After the treatments were repeated for more than 12 months, the serum concentration of carnitine reduced markedly and the serum triglyceride level increased significantly. In contrast, in patients who had been supplemented with commercial amino acids solution, the serum concentrations of carnitine and lipid were within normal ranges and remained unchanged even after repeated hemodialysis treatments. Carnitine administration also reduced the serum triglyceride level to or towards normal. The results suggest that carnitine depletion induced by hemodialysis treatments has a probable causal relationship to hyperlipidemia in uremic patients on long-term maintenance hemodialysis and that supplementation of carnitine or amino acids prevents carnitine depletion and improves hemodialysis hyperlipidemia.