Age-related oxidative stress and antioxidant capacity in heat-stressed broilers.

We aimed to evaluate the effects of acute heat stress (HS) and age on the redox state in broilers aged 21 and 42 days. We evaluated the expression of genes related to antioxidant capacity, the production of hydrogen peroxide (H2O2), and the activity of antioxidant enzymes in the liver, as well as oxidative stress markers in the liver and plasma. The experiment had a completely randomized factorial design with two thermal environments (thermoneutral and HS, 38°C for 24 h) and two ages (21 and 42 days). Twenty-one-day-old animals exposed to HS showed the highest thioredoxin reductase 1 (TrxR1) (P<0.0001) and glutathione synthetase (GSS) (P<0.0001) gene expression levels. Age influenced the expression of the thioredoxin (Trx) (P=0.0090), superoxide dismutase (SOD) (P=0.0194), glutathione reductase (GSR) (P<0.0001) and glutathione peroxidase 7 (GPx7) (P<0.0001) genes; we observed greater expression in birds at 21 days than at 42 days. Forty-two-day-old HS birds showed the highest H2O2 production (222.31 pmol dichlorofluorescein produced/min×mg mitochondrial protein). We also verified the effects of age and environment on the liver content of Glutathione (GSH) (P<0.0001 and P=0.0039, respectively) and catalase (CAT) enzyme activity (P=0.0007 and P=0.0004, respectively). Higher GSH content and lower CAT activity were observed in animals from the thermoneutral environment compared with the HS environment and in animals at 21 days compared with 42 days. Broilers at 42 days of age had higher plasma creatinine content (0.05 v. 0.01 mg/dl) and higher aspartate aminotransferase activity (546.50 v. 230.67 U/l) than chickens at 21 days of age. Our results suggest that under HS conditions, in which there is higher H2O2 production, 21-day-old broilers have greater antioxidant capacity than 42-day-old animals.

Possible association of nicotinic acetylcholine receptor gene (CHRNA4 and CHRNB2) polymorphisms with nicotine dependence in Japanese males: an exploratory study.

INTRODUCTION: Smoking is a leading global cause of avoidable mortality. It has been reported that the nicotinic acetylcholine receptor (CHRNA4 and CHRNB2) genes might be associated with smoking behavior in several ethnic populations. However, no study between the 2 genes and nicotine dependence (ND) using a Japanese population has been reported. METHODS: We examined the association between ND and 5 single nucleotide polymorphisms (SNPs) within the CHRNA4 and 3 SNPs within the CHRNB2 using a well characterized sample of 558 Japanese healthy male workers with a relatively homogeneous background. The Fagerström test for nicotine dependence (FTND) was used to quantify the degree of ND. Additionally, we explored the effect of gene-gene interactions of the 2 genes on ND. RESULTS: We found CHRNB2 rs4845652 genotypes to be associated with FTND scores under an additive genetic model: rs4845652 T-allele carriers had lower ND levels (p=0.038; when adjusted for smoking duration: p=0.052). Furthermore, we demonstrated a possible gene-gene interaction of CHRNA4 and CHRNB2 on ND in a dose-dependent manner: those smokers with CHRNA4 rs1044397 GG or GA genotypes along with CHRNB2 rs4845652 CC genotype are likely to demonstrate higher ND scores. DISCUSSION: These findings suggest that CHRNB2 rs4845652 T-allele carriers may be associated with lower levels of ND, and that certain allelic combinations of CHRNA4 and CHRNB2 might be correlated with higher ND levels. This preliminary study has certain limitations (issues such as sample size/power and multiple testing) that need to be taken into account, and the present work thus has an experimental nature.

Quality of life of patients and medical cost of "half elemental diet" as maintenance therapy for Crohn's disease: secondary outcomes of a randomised controlled trial.

BACKGROUND/AIM: Quality of life (QOL) of the patients and medical costs are important in current medical treatments, especially those for chronic diseases. We have reported the effectiveness of 'half elemental diet (ED)' as maintenance therapy for patients with Crohn's disease (CD). The aim of this study was to evaluate the QOL of CD patients and medical costs of half-ED. METHODS: Fifty-one CD patients in remission were randomly assigned to a half-ED group (n=26) or a free diet group (n=25). The primary outcome measure was the occurrence of relapse during a 2-year period. This time, we investigated the QOL of the patients and medical costs of half-ED, as secondary outcomes. QOL was evaluated using the Japanese version of the IBDQ scoring system, and medical costs were calculated monthly from the receipts. RESULTS: IBDQ score was not significantly different between the two groups at 1 and 13 months after the start of maintenance treatment. Medical costs were not significantly different between them either. This study showed that half-ED therapy did not affect the treatment of CD patients, neither regarding their QOL nor medical costs. CONCLUSION: This study has confirmed this half-ED therapy is beneficial for patients with Crohn's disease.

Effectiveness of an 'half elemental diet' as maintenance therapy for Crohn's disease: A randomized-controlled trial.

BACKGROUND: Although thiopurines have a proven role in maintenance therapy for Crohn's disease, an alternative therapy is needed for patients intolerant or resistant to thiopurines. AIM: To evaluate the effectiveness of home enteral nutrition as a maintenance therapy regimen in which half of the daily calorie requirement is provided by an elemental diet and the remaining half by a free diet. We refer to this home enteral nutrition therapy as 'half elemental diet'. METHODS: Between 2002 and 2005, 51 patients in remission from two hospitals were randomly assigned to a half elemental diet group (n = 26) or a free diet group (n = 25). The primary outcome measure of this study was the occurrence of relapse over the 2-year period. RESULTS: The relapse rate in the half elemental diet group was significantly lower [34.6% vs. 64.0%; multivariate hazard ratio 0.40 (95% CI: 0.16-0.98)] than that in the free diet group after a mean follow-up of 11.9 months. Compliance was similar in the two groups. No adverse event occurred in any of the patients throughout the study. CONCLUSION: This randomized-controlled trial shows the effectiveness of an half elemental diet, which is a promising maintenance therapy for Crohn's disease patients.

Significance of colonoscopy in patients with intestinal graft-versus-host disease after hematopoietic stem cell transplantation.

BACKGROUND AND STUDY AIMS: Patients with intestinal graft-versus-host disease (GVHD) are generally in poor clinical condition. In this study we aimed to establish the clinical significance of endoscopic diagnosis of this condition, observing only the distal section of the large intestine. PATIENTS AND METHODS: Endoscopic and pathological findings at colonoscopy were evaluated retrospectively in 12 patients who were diagnosed with intestinal GVHD after undergoing hematopoietic stem cell transplantation. RESULTS: The main mucosal changes observed at endoscopy were granular change, edema, "spotty redness", and sloughing. These were clearly displayed after enhancement with Indigo carmine staining, and with insertion of the colonoscope only as far as the distal section of the large intestine. A histological diagnosis of intestinal GVHD was made in 50 % of the patients, whose intestinal epithelium specimens showed numerous apoptotic bodies. It was possible to perform total colonoscopy in two patients who were in relatively good condition clinically, but there were no remarkable differences in the endoscopic findings throughout the large intestine, from the terminal ileum to the rectum. In terms of clinical outcomes of the 12 patients, their prognosis was poor in that they all either went on to suffer from chronic GVHD or died. CONCLUSIONS: Endoscopic and histological findings on distal colonoscopy are clinically significant in the diagnosis of intestinal GVHD, and limiting this examination to the distal section of the large intestine avoids causing further clinical deterioration in patients who are already in very poor general condition and the possibility of causing endoscopy-related complications.

Lipoprotein analysis using agarose gel electrophoresis and differential staining of lipids.

We established a strategy to directly measure cholesterol and triglyceride levels of each lipoprotein fraction using a combination of agarose gel electrophoresis and differential staining. The cholesterol and triglyceride levels determined by electrophoresis correlated significantly with those of ultracentrifugation. The correlation coefficients between these methods were, for cholesterol levels 0.975(very low density lipoproteins, VLDL), 0.986(low density lipoproteins, LDL) and 0.965(high density lipoproteins, HDL) and for triglyceride levels 0.994(VLDL), 0.963(LDL) and 0.959(HDL) respectively. Both intra-and inter-assays showed low values of coefficients of variation (CV) (less than 3.57%). We observed a strong linearity between staining and triglyceride concentration. An increased VLDL-cholesterol was observed in type III subjects, a result which enabled distinction between type III and type IIb or type V lipoproteinemia. The method revealed lipoprotein patterns in some samples otherwise unexpected from their corresponding serum lipid parameters. Analyses of these electrophoretic patterns thus provide an effective technique to classify types of hyperlipidemia defined by the WHO. Furthermore, quantitative measurement of chylomicrons, usually difficult, proved to be achievable, providing an additional analysis of postprandial hyperlipidemia and the exact measurement of LDL-cholesterol after diet. Consequently, we recommend this simple and easy method for clinical evaluation of abnormalities in lipoprotein profiles.

Usefulness of tc-99m MIBI SPECT in predicting multidrug resistance gene expression levels in non-small cell lung cancer--a preliminary report.

In this study we investigated whether Tc-99m hexakis 2-methoxy isobutyl isonitnile (Tc-99m MIBI) single-photon emission computed tomography (SPECT) has a correlation with the multidrug resistance (MDRI) and multidrug resistance-associated protein (MRP1) gene expression levels in non-small cell lung cancer (NSCLC). Fifteen patients with NSCLC were studied. SPECT images were obtained 15 (early) and 120 (delayed) min after injection of Tc-99m MIBI. We chose only one transverse section and set the region of interest over the tumor and out of the body. The mean counts in the tumor on early and delayed images were corrected by using those in the background and represented as Te and Td, respectively. Resected tumor specimens were frozen with liquid nitrogen and each positive control cell line was cultured. After the total ribonucleic acid (RNA) was extracted from specimens and cell lines, the complimentary deoxyribonucleic acid (cDNA) was amplified by the reverse transcription-polymerase chain reaction (RT-PCR) method. Each product was electrophoresed and fluorointensity was measured. The gene expression level was represented as the ratio of that of the positive control cell line. Te and Td indicated a significant correlation with the MDR1 gene expression level (p = 0.015 and p = 0.022), but not the gene of MRP1 (p = 0.100 and p = 0.145). In conclusion, Te and Td are useful parameters in predicting the MDRI gene expression level, but not MRPI in NSCLC.

Utility of Tc-99m GSA whole-body scintigraphy in detecting bone metastases from hepatocellular carcinoma.

Recent advances in the treatment of hepatocellular carcinoma (HCC) have prolonged patient survival. However, the number of patients with bone metastases identified during follow-up examinations has increased. Tc-99m Sn-N-pyridoxy-5-methyltryptophan (Tc-99m PMT) has been reported to accumulate at a high rate in HCC lesions and bone metastases. In the patient described here, whole-body scintigraphy showed accumulation of DTPA galactosyl human serum albumin (Tc-99m GSA) and Tc-99m PMT in bone metastases from HCC. The authors suggest that asialoglycoprotein receptors may be present in bone metastases from well-differentiated HCC. Tc-99m GSA whole-body imaging can be used to detect bone metastases from HCC and to evaluate hepatic reserve.

Stimulation of catecholamine synthesis in cultured bovine adrenal medullary cells by leptin.

Recently, we characterized leptin receptors in bovine adrenal medullary cells (Yanagihara et al. 2000). Here we report the stimulatory effect of leptin on catecholamine synthesis in the cells. Incubating cells with leptin (10 nM) for 20 min increased the synthesis of 14C-catecholamines from [14C]tyrosine, but not from L-3,4-dihydroxyphenyl [3-14C]alanine. The stimulation of catecholamine synthesis in the cells by leptin was associated with the phosphorylation and activation of tyrosine hydroxylase, the rate-limiting enzyme of catecholamine biosynthesis. The incubation of cells with leptin resulted in a rapid activation of the mitogen-activated protein kinases (MAPKs). An inhibitor of MAPK kinase, U0126, nullified the stimulatory effect of leptin on the synthesis of 14C-catecholamines. Leptin potentiated the stimulatory effect of acetylcholine on 14C-catecholamine synthesis, whereas leptin failed to enhance the phosphorylation and activation of tyrosine hydroxylase induced by acetylcholine. These findings suggest that leptin stimulates catecholamine synthesis via the activation of tyrosine hydroxylase by two different mechanisms, i.e., one is dependent on tyrosine hydroxylase phosphorylation mediated through the MAPK pathway and the second is independent of enzyme phosphorylation.

Protective effect of gamma-glutamylethylamide (theanine) on ischemic delayed neuronal death in gerbils.

We examined the protective effect of gamma-glutamylethylamide (theanine) on ischemic delayed neuronal death in field CA1 of the gerbil hippocampus. One microliter of theanine from each three concentrations (50, 125 and 500 microM) was administered through the lateral ventricle 30 min before ischemia. Transient forebrain ischemia was induced by bilateral occlusion of the common carotid arteries for 3 min under careful control of brain temperature at approximately 37 degrees C. Seven days after ischemia, the number of intact CA1 neurons in the hippocampus was assessed. Ischemia-induced neuronal death in hippocampal CA1 region was significantly prevented in a dose-dependent manner in the theanine-pretreated groups. These findings indicate that theanine might be useful clinically for preventing ischemic neuronal damage.

Neovascularization with blood-brain barrier breakdown in delayed neuronal death.

Various kinds of acute pathological events in the central nervous system, such as ischemia, hemorrhage, and trauma, often cause brain edema. The edema may advance for days or weeks while inducing extensive damage in neural function, regardless of the extent of the original damage, and often results in death. Delayed edema is thought to be vasogenic; however, the mechanism underlying edema induction remains unknown. We found delayed vascular cell proliferation with a blood-brain barrier breakdown in and around the gerbil CA1 hippocampus, a region known to be involved in delayed apoptotic neuronal death 2-6 days after transient ischemia. Vascular cell proliferation, assessed by (3)H-thymidine incorporation, was most prominent 4-6 days after ischemia, and extravasation of exogenously applied dye or endogenous serum albumin from blood vessels was observed concomitantly. We propose neovascularization in delayed neuronal death as a cause of brain edema advancing days after neurological events.

Intraductal ultrasonographic assessment of coagulation depth during endoscopic microwave coagulation therapy in a canine model.

In order to examine the feasibility of endoscopic microwave coagulation therapy (MCT) for the treatment of common bile duct cancer in humans, we investigated the safety and effectiveness of endoscopic MCT with the assistance of intraductal ultrasonography (IDUS). We performed MCT in the canine small intestine as a model because of its similarity in size to the obstructed human common bile duct. Multiple coagulation lesions were made in anesthetized dogs, with the endoscope inserted through a jejunostomy. Under condition "A", the tip of the MCT probe was kept in a fixed position in contact with the mucosa of the intestine, and coagulation was performed with an output of 30 or 50 W for 30 or 10 s, respectively. Under condition "B", the tip of the MCT probe was moved along the intestinal mucosa, and coagulation was performed with an output of 50, 40, or 30 W, all for 10 s. After coagulation, a thin-caliber ultrasonic probe was inserted endoscopically for observation of the effects of coagulation. Each coagulation under condition "A" resulted in degeneration of all layers of the intestine. Coagulation under condition "B" resulted in more localized degeneration, extending from the mucosal layer to the proper muscle layer. By comparison with histologic observations, the accuracy of IDUS in assessing depth of degeneration was 87.5%. Endoscopic microwave coagulation was considered safe and effective, and IDUS was able to accurately assess the effects of microwave coagulation.

A case of local recurrent pheochromocytoma: usefulness of I-123 MIBG early SPECT and maximum intensity projection images.

It has been reported that 10% of patients with pheochromocytoma experience recurrence despite surgery. The authors administered I-123 MIBG to patients with recurrent pheochromocytoma and found that early images are more useful than delayed images to detect recurrence. Early imaging with I-123 MIBG is useful for postoperative follow-up studies in patients with pheochromocytoma and those with possible tumor recurrence.

Characterization and functional role of leptin receptor in bovine adrenal medullary cells.

We report here the characterization and functional roles of the leptin receptor (ObR) in bovine adrenal medullary cells. The plasma membranes isolated from bovine adrenal medulla showed a single class of specific binding sites of (125)I-leptin with an apparent K(d) of 6.6 nM and B(max) of 62 fmol/mg protein. ObRa but not ObRb mRNA was detected in bovine adrenal medulla by reverse transcriptase-polymerase chain reaction. Incubation of cultured adrenal medullary cells with leptin (3-30 nM) for 20 min resulted in a significant increase in [(14)C]catecholamine synthesis from [(14)C]tyrosine without any change in catecholamine secretion. These findings suggest that leptin stimulates catecholamine synthesis through its receptors in bovine adrenal medullary cells.

Comparison of the accumulation and efflux kinetics of technetium-99m sestamibi and technetium-99m tetrofosmin in an MRP-expressing tumour cell line.

The potential clinical use of technetium-99m labeled sestamibi (Tc-MIBI) and tetrofosmin (Tc-Tfos) to image tumours is currently being evaluated. In this study. the accumulation and efflux of Tc-MIBI and Tc-Tfos in the nasopharyngeal carcinoma cell line CNE-1 were examined in the presence or absence of various inhibitors of P-glycoprotein (PGP) and/or multidrug resistance associated protein (MRP) activity [GG918, PSC833, verapamil (Vrp), cyclosporin A (CsA) and buthionine sulfoximine (BSO)]. Reverse-transcriptase polymerase chain reaction analysis and immunodetection of the CNE-1 cells detected expression of MRP, MRPI and MRP2 but not PGP. Tc-MIBI and Tc-Tfos accumulation was increased (P < 0.0001) and efflux decreased (P < 0.05) in the presence of BSO, CsA, Vrp and PSC833 but not GG918, which is a specific inhibitor of PGP. The absolute accumulation of Tc-MIBI was approximately twofold higher than that seen with Tc-Tfos, whereas the addition of inhibitors caused a much greater suppression of Tc-Tfos transport (>2 times greater than for Tc-MIBI). However, no qualitative differences in inhibitors were seen between Tc-MIBI and Tc-Tfos. These results suggest that both Tc-MIBI and Tc-Tfos are substrates for the MRP transporter and that PSC833, Vrp, CsA and BSO but not GG918 can inhibit MRP activity. These results indicate that Tc-MIBI and Tc-Tfos may be suitable imaging agents for detecting MRP-mediated drug resistance in human cancers.

Technetium-99m MIBI single photon emission computed tomography as an indicator of prognosis for patients with lung cancer-preliminaly report.

PURPOSE: We performed technetium-99m hexakis-2-methoxyisobutylisonitrile (Tc-99m MIBI) single photon emission computed tomography (SPECT) in 23 patients with primary lung cancer between July 1993 and March 1996. We evaluated the relationships among the uptake ratio, retention index and the prognosis after radiation therapy and/or chemotherapy. MATERIALS AND METHODS: Tc-99m MIBI SPECT was performed at 30 minutes and at 3 hours after intravenous injection of 600 MBq of Tc-99m MIBI with three gamma camera detectors (GCA-9300A/HG) on transverse SPECT images. Regions of interest were set in the area of abnormal uptake of Tc-99m MIBI and in the contralateral normal lung. The ratio of uptake in the lesion to that in the contralateral normal lung was obtained on early images (early ratio; ER) as well as delayed images (delayed ratio: DR). The retention index (RI) was calculated as follows: RI = (DR - ER)/ER x 100. The ratio was compared with survival time and prognostic factors. RESULTS: There was no correlation between ER and DR. The patients with high RI survived longer than those with low RI (median survival, 19.4 months vs. 9.4 months; p = 0.0104 by the Mantel-Cox test). CONCLUSION: These results suggest that RI is the most useful among Tc-99m MIBI indices of primary lung cancer in predicting prognosis.

Use of iodine-123 metaiodobenzylguanidine scintigraphy to assess cardiac sympathetic denervation and the impact of hypertension in patients with non-insulin-dependent diabetes mellitus.

The objectives of this clinical study using iodine-123 metaiodobenzylguanidine (MIBG) scintigraphy were (a) to evaluate cardiac sympathetic denervation in non-insulin-dependent diabetes mellitus (NIDDM) patients with and without hypertension and (b) to investigate the relation between cardiac sympathetic denervation and prognosis in NIDDM patients. We compared clinical characteristics and MIBG data [heart to mediastinum (H/M) ratio and % washout rate (WR)] in a control group and NIDDM patients with and without hypertension. MIBG scintigraphy was performed in 11 controls and 82 NIDDM patients without overt cardiovascular disease except for hypertension (systolic blood pressure >/=140 and/or diastolic blood pressure >/=90 mmHg). After MIBG examination, blood pressure was measured regularly in all NIDDM patients. There were significant differences between 65 normotensive and 17 hypertensive NIDDM patients with respect to age (55+/-11 vs 63+/-12 years, respectively, P<0.05), prevalence of diabetic retinopathy (12% vs 35%, respectively, P<0.05) and systolic blood pressure (120+/-12 vs 145+/-16 mmHg, respectively, P<0.001). The H/M ratio in hypertensive NIDDM patients was significantly lower than in the control group (1. 81+/-0.29 vs 2.27+/-0.20, respectively, P<0.01). During the follow-up period (18+/- 12 months), 17 NIDDM patients newly developed hypertension after MIBG examination. There were no significant differences in their clinical characteristics compared with persistently normotensive or hypertensive NIDDM patients. %WR in patients with new onset hypertension was significantly higher than in the control group (30.88%+/-16.87% vs 12.89%+/-11.94%, respectively, P<0.05). Moreover, in these patients %WR correlated with duration from the date of MIBG scintigraphy to the onset of hypertension (r=-0.512, P<0.05). Five NIDDM patients died during the follow-up period (four newly hypertensive patients and one normotensive patient). There were significant statistical differences between the control group and non-survivors in terms of age (54+/-11 vs 73+/-11 years, respectively, P<0.01), H/M ratio (2. 27+/- 0.20 vs 1.64+/-0.36, respectively, P<0.01) and %WR (12. 89%+/-11.94% vs 42.52%+/-22.39%, respectively, P<0.01). In conclusion, cardiac sympathetic denervation using MIBG scintigraphy observed in hypertensive NIDDM patients, and was more profound in non-survivors. MIBG scintigraphy proved useful for the evaluation of NIDDM patients with new onset hypertension, and it was found that NIDDM patients with abnormalities on MIBG scintigraphy needed to be observe carefully.