Plumbagin suppresses tumor cell growth in oral squamous cell carcinoma cell lines.

OBJECTIVES: Plumbagin (PL), a naturally occurring quinoid, exerts antitumoral effects in diverse types of cancer cells. However, the effect of PL on tumor cell proliferation in oral squamous cell carcinoma (OSCC) remains poorly understood. In this study, we assessed the efficacy of PL, in human OSCC cells. METHODS: The effect of PL on the cell growth and apoptosis of OSCC cell lines was evaluated using MTT and Annexin V assays, respectively. The effect of PL on mitochondrial membrane potential loss and reactive oxygen species (ROS) generation was evaluated using flow cytometry analysis. RESULTS: MTT assay showed that PL dose-dependently suppressed OSCC cell growth, with IC50 values ranging from 3.87 to 14.6 µM. Flow cytometry analysis revealed that PL treatment resulted in a significant decrease in mitochondrial membrane potential and an increase in the number of apoptotic cells. Notably, ROS generation was significantly elevated after PL treatment. Furthermore, a ROS scavenger, N-acetylcysteine (NAC), clearly suppressed the decrease in mitochondrial membrane potential, increase of caspase-3/7 activity, and apoptosis after PL treatment. CONCLUSION: This study provides the considerable evidence of the tumor-suppressive effect of PL, thereby highlighting its therapeutic potential for OSCC treatment.

Oxidative stress and micronutrient therapy in malaria: an in vivo study in Plasmodium berghei infected mice.

Free radical production from oxidative stress induced by malaria infection plays a major role in the pathogenesis of malaria. However, the use of agents with antioxidant activity may interfere with malaria progression. The study involves an in vivo evaluation of the role of some antioxidant micronutrients in the modulation of malaria infection. Rodent malaria model using Plasmodium berghei NK-65 strain (chloroquine sensitive) was used for the study. Forty five mice of either sex weighing 20.05 +/- 0.02 g were procured for the study. Forty mice were inoculated intraperitoneally with 1 x 10(7) million Plasmodium berghei infected erythrocyte and were administered with 0.2 mL of distilled water, 0.2 mL of vehicle; Tween 80 (control and vehicle group), chloroquine 25 mg kg(-1) and artesunate 4 mg kg(-1) (standard drug group), vitamin A 60 mg kg(-1), vitamin E 100 mg kg(-1), selenium 1 mg kg(-1), zinc 100 mg kg(-1) (test group F, G, H and I, respectively) 72 hours post inoculation. Antioxidant micronutrients demonstrated significant (p < 0.05) schizonticidal activity when compared with negative control during the 4 day curative test. Erythrocyte membrane disability was most markedly elevated in the tween 80 group (426.15%), followed closely by the chloroquine (373.85%) treated group and artesunate group (329.23%) and least in the zinc treated group (32.31%). There was no significant (p > 0.05) difference in MCFI values (0.115 +/- 0.002; 0.114 +/- 0.002 g dL(-1)) between vitamin A treated group and selenium treated group respectively. However, this was significant (p < 0.05) between the micronutrient treated groups and the control (negative, positive and vehicle). Conclusively, antioxidant micronutrients have antimalarial activity which may be due potentiation of erythrocyte membrane stabilization.

Randomised clinical trial: evaluation of the efficacy of mesalazine (mesalamine) suppositories in patients with ulcerative colitis and active rectal inflammation -- a placebo-controlled study.

BACKGROUND: Mesalazine suppositories are recommended and widely used as the standard therapy in induction and maintenance of remission for proctitis. AIM: To evaluate the efficacy of mesalazine suppositories in patients with ulcerative colitis (UC) and rectal inflammation; and in patient groups categorised by the extent of lesions. METHODS: This study was a phase III multicentre, randomised, double-blind, placebo-controlled, parallel-group study. Mild-to-moderate UC patients with rectal inflammation were randomly assigned either a 1 g mesalazine or placebo suppository. The suppository was administered in the rectum once daily for 4 weeks. The primary efficacy end point was the rate of endoscopic remission (mucosal score of 0 or 1) after 4 weeks. RESULTS: The endoscopic remission rates after 4 weeks in the mesalazine and placebo suppository groups were 81.5% and 29.7%, respectively, and the superiority of mesalazine to placebo was confirmed (P < 0.0001, chi-squared test). For proctitis, the endoscopic remission rates after 4 weeks were 83.8% and 36.1% in the mesalazine and placebo suppository groups, respectively, and the corresponding rates for all other types of UC were 78.6% and 21.4%, respectively. The superiority of mesalazine to placebo was confirmed in both subgroups (P < 0.0001, Fisher's exact test). The percentage of patients without bleeding was significantly higher in the mesalazine group than the placebo group from Day 3 of treatment (P = 0.0001, Fisher's exact test). CONCLUSIONS: The effectiveness of mesalazine suppositories in all types of UC patients with rectal inflammation was confirmed for the first time in a double-blind, placebo-controlled, parallel-group study (JapicCTI- 111421).

Interface pressure is affected by slippage of bandages at thigh.

AIM: Very little is known about how compression bandages lose interface pressure. We hypothesized that the loss of interface pressure is correlated with the slippage of the bandages, and studied the interface pressure and slippage of three bandages over 8 hours. METHODS: Twenty-Seven legs from 27 healthy volunteers were bandaged with short stretch bandages (SS), cohesive short stretch bandages (CS), and long stretch bandages (LS). Pressure sensors were placed above the ankle (B1), below the knee (D), and mid thigh (F). Interface pressures in a sitting position were recorded at the beginning, and 4 and 8 hours later. In 17 legs, the pressure sensor sites were marked, and their heights were measured on standing upright. RESULTS: SS and CS lost interface pressure quickly, but LS maintained pressure better than SS and CS at all sites. There was no pressure difference between SS and CS at the lower leg. However, CS maintained pressure better than SS at the mid thigh (44.6% vs. 54.4% pressure loss at 8 hours, respectively. P=0.037). There was a tendency toward less slippage with CS than SS at the mid thigh. In CS and LS, there was a linear correlation between the slippage of bandages and the interface pressure at the mid thigh (P <.01, in both). CONCLUSION: The interface pressure may be affected by the slippage of bandages at the thigh, but not at the lower leg. Cohesive short stretch bandages may exert their beneficial impact at the thigh.

Usefulness of magnifying endoscopy with narrow-band imaging for determining the horizontal extent of early gastric cancer when there is an unclear margin by chromoendoscopy (with video).

BACKGROUND: Magnifying endoscopy (ME) with narrow-band imaging (NBI) may allow reliable delineation of the horizontal extent of early gastric cancers before endoscopic submucosal dissection (ESD). However, the advantages of ME with NBI over standard endoscopy with dye spraying (chromoendoscopy [CE]) have yet to be elucidated. OBJECTIVE: To investigate the usefulness and limitations of ME with NBI when CE is unsuccessful for determining the horizontal extent of early gastric cancer. DESIGN: Case series. SETTING: Single tertiary referral center. MATERIALS: Series of 350 consecutive early gastric cancers resected en bloc using ESD. INTERVENTION: ME with NBI for cancers with unclear margins by CE. MAIN OUTCOME MEASUREMENTS: The rate of successful delineation by ME with NBI for cancers that had demonstrated unclear margins using CE. RESULTS: The proportion of cancers showing unclear margins using CE was 18.9% (66/350). Of these, 62 of 66 cancers were examined using ME with NBI, with the entire margins successfully delineated in 72.6% (45/62) of the lesions that had shown unclear margins using CE. The success rate was 0% for undifferentiated cancers, significantly lower than that for differentiated lesions (P < .00001). LIMITATIONS: Even by using ME with NBI, endoscopic delineation remains difficult for undifferentiated lesions. CONCLUSIONS: ME with NBI is an excellent modality for identifying the entire margin of early gastric cancers, when the margins are unclear using CE.

Cross-city comparison of indoor air temperatures in air-conditioned spaces.

Field measurements were collected through physical measurements and observations in the cities of Seoul, Korea; Eugene and Portland, Oregon; and Yokohama, Japan, during the fall, winter, and summer seasons from 2005 to 2006. A total of 1733 data sets were collected (Seoul - 713; Oregon - 807; Yokohama - 213) in variety of multi-use buildings with the goal of examining operative temperatures and conditions encountered during everyday life. Of the four cities measured, winter and autumn indoor operative temperatures were highest in Seoul and lowest in Yokohama when normalized to outdoor conditions. In contrast, summer indoor operative temperatures were highest in Yokohama and lowest in Oregon. Clothing levels changed seasonally, and differences were observed between 'long-term occupants', 'residents', and 'transients.'

Feasibility and efficiency of indirect lifestyle interventions in offspring of type 2 diabetic patients.

INTRODUCTION: Individuals genetically predisposed to type 2 diabetes represent an important target for preventive strategies. Genetic screening, based on information about individual genetic variants, will be possible technically, but translational research in this field is still insufficient. Family history thus represents a useful tool for detecting genetically high-risk populations in this post-genomic era. OBJECTIVES: The purpose of this pilot study was to investigate the feasibility and efficiency of indirect lifestyle interventions in offspring of type 2 diabetic patients. METHODS: Offspring were recruited from 74 diabetic (Group 1) and 39 non-diabetic (Group 2, control group) patients. A lifestyle intervention was conducted by mail, a total of 3 times, every 3 months. Lifestyle related to diet and physical activity was assessed using a self-administered questionnaire. RESULTS: Ten offspring of type 2 diabetic and 6 of non-diabetic patients participated in this study. Total energy intake decreased after 3 interventions in both of the groups (Group 1: 305 ± 228.8 kcal/day, p = 0.004; Group 2: 82 ± 65.6 kcal/day, p = 0.04); however, the effect of intervention was significantly greater in Group 1 compared to Group 2 (p = 0.021). Physical activity and other physical outcomes were stable in normal levels during the study period in both of the groups. CONCLUSIONS: The intervention program helped to reduce total energy intake in offspring of type 2 diabetic patients more than in the control group, but the acceptance rate of the intervention program was disappointingly low. Further consideration is required to access and motivate offspring to develop precautionary lifestyle principles.

Microsomal prostaglandin E synthase-1 contributes to ischaemic excitotoxicity through prostaglandin E2 EP3 receptors.

BACKGROUND AND PURPOSE: Although microsomal prostaglandin E synthase (mPGES)-1 is known to contribute to stroke injury, the underlying mechanisms remain poorly understood. This study examines the hypothesis that EP(3) receptors contribute to stroke injury as downstream effectors of mPGES-1 neurotoxicity through Rho kinase activation. EXPERIMENTAL APPROACH: We used a glutamate-induced excitotoxicity model in cultured rat and mouse hippocampal slices and a mouse middle cerebral artery occlusion-reperfusion model. Effects of an EP(3) receptor antagonist on neuronal damage in mPGES-1 knockout (KO) mice was compared with that in wild-type (WT) mice. KEY RESULTS: In cultures of rat hippocampal slices, the mRNAs of EP(1-4) receptors were constitutively expressed and only the EP(3) receptor antagonist ONO-AE3-240 attenuated and only the EP(3) receptor agonist ONO-AE-248 augmented glutamate-induced excitotoxicity in CA1 neurons. Hippocampal slices from mPGES-1 KO mice showed less excitotoxicity than those from WT mice and the EP(3) receptor antagonist did not attenuate the excitotoxicity. In transient focal ischaemia models, injection (i.p.) of an EP(3) antagonist reduced infarction, oedema and neurological dysfunction in WT mice, but not in mPGES-1 KO mice, which showed less injury than WT mice. EP(3) receptor agonist-induced augmentation of excitotoxicity in vitro was ameliorated by the Rho kinase inhibitor Y-27632 and Pertussis toxin. The Rho kinase inhibitor HA-1077 also ameliorated stroke injury in vivo. CONCLUSION AND IMPLICATIONS: Activity of mPGES-1 exacerbated stroke injury through EP(3) receptors and activation of Rho kinase and/or G(i). Thus, mPGES-1 and EP(3) receptors may be valuable therapeutic targets for treatment of human stroke.

Microsomal prostaglandin E synthase-1 and cyclooxygenase-2 are both required for ischaemic excitotoxicity.

BACKGROUND AND PURPOSE: Although both microsomal prostaglandin E synthase (mPGES)-1 and cyclooxygenase (COX)-2 are critical factors in stroke injury, but the interactions between these enzymes in the ischaemic brain is still obscure. This study examines the hypothesis that mPGES-1 activity is required for COX-2 to cause neuronal damage in ischaemic injury. EXPERIMENTAL APPROACH: We used a glutamate-induced excitotoxicity model in cultures of rat or mouse hippocampal slices and a mouse middle cerebral artery occlusion-reperfusion model in vivo. The effect of a COX-2 inhibitor on neuronal damage in mPGES-1 knockout (KO) mice was compared with that in wild-type (WT) mice. KEY RESULTS: In rat hippocampal slices, glutamate-induced excitotoxicity, as well as prostaglandin (PG) E(2) production and PGES activation, was significantly attenuated by either MK-886 or NS-398, inhibitors of mPGES-1 and COX-2 respectively; however, co-application of these inhibitors had neither an additive nor a synergistic effect. The protective effect of NS-398 on the excitotoxicity observed in WT slices was completely abolished in mPGES-1 KO slices, which showed less excitotoxicity than WT slices. In the transient focal ischaemia model, mPGES-1 and COX-2 were co-localized in the infarct region of the cortex. Injection of NS-398 reduced not only ischaemic PGE(2) production, but also ischaemic injuries in WT mice, but not in mPGES-1 KO mice, which showed less dysfunction than WT mice. CONCLUSION AND IMPLICATIONS: Microsomal prostaglandin E synthase-1 and COX-2 are co-induced by excess glutamate in ischaemic brain. These enzymes are co-localized and act together to exacerbate stroke injury, by excessive PGE(2) production.

Role of N-terminus of tyrosine hydroxylase in the biosynthesis of catecholamines.

Tyrosine hydroxylase (TH) catalyzes the conversion of L: -tyrosine to L: -dopa, which is the initial and rate-limiting step in the biosynthesis of catecholamines [CA; dopamine (DA), noradrenaline, and adrenaline], and plays a central role in the neurotransmission and hormonal actions of CA. Thus, TH is related to various neuro-psychiatric diseases such as TH deficiency, Parkinson's disease (PD), and schizophrenia. Four isoforms of human TH (hTH1-hTH4) are produced from a single gene by alternative mRNA splicing in the N-terminal region, whereas two isoforms exist in monkeys and only a single protein exist in all non-primate mammals. A catalytic domain is located within the C-terminal two-thirds of molecule, whereas the part of the enzyme controlling enzyme activity is assigned to the N-terminal end as the regulatory domain. The catalytic activity of TH is end product inhibited by CA, and the phosphorylation of Ser residues (Ser(19), Ser(31), and especially Ser(40) of hTH1) in the N-terminus relieves the CA-mediated inhibition. Ota and Nakashima et al. have investigated the role of the N-terminus of TH enzyme in the regulation of both the catalytic activity and the intracellular stability by producing various mutants of the N-terminus of hTH1. The expression of the following three enzymes, TH, GTP cyclohydrolase I, which synthesizes the tetrahydrobiopterin cofactor of TH, and aromatic-L: -amino acid decarboxylase, which produces DA from L: -dopa, were induced in the monkey striatum using harmless adeno-associated virus vectors, resulting in a remarkable improvement in the symptoms affecting PD model monkeys Muramatsu (Hum Gene Ther 13:345-354, 2002). Increased knowledge concerning the amino acid sequences of the N-terminus of TH that control enzyme activity and stability will extend the spectrum of the gene-therapy approach for PD.

Effect of peripherally administered lipopolysaccharide (LPS) on GTP cyclohydrolase I, tetrahydrobiopterin and norepinephrine in the locus coeruleus in mice.

Lipopolysaccharide (LPS), an endotoxin released from the outer membranes of Gram-negative bacteria, triggers cells to synthesize and release inflammatory cytokines that may progress to septic shock in vivo. We found that LPS enhances tetrahydrobiopterin (BH4) biosynthesis by inducing the biosynthetic enzyme GTP cyclohydrolase I (GCH) in vitro in the mouse neuroblastoma cell line N1E-115. Furthermore, we observed that gene expression of GCH in the locus coeruleus (LC) in mice was enhanced by peripheral administration of LPS, resulting in increased concentrations of BH4, and norepinephrine, and its metabolite 4-hydroxy-3-methoxyphenylglycol (MHPG). These results suggest that tyrosine hydroxylase (TH) activity is increased by increased content of BH4 due to enhanced mRNA expression of GCH in the LC resulting in the increase in norepinephrine in the LC during endotoxemia. LPS in blood may act as a stressor to increase norepinephrine biosynthesis in the mouse LC.

Effect of peripheral lipopolysaccharide injection on dopamine content in murine anterior olfactory nucleus.

Norepinephrine turnover rate in the murine locus coeruleus (LC) is known to be enhanced by the intraperitoneal (i.p.) injection of lipopolysaccharide (LPS). Approximately 40% of LC neurons are also known to project to the olfactory bulb (OB) and the anterior olfactory nucleus (AON). Therefore, we investigated whether an i.p. injection of 500 microg LPS could modulate the catecholamine biosynthesis in these sites in 8-week-old C3H/HeN male mice. Unexpectedly, the content of norepinephrine was not elevated in both sites during 6-h-observation after LPS injection. The contents of dopamine and its metabolites in the AON were highly increased at 4 h after LPS injection, whereas those in the OB were not elevated during 6-h-observation. Although the AON has been considered not to belong to the dopaminergic neuron system, our report is the first to show an elevated dopamine content in the AON under a stressful condition such as endotoxemia.

Peripheral administration of lipopolysaccharide enhances the expression of guanosine triphosphate cyclohydrolase I mRNA in murine locus coeruleus.

GTP cyclohydrolase I is the first and rate-limiting enzyme for the de novo biosynthesis of tetrahydrobiopterin, which is the cofactor for tyrosine hydroxylase. Lipopolysaccharide can modulate tetrahydrobiopterin production by upregulating GTP cyclohydrolase I protein expression in the locus coeruleus in the mouse brain. The increased supply of tetrahydrobiopterin in the locus coeruleus leads to increased tyrosine hydroxylase activity without affecting the level of tyrosine hydroxylase protein expression, resulting in an increase in norepinephrine turnover at the site. This study was performed to address whether the increase in GTP cyclohydrolase I protein is dependent on the de novo synthesis of GCH in the locus coeruleus. After i.p. administration of lipopolysaccharide, the mRNA expression of GTP cyclohydrolase I was examined. The expression level increased within 2 h, and reached to maximum level at 4 h after the lipopolysaccharide administration. However, the mRNA expression level of 6-pyruvoyl-tetrahydropterin synthase and sepiapterin reductase, both of which are involved successively after GTP cyclohydrolase I in tetrahydrobiopterin biosynthesis, were not affected by the lipopolysaccharide administration. These results suggest that GTP cyclohydrolase I upregulation alone is enough to modulate tetrahydrobiopterin production in the locus coeruleus. In addition, the mRNA level of tyrosine hydroxylase was also not affected by the lipopolysaccharide administration. Taken together, the data indicate that GTP cyclohydrolase I plays a crucial role in regulating norepinephrine biosynthesis by a pathway the activity of which is triggered by lipopolysaccharide i.p. administration.

Determination of tetrahydrobiopterin in murine locus coeruleus by HPLC with fluorescence detection.

Tetrahydrobiopterin in the murine locus coeruleus was measured as its fully oxidized form, biopterin, using a HPLC coupled to a fluorescence detector, because tetrahydrobiopterin itself cannot be detected by such means. The differential oxidization method distinguished tetrahydrobiopterin-derived biopterin and dihydrobiopterin-derived biopterin. The protocol reported here is a rapid and sensitive method that facilitates the measurement of tissue and/or cellular tetrahydrobiopterin. Using this assay protocol, we were able to detect and quantify variations in the tetrahydrobiopterin content in the murine locus coeruleus.

A new reconstructive procedure after segmental pancreatectomy: an experimental study of pancreatic end-to-end (duct-to-duct) anastomosis.

We produced experimental models of pancreatic end-to-end anastomosis, including ductal end-to-end anastomosis (with or without stent) and pancreaticojejunostomy, using mongrel dogs, with a view to evaluating reconstructive procedures after segmental pancreatectomy. We examined macroscopic findings, pancreatograms, and microangiographic and histopathological findings to determine whether pancreatic end-to-end anastomosis was as practicable as pancreaticojejunostomy. Macroscopic findings showed no suture failure in any animal in the end-to-end anastomosis group. Pancreatography revealed obstruction of the stent tube in the stent subgroup, but good patency in the no-stent subgroup. On the imaging of the microvasculature in the end-to-end anastomosis group, proliferation of neovascular vessels and formation of communicating vessels were detected. Histopathologically, no suture failure was detected, and the viability of the pancreatic end-to-end anastomosis was confirmed. From this experiment, we concluded it that it was possible to employ pancreatic end-to-end anastomosis after segmental pancreatectomy in the clinical situation.