Anti-inflammatory effects of simvastatin in nonsteroidal anti-inflammatory drugs-induced small bowel injury.

Small bowel injury can occur as the result of a multifaceted process that includes increased acid secretion, generation of reactive oxygen species, and cyclooxygenase inhibition. However, no effective medication for small bowel ulceration is available. Simvastatin is an important lipid-lowering agent with anti-inflammatory activity. We aimed to validate the effects of simvastatin in vitro and in vivo. In presence or absence of simvastatin, IEC-6 small bowel cell line with 50 ng/ml of tumor nectosis factor α (TNF-α) was investigated by western blotting, qRT-PCR, and DCF-DA assay. In addition, an in vivo study of nonsteroidal anti-inflammatory drugs (NSAID)-induced small bowel inflammation was performed using 7-week-old specific-pathogen-free (SPF) male C57BL/6 mice. Simvastatin treatment reduced the mRNA levels of interleukin-6 and interleukin-8 by approximately 50% in TNF-α-stimulated IEC-6 cells. Treatment with a combination of 50 ng/ml TNF-α and µM simvastatin decreased activation of Akt, IκBα, and nuclear factor-κB p65 level in IEC-6 cells. By DCF-DA staining, intracellular reactive oxygen species (ROS) production was increased in TNF-α-stimulated cells, and treatment with simvastatin decreased the level of ROS. In addition, in vivo mouse model of NSAID-induced small bowel inflammation, the administration of simvastatin reduced the number of small bowel hemorrhagic lesions and the level of ROS production as determined by gross examination and 8-hydroxydeoxyguanosine immunohistochemistry of small bowel tissue, respectively. Simvastatin reduced NSAID-induced injuries by both suppression of ROS generation and modulation of inflammatory cytokines in vitro and in vivo. Therefore, simvastatin, an HMG-CoA reductase inhibitor, has potential as a prophylactic and therapeutic agent for NSAID-induced small bowel injury.

A dual role for the RhoGEF Ephexin5 in regulation of dendritic spine outgrowth.

The outgrowth of new dendritic spines is closely linked to the formation of new synapses, and is thought to be a vital component of the experience-dependent circuit plasticity that supports learning. Here, we examined the role of the RhoGEF Ephexin5 in driving activity-dependent spine outgrowth. We found that reducing Ephexin5 levels increased spine outgrowth, and increasing Ephexin5 levels decreased spine outgrowth in a GEF-dependent manner, suggesting that Ephexin5 acts as an inhibitor of spine outgrowth. Notably, we found that increased neural activity led to a proteasome-dependent reduction in the levels of Ephexin5 in neuronal dendrites, which could facilitate the enhanced spine outgrowth observed following increased neural activity. Surprisingly, we also found that Ephexin5-GFP levels were elevated on the dendrite at sites of future new spines, prior to new spine outgrowth. Moreover, lowering neuronal Ephexin5 levels inhibited new spine outgrowth in response to both global increases in neural activity and local glutamatergic stimulation of the dendrite, suggesting that Ephexin5 is necessary for activity-dependent spine outgrowth. Our data support a model in which Ephexin5 serves a dual role in spinogenesis, acting both as a brake on overall spine outgrowth and as a necessary component in the site-specific formation of new spines.

Three-dimensional characterization of in vivo intervertebral disc degeneration using EPIC-µCT.

OBJECTIVE: Small animal models are commonly employed to study progression of and potential treatment techniques for degenerative disc disease (DDD), but assessment using conventional imaging techniques is challenging due to resolution. The objective of this study was to employ equilibrium partitioning of an ionic contrast agent micro computed tomography (EPIC - µCT) to map three-dimensional (3D) degenerative changes in the rabbit intervertebral disc (IVD). MATERIALS AND METHODS: In vivo degeneration was induced surgically in 12 New Zealand White rabbits via percutaneous annular puncture and percutaneous nucleotomy. IVDs were harvested after 3 and 6 weeks. EPIC-µCT imaging was performed on fresh, IVDs before and after formalin fixation, and 3D IVD volumes were segmented. IVDs were histologically stained with Safranin-O/Fast-Green and Hematoxylin & Eosin (H&E). EPIC-µCT attenuation and 3D morphological measurements were assessed in healthy and degenerate IVDs and compared to qualitative grading and disc height measurement from histology. RESULTS: EPIC-µCT caused pronounced contrast enhancement of the IVD. Annular puncture and nucleotomy produced mild and severe degenerative changes, respectively. IVD attenuation following contrast enhancement increased significantly in nucleotomized discs at 3 and 6 weeks. IVD attenuation correlated significantly with histologic score and disc height measurements. Disc height decreased most extensively in the posterior and lateral aspects of the IVD. 3D morphological measurements correlated strongly to IVD attenuation and were more sensitive to degenerative changes than histologic measurements. Formalin fixation reduced the attenuation of IVDs by ∼10%. CONCLUSION: EPIC-µCT is sensitive to in vivo DDD induced by nucleotomy and provides a high resolution 3D method for mapping degenerative changes in rabbit IVDs.

Prognostic significance of glycaemic control in patients with HBV and HCV-related cirrhosis and diabetes mellitus.

AIMS: Diabetes mellitus (DM) is frequently observed in patients with cirrhosis, particularly that due to hepatitis C virus (HCV) infection. However, no studies have focused on the clinical significance of glycaemic control in cirrhotic patients because of their short life expectancy and poor hepatic function. The aim of this study was to evaluate the prognostic impact of glycaemic control in patients with hepatitis B virus (HBV) and HCV-related cirrhosis and DM. METHODS: A total of 434 patients with HCV-related (HCV group, n = 88) or HBV-related (HBV group, n = 346) cirrhosis were studied retrospectively. We determined the prevalence of DM and treatment methods for hyperglycaemia and status of glycaemic control, and the patients' outcome. RESULTS: The prevalence of DM was 43.2% (38/88) in the HCV group and 19.7% (68/346) in the HBV group. Patients in the HCV group were older with a female preponderance. DM was detected before the diagnosis of cirrhosis or simultaneously in 92% and 79% in the HCV and HBV groups, respectively. Most patients were treated with insulin or oral hypoglycaemic agents. However, blood glucose levels were maintained within the normal range in 34.2% of the HCV group and in 23.5% of the HBV group. Forty-six patients died during the observation period in both groups. Hepatic failure was the most common cause of death, and sepsis and variceal bleeding were more frequent in the HCV group than in the HBV group. Multivariate analysis showed that Child-Pugh class was the most important factor for survival in both groups. In the HCV group, the status of glycaemic control was a significant independent factor of survival (P = 0.018). In the HBV group, age and the development of spontaneous bacterial peritonitis were significant. CONCLUSION: DM is more frequent in patients with HCV-related cirrhosis than in patients with HBV. Strict control of blood glucose levels could improve survival in HCV patients. A precise assessment of the risks and benefits of glycaemic control is required to reduce the mortality and morbidity of patients with cirrhosis and DM.

A phase II trial of oral temozolomide in patients with metastatic renal cell cancer.

PURPOSE: To determine the activity of temozolomide, an oral imidazotetrazine alkylating agent that has exhibited broad antitumor activity in preclinical studies, in renal cell cancer (RCC) patients. METHODS. Metastatic RCC patients were treated with temozolomide, 200 mg/m(2) per day orally, and traditional radiologic response endpoints were assessed. O(6)-Alkylguanine-DNA alkyltransferase (AGT) activity was measured in four pretreatment biopsies. RESULTS: Among 12 patients, there were no responses. High AGT activity was observed in all four biopsies analyzed. CONCLUSIONS: Temozolomide is not active against RCC and this clinical observation may be due to high levels of AGT in this tumor.

Successful treatment of Boerhaave's syndrome with endoscopic insertion of a self-expandable metallic stent: report of three cases and a review of the literature.

Boerhaave's syndrome is a rare but life-threatening condition which presents a diagnostic as well as therapeutic challenge. Early diagnosis and prompt surgical repair are critical for survival; however late recognition of esophageal rupture is not unusual. A variety of nonsurgical approaches have been proposed, particularly in the case of delayed diagnosis. In recent years, the insertion of a metallic stent has been described as a promising modality in the treatment of spontaneous esophageal rupture. We report three cases of Boerhaave's syndrome treated with self-expandable metallic stents and review previously published cases.

Effect of dietary restriction on age-related increase of liver susceptibility to peroxidation in rats.

Dietary restriction (DR) increases life span and decreases age-related diseases in experimental animals. It has received a great deal of attention in connection with the relationship between aging, nutrition, and oxidative stress because oxidative injury in several organ systems is a prominent feature in aging. We investigated the possibility that DR can protect vulnerable liver lipids against age-related increases of peroxidation. Male Fischer 344 rats fed ad libitum (AL) or dietarily restricted (maintained on 60% of AL food intake) were killed by decapitation at 4 (young) or 12 mon (adult) of age. Phosphatidylcholine hydroperoxide (PCOOH) concentration of liver was determined using a chemiluminescent high-performance liquid chromatographic method. Liver PCOOH increased with age in adult rats, but less of an increase of PCOOH was seen in DR rats, which is consistent with results on production of thiobarbituric acid-reactive substances and oxygen-derived free radicals. No significant differences were found in liver superoxide dismutase and catalase activity between AL and DR groups of young and adult rats. Liver triglyceride and cholesterol contents were lower in DR than AL rats at 12 mon. Fatty acid compositions of phosphatidylcholine and phosphatidylethanolamine indicated that the ratio of (20:3n-6 + 20:4n-6)/18:2n-6, an index of linoleic acid (18:2n-6) desaturation, was lower in DR than in AL rats. We concluded that DR suppresses age-related oxidative damage in liver by modulating the amount of lipid as well as fatty acid composition.

Automatic identification of human helminth eggs on microscopic fecal specimens using digital image processing and an artificial neural network.

In order to automate routine fecal examination for parasitic diseases, we propose in this study a computer processing algorithm using digital image processing techniques and an artificial neural network (ANN) classifier. The morphometric characteristics of eggs of human parasites in fecal specimens were extracted from microscopic images through digital image processing. An ANN then identified the parasite species based on those characteristics. We selected four morphometric features based on three morphological characteristics representing shape, shell smoothness, and size. A total of 82 microscopic images containing seven common human helminth eggs were used. The first stage (ANN-1) of the proposed ANN classification system isolated eggs from confusing artifacts. The second stage (ANN-2) classified eggs by species. The performance of ANN was evaluated by the tenfold cross-validation method to obviate the dependency on the selection of training samples. Cross-validation results showed 86.1% average correct classification ratio for ANN-1 and 90.3% for ANN-2 with small variances of 46.0 and 39.0, respectively. The algorithm developed will be an essential part of a completely automated fecal examination system.

Acid secretion from a heterotopic gastric mucosa in the upper esophagus demonstrated by dual probe 24-hour ambulatory pH monitoring.

Heterotopic gastric mucosa in the upper esophagus is frequently found during endoscopic examination. Although most patients with heterotopic gastric mucosa of the upper esophagus, referred as inlet patch, are asymptomatic, symptomatic patients with complications resulting from this ectopic mucosa have also been reported. Acid secretion by the inlet patch has been suggested in some reports. We report a case of heterotopic gastric mucosa in the upper esophagus, with secretion of acid, demonstrated by continuous ambulatory pH monitoring, and the improvement of pharyngeal symptoms after the use of a proton pump inhibitor.

A case of primary adenosquamous carcinoma of the liver presented with liver abscess.

Primary adenosquamous carcinoma of the liver is a very rare type of cholangiocarcinoma and is defined as a cancer containing both squamous and adenomatous components in the same lesion. Recently, we experienced a primary adenosquamous carcinoma of the liver presented as liver abscess. A 63-year-old man was presented with a 4-day history of fever and chill. The radiologic study showed a 4 cm-sized, central hypoattenuated mass with peripheral rim enhancement in the left lobe of the liver. Ultrasonography-guided aspiration and biopsy suggested an adenocarcinoma with abscess in the liver. At laparotomy, the tumor occupied the left lobe of the liver and invaded the right diaphragm. An extended left lobectomy and a partial excision of the involved diaphragm were done. Grossly, the tumor was 6 x 5 x 5 cm in size and had an eccentric necrosis. Microscopically, the tumor was composed of adenocarcinoma and squamous cell carcinoma with a transitional area.

The effects of ascorbic acid on dopamine-induced death of PC12 cells are dependent on exposure kinetics.

The role of ascorbic acid on dopamine (DA) oxidation-mediated cytotoxicity was studied using the PC12 cell line. DA cytotoxicity was slightly attenuated by ascorbic acid, whereas the cytotoxicity of 6-hydroxydopamine (6-OHDA), a DA oxidation product, was markedly potentiated. To elucidate the relationship between the ascorbic acid effect and the degree of DA oxidation, ascorbic acid was added in a time-dependent fashion after DA treatment. We found greater cell death the later ascorbic acid was applied. Treatment of cells with glutathione alleviated DA- and 6-OHDA-induced cell death, while L-buthionine sulfoximine potentiated DA and 6-OHDA cytotoxicity. Ascorbic acid combined with glutathione eliminated the toxicity of DA and 6-OHDA. These results suggest that the interaction between DA and ascorbic acid is dependent upon the degree of DA oxidation and glutathione availability.

Choline acetyltransferase-immunoreactive neurons in the developing rat retina.

The development of cholinergic cells in the rat retina has been examined with immunocytochemistry by using antisera against choline acetyltransferase (ChAT). ChAT-immunoreactive (IR) cells were first detected at embryonic day 17 (E17) in the transitional zone between the neuroblastic layer (NBL) and ganglion cell layer (GCL). At E20, ChAT-IR cells are located exclusively in the GCL. At postnatal day 0 (P0), ChAT immunoreactivity appeared for the first time in cells at the distal margin of the NBL. Two prominent bands of labeled processes were first visible at P3, and by P15, these two bands resembled those of the adult retina. In addition, ChAT immunoreactivity appeared transiently in horizontal cells from P5 to P10. The number of ChAT-IR cells increased steadily up to P15. This resulted in a 93.8-fold increase between E17 and P15 (680-63,800 cells). However, after P15, the number declined by 19% from 63,800 cells at P15 to 51,800 in the adult. At all ages, the spatial density of each ChAT-IR cell population in the central retina was higher than in the periphery. In both central and peripheral regions, the peak density of ChAT-IR cells in the GCL was attained at E20. However, in the INL, the peak densities occurred at P3 in the central region and at P5 in the peripheral region. Up to P15, the soma diameter of ChAT-IR cells in the INL and GCL in each region increased continuously, reaching peak values at P15. Our results demonstrate that ChAT immunoreactivity is expressed in early developmental stages in the rat retina, as in other mammals, and that acetylcholine released from ChAT-IR cells may have neurotrophic functions in retinal maturation.

Nitric oxide is involved in sustained and delayed cell death of rat retina following transient ischemia.

We have investigated the role of nitric oxide (NO) in the rat retina following ischemic injury induced by transient increase of intraocular pressure. The thickness of both the inner plexiform layer and inner nuclear layer decreased during early postischemic stages (up to 1 week). In late postischemic stages (2-4 weeks), the thickness of the outer nuclear layer (ONL) decreased markedly. Thus, mechanisms other than excitotoxic ones may contribute to postischemic retinal cell death. Treatment of rats with N(G)-nitro-L-arginine methyl ester, a nitric oxide synthase (NOS) inhibitor, significantly reduced ischemic damage. Our findings suggest that NO is involved in the mechanism of ischemic injury, and plays a key role in the delayed and sustained cell death in the ONL following transient retinal ischemia.

Application of flow injection--chemiluminescence to the study of radical scavenging activity in plants.

Chemiluminescence (CL) was observed during the oxidation of luminol (2 mg/L). mediated by 0.06% hydrogen peroxide (H(2)O(2)) and cytochrome c (10 mg/L). CL intensity was decreased by the presence of radical scavengers and the reduction was linearly proportional to the concentration and ability of scavengers; butylated hydroxytoluene (BHT), caffeic acid and gallic acid. The order of effectiveness as radical scavengers was gallic acid > caffeic acid > BHT, which shows that the number of hydroxyl groups (OH) in the B-ring of flavonoids plays a key role in a good radical scavenging activity. Of eight catechins obtained from green tea extracts, (-)-catechin was the least effective and (-)-epigallocatechin gallate (EGCg) showed the strongest activity. This result indicates that the stereoscopic structure between the C-3 group and the B ring of flavonoids as well as substituents at the C-3 position make a contribution to radical scavenging activity. Of the tested Chinese herbal ingredients, five species of ingredients represented more than 90% of the radical scavenging activity.

Topographic expression of p21WAF1/SDI1/CIP1, bcl2, and p53 is altered at the early stage of colorectal carcinogenesis.

We analyzed the expression of p21, bcl2, and p53 in normal and different pathologic mucosa of the human colorectum using immunohistochemistry and cold polymerase chain reaction-single strand conformation polymorphism. The topography of normal mucosa showed; bcl2 and p53 expression restricted to basal epithelial cells and p21 expressed only in superficial epithelial cells. This topographic expression was altered in hyperplastic polyps and adenomas. Hyperplastic polyps revealed absence of or weak bcl2 expression and strong p21 expression without topography. In adenomas, whereas bcl2 expression increased and extended to parabasal and superficial dysplastic epithelium, the increase of p21 expression was limited to surface dysplastic epithelium. p53 was weakly expressed throughout the full thickness of dysplastic epithelium. Bcl2 expression in adenomas was stronger than in carcinomas; p53 expression was converse and p21 expression was variable. In carcinomas, this topographic expression was largely abrogated but p53 mutation (36%) was more frequent than in adenomas (2%). In carcinomas, p21 and p53 expression correlated inversely, but there was no relationship with bcl2. These results suggest that there is precisely ordered topographic pattern of p21, bcl2, and wild p53 expression in normal colorectal cells, but this becomes disordered during the early stage of colorectal carcinogenesis.

The antioxidative effect of ganhuangenin against lipid peroxidation.

The antioxidative effect of ganhuangenin (GHG), isolated from Scutellaria baicalensis Georgi, was examined by measuring its ability to suppress the formation of phospatidylcholine hydroperoxide (PCOOH). The results show that a pretreatment with GHG effectively suppressed PCOOH formation, which was initiated by the peroxyl-generating oxidant, AAPH (2,2'-azobis-2-aminopropane hydrochloride). The protective action of GHG against the formation of the PCOOH was observed in liver, lung, and kidney. When compared with other known antioxidants, we found the antioxidative potency of GHG to be greater than that of alpha-tocopherol. Our data strongly indicate that GHG is a powerful antioxidant against lipid peroxidation and is, therefore, responsible for this prophylactic effect.

Horizontal cells of the rat retina show choline acetyltransferase- and vesicular acetylcholine transporter-like immunoreactivities during early postnatal developmental stages.

We examined cholinergic neurons in the developing rat retina; an antiserum against choline acetyltransferase (ChAT) and an antiserum against vesicular acetylcholine transporter (VAChT) were used. From postnatal day 4 (P4) to P10, ChAT- and VAChT-like immunoreactivities were seen in cells which were located in the outer part of the inner nuclear layer. These cells had relatively large cell bodies and extended several transversely oriented processes. Double fluorescence immunohistochemistry using an antiserum against calbindin D-28K, a specific marker for the horizontal cells, revealed that all of ChAT- or VAChT-labeled cells showed calbindin D-28K-like immunoreactivity. These cells were no longer immunostained after P11. Thus, acetylcholine was considered to be transiently synthesized in the horizontal cells during early postnatal developmental stages in the rat retina.