Protein Transduction Method for Cerebrovascular Disorders

Many studies have shown that a motif of 11 consecutive arginines (11R) is one of the most effective protein transduction domains (PTD) for introducing proteins into the cell membrane. By conjugating this &#34;11R&#34;, all sorts of proteins can effectively and harmlessly be transferred into any kind of cell. We therefore examined the transduction efficiency of 11R in cerebral arteries and obtained results showing that 11R fused enhanced green fluorescent protein (11R-EGFP) immediately and effectively penetrated all layers of the rat basilar artery (BA), especially the tunica media. This method provides a revolutionary approach to cerebral arteries and ours is the first study to demonstrate the successful transductionof a PTD fused protein into the cerebral arteries. In this review, we present an outline of our studies and other key studies related to cerebral vasospasm and 11R, problems to be overcome, and predictions regarding future use of the 11R protein transduction method for cerebral vasospasm (CV).</p

Diagnostic Validity of DNMT-1 and 3b Immunoreactivity in Non-neoplastic Epithelium of UC Patients with and Without Neoplasia

It is important to improve the efficacy of surveillance in UC patients with neoplasia.In the present study, we assessed the cut-off value of expression of DNMT-1 and DNMT-3b expression in the non-neoplastic rectal epithelium of patients with long-standing and extensive UC. Sixty patients with long-standing and extensive UC participated in this study( 30 with colorectal neoplasia and 30 without). Immunohistochemical analysis was performed to determine the expression of DNMT-1 and 3b in non-neoplasticrectal epithelium of UC patients without neoplasia, and in non-neoplastic rectal epithelium of UC patients with neoplasia. The level of immunoreactive DNMT-1 and DNMT-3b expression was determined as the percentage of positive cells relative to the total number of cells counted under high power magnification.DNMT-1 and 3b expression in non-neoplastic rectal epithelium of UC patients with neoplasia (0.57, range0.53-0.63)(0.32, range 0.18-0.67) was higher than in the non-neoplastic epithelium of UC patients without neoplasia (0.41, range 0.25-0.54, P=.001)(0.0, range 0.0-0.13, P<.001). ROC curve analysis confirmed0.53 and 0.07 as the best DNMT-1 and DNMT-3b cut-off values for identifying individuals at increased risk of neoplasia( area under the curve=0.798 and 0.842, respectively). The cut-off value for DNMT-1 andDNMT-3b expression in non-neoplastic rectal epithelium is therefore an efficient predictor for the increased risk of UC-associated neoplasia

Immunohistochemical Localization of REG Ia Protein in Salivary Gland Tumors

The regenerating gene( Reg) Ia protein has a trophic effect on gastric epithelial cells, and its overexpressionis reported in gastrointestinal cancers. The salivary gland is a component of the digestive system, andtherefore, REG Ia protein may play some role in the pathophysiology of salivary gland tumors. In the presentstudy, we determined the immunohistochemical localization of REG Ia protein in salivary gland tumorsand moreover investigated its relationship to clinicopathological features. Twenty-eight patients with salivarygland tumor were enrolled. The specimens resected by surgery from those patients were examinedusing immunohistochemistry for REG Ia protein and Ki67. Five of the 16 pleomorphic adenomas (31.3%)were positive for REG Ia protein. Regarding salivary gland carcinomas, four of five mucoepidermoid carcinomas(80%), three of five adenoid cystic carcinomas (60%), one of two polymorphous low-grade adenocarcinomas(50%) were also positive for REG Ia protein. However, no relationships were found betweenREG Ia protein expression and clinicopathological features. Regarding the Ki67 expression, strong signalwas observed in the tumor cells of patients with salivary gland adenoma as well as carcinoma. REG Ia proteinis expressed not only in adenocarcinoma but also precancerous adenoma cells proliferating actively,suggesting that REG Ia protein may play a role at least in part in the development of salivary gland tumors

A Novel Approach to Endoscopic Submucosal Dissection Using Bipolar Current Needle Knife for Colorectal Tumors

Objective: To completely and safely remove a large colorectal lesion in a single fragment, we have developed an endoscopic electrosurgical knife( B-Knife) for a more effective bipolar cutting adevelopedannd coagulation system.The aim of this study was to evaluate the effectiveness and safety of the B-Knife in patients with large colorectal tumors.Methods:Endoscopic submucosal dissection (ESD) using the B-Knife was performed initially in 3 patients with large colorectal tumors in a pilot study. Subsequently, we examined the clinical outcomes of ESDusing the B-Knife in 25 patients with colorectal tumors.Results:During initial clinical use of the B-Knife, en bloc resection was achieved in all 3 cases, and themean operating time was 43 minutes. All margins of resected material were histologically free of neoplasia.There were no cases of delayed bleeding or perforation. In a series of 25 ESD cases, which consisted of 8adenomas, 15 intramucosal carcinomas, one slightly submucosal invasive carcinoma, and one massive submucosalinvasive carcinoma, the en bloc resection rate was 84% . The mean operation time was 91.6 minutes and the mean size of resected specimens was 36.4 mm (range:19-80 mm). Perforations occurred in one( 4%) case, but delayed bleeding did not occur in any of the cases. Additional surgery was required for2 cases( 8%).Conclusions:ESD using the B-Knife is reliable and safe for the complete resection of select large flat lesions in the colorectum

Validation of Pyrosequencing for the Analysis of KRAS Mutations in Colorectal Cancer

The use of antibodies against epidermal growth factor receptor( EGFR) in conjunction with conventionalchemotherapy for metastatic colorectal cancer (CRC) in patients with KRAS wild-type tumors has beenproven to be efficacious. Recently, KRAS testing prior to anti-EGFR therapy has become mandatory formetastatic CRC patients. Although newly developed pyrosequencing is expected to be one of the highthroughput procedures detecting such mutations, the accuracy of the procedure has not been well evaluated.In the present study, we aimed to validate the accuracy, especially the potential for a false-negative result,in detecting KRAS mutations by pyrosequencing using cultured tumor cells. DNA extracted from cultured&igrave;NOZ&icirc; gallbladder cancer cells( known to contain KRAS mutation G12V) at concentrations of 1%, 5%, 10%, and 25%, as well as 2 DNA samples extracted from a resected CRC specimen( known to contain anotherKRAS mutation, G12C) at concentrations of 5% and 25%, were prepared. We analyzed KRAS mutationalstatus and nonexistent and/or nonfunctional mutations of these 6 samples using pyrosequencing. TheKRAS mutation detection rates in the 4 NOZ samples( 1%, 5%, 10%, and 25%) were 0.37%, 2.79%, 5.28%,and 13.85%, respectively. Some artifacts of KRAS mutations unlikely to be present were detected in 1%samples of NOZ at a rate similar to that of the G12V mutation( G12C, 0.29%;G13C, 0.42%). Although theKRAS mutation G12C was detected at rates of 1.26% and 6.49% in samples with 5% and 25% DNA extractedfrom resected CRC specimen, respectively, no other type of KRAS mutation was detected in suchsamples. Pyrosequencing could not detect KRAS mutations correctly in the sample containing 1% DNA.This might cause false negatives. A sample mutated DNA concentration of at least 5% was necessary forprecise analyses by this procedure