Reactivation of heat-inactivated Ku proteins by heat shock cognate protein HSC73

Purpose: Mouse double-stranded DNA-dependent protein kinase (DNA-PK) activity is heat sensitive. Recovery of heat-inactivated DNA repair activity is a problem after combination therapy with radiation and heat. We investigated the mechanism of recovery of heat-inactivated DNA-PK activity.

Methods: Hybrid cells containing a fragment of human chromosome 8 in scid cells (RD13B2) were used. DNA-PK activity was measured by an in vitro assay. Immunoprecipitation of the nuclear extract was performed with an anti-Ku80 antibody. Proteins co-precipitated with Ku80 were separated by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis and detected by Western blotting using anti-heat shock protein (HSP)72 and anti-heat shock cognate protein (HSC)73 antibodies. HSC73 was overexpressed with the pcDNA3.1 vector. Short hairpin (sh)RNA was used to downregulate HSC73 and HSP72.

Results: The activity of heat-inactivated DNA-PK recovered to about 50% of control during an additional incubation at 37?°C after heat treatment at 44?°C for 15?min in the presence of cycloheximide (which inhibits de novo protein synthesis). Maximal recovery was observed within 3?h of incubation at 37?°C after heat treatment. Constitutively expressed HSC73, which folds newly synthesized proteins, reached maximal levels 3?h after heat treatment using a co-immunoprecipitation assay with the Ku80 protein. Inhibiting HSC73, but not HSP72, expression with shRNA decreased the recovery of DNA-PK activity after heat treatment.

Conclusions: These results suggest that de novo protein synthesis is unnecessary for recovery of some heat-inactivated DNA-PK. Rather, it might be reactivated by the molecular chaperone activity of HSC73, but not HSP72.     

Easier node dissection after chemoradiotherapy for lung cancer with collagen insertion at mediastinoscopy

Objective: Induction chemoradiotherapy followed by anatomical resection is a current therapeutic strategy for non-small-cell lung cancer with mediastinal node involvement. Dense peritracheal fibrosis and sclerosis after chemoradiotherapy cause difficult mediastinal node dissection. We evaluated a novel technique to make the mediastinal node dissection easier after induction therapy. Methods: At the end of mediastinoscopic node biopsy for staging of lung cancer, cotton-type collagen was inserted anterior and lateral to the trachea in patients with pathologically confirmed mediastinal node involve-ment (n=45). The induction therapy consisted of concurrent use of platinum-based chemotherapy and hyperfractionated radiotherapy. After the chemoradiotherapy all patients underwent a pulmonary resection with complete mediastinal node dissection 7–12 weeks after the collagen insertion. Surgical findings of the mediastinum and the time for node dissection were compared with those without collagen insertion at mediastinoscopy after chemoradiotherapy (n=5). Results: All five patients without collagen insertion showed sclerotic and fibrotic change of mediastinal nodes with severe adhesion to the trachea. In 42 of 45 patients with collagen insertion (93.3%) the collagen remained unabsorbed and separated the mediastinal nodes from the trachea. Mediastinal node dissection was easily accomplished by removing mediastinal tissues lateral and anterior to the collagen. The rate of mediastinal node separation was significantly higher with collagen insertion than without (p< 0.0001). The times for node dissection in patients with and without collagen insertion showed no significant difference. Conclusion: Cotton-type collagen insertion at staging mediastinoscopy for lung cancer separates the mediastinal nodes from the trachea and makes the node dissection easier after induction chemoradiotherapy.     

Genetic linkage analysis of pulmonary fibrotic response to silica in mice.

Inter-individual variations in the development of silicosis, even within the same environments, have been reported, which suggest the contribution of genetic factors in silicosis aetiology. The aim of the present study was to determine whether there is any significant genetic influence on the development of silicosis. Furthermore, which genetic loci are responsible for the pulmonary response to silica exposure? Eight strains of inbred mice were used to examine the genetic influence on the lung fibrotic response to silica exposure. After intercross-breeding between the most susceptible and most resistant strains, a genome-wide linkage analysis of quantitative trait loci (QTL) was performed. Hydroxyproline was applied as an index, and genotypes of 167 marker genes were analysed by fragment analysis using a capillary-type sequencer. There was significant inter-strain difference in the mean concentration of hydroxyproline contents among the eight strains of mice. Breeding studies were conducted between the most susceptible, C57BL/6J, and the most resistant strain, CBA/J. A genome-wide linkage analysis of silica-exposed intercrossed cohorts identified significant QTL on chromosome 4 and suggestive QTL on chromosomes 3 and 18. The present study demonstrates that genetic factors may play a significant role in fibrotic-lung responses to silica; one significant and two suggestive quantitative trait loci were identified.     

ENDOSCOPIC NASOBILIARY DRAINAGE: CURRENT INDICATIONS AND EVALUATION OF THE PRODUCTS

Endoscopic nasobiliary drainage (ENBD) is a well established mode of biliary decompression. Although ENBD is certainly an uncomfortable procedure with the potential risk of spontaneous dislocation or removal of the drainage catheter by disoriented patients, it has several advantages over endoscopic biliary drainage (EBD) using an indwelling stent. The current indications for ENBD are: (i) temporary drainage to treat obstructive jaundice and cholangitis caused by malignant or benign biliary stricture; (ii) urgent drainage to treat suppurative cholangitis primarily caused by common bile duct stones; (iii) temporary drainage after stone removal in patients with suspected incomplete clearance and/or with cholangitis; and (iv) biliary leaks that occur primarily after surgery, as well as other indications. Different types of nasobiliary catheters are currently available that have been designed with various diameters, shapes, and materials. However, the current catheters are not considered by most endoscopists to be sufficient. Further improvements are needed to achieve better drainage and better maneuverability.     

Immunohistochemical analysis of TrkA neurotrophin receptor expression in human non-neuronal carcinomas

The Trk family of tyrosine protein kinase receptors plays a significant role in the development and maintenance of neural tissues. It has been recently shown that Trk receptors are also expressed by a wide range of normal non-neuronal tissues in humans in a cell type-specific manner. In the present study, the expression patterns of TrkA in 337 non-neuronal invasive carcinomas of 15 different human tissues were investigated immunohistochemically. Overall, 133 (39%), 101 (30%) and 103 (31%) tumors exhibited strong, moderate and no TrkA Immunoreactivity, respectively. Esophageal and thyroid carcinomas expressed high levels of TrkA, whereas the levels in gastric and colon cancers were low. TrkA expression was detected not only in carcinomas originating from TrkA-positive normal counterpart tissues, Including the esophagus, breast, lung and uterus, but also in those from TrkA-negative tissues/cells of the thyroid, liver and ovary. Immunostaining for nerve growth factor-β, the specific ligand for TrkA, in esophageal and breast carcinomas demonstrated its immunoreactivity in stromal fibroblasts and some TrkA-expressing tumor cells. These results suggest that paracrine/autocrine regulation via stromal/tumoral NGF-tumoral TrkA interaction may be involved In the growth of certain non-neuronal carcinomas.     

Experience of expandable metallic stents for central airway obstruction

The aim of this study was to investigate retrospectively the efficacy of expandable metallic stents (EMSs) for severe respiratory distress in patients with central airway obstruction. Twenty patients with central airway obstructions were treated with an EMS. An intraluminal stricture was present in 15 and an extraluminal stricture in 5 patients. Of the 15 patients with intraluminal stenosis, 11 exhibited symptomatic improvement. All 11 patients had tumor infiltration occupying less than 50% of the endoluminal diameter. The other four patients with intraluminal stenosis had tumor infiltration occupying > 50% of the endoluminal diameter and demonstrated no improvement. All five patients with extraluminal stenosis were improved. EMS is useful for an extraluminal stricture in the central airway and the effect of EMS for intraluminal stenosis is related to the degree of infiltration and of tumor progression itself.      

Availability of 4'galactosyllactose (O-beta-D-galactopyranosyl-(1----4)-O-beta-D-galactopyranosyl-(1----4)- D-glucopyranose) in rat

O-beta-D-Galactopyranosyl-(1----4)-O-beta-D-galactopyranosyl-(1----4)-D- glucopyranose (designated as 4'GL) is produced from lactose by Cryptococcus laurentii. The influence of chronic ingestion of 4'GL on body weight gain, organ weight, serum lipids, and liver lipids was investigated in rats. The body weight gains of the 5% and 10% 4'GL-diet groups were higher than that of the control group. Food intake and fecal dry weight were significantly increased (p less than 0.05) by 4'GL feeding. The 4'GL diet produced a significant increase (p less than 0.01) in the wet weight and contents of both the cecum and the colon. However, no significant increase was observed in the weight of the stomach, small intestine, liver, or other organs. The effects of 4'GL on serum and liver lipid levels were not observed in this experiment. The digestion of 4'GL was measured in vitro using the artificial gastric juice, alpha-amylase of human saliva, alpha-amylase of hog pancreas, and mucosa of rat intestine. 4'GL was not hydrolyzed by these enzymes. Long-term ingestion of 4'GL did not cause any induction of 4'GL hydrolyzing enzyme activity in the rat small intestine.     

Reduced Expression of nm23 Is Associated with Metastasis of Human Gastric Carcinomas

Reduced expression of nm23 gene is implicated in high metastatic potential In a variety of malignancies. To elucidate the role of nm23 in human gastric carcinomas, we examined loss of heterozygosity (LOH) of nm23 gene by Southern blotting, nm23 mRNA expression by Northern blotting and nm23 protein expression by Western blotting as well as immunohistochemistry in both primary and metastatic tumors. LOH of nm23 gene was found in 2 (8%) out of the 23 informative gastric carcinomas. Twenty-two (84%) out of the 26 cases expressed nm23 mRNA at higher levels in primary tumor tissue than in corresponding non-neoplastic mucosa. No obvious correlation was observed between clinico-pathological features and LOH of nm23 gene or nm23 mRNA expression. On the other hand, 52% of the gastric carcinomas showed reduction of nm23 immunoreactivity in the metastatic tumor of regional lymph nodes, as compared to the primary tumor. Interestingly, 71% of the gastric carcinomas showed weaker nm23 immunoreactivity in the liver metastasis than in the primary tumor. These results suggest that nm23 overexpression is linked with development of gastric carcinomas and the decrease in expression of nm23 participates in metastasis.     

Practical guidelines for physicians in the management of febrile seizures

Recent studies have shown that adequate medication can prevent the recurrence of febrile seizures (FS). It has also been clarified that the vast majority of, though not all, FS patients follow a benign course. Then, questions arise as to whether or not FS should be prevented, particularly in light of the risks of side effects from drugs. Which kinds of FS can be prevented, if necessary? The guidelines presented here are aimed primarily at helping general practitioners in considering how to manage FS most appropriately. The guidelines stress that judgements should be individualized, while referring to a few specific ‘warning factors’. The guidelines follow a ‘laissez-faire’ principle for the majority of FS cases, whereas intermittent therapy with diazepam and continuous medication with either phenobarbital or valproate are indicated in other limited cases meeting respective definite criteria.