Evaluation of groEL gene analysis for identification of Borrelia burgdorferi sensu lato

The nucleotide sequences of the groEL genes, the flagellin genes, and the 16S rRNA genes from 22 reference strains of Borrelia were compared. groEL sequence analysis is useful not only in interspecies differentiation but also in intraspecies differentiation of Borrelia afzelii and Borrelia garinii isolates.     

Burkitt's lymphoma representing periportal infiltrating mass on CT

Imaging findings of secondary hepatic lymphoma have been reported as variable, ranging from single or multiple small nodules to diffuse infiltrative tumor patterns. We hear present a rare case report concerning aggressive B cells, secondary Burkitt's lymphoma in non-AIDS demonstrating a surprising periportal lymphoma infiltration, without upper abdominal lymphadenopathy or splenomegaly on the sonography and CT scans. Clinically, the case was characterized by atypical and highly aggressive course, with the patient presenting an abruptly developed obstructive jaundice with rapidly deteriorating hepatic function that could be indicative of cholestatic hepatitis, which differs in its clinical manifestations from hepatic lymphoma without functional deterioration in respect of its non-tissue destructive growth pattern. We suggest that hepatic lymphoma can sometimes be consistent with periportal infiltrating homogeneous mass, with no lymphadenopathy or splenomegaly on the imaging examination, with a predictable aggressive clinical course of the disease and poor prognosis.     

Altered expression of KCNK9 in colorectal cancers

K(+) channels have been reported to be involved in the proliferation of many types of cells, including some human carcinoma and tumor cell lines. KCNK9, a TASK channel, is amplified and overexpressed in several types of human cancer. In the present study, we examined the expression and somatic mutations of KCNK9 in 124 colorectal cancers by immunohistochemistry using tissue microarray and PCR-SSCP. Immunopositivity was observed in 57 (46.0%) of 124 colorectal cancers. Clinically, KCNK9 was immunopositive in 4 (30.7%) of 13 cases which were stage A, 26 (55.3%) of 47 which were stage B, 23 (41.1%) of 56 which were stage C, and 4 (50%) of 8 which were stage D. Statistically, KCNK9 protein expression was not related to tumor stage (Bartholomew test, p>0.05) and lymph node metastasis (Chi-Square test, p=0.8338). In the mutation study of the KCNK9 gene, we found only one sequence variation (ACG-->ACC, Thr-->Thr) at codon 170 both in corresponding normal and tumor DNAs. These results indicate that overexpression rather than mutation of the KCNK9 gene may contribute to the development of colorectal cancers and suggest that the development of KCNK9-targeted agents may provide new possibilities in the treatment of colorectal cancer.     

A neural network method for identification of RNA-interacting residues in protein

Identification of the most putative RNA-interacting residues in protein is an important and challenging problem in a field of molecular recognition. Structural analysis of protein-RNA complexes reveals a strong correlation between interaction residues and their structure. Building on this viewpoint, we have developed a neural network predictor to correctly identify residues involved in protein-RNA interactions from protein sequence and its structural information. The system has been exhaustedly cross-validated with various strategies differing in input encoding, amount of input information, and network architectures. In addition, we have evaluated performance among functional subsets of complexes. Finally, to reflect the properties of protein-RNA complexes in our dataset, two kinds of post-processing method are adopted. The experimental result shows that our system yields not-trivial performance although the residues in interaction sites are too scarce.     

Disposition mechanisms of raloxifene in the human intestinal Caco-2 model

The purpose of this study was to determine the mechanisms responsible for transport of raloxifene and its hydrophilic conjugates. Human intestinal Caco-2 cell culture model and Caco-2 cell lysate were used for the studies. The results indicated that absorptive permeability (PAB) of raloxifene was lower than its secretory permeability (PAB). As the concentration increased, the efflux ratio (PBA/PAB) decreased, but PBA increased. PAB was also increased in the presence of verapamil and cyclosporine A, two P-glycoprotein inhibitors. Raloxifene was extensively metabolized into sulfated and glucuronidated conjugates. The extent of metabolism or clearance was decreased as the concentration increased from 3.4 (96%) to 30 (22%) microM. Multidrug resistance-related protein inhibitors MK-571 (C26H26ClN2O3S2) and leukotriene C4 significantly decreased (maximal 80%) apical efflux of both conjugates. They also significantly decreased (maximal 85%) basolateral efflux of glucuronides but not sulfates. On the other hand, organic anion transporter (OAT) inhibitor estrone sulfate and estrone glucuronide significantly decreased (maximal 50%) the efflux of sulfate from both sides but had variable effects on glucuronide efflux. Inhibition of conjugate efflux with the OAT inhibitor estrone sulfate was concentration dependent, which resulted in increased transport of intact raloxifene (maximal 90%). This increase in raloxifene transport was also observed in the presence of another OAT inhibitor estrone glucuronide (70%). In conclusion, this is the first report that inhibition of an efflux transporter responsible for the transport of metabolites can result in increase in the transport of the intact compound. It also provides additional explanation why raloxifene has low bioavailability but a long half-life.     

Evaluation of four DNA extraction methods for the detection of Mycobacterium avium subsp. paratuberculosis by polymerase chain reaction

Polymerase chain reaction (PCR) has been widely used due to its high specificity, sensitivity, and rapid turn-around time. However, inhibitory factors may be co-extracted with the target nucleic acid that will hinder the performance of PCR. In this study, DNA extraction methods for Mycobacterium avium subsp. paratuberculosis were evaluated including rapid lysis, organic extraction, silica-based and magnetic particle-based (MagaZorb) technologies on bacterial cells, and spiked bovine feces. Efficiency of the extraction was determined by PCR end point titration with primers targeting the insertion sequence, IS900. Results of the end point titrations are identical for bacterial cells and spiked feces. Inhibition was observed in PCR with DNA isolated from spiked feces, and a 1/100 dilution was able to alleviate this problem with DNA extracted by MagaZorb. A 1/1000 dilution was required for the other three methods. MagaZorb proved to be more efficient at removing inhibitory factors and required the least labor and completion time. Further evaluation is required for its utilization in other clinical specimens.     

Skf 525-A induces cocaine N-demethylase, ethoxyresorufin O-deethylase, and pentoxyresorufin O-dealkylase activities by induction of cytochrome p-450 2B in female B6C3F1 mice

Studies demonstrated that cocaine-induced immunosuppression is mediated by metabolites of cocaine. Although SKF 525-A inhibited cocaine N-demethylation in liver S9 fractions isolated from female B6C3F1 mice, our study showed that pretreatment of mice with SKF 525-A potentiated cocaine-induced suppression of the antibody response to sheep red blood cells. An increase in formaldehyde generation was subsequently shown following incubation of cocaine with the S9 fractions prepared from SKF 525-A-treated mice, indicating the possibility of cytochrome P-450 (CYP) induction. Therefore, the inductive effects of SKF 525-A on CYP enzyme activities and proteins were investigated in female B6C3F1 mice to elucidate the potentiation of cocaine-induced immunosuppression by SKF 525-A. When SKF 525-A was administered at 10, 20, or 40 mg/kg/d intraperitoneally for 7 consecutive days, both ethoxyresorufin O-deethylase and pentoxyresorufin O-dealkylase activities were induced dose-dependently. Furthermore, the induction of enzymatic activity was time dependent. Meanwhile, when the type of isozyme induced by SKF 525-A was analyzed by Western immunoblotting with monospecific anti-CYP 1A and anti-CYP 2B antibodies, only the CYP 2B appeared to be induced. From in vitro inhibition studies with monoclonal antibodies, it was confirmed that the induced activity of ethoxyresorufin O-deethylase by SKF 525-A was due to increased levels of CYP 2B proteins. Our present results provide an explanation for the potentiation of cocaine-induced immunosuppression by repeated exposure to SKF 525-A. Our results also indicate that ethoxyresorufin O-deethylase, a selective substrate for CYP 1A, may also be catalyzed by CYP 2B.