Emergence and scattering of multiple neurofibromatosis (NF1)-related sequences during hominoid evolution suggest a process of pericentromeric interchromosomal transposition

Type 1 neurofibromatosis (NF1) gene encodes for a member of the GTPase activating protein family and is considered to be a tumor suppressor gene. Its very high rate of de novo mutation in humans led us to study a specific feature of this gene: the presence of numerous NF1-related sequences. According to our results, the human genome contains at least 11 NF1-related sequences, nine of which are scattered near centromeric sequences of seven different chromosomes. These NF1-related sequences, whose extent is quite varied according to loci, are unprocessed copies of the NF1 gene, and bear numerous mutations. A phylogenetic analysis of the six largest sequences indicates that they are all derived from a common ancestor, which would have appeared 22-33 million years ago, and was subsequently duplicated several times during hominoid evolution. The most recent duplication and interchromosomal transposition occurred in the last million years suggesting that the process could still be ongoing. Intriguing similarities between the evolution of alpha- satellite DNA and NF1-related sequences suggest the involvement of a common genetic mechanism for the generation and pericentric spreading of these NF1 partial copies.      

Influence of sugars on the activity of cellulase system from two basidiomycetes cultures

Influence of some sugars was studied on activity of purified cellulase enzymes produced by two white-rot basidiomycetes cultures in submerged fermentation. Exo- and endo-glucanases were found to be stimulated by glucose, sucrose and xylose, but β-glucosidase was inhibited by all sugars tested except sucrose being neutral. Cellobiose severely inhibited all three cellulase activities at all three concentrations taken in reaction mixture.     

Homozygosity mapping of achromatopsia to chromosome 2 using DNA pooling

Achromatopsia is an autosomal recessive disease of the retina, characterized clinically by an inability to distinguish colors, impaired visual acuity, nystagmus and photophobia. A genome-wide search for linkage was performed using an inbred Jewish kindred from Iran. To facilitate the genome-wide search, we utilized a DNA pooling strategy which takes advantage of the likelihood that the disease in this inbred kindred is inherited by all affected individuals from a common founder. Equal molar amounts of DNA from all affected individuals were pooled and used as the PCR template for short tandem repeat polymorphic markers (STRPs). Pooled DNA from unaffected members of the kindred was used as a control. A reduction in the number of alleles in the affected versus control pool was observed at several loci. Upon genotyping of individual family members, significant linkage was established between the disease phenotype and markers localized on chromosome 2. The highest LOD score observed was 5.4 (theta = 0). When four additional small unrelated families were genotyped, the combined peak LOD score was 8.2. Analysis of recombinant chromosomes revealed that the disease gene lies within a 30 cM interval which spans the centromere. Additional fine-mapping studies identified a region of homozygosity in all affected individuals, narrowing the region to 14 cM. A candidate gene for achromatopsia was excluded from this disease interval by radiation hybrid mapping. Linkage of achromatopsia to chromosome 2 is an essential first step in the identification of the disease-causing gene.      

Inhibition of hepatitis C virus nonstructural protein, helicase activity, and viral replication by a recombinant human antibody clone

Hepatitis C virus (HCV) nonstructural protein 3 (NS3), with its protease, helicase, and NTPase enzymatic activities, plays a crucial role in viral replication, and therefore represents an ideal target for the development of anti-viral agents. We have developed a recombinant human antibody (Fab) that reacts with the helicase domain of HCV NS3. The affinity-purified Fab antibody completely inhibited the helicase activity of HCV NS3 at equimolar concentration. To evaluate the effect of the Fab on HCV replication, the clone encoding the Fab gene was put into an expression vector, which converts Fab into a complete IgG1 antibody. Using a DNA-based transfection model, we demonstrated that intracellular expression of this antibody resulted in significant reduction of HCV-negative strand RNA synthesis. Intracellular expression of this antibody into either a stable cell line replicating subgenomic RNA, or a transient full-length HCV replication model, reduced both HCV RNA and viral protein expression. These results support the use of recombinant antibody fragments to inhibit NS3 enzyme as a novel, feasible, and effective approach for inhibiting HCV replication.     

Toxic Clostridioides (formerly Clostridium) difficile colitis: No longer a diarrhea associated infection

BackgroundClostridioides difficile infection (CDI) is traditionally taught to be an antibiotic associated diarrheal infection. This diagnosis is based on the presence of clinical symptoms (usually defined as more than 3 watery, loose or unformed stool within 24 h) coupled with a diagnostic test. There is now a new presentation of CDI, including progression to toxic megacolon, in patients without diarrhea.MethodsWe report a case series of 9 surgical patients from a single institution who developed CDI without preceding diarrhea.ResultAll 9 patients had CDI with positive laboratory testing for C. difficile toxin. They, however, presented with a lack of or minimal bowel movements. Six patients had rapid development of abdominal distention, 1 patient had a single episode of watery stool in 3 days, while the other 2 patients presented with constipation. Seven patients received stool softeners, suppositories and/or enemas for presumed constipation. Four patients had a mild course of infection and were successfully treated medically. The other 5 patients developed toxic megacolon, and eventually required total abdominal colectomy. Out of the 5 patients that required total colectomy, 2 expired.ConclusionCDI must be suspected in patients who rapidly develop abdominal distention, vague abdominal complaints or change in bowel function even in the absence of diarrhea, especially if coupled with multi-system organ failure.     

Adenovirus-mediated p53 gene therapy in nasopharyngeal cancer

EBV-associated nasopharyngeal cancer (NPC) occurs with high frequency in China and is a major cause of morbidity and mortality. To explore the potential use of adenovirus-mediated tumor suppressor p53 gene therapy In NPC, we first examined the in vitro effects of p53 introduced into the NPC cell lines RPMI 2650, Fadu and Detroit 562. p21(WAF1/CIP1) induction by chemotherapy was used as a functional assay which revealed that RPMI 2650 expresses wild-type p53 whereas Fadu and Detroit 562 encode mutant p53. Infection with p53-expressing adenovirus (Ad-p53) induced apoptosis and inhibited cell growth in all three NPC cell lines, regardless of the endogenous p53 status. Adenovirus infectivity was greatest in RPMI 2650 cells, with 100% of the cells expressing beta-galactosidase following Ad-LacZ infection using an MOI of 100, as compared to 20-30% infectivity with the other NPC lines. Using RPMI 2650 cells injected into nude mice, we developed an animal model for nasopharyngeal cancer. Established tumors (0.6-0.8 cm) were injected with 5x10(9) PFU Ad-LacZ, Ad-p53 or PBS in a 100 mu l volume. We found evidence for in vivo expression of beta-galactosidase or p53 and p21 up to two weeks following Ad-LacZ or Ad-p53 virus injection respectively. Objective regression of tumor size was observed at two weeks in 4/6 Ad-p53-treated tumors, but not in Ad-LacZ or PBS-treated tumors. The results provide an animal model for human nasopharyngeal cancer, and indicate a potential use of p53 in its therapy in vivo.     

Interventional radiology placement of twin Tesio catheters for dialysis access: review of 75 patients

PURPOSE: To evaluate the safety and efficacy of modern interventional radiology techniques and imaging guidance for placement of jugular vein twin Tesio hemodialysis catheters. MATERIALS AND METHODS: Eighty-two sets (75 patients) of twin Tesio catheters were percutaneously placed in the right (n = 70) and left (n = 12) internal jugular veins with use of ultrasound (US) and fluoroscopic guidance. Immediate procedural and late complications were recorded. The efficacy of the Tesio system was also evaluated. RESULTS: With US and fluoroscopic guidance, the technical success for access and catheter placement was 100%. Measured dialysis blood flow rate of greater than 375 mL/min was obtained in 95% of the patients and recirculation averaged 4.6% +/- 5%. An inadvertent common carotid artery puncture occurred in one (0.6%) patient and prolonged exit site bleeding occurred in another five patients (3%). Each of these was successfully controlled with compression. More chronically, catheter thrombosis and exit site infection occurred each at the rate of 0.16 episodes per 100 catheter days. All thrombosis and exit site infections responded to local thrombolysis and antibiotic therapy, respectively. Bacteremia occurred in 20 patients and required catheter removal in five patients. There was no clinical evidence of upper extremity or superior central vein thrombosis. CONCLUSION: Placement of internal jugular, twin Tesio catheters with use of imaging and interventional techniques provides a safe and efficacious means of either short or long-term hemodialysis.