Chromosome breakpoint distribution in nonmelanoma skin cancers

Human herpesvirus 8 (HHV-8) DNA sequences were examined in peripheral blood mononuclear cell (PBMC) DNA samples of 56 children, 15 healthy adults, and 10 renal transplant patients by the polymerase chain reaction (PCR). The PCR amplification was carried out using the published KS330(233) primer pairs to amplify HHV-8 DNA sequences. The PCR-amplified products were confirmed by Southern blot hybridization with radiolabeled 233 bp HHV-8 DNA fragment, which was cloned and sequenced from the PCR-amplified product of Kaposi's sarcoma tissue. Six PCR-amplified product of four children and two renal transplant patients were cloned and sequenced. HHV-8 DNA sequences were detected in 36 of 56 (64%) normal children, in 12 of 15 (80%) healthy adults, and in all 10 renal transplant patients by Southern blot hybridization of PCR-amplified products. Six PCR-amplified products were confirmed by sequencing. These results suggest that HHV-8 infection is prevalent in the Japanese population with infection occurring in early childhood.     

A functional telomerase RNA swap in vivo reveals the importance of nontemplate RNA domains

The ribonucleoprotein (RNP) enzyme telomerase is required for replication of eukaryotic chromosomal termini. The RNA moiety of telomerase is essential for enzyme function and provides the template for telomeric DNA synthesis. However, the roles of its nontemplate domains have not been explored. Here we demonstrate that a novel interspecies telomerase RNA swap in vivo creates a functional but aberrant telomerase. Telomerase RNA from the ciliate Glaucoma chattoni was expressed in Tetrahymena thermophila cells. The telomerase RNAs from these two species have almost superimposable secondary structures. The template region base sequence is identical in the two RNAs, but elsewhere their sequences differ by 49%. This hybrid telomerase RNP was enzymatically active but added only short stretches of telomeric repeat tracts in vivo and in vitro. This new enzyme also had a strong, aberrant DNA cleavage activity in vitro. Thus, molecular interactions in the RNP involving nontemplate RNA domains affect specific aspects of telomerase enzyme function, raising the possibility that they may regulate telomerase activity.     

Structural polymorphism of the RecA protein from the thermophilic bacterium Thermus aquaticus

The Escherichia coli RecA protein has served as a model for understanding protein-catalyzed homologous recombination, both in vitro and in vivo. Although RecA proteins have now been sequenced from over 60 different bacteria, almost all of our structural knowledge about RecA has come from studies of the E. coli protein. We have used electron microscopy and image analysis to examine three different structures formed by the RecA protein from the thermophilic bacterium Thermus aquaticus. This protein has previously been shown to catalyze an in vitro strand exchange reaction at an optimal temperature of about 60 degrees C. We show that the active filament formed by the T. aquaticus RecA on DNA in the presence of a nucleotide cofactor is extremely similar to the filament formed by the E. coli protein, including the extension of DNA to a 5.1-A rise per base pair within this filament. This parameter appears highly conserved through evolution, as it has been observed for the eukaryotic RecA analogs as well. We have also characterized bundles of filaments formed by the T. aquaticus RecA in the absence of both DNA and nucleotide cofactor, as well as hexameric rings of the protein formed under all conditions examined. The bundles display a very large plasticity of mass within the RecA filament, as well as showing a polymorphism in filament-filament contacts that may be important to understanding mutations that affect surface residues on the RecA filament.     

The cellular trafficking and zinc dependence of secretory and lysosomal sphingomyelinase, two products of the acid sphingomyelinase gene

The acid sphingomyelinase (ASM) gene, which has been implicated in ceramide-mediated cell signaling and atherogenesis, gives rise to both lysosomal SMase (L-SMase), which is reportedly cation-independent, and secretory SMase (S-SMase), which is fully or partially dependent on Zn2+ for enzymatic activity. Herein we present evidence for a model to explain how a single mRNA gives rise to two forms of SMase with different cellular trafficking and apparent differences in Zn2+ dependence. First, we show that both S-SMase and L-SMase, which contain several highly conserved zinc-binding motifs, are directly activated by zinc. In addition, SMase assayed from a lysosome-rich fraction of Chinese hamster ovary cells was found to be partially zinc-dependent, suggesting that intact lysosomes from these cells contain subsaturating levels of Zn2+. Analysis of Asn-linked oligosaccharides and of N-terminal amino acid sequence indicated that S-SMase arises by trafficking through the Golgi secretory pathway, not by cellular release of L-SMase during trafficking to lysosomes or after delivery to lysosomes. Most importantly, when Zn2+-dependent S-SMase was incubated with SMase-negative cells, the enzyme was internalized, trafficked to lysosomes, and became zinc-independent. We conclude that L-SMase is exposed to cellular Zn2+ during trafficking to lysosomes, in lysosomes, and/or during cell homogenization. In contrast, the pathway targeting S-SMase to secretion appears to be relatively sequestered from cellular pools of Zn2+; thus S-SMase requires exogeneous Zn2+ for full activity. This model provides important information for understanding the enzymology and regulation of L- and S-SMase and for exploring possible roles of ASM gene products in cell signaling and atherogenesis.     

Colorectal cancer: comparison of laparoscopic with open approaches

PURPOSE: We compared laparoscopic with open colectomy for treatment of colorectal cancer. METHODS: We performed a retrospective review of patients undergoing colectomy for colorectal cancer between January 1991 and March 1996 at a large private metropolitan teaching hospital. Operative techniques included open (n=90) and laparoscopic (n=80) colectomy. Laparoscopic colectomy was further subdivided into the following groups: facilitated (n=62), with extracorporeal anastomosis; near-complete (n=9), with small incision for specimen delivery only; complete (n=3), with specimen removal through the rectum; and converted to an open procedure (n=6). Main outcome measures included operative time, blood loss, time to oral intake, length of postoperative hospitalization, morbidity, lymph node yield, recurrence, survival, and costs. RESULTS: Operative time was equivalent in the laparoscopic and open groups (laparoscopic, 161 minutes; open, 163 minutes; P=0.94). Blood loss was less for the laparoscopic group (laparoscopic, 104 ml; open, 184 ml; P=0.001), and resumption of oral intake was earlier (laparoscopic, 3.9 days; open, 4.9 days; P=0.001), but length of hospitalization was similar. Mean lymph node yield in the laparoscopic group was 12 compared with 16 in the open group (P=0.16). Rates of morbidity, recurrence, and survival were similar in both groups. No port-site recurrences occurred. CONCLUSIONS: Laparoscopic and open colectomy were therapeutically similar for treatment of colorectal cancer in terms of operative time, length of hospitalization, recurrence, and survival rates. The laparoscopic approach was superior in blood loss and resumption of oral intake.     

Aerobic and anaerobic microbiology of empyema. A retrospective review in two military hospitals

The microbiology and clinical features of empyema were studied retrospectively in 197 patients whose specimens yielded bacterial growth after inoculation for aerobic and anaerobic bacteria. Three hundred forty-three organisms (216 aerobic or facultative and 127 anaerobic organisms) were isolated. Aerobic bacteria were isolated in 127 (64 percent) patients, anaerobic bacteria in 25 (13 percent), and mixed aerobic and anaerobic bacteria in 45 (23 percent). The predominant aerobic or facultative organisms were Streptococcus pneumoniae (70 isolates), Staphylococcus aureus (58), Escherichia coli (17), Klebsiella pneumoniae (16), and Haemophilus influenzae (12). The predominant anaerobes were pigmented Prevotella and Porphyromonas species (24), Bacteroides fragilis group (22), anaerobic cocci (36), and Fusobacterium species (20). beta-Lactamase-producing organisms were recovered in 49 (38 percent) of 128 tested specimens. These included all 42 tested S aureus and 15 B fragilis group, 4 of 9 K pneumoniae, 3 of 9 H influenzae, 3 of 8 pigmented Prevotella and Porphyromonas species, and 2 of 6 E coli. Most patients from whom S pneumoniae and H influenzae were recovered had pneumonia, and most patients with S aureus had pneumonia, aspiration pneumonia, and lung abscesses. The recovery of anaerobic bacteria was mostly associated with the concomitant diagnosis of aspiration pneumonia, and lung, subdiaphragmatic, dental, and oropharyngeal abscesses. These data highlight the importance of anaerobic bacteria in selected cases of empyema.     

Reversal and prevention of cerebral vasospasm by intracarotid infusions of nitric oxide donors in a primate model of subarachnoid hemorrhage

OBJECTIVE: To determine the frequency of adverse reactions, particularly the occurrence of apnea, among preterm infants after immunization with diphtheria and tetanus toxoids and whole cell pertussis vaccine adsorbed (DTP) and Haemophilus influenzae type b conjugate (HibC) vaccine in the neonatal intensive care unit. STUDY DESIGN: After the occurrence of apnea in two preterm infants following immunization with DTP and HibC, a prospective surveillance of 97 preterm infants younger than 37 weeks of gestation who were immunized with DTP (94 also received HibC at the same time) in the neonatal intensive care unit was performed to assess the frequency of adverse reactions and in particular, the occurrence of apnea. For each infant, data were recorded for a 3-day period before and after receipt of the immunization. RESULTS: The majority of preterm infants tolerated immunizations with DTP and HibC without ill effects. However, 12 (12%) infants experienced a recurrence of apnea, and 11 (11%) had at least a 50% increase in the number of apneic and bradycardic episodes in the 72 hours after immunization. This occurred primarily among smaller preterm infants who were immunized at a lower weight (p = 0.01), had experienced more severe apnea of prematurity (p = 0.01), and had chronic lung disease (p = 0.03). CONCLUSION: The temporal association observed between immunization of preterm infants and a transient increase or recurrence of apnea after vaccination merits further study. Cardiorespiratory monitoring of these infants after immunization may be advisable.