A system for assaying homologous recombination at the endogenous human thymidine kinase gene.

A system for assaying human interchromosomal recombination in vitro was developed, using a cell line containing two different mutant thymidine kinase genes (TK) on chromosomes 17. Heteroalleles were generated in the TK+/+ parent B-lymphoblast cell line WIL-2 by repeated exposure to the alkylating nitrogen mustard ICR-191, which preferentially causes +1 or -1 frameshifts. Resulting TK-/- mutants were selected in medium containing the toxic thymidine analog trifluorothymidine. Mutations were characterized by exon-specific polymerase chain reaction amplification and direct sequencing. In two lines, heterozygous frameshifts were located in exons 4 and 7 of the TK gene separated by approximately 8 kilobases. These lines undergo spontaneous reversion to TK+ at a frequency of less than 10(-7), and revertants can be selected in cytidine/hypoxanthine/aminopterin/thymidine medium. The nature and location of these heteroallelic mutations make large deletions, rearrangements, nondisjunction, and reduplication unlikely mechanisms for reversion to TK+. The mode of reversion to TK+ was specifically assessed by DNA sequencing, use of single-strand conformation polymorphisms, and analysis of various restriction fragment length polymorphisms (RFLPs) linked to the TK gene on chromosome 17. Our data suggest that a proportion of revertants has undergone recombination and gene conversion at the TK locus, with concomitant loss of frameshifts and allele loss at linked RFLPs. Models are presented for the origin of two recombinants.     

Enhancement of benzo[a]pyrene-induced sister chromatid exchanges in lymphocytes from cigarette smokers occupationally exposed to asbestos

The frequencies of base-line and benzo[a]pyrene [(BP) CAS 50-38-8]-induced sister chromatid exchanges (SCE) were measured in peripheral blood lymphocytes from 22 male asbestos-exposed workers and 10 nonexposed workers of comparable age. A clear association between cigarette smoking and asbestos exposure in the sensitivity of lymphocytes to BP was observed. Among asbestos-exposed workers, lymphocytes from those who smoked cigarettes were significantly more susceptible to the induction of SCE by in vitro exposure to BP (P = .01) than were lymphocytes from nonsmokers. Active smoking elevated the base-line SCE frequency in both asbestos-exposed and nonexposed workers (P = .001), and an interaction between smoking and asbestos in the production of base-line SCE was suggested (P = .07). Asbestos exposure alone was not associated with an enhanced susceptibility to the induction of SCE by BP or with an elevation of base-line SCE. Increased age was associated with an increase in SCE inducibility by BP (P = .01), and a history of smoking was marginally associated with SCE inducibility by BP (P = .07). These findings support the hypothesis that an increased susceptibility of asbestos-exposed individuals to polyaromatic hydrocarbon-induced cancer results from an enhanced sensitivity to the induction of genetic damage rather than to an asbestos-induced differential cellular metabolic capacity.     

Lack of acute d-amphetamine effects on thyrotropin release

Case reports have suggested that amphetamine abuse causes excessive secretion of thyrotropin (TSH) and thyroxine (T4). Such an amphetamine-induced effect might be noradrenergic-mediated in the hypothalamus. The current controlled study examined oral d-amphetamine effects on the hypothalamic-pituitary-thyroid axis in normal humans. No acute effects were seen on TSH, T3 or T4 levels. d-Amphetamine elevated cortisol levels at 180 min, as previously reported.     

Infection of mesothelial cells with human herpes virus 8 in human immunodeficiency virus-infected patients with Kaposi's sarcoma, Castleman's disease, and recurrent pleural effusions.

Recurrent pleural effusions are common complications of hospitalized patients with human immunodeficiency virus (HIV) infection and may pose difficult diagnostic dilemmas. A common cause of recurrent pleural effusions in up to 30% of HIV-seropositive patients is pulmonary involvement by Kaposi's sarcoma, a human herpesvirus 8 (HHV 8)-related neoplasm. The pathogenesis of these effusions is unclear. These recurrent effusions, although benign, have shown significant mesothelial atypia/reactive changes of uncertain etiology. We attempted to evaluate these effusions morphologically and molecularly for the presence of HHV 8, with particular attention to mesothelial cells. All recurrent pleural effusions, as defined by any effusion tapped for cytological examination on more than two occasions, in HIV-positive patients at the National Institutes of Health were examined from 1998 to the present. Cases were stratified according to patients with and without histologically confirmed HHV 8 disease manifestations. Five patients with HHV 8 diseases (four with disseminated Kaposi's sarcoma and one with Castleman's disease) were identified. As a control group, five effusions from HIV-seropositive patients without known HHV 8-related diseases were identified. Cytological examination of effusions in patients with HHV 8-related diseases demonstrated atypical/markedly reactive mesothelial cells accompanied by a polymorphous background of lymphocytes. Molecular studies for B- and T-cell clonality in microdissected whole samples showed no definitive clones in these cases. Conversely, polymerase chain reaction (PCR) studies for the HHV 8 virus was positive in these samples. PCR studies on pure populations of microdissected mesothelial cells from the HHV 8-related effusions were positive for HHV 8 sequences, whereas those from HIV patients with non-HHV 8 related diseases were negative. Immunohistochemistry for HHV 8 (monoclonal antibody to latent nuclear antigen (LNA-1; ORF-73) on cellblock material demonstrated scattered positive mesothelial cells in three of the five cases of HHV 8-associated effusions. HHV 8 has been recently implicated in the pathogenesis of Kaposi's sarcoma and primary effusion lymphoma. Mesothelial cells in recurrent pleural effusions from patients with Kaposi's sarcoma and Castleman's disease appear to be infected with HHV 8. Additional studies need to be done to define the role of mesothelial cell infection in the pathogenesis of these HHV 8-associated effusions and define the prognostic significance.     

Making the most of a teachable moment: a smokeless-tobacco cessation intervention in the dental office.

OBJECTIVES. Primary care medical clinics are good settings for smoking interventions. This study extends this strategy with a smokeless tobacco intervention delivered by dentists and dental hygienists in the course of routine dental care. METHODS. Male users of moist snuff and chewing tobacco (n = 518) were identified by questionnaire in clinic waiting rooms and then randomly assigned to either usual care or intervention. The intervention included a routine oral examination with special attention to the part of the mouth in which tobacco was kept and an explanation of the health risks of using smokeless tobacco. After receiving unequivocal advice to stop using tobacco, each patient viewed a 9-minute videotape, received a self-help manual, and was briefly counseled by the dental hygienist. RESULTS. Long-term success was defined as no smokeless tobacco use at both 3- and 12-month follow-ups, with those lost to follow-up counted as smokeless tobacco users. The intervention increased the proportion of patients who quit by about one half (12.5% vs 18.4%, P < .05). CONCLUSIONS. These results demonstrate the efficacy of a brief dental office intervention for the general population of smokeless tobacco users.     

A patient with Werner's syndrome and osteosarcoma presenting as scleroderma.

Werner's syndrome (WS) is often mistaken for scleroderma. We describe a patient with WS who presented with an enlarging, painful mass of the right knee that proved to be a juxtaarticular osteosarcoma of the distal femur. Recognition of WS and prompt investigation of any painful, enlarging masses to exclude sarcomatous degeneration will benefit these patients.     

FRANTZ'S TUMOUR: PAPILLARY AND CYSTIC CARCINOMA OF THE PANCREAS

Frantz's tumour (papillary and cystic tumour) of the pancreas is a rare neoplasm usually seen in young women. It is of low grade malignancy and deserves special note among pancreatic malignancies as it is frequently amenable to local resection and has a good long-term survival rate after excision. Three such cases have been treated at Westmead Hospital, one young male and two females. In two the disease was confined to the pancreas. In one, local invasion outside the pancreas and trans-coelomic spread to the ovaries was present at the time of diagnosis. Complete surgical removal of macroscopic disease was achieved in all three and all remain disease free between 2 and 4 years post-surgery. All have good exocrine and endocrine pancreatic function. These cases are discussed in detail. The need to be aware of this uncommon variant of pancreatic cancer is stressed. Investigation and treatment options are reviewed. The role of cytology studies in diagnosis and the potential for long-term surgical control of this tumour are highlighted on the basis of our limited experience and that presented in recent surgical literature.