Responses to ultraviolet-B in cell lines from hereditary melanoma kindreds.

Ultraviolet-B (UV-B) triggers a cascade of events involving cell cycle control genes leading ultimately to DNA repair or apoptosis. The hypothesis examined here is that the genetic abnormality predisposing to melanoma affects the ability of the cell to respond appropriately to UV-B, so favouring mutagenesis. Epstein-Barr virus-transformed lymphoblastoid cell lines from hereditary melanoma kindreds were irradiated with UV-B, and changes in p53, p21 and Bcl-2 expression and cell cycle phase distribution were analysed. Twenty-two cell lines were tested: 12 carriers of melanoma susceptibility and 10 non-carriers (unaffected first degree relatives). At 24 h after irradiation with 50 J/m2, 15 of the 22 cell lines showed a rise in G2/M. After 400 J/m2, all the cell lines showed a reduction or loss of G2/M and 17 of the 22 showed an S phase delay. More carriers than noncarriers of melanoma susceptibility showed significant S phase delay after 50 J/m2 (seven out of 12 carriers versus two out of 10 non-carriers). Six of the 10 pairs (carrier versus non-carrier) tested showed discordant cell cycle responses; however the nature of the difference was not universal. Bcl-2 reduction was seen 4 h post-irradiation in all the carriers and non-carriers. The p53 and p21 responses, although showing some individual variations, were not related to carrier status. These results show individual variations in response to UV-B irradiation among cell lines from the members of hereditary melanoma kindreds, but no consistent differences between carriers and non-carriers of melanoma susceptibility.

Normal repair of ultraviolet radiation-induced DNA damage in familial melanoma without CDKN2A or CDK4 gene mutation.

Excessive sun exposure and family history are strong risk factors for the development of cutaneous melanoma. Inherited susceptibility to this type of skin cancer could therefore result from constitutively impaired capacity to repair ultraviolet (UV)-induced DNA lesions. While a proportion of familial melanoma kindreds exhibit germline mutations in the cell cycle regulatory gene CDKN2A (p16INK4a) or its protein target, cyclin-dependent kinase 4 (CDK4), the biochemical basis of most familial melanoma is unknown. We have examined lymphoblastoid cell lines from melanoma-affected and unaffected individuals from large hereditary melanoma kindreds which are not attributable to CDKN2A or CDK4 gene mutation. These lines were tested for sensitivity of clonogenic growth to UV radiation and for their ability to repair transfected UV-damaged plasmid templates (host cell reactivation). Two of seven affected-unaffected pairs differed in colony survival after exposure to UVB radiation; however, no significant differences were observed in the host-cell reactivation assays. These results indicate that melanoma susceptibility genes other than CDKN2A and CDK4 do not impair net capacity to repair UV-induced DNA damage.

Multiple abnormalities of the p16INK4a-pRb regulatory pathway in cultured melanoma cells.

The retinoblastoma protein (pRb) pathway is critical in regulating the G1 phase of the cell cycle and it is frequently disrupted in human cancers. Components of the pRb pathway which are often altered in tumour progression include the INK4 cyclin-dependent kinase (CDK) inhibitors p16INK4a/ CDKN2A and p15INK4b/CDKN2B, CDK4, D-type cyclins and pRb. Several of these components were studied in a series of cultured melanoma cell lines in order to determine the frequency and spectrum of genetic alterations and to define targets for potential gene transfer studies. Also studied were the p16INK4a alternate transcript (p14ARF) and the p21(waf1) CDK inhibitor. The majority of the melanoma cell lines tested (13 out of 17; 76%) carried mutated (two), deleted (nine) or silenced (two) p16(INK4a). CDK4 was mutated or overexpressed in two melanoma cell lines with homozygously deleted CDKN2A and CDKN2B genes. This suggests that the selective growth advantages afforded by CDKN2A inactivation and CDK4 insensitivity are distinct and may involve the mediation of other CDK inhibitors or CDKs.

Src phosphorylates the insulin-like growth factor type I receptor on the autophosphorylation sites. Requirement for transformation by src.

The insulin-like growth factor type I (IGF-I) receptor can become tyrosine phosphorylated and enzymatically activated either in response to ligand or because of the activity of the Src tyrosine kinase (Peterson, J. E., Jelinek, T., Kaleko, M., Siddle, K., and Weber, M. J. (1994) J. Biol. Chem. 269, 27315-27321). The goal of the present study was to analyze the mechanistic basis and functional significance of the Src-induced phosphorylation and activation of the IGF-I receptor. 1) We mapped the sites of IGF-I receptor autophosphorylation to peptides representing three different receptor domains: tyrosines 943 and 950 in the juxtamembrane region; tyrosines 1131, 1135, and 1136 within the kinase domain; and tyrosine 1316 in the carboxyl-terminal domain. The juxtamembrane and kinase-domain peptides were phosphorylated both in vivo and in vitro. The carboxyl-terminal site, although phosphorylated in vitro and in src-transformed cells, was not a major site of ligand-induced phosphorylation in vivo. 2) We determined that the sites of Src-induced phosphorylation of the IGF-I receptor are the same as the ligand-induced autophosphorylation sites and that the Src kinase can catalyze these phosphorylations directly. 3) We showed that cells cultured from mice in which the IGF-I receptor has been knocked out by homologous recombination are defective for morphological transformation by src. Thus, the Src kinase can substitute for the receptor kinase in phosphorylating and activating the IGF-I receptor, and this receptor phosphorylation and activation are essential for transformation by src.

Bone presentation of non-Hodgkin's lymphoma: experience at the Royal North Shore Hospital, Sydney; highlighting primary bone lymphoma.

BACKGROUND: Primary lymphoma of bone (PLB) is a rare form of extranodal lymphoma. Between 1975 and 1992 39 patients with lymphoma presenting in bone were seen at the Royal North Shore Hospital (RNSH), Sydney. Of these, 12 (31%) had truly localised disease (Stage IE). AIMS: Patients were studied retrospectively to determine the prognostic significance of bony involvement per se versus involvement of a single bony site, and to determine the impact of treatment modality on outcome. METHODS: The 39 patients were divided into three groups according to extent of disease; single osseous site (Stage IE), multifocal bone, and bone plus visceral and/or nodal disease. Kaplan-Meier survival curves were constructed, and five year actuarial survival stated. Cox regression analysis was used to determine hazard ratios. Overall survival was used as the end-point. RESULTS: A trend for better survival was noted with Stage IE disease. Multifocal and disseminated disease appeared to have a poorer outcome when assessed by hazard ratio, with a value of 3 (95% CI 0.87-10.4; p = 0.08), compared to unifocal disease. Radiotherapy alone was as effective as combined modality treatment although patient numbers were too small for statistical confirmation. CONCLUSIONS: The stage of lymphoma, rather than bony involvement per se, seems to have more prognostic importance. Radiotherapy alone offered equivalent results to combined modality treatment in this series.

Predictive value of early electrophysiologic testing in determining long-term outcome with amiodarone treatment in patients with sustained ventricular tachycardia.

This prospective study sought to determine whether programmed ventricular stimulation before hospital dismissal in patients who had received a loading dose of amiodarone would identify those at risk for recurrent ventricular arrhythmias. Between January 1985 and January 1989, 64 patients (55 men and 9 women; mean age, 64 years) with a history of sustained ventricular tachycardia (VT) or ventricular fibrillation were referred to our institution for electrophysiologic testing. Of these patients, 52 had coronary artery disease, 11 had dilated cardiomyopathy, and 1 had hypertrophic cardiomyopathy. Of the 64 patients, 47 had baseline tests while no drugs were administered and repeated electrophysiologic testing after 10 days of amiodarone loading (1.2 g/day). The other 17 patients had no baseline study because of instability of their arrhythmias but underwent electrophysiologic testing after amiodarone loading. Follow-up ranged from 7 to 1,536 days (mean, 652 days). During the follow-up period, recurrent arrhythmias were detected in 22 patients. Of the 64 patients, 14 had suppression of VT. Of 43 patients in whom the cycle lengths of VT were determined both at baseline and after amiodarone therapy, 20 had an increase of 100 ms or more, and 23 had no substantial change. The mean ejection fraction was 31%. Of a total of 16 deaths in the series, 8 were sudden. Suppression of VT during amiodarone therapy suggested a lower rate of fatal and nonfatal recurrent arrhythmias, but the difference was not statistically significant. An increase in the cycle length of VT did not predict an improved outcome. The age of the patient and the presence of a left ventricular aneurysm were slightly predictive of mortality.(ABSTRACT TRUNCATED AT 250 WORDS)

Abnormal adrenal gland metabolism in opioid addicts: implications for clinical treatment.

Adrenal gland metabolism is markedly altered in heroin addicts. During daytime hours, the addict may suffer corticoid deficiency of the addisonian type, and in the evening, an excess of the cushingoid type. The high plasma levels of cortisol that are found in the evening in addicts antagonize endogenous opioids in a manner similar to naloxone. In the present study, 72% of the heroin addicts who sought treatment demonstrated reduced adrenal cortisol reserve. Effective immune and stress responses are dependent on adrenal cortisol reserve. This finding provides an explanation for the heroin addict's vulnerability to AIDS and other infectious diseases. One of methadone's greatest attributes is that it helps normalize adrenal metabolism. Clinical methods to at least partially correct adrenal metabolism may enhance current opioid addiction treatment modalities.

The National Institutes of Health: some critical years, 1955-1957.

It has been my pleasure to participate in the conversion of a small but superb federal institution into a driving force for the development of excellence in the nation's biomedical sciences. The initial step was the establishment of an adequate science base for the developing enterprise. Given this, it was agreed that the nation's medical establishment could use the evolving support system effectively. What follows is a brief consideration of some events contributing to the reduction of a possibility to reality. Much of the material that follows was derived from a presentation to a presidential commission established by President Gerald Ford in 1975 and later published as a supplement to the Journal of Medical Education. The latter encompassed what happened during the critical years of development. But it seemed too recent at that time to discuss with grace the "how" of program changes. It is the "how of things" that will be treated in this article.

Detoxification from methadone maintenance: double-blind comparison of two methods.

A well-rehabilitated group of 22 methadone maintenance patients were detoxified under ambulatory and double-blind conditions, over 42 days, either by decreasing the daily dose of methadone and supplementing it by propoxyphene napsylate (M--PN), or by decreasing the daily dose of methadone administered in two equal doses given 12 hours apart (M--M). All of 12 (100%) M--M and 5 of 10 (50%) M--PN subjects were able to temporarily withdraw from methadone maintenance, although two of the M--PN patients required maintenance with propoxyphene napsylate to cease methadone maintenance. In our experience both detoxification methods employed appear superior to the standard withdrawal technique of simply reducing the daily methadone maintenance dose in a step-wise fashion.

Medicinal use of hypodermic needles in an addict patient population.

Narcotic addicts may become so conditioned to the use of hypodermic needles that self-injection without a drug may become pleasurable. This phenomenon has raised the question as to whether addicts in treatment should be administered any medication by hypodermic needle for fear of precipitating withdrawal or inducing a craving for heroin. In this study addicts in treatment were given a medicinal needle challenge but there were no documented deleterious effects.