Efficient and faithful in vitro translation of natural and synthetic mRNA with human ribosomes

Efficient systems for in vitro translation are of importance for biochemical and gene expression studies as well as for biotechnological developments. We optimized a cell-free translation system using subcellular fractions from human placenta and high quality placental tRNAs isolated using a simple and fast procedure. The postmitochondrial fraction or a reconstituted system containing soluble proteins plus polysomes were able to efficiently translate endogenous and exogenous mRNAs. Optima for ions, enzymes, tRNA and energy mix components were determined for a poly(U)-directed poly(Phe) synthesis test. The use of homologous tRNAPhe, omission of commercial creatine kinase, and addition of 3.5 mM spermidine at near physiological magnesium concentration (2.5 mM), were the most significant improvements. Under optimal conditions, poly(Phe) synthesis proceeded at a maximal initial rate of 1.2 Phe/80S/min at 37 degrees C, while natural mRNA translation by S-30 started at a near in vivo estimated rate of 0.3-0.5 amino acid/80S/sec. Furthermore, natural mRNA directed the synthesis of a family of polypeptides closely resembling the pattern of cytoplasmic proteins in both, molecular weight and relative amounts. This efficient and faithful system is of interest for biochemical studies of the human translational machinery, as well as a basis for screening new drugs affecting protein synthesis in pathogenic microorganisms.     

Human disorganization complex, as a polytopic blastogenesis defect: a new case

We describe a baby girl of 4,000 g and 55 cm with supernumerary, malformed, and partially duplicated lower limbs, malformed and partially duplicated pelvis, spina bifida, coccygeal dermal sinus, ectopic anus located in the right buttock, duplicated internal genitalia, rectovaginal fistula, ileal atresia, Meckel diverticulum, and various renal system anomalies. We think that this phenotype is a new case of disorganization in humans (DsH) and postulate that this condition constitutes a polytopic defect of the blastogenesis. In this case, the presence of a malformation pattern involving structures in different parts of the body and organs derived from all of the germ layers, suggests that the pathogenetic event most probably occurred during blastogenesis affecting various progenitors fields.     

Response rates to a questionnaire 26 years after baseline examination with minimal interim participant contact and baseline differences between respondents and nonrespondents

Research on response rates to surveys mailed to study participants decades after baseline examination, with minimal interim contact, is limited. This paper documents response rates to a 26-year follow-up survey of surviving participants from a large cohort in Illinois and compares baseline characteristics of nonrespondents and respondents. Mortality follow-up of the Chicago Heart Association Detection Project in Industry 1967-1973 cohort involved minimal or no participant contact since baseline. In 1996, a 26-year follow-up questionnaire was mailed to all surviving participants aged 65 years or older. Current addresses were obtained from the Health Care Financing Administration for 96.5 percent of 12,409 participants in our analyses. Total response rates were 59.8 percent and, for participants for whom Health Care Financing Administration addresses were available, 60.8 percent. A higher response rate was obtained for younger recipients, men, Whites, more-educated persons, nonsmokers, and those with a better cardiovascular risk profile at baseline. A graded negative relation was found between number of cardiovascular risk factors at baseline and response rates obtained in 1996. Use of Health Care Financing Administration records as an additional follow-up method and factors that influence response rates are discussed. In conclusion, long-term follow-up of older surviving participants is feasible if current addresses can be obtained from standardized sources.     

Intensive chemotherapy with and without cranial radiation for Burkitt leukemia and lymphoma: final results of Cancer and Leukemia Group B Study 9251

BACKGROUND: The objective of the current study was to evaluate the efficacy of intensive chemotherapy with and without cranial radiation for central nervous system (CNS) prophylaxis in adults with Burkitt leukemia or lymphoma. METHODS: Patients received 18 weeks of therapy. Prophylactic cranial radiation (2400 centigrays) and 12 doses of triple intrathecal chemotherapy were administered to the first cohort of patients. A subsequent cohort received the same therapy, with the exceptions that intrathecal therapy was reduced to six doses and radiotherapy was administered only to high-risk individuals. RESULTS: The median follow-up durations were 6.8 years in Cohort 1 and 4.1 years in Cohort 2. Three occurrences of transverse myelitis, 2 severe neuropathies, 3 cases of aphasia, and 1 case of blindness were documented in the first cohort of 52 patients (Cohort 1). In the subsequent cohort of 40 patients (Cohort 2), none of these occurrences were observed, and patients experienced less neurologic toxicity overall (61% vs. 26%; P=0.001). Responses were similar, and the 3-year event-free survival rate was 0.52 (95% confidence interval, 0.38-0.65) for Cohort 1 and 0.45 (0.29-0.60) for Cohort 2. CONCLUSIONS: Intensive, short-duration chemotherapy with less intensive CNS prophylaxis led to control at this sanctuary site with little neurotoxicity and may be curative for adults with Burkitt leukemia or lymphoma.     

Type I interferon and TNFalpha cooperate with type II interferon for TRAIL induction and triggering of apoptosis in SK-N-MC EWING tumor cells

Ewing's sarcoma is the second most common human bone tumor in childhood. Here, we investigated the sensitivity of the Ewing tumor cell line, SK-N-MC, to the apoptotic effect of type I (IFNalpha) and type II (IFNgamma) interferons and TNFalpha. We demonstrate that although IFNalpha and TNFalpha alone are unable to induce cell death, they act in synergy with IFNgamma to induce SK-N-MC cell apoptosis. The synergistic induction of apoptosis correlated with the synergistic induction of TNFalpha-related apoptosis-inducing ligand (TRAIL) mRNA and TRAIL protein synthesis as well as of TRAIL secretion. Preparations of inducer-free supernatants from SK-N-MC cells stimulated with combinations of cytokines were shown to be cytotoxic for untreated SK-N-MC cells. This cytotoxicity was partially inhibited by addition of TRAILR2/Fc fusion protein, indicating that the secreted TRAIL mediates, at least in part, the apoptotic effect displayed by the supernatants of stimulated SK-N-MC cells. We have shown that the presence of IFNgamma is required to allow the sustained expression of IRF1 in SK-N-MC cells stimulated by addition of IFNalpha or TNFalpha suggesting that IRF1 plays a role in the synergistic induction of apoptosis by combinations of cytokines. Furthermore, we have shown that inhibition of NF-kappaB activation contributes to the IFNgamma-mediated sensitization to the apoptotic effect of TNFalpha. To our knowledge, this is the first report showing that interferon/cytokine combinations are able to induce TRAIL gene expression and TRAIL protein synthesis and secretion in Ewing sarcoma-derived cells. We believe that the observations reported here might contribute to the development of alternative new approaches to the treatment of Ewing tumors resistant to conventional therapy.     

Retrospective review of modafinil use for cerebral palsy

A retrospective review was undertaken at Texas Tech University Health Sciences Center regarding the use of modafinil for the treatment of spasticity associated with cerebral palsy. Neurology clinic records were reviewed from January 1, 2000, until October 1, 2001. Thirty pediatric patients with cerebral palsy were identified who were treated empirically with modafinil during this time period. Twenty-three (76%) patients reported diminished spasticity with treatment, which was confirmed by physical examination; these patients had improved joint mobility. Seventeen (56%) patients continued treatment with modafinil by the end of the formal review period (September 30, 2001). Twenty-three percent (seven) of the patients stopped taking modafinil during the study owing to one of the following: decreased sleep time (four), decreased appetite (one), hyperactivity, and irritability (two). Thus, this retrospective review shows a reduction in spasticity from cerebral palsy, with only minor and reversible side effects noted from modafinil. A blinded, crossover study using modafinil for spastic cerebral palsy is planned.     

Neuropsychiatry: a management model for academic medicine

Neuropsychiatry has become the subject of a number of editorials and position papers in recent years. Historical, philosophical, clinical, scientific, and educational dimensions of neuropsychiatry have been discussed in these papers. The potential business aspects of this topic, however, have received little, if any, comment. In this paper, the authors describe the business performance characteristics of an integrated neuropsychiatry department, formed through the merger of two traditional departments of psychiatry and neurology. The merger of neurology and psychiatry to create an integrated neuropsychiatry department according to the model described produced significant improvement in financial performance.