BACKGROUND: Genetically modified T-lymphocytes are potential therapeutic agents for the treatment of various disorders. Successful retroviral infection of primary murine T-lymphocytes is a prerequisite to the study of adoptive cell therapies in a small animal model. The definition of factors controlling retroviral infection of T-lymphocytes would also be useful to better understand retroviral diseases. However, retroviral-mediated gene transfer into murine primary T-cells has remained elusive. METHODS: In order to define the requirements for stable and efficient gene transfer in primary murine T-lymphocytes, we investigated factors capable of affecting retroviral infection. These include activation conditions (using mitogens or monoclonal antibodies), culture conditions (including media composition and cytokine addition), timing of viral exposure, retroviral receptor selection (ecotropic, amphotropic, or vesicular stomatitis virus G (VSV-G) glycoprotein receptor), and viral titer. RESULTS: We show that efficient gene transfer can be achieved in murine T-lymphocytes, provided that a number of favorable conditions are met, in particular optimized T-cell activation conditions, optimal timing of infection, adequate interleukin-2 concentration and T-cell density, and a high viral titer. On a particulate basis, we find that ecotropic particles are more effective than amphotropic or VSV-G-pseudotyped particles, and recommend the use of a specifically selected retroviral packaging cell line. CD4+ T-cells are equally or more infectible than CD8+ lymphocytes, depending on the activation conditions. The Th1 and Th2 subsets are comparably susceptible to retroviral infection, in contrast to what has been reported in some instances of human T-cell infection by human immunodeficiency virus (HIV). CONCLUSIONS: We establish conditions that enable efficient retroviral-mediated gene transfer in murine primary T-lymphocytes. T-lymphocytes are not uniformly susceptible to retroviral infection depending on the mode of T-cell activation. These findings have implications for devising approaches to the study of T-cell biology, adoptive cell therapies, and the pathophysiology of retroviral diseases in mouse models.

Animals↗

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Multifactorial analysis of survival in primary extremity liposarcoma.

The treatment of extremity soft tissue sarcoma is now directed at limb preservation with the addition of various adjuvant therapies to improve treatment results. To achieve this goal, a knowledge of prognostic factors for extremity soft tissue sarcoma becomes increasingly critical. The object of this study was to analyze prognostic factors for survival in patients with extremity liposarcoma. Eighty-three patients with primary localized extremity liposarcoma, admitted from 1968 to 1978, were retrospectively reviewed. Surgical resection was the primary mode of treatment. Eleven prognostic factors were analyzed. Tumor factors included: histologic subtype, tumor grade, size, depth, invasion of vital structures, and site; operative factors included: type of operation, and surgical margins; and patient factors included: symptoms, age at diagnosis, and sex. Kaplan-Meier survival curves, and univariate and stratified log-rank tests of association were performed. Independent factors for predicting survival were identified using the Cox model stepwise regression technique. In univariate analysis of the entire group of patients, 5 factors were significant for tumor mortality: tumor grade (p = 0.00005), histologic subtype (p = 0.00025), tumor size greater than or equal to 5 cm (p = 0.005), type of surgery/margin (p = 0.0001), and invasion of vital structures (p = 0.008). When associations among all factors were analyzed, it was found that many factors were interdependent. The independent prognosticators were, therefore, determined according to the Cox model technique. For the multivariate analysis, well-differentiated and lipoblastic liposarcomas were excluded because of lack of survival variation within each group. The former group had no tumor-related deaths and the latter group showed 80% tumor mortality.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance↗

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Evaluation of four warming procedures to minimise heat loss induced by anaesthesia and surgery in dogs.

OBJECTIVE: To evaluate the efficiency of four warming procedures, introduced after anaesthetic induction and continued during surgery, in minimising heat loss in anaesthetised dogs. DESIGN: Dogs were paired. One of each pair was a control; the other was subjected to one of four warming procedures. METHODS: Ninety-six dogs were involved in total. Pairs of dogs were matched for breed, hair length, and type of surgical procedure and placed adjacent to each other in a large temperature-controlled surgical theatre. One dog within each pair was assigned to one of four warming procedures that commenced immediately after anaesthetic induction. Group 1 (11 pairs) were placed on a purpose-designed prewarmed (41 degrees C) electrically heated pad. Group 2 (18 pairs) were placed on a prewarmed electric heat pad (41 degrees C), cocooned by four wrapped water bottles (initially 41 degrees C) and subjected to radiant heat (150 watt lamp placed 50 cm away from the head of the dog). Group 3 (11 pairs) were surrounded by a forced air warming mattress (set at 43 degrees C). Group 4 (8 pairs) were connected via the anaesthetic breathing circuit to a heater/humidifier (set at 41 degrees C). Rectal temperature measurements were recorded every 15 min for the first 3 h of anaesthesia. The fall in rectal temperature of the control dog was subtracted from the fall in temperature of the treatment dog and this measurement was used to assess the efficacy of the various warming procedures. RESULTS: The mean rectal temperature of unheated 'control' dogs decreased 1.9 +/- 0.6, 1.4 +/- 0.4 and 1.1 +/- 0.4 degrees C over the first, second and third hour respectively. After 3 h the temperature fall differential for all groups were 0.7 +/- 0.7 (Group 1), 3.1 +/- 1.1 (Group 2), 2.4 +/- 1.1 (Group 3) and 1.0 +/- 1.1 degrees C (Group 4). Thus the group 2 procedure was the most successful in preventing a drop of temperature followed by groups 3, 4 and 1. CONCLUSION: Large dogs undergo significant reduction in core body temperature especially during the first 2 h of anaesthesia and surgery. Supplementary warming substantially reduces this fall in body temperature, although certain warming procedures were found to be more effective than others.

Anesthesia, General↗

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Clinical findings of West Nile virus infection in hospitalized patients, New York and New Jersey, 2000.

Outbreaks of West Nile (WN) virus occurred in the New York metropolitan area in 1999 and 2000. Nineteen patients diagnosed with WN infection were hospitalized in New York and New Jersey in 2000 and were included in this review. Eleven patients had encephalitis or meningoencephalitis, and eight had meningitis alone. Ages of patients ranged from 36 to 87 years (median 63 years). Fever and neurologic and gastrointestinal symptoms predominated. Severe muscle weakness on neurologic examination was found in three patients. Age was associated with disease severity. Hospitalized cases and deaths were lower in 2000 than in 1999, although the case-fatality rate was unchanged. Clinicians in the Northeast should maintain a high level of suspicion during the summer when evaluating older patients with febrile illnesses and neurologic symptoms, especially if associated with gastrointestinal complaints or muscle weakness.

Adult↗

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Coronary artery bypass grafting in patients with left main coronary artery stenosis. Improved survival in a predominantly Oriental population.

Over the last 10 years, there has been a reduction of surgical mortality in coronary bypass grafting for left main coronary artery disease. Most of the available literature reports are of Caucasian populations. We report our experience of the surgical treatment of this disease in a predominantly Oriental population. From 1 January 1987 to 30 June 1991, 86 consecutive patients had coronary artery bypass surgery for left main stenosis. There were 65 men and 21 women with an age range of 34 to 80 years (mean age: 58 years SD +/- 8.63). There were no operative deaths and 1 hospital death (1.2%) from a non-cardiac cause. The perioperative myocardial infarction rate was 3.5% (3/86). These results are better than some earlier series in which hospital mortality was reported as high as 12.6% and the preoperative myocardial infarction rate 11.8%. In this series where the patients are predominantly Oriental, the results are comparable if not better than in series reported in centres where the population is predominantly Caucasian.

Age Factors↗