Role of p21 RAS in p210 bcr-abl transformation of murine myeloid cells

The p21 RAS product has been implicated as part of the downstream signaling of certain nonreceptor tyrosine kinase oncogenes and several growth factor receptor-ligand interactions. We have reported that the chronic myelogenous leukemia oncogene p210 bcr-abl transforms a growth- factor-dependent myeloid cell line NFS/N1.H7 to interleukin-3 (IL-3) independence. In these p210 bcr-abl-transformed cells (H7 bcr-abl.A54) and in two other murine myeloid cell lines transformed to IL-3 independence by p210 bcr-abl, endogenous p21 RAS is activated as determined by an elevated ratio of associated guanosine triphosphate (GTP)/guanosine diphosphate (GDP), assayed by thin-layer chromatography of the nucleotides eluted from p21 RAS after immunoprecipitation with the Y13-259 antibody. Treatment of p210 bcr-abl-transformed cells with a specific tyrosine kinase inhibitor herbimycin A resulted in diminished tyrosine phosphorylation of p210 bcr-abl and associated proteins, without major reduction in expression of the p210 bcr-abl protein itself. Inhibition of p210 bcr-abl-dependent tyrosine phosphorylation resulted in a reduction of active p21RAS-GTP complexes in the transformed cells, in diminished expression of the nuclear early response genes c-jun and c-fos, and in lower cellular proliferation rate. To further implicate p21 RAS in these functional events downstream of p210 bcr-abl tyrosine phosphorylation, we targeted G- protein function directly by limiting the availability of GTP with the inosine monophosphate dehydrogenase inhibitor, tiazofurin (TR). In p210 bcr-abl-transformed cells treated for 4 hours with TR, in which the levels of GTP were reduced by 50%, but GDP, guanosine monophosphate, and adenosine triphosphate (ATP) were unaffected, p210 bcr-abl tyrosine phosphorylation was at control levels. However, expression of c-fos and c-jun nuclear proto-oncogenes were strongly inhibited and p21 RAS activity was downregulated. These findings show that p210 bcr-abl transduces proliferative signals, in part, through downstream activation of p21 RAS. Furthermore, p21 RAS activity is linked to pathways that regulate c-jun and c-fos expression.     

Activation of apoptosis associated with enforced myc expression in myeloid progenitor cells is dominant to the suppression of apoptosis by interleukin-3 or erythropoietin

The inappropriate expression of c-myc in cells deprived of growth factors has recently been implicated in the activation of programmed cell death (apoptosis). The studies described here examine the ability of interleukin-3 (IL-3) or erythropoietin (Epo) to suppress apoptosis that occurs in association with enforced myc expression during cell cycle arrest of a murine IL-3-dependent myeloid progenitor cell line, 32D. G1 arrest was observed when culturing 32D cells to high density in medium supplemented with IL-3, or at subconfluent densities in medium supplemented with Epo. Under both conditions, endogenous c-myc expression was downregulated and viability was maintained. In clones of cells in which c-myc is constitutively expressed from a retroviral vector, enforced c-myc expression was associated with the activation of apoptosis at high cell densities. Similarly, enforced c-myc expression was deleterious to cell survival when these cells were cultured in Epo, as apoptosis was evident within 6 hours. The results support the concept that inappropriate c-myc expression activates apoptosis and that neither IL-3 nor Epo can suppress this program under these conditions.     

Accumulation of methotrexate and methotrexate polyglutamates in lymphoblasts at diagnosis of childhood acute lymphoblastic leukemia: a pilot prognostic factor analysis

Lymphoblasts in bone marrow samples, obtained from 43 children with acute lymphoblastic leukemia at diagnosis, were incubated with 1.0 mumols/L [3H] methotrexate for 24 hours in vitro. Nonexchangeable methotrexate and methotrexate polyglutamates were separated and quantitated. Event-free survival at 5 years was 38% +/- 9% for all 43 patients (27 failures), and 44% +/- 10% for the 35 with non-T, non-B- cell acute lymphoblastic leukemia (20 failures). Of these 35 children, those whose lymphoblasts accumulated more than 100 pmol methotrexate and 500 pmol methotrexate polyglutamates per billion cells experienced better 5-year event-free survival than those whose lymphoblasts did not (65% +/- 12% v 22% +/- 9%, P = .010). This difference characterized "good-risk" patients who were female (P = .014), less than age 7 at diagnosis (P = .005), or had low initial white blood cell counts (less than 20 X 10(9)/L, P = .018). Findings were similar for the 43 children with acute lymphoblastic leukemia and for the "good-risk" children in this total group. Thus, the ability of lymphoblasts to accumulate methotrexate and form methotrexate polyglutamates may be important to the curative properties of current therapy of acute lymphoblastic leukemia in children, particularly for "good-risk" patients. In such patients, inherent rather than acquired drug resistance may be the initial event leading to treatment failure.     

Bone marrow transplantation for patients with Philadelphia chromosome- positive acute lymphoblastic leukemia

We report the treatment outcome of allogeneic bone marrow transplantation in ten patients with Philadelphia chromosome-positive acute lymphoblastic leukemia. Six patients are alive and well for 6 to 30 months (median 19 months) after transplantation. Four patients died with transplant related complications. In view of the poor prognosis associated with this disease, marrow ablation followed by allogeneic or syngeneic marrow grafting may be the preferred treatment modality if a suitable marrow donor is available.     

Fractionated total-body irradiation and high-dose etoposide as a preparatory regimen for bone marrow transplantation for 94 patients with chronic myelogenous leukemia in chronic phase

Ninety-four consecutive patients with chronic myelogenous leukemia in first clinical chronic phase, median age of 34.0 years (range, 6.8 to 52.4 years), with a histocompatible sibling donor, were treated with fractionated total body irradiation (1,320 cGy) and high-dose etoposide (60 mg/kg) followed by allogeneic bone marrow transplantation (BMT). The median time from diagnosis to BMT was 7.0 months (range, 2.3 to 72.0 months). Sixty patients were treated before BMT with hydroxyurea alone, four patients with busulfan alone, one patient with interferon alone, and the other 29 patients were treated with various combinations of these drugs. Cumulative probabilities of overall survival, event- free survival, and relapse at 5 years were 73%, 64%, and 14%, respectively. The median follow-up time for surviving patients was 38 months, ranging from 12 to 88 months. By stepwise Cox regression analysis, significant prognostic variables were age at transplant, acute graft-versus-host disease > or = grade II, cytomegalovirus- associated interstitial pneumonitis, and years from diagnosis to BMT.     

Central regulation of intestinal basal and stimulated water and ion transport by endogenous opiates in dogs

The influence of intracerebroventricular (ICV) vs intravenous (IV) administration of (D-Ala2, Met5) enkephalinamide (Dalamide) on normal and stimulated (cholera toxin) jejunal fluxes of water, Na+, and K+ were investigated in dogs prepared with a Thiry-Vella (TV) loop. Intestinal transport in the TV loop and concomitant transit time were measured during an infusion (2 ml/min) of an isotonic electrolyte solution alone, or containing 0.4 micrograms/ml of cholera toxin (CT). Basal net water absorption was slightly, but significantly (P less than 0.05), increased during an ICV infusion of Dalamide at 0.5 ng/kg/min, while the secretory effects of cholera toxin were markedly reduced by nearly 75%. Similar effects were observed for Na+ and K+ movement. In contrast, Dalamide infused intravenously at a five times higher dose, ie, 2.5 ng/kg/min did not affect the control and CT-stimulated water and electrolyte movements. The jejunal loop transit times were halved during CT infusion. Similar values were observed under Dalamide ICV administration as well as during a five times higher dose of Dalamide administered intravenously. It was concluded that (1) Dalamide administered into the CNS, but not peripherally, increased the absorption of water, Na+, and K+, causing a net reduction in their secretion induced by cholera toxin; and (2) these effects did not result from changes in transit time. These results also suggest that Met-enkephalin can act in the brain to affect the intestinal transport of water and electrolytes in dogs.     

Lipomatous tumors of the liver: evaluation with CT and US

Seven cases of lipomatous masses within the liver parenchyma were demonstrated with computed tomography (CT). Five of these cases were obtained from a retrospective review of 50 cases of renal angiomyolipoma in which the liver was adequately demonstrated. The other two cases were from the files of the Armed Forces Institute of Pathology and had no associated renal lesions. Three of the five cases were associated with tuberous sclerosis. In all seven cases, the fatty tumors appeared on CT scans as a well-defined, 0.8-13-cm mass, with attenuation coefficients of less than -30 HU. On ultrasound studies, the lesions were well circumscribed, highly echogenic, and similar to hemangiomas. While distinctly rare lesions, these lipomatous masses are not as unusual as the literature would indicate. One may anticipate such masses in patients with renal angiomyolipomas and in a relatively high percentage of those with tuberous sclerosis.     

Integration of local inputs in visual cortex

In mammalian visual cortex, local connections are ubiquitous, extensively linking adjacent neurons of all types. In this study, optical maps of intrinsic signals and responses from single neurons were obtained from the same region of cat visual cortex while the effectiveness of the local cortical circuitry was altered by focally disinhibiting neurons within a column of known orientation preference. Maps of intrinsic signals indicated that local connections provide strong and functional subthreshold inputs to neighboring columns of other orientation preferences, altering the observed orientation preference to that of the disinhibited column. However, measuring the suprathreshold response using single-cell recordings revealed only mild changes of preferred orientation over the affected region. Because strongly tuned subthreshold inputs from cortex only marginally affect the tuning of a cortical cell's output, it is concluded that local cortical inputs are integrated weakly compared to geniculate inputs. Such circuitry potentially allows for the normalization of responses across a wide range of input activity through local averaging.      

Incidence of injury in elite junior Rugby Union: a prospective descriptive study

The high incidence of injury in Rugby Union is well documented, particularly at elite levels of competition. This article describes the incidence and nature of all injuries sustained by elite Western Australian junior Rugby Union players during the 26 weeks up to and including the 1997 National Championship campaign. Informed consent was gained for each participant (n = 44) prior to completion of an extensive baseline questionnaire. Exposure and injury data were collected at each training session and game. The injury incidence rate over the 26 week period was 13.26/1000 player hours. Injury data were analysed by phase of play, position, severity and if occurred at games or training. The incidence of injury was significantly associated with the position played (chi2 = 67.49, p value = 0.008) and the phase of play in which the injury occurred (chi2 = 8.07, p value = 0.042). Tackling was the most dangerous phase of play (52% of injuries) and the most common site of injury was the lower limb (37%). Most injuries occurred during games (56%) and the flanker was the position most at risk of injury (12%). Further research is needed to identify the aetiology of injury at all levels of competition and to use these findings to develop effective injury prevention strategies in this sport. Position-specific risk factors should also be investigated, as should the mechanism of injury associated with tackling which is the phase of play in which significantly more injuries occur in rugby.     

Prevention of transfusion-associated graft-versus-host disease: selection of an adequate dose of gamma radiation

To determine the optimal dose of gamma radiation necessary to inhibit T- lymphocyte function and prevent transfusion-acquired graft-versus-host disease (TA-GVHD), a donor plateletpheresis component was initially divided into ten 20-mL samples. One sample was not irradiated, while the other nine samples were treated with gamma radiation at doses ranging from 500 to 4500 cGy. T-lymphocyte function was subsequently measured by mixed lymphocyte cultures and mitogen stimulation assays. The results were assessed in each test by calculating the percentage of inhibition of each irradiated sample as compared to that of the unirradiated sample. The accuracy of the delivered dose of gamma radiation was measured with thermoluminescent dosimeters. It was concluded that a nominal dose of 3000 cGy (actual dose delivered, 2898 cGy) is the appropriate amount of gamma radiation needed to eliminate T- lymphocyte-mediated graft-versus-host disease.