Oncogenic K-RAS subverts the antiapoptotic role of N-RAS and alters modulation of the N-RAS:gelsolin complex.

Activating mutations in members of the RAS family of genes are among the most common genetic events in human tumorigenesis. Once thought to be functionally interchangeable, it is increasingly recognized that the classical members of this protein family (H-RAS, N-RAS and K-RAS4B) exhibit unique and shared functions that are highly context-dependent. Herein, we demonstrate that the presence of an oncogenic KRAS allele results in elevated levels of GTP-bound N-RAS (N-RAS.GTP) in two human colorectal cancer cell lines, HCT 116 and DLD-1, compared to their isogenic counterparts in which the mutant KRAS allele has been disrupted by homologous recombination. N-RAS subserves an antiapoptotic role in cells expressing wild-type K-RAS; this function is compromised, however, by the presence of mutant K-RAS, and these cells display increased sensitivity to apoptotic stimuli. We additionally identify a physical interaction between N-RAS and gelsolin, a factor that has been shown to promote survival and show that the N-RAS:gelsolin complex is modulated differently in wild-type and mutant K-RAS environments following apoptotic challenge. These findings represent the first biochemical evidence of a functional relationship between endogenous RAS proteins and identify a dynamic physical interaction between endogenous N-RAS and gelsolin that correlates with survival.

Teaching children with autism to ask questions about hidden objects.

We taught 2 4-year-old children with autism to ask questions of an adult who held a closed box with a toy inside. The treatment package (modeling, prompting, and reinforcement) was evaluated with a multiple baseline design across the three question forms during training, generalization, and follow-up evaluations. The first question form ("What's that?") produced the name of the hidden item. The second form ("Can I see it?") produced sight of it, and the third form ("Can I have it?") produced the item itself. Both children learned to ask questions about hidden objects.

Crystal structures of dialkylglycine decarboxylase inhibitor complexes.

The crystal structures of four inhibitor complexes of dialkylglycine decarboxylase are reported. The enzyme does not undergo a domain closure, as does aspartate aminotransferase, upon inhibitor binding. Two active-site conformations have been observed in previous structures that differ in alkali metal ion content, and two active-site conformations have been shown to coexist in solution when a single type of metal ion is present. There is no indication of coexisting conformers in the structures reported here or in the previously reported structures, and the observed conformation is that expected based on the presence of potassium in the enzyme. Thus, although two active-site conformations coexist in solution, a single conformation, corresponding to the more active enzyme, predominates in the crystal. The structure of 1-aminocyclopropane-1-carboxylate bound in the active site shows the aldimine double bond to the pyridoxal phosphate cofactor to be fully out of the plane of the coenzyme ring, whereas the Calpha-CO2(-) bond lies close to it. This provides an explanation for the observed lack of decarboxylation reactivity with this amino acid. The carboxylate groups of both 1-aminocyclopropane-1-carboxylate and 5'-phosphopyridoxyl-2-methylalanine interact with Ser215 and Arg406 as previously proposed. This demonstrates structurally that alternative binding modes, which constitute substrate inhibition, occur in the decarboxylation half-reaction. The structures of d and l-cycloserine bound to the active-site show that the l-isomer is deprotonated at C(alpha), presumably by Lys272, while the d-isomer is not. This difference explains the approximately 3000-fold greater potency of the l versus the d-isomer as a competitive inhibitor of dialkylglycine decarboxylase.

The impact of "managed care" on the practice of psychological testing: preliminary findings.

Although the impact of managed care constraints on assessment practices has received recent attention, a review of the literature found no data-based articles that address this issue. We report survey data on 137 members of the National Register of Health Service Providers in Psychology (Council for the National Register of Health Service Providers in Psychology, 1996) on current testing practices. The majority (72%) reported that their use of tests has changed in the last 5 years due to managed care directives. These clinicians are doing less testing overall and restrict their pool of assessment instruments. The Rorschach inkblot technique (Rorschach, 1942), the Thematic Apperception Test (Murray, 1943), and the Wechsler Intelligence scales (Matarazzo, 1972) were the instruments most noted for disuse. Apparently, practitioners are relying more on short, brief self-report measures that tap targeted symptoms or problem areas, and less on tests that demand considerable clinicians' time. Implications and limitations of the findings are discussed.

Resistance exercise prevents glucocorticoid-induced myopathy in heart transplant recipients.

PURPOSE: To determine the effect of resistance exercise training (ET) on glucocorticoid-induced myopathy in heart transplant recipients (HTR), 14 male HTR were randomly assigned to a ET group that trained for 6 months (54 +/- 3 yr old; mean +/- SD) or a control group (51 +/- 8 yr old; mean +/- SD). METHODS: Fat mass, fat-free mass, and total body mass were measured by dual-energy x-ray absorptiometry before and 2 months after transplantation (Tx), and after 3 and 6 months of ET or control period. The exercise regimen consisted of lumbar extension (MedX) performed 1 d.wk-1 and variable resistance exercises (Nautilus) performed 2 d.wk-1. PreTx body composition did not differ between groups. RESULTS: At 2 months after Tx, fat-free mass was significantly decreased below baseline in both control (-3.4 +/- 2.1%) and ET groups (-4.3 +/- 2.4%). Fat mass was significantly increased at 2 months after Tx in both the control (+8.3 +/- 2.8%) and ET groups (+7.3 +/- 4.0%). Six months of ET restored fat-free mass to levels 3.9 +/- 2.1% greater (P < or = 0.05) than before Tx. Fat-free mass of the control group decreased progressively to levels that were 7 +/- 4.4% lower than preTx values (P < or = 0.05). Both groups increased knee extension, chest press, and lumbar extensor strength, but improvements in the ET group were four- to six-fold greater (P < or = 0.05). CONCLUSION: Our results demonstrate that glucocorticoid-induced changes in body composition in HTR occur early after Tx. However, 6 months of specific ET restores fat-free mass to levels greater than before Tx and dramatically increases skeletal muscle strength. Resistance exercise, as part of a strategy to prevent steroid-induced myopathy, appears to be safe and should be initiated early after heart Tx.

Resistance exercise training restores bone mineral density in heart transplant recipients.

OBJECTIVES: This was a prospective, randomized, controlled study designed to determine the effect of resistance exercise training on bone metabolism in heart transplant recipients. BACKGROUND: Osteoporosis frequently complicates heart transplantation. No preventative strategy is generally accepted for glucocorticoid-induced bone loss. METHODS: Sixteen male heart transplant recipients were randomly assigned to a resistance exercise group that trained for 6 months (mean [+/- SD] age 56 +/- 6 years) or a control group (mean age 52 +/- 10 years) that did not perform resistance exercise. Bone mineral density (BMD) of the total body, femur neck and lumbar spine (L2 to L3) was measured by dual-energy X-ray absorptiometry before and 2 months after transplantation and after 3 and 6 months of resistance exercise or a control period. The exercise regimen consisted of lumbar extension exercise (MedX) performed 1 day/week and variable resistance exercises (Nautilus) performed 2 days/week. Each exercise consisted of one set of 10 to 15 repetitions performed to volitional fatigue. RESULTS: Pretransplantation baseline values for regional BMD did not differ in the control and training groups. Bone mineral density of the total body, femur neck and lumbar vertebra (L2 to L3) were significantly decreased below baseline at 2 months after transplantation in both the control (-3.3 +/- 1.3%, -4.5 +/- 2.8%, -12.7 +/- 3.2%, -14.8 +/- 3.1%, respectively). Six months of resistance exercise restored BMD of the whole body, femur neck and lumbar vertebra to within 1%, 1.9% and 3.6% of pretransplantation levels, respectively. Bone mineral density of the control group remained unchanged from the 2-month posttransplantation levels. CONCLUSIONS: Within 2 months after heart transplantation, approximately 3% of whole-body BMD is lost, mostly due to decreases in trabecular bone (-12% to -15% of lumbar vertebra). Six months of resistance exercise, consisting of low back exercise that isolates the lumbar spine and a regimen of variable resistance exercises, restores BMD toward pretransplantation levels. Our results suggest that resistance exercise is osteogenic and should be initiated early after heart transplantation.

Anatomic variation of extrahepatic biliary tree structures: importance in treatment planning for radiation therapy.

PURPOSE: To demonstrate the variable location of biliary tree components and to assess two treatment planning rules of thumb used for locating porta hepatic nodes for radiation therapy. MATERIALS AND METHODS: The distance of the common hepatic duct bifurcation and duodenal papilla from adjacent vertebral bodies was measured on 34 cholangiograms, and their relationships to particular vertebral body levels were recorded. Adequacy of treatment volume to encompass porta hepatic lymph nodes was evaluated in 30 of these cholangiograms by constructing radiation portals according to the two rules of thumb. RESULTS: Location of the common hepatic duct bifurcation and duodenal papilla varied widely. The treatment portal encompassed 13% (four of 30) of cases when the first rule of thumb was used and 80% (24 of 30) of cases when the second rule of thumb was used. CONCLUSION: The wide variability of location of extrahepatic biliary tree structures suggests the need for individualized treatment planning so that sensitive and dose-limiting healthy tissues can be optimally excluded from radiation treatment portals. Computed tomography and other imaging modalities enable precise location of biliary tree components for radiation treatment planning.

A totally integrated simulation technique for three-field breast treatment using a CT simulator.

A method was devised to simulate patients with breast cancer in the actual treatment position utilizing a diagnostic CT spiral scanner, 3-D Image Workstation for virtual simulation, and a laser coordinate system to transfer planning parameters to the patient's skin. It was desired to produce non-divergent tangential beams through the lung as well as a matched line for tangential and supraclavicular fields. The patients were immobilized in an Alpha CradleTM cast. Radio-opaque markers were placed on the superior, inferior, medial, and lateral margins of the field so as to afford appropriate initial field set-up approximations. The patient was scanned. The data set was then transferred to the workstation where an isocenter was chosen. The patient was marked. Virtual simulation was then performed. This method employed a half beam technique for the posterior edge of the tangential fields. Table rotation and blocking of the superior margin of the tangential fields were used to produce a vertical edge to match a supraclavicular field. Using a beam's eye view the lateral tangent was matched to the medial exit. A digitally reconstructed radiograph was created to define the tangent fields and place the supraclavicular block. Our initial experience with 50 patients verifies that this is a reproducible and accurate technique. Time required for immobilization and tangential field simulation is approximately 30 minutes. Data is available for 3-D treatment planning or 2-D treatment planning on a reconstructed transverse slice angled to match the collimator angle through the patient. Using a CT simulator for simulation of breast cancer affords accuracy of at least equal magnitude to conventional simulators as determined by beam films and ease of set-up. This technique also affords greater ease in changing treatment parameters without having to resimulate the patient.

Prescription privileges and training issues: a pilot study.

Despite all the rhetoric on the controversial issue of prescriptive authority for psychologists, we posit that the major factor will be one of adequate training standards. As several training models have involved lengthy and rigorous curriculum formats, we conducted a brief pilot study on the views of practicing psychologists. Only one of the 20 licensed psychologists in our sample was willing to undertake an intensive one- to two-year program in psychopharmacology at this stage of their careers.

Definitive radiation therapy for extrahepatic bile duct carcinoma.

PURPOSE: To evaluate retrospectively the role of radiation therapy for extrahepatic bile duct (EHBD) carcinoma. MATERIALS AND METHODS: Twenty-seven patients with local-regional EHBD carcinomas were treated with definitive radiation therapy. Radiation therapy was delivered by means of external beam radiation therapy (EBRT) alone or combined with transcatheter iridium-192 brachytherapy. The median total dose was 54 Gy (range, 30-144 Gy). Survival rates were calculated and compared by using the log-rank test. Possible prognostic factors affecting survival were evaluated by means of univariate analysis. RESULTS: The median survival of all patients was 13 months, with 1- and 2-year actuarial survival rates of 52% and 10%, respectively. Univariate analysis revealed that men, patients with tumors limited to the bile duct, and patients receiving EBRT doses of at least 45 Gy had significantly better outcomes. Local-regional recurrence was the main cause of treatment failure (82%). Two patients developed gastric outlet obstruction. CONCLUSION: Patients with locally advanced EHBD carcinomas have a low survival rate. Certain factors, however, appear to have prognostic significance.

Use of dreams in therapy: a survey of clinicians in private practice.

A literature review indicated a dearth of research on how dreams are used in therapeutic settings so a questionnaire was sent to a random sample of 500 members of the Florida Psychological Association to assess (a) the extent of dream use in therapy, (b) theoretical approaches applied to dream interpretation, and (c) basis for experience and expertise in dream work. Of the 500 potential clinicians, 228 returned survey forms for a response rate of 46%. Analysis indicated that 83% of the respondents used dream material at least occasionally in their practice and that Freudian and Gestalt approaches were most often used in dream interpretation. Interestingly, most respondents gained their 'experience' in dream work through self-study and continuing education workshops and seminars.

Structural and mechanistic analysis of two refined crystal structures of the pyridoxal phosphate-dependent enzyme dialkylglycine decarboxylase.

Two refined structures, differing in alkali metal ion content, of the bifunctional, pyridoxal phosphate-dependent enzyme dialkylglycine decarboxylase (DGD) are presented in detail. The enzyme is an alpha 4 tetramer, built up as a dimer of dimers, with a subunit molecular mass of 46.5 kDa. The fold of DGD is similar to those of aspartate aminotransferase, omega-amino acid aminotransferase and tyrosine phenol-lyase. The structure has two binding sites for alkali metal ions. DGD with potassium in site 1 (near the active site) and sodium in site 2 (at the surface of the molecule) has been refined against 2.6A resolution data (R-factor = 17.6%), and DGD with sodium at both sites has been refined against 2.1 A resolution data (R-factor = 17.8%). The proximity of site 1 to the active site accounts for the dependence of enzyme activity on potassium ions, and the observed active site structural changes caused by ion exchange at this site explain the inhibition of activity by sodium. DGD catalyzes both the decarboxylation of dialkylglycine species and the transamination of L-amino acids in its normal catalytic cycle. The active site structure of DGD is moderately homologous to that of aspartate aminotransferase, which catalyzes only transamination; both the differences and similarities provide mechanistic guidelines for the DGD-catalyzed reactions. Models of the L-isovaline and L-alanine external aldimine intermediates suggest mechanisms by which the decarboxylation and transamination reactions could be accomplished within the single active site. Decarboxylation is proposed to be at least partially catalyzed by stereoelectronic activation of the C alpha-carboxylate bond achieved by orienting this bond perpendicular to the plane of the pyridinium ring in the dialkylglycine external aldimine intermediate. Transamination is proposed to be catalyzed by a similar effect on the C alpha-H bond of the L-amino acid external aldimine intermediate, combined with general base catalysis provided by Lys272, in analogy to the mechanism of aspartate aminotransferase.

Effect of low-dose testicular irradiation on sperm count and fertility in patients with testicular seminoma.

The treatment of seminoma with radiation therapy risks transient infertility. We have prospectively followed eight patients with stage I seminoma of the testicle. All patients underwent radical orchiectomy of the affected testis. The mean age of the patients was 32.9 years (range 24-40). Each patient was treated with megavoltage radiation with a 10- or 18-MV linear accelerator. The remaining testicle was shielded using a standard lead enclosure, and the mean testicular dose was 44 cGy (range 20.8-78.2). Semen specimens were delivered to the lab within 30 minutes of ejaculation. All specimens were analyzed using a computer-assisted sperm analyzer. Pretreatment parameters were within normal limits for all but one patient; one patient presented with a borderline normal sperm count at 18 and 22 x 10(6)/ml. Following treatment, there was a decrease in sperm count, detected at 3 months, to < 10 x 10(6)/ml (range 4.4- 8.6 x 10(6)) in all patients except one, who presented with an initial pretreatment count of 189 x 10(6)/ml, which decreased to 58 x 10(6)/ml at 3 months, 32 x 10(6)/ml at 6 months, and rose to 325 x 10(6)/ml by 12 months following treatment. Although the sperm count for this patient (D.L.) was within the normal range, the post-radiation sperm count was less than 20% of the pretreatment count. There was no difference in the motility at 3 months, the mean of which was 51.3%. One patient's (F.C.) wife conceived at 9 months following treatment, one at 12 months (J.R.), and one (J.S.) at 14 months, and all have delivered normal infants.(ABSTRACT TRUNCATED AT 250 WORDS)

An alkali metal ion size-dependent switch in the active site structure of dialkylglycine decarboxylase.

The pyridoxal 5'-phosphate-dependent enzyme dialkylglycine decarboxylase (DGD) is activated by K+ and Rb+ ions, whereas Li+ and Na+ ions are inhibitory. A binding site for alkali metal ions close to the active site (site 1) was discovered in the crystal structure of DGD, and an exchange of K+ for Na+ at this site was shown to affect the conformation of two active site residues [Toney, M. D., Hohenester, E., Cowan, S. W., & Jansonius, J. N. (1993) Science 261, 756-759]. We have investigated the effects of alkali metal ions on DGD activity and have determined the crystal structures at 2.8 A resolution of DGD with Li+ and Rb+ bound at site 1. Due to the weak scattering of the Li+ ion, its position had to be modeled using information from small molecule structures. A comparison of the DGD structures with Li+, Na+, K+, and Rb+ bound at site 1 reveals a striking correlation between active site structure and enzymatic activity. The small, inhibitory ions Li+ and Na+ are accommodated by replacing two protein-derived ligands of the larger, activating ions K+ and Rb+ by a single water molecule. This actuates a two-state structural switch between active and inactive enzyme that involves a concerted reorientation of the active site residues Ser80 and Tyr301 and a small change in the quaternary structure of the DGD tetramer. An important role of the essential K+ ion in both cofactor binding and the organization of a catalytically competent active site structure is proposed. In the structure of DGD with Rb+ bound at site 1, a second Rb+ ion has partially replaced the structural Na+ ion at metal binding site 2 on the surface of the DGD molecule, without significantly altering the protein structure. In contrast to Na+, the Rb+ ion is bound with unfavorable geometry, and it is proposed that the rigid site 2 structure results in a pronounced selectivity for Na+ ions.

Projective techniques: an international perspective.

This is a review of findings from four recent surveys on use of clinical tests in the United States, The Netherlands, Japan, and Hong Kong. The preliminary analysis indicates that projective techniques are popular in the assessment of personality worldwide. Obviously, projective tests are 'universal' in that unstructured stimuli serve as the basis for assessment and do not pose a language barrier. Also, reliance on projective methods may reflect problems in access to adequately translated and standardized objective tests. More data are needed from developed countries before firm conclusions on the international status of projective techniques can be affirmed.

Nondetectable prostate-specific antigen in moderately differentiated adenocarcinoma of prostate.

Prostate-specific antigen (PSA) is a glycoprotein derived from prostatic ductal and acinar epithelial cells. The main clinical use of PSA is as a marker of prostate tumor progression/recurrence. We present a case of a sixty-nine-year-old patient with recurrent endometrioid carcinoma of the prostate (status post-radical prostatectomy, hormonal therapy, and external beam radiation therapy) with normal serum PSA.

The prognostic significance of race and survival from prostate cancer based on patients irradiated on Radiation Therapy Oncology Group protocols (1976-1985).

A number of studies have identified race as a prognostic factor for survival from prostate cancer. To evaluate the prognostic significance of race in a controlled setting, we evaluated 1294 patients treated on three prospective randomized trials conducted by the Radiation Therapy Oncology Group between 1976 to 1985. One-hundred and twenty (9%) of the patients were coded as black, while 1077 (83%) of the patients were coded as white. Protocol 7506 included 607 patients with clinical Stage T3-T4Nx or T1b-T2N1-2. Protocol 7706 included 484 patients with clinical Stage T1b or T2 who were node negative. Protocol 8307 included 203 Stage T2b-T4 patients with no lymph node involvement beyond the pelvis. Univariate and multivariate analyses were used to assess the possible independent significance of race and other prognostic factors, including Gleason score, serum acid phosphatase, nodal status, and hormonal status. Protocols 7706 and 8307 revealed that race was not of prognostic significance for disease-free or overall survival by either univariate or multivariate analysis. Univariate analysis of Protocol 7506 revealed that the median survival for blacks was somewhat shorter (5.4 years vs. 7.1 years, p = 0.02). This difference persisted after a multivariate analysis. A higher percentage of blacks treated on 7506 had an abnormally elevated serum acid phosphatase compared to whites (p = 0.006), and the time to distant failure tended to be shorter (p = 0.07). These findings suggest that blacks treated on 7506 may have had more extensive disease at presentation. Based on these prospective randomized trials, it is most likely that the lower survival noted for black Americans with prostate cancer reflects the tendency for blacks to present with more advanced disease. Differences in access to care, the quality of care received, and the impact of co-morbid conditions may explain the lower survival reported for black Americans elsewhere in the literature.

Crystallization and preliminary X-ray diffraction studies of dialkylglycine decarboxylase, a decarboxylating transaminase.

The pyridoxal phosphate-dependent enzyme dialkylglycine decarboxylase (E.C. 4.1.1.64) has been crystallized by vapor diffusion from a 15% polyethyleneglycol solution with sodium pyruvate as coprecipitant. The space group of the crystals is either P6(2)22 or the enantiomorph, P6(4)22, with one subunit of 46,500 Da per asymmetric unit. The unit cell has dimensions a = b = 152.7 A, c = 86.6 A, alpha = beta = 90 degrees, gamma = 120 degrees, and a solvent content of approximately 61%. diffraction extends to 2.3 A resolution.