Overexpression of p53 in prostate carcinoma is associated with improved overall survival but not predictive of response to radiotherapy

p53 gene mutations are among the most common specific genetic alterations in human cancer. Inactivation of p53 and subsequent protein accumulation has been implicated in a variety of human malignancies and associated with prostate cancer progression. In this study, we assessed p53 protein overexpression and gene mutations in prostate carcinoma and investigated associations between p53 alterations and clinicopathological parameters, survival, and response to radiotherapy. We evaluated 58 archival formalin-fixed, paraffin-embedded prostate carcinomas to detect abnormal p53 nuclear protein accumulation using immunohistochemistry. p53 mutational status of tumor DNA was evaluated using polymerase chain reaction-single-strand conformation polymorphism analysis of exons 5-9 and confirmed by direct DNA sequencing. Univariate and multivariate statistical analysis was used to determine the association of p53 status with clinical characteristics and response to radiotherapy. Overexpression of p53 was detected in 42 (72%) of 58 primary prostate carcinomas, but was undetectable in 7 samples of benign prostatic hyperplasias or 5 samples of normal prostate tissue. p53 exon 5-9 mutations were detected in 8 (14%) of 58 patient specimens. p53 mutational status, but not overexpression, was associated with higher Gleason scores (p=0.0145). Neither p53 overexpression nor mutation was associated with clinical stage, biochemical disease-free probability, or predictive of response to radiotherapy. p53 protein accumulation was inversely associated with improved overall survival (p=0.0108). Our studies demonstrate that p53 protein accumulation is a frequent alteration in prostate cancer. The disparity between p53 protein overexpression and p53 exon 5-9 mutations suggests the possibility of mutations outside this region or stabilization of wild-type p53 by alternative mechanisms. In our patient population, p53 protein overexpression or mutational status was not predictive of outcome in patients treated with radiation therapy. Additional studies are needed to further evaluate the association between p53 protein overexpression and improved overall survival.     

Optimization and standardization of an enzyme-linked immunosorbent assay protocol for serodiagnosis of Actinobacillus pleuropneumoniae serotype 5.

An indirect enzyme-linked immunosorbent assay protocol has been optimized with special emphasis given to assay standardization and quality control. Technical aspects such as choice of a microplate, antigen immobilization, buffer composition, optimal screening dilution of sera, and kinetics of the enzymatic reaction were studied and evaluated in order to design a standard protocol offering maximal analytical sensitivity and specificity, as well as to obtain minimal within- and between-plate variability. Among the 27 plates tested, the Nunc 475-094 and 269-620 immunoplates were found to be the best in terms of high positive-to-negative ratio and low variability. No significant differences in antigen immobilization were found by using buffers of various compositions or pHs; however, the presence of magnesium ions (Mg2+; 0.02 M) resulted in a twofold increase in nonspecific background. An optimal screening dilution of sera was established at 1:200. A 1-h incubation period for test serum was found to be optimal. Maximum enzymatic activity for peroxidase was obtained by adjusting both substrate (H2O2) and hydrogen donor [2,2' -azinobis(3-ethylbenz-thiazoline sulfonic acid)] concentrations to 4 and 1 mM, respectively. To control between-plate variability, a timing protocol was adopted. Within-plate variability was also controlled by using a sample placement configuration pattern. Sliding scales were determined by repeated testing of a cross section of samples to set acceptance limits for both within- and between-plate variability. These limits were used in a quality control program to monitor assay performance. The results obtained suggest that this standardized protocol might be useful in the serodiagnosis of Actinobacillus pleuropneumoniae serotype 5.     

The dual-task methodology and assessing the attentional demands of ambulation with walking devices.

The purposes of this article are (1) to provide a preliminary examination of the attentional demands of ambulating with two commonly prescribed walking aids (a standard walker and a rolling walker) and (2) to introduce the dual-task methodology to the physical therapy community. Five subjects familiar with the appropriate use of the walkers and five subjects uninformed as to the correct use of the walkers participated in the study. Each subject completed the three phases of the experiment: (1) performing the reaction time (RT) task only; (2) performing each of the walking tasks only; and (3) performing each of the walking tasks in conjunction with the RT task, which constituted the dual-task conditions. The findings indicated that walking aided by either the rolling walker or the standard walker was highly attention demanding. More importantly, it appears that greater attentional demand was required when ambulating with the standard walker. These results are discussed with respect to the gait modifications and accuracy demands required when using these walkers. The usefulness of the dual-task methodology as a research tool for addressing clinically oriented questions is emphasized, and some potential applications of this methodology for the therapist within the clinic are discussed.     

Computational methods for determining protein structures from NMR data

The general procedures by which solution structures of proteins may be deduced from distance and angular constraints derived from NMR are reviewed, with an emphasis on practical aspects of the calculations. In addition, novel methods based on chemical shift calculations and on quantitative fits to nuclear Overhauser effect intensities are presented; these should provide improved understanding of the limits of our ability to simulate complex spectra, and may permit higher precision structures to be determined.     

Frontal sinus fractures: evaluation of CT scans in 132 patients.

PURPOSE: To determine the frequency of detection of frontal sinus fractures on initial CT scans of patients with intracranial injuries, and to characterize associated injuries. METHODS: The initial head CT scans in 132 patients with clinical or radiographic evidence of a frontal sinus fracture were retrospectively reviewed to further characterize the fracture. Additional radiographic studies and medical records were reviewed to determine associated injuries, therapy, clinical outcome, and complications. RESULTS: In 90% (124) of the patients, the frontal sinus fractures were visualized on initial head CT scans that were obtained to evaluate suspected intracranial injury. Complex fractures involving both the anterior and posterior wall of the sinus accounted for 65% of cases (86 patients), whereas fractures of the anterior wall only or posterior wall only occurred in 24% (32) and 11% (14) of patients, respectively. Significant intracranial hemorrhage occurred in over 90% of patients with fractures involving the posterior wall. CONCLUSIONS: In general, fractures that involved the posterior wall had more complications and a worse clinical outcome than fractures that only involved the anterior wall; nearly all frontal sinus fractures can be detected on head CT studies in patients with intracranial injuries.     

The usefulness of clinical measurements of cell proliferation in gynecological cancer.

This article critically reviews the available methods for the assessment of cell proliferation in clinical practice, and evaluates their applications as diagnostic and prognostic variables in gynecological cancer. The mitotic count is not a standardised method and should not be considered to have inherent value as a measurement of cell proliferation in the clinical situation. In distinguishing benign and malignant uterine smooth muscle tumors, the degree of mitotic activity can only be interpreted in the context of the other pathological and clinical findings. Cytological atypia and the absence of stromal differentiation appear to be more accurate than mitotic activity in delineating high-grade endometrial stromal sarcomas. The potential biological behavior of placental site trophoblastic tumor cannot be reliably predicted on the basis of mitotic counts, and hysterectomy remains the treatment of choice for this condition. Flow cytometry produces accurate cell kinetic data and provides useful prognostic information in ovarian and endometrial tumors. The value of flow cytometry in cervical cancer is questionable and needs clarification. Complete and partial hydatidiform moles are generally distinguishable on the basis of ploidy studies. However, in view of the existence of a diploid partial mole, serial human chorionic gonadotrophin measurement is the only reliable method of establishing or excluding a diagnosis of persistent trophoblastic disease. The other clinical markers of cell proliferation do not have an established role as diagnostic or prognostic parameters in gynecological cancer and merit further critical evaluation. Estimation of mitotic activity is a simple, readily accessible method of assessing cell proliferation in routine histopathological practice and the role of an accurate mitotic index in diagnosis and prognosis must be investigated to determine the true value of counting mitotic figures in histological sections.     

Nonprostanoid prostacyclin mimetics. 2. 4,5-Diphenyloxazole derivatives.

4,5-Diphenyl-2-oxazolenonanoic acid (18b) was synthesized and found to inhibit ADP-induced aggregation of human platelets with an IC50 of 2.5 microM. Acid 18b displaced [3H]iloprost from human platelet membranes in a concentration-dependent fashion, consistent with 18b inhibiting platelet function by acting as a prostacyclin mimetic. By inserting a phenoxy ring into the side-chain moiety of 18b and systematically varying the pattern of substitution and length of the tethers, more potent inhibitors of platelet aggregation were identified. A phenoxy ring inserted centrally in the side chain proved to be the optimal arrangement but significant activity was observed when the aromatic ring was bound directly to the 2 position of the heterocycle. The meta-substituted cis-(ethenylphenoxy)acetic acid 37 is the most potent platelet aggregation inhibitor synthesized as part of this study with an IC50 of 0.18 microM. Acid 37 displaces [3H]iloprost from human platelet membranes with an IC50 of 6 nM. The trans-olefinic isomer of 37 (25p) is 72-fold weaker as an inhibitor of ADP-induced platelet aggregation, but the saturated derivative 25w (BMY 42393) is intermediate in potency. Structure-activity studies using 25w as a template focused on modification of the tethers intervening between the side-chain phenyl ring and the oxazole and carboxylate termini and substitution of the phenyl ring. These studies revealed that biological activity was sensitive to both the identity of the concatenating atoms and the pattern of ring substitution. The structure-activity relationships provide insight into the topographical relationship between the diphenylated oxazole ring and the carboxylic acid terminus that comprise the nonprostanoid prostacyclin mimetic pharmacophore.     

Biotic Factors Modifying Acute Toxicity of Aqueous Cadmium to Estuarine Amphipod Leptocheirus plumulosus

A 96-h exposure to aqueous cadmium (Cd) is the recommended reference toxicity test for 10-day sediment bioassays with the estuarine amphipod, Leptocheirus plumulosus (US EPA 1994). This water-only test was used to assess the influence of organism size, sex, and nutritional status on the sensitivity of laboratory-cultured L. plumulosus to Cd. In addition, the response of field-collected amphipods was compared to similarly sized laboratory animals to assess potential seasonal changes in Cd sensitivity. Lipid content of test organisms was measured in these seasonal experiments and those evaluating effects of nutritional status because of its potential as an indicator of physiological condition. LC₅₀ values of laboratory animals size-sorted on nested 500-, 710-, and 1000-μm mesh sieves, increased with size class: 0.36, 0.65, and 0.88 mg Cd/L, respectively. Gravid females were less sensitive than males or mature females to aqueous Cd. Studies on the influence of the molt cycle on Cd toxicity indicated enhanced sensitivity of immediate postmolt animals that may explain some of the observed differences in Cd tolerance. Nutritional effects were investigated by comparing the sensitivity of fed and starved laboratory-reared amphipods. Starved juveniles and adults were significantly smaller than their fed counterparts and exhibited a 28–43% reduction in lipid content, respectively. However, comparison of LC₅₀ values indicated no significant differences in sensitivity to Cd between starved and fed juveniles (0.23 vs 0.30 mg Cd/L) or adults (0.37 vs 0.52 mg Cd/L). Field-collected amphipods were typically more sensitive to Cd than laboratory animals, regardless of the season, although their lipid content varied, ranging from 6.6% in August to 13.7% in November. Results are discussed with respect to the use and interpretation of toxicity tests with this species. Received: 18 February 1997/Accepted: 5 May 1997