Development of a microassay for estradiol receptors

The calculation of equilibrium binding constants for a specific receptor-hormone interaction requires the exact determination of the unbound ligand concentration and the specifically bound concentration at equilibrium. These determinations usually require corrections for the contribution of non-specific binding. We introduce several parameters allowing equilibrium concentrations to be calculated from experimental concentration values; these parameters can be measured for the particular receptor assay procedure used. These parameters are used in a microassay of estradiol-receptor complex by selective retention on DE-81 cellulose paper.     

Deletion of the central proline-rich repeat domain results in altered antigenicity and lack of surface expression of the Streptococcus mutans P1 adhesin molecule

Members of the family of surface adhesins of oral streptococci, including P1 of Streptococcus mutans, contain two highly conserved repeat domains, one rich in alanine (A region) and the other rich in proline (P region). To assess the contribution of the P region to the biological properties of P1, an internal deletion in spaP was engineered. In addition, the P region was subcloned and expressed as a fusion partner with the maltose binding protein of Escherichia coli and liberated by digestion with factor Xa. Results of Western blot experiments in which recombinant polypeptides were probed with a panel of 11 monoclonal antibodies indicated that the P region is a necessary component of conformational epitopes within the central portion of P1. Antibodies reactive with the P region were detected in a polyclonal rabbit antiserum generated against whole S. mutans cells but not in two rabbit antisera generated against purified P1 (Mr approximately 185,000), suggesting that this domain is immunogenic on the surface of intact bacteria but not as part of a soluble full-length molecule. Finally, transformation of a spaP-negative mutant with a shuttle vector containing an internally deleted spaP lacking P-region DNA resulted in a complete absence of surface-localized P1 and substantially less P1 in sonicated cells compared to the case for the mutant complemented with the full-length gene. These results suggest that the P region is an integral component contributing to the conformation of the central region of P1 and indicate that its presence is necessary for surface expression of the molecule on S. mutans.     

5-hydroxytryptamine1A receptor-mediated effects on adenylate cyclase and nitric oxide synthase activities in rat ventral prostate

AIM: To investigate the patterns of expression of transforming growth factor alpha (TGF-alpha) and epidermal growth factor receptor (EGFR) in squamous metaplasia and squamous cell carcinomas of the urinary bladder with and without schistosomiasis. METHODS: Immunohistochemical study of the expression of TGF-alpha and EGFR in squamous metaplasias (n = 12) and various grades of squamous cell carcinomas (n = 21) of the bladder with and without schistosomiasis. RESULTS: Focal cytoplasmic and membranous positivity for EGFR and TGF-alpha was seen in all cases of squamous metaplasia. The markers were diffusely coexpressed in a concordant pattern in areas of hyperplastic keratinising squamous metaplasia. A similar pattern of positivity was seen in verrucous carcinomas (n = 2) and well differentiated squamous carcinomas (n = 6). Progressive loss of differentiation was associated with increasing loss of EGFR staining while TGF-alpha staining was retained. Squamous cell carcinoma in situ (n = 2) showed focal positivity for TGF-alpha and EGFR. There were no differences in staining patterns between cases with and without schistosomiasis. CONCLUSIONS: The coexpression of TGF-alpha and EGFR by well differentiated squamous cell carcinomas and hyperplastic keratinising squamous metaplasia is consistent with the active regulatory role exerted by this autocrine loop. There is regional absence of expression of EGFR but not of TGF-alpha in squamous cell carcinomas of lesser differentiation, suggesting heterogeneity of such control in these tumours. The focal expression of the two markers in squamous cell carcinomas in situ indicates a possible second pathway of oncogenesis for less differentiated tumours. These observations may have important implications for the effectiveness of putative growth factor based treatments.     

Ocular chrysiasis correlated with gold concentrations in the crystalline lens during chrysotherapy

The eyes of 11 patients with rheumatoid arthritis (RA) who received extended chrysotherapy (mean cumulative dose greater than 7 grams during a mean 6-year period) were examined biomicroscopically. Minute reddish-purple particles were seen in the cornea (corneal chrysiasis) in 5 and in the lens (lens chrysiasis) in 4 patients. Particulate deposits were absent in 11 other RA patients who had not received gold treatment. Seven crystalline lenses from 5 gold-treated patients were removed surgically because of incidental cataract formation and analyzed for gold content using neutron activation analysis. Although the mean lens gold concentration was higher in these patients than in non-gold-treated controls without RA (0.0073 microgram/grams versus 0.001 microgram/grams), the absolute gold level was markedly lower than that found in 25 diverse tissue analyzed previously. This finding is compatible with the absence of clinical gold-related lens disease or visual impairment.     

Concurrent mapping of brain sites for sensitivity to the direct application of morphine and focal electrical stimulation in the production of antinociception in the rat

The effect of morphine microinjection (5 microgram/0.5 microliter) and focal electrical stimulation on the animal's response to radiant heat and noxious pinch was studied concurrently at 117 brain loci extending from the medial thalamus caudally to the periaqueductal gray area (PAG). Three populations of brain sites were discernible based on their responsiveness to focal electrical stimulation and morphine microinjection in the production of antinociception: (a) sites which support stimulation-produced analgesia (SPA, n = 24), (b) sites which were sensitive to the direct application of morphine (n = 8), (c) sites responsive to both manipulations (n = 8). With a few exceptions, all morphine sensitive sites were located within the anatomical boundaries of the PAG while sites supporting SPA were located not only within the PAG but also in the brain regions peripheral to this structure. Sites responsive to both manipulations were generally distributed throughout thf lateral aspect of the posteroventral PAG. Stimulation strength-effect curves for sites subserving SPA were also obtained. No differences were discovered between curves obtained from morphine-sensitive and -insensitive brain loci.     

A study of thyroid disease in family practice

Thyroid disease is relatively common in family practice, yet is often undiagnosed or poorly managed. This study examines several aspects of thyroid disease in a large, semirural family practice setting and exemplifies the type of practical clinical research that can be done in family medicine. An overall prevalence of approximately one percent was determined for thyroid disease in this practice. In a series of 85 patients, the ratio of hypothyroidism:hyperthyroidism:euthyroid goiter was 9:2:1 respectively. Initial signs and symptoms recorded for these patients conformed closely to the findings in other large series. Eighty percent of the patients with idiopathic hypothyroidism never had enlarged glands, whereas 100 percent of the patients with hypothyroidism associated with Hashimoto's thyroiditis had enlarged glands. Laboratory aids such as serum thyroid stimulating hormone (TSH), anti-thyroid antibodies, and radioactive iodine uptake (RAIU) and scans were inadequately utilized. Medical and/or surgical consultation was obtained in 17.5 percent of patients with hypothyroidism, 80 percent of patients with hyperthyroidism, and 63 percent of those with euthyroid goiter. Currently 95 percent of the hypothyroid patients and 100 percent of the hyperthyroid patients are euthyroid.