Effects of manipulating catecholamines on the incidence of the preovulatory surge of of luteinizing hormone and ovulation in the rat: evidence for a necessary involvement of hypothalamic adrenaline in the normal or 'midnight' surge

The preovulatory surge of luteinizing hormone reaches a maximum at 18.00 h on the day of pro-oestrus in female rats maintained with regular lighting from 06.00 to 20.00 h. This surge is initiated by a discharge of luteinizing hormone-releasing hormone into hypophysial portal blood. In this study, drugs which affect catecholamine-mediated neurotransmission were administered on the day of pro-oestrus and the effects on serum concentrations of luteinizing hormone and on subsequent ovulation were observed. alpha-Methyl-p-tyrosine, diethyldithiocarbamate and SKF 64139 inhibit catecholamine synthesis at the level of tyrosine hydroxylase, dopamine beta-hydroxylase and phenylethanolamine N-methyltransferase, respectively. Although alpha-methyl-p-tyrosine suppressed ovulation, it had a negligible effect on the incidence of the preovulatory surge. In contrast, the various treatments with diethyldithiocarbamate and SKF 64139 resulted in a minimal occurrence of the 18.00 h surge; at relatively low doses, however, these drugs frequently elicited a surge at 22.00 or 24.00 h which invariably resulted in ovulation. The failure of the surge after diethyldithiocarbamate or SKF 64139 was not associated with a loss of pituitary sensitivity to luteinizing hormone-releasing hormone. In terms of the hypothalamic concentration of dopamine, noradrenaline, adrenaline and 5-hydroxytryptamine at 18.00 h on pro-oestrus, the only common effect of diethyldithiocarbamate and SKF 64139, given in a dose which blocks the surge, was a severe depletion of adrenaline; alpha-methyl-p-tyrosine failed to produce this effect despite inducing a marked depression of dopamine and a moderate loss of noradrenaline. Neither the increase in hypothalamic dopamine after diethyldithiocarbamate, nor the alpha 2 receptor blocking properties of SKF 64139 appear to be relevant in this context since injections of L-dopa or piperoxane, an alpha 2 receptor antagonist, were without effect on the surge or ovulation. The failure of the surge after prazosin, an alpha 1 receptor antagonist, indicates that the function of adrenaline may be mediated postsynaptically by alpha 1 receptors. Clonidine, an alpha 2 receptor agonist which reduces the turnover rate of hypothalamic adrenaline, had effects of the surge and ovulation which were comparable to those of diethyldithiocarbamate and SKF 64139, the relatively low doses causing some of the surges to occur at 24.00 instead of 18.00 h and higher doses suppressing the surge at both times and thus preventing ovulation.(ABSTRACT TRUNCATED AT 400 WORDS)     

Idiotypic characterization of antibody-induced antibody responses

Anti-idiotypic antisera were produced in syngeneic (C57BL/6) mice against a monoclonal anti-Dextran B512 (Dex) antibody (38-13). In radioimmunoassays, anti-idiotypic antibodies were shown to react with the homologous idiotype, while failing to recognize another monoclonal anti-Dex antibody, independently derived from C57BL/6 mice (D-16). Plaque inhibition tests confirmed the specificity of the anti-idiotypic antibodies and revealed that the 38-13 idiotype is expressed by about half of all anti-Dex antibodies produced in C57BL/6, but not in CBA mice. Injection of normal (but not athymic) C57BL/6 mice with low doses of 38-13 monoclonal antibodies, contained culture supernatants or ascitic fluids, resulted in a 10-20 fold increase in the numbers of anti-Dex PFC detected in the spleen 5 days later, the majority of which carried the 38-13 idiotype.     

Tubal metaplasia: A cytologic study with comparison to other neoplastic and non-neoplastic conditions of the endocervix

Tubal metaplasia of the endocervix (TME), a condition that may be con/used morphologically with glandular neoplasia, is frequently found in cone or hysterectomy specimens. To determine the frequency of detecting TME in cytologic smears, we retrospectively reviewed 28 Papanicolaou (Pap) smears from 22 women (mean age 39.1 yr; range 25-60 yr) with histologically proven TME. Our criteria for TME were the presence of two cell types in addition to endocervical secretory cells, i.e., peg cells (cells with dark and granular cytoplasm and elongate nuclei) and ciliated cells. All women had cervical cytology specimens obtained with an endocervical brush shortly before the procedures in which TME was diagnosed, and five also had at least one post-procedure smear. Of 20 smears with an adequate, non-neoplastic endocervical component, TME was found in 2 (10%). In these two, TME cells constituted 10% and < 5% of all the glandular cells, respectively, and the percentage of ciliated cells in the TME was approximately 25% and 75%. In conclusion, TME was noted infrequently (10%) on the cervical cytosmears of women with histologically-proven TME. This result corresponds to the histologic finding that TME typically involves the upper endocervix and glandular epithelium, with only 13% of the women having TME on the surface of the lower endocervix. Atypical glandular cells on cervical cytology are a problem for clinicians and pathologists alike. The differential diagnosis of such atypia, including TME, cells of the lower uterine segment, squamous intraepithelial lesion in glands and glandular neoplasia, is discussed.     

Polarized expression of Na+/H+ exchange activity in LLC-PK1/PKE20 cells: II. Hormonal regulation

LLC-PK₁/PKE₂₀ cells (a continuous epithelial cell line) has two different Na/H exchange activities: Na/H-1 located in the basolateral membrane and Na/H-2 located in the apical membrane [Casavola et al. (1989) Biochem Biophys Res Commun 165:833–837; Haggerty et al. (1988) Proc Natl Acad Sci USA 86:6797–6801]. In the present report we have studied hormone regulation of these exchange activities by measuring Na-dependent recovery of pH_i from an acid load (by using microspectrofluorometry and 2,7-bis(carboxyethyl)-5,6-carboxyfluorescein) in response to activation of regulatory cascades by either pharmacological agents or by vasopressin or calcitonin. Agents leading to activation of protein kinase A (cAMP-dependent), such as forskolin (10 M), 8-Br-cAMP (0.25 mM), and isobutylmethylxanthine (0.5 mM), inhibited Na/H-2 and Na/H-1 by an average of 49%. Stimulation of protein kinase C by a phorbol ester (phorbol 12-myristate 13-acetate, TPA, 100 nM) inhibited Na/H-2 (by an average of 48%) and stimulated Na/H-1 (by an average of 38%); these effects of TPA were also observed in the presence of forskolin (100 M). Addition of either vasopressin (2 M) or calcitonin (0.3 M) onto both sides of the monolayer decreased the activity of Na/H-2 by an average of 26.3% and 27.7% respectively, and stimulated the activity of Na/H-1 by an average of 17.4% and 38.7% respectively; exposure of cells to either hormone stimulated production of cAMP and inositol trisphosphate, respectively. Separate hormone additions to either the apical or basolateral cell surface led to effects similar to those produced by simultaneous hormone additions onto both cell surfaces, although the relative response of Na/H exchangers to either agonist is variable. In summary, these results suggest that in LLC-PK ₁/PKE₂₀ cells, vasopressin and calcitonin can act via receptor systems coupled either to adenylate cyclase or to phospholipase C. Activation of these receptor systems can lead to inhibition of Na/H-2 and stimulation of Na/H-1.     

Hepatitis B surface antigen particles of subtypes adw and adr, and compound subtype (adwr) in symptom-free carriers in Japan.

Of sera from 1,878 Japanese blood donors who carried hepatitis B surface antigen (HBsAg), 420 were subtyped as adw (22.4%) and 1,443 as adr (76.8%); only 15 (0.8%) contained HBsAg of subtype ayw or ayr. Sera with HBsAg/adr had higher HBsAg titres than those with HBsAg/adw (geometric mean of haemagglutination titre: 10.1 +/- 2.4 vs. 9.7 +/- 2.4, p less than 0.01), and a higher prevalence of hepatitis B e antigen (24% vs. 13%, p less than 0.001). Carriers of HBsAg/adr progressively predominated over those of HBsAg/adw with increasing age. Of sera from 1,863 carriers of HBsAg/adw or HBsAg/adr, 182 (9.8%) contained HBsAg particles with both subtypic determinants in the w/r allele. The presence of w and r determinants on the same particles was ascertained by sandwiching them between monoclonal antibody with the specificity for w and that with the specificity for r. HBsAg particles of compound subtype (adwr) were found more often in sera with hepatitis B e antigen than those without it (145/403 [36.0%] vs. 37/1,460 [2.5%], p less than 0.001). Sera with HBsAg/adwr particles had HBsAg titres higher than those without them (12.4 +/- 1.9 vs. 9.7 +/- 2.3, p less than 0.001). HBsAg/adwr particles arise from phenotypic mixing of the S-gene product of wild-type virus and that of mutants with point mutations for subtypic changes. The results obtained indicated that HBV strains of subtype adr have a higher replicative activity than those of adw, and suggested that mutations in the S gene for subtypic changes would be associated with an active replication of hepatitis B virus.     

Gestörte alpha2-Adrenozeptorfunktion bei Hämodialysepatienten mit renaler Anämie — eine mögliche Ursache der Blutdrucksteigerung unter rekombinantem humanem Erythropoietin?

Summary Nine patients on maintenance hemodialysis and transfusion-demanding renal anemia (group A) were treated with rHuEPO 120 IU/kg i.v. three times per week. Hemoglobin-content was raised from 7.2±0.9 to 10.4±0.8 g/dl. In all patients blood pressure rose, three patients developed arterial hypertension. Mean diastoloic blood pressure was 66±12 and 78±16 mmHg (p<0.001) before and after rHuEPO. Rise in blood pressure was accompanied by a significant fall in plasma-noradrenaline-levels (from 498±100 to 383±75 pg/ml;p<0.05) and alpha₂-adrenoceptor-density (from 574±76 to 384±49;p<0.05). Compared to nine patients on maintenance hemodialysis and hematocrit over 30% (group B), patients with severe renal anemia (group A before treatment) had higher densities of alpha₂-adrenoceptors (574±76 vs. 218±32;p<0.001) despite higher plasma-noradrenaline-levels (498±100 vs. 399±63; n.s.). We suppose a anemia-related disturbance of alpha₂-receptor-function with the result of abolished receptor down-regulation and impaired vascular reagibility to vasoconstricting stimuli. With the correction of anemia receptor-function improves, receptor down-regulation as well as vascular reagibility is re-established resulting in augmented vascular resistance and higher blood pressure.

---

Abkürzungen rHuEPO rekombinantes humanes Erythropoietin - teMAP mittlerer arterieller Blutdruck - RR Blutdrucknach RIVA-ROCCI - RBF regionaler Blutfluß - RPR regionaler peripherer Widerstand     

The value of deletion analysis for carrier detection in Duchenne muscular dystrophy (DMD)

We performed genetic analysis for carrier detection for several at-risk females in a four-generation Duchenne muscular dystrophy (DMD) pedigree using deletion analysis. We demonstrated that dosage analysis is a suitable alternative method to determine the carrier status of female relatives of DMD patients shown to have a deletion within the DMD gene. Subsequently, we diagnosed an affected male fetus for an at-risk female shown to be a DMD carrier by deletion analysis. The usefulness of deletion and linkage analysis are compared. In this family, linkage analysis was complicated by the unavailability of key family members, two recombination events and by previously undisclosed nonpaternity. We found that dosage analysis was more efficient than linkage for carrier evaluation in this family.     

Improvement of Accuracy in a High-Capacity, Six Degree-of-freedom Load Cell: Application to Robotic Testing of Musculoskeletal Joints

This study investigated a previously unaccounted for source of error in a high-capacity, six degree-of-freedom load cell used in multi-degree-of-freedom robotic testing of musculoskeletal joints, an application requiring a load cell with high accuracy in addition to high load capacity. A method of calibration is presented for reducing the error caused by changes in universal force-moment sensor (UFS) orientation within a gravitational field. Uncorrected, this error can exceed a magnitude of 1% of the full-scale load capacity—the manufacturer-stated accuracy of the UFS. Implementation of the calibration protocol reduced this error by approximately 75% for a variety of loading conditions. This improvement in load cell accuracy (while maintaining full load capacity) should improve both the measurement and control of specimen kinetics by robotic/UFS and other biomechanical testing systems. © 1999 Biomedical Engineering Society. PAC99: 8719Rr, 8780Vt, 0620Fn, 0620Dk, 8719Ff     

Infrequent genetic alterations of the PTEN gene in Japanese patients with sporadic prostate cancer

Prostate cancer is a major cause of cancer death among elderly men in America, Europe, and Japan. However, the molecular mechanism of carcinogenesis is not yet well characterized. Frequent loss of heterozygosity (LOH) on chromosome 10q was reported in prostate cancer, and a candidate tumor suppressor gene, PTEN, was isolated on chromosome band 10q23.3. To investigate the genetic alterations of PTEN, we examined 45 primary prostate cancer specimens. LOH at the PTEN locus was observed in two (11.1%) of 18 tumors. However, no mutations were observed in any of the primary prostate cancers. These data suggest that mutation of the PTEN gene does not play a major role in prostate carcinogenesis of Japanese patients. Received: February 6, 1998 / Accepted: July, 3, 1998     

Increased poly(ADP-ribose) polymerase activity in lymphoblastoid cell lines from centenarians

 Poly(ADP-ribosyl)ation is a posttranslational modification of nuclear proteins which is catalyzed by poly(ADP-ribose) polymerase and represents an immediate response of eukaryotic cells to oxidative and other types of DNA damage. Previously a strong correlation had been detected between maximal poly(ADP-ribose) polymerase activity in permeabilized mononuclear leukocytes of various mammalian species and species-specific life span. To study a possible relation between longevity and poly(ADP-ribosyl)ation in humans we measured maximal oligonucleotide-stimulated poly(ADP-ribose) polymerase activity in permeabilized, Epstein-Barr virus transformed lymphoblastoid cell lines from a French population of 49 centenarians and 51 controls aged 20–70 years. Maximal enzyme activity was significantly higher in centenarians than in controls [median of controls: 9035 cpm/10⁶ cells (lower quartile: 6156; upper quartile: 11,410); median of centenarians: 10,380 cpm/10⁶ cells (lower quartile: 7994; upper quartile: 12,991); P=0.031 by Mann-Whitney U test]. In a subset of 16 controls and 24 centenarians, cellular poly(ADP-ribose) polymerase content was determined by quantitative western blotting, thus allowing the calculation of specific enzyme activity. The latter was significantly higher in centenarians (P=0.006), the median value for centenarians being about 1.6-fold that of controls. Specific poly(ADP-ribose) polymerase activity was a more powerful parameter for differentiating between centenarians and controls than enzyme activity relative to cell number. In addition, in a genetic association study we analyzed 437 DNA samples (239 centenarians and 198 controls) by PCR amplification of a polymorphic dinucleotide repeat located in the promoter region of the poly(ADP-ribose) polymerase gene in an attempt to detect an association between this polymorphic marker and variability of enzyme activity or human longevity. However, this genetic analysis revealed no significant enrichment of any of the alleles or genotypes identified among centenarians or controls, but its power was limited by the relatively weak hetero-zygosity of this polymorphic marker in our population (51%). Viewed together with previous results on poly(ADP-ribose) polymerase activity in various mammalian species, the present data provide further evidence for the notion that longevity is associated with a high poly(ADP-ribosyl)ation capacity. Received: 5 September 1997 / Accepted: 10 November 1997