Impact of primary surgical approach in the management of the impalpable testis.

There is no univoque opinion about the place of preoperative studies in non-palpable testes. During a 6.5-year period, we operated on 296 impalpable testes in prepubertal boys. A combined inguinal-abdominal approach was used in all cases verifying the eventual abdominal testis and its exact vascular anatomy before any manipulation of the cord was undertaken. Forty-five testes (15.2%) were canalicular, 142 (48%) were abdominal, 5 (1.7%) dysgenetic and 104 (34.1%) absent (agenesis or vanishing testis). Of the abdominal testes, 122 underwent a standard orchidopexy in dartos pouch, 11 a staged repair, 8 a Fowler-Stephens operation and 1 orchiectomy. All means of investigation for impalpable testes are either unreliable, too expensive or too invasive for routine use, and in most cases, a surgical exploration has to be performed anyway. The primary surgical approach has the most favorable cost/benefit ratio, being diagnostic and therapeutic at one time. Provided the exploration is performed correctly, all the advantages of previous laparoscopy can be achieved with surgery alone.


Correlation between the expression of P-glycoprotein and multidrug-resistant phenotype in transitional cell carcinoma of the urinary tract.

The expression of a multidrug-resistant (MDR) gene product, P-glycoprotein, was examined immunohistochemically in 41 transitional cell carcinomas (TCCs) of the urinary tract. In 23 of these, chemosensitivity to adriamycin (ADM) and vinblastine (VBL) was also assessed by a microtiter succinate dehydrogenase inhibition test and the correlation between the expression of P-glycoprotein and MDR phenotype was investigated. P-glycoprotein was detected in 13 (72.2%) of the 18 untreated TCCs of the upper urinary tract (UUT), 6 (31.6%) of the 19 untreated TCCs of the bladder, and all of the 4 TCCs treated with M-VAC chemotherapy, respectively. Fourteen (87.5%) of the 16 TCCs with a positive expression of P-glycoprotein were resistant to ADM and VBL, whereas all of the 4 TCCs sensitive to both drugs were negative in the expression of P-glycoprotein. The succinate dehydrogenase activity of TCCs with a positive expression of P-glycoprotein was significantly higher than that of TCCs with a negative expression of P-glycoprotein (P < 0.05). Thus, there was a good correlation between the expression of P-glycoprotein and MDR phenotype in the chemosensitivity test. These results suggest that intrinsic MDR exists in some TCCs of the urinary tract, particularly UUT, and that the immunohistochemical investigation of P-glycoprotein may be useful for predicting the MDR phenotype in TCCs of the urinary tract.


Early diagnosis of the multiple endocrine neoplasia type 2 syndrome: consensus statement. European Community Concerted Action: Medullary Thyroid Carcinoma.

The diagnosis of medullary thyroid carcinoma by biochemical and genetic testing is possible in families with multiple endocrine neoplasia type 2. At an early stage total thyroidectomy usually cures the patient. As the clinical penetrance of the autosomal dominant, transmitted, multiple endocrine neoplasia type 2 gene is not complete, family screening is indicated for every new patient who presents with apparently sporadic medullary thyroid carcinoma. Problems related to a screening programme and early diagnosis have led the members of the European Community Concerted Action: Medullary Thyroid Carcinoma group to formulate a consensus on biochemical and genetic screening. For biochemical screening, measurement of basal and pentagastrin and/or calcium stimulated serum levels of calcitonin by radioimmunoassay are essential starting at the age of three and continuing annually until 35 years of age. Furthermore, annual screening for pheochromocytoma by measuring the urinary excretion of catecholamines and for hyperparathyroidism by serum calcium determination is indicated. Genetic screening using linked markers can be done with a 95% accuracy in informative families when DNA is available from at least two family members proven to be affected. Biochemical screening can thus be reserved for gene carriers, while those at low risk can be reassured. Combined biochemical and genetic screening for multiple endocrine neoplasia type 2 is important and effective for the cure of medullary thyroid carcinoma.


Noradrenergic mediation of spinal antinociception by 5-hydroxytryptamine: characterization of receptor subtypes.

The present study examined the involvement of spinal noradrenergic mechanisms in spinal antinociception by the 5-hydroxy-tryptamine (5-HT) receptor-selective agonists CGS 12066B (5-HT1B; 7-trifluoromethyl-4(4-methyl-1-piperazinyl)-pyrrolo[1,2-a]quinoxaline), TFMPP (5-HT1C; M-trifluoromethylphenyl-piperazine) and DOI (5-HT2; 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane) using the rat hot plate test. Effects of alpha-adrenoreceptor antagonists (phentolamine, yohimbine), the adrenergic neurotoxin 6-hydroxydopamine, and the selective noradrenergic uptake blocker desipramine were determined. CGS 12066B, TFMPP and DOI produced dose-related antinociception. The antinociceptive effect of each agent was reduced by pretreatment with both phentolamine and yohimbine (15-60 micrograms). Pretreatment with 6-hydroxydopamine (100 micrograms, intrathecal) for 7-10 days, which reduced spinal cord levels of noradrenaline by 87%, inhibited the action of TFMPP (and 5-HT), but not CGS 12066B or DOI. Pretreatment with desipramine (25 mg/kg, systemic) potentiated the action of TFMPP but not CGS 12066B or DOI (or 2-methyl-5-HT). These results suggest that antinociception by TFMPP is dependent on release of endogenous noradrenaline from the spinal cord, while that produced by CGS 12066B and DOI is not. As TFMPP exhibits a close similarity to 5-HT in these experiments, the 5-HT receptor subtype being activated to induce noradrenaline release may either be a 5-HT1C or a 5-HT1S subtype. Other mechanisms account for the observed blockade of the action of CGS 12066B and DOI by alpha-adrenoreceptor antagonists.


5-HT2 receptors exert a state-dependent regulation of dopaminergic function: studies with MDL 100,907 and the amphetamine analogue, 3,4-methylenedioxymethamphetamine.

The highly selective 5-HT2 receptor antagonist, MDL 100,907, was used to explore the role of serotonin in the stimulation of dopaminergic function produced by the amphetamine analogue 3,4-methylenedioxymethamphetamine (MDMA). MDL 100,907 blocked MDMA-stimulated dopamine synthesis in vivo without affecting basal synthesis. The long-term deficits in 5-HT concentrations believed to be a consequence of MDMA-induced dopamine release were also blocked by MDL 100,907 over the same dose range. In vivo microdialysis confirmed that 5-HT2 receptor blockade with MDL 100,907 attenuated MDMA-induced increases in extracellular concentrations of striatal dopamine. In contrast to its effect on MDMA-induced synthesis, MDL 100,907 did not alter dopamine synthesis stimulated by haloperidol or reserpine. In vivo dopamine release produced by haloperidol was also unaffected by MDL 100,907. The results suggest a permissive role for 5-HT2 receptors in the activation of the dopamine system which occurs during states of high serotonergic activity or during conditions of elevated dopamine efflux with high D2 receptor occupancy.


Suppression by cetirizine of pleurisy triggered by antigen in actively sensitized rats.

The efficacy of cetirizine in comparison with meclizine, another piperazine H1 receptor antagonist, in rat pleurisy caused by allergen or autacoid was investigated. Sensitization was achieved by subcutaneous injection of a mixture of ovalbumin and aluminium hydroxide. Fourteen days later, the animals were challenged with an intrathoracic injection of ovalbumin (12 micrograms/cavity), which caused drastic mast cell degranulation, followed by pleural oedema and leucocyte influx. Cetirizine and meclizine (2.5-30 mg/kg i.p.), 1 h before challenge, inhibited the exudatory response evoked by antigen, under conditions where neutrophil and eosinophil accumulation was affected only by the former. When administered intrathoracically 22 h after allergen, i.e. using a curative approach, cetirizine (15 micrograms/cavity) drastically reduced the pleural eosinophilia noted 24 h post-challenge, indicating that this drug can reverse an already established eosinophilia. Cetirizine (15 mg/kg i.p.) also restored, to about 39% (P < 0.001), the number of uninjured mast cells recovered from the pleural cavity following allergen stimulation. In normal rats, cetirizine (5-15 micrograms/cavity) completely inhibited the pleural exudation elicited by histamine and only partially the exudation caused by 5-hydroxytryptamine or bradykinin, but was quite inactive against platelet-activating factor. We conclude that the pleural exudation triggered by allergen, vasoactive amines or bradykinin is clearly sensitive to cetirizine. In addition, the ability of the drug to interfere with pleural neutrophil or eosinophil mobilization and mast cell degranulation seems not to be associated with its ability to block the histamine H1 receptor.


Cardiovascular effects of the 5-HT1A receptor ligand, MDL 73005EF, in conscious spontaneously hypertensive rats.

The effects of pretreatment with the potent and selective 5-HT1A receptor ligand, MDL 73005EF, on the cardiovascular responses to administration of the 5-HT1A receptor agonists, 8-hydroxy-2-(di-n- propylamino)tetralin (8-OH-DPAT), flesinoxan and 5-methylurapidil were studied in conscious spontaneously hypertensive rats (SHR) and compared with those of putative 5-HT1A receptor antagonists. MDL 7300EF (0.1-3 mg/kg) induced a dose-dependent but transient decrease in mean arterial pressure (MAP). Pretreatment with doses of 1 or 3 mg/kg MDL 73005EF significantly inhibited the hypotensive and bradycardiac effects of 8-OH-DPAT (0.03-1 mg/kg). Pretreatment with 1 mg/kg MDL 73005EF similarly reduced the hypotensive actions of flesinoxan (0.3-1 mg/kg) and 5-methylurapidil (0.1 mg/kg). In contrast, MDL 73005EF did not significantly affect the decrease in blood pressure induced by administration of 0.01 mg/kg clonidine, 0.3 mg/kg hydralazine or 0.2 mg/kg nifedipine. The effect of 8-OH-DPAT (0.1 mg/kg) on MAP was also reduced by pretreatment with 1 mg/kg BMY 7378, buspirone or pindolol, but not NAN 190 or spiperone. BMY 7378, NAN 190, pindolol and spiperone induced a significant decrease in blood pressure. To rule out the possibility that the reduced baseline may have influenced responses to 8-OH-DPAT, we showed that pretreatment with the vasodilator, hydralazine (0.3 mg/kg), had no effect on the MAP response to 8-OH-DPAT although it significantly reduced MAP. We conclude that MDL 73005EF acts as a mixed agonist/antagonist at 5-HT1A receptors since it caused a decrease in blood pressure, but also reduced the cardiovascular responses to the 5-HT1A receptor agonists, 8-OH-DPAT, flesinoxan and 5-methylurapidil.


Effect of (+)-niguldipine on myocardial alpha 1-adrenoceptors in the rabbit.

The influence of the alpha 1A-adrenoceptor subtype-selective antagonist (+)-niguldipine on the alpha 1-mediated positive inotropic effect was assessed in the isolated rabbit ventricular myocardium. (+)-Niguldipine displaced the specific binding of [3H]prazosin to a membrane fraction derived from rabbit ventricular muscle with high (Ki = 64.6 pmol/l; RH = 23%) and low (Ki = 7.08 nmol/l) affinity. (+)-Niguldipine displaced specific [3H]CGP-121177 binding only at very high concentrations (Ki = 118 nmol/l). (+)-Niguldipine at 0.1 pmol/l and higher shifted the concentration-response curve for the alpha 1-mediated positive inotropic effect downwards, but at higher concentrations (up to 10 and 100 nmol/l) it did not cause a further shift of the curve. (+)-Niguldipine (1-100 nmol/l) did not affect the beta-mediated positive inotropic effect and the basal force of concentration. (-)-Niguldipine also showed a selective inhibitory action on the alpha 1-adrenoceptor-mediated positive inotropic effect, but its affinity and potency were approximately 1-2 log units lower than those of (+)-niguldipine. The present results indicate that the alpha 1A-adrenoceptor subtype is involved in the alpha 1-mediated positive inotropic effect. (+)-Niguldipine (or (-)-niguldipine with lower affinity) is able to antagonize selectively the cardiac alpha 1A-adrenoceptor-mediated positive inotropic effect. The magnitude of the alpha 1A-mediated inotropic effect, however, may be much less than that mediated by the alpha 1B-subtype in the rabbit ventricular myocardium.


Pharmacological characterization of a 5-HT receptor in locust nervous tissue.

A 5-HT receptor in the nervous tissue of the desert locust (Schistocerca gregaria Forsk.) was investigated, using [3H]LSD (lysergic acid diethylamide) as the radioligand. [3H]LSD labels in addition a putative dopamine receptor whose specific [3H]LSD binding nevertheless could easily be diminished by co-incubation with 1 microM dopamine. The binding site was characterized by a KD of 1.64 nM, and a maximal concentration of binding sites of 79.8 fmol/mg protein. Pharmacological investigation revealed a relatively low affinity for the putative natural agonist, serotonin (KI = 0.209 microM). In contrast to the high affinity of classical serotonergic antagonists (e.g. dihydroergotamine or (+)-butaclamol) substances with subtype specificity such as 8-OH-DPAT (8-hydroxyl-1-(N,N-dipropyl)-aminotetralin) or ketanserin have only moderate affinities. Quantitative comparison of the pharmacological data demonstrated that there is obviously no pharmacological homology with vertebrate 5-HT receptors characterized so far. The only receptors with a close pharmacological relationship to the 5-HT receptor of locusts are the 5-HTdro1 receptor expressed in Drosophila nervous tissue and a 5-HT receptor in snail nervous tissue which might be homologous to that of locusts. The 5-HT receptor investigated, was shown to be G-protein-coupled, as addition of stable GTP analogues or depletion of Mg2+ ions from the incubation medium led to agonist-specific lowering of the affinity.


Prognostic significance of immunohistochemical c-erbB-2 proto-oncogene expression and nuclear DNA content in human breast cancer.

Immunoreactivity of the c-erbB-2 proto-oncogene product and nuclear DNA content were assessed in specimens from 211 breast cancer patients with a mean follow-up of 16 years (range 13-19 years). A routine immunoperoxidase technique was used and cytometrical DNA assessments were performed on cytodiagnostically identified tumour nuclei, using image analysis. C-erbB-2 cell membrane staining was observed in 29% of the cases and was found to be related to tumour size (P = 0.02), histopathological grade (P = 0.02) and nuclear DNA content (P < 0.01). In univariate analysis immunohistochemical c-erbB-2 expression was of prognostic significance among node-positive patients (P = 0.02), but not among women with node-negative disease. This prognostic ability was reduced by multivariate analysis and was no longer significant. In contrast, nuclear DNA content was significantly related to distant recurrence-free survival even in multivariate analysis after adjustment for nodal status and tumour size (P < 0.01). In conclusion, the findings of the present study indicate that c-erbB-2 expression is of limited prognostic value in a subgroup of patients, whereas nuclear DNA content seems to provide significant prognostic information even in node-negative patients.


[Beta blocking agents in the treatment of dilated cardiomyopathy: review of the literature and clinical experience with 67 patients].

Several reports suggest that chronic beta blockade, most often with the beta 1 selective agent metoprolol, may improve hemodynamic and clinical function in patients with idiopathic dilated cardiomyopathy. However, controlled trials are limited and some studies have not shown beneficial effects in short term trials. Mechanisms of effectiveness are still debated and probably concern the capacity to avoid toxic myocardial damage by catecholamines, to induce receptor up-regulation, to contribute to the control of arrhythmias, to improve diastolic relaxation and other mechanisms. After a revision of the literature, a preliminary clinical experience with metoprolol in dilated cardiomyopathy diagnosed according to the WHO definition is reported. Sixty-seven patients symptomatic for congestive heart failure or with complex ventricular arrhythmias associated with severe left ventricular dysfunction were submitted to test dose with metoprolol 5 mg bid for 2-7 days. All patients were completely studied, including coronary angiography and endomyocardial biopsy to exclude ischemic heart disease and active myocarditis. Four pts (6%) did not tolerate the first test dosage of metoprolol and twenty-two patients were excluded from analysis because of inadequate follow-up or because they were enrolled in an international trial. Forty-one patients underwent long-term treatment with metoprolol at a final mean dosage of 150 mg a day (range 50-200 mg) and are presently analyzed. The dosage was gradually increased during the first seven weeks. After 6 +/- 2 months and 12 +/- 2 months, 34 patients were stable or ameliorated (Group 1) and experienced an overall significant improvement of functional class (all pts in class I-II NYHA), of left and right ventricular ejection fraction (from 28 +/- 8.8% to 35.8 +/- 13.7% to 33.2 +/- 12.3% and from 38.6 +/- 11.8% to 42.4 +/- 5.8% to 45.2 +/- 12.2% respectively), of clinical signs of congestive heart failure, of cardiothoracic index, of left ventricular diameters and of arrhythmic pattern. Furthermore, the rate of ventricular couplets > 20/24h and of non-sustained ventricular tachycardia changed respectively from 46% and 54% to 4% and 21% at 12 +/- 2 months. None in Group 1 died nor is any waiting for heart transplantation. Eleven patients (Group 2) did not tolerate the drug acutely (4 pts) or deteriorated during the first 6 +/- 2 months (7 pts) of the treatment. In this group a worsening or an insignificant variation of all clinical and instrumental parameters was observed. During follow-up four patients of this group underwent heart transplantation (one died shortly after the operation because of infective complications), one died while waiting, two are currently waiting for heart transplantation, and three are still in heart failure (class III NYHA). No cases of sudden death occurred in any group of patients (15 pts with follow-up > 12 mo). Our uncontrolled study seems to confirm the beneficial effect of betablockers in a subgroup of patients with idiopathic dilated cardiomyopathy. The characterization of responders to this therapy is still undefined and will constitute the aim of future analyses.


Evidence for the kidney as an important source of 5'-monodeiodination activity and stimulation by somatostatin in Oreochromis niloticus L.

Radioimmunoassay of 3,5,3'-triiodothyronine (T3) and thyroxine (T4) in the thyroidal region of mature male Oreochromis niloticus revealed stores of T4 but negligible levels of T3, yielding a very low T3/T4 ratio (0.3%). 5'-Deiodination (5-D) of T4 into T3 was examined in liver and kidney homogenates in vitro by radioimmunoassay of T3 with T4 as substrate. In both organs, the 5'-D activity was temperature dependent: at 4 degrees C, T3 production was below the level of detection and maximal in both tissues at 37 degrees C; and at 45 degrees C, the enzymatic activity was reduced. T3 production seemed to reach a plateau after 60 min of incubation. The reaction required exogenous thiol cofactor (dithiothreitol) and was inhibited partially or completely by propylthiouracil depending on the concentrations used. Hepatic and renal 5'-D activities were stimulated by somatostatin (SRIF) within 4 hr, but a subsequent increase in plasma T3 was observed only when SRIF was injected together with T4, while the magnitude of rT3 production decreased. It is concluded that almost all the circulating T3 is provided by peripheral T4 to T3 conversion since T3 RIA in thyroidal follicles demonstrated insignificant T3 production. The kidney may contain the large part of the functional deiodinase which converts T4 into T3. As in mammals and unlike in other fishes, there is not only 5'-D activity, but also 5-D activity, and both may be influenced by SRIF.


Regulation of nitrogen fixation in Azospirillum brasilense Sp7: involvement of nifA, glnA and glnB gene products.

The expression of nifA-, niH- and nifB-lacZ fusions was examined in different mutants of Azospirillum brasilense. Mutations in nifA, glnA and glnB severely impaired the expression of nifH- and nifB-lacZ fusions. By contrast, a nifA-lacZ fusion was not affected in a nifA or a glnB background and was only partially impaired in glnA mutants. It is proposed that in A. brasilense, the PII protein and glutamine synthetase are involved in a post-translational modification of NifA.


The autolytic ('suicidase') system of Enterococcus hirae: from lysine depletion autolysis to biochemical and molecular studies of the two muramidases of Enterococcus hirae ATCC 9790.

Autolysis of Enterococcus hirae ATCC 9790 is the result of the action of endogenous enzymes that hydrolyze bonds in the protective and shape-maintaining cell wall peptidoglycan. It is thought that these potentially suicidal enzymes play a positive role(s) in wall growth and division and are expressed as autolysins when cell wall assembly and/or repair are inhibited. E. hirae possesses two potentially autolytic enzymes, both of which are muramidases. Although they hydrolyze the same bond as hen egg-white lysozyme, both are high-molecular-mass, complex enzymes. Muramidase-1 is synthesized as a zymogen, requiring protease activation. It is a glucoenzyme that is also multiply nucleotidylated with an unusual nucleotide, 5-mercaptouridine monophosphate. Muramidase-2 is almost certainly a product of a separate gene. The deduced amino acid sequence of a cloned gene for extracellular muramidase-2 showed several unusual features. It appears to be a two-, or perhaps three-domain protein with a putative glycosidase-active site near the N-terminal end and six 45-amino-acid-long repeats at the C-terminal end which are presumed to be involved with high-affinity binding to the insoluble peptidoglycan substrate. Muramidase-2 binds penicillin with low affinity. The presence of several amino acid groupings characteristic of serine-active site beta-lactam-interactive proteins is consistent with the possible presence of a penicillin-binding, third domain. Indirect evidence consistent with a role(s) for these enzymes in cell wall growth and division has been obtained. However, proof of such role(s) awaits modern genetic, molecular, and biochemical analyses.


Penetration of fimbriate enteric bacteria through basement membranes: a hypothesis.

A mechanism for penetration of basement membranes by Escherichia coli is presented. The mechanism is based on the ability of the S fimbriae of meningitis-associated E. coli to bind to vascular endothelium and choroid plexuses in brain and to basement membranes. On the other hand, the S and the type 1 fimbriae of E. coli immobilize plasminogen and tissue-type plasminogen activator; this process generates proteolytic plasmin activity on the surface of fimbriate cells. Our hypothesis is that bacterium-bound plasma activity, directed to basement membranes through fimbrial binding, promotes bacterial penetration through basement membranes.


Bacterial flagellar diversity and significance in pathogenesis.

Bacterial flagella are structurally diverse, ranging from the thoroughly investigated model examples found in Escherichia coli and Salmonella typhimurium to the more exotic sheathed flagella of, for example, Helicobacter pylori, and the complex multi-flagellin endoflagella found in many spirochaetes. We summarize some of the emerging structural and genetic findings relating to these more novel flagellar types, and outline their possible significance in the pathogenicity of some medically important bacteria.


Structural analysis and biological significance of the cell wall lytic enzymes of Streptococcus pneumoniae and its bacteriophage.

The development of an appropriate technique for the identification of autolysin-defective mutants of pneumococcus has been a fundamental step to carry out studies on the molecular characteristics of the lytic enzymes of Streptococcus pneumoniae and its bacteriophage. Our results show that the principal pneumococcal autolysin (an amidase) is responsible for the separation of the daughter cells at the end of the cell division. On the other hand, this system provides a reliable experimental model to support the extended idea concerning the modular organization of most proteins. The comparative analyses of the deduced amino acid sequences of these enzymes, as well as the construction of functional chimeric phage-bacterial enzymes, demonstrate that the C-terminal domain, which contains a large number of repeated amino acid motifs, is the substrate-binding domain, whereas the N-terminal domain provides enzymatic specificity. We propose that the pneumococcal lytic enzymes have evolved by modular exchange providing examples of the types of novel genes that the bacteria or the phage might create to allow them to become adapted to new environmental situations.


[Hygienic evaluation of the work of technical school students on the video display terminals].

Psychologic and hygienic examination of students working on display calculating complexes (DCC) DCC-2 and DCC-3 during the extra study time revealed the features of their functional status caused by the DCC design and work schedule. Positive role of eye exercises during the work was stressed. 6-hours duration of students' work was considered as unfavourable. The duration of work must be reduced to 3 hours, prophylactic measures (eye and physical exercises) are to be added.


AIDS and hemophilia: experience in the La Paz Hemophilia Center.

435 hemophiliacs are usually being attended in the La Paz hemophilia Center (Madrid, Spain). 257 (59%) of these patients have been infected by the human immunodeficiency virus (HIV-1) because of human plasma derivate substitution therapy. The infection has been more frequent among the severely affected patients and among the most treated patients. 82% of the infected patients are between 14 and 40 years old. By December 1991, 95 (37%) of 257 seropositive patients have developed full-blown AIDS. The most frequent opportunistic infection they had suffered was esophageal candidiasis. Looking for an evolution marker, we can point that the patients older than 35 years with CD4 levels below 200/mm3 had the worst prognosis. There was no difference in the evolution among the patients aged below 17 and those aged between 17 and 35 years. The amount of concentrate used between 1980 and 1984 did not hold any relation to the evolution. 49 patients (51%) of the 95 suffering from AIDS had died by December 1991. The evolution to the death was unrelated to the patient age, CD4 lymphocyte levels, and amount of substitution therapy. In our opinion, the most valuable marker could be the kind of opportunistic infection or tumor the patient suffers from. Finally, Retrovir has demonstrated to be useful in increasing the survival rate of the patients, but after 36 months of treatment, only 33% of those AIDS patients who began taking it remained alive. Retrovir was also used in asymptomatic patients, and during an average period of time of 15 months, a lesser bone marrow toxicity and a stabilization in CD4 lymphocyte levels could be observed, but this was unable to modify the disease progression in those patients who presented circulating p24 antigen.


Plasma protein C as a marker of hepatocellular damage in alcoholic liver disease.

The synthesis of a number of clotting factors takes place in a hepatocyte. Therefore, measurement of these factors in the blood have proved to be of additional value in the diagnosis and follow-up of liver diseases. In the present study we evaluated protein C level in the plasma of patients with liver cell damage due to chronic alcohol consumption. A decrease in plasma protein C concentration which correlated with clinical performance of the patients was found. Significant correlations between the level of protein C and antithrombin III, one-stage prothrombin time, factors VII and X and some biochemical tests reflecting liver cell damage were also stated. The obtained data indicate that plasma protein C level may constitute a useful marker of hepatocellular disease in alcoholics.


Class I gene contraction within the HLA-A subregion of the human MHC.

Individuals expressing either the HLA-A24 or the HLA-A23 histocompatibility antigens have been found to possess an HLA-A class I subregion approximately 50 kb smaller in size than those studied from individuals expressing other HLA-A haplotypes. This originally manifested itself as a haplotype-associated size variation in the NotI and MluI megabase fragments observed on pulsed-field electrophoresis gels after blotting and probing with HLA-A subregion-specific genomic probes. The contracted region falls between the HLA-A and the HLA-G class I genes and specifically includes the novel HLA-A-related pseudogene, HLA-H, as well as the adjacent deteriorated class I pseudogene, 7.0 p. The intactness of locus D6S128, defined by probe pMC6.7 located telomeric to the HLA-H gene, demonstrates that the distal rearrangement point falls within a 20-kb stretch of DNA separating HLA-H from pMC6.7. This extends a previous report regarding variation in class I gene number within the human major histocompatibility complex and precisely localizes the genomic residence of sequences that may define a recombination hot spot. Because the size variation maps to a recombinogenic area, its characterization may ultimately reveal important biological information relevant to the events that shaped the organization of the human HLA class I multigene family.


Characterization of the murine Icam-1 gene.

Intercellular adhesion molecule-1 (ICAM-1, CD54) is a cell adhesion molecule that interacts with the leukocyte beta 2 integrins, LFA-1 and Mac-1. Murine inflammatory models are being utilized increasingly to define the role that ICAM-1 induction plays in the initiation of inflammation. We have isolated murine genomic clones that contain the Icam-1 gene including over 2 kb of 5' flanking sequence. The gene for murine Icam-1 spans over 13 kb and is composed of seven exons and six introns. Each of the extracellular immunoglobulin-like domains of ICAM-1 is encoded by a single exon that ends with the first base of the next codon. Examination of ICAM-1 expression in vivo shows that mRNA levels of ICAM-1 are low in all organs except for the lung but increase markedly in multiple organs at 3 h after administration of endotoxin. The 5' flanking region of the murine gene contains a putative TATA box and potential SP-1, AP-1, and AP-3 sites in positions nearly identical to those in the human gene.


Genetic mapping of a new homeobox gene to mouse chromosome 7.

A newly identified homeobox gene designated Dbx has been mapped to mouse Chromosome (Chr) 7. This gene is expressed in a restricted manner in developing mouse brain and spinal cord and has amino acid sequence similarities with members of the homeobox gene family such as Drosophila H2.0 and mouse Hlx. Using a fragment of the Dbx cDNA as a probe, a PstI restriction fragment length polymorphism was used to determine genotypes of 144 progeny from an interspecific backcross. Segregation analysis revealed linkage of Dbx with six prepositioned reference loci on mouse Chr 7. No recombination was observed between Dbx and Odc-rs6, indicating that Dbx lies approximately 25 cM distal to the Chr 7 centromere in a region that has conserved linkage relationships with regions of human Chrs 11 and 19.


The gene for dominant white color in the pig is closely linked to ALB and PDGRFRA on chromosome 8.

White is a widespread coat color among domestic pig breeds and is controlled by an autosomal dominant gene I. The segregation of this gene was analyzed in a reference pedigree for gene mapping developed by crossing the European wild pig and a Large White domestic breed. The gene for dominant white color was shown to be closely linked to the genes for albumin (ALB) and platelet-derived growth factor receptor alpha (PDGFRA) on chromosome 8. An unexpected phenotype with patches of colored and white coat was observed among the F1 and F2 animals. The segregation data indicated that the phenotype was controlled by a third allele, denoted patch (Ip), most likely transmitted by one of the Large White founder animals. It is shown that the ALB, PDGFRA, I linkage group shares homologies with parts of mouse chromosome 5, human chromosome 4, and horse linkage group II, all of which contain dominant genes for white or white spotting. Candidate genes for the dominant white and patch mutations in the pig are proposed on the basis on these linkage homologies and the recent molecular definition of the dominant white spotting (W) and patch (Ph) mutations in the mouse.


Enhanced proliferative cellular responses to HIV-1 V3 peptide and gp120 following immunization with V3:Ty virus-like particles.

The induction of CD4+ T-helper (Th) cell responses is likely to be an important requirement of vaccine candidates designed to prevent or moderate human immunodeficiency virus-1 (HIV-1) infection. We have investigated the ability of hybrid Ty virus-like particles carrying the V3 loop region of the HIV-1 IIIB envelope gp120 (V3:Ty-VLP) to elicit V3-specific proliferative responses. Significant proliferation in response to stimulation in vitro with homologous IIIB V3 peptide was observed following immunization of mice with V3:Ty-VLP either as an aluminium hydroxide precipitate or without adjuvant. Responses to MN V3 peptide were also observed in certain mouse haplotypes. To assess the effect of presenting the V3 loop in this particulate form, we compared the responses induced by V3:Ty-VLP with those obtained with two non-particulate immunogens, recombinant gp120 (rgp120) and V3 peptide conjugated to albumin. V3-specific responses to V3 peptide in vitro were reproducibly higher following immunization with V3:Ty-VLP than with either rgp120 or V3-albumin coagulate (V3-alb). The data indicate that immunization with the V3 loop as a hybrid Ty-VLP results in enhanced proliferative responses to V3 peptide and recognition of rgp120 in vitro. Some cross-reactivity of Th cells for V3 sequences from different isolates was also observed.


Subpopulations of guinea-pig T lymphocytes defined by isoforms of the leucocyte common antigen.

This report presents the characterization of three mouse monoclonal antibodies (mAb) reactive with the guinea-pig leucocyte common antigen (LCA); CD45 in the human nomenclature. One, IH-1, reacted with LCA on all leucocytes. The other two were more restricted: IH-2 recognized only the 220,000, 210,000 and 195,000 MW isoforms, and IH-4 the 220,000, 210,000 MW isoforms. Both IH-2 and IH-4 reacted with all B cells and all Kurloff cells [the putative guinea-pig natural killer (NK) cell]. IH-2, but not IH-4, reacted with monocytes and macrophages. Neither reacted with neutrophils. Most thymocytes expressed low levels of the IH-2 and IH-4 epitopes, with those expressing high levels located predominantly within the medulla. Most (90%) CD4+ T cells from newborn guinea-pigs expressed high levels of the IH-2 and IH-4 epitopes; this percentage decreased with age to 70% in 2-year-old animals. We have demonstrated that CD4+ T cells which express low levels of the IH-2 epitope also express low levels of the IH-4 epitope. CD8+ T cells can be divided into two subsets by IH-4 but not IH-2. The reactivities of IH-2 and IH-4 are remarkably similar to those of human anti-CD45RB and anti-CD45RA antibodies respectively. Analogies with man and other species suggest important functional differences for subpopulations of guinea-pig T lymphocytes defined by anti-CD45R antibodies.


Studies on the susceptibility to NK-mediated lysis and the simultaneous expression of various surface molecules in anthracyclin-treated K562 cells and in four K562 cell clones.

Target molecules for NK cells are unknown. Numerous studies have proposed putative target molecules, but have examined their role in the modulation of sensitivity to NK-mediated lysis one independently of each other. We examined the simultaneous expression of various surface molecules and the susceptibility of K562 cells to NK attack. We have previously shown that adriamycin (40 nM) and aclacinomycin (15 nM) can induce, in vitro, an increase of glycophorin A (GPA) on K562 cells, a modulation of transferrin receptor (TfR) and CD15 antigen expression and a significant resistance of cells to NK-mediated lysis. In the present work, Fc gamma receptor II (CD32) expression at the K562 cell membrane was clearly decreased after aclacinomycin-treatment but was unaltered by adriamycin-treatment. Four K562 cell clones were studied. Two clones (F and G) expressed a higher level of CD32 at the membrane (62% and 70% of erythrocyte antibody (EA) rosettes respectively) and two clones (9 and 19) expressed lower a level (18% and 7% EA rosettes respectively) than the original population (43%). The sensitivity to lysis by NK cells was increased in clones F, G and 9 but decreased in clone 19 (without alteration in the binding capacity). Relationships between the sensitivity to NK attack and the levels of simultaneous expression of CD32, TfR, CD15, glycophorin A (GPA) and MHC class I monomorphic antigens were studied. In addition, the presence at the membrane of some cellular adhesion molecules (CD54, CD58, CD29, CD18, CD56) was examined in anthracyclin-treated cells and in the four clones. The difference in the sensitivity of target cells to NK attack is not strictly related to variation of one or other of these molecules. Our previous and present data suggest that the resistance of K562 cells to NK cells may correlate with the level of erythroid maturation at the cell membrane, involving simultaneous variations in expression of several molecules such as a decrease of TfR, CD15 and CD32 and an increase of GPA.


Oral contraceptives: an epidemiological perspective.

Oral contraceptives (OCs) containing a fixed dose of estrogen and progestogen in a 21-day regimen initially were approved for unrestricted use in the 1960s in the United States. OCs have been used and studied extensively for more than 30 years. They have always been seen as providing excellent efficacy. However, estrogens were associated with major risks in the use of OCs. In the early 1960s, case reports of thromboembolism in women using OCs led to epidemiological studies suggesting a correlation between deep vein thrombosis and the estrogen content of the pill. Research focused on decreasing the estrogen dose. In the evolution process, the combination monophasics of the early 1960s gave way to the sequential pill and the "mini pill" of the 1970s. Sex steroids given in the form of OCs were shown to alter lipoprotein and carbohydrate metabolism. This, together with the increased risk of thromboembolism, led to increased risk of cardiovascular disease, e.g., myocardial infarction. Little consideration (in the early 1960s) was given to the effects of potent progestogens and their interaction with estrogens. Hence, the recent focus of clinical research is the development of new and improved progestational agents. These third-generation gonane types appear to have little impact on carbohydrate or lipoprotein metabolism while maintaining excellent efficacy and cycle control. They should reduce the risk of serious side effects. However, to understand how and why these agents evolved, tracing the history of OC development is valuable. It might allow us to determine the baseline dose where we will begin to lose some of the OCs' benefits.


The underrated benefits of oral contraception: consequences of pregnancy and induced abortion in teenagers.

If complications occur within a pregnancy planned and brought to term, they often can be dealt with and accepted. They are even more traumatic when they occur in an unwanted pregnancy that could have been prevented through contraception. Teenagers, because of their physical and psychological immaturity and also because of their social environment, seem to suffer with undue frequency from the complications of induced abortion. Its result, for the teenager, is a handicapped future in comparison to other women. Hence, access to contraception is important for all women, and especially for teenagers, in order to avoid such prejudicial situations. It is important, then, to prescribe oral contraception for its efficacy and its short- and long-term innocuousness. Because of her immaturity, the pregnant teenager is at risk: of spontaneous abortion, pre-eclampsia, anemia, hemorrhage, and prematurity. She is also at risk because of the social difficulties she will be facing. This is particularly true in families from developing countries. From birth, the child is also at risk: of low birth weight for the term, mortality in the first year of life, and all risks linked to abandonment, or education by a third party. In a proportion of 13 to 30% in western countries and in a proportion of 3% in East Asia or in Northwest Africa (Maghreb), induced abortions are a reflection of the following: early sexual activity without contraception even if fertility is still low in very young teenagers, absence of social protection or social independence, refusal of forced marriage, and presence or absence of liberal legislation.(ABSTRACT TRUNCATED AT 250 WORDS)


The androgenicity of oral contraceptives: the young patient's concerns.

Sexual activity is quite common among women aged 14 to 20 in developed countries, averaging perhaps 10% at age 15 to about 70% at 19. Thus, the need for contraception may begin quite early in life and will continue for as long as 30 years. One of the best candidates for long-term contraception for young sexually active females is the oral contraceptive (OC), which provides health benefits besides contraception. Long-term benefits include lowered rates of ovarian and endometrial cancer, as well as of benign breast disease and ovarian cysts. Another benefit is protection against upper-tract sequelae of sexually transmitted diseases. Short-term benefits are correction of menstrual irregularity, reduction in menstrual flow, and diminished premenstrual syndrome and dysmenorrhea. Recent OC formulations contain only one-third the estrogenic potency of older OCs and therefore are associated with dramatic decreases in what were always the major side effects of OCs: heart attack, stroke, and pulmonary embolism. Other side effects of OCs have been most closely associated with the progestogenic component, and are related to the androgenic effects of progestins, particularly some synthetic progestins. However, some new synthetic progestins have been found to have minimal androgen receptor activity in preclinical testing and to cause minimal or no androgen-related side effects in clinical trials. One of these new progestins having a favorable androgenic profile is norgestimate. Its efficacy and safety in combination with low doses of ethinyl estradiol have been documented in the European and the American literature.(ABSTRACT TRUNCATED AT 250 WORDS)


Long-term profile of a new progestin.

Major complications attributable to oral contraceptives (OCs) may occur in the circulatory system. The inherent risk factors, such as race and family history, are unchangeable. Others may be altered by patient counseling and subsequent adjustment of certain behaviors. Hypercoagulability is estrogen dose related. Older, high-dose-estrogen OC users were at 40% increased risk of mortality from thromboembolic phenomena. Reduction in estrogen content has unmasked the androgenic effects of some synthetic progestogens. These effects may include progression of atherogenesis, effected through changes in cholesterol and lipoproteins; reduction in sex hormone binding globulin (SHBG), which enhances the androgenic effect; and changes in carbohydrate metabolism. This review of clinical findings is based on four studies; three had prospective cohort designs, and one was a prospective randomized comparison of a norgestimate-containing OC with a norgestrel-containing one. Numbers of subjects ranged from 20 to 59,701; the largest evaluated 343,348 cycles of treatment. Study intervals were from 4 to 24 months. The findings reported here are from the individual studies. 1. The normal value for cholesterol is less than 200 mg/dL. Of 2,197 women who met this cut-off point, 95% remained below it after 6 months of treatment. Of 756 who initially exceeded this value, 25% were below after 6 months and 75% remained above it. All studies to date have demonstrated that norgestimate produces consistent and significant elevations in high-density lipoprotein levels and variable change in low-density lipoproteins. A similar effect was noted on serum triglyceride values.(ABSTRACT TRUNCATED AT 250 WORDS)


Occurrence and specificities of alpha 3-fucosyltransferases.

The Le(x) (CD15) carbohydrate antigen and sialylated and oligomeric derivatives thereof have been implicated in cell adhesion processes. Expression of these antigens is developmentally regulated and (re)occurrence of several members of this group has been reported in malignant transformation of cells. Studies on the enzymology and genetics of alpha 3-fucosyltransferases, glycosyltransferases that play a key role in the biosynthesis of these antigens, would yield insight in the regulation of expression of these carbohydrate structures. In this paper the existing literature on these enzymes is reviewed and placed in the context of cell adhesion and malignancy.


Le(X) and related structures as adhesion molecules.

Le(x) (alpha 1-->3 fucosylated type 2 chain) functions as an adhesion molecule capable of Ca(2+)-mediated homotypic binding. Cells with high surface expression of Le(x) therefore exhibit strong self-aggregation (based on Le(x)-Le(x) interaction) in the presence of Ca2+. In this review, I have summarized several lines of supporting data for this concept, and the role of Le(x)-Le(x) interaction in the process of embryo compaction and autoaggregation of F9 teratocarcinoma cells. In general, cell adhesion events based on Le(x)-Le(x) interaction may be followed and reinforced by integrin- or Ig receptor-based adhesion systems. SLe(x), the 2-->3 sialosyl derivative of Le(x), and its positional isomer SLe(a), have been identified as the target molecules for selectin-dependent cell adhesion. Adhesion of leukocytes or tumour cells to ECs or platelets, which express E-selectin and P-selectin respectively, is initiated by this process. The target epitopes SLe(x) and SLe(a) are presented mainly on transmembrane glycoproteins having many clusters of O-linked carbohydrate chains. Therefore, inhibition of O-glycosylation may be effective for blocking selectin-mediated cell adhesion. The abundant presence of Le(x) epitope in the central nervous system, and the physiological changes of Le(x) expression as described in this monograph, reflect the adhesive properties of this molecule and its sialyosylated and/or fucosylated derivatives.


Expression of CD15 (FAL) on myeloid cells and chromosomal localization of the gene.

Different CD15 murine monoclonal antibodies were studied. These antibodies appeared to react specifically with the human myeloid-lineage-derived cell types in both peripheral blood and bone marrow. The antigens recognized by these antibodies were immunoprecipitated from lysates of 125I-labelled neutrophilic PMNs of healthy donors and subsequently analysed by electrophoresis on SDS-polyacrylamide gel and autoradiography. All antibodies precipitated the same membrane polypeptides from the membrane-iodinated PMN lysates: 105 and 150-kDa as most prominent, together with 260-, 230-, 67- and 52-kDa polypeptides. Absorption studies were performed with synthesized carbohydrate molecules. Antibody B4.3 appears to be directed against 3-alpha-fucosyl-N-acetyl-lactosamine (FAL). Competition experiments with 125I-labelled B4.3 demonstrated complete inhibition of binding by B4.3 and three other CD15 antibodies (VIM D5, UJ308, MI/N1), and partial inhibition by three additional antibodies (FMC10, FMC12, FMC13), indicating binding to the same antigenic structure. None of the antibodies reacted with monocytes using the immunofluorescence technique, but after neuraminidase digestion of these cells, positive reactions were obtained with all antibodies. Immunoprecipitation with lysates of both native and neuraminidase-digested monocytes showed no polypeptide bands. Monocytic differentiation of the myeloid cell line HL60 by 12-O-tetradecanoylphorbol-13-acetate (TPA) was accompanied by a decrease in reactivity with the antibodies, which could be reversed by neuraminidase digestion. This indicates that 3-alpha-fucosyl-N-acetyl-lactosamine is masked for the detection with antibodies upon monocytic differentiation by sialylation. Human x mouse myeloid cell hybrids were obtained after fusion of human myeloid cells and the HPRT-deficient murine myeloid cell line WEHI-TG.(ABSTRACT TRUNCATED AT 250 WORDS)


The expression of CD15 in dissociated cultured rat dorsal root ganglion cells.

This study describes the presence of CD15 in dorsal root ganglia neurons in five experimental conditions: chemically defined medium and the same medium with added nerve growth factor, retinoic acid or antibodies against insulin or tyrosine phosphate. Positive astrocyte controls were used to differentiate the monoclonal antibodies that did not react with CD15. Those monoclonal antibodies which detected CD15 in this positive control were also used to study CD15 positivity in dorsal root ganglion cells. This study shows: (i) masking of the CD15 antibody, which influences the detection capacity of the monoclonal antibodies used; (ii) that CD15 discerns two subpopulations of DRG neurons: a CD15-positive and a CD15-negative population; (iii) that CD15 expression is not involved in the outgrowth of protrusions or the wrapping by non-neuronal cells of DRG neurons.


Retinoic acid increases CD15 expression in immortalized rat astrocytes.

We have studied the expression of the CD15 (3-fucosyl-N-acetyl-lactosamine) epitope on immortalized astroglial cells derived from embryonic (E 19/20) rat brain. Immortalization was achieved by pulse-treatment of primary culture with 5-azacytidine. Seventy-three permanent cell lines were established by repeated cell cloning. Clones expressing GFAP, A2B5, and vimentin were regarded as immature astrocytes. One of these clones expressing CD15 was selected for manipulation studies. Monoclonal antibody was used for immunocytochemical detection of CD15 epitope and in immunoblot analysis. CD15 expression was visible in about 20% of the cells and was associated with a special morphological appearance. In the presence of retinoic acid the proportion of CD15-positive cells increased in a time-dependent manner, reaching about 90% within four days. Again, this expression was associated with the formation of distinct morphological features, including immunoreactive perinuclear granula, tips of processes and contact sites. After treatment with neuraminidase, all cells showed CD15-positive immunoreaction, revealing the presence of the epitope masked by sialylation. Immunoblot patterns of glycoproteins from trypsinized and mechanically detached cell preparations suggest that proteins, carrying sialylated CD15, might represent intracellular precursors of extracellularly active molecules.


Transient expression of stage-specific embryonic antigen-1 (CD15) in the developing dorsal rat spinal cord.

The localization of CD15 (synonyms: stage-specific embryonic antigen-1 (SSEA-1), 3(alpha)-fucosyl-N-acetyl-lactosamine or FAL), which is implicated in neuronal differentiation, in the developing dorsal rat spinal cord was studied by immunocytochemistry. A embryonal day 9 (E9), SSEA-1 was detected in the neural ectoderm and, at E11, in cells near the ventricle of the matrix layer. This localization indicated that SSEA-1 is present in proliferating premigratory cells of the rat spinal cord. Between E12 and E16, cells of the alar plate expressed SSEA-1. Expression of the antigen was restricted to neuroblasts that will form the dorsal horn. SSEA-1, therefore, can be used at this stage as a marker for a subdivision of the matrix layer. At E14, the dorsal root entrance zone showed SSEA-1. This indicated that SSEA-1 is associated with ingrowing primary afferents. From E16 on, SSEA-1 was present in the dorsal raphe, which suggested a function for SSEA-1 in the guidance of developing fibres. After E17, the antigen was also found within the dorsal mantle layer. SSEA-1 was first present in Rexed's laminae II, IV and V. Later on in development the antigen was detected only in Rexed's laminae II (substantia gelatinosa). These distribution patterns indicated that SSEA-1 is present on migratory and/or postmigratory cells. In addition, SSEA-1 is associated with small-diameter dorsal root fibres, the C fibres and A(sigma) fibres, that terminate within the substantia gelatinosa. After birth, SSEA-1 was present throughout the dorsal horn, probably as a result of the myelination of the fibres.


Expression of CD15 in tumours of the nervous system.

The expression of the CD15 epitope was investigated by immunohistochemistry, western blotting and immuno-thin-layer chromatography on a large series of human nervous system tumours and ethylnitrosourea-induced rat gliomas. Our results show that CD15 is expressed as a glycoprotein- or glycolipid-associated epitope in normal human and rat brain. In contrast, immunoreactivity for CD15 was absent in tumour cells of experimental rat gliomas. In human tumours we found a more complex expression pattern. While intra- and perivascular granulocytes as well as macrophages in necrotic areas of anaplastic tumours were always strongly CD15-positive, immunoreactive tumour cells were detectable only in a fraction of low-grade gliomas. Anaplastic gliomas and glioblastomas consistently did not express the epitope on their tumour cells. In addition to individual low-grade gliomas, we found CD15-positive cases among metastatic carcinomas, craniopharyngeomas, meningiomas, germinomas and malignant melanomas. Our results suggest that immunohistochemistry for CD15 is potentially useful in diagnostic neuropathology as a marker for granulocytes in paraffin sections, as a supplementary tool for the histopathological grading of gliomas, and as an aid for differentiation between anaplastic glioma cells and non-neoplastic glia. Furthermore, it can be speculated that the lack of CD15 expression on anaplastic glioma cells may potentially be responsible for some of their characteristics--such as altered cellular interaction and loss of contact inhibition.


Requirements for lysis of activated T cells by class-II-restricted cytolytic T-lymphocytes.

In the present study, we explored the specific requirements for lysis of human activated T cells by CD4+ CTLs. This was achieved by using human CD4+ T cell lines or clones specific for a peptidic fragment of influenza virus as both CTL effectors and target T cells (TTCs). Our results further establish that human activated T cells expressing HLA-DR molecules can present Ag to and be lysed by CD4+ HLA-DR restricted CTLs. This killing is Ag specific and HLA-DR restricted. It can be observed whether TTCs are heterologous or autologous, CD4+ or CD8+. However, we find that in our model: (a) TTCs are able to present artificially processed peptidic fragments of Ag, but not the corresponding natural Ag in the context of class II determinants, even if they can process whole virus in the context of class I determinants; (b) TTCs must express high density of HLA-DR molecules on their membrane; (c) preincubation of TTCs with high concentrations of peptide is required; and (d) interestingly enough, addition of free peptide at similar concentration during the cytolytic assay to replace TTC preincubation inhibits TTC lysis by at least two different mechanisms, i.e., cold-target inhibition in which CTLs serve as their own cold targets and inhibition at the effector cell level. From these results, one can conclude that stringent conditions are required for lysis of activated T cells by class-II-restricted CTLs.


Measurement of glutamate, aspartate and glycine and its potential precursors in human brain using high-performance liquid chromatography by pre-column derivatization with dimethylaminoazobenzene [correction of diethylaminoazobenzene] sulphonyl chloride.

This paper describes a high-performance liquid chromatographic technique, with dimethylaminoazobenzene sulphonyl chloride derivatization, for the measurement of glutamate, aspartate and glycine and its potential precursors in human brain tissue. The derivatization procedure is simple, sensitive and highly reproducible. The derivatized amino acids are stable and can be analysed by reversed-phase chromatography with visible detection at an absorption wavelength of 436 nm. A preliminary application to the determination of the concentrations of several amino acids in several regions of the human brain is described.


Dipeptidyl peptidase IV as a differentiation marker of the human endometrial glandular cells.

To investigate the involvement of membrane-bound peptidases in the human endometrial function, we examined the expression of dipeptidyl peptidase (DPP) IV and its enzyme activity. Immunohistological studies revealed that DPP IV was detected on human endometrial glandular cells and endometrial surface epithelium, but not on endometrial stromal cells or decidual cells in the first trimester of pregnancy. DPP IV expression on glandular cells and surface epithelium was weak in the proliferative phase, began to increase gradually in the early secretory phase, and was strong in mid-to late secretory phase and in the first trimester of pregnancy. DPP IV enzyme activity was detected histochemically in glandular cells and surface epithelium in the mid-secretory phase, and became stronger in the late secretory phase, but was rarely detected in the proliferative phase and early secretory phase. During the first trimester of pregnancy DPP IV enzyme activity in glandular cells and surface epithelium was slightly weaker than in the late secretory phase. Endometrial stromal cells and decidual cells, however, had no detectable DPP IV enzyme activity at any time throughout the menstrual cycle or during the first trimester of pregnancy. These findings indicate that DPP IV is a differentiation marker for glandular cells and surface epithelium and that active DPP IV is present in both areas during the peri-implantation period and thereafter.


Preimplantation diagnosis of a human beta-globin transgene in biopsied trophectoderm cells and blastomeres of the mouse embryo.

The preimplantation diagnosis of a HbSA-globin transgene in biopsied trophectoderm cells and blastomeres in embryos using a transgenic mouse model for the trait of human sickle-cell anaemia has been undertaken. A sensitive procedure was developed for the amplification of the human beta-globin gene sequence flanking the sickle mutation. Polymerase chain reaction (PCR) assays were undertaken on one to five biopsied trophectoderm cells and isolated blastomeres of the preimplantation mouse embryo. After biopsy the blastocysts were cultured whilst the cells were analysed for the presence of the transgene, and a high proportion (82-91%) were viable as assessed by the presence of a blastocoele cavity within a 5-h period. The majority of the biopsied cultured blastocysts were frozen and used to confirm the diagnosis; 90 biopsied cultured blastocysts were transferred to pseudopregnant recipients and 34% established pregnancy. Material from day 13.5 post-coitum fetuses was also used to confirm the original diagnosis. The time (4-5 h) required to carry out the analysis obviates a need for extended culture or cryopreservation of the biopsied embryo. In individual experiments under optimal conditions, the presence of the transgene in biopsied cells was detected with 100% accuracy, and the PCR analysis was sensitive at the 1-cell level. The overall success rate of diagnosis and confirmation of the presence or absence of the human beta-globin sequence in the biopsied embryo was 70%. Over the entire experimental period (14 months) DNA contamination from a variety of sources did occasionally occur; the methods used to overcome this problem are discussed.


Expression of aminopeptidase N and neutral endopeptidase on the endometrial stromal cells in endometriosis and adenomyosis.

Indirect immunofluorescence staining revealed that endometrial stromal cells (ESC) in the ectopic endometrium of patients with endometriosis or adenomyosis expressed aminopeptidase N/cluster of differentiation (CD) 13 antigen and neutral endopeptidase/CD10 antigen, both of which are expressed on ESC in the normal endometrium throughout the menstrual cycle. Thus, ESC in the ectopic endometrium resembled ESC in the normal endometrium not only morphologically but also antigenically. Both peptidase antigens may be useful markers for the histological diagnosis of endometriosis and adenomyosis.


Placebo-controlled, double-blind study of the effects of proglumide in the treatment of schizophrenia.

A double-blind, placebo-controlled, randomized study was performed to determine whether proglumide added to ongoing neuroleptic medication was efficacious in the treatment of 32 patients with persistent positive and negative schizophrenic symptoms. Patients treated with both proglumide and placebo showed a significant improvement over the 8 weeks of the study, but no significant difference between the patients taking proglumide and those taking placebo could be demonstrated. In addition, proglumide had no effect on plasma homovanillic acid concentrations or neuroleptic drug activity. The results suggest that, at least for the dose of proglumide used in this study (15 mg/day), the addition of this particular cholecystokinin antagonist does not potentiate the antipsychotic efficacy of neuroleptic medication in patients with persistent schizophrenic symptoms.


Kinetics, brain uptake, and receptor binding of tandospirone and its metabolite 1-(2-pyrimidinyl)-piperazine.

Tandospirone is an azaspirodecanedione derivative under investigation as an antidepressant. Metabolism of tandospirone in humans and rodents leads to 1-(2-pyrimidinyl)-piperazine (1-PP), presumed to have pharmacologic activity. To determine the relative contributions of tandospirone and 1-PP after tandospirone administration, we evaluated open-field activity, pharmacokinetics, and receptor binding of tandospirone and 1-PP in a mouse model. Tandospirone significantly reduced open-field activity during 30 minutes at doses of 1-20 mg/kg. 1-PP had no significant effect on activity except for a trend toward reduction at 20 mg/kg. At 30 minutes after administration, plasma and cortex concentrations of tandospirone and 1-PP increased in proportion to dose. Plasma protein binding (free fraction) for tandospirone was 30.4%, and for 1-PP, 87.5%. Receptor binding studies indicated that tandospirone bound with high affinity to serotonin1A sites, and with low affinity to serotonin2, alpha 1, alpha 2, and benzodiazepine sites. 1-PP bound with high affinity to alpha 2 sites and with low affinity to the other sites evaluated. A "receptor occupancy index" of tandospirone cortex concentrations divided by receptor affinity suggests that after acute administration of tandospirone, effects are likely to be due to the parent compound rather than to the metabolite. Similar conclusions are likely to be correct for other azaspirodecanediones, including buspirone.


Topographic organization of cardiovascular responses to electrical and glutamate microstimulation of the parabrachial nucleus in the rat.

We examined the functional organization of the parabrachial complex (PB) by mapping the cardiovascular and respiratory responses to PB microstimulation in anesthetized rats. The PB was explored with 100 microns resolution, at threshold doses of electrical current (5 microA) and glutamate (10-500 pmols), and the locations of stimulation sites were identified by small iontophoretic or pressure injections of biocytin or Phaseolus vulgaris leucoagglutinin. Threshold doses of either L-glutamate or electrical current pulses caused pressor-tachycardic responses that mapped to the outer edge of the external lateral subnucleus while depressor bradycardic responses were elicited from stimuli near the dorsal lateral subnucleus. Pressor responses persisted in paralyzed, ventilated animals and were thus not dependent upon concomitant respiratory changes. Cardiac arrhythmias sometimes occurred during large pressor responses and during augmented breaths that occurred during or following PB stimulation. These observations indicate that the PB contains at least two distinct neuronal systems that are potently and opposingly involved in cardiovascular control. The locations of the sites giving the most potent responses implicate specific ascending and descending pathways as substrates for the cardiovascular responses.


Postnatal development of tyrosine hydroxylase immunoreactive amacrine cells in the rabbit retina: II. Quantitative analysis.

Tyrosine hydroxylase (TH)-immunoreactive (IR) amacrine cells of the rabbit retina mature during the first four postnatal weeks, and their cellular development is described in the preceding paper (Casini, G., and N.C. Brecha, J. Comp. Neurol. 326:283-301, 1992). The present investigation is a quantitative analysis of the postnatal development of the TH-IR amacrine cell population. TH-IR amacrine cells gradually increase in size from birth (soma area of 44.7 +/- 12.4 microns2, mean +/- standard deviation) to adulthood (144.2 +/- 28.0 microns2). Cell density slightly increases from postnatal day (PND) 0 (41.9 +/- 9.5 cells/mm2) to PND 6 (47.2 +/- 7.2 cells/mm2), then markedly decreases from PND 6 to adulthood (17.8 +/- 5.3 cells/mm2) as a consequence of retinal growth. TH-IR cell number almost doubles from PND 0 (about 4,100 cells/retina) to adulthood (about 7,850 cells/retina). The increase in the total number of TH-IR amacrine cells can be explained by the generation of new TH-IR cells in the inner nuclear layer, a delay in the expression of the TH phenotype after neurogenesis by cells committed to be dopaminergic, or the acquisition of this dopaminergic phenotype by uncommitted cells. The development of the TH-IR amacrine cell mosaic was assessed by an evaluation of the distribution of nearest neighbor distances of TH-IR cells. There is a poor correlation between this distribution and a theoretical nonrandom distribution before PND 12. After this age, the nearest neighbor distance distribution shifts towards a nonrandom distribution, and is similar to that of the TH-IR amacrine cell population in the adult retina. The establishment of the TH-IR amacrine cell population mosaic is likely to be achieved through different interacting events, including intrinsic (e.g., genetic) factors, environmental influences, and nonuniform retinal growth. Overall, the population parameters analyzed in the present study approach adult values about the time of eye opening (PND 12) and they reach adult values by PND 26.


Verrucous lesions secondary to DNA viruses in patients infected with the human immunodeficiency virus in association with increased factor XIIIa-positive dermal dendritic cells. The Military Medical Consortium of Applied Retroviral Research Washington, D.C.

Hyperkeratotic lesions caused by varicella-zoster, herpes simplex, or cytomegalovirus occur in patients infected with human immunodeficiency virus type 1 (HIV-1). We have also observed this type of lesion with molluscum contagiosum. These cases were studied to determine whether there are any pathologic changes unique to these lesions. The cases were studied by routine microscopic examination and immunohistochemistry. Each case showed changes diagnostic of the viral infection, which was confirmed by immunohistochemical stains for herpes simplex and cytomegalovirus. In the dermis there were fewer inflammatory cells than expected, but there was an increase in factor XIIIa-positive dendritic cells. Varicella-zoster, herpes simplex virus, cytomegalovirus, and molluscum contagiosum can cause verrucous lesions in HIV-1-infected patients. These lesions may be related to an increase in factor XIIIa-positive dendritic cells.


Stress proteins and cross-protection by heat shock and salt stress in Bacillus subtilis.

Bacillus subtilis induced a set of general stress proteins in response to a salt or heat stress. Cells subjected to a mild heat stress showed a protective response which enabled them to survive otherwise lethal temperatures (e.g. 52 degrees C). In a similar way bacteria were enabled to survive toxic concentrations of NaCl by pretreatment with lower salt concentrations. A mild heat shock induced a cross-protection against lethal salt stress. The pretreatment of cells with low salt, however, was less effective in the induction of thermotolerance than a preceding mild heat stress. Three stress proteins were identified on the basis of their N-terminal amino acid sequences as homologues of GroEL, DnaK and ClpP of Escherichia coli. The role of general and specific stress proteins in the induction of thermotolerance/salt tolerance and cross-protection is discussed.


Computer-assisted pattern recognition model for the identification of slowly growing mycobacteria including Mycobacterium tuberculosis.

We present a computerized pattern recognition model used to speciate mycobacteria based on their restriction fragment length polymorphism (RFLP) banding patterns. DNA fragment migration distances were normalized to minimize lane-to-lane variability of band location both within and among gels through the inclusion of two internal size standards in each sample. The computer model used a library of normalized RFLP patterns derived from samples of known origin to create a probability matrix which was then used to classify the RFLP patterns from samples of unknown origin. The probability matrix contained the proportion of bands that fell within defined migration distance windows for each species in the library of reference samples. These proportions were then used to compute the likelihood that the banding pattern of an unknown sample corresponded to that of each species represented in the probability matrix. As a test of this process, we developed an automated, computer-assisted model for the identification of Mycobacterium species based on their normalized RFLP banding patterns. The probability matrix contained values for the M. tuberculosis complex, M. avium, M. intracellulare, M. kansasii and M. gordonae species. Thirty-nine independent strains of known origin, not included in the probability matrix, were used to test the accuracy of the method in classifying unknowns: 37 of 39 (94.9%) were classified correctly. An additional set of 16 strains of known origin representing species not included in the model were tested to gauge the robustness of the probability matrix. Every sample was correctly identified as an outlier, i.e. a member of a species not included in the original matrix.(ABSTRACT TRUNCATED AT 250 WORDS)


The effect of withdrawal of dopaminergic medication on simple and choice reaction time and the use of advance information in Parkinson's disease.

Eight patients with Parkinson's disease performed simple reaction time (SRT), uncued, partially and fully cued four choice (CRT) tasks. They were tested on two occasions; on their normal dose of dopaminergic medication and following withdrawal of such medication for an average of 14.4 hours. Disability as rated on the Webster scale was greater in the drug reduced state. Although RTs were generally slower when tested in the drug reduced state than when on medication, few differences emerged. Withdrawal of dopaminergic medication had no effect on unwarned SRT and unwarned and uncued CRT performance. Both on and off medication, the patients benefited from a warning signal presented before the imperative stimulus. In both medication states, the speeding up of RT was greatest with a warning signal presented 200 ms before S2. When the imperative stimulus was unwarned, the temporal predictability of its occurrence speeded RT more when on medication than when off. Advance movement parameter information was used by patients to pre-programme responses both on and off medication. In both medication states, the fully cued CRT was the same as SRT only with the 3200 ms S1-S2 interval. Medication state had no effect on movement time or the number of errors. It is suggested that slowness in motor readiness and motor programming may not be specific to striatal dopamine deficiency but rather a nonspecific concomitant of brain damage.


K-dependent inhibition in the dentate-CA3 network of guinea pig hippocampal slices.

1. The occurrence of potassium-dependent inhibitory postsynaptic potentials (K-IPSPs) in relation to burst discharges induced by 4-aminopyridine (4-AP; 30 microM) was studied in CA3, granule and hilar neurons in guinea pig hippocampal slices with the use of paired extra- and/or intracellular recording. 2. Slow small (2-5 mV) and large (up to 30 mV) K-IPSPs were observed in CA3, granule and in some hilar neurons during 4-AP applications in the presence of blockers for fast synaptic transmission, picrotoxin (50 microM), and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 5-10 microM). Amplitudes of K-IPSPs were linearly related to voltage, and they reversed in sign close to -100 mV, as expected for synaptic potentials generated by an increase in K-conductance. 3. In CA3 neurons, 4-AP applied in the presence of picrotoxin elicited burst discharges and K-IPSPs. CNQX blocked the burst discharge activity and increased the amplitude of K-IPSPs. 4. In granule cells, 4-AP applied in the presence of picrotoxin elicited K-IPSPs and only inconsistently small excitatory postsynaptic potentials (EPSPs). The EPSPs were blocked by CNQX, but CNQX application did not affect the K-IPSPs. However, in granule cells it could be observed that blockade of Cl-inhibition by picrotoxin in the presence of CNQX increased the amplitude of K-IPSPs. 5. In hilar neurons, 4-AP applied in the presence of picrotoxin elicited mainly burst discharges. CNQX blocked the burst discharges only in a few cells. In most hilar neurons K-IPSPs were observed at the beginning of the 4-AP effect, but subsequently K-IPSPs were replaced by burst discharges. 6. To determine the type of cells that burst in picrotoxin and 4-AP, neurons were stained intracellularly with horseradish peroxidase. Neurons stained in the granule cell layer did not burst and were morphologically identified as granule cells. Neurons stained in the hilar region burst and were nonpyramidal, nongranule cells. Bursting cells stained in the CA3 area were all pyramidal cells. 7. The hilar neurons varied considerably in size and dendritic organization. They could be classified as aspiny and spiny cells, the latter including mossy cells. 8. We conclude that K-dependent inhibition may explain the long-lasting IPSPs observed in in vivo recordings from hippocampal cells. In a hippocampal lamella, burst discharge activity of hilar neurons including presumed excitatory mossy cells is associated with inhibition of granule cells.(ABSTRACT TRUNCATED AT 400 WORDS)


Role of sympathetic innervation in the feline continence process under natural filling conditions.

1. The effect of the sympathetic innervation on the bladder detrusor muscle was assessed in pentobarbitone-anesthetized cats by measuring the changes in bladder wall tension that occurred during sympathetic ganglion blockade after filling the bladder at a natural rate. 2. At a point 60% of the way through the continence process, systemic sympathetic blockade was produced by intravenous trimethaphan, and selective blockade of postganglionic hypogastric nerve activity was produced by application of trimethaphan to the exposed inferior mesenteric ganglia. The level of blockade was monitored with an in-continuity hypogastric nerve recording. 3. During both systemic and selective ganglion blockade, there was an increase in baseline transmural bladder pressure and a decrease in the amplitude of nonmicturating contractions. 4. Although there was no change in the mean level of transmural bladder pressure during either systemic or selective blockade, there was a significant increase in the mean level of bladder wall mechanoreceptor discharge, suggesting that before the blockade sympathoinhibitory effects were greater than sympathoexcitatory effects. 5. Measurement of bladder wall mechanoreceptor discharge before and during ganglion blockade revealed a net sympathoinhibitory influence on the level of bladder wall tension under natural filling conditions. These results confirm that the detrusor muscle of the feline bladder is under both sympathoinhibitory and sympathoexcitatory influences for a significant portion of the continence process.


Differences of L-myc polymorphic patterns of neuroblastoma in patients under 1 year versus older ages: a preliminary report.

The age of the patient at the onset of symptoms or at diagnosis is generally accepted as one of the most important prognostic factors of neuroblastomas (NBs). Children less than 1 year of age have a better survival rate than older patients, but the reason for this is unknown. Forty-eight unselected NB patients were divided into two groups: less than 1 year (younger NB patient) and over 1 year (older NB patient) of age at diagnosis. Two of 12 younger NB patients and 18 of 36 older NB patients had N-myc amplification in their tumors. To elucidate further the possible genetic difference between younger and older NB patients, studies of restriction fragment length polymorphism (RFLP) of the L-myc gene was carried out in these two groups. The L-myc locus showed 2-allele polymorphism, allele L(10 kb) and S (6.6 kb), after digestion with EcoRI. Patients homozygous for L-band have been reported as individuals having less metastatic potential in some cancers. The allele frequencies of L and S in neuroblastomas of younger NB patients were 0.50 and 0.50, while those of older NB patients were 0.35 and 0.65, respectively. Although we did not determine L-myc RFLP in normal tissue of individual patients, we expect that the distribution of allele L and S is partly affected by possible allelic loss involving the L-myc region. However, the L-myc RFLP patterns in younger NB patients were the same as those of normal individuals and significantly differed from those of older NB patients.(ABSTRACT TRUNCATED AT 250 WORDS)


Receptor-mediated methylprednisolone pharmacodynamics in rats: steroid-induced receptor down-regulation.

Several approaches to receptor down-regulation were examined to extend previous receptor/gene-mediated pharmacokinetic/dynamic models of corticosteroids. Down-regulation of the glucocorticoid receptor was considered as an instantaneous event or as a gradual steroid-receptor-mediated process. Concentrations of plasma methylprednisolone, free hepatic cytosolic receptors, and the activity of hepatic tyrosine aminotransferase (TAT) enzyme were measured for 16 hr following administration of 0, 10, and 50 mg/kg methylprednisolone sodium succinate to 93 adrenalectomized rats. Receptor down-regulation was best described by a fractional decrement in the rate of return of free cytosolic glucocorticoid receptor. Predicted values for free receptor, bound receptor, nuclear bound receptor, and transfer compartments were in accord with the expected rank order values based on the high and low steroid doses. Model parameter estimates were independent of dose and described the rapid depletion of free cytosolic receptor, late-phase return of cytosolic receptor to a new baseline level that was 20-40% lower than control, and the TAT induction/dissipation pattern following steroid dosing. The microscopic association and dissociation constants describing the steroid-receptor interaction were 0.23 L/nmole per hr (k(on)) and 4.74 hr-1 (k(off)) for methylprednisolone compared to previously obtained values of 0.20 L/nmole per hr and 15.7 hr-1 for the related steroid prednisolone. The time course of TAT induction was similar to that observed previously for prednisolone. Efficiency of TAT induction was more closely related to steroid receptor occupancy than plasma methylprednisolone concentrations due to receptor saturability and receptor recycling.


Linkage analysis with chromosome 15q11-13 markers shows genomic imprinting in familial Angelman syndrome.

Angelman syndrome (AS) and Prader-Willi syndrome (PWS) have become the classical examples of genomic imprinting in man, as completely different phenotypes are generated by the absence of maternal (AS) or paternal (PWS) contributions to the q11-13 region of chromosome 15 as a result of deletion or uniparental disomy. Apparently, most patients are sporadic cases. The genetic mechanism underlying familial AS has remained enigmatic for a long time. Recently, evidence has been emerging suggesting autosomal dominant inheritance of a detectable or undetectable defect in a gene or genes at 15q11-13, subject to genomic imprinting. The present report describes an unusually large pedigree with segregation of AS through maternal inheritance and apparent asymptomatic transmission through several male ancestors. Deletion and paternal disomy at 15q11-13 were excluded. However, the genetic defect is still located in this region, as we obtained a maximum lod score of 5.40 for linkage to the GABA receptor locus GABRB3 and the anonymous DNA marker D15S10, which have been mapped within or adjacent to the AS critical region at 15q11-13. The size of the pedigree allowed calculation of an odds ratio in favour of genomic imprinting of 9.25 x 10(5). This family illustrates the necessity of extensive pedigree analysis when considering recurrence risks for relatives of AS patients, those without detectable deletion or disomy in particular.


[Taxol and related diterpenoids of Taxus sp.: important acquisitions in the treatment of cancer].

Phytochemical and pharmacological studies on Taxus sp extracts have resulted in the isolation and the identification of several diterpenoids, and the discovery of the potent antitumor activity of taxol. This natural compound displays a unique mechanism of action as it stabilizes microtubules and inhibits their depolymerization into free tubulin. The emergence of taxol from the screening of natural products shows the benefits and the potent interest of these constituents in the search of drugs exhibiting novel mechanisms of action. The most pressing problem in taxol or taxol derivates commercialization is the drug supply. Alternative sources of taxol are now under investigation, mainly the total synthesis of taxol, the hemisynthesis of taxol or of taxotere from 10-deacetylbaccatine III, the in vitro production of taxol by cell or tissue cultures, and the prospection of new natural sources of taxol.


Association between schizophrenia and homozygosity at the dopamine D3 receptor gene.

Disturbances in dopamine neurotransmission have been postulated to underlie schizophrenia. We report data from two independent studies of a BalI polymorphism in the dopamine D3 receptor gene in patients with schizophrenia. In both studies, more patients than controls were homozygous (p = 0.005, p = 0.008). When pooled data were analysed, this difference was highly significant (p = 0.0001) with a relative risk of schizophrenia in homozygotes of 2.61 (95% confidence intervals 1.60-4.26).


A new mutant transthyretin (Arg 10) associated with familial amyloid polyneuropathy.

We report a new kindred with systemic amyloidosis presenting as peripheral neuropathy in the sixth and seventh decades of life. Polymorphism in exon 2 of the transthyretin (TTR) gene was suggested by single strand conformation polymorphism analysis and determined by direct DNA sequencing. We also developed restriction fragment length polymorphism analysis by polymerase chain reaction using a primer with an induced mutation. The point mutation (cytosine for thymine at position 1038 of the TTR gene) is responsible for substitution of arginine for cysteine at position 10 of the TTR molecule. It is hypothesised that the TTR molecules which have no cysteine have a unique structure in heterozygous TTR polymers and are responsible for amyloid fibril formation.


Quantitative Southern blot analysis in the dystrophin gene of Japanese patients with Duchenne or Becker muscular dystrophy: a high frequency of duplications.

Eighty-four unrelated patients with Duchenne or Becker muscular dystrophy in Japan were studied by quantitative Southern blot analysis with dystrophin cDNA probes. We found partial deletions and duplications in 47 (56%) and 12 (14%) cases respectively by HindIII digestion. The duplications were confirmed by BglII digestion and densitometric scanning. The frequency of duplications in this study is significantly higher than those previously reported. This may be because of the small sample number, the racial difference, or our quantitative methods. Our results suggest that attempts to detect duplications are important for a precise diagnosis. Both deletions and duplications clustered at the two hot spots as reported previously. Six cases were exceptions to the 'reading frame hypothesis'. We detected three types of HindIII RFLP. Based on the results of one duplication case, we propose a revised sequential order of exons in the cDNA10 region of the dystrophin gene.


Presymptomatic diagnosis of von Hippel-Lindau disease with flanking DNA markers.

Von Hippel-Lindau (VHL) disease is a dominantly inherited cancer syndrome characterised by the development of retinal, cerebellar, and spinal haemangioblastomas, renal cell carcinoma, and phaeochromocytoma. The gene for VHL disease has been mapped to chromosome 3p25-p26 and flanking markers identified. We have investigated the usefulness of currently available DNA markers for the presymptomatic diagnosis of VHL disease. In the first part of this investigation, genetic linkage data from two previously published studies were updated and reanalysed to provide accurate estimates of sex specific recombination fractions and to confirm that there is no evidence of locus heterogeneity. In the second part of this study, 14 families containing 23 asymptomatic subjects at 50% prior risk of VHL disease were investigated with closely linked DNA markers (RAF1, D3S18, D3S732). Seventeen subjects were informative with one or more markers, six of whom were informative at markers flanking the VHL disease gene. By combining age related and DNA based risk information the carrier risk for 11 subjects was reduced to < 2%.


Angelman syndrome with a chromosomal inversion 15 inv(p11q13) accompanied by a deletion in 15q11q13.

A family is described in which an inversion of chromosome 15, 15 inv(p11q13), is segregating. All family members are healthy except the proband who is a 10 year old boy with Angelman syndrome. Although the chromosomal inversion has been passed from the grandfather to both his son and his daughter with no ill effect, passage from daughter to grandson has resulted in a deletion of chromosome 15 material which is presumed to be the cause of Angelman syndrome in this boy. The probabilities of an inversion of this type being instrumental in causing the syndrome are discussed.


Myocardial glutathione depletion impairs recovery of isolated blood-perfused hearts after global ischaemia.

This study was performed to determine whether depletion of myocardial glutathione would impair recovery of left ventricular function of blood-perfused, isolated hearts after reversible ischaemic injury. Cats were treated with either vehicle or buthionine sulfoximine (BSO), an inhibitor of gamma-glutamylcysteine synthetase, the rate-limiting enzyme in the synthesis of glutathione. The feline isolated hearts were perfused with the blood of normal donor cats before and after 40 min of global myocardial ischaemia. The myocardial concentration of glutathione of the BSO group, 178 +/- 38 ng/mg tissue, was significantly less than that of the control group, 292 +/- 38 ng/mg tissue (P < 0.05). The peak left ventricular developed pressure (LVDP) 1 h after reperfusion, expressed as a fraction of the peak LVDP before ischaemia, was 0.87 +/- 0.10 for the control group and 0.64 +/- 0.08 for the BSO group (P = 0.05 vs. control). The peak left ventricular dP/dt after reperfusion, expressed as a fraction of the peak dP/dt before ischaemia, was 1.08 +/- 0.14 for the control group and 0.78 +/- 0.09 for the BSO group (P = 0.05 vs. control). The myocardial creatine kinase activity of the BSO group, 1046 +/- 46 U/g tissue, was not significantly different from that of the control group, 1038 +/- 17 U/g tissue (P = 0.87). Thus, depletion of myocardial glutathione resulted in impaired post-ischaemic contractile function that cannot be attributed to a greater extent of irreversible cell injury.


Treatment of dual diagnosis patients: a relapse prevention group approach.

The authors describe the successful use of an adjunctive group psychotherapy for substance-abusing patients with major psychiatric disorders (bipolar, schizophrenia, schizoaffective, psychotic depression, and atypical psychosis). The group utilizes a psychoeducational approach that focuses on substance abuse causes and consequences, principles of recovery, and relapse prevention strategies. Eight patients with prolonged histories of abuse of cocaine, alcohol, marijuana, or other drugs were enrolled in this weekly group treatment at a community mental health center drug treatment program, while continuing in treatment with their current case manager or primary therapist. Six of the eight patients achieved periods of stable abstinence, documented by self-report, urine toxicology screens, continued group attendance, and improved social functioning. Case examples are utilized to illustrate the group process.


Treatment of khat addiction.

The authors present two cases of khat addiction that were successfully treated with bromocriptine. Khat is a bush cultivated in the Mid East because of its highly stimulant effects. Its leaves contain a variety of sympathomimetics. While khat is rarely found in the U.S., American soldiers stationed in the Arabian peninsula may be exposed to it. Because of an alcohol interdiction during the current Persian Gulf crisis, these troops may be tempted to use this plant as an alternative recreational drug.


[Possible role of cellular immunity against tubular basement membrane antigen (TBM) in gold nephropathy in rheumatoid arthritis patients].

Autoantibody formation and lymphocytes proliferative response to tubular basement membrane (TBM) antigen were examined to clarify the pathogenesis of gold nephropathy, in rheumatoid arthritis patients. The existence of tubulopathy was ascertained by urine protein analysis, electrophoresis and urine TBM antigen titration. Circulating antibody to human TBM antigen titrated by enzyme immunoassay was significantly elevated in patients with gold tubulopathy, and mitogenic stimulation with TBM antigen of peripheral lymphocytes specifically responded in the early stage after receiving gold, but then clearly decreased after the cessation of gold. But, when the lymphocytes had been passed through a nylon wool column, the reaction was remarkably high even in the later stage after receiving gold, suggesting that another suppressive population of lymphocytes became trapped in the nylon wool column. This evidence suggests that gold compounds definitely act as initiating and promoting agents, and the development of tubular disorders induced by gold are likely related to the cellular recognition of effector T cells to the TBM antigen, following the strong effect of gold on the cellular immune system.


[A case of microscopic polyarteritis nodosa associated with myeloperoxidase-antineutrophil cytoplasmic autoantibodies (MPO-ANCA)].

We report a case of microscopic polyarteritis nodosa associated with myeloperoxidase-antineutrophil cytoplasmic autoantibodies (MPO-ANCA). A 38 year-old female was admitted to our hospital, because of proteinuria, recurrent pyrexia, polyarthralgia, abdominal pain and purpura. She had a history of severe pulmonary hemorrhage and 4 kg weight loss for 8 months. On admission perinuclear ANCA without cytoplasmic ANCA was detected by indirect immunofluorescence assay and MPO-ANCA was detected by enzyme linked immunosorbent assay. But anti-nuclear antibodies, immune complexes and anti-glomerular basement membrane antibodies were not detected. Renal biopsy showed necrotizing crescentic glomerulonephritis without immune deposits. Skin biopsy revealed leukocytoclastic vasculitis. Diagnosis of microscopic polyarteritis nodosa was made by these clinical and histological evidence of vasculitis. As renal failure progressed after admission, corticosteroid and cyclophosphamide administration were started. Renal function and other symptoms improved paralleled with decreased MPO-ANCA titer to normal values. It is suggested that MPO-ANCA may be closely related to the pathogenesis of microscopic polyarteritis nodosa and it may be a good serological marker for diagnosis and disease activity of this disease.


Effects of morphine on responses of nociceptive ventrobasal thalamic neurons in diabetic rats.

The influence of diabetes on the effects of morphine on the responses of ventrobasal (VB) thalamic neurons to mechanical noxious stimuli were studied in chloral hydrate-anesthetized rats. Animals were rendered diabetic by an injection of streptozotocin (60 mg/kg, i.v.). Morphine (0.3 mg/kg), administered i.v., produced a reduction in the responsiveness of VB thalamic neurons to noxious stimulation in control rats. This effect was reversed by naloxone. In contrast, the inhibitory effects of morphine on the nociceptive responses of VB thalamic neurons were significantly attenuated in diabetic rats, as compared with the controls. However, there were no significant differences in inhibitory potency between diabetic and control rats when morphine (30 nM) was administered intrathecally. It seems likely that these changes in the sensitivity of VB thalamic neurons to morphine are, to some extent, the source of the reduction in the analgesic efficacy of morphine in diabetic rats.


Delayed suppressive effect of a low dose of caerulein on the grooming behavior induced by the D1-receptor agonist SKF 38393.

Caerulein (CLN, 0.8-80 micrograms/kg, s.c.) was administered to male rats 10 min or 24 hr before the injection of SKF 38393 (3 mg/kg, i.p.). The increased mouth movement and grooming behavior by SKF 38393 were suppressed dose-dependently by CLN 10 min before the SKF 38393. CLN at the dose of 0.8 micrograms/kg, given 24 hr before the SKF 38393, suppressed the grooming behavior by SKF 38393. These findings suggest that a low dose of CLN, but not a high dose, had a delayed suppressive effect on the grooming behavior induced by an excess of D1-activity.


[The effects of autonomic agonists on the female rabbits urethra--in vitro isovolumetric and isometric study].

To investigate the characteristics of adrenoceptors of the proximal urethra in female rabbits, we performed the in vitro isovolumetric urethral pressure study and isometric study with three parts of muscle strips of the proximal urethra (whole layers, inner layers and outer layers). Both alpha-1 and alpha-2 agonist caused dose dependent response in in vitro isovolumetric pressure study as well as in vivo study. However, the response of alpha-2 agonist in in vitro was small in magnitude compared to in vivo study, suggesting the influence of permeability of drugs from serosa to mucosa in in vitro whole urethra study. The response of the strips of inner layers to alpha-1 agonist is almost the same as that of outer layers and whole layers, while the response of inner layers to alpha-2 agonist is almost twice as that of other layers. These findings are suggestive of predominance of alpha-2 adrenoceptors mediating contractions in the inner layers of the proximal urethra in female rabbits. Alpha-2 adrenoceptors which probably are distributed in mucosal and submucosal layers may have an important role in the mechanism of urinary continence in female rabbits.


[Microsurgical autotransplantation of abdominal testis].

A 32-year-old man who had bilateral non-palpable testis underwent microsurgical orchidopexy of the right abdominal testis. Inferior epigastric vessels were used for the recipient vessels. Ischemic time during the operation was 40 minutes. Postoperative doppler readings revealed excellent blood flow to the testis, and HCG-stimulating test revealed good response. Now, 2 years after surgery, no sign of atrophy is found.


[Acute pain in surgery: the significance of a neglected problem].

Acute pain represents a significant problem in surgical patients. However, the management of acute pain in Germany is unsatisfactory, mostly because surgeons are not interested in the pain of their patients, and anesthesiologists do not give pain treatment on surgical wards. The aim of this article is therefore to point out the significance of the problem of "acute pain" for surgeons. Basic knowledge of the production, perception and projection of pain as well as the special aspects of acute versus chronic pain is mandatory. Every surgeon should know about the predictors of postoperative pain, including the surgical and anesthesiological factors and patient characteristics. Efficient management of acute pain requires knowledge of the clinical significance of pain and the different methods for assessment. It is not sufficient to know the methods and advantages of appropriate management of acute pain; one must also understand the dangers. Problems caused by the treatment of pain should be recognized from a clinical point of view. Surgeons must take a greater interest in the problem of "pain", which should lead to the establishment of new concepts in the management of acute pain in surgical patients.