[Six semi-sold media methods for detecting motility of gram negative bacilli]. Motility is recognized as a significant biological character of certain bacteria, and is used as a fundamental basis of classification in many taxonomic systems. We compared wet mount method, semi-solid medium method and flagella stain method to evaluate the motility activity of 538 Enterobacteriaceae and 300 glucose non-fermentative gram negative bacilli. The results showed a sensitivity of 100% in flagella stain, Gilardi medium, Mueller Hinton semisolid medium and sulfide-indole-motility (SIM) medium (Difco); 99.6% in SIM (Kyokuto) and SIM (BBL); 99.3% in motility test medium (BBL) and 97% in wet mount. Motile Enterobacteriaceae grow well, and turbidity changes clearly and is easy to interpret. Motile glucose nonfermentative gram negative bacilli grow so lightly and rapidly diffuse throughout the medium and are hardly to interpret. It is better to use colorless Gilardi medium and Mueller Hinton semi-solid medium and to read within 4-8 hours or 24 hours. The advantages of the semi-solid medium method are particularly evident in teaching schedules and routine testing, because the results are cumulative, macroscopic, highly sensitive, and easy to manipulate.


[Meniscal repair with autogenous periosteum and fibrin adhesive system]. It is well recognized that meniscal tear in the avascular portion is difficult to heal. The purpose of this study is to evaluate the ability of periosteum and fibrin adhesive system (FAS) in promoting the meniscal repair in the avascular zone. Full-thickness defects of 1 x 3 mm were created in the avascular portion of lateral menisci in adult mongrels. The control menisci were left without any treatment. The experimental defects were either repaired with autogenous periosteum or the combination of periosteum and FAS. Histologic and ultrastructural evaluation of the reparative tissue were compared at 4, 8, and 16 weeks postoperatively. The reparative tissue in the control group remained fibrous even at week 16, whereas that in the grafted and graft/FAS groups resembled normal adjacent fibrocartilage. Moreover, the regenerated chondrocytes in the graft/FAS group appeared more mature than those in the grafted group. Our results suggest that periosteum is a potential source of primitive cells from which chondrocytes could be differentiated and developed during the reparative process of meniscal fibrocartilage. As to FAS, it not only acted as a hemostatic agent and a fixant, but also could indirectly accelerate the chondrogenesis of periosteum in the meniscal defects. It is sufficient to conclude that the combination of periosteum and FAS in promoting meniscal defects or severe tears in the avascular portion bears its potential in future clinical application.


[The role of total gastrectomy in the management of gastrinoma syndrome]. Total gastrectomy used to be considered as a standard operation in the surgical treatment for gastrinoma syndrome. Although this end organ resection is a good palliation, to cure this ulcerogenic disease, gastrinoma resection is preferred. It remains a question if a total gastrectomy is necessary when a gastrinoma is found and resected. In a period of 12 years, nine out of 17 patients with gastrinoma syndrome were surgically treated. Fifteen gastrinomas were removed without histologic evidence of malignancy. Total gastrectomy was added in three, Whipple's operation was done in one and tumor resection alone in five. Serial examination of the resected stomach didn't find any gastrinoma. Eight gastrinomas of seven patients were found located in the pancreas, duodenum and lymph nodes within the gastrinoma triangle. These patients except one were followed up for six months to five years (mean 32 months). One died of post-operative hepatic failure, and one died of hypophysectomy for a co-existent huge prolactinoma (MEN I) four years later. Cure of the disease in terms of normal fasting serum gastrin levels, symptom free without medication, endoscopic evidence of healed ulcer, and a negative conversion of secretin provocative test was found in 2/3 of the patients with additional total gastrectomy, and 4/6 of the others. All cured patients had their gastrinomas located in the gastrinoma triangle, and six of the seven (85.7%) patients with gastrinomas in the triangle were cured. It is our conclusion that total gastrectomy was retrospectively unnecessary in most of the patients with gastrinomas in the gastrinoma triangle. Staged operations should be planned.(ABSTRACT TRUNCATED AT 250 WORDS)


[Clinical analysis of adenocarcinoma of the stomach in patients with duodenal ulcer]. In 332 patients of adenocarcinoma of the stomach and 7095 patients with duodenal ulcer collected in 5-year period, 21 cases were found to have both disease entities. With regard to clinical manifestations, gross morphology, microscopic differentiation and management of adenocarcinoma of the stomach, the patients with duodenal ulcer showed no significant difference from those without duodenal ulcer. The activities of the duodenal ulcers could be in acute active stage or present as chronic scar only. The ulcer history varied from several years to more than 15 years. We concluded that the frequency of adenocarcinoma of the stomach developed in patients with duodenal ulcer is not so rare, presentation of epigastric discomfort or signs of UGI bleeding in patients with duodenal ulcer history not always indicated recurrence of ulcer disease.


[Evaluation of marrow neutrophilic reserve in normal subjects and aplastic anemia patients]. The widespread use of potent marrow suppressing drugs in malignant diseases has gained increasing importance in the functional assessment of neutrophil system. A peripheral neutrophil count of 500 cells/cumm is considered to be a good indicator of risk of bacterial infection. However, despite extreme neutropenia, infection is rare in chronic benign neutropenia patients. It may be presumed that adequate marrow reserve delivers mature neutrophil during bacterial infection in these patients. Currently available methods such as determination of blood neutrophil concentration, differential counts, estimation of bone marrow cellularity provide a relatively crude estimates of the functional capacity of the neutrophil system. The use of endotoxin, etiocholanolone and steroids have been introduced to determine the marrow neutrophil reserve with greater accuracy. In this study, intravenous hydrocortisone 100 mg was used to evaluate marrow neutrophilic reserve in normal and aplastic anemia subjects. The results showed significant statistical differences (P less than 0.001) between controls and aplastic subjects in 2, 3, 4 and 5th hour groups after steroid injection. The sensitivity and specificity occurring 3 and 4 hours after steroid injection were 78.6% and 100%, respectively. The normal marrow neutrophilic reserve showed more than 1198/cumm of increment neutrophilic counts 4 hours after steroid injection. No complication was found. The present finding suggests that steroid may be useful as an accurate evaluator of marrow neutrophilic reserve.


[Clinical study of Urocit-K: a slow releasing potassium citrate]. Citrate is recognized as one of the inhibitors of urinary stone formation. If the urinary citrate and pH could be elevated by the supply of potassium citrate, the recurrence of stone formation would hopefully be prevented. Urocit-K is a kind of slow releasing preparation of potassium citrate. The pharmacokinetics of Urocit-K is investigated in this study, which included two groups: 1) the hypocitraturia group consisting of twenty patients and 2) the normal volunteer group consisting of ten persons. Urokit-K was administered to patients in 10 mEq doses, t.i.d. for 2 weeks. Comparison of urinary biochemistry was done prior to, and one week after administration of the Urocit-K. In the normal volunteer group, the pharmacokinetics of Urocit-K was determined by urinary pH, K, and citrate. Determinations were made six times a day (every 2-hours for first 8 hours, 7- and 9- hours for the last 16 hours) with Urocit-K for comparison. This study indicated that Urocit-K increases urinary pH, K, and citrate in hypocitraturic patients. In the normal volunteer group, Urocit-K deserves its slow-releasing property shown by a constant increase of urinary pH and K for the first 8 hours; and urinary citrate for almost 24 hours. No apparent complications, such as gastrointestinal upset or cardiopulmonary discomfort were observed. In conclusion, Urocit-K is an excellent preparation of potassium citrate and a good choice for prevention of stone recurrence.


Congenital bronchoesophageal fistula in the adult: a case report. The case of a 50-year-old woman with congenital bronchoesophageal fistula is presented. The patient had symptoms of hemoptysis provoked by ingestion of liquids for 2 years. A barium swallow examination showed a bronchoesophageal fistula communicating between the middle third of the esophagus and right intermediate bronchus. An orifice of the fistula at the esophagus was revealed by esophagoscopy. The clinical and operative features and diagnosis of congenital origin are discussed.


Delayed localized right atrial tamponade by intrapericardial hematoma after aortic valve replacement: a case report. Cardiac tamponade, early or late after cardiac surgery, is an uncommon while a potentially lethal condition in which the classic signs of tamponade may be absent. High index of suspicion and rapid diagnosis are mandatory for life-saving decompressive therapy. We herein reported a case of delayed localized right atrial tamponade caused by loculated intrapericardial hematoma 26 days after aortic valve replacement, which was recognized immediately by transthoracic two-dimensional echocardiography. The clinical status improved dramatically after surgical removal of the hematoma.


Pentazocine addict nephropathy: a case report. Medical complications associated with narcotic addiction include bacterial endocarditis, pneumonia, pulmonary embolism and renal disease. Renal disorders associated with pentazocine abuse are rarely reported. They vary with method of administration, dosage, and duration of abuse. We describe a 33-year-old male addict, using intravenous pentazocine for 6 years. He has nephrotic syndrome with a rapid deterioration of renal function to a uremic stage within 3 weeks. The laboratory data includes: IgG 1270 mg/dl, IgA 369mg/dl, IgM 326mg/dl, C'3 65.2 mg/dl, C'4 16.3 mg/dl, and serum soluble interleukin-2 receptor level (sIL-2R) greater than 6000U/ml. A renal biopsy revealed membranoproliferative glomerulonephritis (MPGN) type I with tubulointerstitial nephritis. Immunofluorescent (IF) study revealed granular deposition of C'3 and IgM in mesangium and the glomerular capillary wall. The pathogenesis of glomerular disease in drug addicts is discussed, and the literature reviewed.


[Cerebrospinal fluid rhinorrhea due to sinus surgery: a case report]. Cerebrospinal fluid rhinorrhea is an uncommon but dangerous disease. Many lethal complications, such as bacterial meningitis and pneumoencephalus, may be the result of cerebrospinal fluid rhinorrhea. Otolaryngologist, neurosurgeons and radiologists must know how to diagnose, how to localize the site of leakage and how to choose the best method of treatment. A case of cerebrospinal fluid rhinorrhea and meningitis due to improper sinus surgery is presented. Satisfactory result, such as avoidance of unnecessary brain tissue damage, can be obtained by extracranial endonasal repairing of the fistula with a composite septal flap.


Vasodilative and anti-adrenergic effects of adenosine in diabetic rat hearts. To determine the vasodilative and negative inotropic effects of adenosine in hearts of diabetic rats, isolated hearts, perfused at constant perfusion pressure (Langendorff technique), were prepared from age-matched control Wistar rats and rats made diabetic 10 weeks prior to study by a single injection of streptozotocin (65 mg.kg-1, i.p.). Adenosine and nitroprusside each increased coronary inflow when administered either as bolus injections or as infusions. Coronary flow responses to nitroprusside were unchanged in diabetic hearts. Coronary flow responses of diabetic hearts to adenosine injections were unchanged, but responses to adenosine infusions tended to be larger than in normal hearts. Diabetes had no significant effect on the EC50 for either vasodilator. Adenosine inhibited the inotropic effect of isoproterenol (enhanced left ventricular (LV) pressure (P) and LV dP/dtmax) in normal hearts, independently of its vasodilative action. This negative inotropic action of adenosine appeared equally strong in diabetic hearts. We conclude that adenosine's coronary vasodilative and anti-beta-adrenergic, negative inotropic effects in the rat heart were not diminished after 10 weeks of streptozotocin-induced diabetes mellitus. Thus, earlier reports of diminished adenosine dilative efficacy in experimental diabetes may have been unique to those particular models.


Prostaglandin E2 receptors in the heart are coupled to inhibition of adenylyl cyclase via a pertussis toxin sensitive G protein. In previous studies we have identified and isolated a prostaglandin E2 (PGE2) receptor from cardiac sarcolemmal (SL) membranes. Binding of PGE2 to this receptor in permeabilized SL vesicles inhibits adenylyl cyclase activity. The purpose of this study was to determine if the cardiac PGE2 receptor is coupled to adenylyl cyclase via a pertussis toxin sensitive guanine nucleotide binding inhibitory (Gi) protein. Incubation of permeabilized SL vesicles in the presence of 100 microM 5'-guanylamidiophosphate, Gpp(NH)p, a nonhydrolyzable analogue of GTP, resulted in a shift in [3H]PGE2 binding from two sites, one of high affinity (KD = 0.018 +/- 0.003 nM) comprising 7.7% of the total available binding sites and one of lower affinity (KD = 1.9 +/- 0.7 nM) to one site of intermediate affinity (KD = 0.52 +/- 0.01 nM) without a significant change in the total number of PGE2 binding sites. A shift from two binding sites to one binding site in the presence of Gpp(NH)p was also observed for [3H]dihydroalprenolol binding to permeabilized cardiac SL. When permeabilized SL vesicles were pretreated with activated pertussis toxin, ADP-ribosylation of a 40- to 41-kDa protein corresponding to Gi was observed. ADP-ribosylation of SL resulted in a shift in [3H]PGE2 binding to one site of intermediate affinity without significantly changing the number of binding sites. In alamethicin permeabilized SL vesicles, 1 nM PGE2 significantly decreased (30%) adenylyl cyclase activity. Pretreatment with activated pertussis toxin overcame the inhibitory effects of PGE2. These results demonstrate that the cardiac PGE2 receptor is coupled to adenylyl cyclase via a pertussis toxin sensitive Gi protein.(ABSTRACT TRUNCATED AT 250 WORDS)


Tyrosine kinase inhibitors and the contractile action of epidermal growth factor-urogastrone and other agonists in gastric smooth muscle. We examined the effects of the tyrosine kinase (TK) inhibitors, genistein, and tyrphostin (RG-50864) on the contractile action of epidermal growth factor - urogastrone (EGF-URO), transforming growth factor-alpha (TGF-alpha), and other agonists in two smooth muscle bioassay systems (guinea pig gastric longitudinal muscle, LM, and circular muscle, CM). We also studied the inhibition by tyrphostin of EGF-URO stimulated protein phosphorylation in identical smooth muscle strips. The selective inhibition by genistein and tyrphostin of EGF-URO and TGF-alpha induced contraction, but not of carbachol- and bradykinin-mediated contraction, occurred at much lower concentrations (genistein, less than 7.4 microM (2 micrograms/mL); tyrphostin, less than 20 microM (4 micrograms/mL)) than those used in previously published studies with these TK inhibitors. In LM tissue, the IC50 values were for genistein 1.1 +/- 0.1 microM (0.30 micrograms/mL; mean +/- SEM) and 3.6 +/- 0.5 microM (0.74 micrograms/mL) for tyrphostin, yielding a molar potency ratio (GS: TP) of 1:3 in the longitudinal preparation. In CM tissue, the IC50 values were 3.0 +/- 0.3 microM (0.81 micrograms/mL) for genistein and 2.4 +/- 0.2 microM (0.49 micrograms/mL) for tyrphostin, yielding a molar potency ratio (GS:TP) of 1.0:0.8 in the circular strips. The inhibition by genistein and tyrphostin of EGF-URO and TGF-alpha mediated contraction was rapid (beginning within minutes) and was reversible upon washing the preparations free from the enzyme inhibitors. In intact tissue strips studied under bioassay conditions, tyrphostin (40 microM) also blocked EGF-URO triggered phosphorylation of substrates detected on Western blots using monoclonal antiphosphotyrosine antibodies.(ABSTRACT TRUNCATED AT 250 WORDS)


Induction of C4-dicarboxylate transport genes by external stimuli in Rhizobium meliloti. We have characterized two mutants of Rhizobium meliloti L5-30 obtained by random mutagenesis using Mu-lacZ that were defective in transport of C4-dicarboxylic acids. These mutants induced ineffective nodules on alfalfa. Mutations in the two strains appeared to be located in a dctA gene. Levels of dctA gene expression were determined under different environmental conditions using dctA-lacZ fusions, and beta-galactosidase activities increased in response to osmotic stress and also when the cells were incubated in medium low in calcium. The transcriptional induction of the dctA gene by environmental signals was decreased by DNA gyrase inhibitors such as novobiocin and coumermycin.


A general method of isolating high molecular weight DNA from methanogenic archaea (archaebacteria). High molecular weight DNA was readily isolated from all methanogens treated, as well as from thermophilic anaerobic eubacteria, by grinding cells frozen in liquid N2, prior to lysis with SDS. DNA can subsequently be purified by the usual phenol-chloroform extractions. The procedure yields DNA readily cut by restriction enzymes and suitable for oligonucleotide probing, as well as for mole percent G + C content determination by thermal denaturation. The method routinely yields DNA of high molecular weight and is an improvement over DNA isolation methods for many methanogens, which often involve an initial breakage of the cells in a French pressure cell.


Herpes simplex virus and human papillomavirus sites correlate with chromosomal breakpoints in human cervical carcinoma. The distribution of 1,912 breakpoints observed in a series of 148 cervical cancers was analyzed. Fifty bands were shown to be nonrandomly involved in chromosome structural rearrangements. One hundred thirty-three breaks were noted in bands known to contain a human papillomavirus integration site, and 454 breaks were noted in bands containing a herpes simplex virus breakage site. We suggest that herpes simplex viruses and, possibly, papillomaviruses play an important role in the carcinogenesis and/or development of cytogenetic abnormalities in cervical cancers.


Chromosome fragility in lymphocytes of women with cervical uterine lesions produced by human papillomavirus. We studied 30 women with cervical lesions that showed human papillomavirus infection (HPV). Cervical HPV infection was diagnosed by cytology, histology, immunohistochemistry, and electron microscopy, as well as by DNA viral hybridization in situ with 6, 11, 16, and 18 HPV types. Three groups of patients were studied: 15 women infected by HPV of 6 and 11 types with koilocytic lesions and benign evolution, 15 women infected by HPV of 16 and 18 types with koilocytic lesions and malignant evolution, and 15 normal women without cervical lesions who served as controls. For each group, chromosome fragility was studied in peripheral blood lymphocytes. Aphidicolin (AP) was used as a clastogenic agent at a concentration of 0.12 microM. There were significant differences (p less than 0.001) between the control population and the patients affected by HPV. There were also significant differences (p less than 0.001) between the two groups infected with HPV. Our findings support the concept that chromosome fragility could serve as a cytogenetic marker to measure evolution, prognosis, and treatment of cervical lesion associated with HPV.


Chromosome losses in tumorigenic revertants of EJ/ras-expressing somatic cell hybrids. Tumorigenic transformation of SV40-immortalized human uroepithelial cells (SV-HUC) after transfection with EJ/ras was previously reported to be a rare event. To test the hypothesis that ras transformation requires loss of suppressor genes, somatic cell hybrids were generated between a rare tumorigenic transformant and an isogeneic nontumorigenic EJ/ras transfectant obtained in the same experiment. Both parental cell lines, as well as all hybrid progeny, expressed mutant p21 ras protein, but injections of three such independent hybrids into athymic nude mice at passage (P) 4 demonstrated that tumorigenicity was suppressed at 20 of 22 sites. Two tumors developed, after a relatively long 17-week latent period, as compared with a 4-week latent period for the tumorigenic parent. All three hybrids produced tumors at P8, but these showed different latent periods (3-14 weeks). Revertant hybrid tumors were high-grade carcinomas. Cell lines derived from these tumors expressed mutant p21 ras and retained at least 1 EJ/ras integration site. Karyotypic analysis of six independent hybrid tumor revertants showed that each had a unique clonal karyotype. Losses of two or more homologues of 1p, 3p, 4, 8, 10p, 11p, 13q, and 18 were identified in one or more tumorigenic revertants. Losses of all these chromosomes were previously associated with transformation of SV-HUC by EJ/ras, but were also associated with chemical transformation of SV-HUC in tumors that did not express mutant ras. Genetic losses involving most of these chromosomes have also been identified in clinical bladder cancers (i.e., 1p, 3p, 8, 11p, 13 and 18q). These data show that expression of EJ/ras does not negate or significantly alter requirements for multiple genetic losses in HUC tumorigenesis.


Characterization of a novel T-cell lymphoma cell line established from a patient with systemic lupus erythematosus-associated lymphoma. A malignant lymphoma developed in a 46-year-old male patient who had had systemic lupus erythematosus (SLE) for 18 years. The lymphoma was at disease stage IV at initial examination, and the patient died shortly thereafter. The lymphoma cells were cultured in vitro, and a continuous cell line, named SMZ-1, was established. The SMZ-1 cells, as well as the parental lymphoma cells, were of helper/inducer T-cell immunophenotype; they were positive for CD2, CD3, and CD4 antigens, and negative for CD8. Expression of CD5 and CD7 antigens was observed in a small percentage of the cells. The activation markers identified by antibodies against CD25, CD71, and HLA-DR antigens were positive. Cytogenetic analysis revealed that the SMZ-1 cells had a characteristic translocation between chromosomes 6 and 14 [t(6;14)(p21.1;q24)]. Southern blot analysis of DNA extracted from the cells demonstrated clonal rearrangement of the T cell receptor beta-chain gene. Integration of the human T-cell lymphotrophic virus type I (HTLV-I) genome was negative. The SMZ-1 cell lines should thus provide a useful model for characterization of peripheral T-cell lymphomas.


The M1 subunit of ribonucleotide reductase refines mapping of genetic rearrangements at chromosome 11p15. We report the first use of the ribonucleotide reductase M1 subunit (RRM1) locus as a marker to assist in defining genetic rearrangements at 11p15. Our sample consisted of 21 Wilms' tumors from 18 patients, and one adrenal adenoma from a patient with Beckwith-Wiedemann syndrome, preexisting chromosome 11 maps being refined by the use of the RRM1 locus in all cases. Significantly, one Wilms' tumor showed loss of heterozygosity at the RRM1 locus only, whereas the adrenal adenoma showed a maintenance of heterozygosity at the RRM1 locus, loss having been previously demonstrated at the c-Ha-ras locus. The relevance of this finding to the location of one or more disease-associated loci at 11p15 is discussed.


Development and characterization of a WEHI-3B D+ monomyelocytic leukemia cell line resistant to novobiocin and cross-resistant to other topoisomerase II-targeted drugs. A novobiocin-resistant subline of WEHI-3B D+ murine monomyelocytic leukemia cells was developed by the continuous exposure of cells to this agent in vitro. Sensitive (WEHI-3B/S) and novobiocin-resistant (WEHI-3B/NOVO) sublines were cloned in vitro. WEHI-3B/NOVO cells were stable in the absence of novobiocin for more than 3 months, and the sensitive and resistant clones displayed the same growth rate, cell cycle distribution, cell size, DNA and protein content, and cloning efficiency. Novobiocin has been shown to compete with ATP for the ATP-binding site of topoisomerase II; therefore, intracellular ATP levels can influence the cellular sensitivity to novobiocin. High-performance liquid chromatographic analysis of total cell extracts demonstrated that no difference exists between WEHI-3B/S and WEHI-3B/NOVO cells in the content of ATP. Furthermore, exposure of both cell lines to novobiocin did not affect intracellular ATP levels. In addition to an approximately 2-fold level of resistance to novobiocin, the WEHI-3B/NOVO subline was also 7- and 11-fold cross-resistant to the topoisomerase II-targeted drugs, teniposide and etoposide (VP-16), respectively. A lower level of cross-resistance, comparable to that of novobiocin, was observed in WEHI-3B/NOVO cells for the intercalating topoisomerase II-reactive drugs, doxorubicin, 4'-(9-acridinylamino)methanesulfon-m-anisidide and aclacinomycin A, while the sensitivity to the cytotoxic action of the non-topoisomerase II-acting agents, camptothecin and vincristine, was not altered. After 3-6 h of exposure to 1 microM VP-16, WEHI-3B/S cells accumulated in the S and G2 + M phases of the cell cycle. Similar changes were detected in WEHI-3B/NOVO cells only after exposure to a 10-fold higher concentration of VP-16. Exposure to 150 microM novobiocin caused an accumulation of WEHI-3B/S cells in the G0-G1 phase of the cell cycle but did not affect the cell cycle distribution of WEHI-3B/NOVO cells, while camptothecin induced the same type and extent of changes in the cell cycle distribution of both cell lines. Although the WEHI-3B/NOVO subline appeared to be less responsive to the differentiation-inducing activity of novobiocin and teniposide, the capacity of WEHI-3B/NOVO cells to respond to the differentiation-inducing agent 13-cis-retinoic acid was not significantly different from that of WEHI-3B/S cells. A slight decrease in the accumulation of VP-16 occurred in the resistant cell line, which did not appear to be of sufficient magnitude to account for the 11-fold increase in the degree of resistance to this agent.(ABSTRACT TRUNCATED AT 400 WORDS)


Antitumor quinolones with mammalian topoisomerase II mediated DNA cleavage activity. Ninety quinolones were evaluated to determine whether their ability to induce mammalian topoisomerase II mediated DNA cleavage in vitro correlated with their antitumor activity in vivo. Ten quinolones generated linear DNA at a yield of more than 10% of substrate supercoiled DNA in the mammalian topoisomerase II mediated DNA cleavage assay. All of these compounds showed a significant increase in life span (greater than 20%) in the murine leukemia P388 model. These antitumor quinolones have closely related structures: two halogens at C-6 and C-8; and cyclopropyl at N-1 of quinolone skeleton. In contrast, many analogues of the above quinolones, as well as new quinolones used clinically as an antibacterial drug, did not induce the cleavable complex in vitro or show antitumor activity in vivo. These findings indicate that quinolone derivatives can be a promising new class of antitumor agent targeting mammalian topoisomerase II.


Multiple oncogenes and tumor suppressor genes are structurally and functionally intact during hepatocarcinogenesis in hepatitis B virus transgenic mice. In the current study we sought to elucidate the molecular mechanisms which might contribute to hepatocarcinogenesis in a hepatitis B virus (HBV) envelope transgenic mouse model in which chronic hepatocellular injury and inflammation lead to regenerative hyperplasia and eventually to the development of chromosomal abnormalities and hepatocellular carcinoma (HCC), thereby reiterating many of the pathophysiological events that occur prior to the development of HCC in chronic HBV infection in humans. We have previously demonstrated that HBV envelope gene expression is decreased in regenerating hepatocytes and preneoplastic nodules early in the disease process and that expression of alpha-fetoprotein and the multidrug transporter gene mdr-III is activated in the tumors that develop in this model, but not prior to tumor development. In the current study, we examined the structure and expression of a large panel of dominant acting oncogenes and tumor suppressor genes in the liver at all stages of the disease process in order to determine the extent to which they contribute to hepatocarcinogenesis in these transgenic mice. To our surprise, no changes were observed in the structure or function of any of these genes, many of which are commonly activated in other rodent models of hepatocarcinogenesis but rarely activated in human HCC. These findings suggest that the HBV transgenic mouse model is different from most other rodent models of hepatocarcinogenesis and that it may relate more closely to the events involved in HBV-induced human hepatocarcinogenesis, where generalized chromosomal abnormalities are common, while structural and functional changes in most of the commonly studied positive-acting oncogenes examined herein are not. Since p53 and RB mutations have recently been reported to be late events in human hepatocarcinogenesis, the structural integrity of the RB locus and the absence of p53 mutations in the HBV transgenic mouse model suggest that they may represent a relatively early stage of hepatocellular tumorigenesis and that further manipulation of this model is warranted in order to more fully reproduce the molecular-genetic events that characterize HBV-induced HCC in humans.


Selective inhibition of phosphatidylinositol phospholipase C by cytotoxic ether lipid analogues. The ether lipid analogue 1-octadecyl-2-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3) has been shown to be a direct inhibitor of Swiss 3T3 fibroblast and BG1 ovarian adenocarcinoma cell cytosolic phosphoinositide selective phospholipase C (PIPLC) using [3H]-phosphatidylinositol-(4, 5)-bisphosphate ([3H]PIP2) as the substrate. The inhibition occurred when ET-18-OCH3 was incorporated into the [3H]PIP2 substrate micelles, with 50% inhibition (IC50) occurring at a ET-18-OCH3: [3H]PIP2 ratio of 0.04, or an assay concentration of 0.4 microM, and when ET-18-OCH3 was added directly to the incubation, with an IC50 of 9.6 microM. Lipid prepared from cells exposed to cytotoxic concentrations of ET-18-OCH3 for 18 h also inhibited PIPLC with an IC50 less than 1 microM. The noncytotoxic analogue 1-O-alkyl-2-hydroxy-sn-glycero-3-phosphocholine inhibited PIPLC when incorporated into the [3H]PIP2 substrate micelles, but lipid from cells grown with 5 microM 1-O-alkyl-2-hydroxy-sn-glycero-3-phosphocholine did not inhibit PIPLC. BG1 cells, which were more sensitive than Swiss 3T3 fibroblasts to growth inhibition by ET-18-OCH3, had a cytosolic PIPLC activity one-third that of Swiss 3T3 cells. NIH 3T3 cells exhibited the same sensitivity to growth inhibition by ET-18-OCH3 as Swiss 3T3 cells and had a similar level of PIPLC. v-sis NIH 3T3 cells were relatively resistant (greater than 3-fold) to growth inhibition by ET-18-OCH3 and had a cytosolic PIPLC activity more than twice that of the wild type cells. ET-18-OCH3 was a weak inhibitor, IC50 greater than 100 microM, of phospholipase D activity in NIH 3T3 cell membranes. In intact NIH 3T3 cells ET-18-OCH3 at cytotoxic concentrations did not inhibit phospholipase D or phosphatidylcholine-selective phospholipase C activity. The results show that the ether lipid analogues at cytotoxic concentrations are selective inhibitors of PIPLC and that the inhibition of PIPLC may be related to the growth inhibitory activity of the ether lipid analogues.


Expression pattern of alpha-protein kinase C in human astrocytomas indicates a role in malignant progression. Protein kinase C (PKC) is a family of isoenzymes which play an important role in regulating cell proliferation and differentiation. Constitutive activation of PKC, either by phorbol esters or overexpression of specific isoenzymes, leads to growth abnormalities in vitro and tumor promotion in vivo. Since stimulation of PKC in cultured astrocytes results in biochemical and morphological alterations associated with the transformed phenotype, we wanted to determine whether abnormal expression of specific isoenzymes of PKC was important in development of human astrocytomas in vivo. We have detected a specific pattern of alpha-PKC expression in human astrocytomas which is noteworthy because the highest transcript levels were detected in well-differentiated (Grade 1) tumors, with intermediate expression in anaplastic (Grade 2) astrocytomas and low or nondetectable levels in glioblastomas (Grade 3 astrocytomas) and normal controls. In comparison, the beta-PKC transcript was not detected in any of the tumors, while the gamma-PKC transcript was present in only one Grade 2 tumor. Immunohistochemistry, using a monoclonal antibody to alpha-PKC, revealed diffuse, positive cytoplasmic signals in most cells of the Grade 1 tumors. Grade 2 tumors exhibited heterogeneity of alpha-PKC expression, although a significant percentage of cells showed positivity. In contrast, only a small number of differentiated cells within Grade 3 tumors were positive for alpha-PKC expression, with the more malignant, dedifferentiated cells uniformly negative. Throughout all tumor grades, the staining pattern of alpha-PKC closely paralleled that of glial fibrillary acidic protein. Taken in conjunction with the established role of PKC in tumor promotion, these results suggest that the alpha-PKC isoenzyme plays a specific role in facilitating clonal expansion of transformed astrocytes in low-grade astrocytomas. Analysis of alpha-PKC may therefore serve as a direct biological marker of malignancy which may serve to enhance the current histopathological grading system.


Hepatocarcinogenicity of dehydroepiandrosterone in the rat. Dehydroepiandrosterone, a major secretory steroid hormone of the human adrenal gland, possesses mitoinhibitory and anticarcinogenic properties. It also induces peroxisome proliferation in the livers of rats and mice. Because peroxisome proliferators exhibit hepatocarcinogenic potential, it is necessary to examine the long term hepatic effects of dehydroepiandrosterone since this hormone is contemplated for use as a potential cancer chemopreventive agent in humans. Dehydroepiandrosterone was administered in the diet at a concentration of 0.45% to F-344 rats for up to 84 weeks. At the termination of the experiment, 14 of 16 rats developed hepatocellular carcinomas. Liver tumors induced by dehydroepiandrosterone lacked gamma-glutamyl transpeptidase and glutathione S-transferase (placental form); these phenotypic properties are identical to the features exhibited by liver tumors induced by other peroxisome proliferators. Dehydroepiandrosterone was also shown to markedly inhibit liver cell [3H]thymidine labeling indices, suggesting that cell proliferation is not a critical feature in liver tumor development with this agent. These results show that although dehydroepiandrosterone exerts anticarcinogenic effects in a variety of tissues, the peroxisome-proliferative property makes it a hepatocarcinogen.


Selective mutation of codons 204 and 213 of the p53 gene in rat tumors induced by alkylating N-nitroso compounds. Kidney and esophageal tumors induced by alkylating N-nitroso compounds in rats contain a high incidence (75-100%) of G----A transition mutations in the p53 gene. These are almost selectively (89%) located in the first base of codon 204 and the second base of 213, leading to amino acid substitutions Glu----Lys and Arg----Gln, respectively. In contrast to human neoplasms, a considerable fraction of rat kidney and esophageal tumors carries multiple p53 mutations. All nephroblastomas induced by transplacental exposure to N-nitrosoethylurea and 56% of esophageal tumors induced by N-nitrosomethylurea showed double mutations in codons 204 and 213 of exon 6. The selective targeting of p53 codons by alkylating nitrosamines may provide a basis for molecular epidemiological studies on this class of chemical carcinogens.


[Personal experience with treatment of recurrent herpes infections using combined nonspecific immunostimulation]. Using combined non-specific immunostimulation by means of a dialyzed leucocyte extract and bacterial vaccine, the authors treated 11 patients with relapsing herpetic infections. They achieved disappearance or marked improvement of clinical manifestations of the disease in the course of 12 months' treatment. The favourable clinical effect was associated with improvement of parameters of cellular immunity. Treatment was well tolerated by the patients. The only adverse factor was pain at the site of administration of the dialyzed leucocyte extract which persisted for 30 to 180 minutes, but due to the very favourable effect of treatment, was evaluated as insignificant by the patients.


[A preliminary epidemiological survey of acute hemorrhagic conjunctivitis in Guangzhou during July to September in 1988]. An epidemic of acute hemorrhagic conjunctivitis (AHC) occurred in Guangzhou from July to September in 1988. The main clinical symptoms were feeling of foreign, smart pain, discharging ege, facial edema and conjunctival congestion. In some cases there were headache and fever. The patients recovered rapidly. Epidemiologic feature were that the incubation period of AHC is short, rapid spread, wide prevalence and high attack rate. Approximately 490000 cases of AHC are estimated to occur in Guangzhou from July to September in 1988, with the attack rate is about 79.5%. Most patients were 20 to 59 age group (72.3%). The highest age of patient was 79 years old. The lowest age of patient was 6 months old. Both etiologic and serologic studies showed that this outbreak of AHC was caused by EV70, and then developed a mixed epidemic of Cox A24 and EV70.


[An analysis of heritability of intra-erythrocytic Na+, K+ in 139 twins]. By means of the twin model contributions of shared genes and shared environments for red blood cells Na+ and K+ were analysed by genetic and epidemiologic methods. Determination of zygosity was mainly based on examination of red blood cells system, including ABO blood group, MN blood group, P blood group, Rh blood group and secretor status of saliva, such that the probability a dizygotic pair being identical for all of these indices was less than 0.05. 139 pairs of twin in Nan Kai district Tianjin were investigated containing 74 pairs of monozygotic twins (MZ) and 65 pairs of dizygotic twins(DZ). Several methods of estimating genetic heritability (h2) were used, such as the three methods proposed by Vogel, the Holzinger's method. Levine's method based on model of variance analysis and path analysis. The degrees of compatibility of various h2 were also analysed. The contributions of shared genes, shared environments and twin's own special environments were similar for RBC Na+, h2 = 0.32, C2 = 0.35, u2 = 0.33. The contributions of shared environments were the main cause for RBC K+, C2 = 0.97.


[Study on diagnosis of hepatitis A by testing salivary IgM anti-HAV]. The use of salivary samples to diagnose acute hepatitis A was investigated. Result indicated that IgM anti-HAV can be reliably detected in saliva of 58 cases with acute hepatitis A and 9 cases with mixed infection of hepatitis A and B. It is not present in saliva of individuals without recent hepatitis A including 84 cases with acute hepatitis B, 30 cases of hepatitis non A non B and 10 healthy individuals. It was suggested that use of salivary samples to diagnose hepatitis A was specific. It's concluded that saliva is a convenient and satisfactory alternative to serum for the clinical diagnosis and epidemiological investigation of hepatitis A infection.


The bride of sevenless and sevenless interaction: internalization of a transmembrane ligand. During Drosophila retinal development, the R8 photo-receptor neuron induces a neighboring cell to assume an R7 cell fate through cell contact. This is mediated by the transmembrane protein bride of sevenless (boss) on the surface of the R8 cell, which binds the sevenless tyrosine kinase receptor (sev) on the surface of the R7 precursor cell. The boss protein, which contains a large extracellular domain, seven transmembrane segments, and a C-terminal cytoplasmic domain, has an exceptional structure for a ligand of a receptor tyrosine kinase. Using a panel of antibodies directed to various cytoplasmic and extracellular epitopes, we demonstrate that the entire boss protein from its extreme N-terminus to its extreme C-terminus is internalized by sev-expressing tissue culture cells and by the R7 precursor cell in the developing eye imaginal disc. The receptor-mediated transfer of a transmembrane ligand represents a novel mechanism for protein transfer between developing cells.


Cooperativity in transactivation between retinoic acid receptor and TFIID requires an activity analogous to E1A. In embryonal carcinoma (EC) cells retinoic acid (RA) strongly induces transcription from the RA receptor beta 2 (RAR beta 2) promoter through an RA response element (RARE) located in close proximity to the TATA box. Here we demonstrate that recombinant human TATA box-binding protein, hTFIID, and RAR functionally cooperate in transactivation of the RAR beta 2 promoter in EC cells in a strictly RA-dependent manner. We demonstrate that the core domain of hTFIID is sufficient to mediate RAR-dependent transcription and that Drosophila, but not yeast, TFIID can substitute for hTFIID. In COS cells ectopic expression of the E1A protein is a prerequisite for hTFIID and RAR to cooperate in transactivation. We propose a model for transcriptional regulation of the RAR beta 2 promoter in EC cells in which RAR, following activation by RA, functionally interacts with hTFIID via an E1A-like activity present in EC cells.


Exclusion and inclusion of alpha and beta T cell receptor alleles. Exclusion and inclusion of T cell receptor (TCR) genes were analyzed in alpha beta TCR transgenic mice. Both transgenes are expressed unusually early on the surface of CD4-8-, HSA+, IL-2R- thymocytes. These progenitor cells give rise to progeny, which at the single-cell level contains endogenous alpha but not beta TCR-RNA as well as protein, in addition to products encoded by the transgenes. Thus, the surface expression of an alpha beta TCR does not prevent further alpha TCR rearrangement in immature thymocytes that still transcribe RAG-1 and RAG-2 genes. Reduced levels of RAG-1 and RAG-2 RNA are detectable only in CD4+8+ TCR high cells, which result from positive selection in the thymus. The results suggest that a developing T cell may try different alpha beta TCRs for binding to thymic MHC ligands, and that recombination at the alpha locus ceases only after positive selection.


Wilms tumour associated with unilateral dysplasia of the iris. We report a case of Wilms tumour associated with unilateral partial aniridia. Although Wilms tumour is known to be associated with bilateral aniridia, there is only one previously reported case associated with dysplasia of the iris.


In vivo and in vitro effects of endotoxin on vascular responsiveness to norepinephrine and signal transduction in the rat. We investigated, after in vitro and in vivo exposure to gram-negative endotoxin, the altered responsiveness of rat aortic smooth muscle to catecholamines. Two hour exposure of aortic rings from normal rats to 100 ng/ml of Escherichia coli 0111:B4 endotoxin in vitro in an artificial medium supplemented with 5% fetal calf serum at 37 degrees C did not effect the basal and norepinephrine (NE)-stimulated (10 microM, 1 hr, 37 degrees C) phosphoinositide (PI) hydrolysis and isometric contractions induced by graded doses (1 nM to 10.0 microM) of NE. Increasing the incubation time with endotoxin to 18 hr did not alter the basal PI hydrolysis but significantly (P less than 0.05) decreased the NE-induced PI hydrolysis (30% inhibition) and contractile sensitivity to NE (increase of EC50 from 20.0 +/- 3.8 to 156.4 +/- 46.7 nM). Qualitatively similar results were obtained in experiments where rats were injected intravenously with buffer or an LD50 dose (10 mg/kg) of endotoxin. In these ex vivo measurements, only an 18 hr exposure to endotoxin caused significant (P less than 0.001) decreases in basal (58% inhibition) and NE-stimulated (75% inhibition) PI hydrolysis and in NE-induced isometric contractions (increase of EC50 from 11.0 +/- 3.3 to 664.1 +/- 280.0 nM). The results show that the endotoxin-induced hyporeactivity to alpha 1-adrenergic receptor stimulation 1) is markedly dependent on the length of endotoxin exposure, 2) does not require (although may be enhanced by) contact with blood cells and plasma, and 3) is paralleled by a decrease in both basal and NE-stimulated PI hydrolysis.(ABSTRACT TRUNCATED AT 250 WORDS)


Marijuana testing in urine: use of a hexadeuterated internal standard for extended linearity, and ion trap vs mass selective detector gas chromatograph/mass spectrometer systems. The use of a hexadeuterated internal standard for the assay of the marijuana carboxy metabolite in urine resulted in two significant improvements. First, the linearity of the procedure was increased considerably because of the minimal chromatographic peak overlap of the internal standard and carboxy metabolite derivatives. Second, again because of minimal peak overlap, the same extract could be analyzed with similar results by both the ion trap detector and mass selective detector gas chromatograph/mass spectrometer systems.


Monoclonal immunoradiometric assay and polyclonal radioimmunoassay compared for measuring neuron-specific enolase in patients with lung cancer. Neuron-specific enolase (NSE) is the most sensitive and specific tumor marker for small-cell lung cancer (SCLC). We evaluated a new monoclonal IRMA (Sangtec) for NSE and compared it with a polyclonal RIA (Pharmacia) in patients with SCLC or other lung cancers (NSCLC). We measured NSE concentrations in 100 healthy subjects (NI group), 100 patients with benign pulmonary diseases (BPD group), and 194 patients with advanced lung cancer (97 SCLC and 97 NSCLC). Intra- and interassay CVs were less than 7% for both assays, and dose-dilution curves paralleled their respective standard curves. Values measured by both assays were highly correlated in all groups. NSE concentrations were significantly (P less than 0.001) lower by IRMA than by RIA in NI and BPD groups. The upper 95th percentile values for NSE in the NI group were 11.7 micrograms/L in the RIA and 9.2 micrograms/L in the IRMA. In NSCLC, the values were significantly (P less than 0.05) lower by IRMA but the percentage of subjects with increased values was higher (vs the NI group, 31% for RIA and 44% for IRMA, P less than 0.005). Diagnostic sensitivity for SCLC was improved with IRMA: 83% of values with RIA and 93% with IRMA were increased above the NI group values (P less than 0.005); the corresponding values for SCLC vs BPD were 81% and 89% (P less than 0.05). NSE values measured in 39 patients with SCLC after chemotherapy were more often increased and were significantly higher with the IRMA than with the RIA (P less than 0.005).


Triphasic waves in a patient with glioblastoma multiforme. A 57-year-old woman with biopsy proven glioblastoma multiforme with greater left-sided involvement, presented with subacute right hemiparesis. Her EEG showed triphasic waves with a left-sided predominance. Possible metabolic causes of triphasic waves were excluded. The association between triphasic waves and primary brain tumor is extremely rare, and the pathogenesis unknown.


Immunodiagnostic aspects of autoantibodies against myeloperoxidase. The antigen specificity of autoantibodies causing perinuclear staining of granulocytes and monocytes (pANCA) was evaluated by analyzing 3000 sera, which were sent to us for screening of anticytoplasmic antibodies (ACPA, synonym: cANCA, anti-proteinase 3). In 620 sera, perinuclear staining was found. Antigen specificity was investigated by a myeloperoxidase ELISA and indirect immunofluorescence with Hep2 cells specific for antinuclear antibodies (ANA). Only 9.8% of the 620 sera showed reactivity with myeloperoxidase (AMPO), while 85.6% contained ANA which induced a pANCA-like staining. A further 4.6% of the 620 sera were neither ANA nor AMPO positive. Therefore, pANCA in general is only an indication that one should look for AMPO or other antilysosomal autoantibodies, when ANA have been excluded. To investigate the disease specificity of AMPO, we examined sera from patients with several well-defined autoimmune diseases. There were only very few positive results in collagen vascular diseases (3/114) (positive/total), primary systemic vasculitis (1/116) and clinically and histologically proven Wegener's granulomatosis (2/213). On the other hand, AMPO were present in patients with different forms of glomerulonephritis (45/192), especially crescentic glomerulonephritis (CGN) (34/79) without immune deposits in their biopsy specimen (3/30 showed trace deposits of IgM). There were, however, additionally 11 patients with symptoms resembling WG (who were cANCA negative, w/o characteristic WG biopsy), who had no obvious renal symptoms. These findings indicate that AMPO are primarily associated with idiopathic GN, especially CGN. Together with anti-proteinase-3 antibodies and anti-glomerular basement membrane antibodies they are an essential serologic parameter in the diagnosis of unclear systemic diseases with renal involvement.


Symptomatic accidental introduction of disinfectant electrolytic chloroxidizer solution into the peritoneal cavity of CAPD patients. Incidence and long-term effects on ultrafiltration. In recent years the Y-set with disinfectant [Buoncristiani et al. 1983] has been used in several continuous ambulatory peritoneal dialysis (CAPD) centers. This system has considerably improved the short- and long-term results of CAPD [Maiorca et al. 1983, Scalamogna et al. 1990]. Accidental introduction of disinfectant during the exchange is a risk associated with this system, but at the present time we have little data pertaining to incidence complication and its possible effects on peritoneal function. The aim of the present study was to assess the incidence of acute symptomatic accidental introduction of disinfectant in CAPD patients on the Y system and to establish whether this complication is followed by any long-term changes in peritoneal ultrafiltration.


Expression of gelatinase/type IV collagenase in tumor necrosis correlates with cell detachment and tumor invasion. We have previously observed that acellular extracts from necrotic areas (NE) of the non-metastatic murine mammary adenocarcinoma M3, enhance in vitro cell detachment and spontaneous lung metastases. In the present study, using different proteinase inhibitors along with NE, only the calcium chelator EDTA could significantly abrogate the enhanced cell detachment from M3 produced by NE. The typical cleavage products of type IV collagenase were detected inside the tumor necrotic area, mainly in association with necrobiotic cells, as evaluated by Western blot analysis and immunohistochemical assays. Zymography revealed the presence of 72- and 92-kDa gelatinase/type IV collagenase in NE. Moreover, NE increased the in vitro invasive ability of cultured M3 cells. The use of specific antibodies against both 72- and 92-kDa type IV collagenases in the invasion assay showed that only the latter was able to revert the enhanced invasiveness to the baseline. It can be concluded that tumor necrosis is an important source of gelatinase/type IV collagenase, mainly in its 92 kDa form, and plays a major role in tumor invasion.


Intracellular signal transduction in proliferation of synovial cells. We investigated some steps in the signal transduction pathway leading to the proliferation of synovial cells. 12-o-tetradecanoyl phorbol-13-acetate (TPA) which is known to stimulate phospholipid- and Ca(2+)-dependent protein kinase (C-kinase) enhanced the proliferation of synovial cells. The proliferation of synovial cells induced by interleukin-1 beta. Tumour necrosis factor alpha and granulocytes/macrophages colony stimulating factor, was inhibited by a potent C-kinase inhibitor, H7. These findings strongly suggested that the signal transduction pathway leading to proliferation of synovial cells is transmitted via C-kinase activation. Prostaglandin E2, which is known to stimulate adenylate cyclase, leading to the elevation of intracellular c-AMP level, inhibited the proliferation of synovial cells. This effect could also be mimicked by the addition of a cell permeable c-AMP analog, dibutyryl c-AMP or theophylline. Studies suggest that the feedback signal for proliferation of synovial cells was transmitted through c-AMP. We therefore conclude that signals for stimulation and inhibition of synovial cell proliferation are transmitted through different pathways.


Issues in the management of chest malignancies. Opinions can vary as to the best method of treatment for specific types of chest malignancies. If the pulmonologist is to make the primary treatment decision regarding a complicated chest malignancy, he or she must be aware of both the surgical and nonsurgical treatment options. This awareness includes knowledge of applicable surgical procedures, complications, and expected results. Chest malignancies in this category include clinically advanced lung cancer, superior sulcus tumors, chest wall invasion by lung cancer, and malignant mesothelioma. The pulmonologist should also be familiar with the indications and results of limited resection for malignancy.


Fungal scleritis after cataract surgery. Successful outcome using itraconazole. We report the development of fungal scleritis in a 53-year-old man after uncomplicated cataract surgery. Histopathology and culture identified the organism as Aspergillus flavus. Clinically, the patient worsened on treatment with oral ketoconazole and topical amphotericin B with progression of multifocal scleral nodules and necrosis. Resolution of inflammation was achieved using oral itraconazole, a new triazole antifungal agent. The patient achieved 20/15-2 visual acuity and remains free of symptoms and signs for greater than 2 years after discontinuation of all treatment.


Comparative in vitro activity of enoxacin and other fluoroquinolones against multi-resistant strains of Salmonella typhi. The in vitro activities of enoxacin, lomefloxacin, norfloxacin, ofloxacin, and pefloxacin against 274 strains of Salmonella typhi isolated from suspected typhoid fever patients (137 multi-resistant strains and 137 strains sensitive to chloramphenicol, ampicillin and/or co-trimoxazole) were determined using disk diffusion and agar dilution techniques. In vitro, enoxacin was active against all tested strains with a MIC90 and inhibition zone size against multi-resistant strains of 0.12 mg/l and 34 mm diameter, respectively. Similar results were found with the other fluoroquinolones. Enoxacin and other fluoroquinolones may be the therapy of choice in cases of typhoid fever caused by organisms resistant to the standard therapy, chloramphenicol.


Post-surgical deep vein thrombosis prevention: evaluation of the risk/benefit ratio of fractionated and unfractionated heparin. An open controlled study was carried out to assess the efficacy and tolerance of a new low molecular weight heparin for the prevention of post-surgical deep vein thrombosis and pulmonary embolism. Forty-five patients undergoing abdominal surgery mainly for neoplasm, gallstones and gastric ulcers were administered 7,500 AXaU of low molecular weight heparin subcutaneously, 2 hours before surgery and once a day for 7 days after. Heparin calcium (15,000 IU subcutaneously per day) was used as a comparison drug in 45 control subjects, matched for age, sex and type of operation. Deep vein thrombosis was identified with clinical parameters, radio-labelled fibrinogen uptake test, echo-doppler and venography; pulmonary embolism with clinical examination, chest X-rays and/or scintigraphy. No episodes of deep vein thrombosis occurred in the low molecular weight heparin-treated patients, whilst there was 1 episode, without pulmonary embolism, in the control group. The consumption of blood and haemoderivatives for transfusions was higher in the heparin calcium group. Only in this group, furthermore, did 5 patients have to suspend antithrombotic treatment due to severe haemorrhages. General tolerance of the two drugs was identical and very good.


Prevention of deep vein thrombosis in orthopaedic surgery. Comparison of two different treatment protocols with low molecular weight heparin ('Fluxum'). The efficacy of two treatment protocols with a low molecular weight heparin used for the prevention of post-operative deep vein thrombosis was compared in 40 patients undergoing hip replacement surgery. The aim was to assess whether the different timing--2 hours before (Group A, 19 patients) or 2 hours after (Group B, 21 patients) the surgical operation--of the first dose administered (15,000 aXaU) of a therapeutic cycle of 7 days could affect the results, both with respect to the preventive efficacy and to the risk of haemorrhage often connected with antithrombotic therapy. The results showed that the incidence of deep vein thrombosis was very similar and extremely low in the two groups, only 1 patient in each group having a positive diagnosis on phlebography. The effectiveness of a single daily dose of 15,000 aXaU in orthopaedic surgery was also confirmed. This dose guaranteed effective prophylaxis against the onset of deep vein thrombosis and did not appear to have any local side-effects (such as burning or pain at the site of injection) which could diminish patient compliance.


Late survival of non-small cell lung cancer patients with brain metastases. Influence of treatment. The presence of brain metastasis in lung cancer patients is a highly unfavorable event that usually allows only palliative treatment. A retrospective study was conducted to evaluate the prognostic factors in patients with non-small cell lung cancer (NSCLC) associated with brain metastases. From July 1984 through June 1990, a total of 50 patients with NSCLC associated with symptomatic brain metastasis seen at National Taiwan University Hospital were included. Patients who had incomplete cancer staging workup or loss of follow-up were excluded. Several possible prognostic variables were analyzed initially with univariate analysis and subsequently with multivariate analysis with maximal partial likelihood ratio test in the Cox model. In the univariate analysis, several factors, including number of brain metastases, treatment for brain metastasis with brain tumor resection (BTR) or whole brain radiation therapy (WBRT), and chemotherapy (C/T) after brain metastasis were found to have significant influence on the survival. However, in the multivariate analysis, patients receiving BTR, WBRT, and/or C/T lived significantly longer. The median survival of patients treated with BTR was nine months, eight months in patients with C/T, and seven months in patients with WBRT. Taken together, these patients had a median survival of seven months, which was significantly longer than patients treated with supportive care only (with a median survival of two months). Treatment of brain metastases with WBRT, BTR, C/T, or in combinations also improved the quality of life. We conclude that NSCLC patients with brain metastases should be more aggressively treated with WBRT, BTR, C/T, or in combinations than supportive care only.


Hepatoblastoma in two cousins in a family with adenomatous polyposis. Report of two cases. Two cases of hepatoblastoma in cousins in a family with familial adenomatous polyposis (FAP) are reported. Twenty-five cases of hepatoblastoma with family history of FAP have been documented in the literature, but there has never been a report of two cases of hepatoblastoma in the same polyposis family.


A comparison between the nutritive value of short-cutting cycle, high temperature-dried alfalfa and timothy hay for horses. The objective was to evaluate the nutritive value of short-cutting cycle, high temperature-dried (SCCHTD) alfalfa compared to timothy hay. This was achieved by carrying out 4 x 4 Latin Square digestibility trial using 4 Thoroughbred (one three-quarter Thoroughbred) horses (mean liveweight, 531 kg). The four dietary treatments were 0AA (timothy hay only), 33AA (0.33 alfalfa: 0.67 timothy hay), 67AA (0.67 alfalfa: 0.33 timothy hay) and 100AA (alfalfa only). Digestibility data were obtained by using acid-insoluble ash to estimate apparent digestibility coefficients of nutrients. Rate of passage of the feedstuff was determined using chromium-mordanted hay. Plasma triglyceride and cholesterol concentrations were estimated. The digestibilities of organic matter (0.63), energy (0.57) and crude protein (0.74) of the alfalfa were significantly (P less than 0.001) higher than those for the hay (0.45, 0.43 and 0.36 respectively). The fibre components of alfalfa and hay were digested to the same extent but the ether extract of alfalfa was less well digested. Alfalfa saponins had no consistently significant effects on plasma cholesterol and triglyceride values but may contribute to the negative digestibility of alfalfa ether extract. We conclude that SCCHTD alfalfa is of much higher nutritive value than timothy hay when fed to Thoroughbred horses.


Effects of induced alkalosis on performance in thoroughbreds during a 1,600-m race. There is considerable debate regarding the ergogenic effects of sodium bicarbonate (NaHCO3) on racing performance in horses. Anecdotal evidence suggests that NaHCO3 improves performance by increasing the buffering capacity of the blood and delaying the onset of hydrogen ion-induced fatigue. In a cross-over study, 16 Thoroughbred racehorses were given an aqueous solution of NaHCO3 (0.4 g/kg in 1 litre H2O) or a control treatment (1 litre H2O) before a 1600-m race. Treatments were administered 3 h before the race, which was the time to peak buffering capacity (2.5-3.0 h) determined in a separate study. Before the race, there was a significant increase in venous HCO3- and pH in the NaHCO3-treated horses. After the race, there was a significant increase in venous blood pH and lactate in the NaHCO3-treated horses. Collectively, the data suggest an improved buffering capacity of the blood after NaHCO3 treatment. However, there was no change in race times or venous partial pressure of carbon dioxide. Therefore, the administration of NaHCO3 provided no ergogenic benefit to horses competing in a 1,600-m race.


Mutation of a cysteine in the first transmembrane segment of Na,K-ATPase alpha subunit confers ouabain resistance. The cardiac glycoside ouabain inhibits Na,K-ATPase by binding to the alpha subunit. In a highly ouabain resistant clone from the MDCK cell line, we have found two alleles of the alpha subunit in which the cysteine, present in the wild-type first transmembrane segment, is replaced by a tyrosine (Y) or a phenylalanine (F). We have studied the kinetics of ouabain inhibition by measuring the current generated by the Na,K-pump in Xenopus oocytes injected with wild-type and mutated alpha 1 and wild-type beta 1 subunit cRNAs. When these mutations, alpha 1C113Y and alpha 1C113F [according to the published sequence [Verrey et al. (1989) Am. J. Physiol., 256, F1034] were introduced in the alpha 1 subunit of the Na,K-ATPase from Xenopus laevis, the inhibition constant (Ki) of ouabain increased greater than 1000-fold compared with wild-type. A more conservative mutation, serine alpha 1C113S did not change the Ki. We observed that the decreased affinity for ouabain was mainly due to a faster dissociation, but probably also to a slower association. Thus we propose that an amino acid residue of the first transmembrane segment located deep in the plasma membrane participates in the structure and the function of the ouabain binding site.


The effect of tyrosine-specific protein phosphorylation on the assembly of adherens-type junctions. Adherens-type junctions (AJs) are major subcellular targets for tyrosine specific protein phosphorylation [Volberg et al. (1991) Cell Regul., 2, 105-120]. Here we report on the apparent effect of such phosphorylation events on the assembly and integrity of AJs. We show that incubation of MDCK cells with potent inhibitors of tyrosine-specific phosphatases (PTP), namely H2O2 and vanadate, leads to a dramatic increase in AJ-associated phosphotyrosine which was apparent already within 2-5 min of treatment and progressed upon further incubation. Examination of H2O2 vanadate treated cells at later time points indicated that intercellular AJs rapidly deteriorated, concomitantly with a marked increase in the number and size of vinculin and actin containing focal contacts. In parallel, major changes were observed in cell structure and topology, as revealed by electron microscopy. These were manifested by rapid rounding-up of the cells followed by reorganization of the cell monolayer. Other intercellular junctions, including desmosomes and tight junctions, visualized by staining with desmoplakin and ZO-I antibodies, were not significantly affected. To verify that modulation of AJs was indeed related to tyrosine phosphorylation, we have carried out reciprocal experiments in which Rovs Sarcoma virus (RSV) transformed chick lens cells, expressing high levels of pp60src kinase, were treated with inhibitors of tyrosine kinases, (tyrphostins). We show that following such treatment, intercellular AJs which were deteriorated in the transformed cells, were reformed. Based on these observations, we propose that specific tyrosine phosphorylation of AJ components is involved in the downregulation of these cellular contacts.


Cyclin A- and cyclin B-dependent protein kinases are regulated by different mechanisms in Xenopus egg extracts. Cyclins are proteins which are synthesized and degraded in a cell cycle-dependent fashion and form integral regulatory subunits of protein kinase complexes involved in the regulation of the cell cycle. The best known catalytic subunit of a cyclin-dependent protein kinase complex is p34cdc2. In the cell, cyclins A and B are synthesized at different stages of the cell cycle and induce protein kinase activation with different kinetics. The kinetics of activation can be reproduced and studied in extracts of Xenopus eggs to which bacterially produced cyclins are added. In this paper we report that in egg extracts, both cyclin A and cyclin B associate with and activate the same catalytic subunit, p34cdc2. In addition, cyclin A binds a less abundant p33 protein kinase related to p34cdc2, the product of the cdk2/Eg1 gene. When complexed to cyclin B, p34cdc2 is subject to transient inhibition by tyrosine phosphorylation, producing a lag between the addition of cyclin and kinase activation. In contrast, p34cdc2 is only weakly tyrosine phosphorylated when bound to cyclin A and activates rapidly. This finding shows that a given kinase catalytic subunit can be regulated in a different manner depending on the nature of the regulatory subunit to which it binds. Tyrosine phosphorylation of p34cdc2 when complexed to cyclin B provides an inhibitory check on the activation of the M phase inducing protein kinase, allowing the coupling of processes such as DNA replication to the onset of metaphase. Our results suggest that, at least in the early Xenopus embryo, cyclin A-dependent protein kinases may not be subject to this checkpoint and are regulated primarily at the level of cyclin translation.


Relocation and distinct subcellular localization of p34cdc2-cyclin B complex at meiosis reinitiation in starfish oocytes. M phase promoting factor (MPF) is a major element controlling entry into the M phase of the eukaryotic cell cycle. MPF is composed of two subunits, p34cdc2 and cyclin B. Using indirect immunofluorescence staining with specific antibody against starfish cyclin B, we monitored the dynamics of the subcellular distribution of MPF during meiosis reinitiation in starfish oocytes. We found that all of the cyclin B is already associated with p34cdc2 in immature oocytes arrested at the G2/M border and that this inactive complex is present exclusively in the cytoplasm. After its activation, part of the p34cdc2-cyclin B complex moves into the germinal vesicle before nuclear envelope breakdown, independently of either microtubules or actin filaments. Thereafter, some part of the complex accumulates in the nucleolus and condensed chromosomes. Another portion of the complex accumulates on meiotic asters and spindles, while the rest is still present throughout the cytoplasm. As these patterns of localization are detected in the detergent-extracted oocytes, we propose at least four distinct subcellular states of the p34cdc2-cyclin B complex: freely soluble, microtubule-associated, detergent-resistant cytoskeleton-associated and chromosome-associated. Thus, in addition to the intramolecular modification of p34cdc2-cyclin B complex, its intracellular relocation plays a key role in promoting the M phase.


The Cln3-Cdc28 kinase complex of S. cerevisiae is regulated by proteolysis and phosphorylation. In Saccharomyces cerevisiae, several of the proteins involved in the Start decision have been identified; these include the Cdc28 protein kinase and three cyclin-like proteins, Cln1, Cln2 and Cln3. We find that Cln3 is a very unstable, low abundance protein. In contrast, the truncated Cln3-1 protein is stable, suggesting that the PEST-rich C-terminal third of Cln3 is necessary for rapid turnover. Cln3 associates with Cdc28 to form an active kinase complex that phosphorylates Cln3 itself and a co-precipitated substrate of 45 kDa. The cdc34-2 allele, which encodes a defective ubiquitin conjugating enzyme, dramatically increases the kinase activity associated with Cln3, but does not affect the half-life of Cln3. The Cln--Cdc28 complex is inactivated by treatment with non-specific phosphatases; prolonged incubation with ATP restores kinase activity to the dephosphorylated kinase complex. It is thus possible that phosphate residues essential for Cln-Cdc28 kinase activity are added autocatalytically. The multiple post-translational controls on Cln3 activity may help Cln3 tether division to growth.


Casein kinase II phosphorylates the eukaryote-specific C-terminal domain of topoisomerase II in vivo. The decatenation activity of DNA topoisomerase II is essential for viability as eukaryotic cells traverse mitosis. Phosphorylation has been shown to stimulate topoisomerase II activity in vitro. Here we show that topoisomerase II is a phosphoprotein in yeast and that the level of incorporated phosphate is significantly higher at mitosis than in G1. Comparison of tryptic phosphopeptide maps reveals that the major phosphorylation sites in vivo are targets for casein kinase II. Incorporation of phosphate into topoisomerase II is nearly undetectable at the non-permissive temperature in a conditional casein kinase II mutant. The sites modified by casein kinase II are located in the extreme C-terminal domain of topoisomerase II. This domain is absent in prokaryotic and highly divergent among eukaryotic type II topoisomerases, and may serve to regulate functions of topoisomerase II that are unique to eukaryotic cells.


A confined variable region confers ligand specificity on fibroblast growth factor receptors: implications for the origin of the immunoglobulin fold. Binding of cellular growth factors to their receptors constitutes a highly specific interaction and the basis for cell and tissue-type specific growth and differentiation. A unique feature of fibroblast growth factor (FGF) receptors is the multitude of structural variants and an unprecedented degree of cross-reactivity between receptors and their various ligands. To examine receptor-ligand specificity within these families of growth factors and receptors, we used genetic engineering to substitute discrete regions between Bek/FGFR2 and the closely related keratinocyte growth factor receptor (KGFR). We demonstrate that a confined, 50 amino acid, variable region within the third immunoglobulin-like domain of Bek and KGFR exclusively determines their ligand binding specificities. Replacing the variable region of Bek/FGFR2 with the corresponding sequence of KGFR resulted in a chimeric receptor which bound KGF and had lost the capacity to bind basic FGF. We present evidence that the two variable sequences are encoded by two distinct exons that map close together in the mouse genome and follow a constant exon, suggesting that the two receptors were derived from a common gene by mutually exclusive alternative mRNA splicing. These results identify the C-terminal half of the third immunoglobulin-like domain of FGF receptors as a major determinant for ligand binding and present a novel genetic mechanism for altering receptor-ligand specificity and generating receptor diversity.


The mouse mammary tumour virus long terminal repeat encodes a type II transmembrane glycoprotein. Superantigens are products of bacterial or viral origin which stimulate large numbers of T cells as a consequence of the interaction of particular V beta chains of the T cell receptor with class II major histocompatibility complex (MHC) molecules and superantigen on the stimulating cell. The Minor lymphocyte stimulatory (Mls) antigens, originally discovered as strong lymphocyte stimulatory determinants in vitro and subsequently shown to delete T cells expressing specific V beta chains during development, have recently been shown to be genetically linked to endogenous mouse mammary tumour viruses (MTVs). This stimulation is effectuated by an unidentified product encoded by an open reading frame (orf) present in the 3' long terminal repeat (LTR) of MTVs. Using in vitro translation in the presence of rough microsomal vesicles, we show that (i) the orf of MTV encodes a type II transmembrane glycoprotein (N-terminus intracellular, C-terminus extracytoplasmic), and (ii) a cotranslationally secreted orf protein is not produced. We have also isolated and sequenced several endogenous MTV orfs (MTV-1, MTV-6 and MTV-13) which are involved in the deletion of V beta-bearing T cells; each of these sequences are nearly identical to each other. These observations, together with sequence comparisons of several orf genes, lead to a model of action of viral superantigens.


Existence in Escherichia coli of a mechanism that generates 'staggered' head-to-head dimers of plasmid DNA; possible involvement of the Tn3 transposase. The structures of two plasmids, one found in one of the transformants with putative products of an in vitro DNA rearrangement reaction containing the Tn3 transposase, and another found as a spontaneous tnpA negative derivative of an overproducer of the Tn3 transposase, have been elucidated. They both had shown non-conventional results in restriction enzyme analysis. For the determination of the structures, we used restriction enzyme analysis, denaturation and renaturation experiments, and DNA nucleotide sequencing. The structures turned out to be 'staggered' head-to-head dimers of the original monomer plasmids, containing gigantic inverted repeats separated by two identical spacers which are also in inverted orientations themselves. Two alternative models for the mode of origin of such a structure, a bimolecular model and a unimolecular one, are discussed. The circumstances in which these two plasmids occurred suggest possible involvement of the Tn3 transposase in their generation.


Increase of glomerular filtration rate and renal plasma flow by insulin-like growth factor-I during euglycaemic clamping in anaesthetized rats. Renal function was studied in anaesthetized rats receiving i.v. infusions of recombinant human insulin-like growth factor-I (IGF-I) under euglycaemic clamp conditions. IGF-I increased glomerular filtration rate up to 35% and renal plasma flow up to 100%, this increase being dose dependent with half-maximal stimulation at serum IGF-I concentrations of about 24 pmol ml-1. Renal vascular resistance was reduced up to 50%, filtration fraction decreased up to 30% and urine flow increased up to three fold while arterial blood pressure was unchanged. Renal haemodynamics were affected at serum IGF-I concentrations that did not stimulate total body glucose disposal during euglycaemic clamping. IGF-I seemed to regulate renal function through IGF-I receptors apparently independent of acute changes of glucose metabolism.


Enhancement of regeneration by Org 2766 after nerve crush depends on the type of neural injury. The neurotrophic effects of the adrenocorticotropin (ACTH)-(4-9) analog Org 2766 (Met(O2)-Glu-His-Phe-D-Lys-Phe) were studied in rats recovering from a sciatic nerve crush. Org 2766 (10 micrograms/rat s.c., every 48 h) increased the number of myelinated axons reinnervating a previously denervated sciatic nerve by 32% (P less than 0.01), as assessed 13 days after crush lesioning, and facilitated recovery of sensorimotor functioning by 14% (P = 0.05), as measured by foot withdrawal after stimulation of the footsole with hot air. However, these facilitating effects were only seen if the nerve was lesioned using forceps with grooved jaws and not if forceps were used with cross-hatched jaws. Endoneural tubes and Schwann cells of the sciatic nerve appeared to be better preserved after crushing with grooved rather than cross-hatched jaws. Our data indicate that the regeneration-enhancing effects of Org 2766 are dependent on the type of injury applied to the endoneurium and endoneural tubes of the sciatic nerve and suggest that endoneural tissue may mediate the neurotrophic properties of Org 2766.


The tail-flick inhibition induced by beta-endorphin administered intrathecally is mediated by activation of kappa- and mu-opioid receptors in the mouse. The inhibition of the tail-flick response induced by beta-endorphin given i.c.v. has been demonstrated to be mediated by the stimulation of epsilon- but not mu-, delta- or kappa-opioid receptors. beta-Endorphin given i.t. also inhibited the tail-flick response. The present studies were designed to determine what types of opioid receptors in the spinal cord were involved in i.t. beta-endorphin-induced tail-flick inhibition. Blockade of kappa-opioid receptors by coadministration of nor-binaltorphimine or Win 44,441-3 with beta-endorphin given i.t. dose dependently inhibited i.t. beta-endorphin-induced inhibition of the tail-flick response. Blockade of mu-opioid receptors by i.t. coadministration of D-Phe-Cys-Tyr-D-Try-Orn-Thr-Pen-Thr-NH2 with beta-endorphin blocked i.t. beta-endorphin-induced inhibition of the tail-flick response. I.t. injection of delta-opioid receptors antagonists, ICI 174,864 and naltrindole, or epsilon-opioid receptor antagonist, beta-endorphin-(1-27), did not affect inhibition of the tail-flick response induced by beta-endorphin given i.t. Blockade of alpha 2-adrenoceptors and 5-HT receptors by i.t. injection of yohimbine and methysergide, respectively, also did not affect inhibition of the tail-flick response induced by beta-endorphin given i.t. The results indicate that the inhibition of the tail-flick response induced by beta-endorphin given i.t. is mediated by the stimulation of kappa- and mu-opioid receptors but not delta- and epsilon-opioid receptors, alpha 2-adrenoceptors or 5-HT receptors.


Influence of benzodiazepines on the response of the guinea-pig isolated trachea to the contractile action of adenosine. The effects of diazepam and other agonists of central or peripheral benzodiazepine receptors were studied on the contractile action of adenosine, 2-chloroadenosine and R-PIA (N6-(L-2-phenylisopropyl)-adenosine) on the guinea-pig isolated trachea. These effects were compared to those of dipyridamole. Diazepam 10(-7) to 10(-5) M potentiated the efficacy of adenosine; the maximal contractile effect of adenosine (% vs. acetylcholine 10(-3) M) was 20.4 +/- 4.2 (n = 21) in control conditions and 45.5 +/- 3.7 (n = 6; P less than 0.001) in the presence of diazepam 10(-5) M. Ro5-4864 (10(-7) to 10(-5) M) or alpidem (10(-7) to 10(-5) M), both agonists of peripheral benzodiazepine receptors, potentiated the contractile effects of adenosine to the same extent as diazepam. Clonazepam and zopiclone, both agonists of central benzodiazepine receptors, did not modify these effects. Antagonists of central (flumazenil) or peripheral (RP 52028) benzodiazepine receptors had no influence on the interaction between diazepam or Ro5-4864 and adenosine. Conversely, dipyridamole significantly reduced (10(-7) M) or suppressed (10(-6) M) the contractile effects of adenosine. The contractile effects of 2-chloroadenosine and R-PIA were weakly affected in presence of high concentrations of diazepam and dipyridamole. Epithelium removal potentiated the contractile effect of adenosine on the guinea-pig isolated trachea and increased the potentiating effect of diazepam. It is concluded that benzodiazepines and related compounds can potentiate the contractile effect of adenosine on the guinea-pig isolated trachea through the activation of a peripheral receptor for the benzodiazepines and the resulting inhibition of adenosine uptake.


The density of GABAB binding sites in the substantia nigra is greater in rat pups than in adults. Baclofen-displaced [3H]GABA (gamma-aminobutyric acid) binding to GABAB sites was assayed in brain regions obtained from adult and 14- to 17-day-old rats. The substantia nigra of rat pups exhibited a 3- to 5-fold higher density (Bmax) of GABAB binding sites than the nigra of adult rats. In contrast, the density of GABAB sites was lower in hippocampal and cerebellar tissue from pups, both approximately two-thirds the density in corresponding adult structures. This anatomically selective overproduction of GABAB receptors in immature substantia nigra is consistent with the increased anticonvulsant efficacy of the GABAB agonist baclofen observed in rat pups.


Effects of 5-HT1A receptor agonists on the circadian rhythm of wheel-running activity in hamsters. The effects of 5-HT1A receptor agonists 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), buspirone and ipsapirone on wheel-running activity in hamsters were investigated in comparison with those of GABAA receptor agonist muscimol and benzodiazepine triazolam. Intraperitoneal administration of 8-OH-DPAT, buspirone, ipsapirone, muscimol and triazolam at circadian time (CT) 8 (CT 12; onset of activity) induced a significant phase advance of wheel-running activity under constant light conditions. However, administration of these drugs at other CT points did not induce phase changes. The administration of trifluoromethylphenylpiperazine (TFMPP), a 5-HT1B receptor agonist, at CT8 produced a small phase advance. The phase advance induced by 8-OH-DPAT was blocked by pretreatment with (-)-pindolol, a 5-HT1A receptor antagonist. In addition, 8-OH-DPAT, buspirone and SM3997 accelerated the rate of re-entrainment to an 8-h phase advance in the light-dark cycle. These observations suggest that 5-HT1A receptors in the brain participate in the regulation of the circadian rhythm of wheel-running activity in hamsters.


Evidence for more than two central benzodiazepine receptors in rat spinal cord. Competitive binding assays were performed in rat spinal cord membranes at 0 degrees C against [3H]Ro15-1788. The displacement curves of Ro15-1788 and alpidem produced pseudo-Hill slopes of 1.0 and 0.5, respectively. In addition, 5-10% of the specifically bound [3H]Ro15-1788 could not be displaced by greater than 100 microM of alpidem. These two pieces of evidence strongly suggest the presence of at least three central benzodiazepine binding sites in the spinal cord.


Genetic relationships among strains of Salmonella enteritidis in a national epidemic in Switzerland. A collection of Salmonella enteritidis strains isolated in Switzerland (1965-90) was characterized. The phage type and plasmid profile of isolates were compared with the copy number and insertion loci of the DNA insertion element IS200. Three clonal lines of S. enteritidis were identified by IS200 profile; the various phage types were subtypes reproducibly associated with one of these lines. All human and poultry isolates contained a 38 Mda plasmid which hybridized with a mouse virulence-associated gene probe. In S. enteritidis, the IS200 profile is a race-specific molecular marker of the chromosome, and may be particularly applicable for studying the epidemiology of less common serovars.


Methods for estimating the incidence of primary infection in pregnancy: a reappraisal of toxoplasmosis and cytomegalovirus data. Accurate incidence information is required to plan and evaluate screening programmes which have been proposed for the detection of primary toxoplasmosis and cytomegalovirus infection in pregnancy. Appropriate statistical methods are described for deriving incidence rates and their confidence intervals from three types of data: change in age-specific seroprevalence, seroconversion, and IgM studies. These methods are applied to seven published studies on toxoplasmosis and cytomegalovirus carried out in the UK. In these publications only one estimate of the infection rate per pregnancy was correctly derived, and none were accompanied by confidence intervals. Using the proposed methods, most estimates of the primary toxoplasmosis rate in these studies were between 2.5 and 5.5 per 1000 pregnancies, compared to the 2 per 1000 usually cited. Most cytomegalovirus incidence estimates were between 4 and 10 per 1000 pregnancies.


Purification of platelet-derived endothelial cell growth inhibitor and its characterization as transforming growth factor-beta type 1. In 1986, Brown and Clemmons (Proc. natl Acad. Sci. USA 83 (1986) 3321) showed that platelets contain a substance, platelet-derived growth inhibitor (PDGI), that inhibits in vitro endothelial cell replication. Although platelets are rich in transforming growth factor beta (TGF-beta), PDGI was considered not to be related to TGF-beta, on the basis of its reported properties (extraction from platelets at neutral pH, binding to heparin-Sepharose). However, we purified PDGI to near homogeneity and showed that on the basis of HPLC retention behavior, in vitro growth inhibitory activities with several cell types, receptor binding, and immunoneutralization of growth inhibitory activity with specific anti-TGF-beta type 1 antibodies, PDGI is most probably identical with TGF-beta type 1.


Deficits in social behavior in autism and their modification by a synthetic adrenocorticotrophic hormone (4-9) analog. When charting the structure of the social behavior of autistic children by means of an ethologically analyzed playroom session, deficits appeared in the reciprocity of eye-contact and in the location of verbal initiatives. These deficits in social behavior were beneficially influenced by treatment with the adrenocorticotrophic hormone (4-9) analog ORG 2766.


Hepatic chemoinfusion of 5-FU in metastasis of gastrointestinal cancer and advanced primary hepatocellular carcinoma. From 1 January 1983 to 1 January 1988, 38 patients were treated for hepatic cancer in the HEINZ-KALK-Hospital. Thirty-one of these had liver metastases due to gastrointestinal cancer and seven had advanced primary hepatocellular cancer. In all patients more than 50% of the liver volume was involved with the tumour or the metastases. Eleven patients with liver metastases of gastrointestinal cancer (excepting colorectal cancer) were treated by intra-arterial hepatic bolus infusion of 750-1000 mg 5-fluorouracil (5-FU) by selective catheterisation of the hepatic or superior mesenteric artery after puncture of the right or left femoral artery. The median survival was 13.4 months. In seven patients with advanced primary hepatocellular carcinoma the same therapeutic regime was used. The median survival was 10 months. In the 21 patients with disseminated metastases of previously resected colorectal cancer a catheter was inserted into the gastro-duodenal artery and connected to a subcutaneously placed port. Brief infusions of 750-1000 mg 5-FU were administered for 14 days with a day interruption and thereafter 2 month interruption. There were few side effects and 80% of the patients continued to work or carry on a normal life. The median survival was 14.4 months. Based on this experience we consider hepatic chemoinfusion with 5-FU in gastrointestinal cancer and advanced primary hepatocellular carcinoma is capable of improving quality of life and possibly expectancy.


Biodistribution of lipiodol following hepatic arterial injection. Lipiodol, a derivative of poppy seed oil, has been used angiographically to improve visualisation of small liver tumours. We have utilised this finding to determine whether intrahepatic arterial injection of lipiodol can be used as a vehicle to deliver selectively 131I into liver tumours. Two groups of rats were studied. Group 1 (control, no liver tumour) received 0.1 ml 131I-lipiodol (1 microCi) into the hepatic artery. Animals were killed at regular time intervals over 30 days and organs were submitted to well-counting. Over 90% of activity remained in the liver at 6 h. Eighty per cent activity was lost from the normal liver, to be excreted in the urine over 30 days. Group 2 animals received intraportal injections of 7.5 x 10(5) MC28 sarcoma cells. Multiple liver metastases were present after 14 days. Animals were similarly studied at each time interval and samples from tumour and normal liver were submitted to well-counting. Lipiodol was selectively retained within tumour and cleared from normal liver. 131I-lipiodol may prove valuable as a delivery agent for radio/chemotherapy to liver metastases.


[The calcium messenger system].

[The calcium messenger system]. Calcium ion acts as an intracellular messenger of various types of extracellular signals including hormones, neurotransmitters, cytokines and growth factors. Recent advances in the research of the calcium messenger system have provided informations as to the structure and function of many proteins involved in the messenger system. Also, technical advances in the measurement of the dynamics of calcium ion in a small cell have made it possible to better understand the role of calcium in the activation of the cell. In the present review, a picture of the calcium messenger system is presented.


Possible involvement of myeloperoxidase in lipid peroxidation. 1. Exposure of liposomes to the MPO-H2O2-Cl- system results in oxidation of lipids. Malondialdehyde and 4-hydroxynonenal are formed. 2. Oxidation of liposomes by stimulated rat neutrophils, assessed by malondialdehyde formation, is inhibited by KCN. This indicates involvement of MPO in the process. 3. The MPO-H2O2 system oxidizes mildly LDL but in the presence of chloride a propagation phase, with a rapid increase of conjugated diene formation, was observed.


Ontogenetic changes in porcine adrenocortical adrenocorticotropic hormone receptors. 1. Binding of [125I]ACTH(1-38) analog to adrenal receptors was measured in fetal pigs (Sus domesticus) at 15-day intervals from midpregnancy (60 days) to near term (105 days; pregnancy length 114 days). 2. Binding was greatest at day 60 (0.42 +/- 0.03 fmol/200 micrograms protein or 0.50 +/- 0.08 fmol/50 micrograms DNA), and least at day 105 (0.13 +/- 0.03 fmol/200 micrograms protein or 0.16 +/- 0.04 fmol/50 micrograms DNA). Total adrenal binding was constant (0.61 +/- 0.02 fmol/paired adrenals). 3. Scatchard analyses at day 60 and day 105 showed comparable apparent affinities of ACTH receptors (Ka day 60 = 1.51 +/- 0.72 x 10(9) M-1 vs Ka day 105 = 1.94 +/- 0.78 x 10(9) M-1). 4. DNA per paired adrenals and membrane-associated protein increased 1.6-fold, providing a constant protein: DNA ratio. Concentrations of adrenal cortisol were constant from 60 to 90 days of gestation age but increased dramatically by day 105. 5. These data suggest that during 60-105 days of gestation age the number of ACTH receptors per cell is reduced.


Studies on carbohydrate-binding proteins using liposome-based systems--I. Preparation of neoglycoprotein-conjugated liposomes and the feasibility of their use as drug-targeting devices. 1. Five types of neoglycoprotein-coupled liposomes were prepared in order to investigate their potential utility as new types of drug-targeting devices which exploit cellular functions of carbohydrate-binding proteins. 2. These preparations were shown to be stable at 37 degrees C for 24 hr and at 7 degrees C over 4 months. 3. An inhibition assay in an in vitro system using human adenocarcinoma cells indicated the high affinity binding of neoglycoprotein-conjugated liposomes. The inhibitory potency correlated with both the type and the amount of immobilized neoglycoproteins on liposomes. 4. A tissue distribution assay in an in vivo system using Ehrlich solid tumor-bearing mice showed the feasibility of the application of [125I]neoglycoprotein-conjugated liposomes as drug-targeting devices, based on carbohydrate-protein interactions.


[Familial adenomatous polyposis: early diagnosis by genetic mapping]. The use of probes detecting polymorphic loci within the human population has enabled accurate localization of the genetic defect responsible for familial adenomatous polyposis on chromosome 5. This was used to screen two families for the presymptomatic diagnosis in children of an affected parent. In both cases, the use of 8 polymorphic probes located on either side of the gene provided information which could be used in the management of children born from the patients at risk. The set of probes used in this work should be informative in most of the affected adenomatous polyposis families.


[Value of screening of familial adenomatous polyposis for the prevention of colorectal cancer]. Familial adenomatous polyposis coli is a hereditary autosomal dominant disease which spontaneously and inevitably leads to degeneration of colorectal adenomas and requires preventive surgical treatment. The aim of this study was to evaluate the age of colorectal degeneration and the need for a screening technique in family members. Between 1983 and 1989, 141 patients were treated for familial adenomatous polyposis in our surgical center. Mean age at surgery was 32 years and 64 patients (45.4 percent) had a colorectal carcinoma. Thirty had an in situ tumor (mean age: 30 years) and 34 had an invasive adenocarcinoma (mean age: 45 years), 7 of whom died of their cancer. No colonic cancer was found in patients younger than 20. Thirty-eight percent of the patients under 40 years of age, 73 percent of the patients older than 40 years and 81 percent of those older than 50 had an adenocarcinoma. Fifty percent of the patients with carcinoma were younger than 40 years and 7 percent were less than 25 years old. Seventy-one patients were symptomatic at the time of operation (mean age: 40 years), 32 (45 percent) had a colonic cancer. In 70 patients, familial adenomatous polyposis was detected by screening (mean age: 24) and 2.8 percent had a colonic carcinoma. We conclude that the age-related risk of developing colonic carcinoma requires prophylactic surgery in asymptomatic patients before 20 years of age, and that routine familial screening would be of some benefit.


[Cytomegalovirus infections in patients after liver transplantation: determination of risk groups]. The goal of this study was to identify high-risk groups for cytomegalovirus infection after liver transplantation. Sixty-one patients were evaluated. Twenty-five patients (41 percent) had infection. Among the 16 patients who were seronegative for the virus before transplantation, 11 received a liver graft and blood products from seronegative donors and none of them developed infection. All seronegative recipients of a liver from seropositive donors (5/5) developed primary infection. Among the 45 patients seropositive before transplantation, 20 developed a cytomegalovirus infection, whatever the donor serologic status. The incidence of symptomatic reactivation or reinfection was high (14/20), and, for 12/14 of them, associated with early acute rejection. Two high-risk groups of patients, eligible for cytomegalovirus prophylaxis, were identified: seronegative recipients of seropositive donors and seropositive recipients with early acute rejection.