Fecal bile acid metabolic pattern after administration of different types of bread. Increased concentrations of fecal bile acids have been suggested to be associated with increased risk of colorectal cancer. Fecal bile acid profiles were determined in 12 healthy Finnish women who included in their normal diets for 2-week periods in turn three different types of bread, 200-300 g/day. The breads contained either low-fiber wheat, whole-meal wheat, or whole-grain rye. During consumption of rye bread, the total mean concentration of fecal free bile acids was 4.77 +/- 0.90 mumol/g of dry feces (mean +/- SEM), which was much lower than with the normal omnivorous diet (8.05 +/- 1.56 mumol/g) or during administration of the low-fiber wheat bread (8.83 +/- 1.56 mumol/g) or the whole-meal wheat bread (7.88 +/- 1.34 mumol/g) (P less than 0.05). This decrease was mainly caused by increased proportions of saponifiable bile acids (P less than 0.01). During intake of the whole-grain rye bread, 46% +/- 3% of the fecal bile acids were in their saponifiable forms; this percentage was 30% +/- 3% during the control period, 30% +/- 4% during the low-fiber wheat bread period, and 27% +/- 4% during the whole-meal wheat bread period. It is concluded that the type of bread significantly effects concentrations of cocarcinogenic and comutagenic free lithocholic and deoxycholic acids by changing modes of conjugation in the gut.


Hepatitis C and B viruses in hepatitis B surface antigen-negative hepatocellular carcinoma. The relative role of hepatitis C virus and hepatitis B virus in hepatitis B surface antigen-negative hepatocellular carcinoma was evaluated by polymerase chain reaction in 31 patients from Taiwan. Twenty-one were positive for antibody to hepatitis C virus (group 1) and 10 were negative (group 2). Of the group 1 patients, hepatitis C viral RNA was detected in the serum by polymerase chain reaction in 16 and in the liver tissue in 17, whereas hepatitis B viral DNA was found in the liver tissue in only 4, and none were found in the serum. In group 2 patients, hepatitis C viral RNA was detected in the serum of 1 and in the liver tissue of another. In contrast, hepatitis B viral DNA was found in the serum of 4 patients and in the liver tissues of 5. It was concluded that hepatitis C virus plays an important role in hepatocarcinogenesis in hepatitis B surface antigen-negative patients in Taiwan, especially in those who had antibody to hepatitis C virus; in those without antibody to hepatitis C virus, hepatitis B virus might still be associated with the development of hepatocellular carcinoma in a significant proportion of such patients.


Infectious agents in the etiology of esophageal cancer. Extensive epidemiological and experimental studies have suggested that some chemical agents, nutritional deficiencies, and physical factors are associated with the development of esophageal cancer (EC). Recent evidence also suggests an etiologic role of certain microorganisms in esophageal carcinogenesis either by producing carcinogens or promotors or by acting directly on the host cells. The mutagenic and carcinogenic effects of several fungi and bacteria isolated from the grains and foodstuffs in high-risk areas have been shown by in vitro and in vivo studies. Certain viruses, e.g., human papillomavirus, herpes simplex virus, cytomegalovirus, and Epstein-Barr virus, have been implicated in the pathogenesis of a variety of human cancers, and all of them are known to produce tumors in animals and cell transformation in vitro. These viruses also have been shown to infect the esophageal epithelium. Therefore, although many of the key issues of their mechanisms of action are unclear as yet, they should be considered potential etiologic agents of EC. The present review summarizes the data available on the etiology of EC, emphasizing the current evidence implicating an etiologic role of microorganisms in the pathogenesis of this malignancy.


[Colposcopy, histology, cytology and HPV findings in patients with condylomata of the vulva]. The presence of vulvar condylomata acuminata shows evidence HPV infection of the vulva. 49 patients with such lesions came to our hospital for laser treatment. At that time, none of the patients were aware of their cervical disease. Colposcopy and biopsy of the cervix revealed in eight (16%) of the 49 patients CIN I and CIN II in one patient (2%). Condylomata or other HPV-associated lesions were found in seven patients (14%) and cervicitis in two (4%). HPV were detected in the cervix in 23 patients (46%). HPV types 16 and 18 were found in seven patients (14%) and HPV types 31, 33 and 35 in four patients (8%). It is concluded, that patients with condylomata acuminata of the vulva have a high risk of developing CIN and should undergo colposcopy.


Endoscopic needle aspiration biopsy at ERCP in the diagnosis of biliary strictures. We have developed a ball-tipped catheter with a retractable 22-gauge, 7-mm long needle to perform endoscopic needle aspiration (ENA) for cytology and compared this technique to brush cytology of malignant-appearing biliary strictures during ERCP. Of 31 patients, 26 had proven malignant strictures involving the common bile duct and 5 had benign lesions. All 31 patients had ENA and 29 were brushed. Positive ENAs were obtained in 16 of 26 patients (61.5%) and positive brushings in 2 of 24 (8.3%). With the addition of two suspicious ENAs for pancreatic adenocarcinoma, 73% of patients had positive or suspicious cytology for malignancy by combined ENA and brush with a specificity of 100%. Although ENA appeared to be more sensitive in diagnosing cholangiocarcinoma, it proved to be most effective in the diagnosis of pancreatic adenocarcinoma when compared with brush cytology. One patient with cholangiocarcinoma in our series was diagnosed by brush cytology only, with a negative ENA, supporting our recommendation of using both endoscopic brushings and ENA for cytology when evaluating biliary strictures.


Early detection of hepatocellular carcinoma by laparoscopy: yellow nodules as diagnostic indicators. The diagnostic value in cirrhotic livers of yellow nodules undergoing fatty change was evaluated as an early indicator of hepatocellular carcinoma. Laparoscopic biopsy specimens were obtained from 22 yellow nodules in 21 patients with cirrhosis and from 1 non-cirrhotic patient. Six particular findings were recorded: at laparoscopy, (1) on the surface of the liver, the sporadic presence of yellow nodules exceeding 10 mm in diameter, or a scattering of yellow nodules smaller than 10 mm; (2) neovascularity or excessive focal vascularity; in histologic sections, (3) fatty changes; (4)"nodule-within-nodule" formation; (5) increased nuclear to cytoplasmic ratio; and (6) hypercellularity. The greater number of these findings that were observed, the more reliable was the presumed diagnosis of concomitant hepatocellular carcinoma. We conclude that the appearance of yellow nodules on the surface of the liver is a sensitive indicator of hepatocellular carcinoma and that laparoscopy thus can be of distinct value in early diagnosis.


[In vivo and in vitro effects of GnRH analogs on ovarian Leydig cell tumor]. A 65-year old patient, suspected to be suffering from an androgen producing ovarian tumour, was treated preoperatively with the GnRH agonist triptorelin (500 micrograms/day s.c.) for 7 days. After an initial rise, gonadotrophin levels were suppressed under this treatment. The elevated serum testosterone concentrations were reduced by approx. 50% by the triptorelin injections. After the extirpation of the tumour (histologically a Leydig cell tumour of the ovary without signs of malignancy), primary cell cultures which secreted testosterone and androstenedione were prepared. Coincubation of the tumour cells with the GnRH agonist triptorelin had no effect on their androgen secretion. Treatment of the tumour cells with high concentrations (10(-5) M) of a GnRH antagonist, however, resulted in a 100% increase of their testosterone and androstenedione secretion. GnRH-binding sites of low affinity (Ka = 0.54 x 10(5) M-1) and high capacity (B max = 1364 x 10(-12) M/mg membrane protein) were identified in the tumour. These findings suggest that GnRH analogues might modify androgen secretion of sex-cord stromal tumours of the ovary via the suppression of endogenous gonadotrophin secretion and possibly also via direct effects on the tumour cells.


[Magnetic resonance tomography in follow-up of hydatidiform mole]. The value of magnetic resonance imaging (MRI) was studied in a case of a gestational trophoblastic tumour. In addition to the tumour size, MRI made it possible to measure necrotic areas, which are assumed to be a sign of response to chemotherapy. Moreover, uterine zonal anatomy and subserosal uterine veins yielded important information on tumour biology. Complete remission was induced in high-risk patients by etoposide, methotrexate, and actinomycin D treatment.


[A rare clinical manifestation of a combination tumor of cystosarcoma phylloides malignum and an intraductal cancer]. A combination of cystosarcoma phylloides and ductal carcinoma in the breast of a 47-year old woman is reported. Only few cases of this simultaneous tumour formation is described in the literature. Therapy, prognosis and follow-up are discussed. Current standard therapy is ablatio mammae, modified according to Patey, with axillary lymphnode dissection. Decision in respect of chemotherapy depends on the results of staging. Follow-up control of the patients for early detection and therapy of metastasis is recommended. The reported case underlines, that histological examination of every breast node is indicated, even if there is no sign of malignancy.


Chronic ethanol intake inhibits both the vasoactive intestinal peptide binding and the associated cyclic AMP production in rat enterocytes. 1. Chronic ethanol intake during 6, 8, 10 or 12 weeks resulted in a decrease of 125I-vasoactive intestinal peptide (VIP) binding to rat enterocytes. 2. Native peptide displaced 125I-VIP binding to enterocytes, exhibiting a IC50 at about 4 nM native VIP in control and ethanol-treated animals. 3. The number of binding sites in ethanol-treated animals were significantly diminished when compared to control animals. This reduction is observed in both the high-affinity and the low-affinity binding sites. 4. Increasing concentrations of native VIP produced a similar cyclic AMP rise in enterocytes from control or ethanol-treated rats during 6 weeks. However, after 8 weeks of ethanol treatment, a significant decrease in cyclic AMP production stimulated by VIP was observed.


Rabbit isolated vas deferens possess A1 and A2 adenosine receptors. 1. The effects of adenosine, 5'-N-ethylcarboxamidoadenosine (NECA), 2-chloroadenosine, N6-phenylisopropyladenosine (L-PIA and D-PIA) and N6-cyclohexyladenosine (CHA) were examined on the rabbit isolated vas deferens. 2. All the analogues in a concentration-dependent manner inhibited contractile response to electrical stimulation. 3. 8-Phenyltheophylline caused a rightward shift of all the adenosine and its analogues' concentration-response curves. 4. The order of potency for the adenosine and its analogues on the rabbit isolated vas deferens was: CHA = NECA greater than L-PIA greater than 2-chloroadenosine greater than D-PIA greater than adenosine. 5. It is concluded that CHA, NECA, L-PIA, 2-chloroadenosine, D-PIA and adenosine mediate their inhibitory effects on the rabbit isolated muscle via both A1 and A2 adenosine receptors.


Pharmacological profile of the 5-hydroxytryptamine receptor that mediates relaxation of rat oesophageal smooth muscle. 1. The pharmacological profile of the inhibitory 5-hydroxytryptamine (5-HT) receptor in rat oesophageal smooth muscle has been characterized by means of a series of agonists active at 5-HT1-, 5-HT2-, 5-HT3- and 5-HT4-receptor sites, and a broad range of antagonists. The possible involvement of cyclic nucleotides in the 5-HT response was also examined. 2. Under conditions of tone induced by muscarinic receptor activation, the upper two-thirds (proximal segment) of the oesophageal smooth muscle tunic was more sensitive to the inhibitory effects of 5-HT receptor agonists when compared with the distal region. 3. The inhibitory response to 5-HT was blocked by MDL 72222 (5-HT3 antagonist) and ICS 205-930 (5-HT3/5-HT4 antagonist) but not by antagonists active at 5-HT1- or 5-HT2-receptors. 4. The phosphodiesterase inhibitor, 3-isobutyl-methyl-xanthine (IBMX) enhanced oesophageal smooth muscle inhibitory response to 5-HT, isoprenaline and forskolin, but not that elicited by the potassium channel opener, BRL 34915. 5. 5-HT increased tissue cyclic AMP content over basal levels in proximal and distal segments of oesophageal smooth muscle. However, 5-HT had no significant effect on basal cyclic GMP levels in both segments. 6. We conclude that the inhibitory 5-HT receptor in rat oesophageal smooth muscle may represent a high affinity subtype which is sensitive to 5-HT3/5-HT4 antagonists and is coupled to the cyclic AMP pathway.


Modulation of Na+/K+ pump in intact erythrocytes by cardioglycosides, steroid hormones and ouabain-like compounds. 1. Pure erythrocytes preparations, free from platelets and white cells, were incubated for a long time without hemolysis. 2. Dose-response experiments performed with (a) cardioglycosides (ouabain and K-strophantoside), (b) steroid hormones and their glucuronides (tetrahydrocortisol, oestradiol and the respective 3-glucuronic derivatives) and (c) ouabain-like compounds purified in our laboratory (0.7 kDa and 2-4 kDa respectively) emphasise a modulatory effect [activation of Na+ efflux rate and K+ uptake at very low ligand concentrations, inhibition at higher levels; maximum enhancement of cation transport: (a) and (b) 10-0.1 nM (+40-50%), (c) 1-0.01 nM (2.5-fold)]. 3. Binding experiments show upward-curved Scatchard graphs, with the Kd values of 50 nM and 18 microM and the Bmax values of 10.2 and 984.5 fmol/100 microliters RBC (red blood cells) respectively.


The effect of chronic and acute administration of deuterium oxide (D2O) on vascular smooth muscle contraction in spontaneously hypertensive and Wistar-Kyoto rats. 1. Oral administration of 25% D2O for 12 days reduced blood pressure of spontaneously hypertensive rats (SHR) to the level of Wistar-Kyoto (WKY) controls. 2. However, the chronic D2O treatment appeared to have little effect on the phenylephrine and potassium chloride induced dose-response curves of SHR and WKY rats, producing a decreased maximal contraction of the potassium chloride dose-response curve of SHR only. 3. Further acute studies revealed that desensitization results from chronic exposure to D2O such that 60% D2O produces a significant depression of contraction only in aortic rings obtained from SHR and WKY which had not been chronically treated with 25% D2O.


Renin and angiotensin converting enzyme in elasmobranchs. Renin-like activity (RLA) and angiotensin I converting enzyme-like activity (ACELA), the two key enzymes of the renin-angiotensin system (RAS), were sought in the elasmobranch Scyliorhinus canicula. Renal extracts were desalted in a G-25 and eluted in a G-100 Sephadex column (calibration 15,000-70,000). The fractions were concentrated in a vacuum device. A 48,000-MW fraction incubated with synthetic and porcine angiotensiongen generated angiotensin I estimated by RIA. This same fraction was vasopressor in rats and dogfish. ACELA was sought in gill, heart, liver, spleen, pancreas, intestine, kidney, gonads, brain, skin, and muscle of dogfish using a spectrophotometric assay. The highest level of ACELA was found in the gills followed by spleen, kidney, and brain (33.79 +/- 2.3, 29.56 +/- 1.0, 14.62 +/- 1.0, and 13.80 +/- 2.3 nmol hippurate/min/mg protein, respectively). Intestine, gonads, skin and muscle contained no measurable amounts of ACELA. Captopril inhibited enzymatic activity from all ACELA containing tissues.


In vitro study of the effect of adenosine on frog adrenocortical cells. Previous reports have shown that adenosine in rat inhibits both spontaneous and ACTH-induced release of corticosteroids through activation of adenosine A1 receptors. In the present study, we have investigated the possible effect of adenosine in the secretion of corticosteroids in amphibians using a perfusion technique for frog adrenocortical slices. Infusion of adenosine, at concentrations ranging from 10(-7) to 10(-4) M, had no effect on the basal output of corticosterone and aldosterone by frog interrenal cells. Similarly, adenosine did not affect the response of frog adrenocortical slices to ACTH, vasoactive intestinal peptide, or angiotensin II. The stable adenosine A1 receptor agonist N6-phenylisopropyl adenosine (PIA) was also totally devoid of effect on the spontaneous or ACTH-induced release of corticosteroids. These results show that in amphibians, adenosine does not modulate adrenal steroidogenesis.


Detection and partial characterization of proopiomelanocortin-related end-products from the pars intermedia of the toad, Bombina orientalis. Steady-state analyses were performed on the proopiomelanocortin (POMC)-related end-products present in acid extracts of the pars intermedia of the anuran amphibian, Bombina orientalis. Sephadex G-75 gel filtration chromatography indicated that immunoreactive alpha-MSH-sized material and N-acetylated beta-endorphin-related material are the major POMC-related products present in this tissue. The alpha-MSH-sized immunoreactivity was further fractionated by reversed phase HPLC. The major peak of immunoreactivity isolated by this procedure eluted with the same retention time as synthetic ACTH(1-13)amide. Cation exchange chromatography supported the conclusion that the major storage form of alpha-MSH in the pars intermedia of Bombina is ACTH(1-13)amide. Analysis of Bombina pars intermedia in culture indicated that mono-acetylated and di-acetylated alpha-MSH were the major forms of alpha-MSH secreted into the medium. The major peak of N-acetylated beta-endorphin-related material was further analyzed by cation exchange chromatography and Sephadex G-25 gel filtration column chromatography. The major storage form of beta-endorphin in this tissue is N-acetylated, has a net positive charge at pH 2.75 of +1, and has an apparent molecular weight of 1.2K. The beta-endorphin present in the pars intermedia of this tissue does not undergo further N-acetylation at the time of secretion. These results indicate that in the pars intermedia of the archaeobatrachian, Bombina orientalis, the N-acetylation of alpha-MSH is a cosecretory processing event, whereas N-acetylation of beta-endorphin is a post-translational processing event. These results are compared to other archaeobatrachian and neobatrachian pituitary POMC systems that have been analyzed.


Evidence for ultra-short-loop feedback in ACTH-induced interrenal steroidogenesis in coho salmon: acute self-suppression of cortisol secretion in vitro. Interrenal tissues from coho salmon (Oncorhynchus kisutch) were incubated in a defined medium under blood-gas atmosphere at 17 degrees. Rates of cortisol secretion by tissues incubated in media containing 50 mU/ml porcine-ACTH were initially much greater than those of resting tissues in hormone-free media, but after 3 to 6 hr returned to resting rates. The time course of cortisol accumulation in ACTH-containing media was the same when tissues were incubated in different volumes; the final concentrations of cortisol in these incubations were similar to each other and resembled peak in vivo concentrations in juvenile coho subjected to acute stress. Cortisol secretion rates of tissues sequentially transferred to fresh ACTH-containing media every 6 hr did not return to resting levels but remained elevated for at least 24 hr. Cortisol secretion in response to ACTH was attenuated or completely abolished in tissues incubated in media containing exogenous cortisol; this effect was reversible and dose-dependent. Our results suggest that in coho salmon, cortisol may exert ultra-short-loop negative feedback directly at the level of the interrenal gland to effect self-suppression.


Use of long-chain fatty acid-CoA ligase (AMP-forming) from Pseudomonas fragi for the 'in vitro' synthesis of natural penicillins. Five different naturally occurring penicillins containing as side chains hexanoic, trans-3-hexenoic, heptanoic, octanoic or trans-3-octenoic acids have been synthesized 'in vitro' by coupling long-chain fatty acid-CoA ligase (AMP-forming) (EC 6.2.1.3) from Pseudomonas fragi (LFCoA-L) with acyl-CoA: 6-aminopenicillanic acid acyltransferase (AT) from Penicillium chrysogenum. The quantity of penicillin produced was directly related with the carbon length of the side chain precursor tested, being maximal with octanoic acid. Fatty acids with a lower length than C5 were not recognized as substrates and nor were certain aromatic molecules.


De novo glucan synthesis by mutants streptococcal glucosyltransferases present in pellicle promotes firm binding of Streptococcus gordonii to tooth surfaces. Adherence of 3H-labelled cells of Streptococcus gordonii and Streptococcus milleri to artificial pellicles prepared from saliva supplemented with glucosyltransferases from mutants streptococci was examined using a new assay for sucrose-dependent cell-to-pellicle attachment. Results indicate that S. gordonii, but not S. milleri, could attach tightly to hydroxylapatite surfaces through de novo glucan synthesis by mutants streptococcal glucosyltransferases present in the experimental salivary pellicles.


The mechanism of inhibition by EDTA and EGTA of methanol oxidation by methylotrophic bacteria. Ethyleneglycol (aminoethylether) tetra-acetic acid (EGTA) was shown to be a potent competitive inhibitor of electron transfer between methanol dehydrogenase (MDH) and its electron acceptor cytochrome cL. Addition of Ca2+ ions relieved the inhibition by removal of the inhibitory EGTA. Removal of EGTA by gel filtration completely relieved the inhibition. EGTA did not remove the tightly bound Ca2+ present in the MDH. Indo-1, a fluorescent analogue of EGTA, bound tightly to MDH in a 1:1 ratio but not to cytochrome cL; binding was prevented by EGTA. It was concluded that EGTA inhibits methanol oxidation by binding to lysyl or arginyl residues on MDH thus preventing docking with cytochrome cL.


Analysis of mitotic and meiotic defects in Saccharomyces cerevisiae SRS2 DNA helicase mutants. The hyper-gene conversion srs2-101 mutation of the SRS2 DNA helicase gene of Saccharomyces cerevisiae has been reported to suppress the UV sensitivity of rad18 mutants. New alleles of SRS2 were recovered using this suppressor phenotype. The alleles have been characterized with respect to suppression of rad18 UV sensitivity, hyperrecombination, reduction of meiotic viability, and definition of the mutational change within the SRS2 gene. Variability in the degree of rad18 suppression and hyperrecombination were found. The alleles that showed the severest effects were found to be missense mutations within the consensus domains of the DNA helicase family of proteins. The effect of mutations in domains I (ATP-binding) and V (proposed DNA binding) are reported. Some alleles of SRS2 reduce spore viability to 50% of wild-type levels. This phenotype is not bypassed by spo13 mutation. Although the srs2 homozygous diploids strains undergo normal commitment to meiotic recombination, this event is delayed by several hours in the mutant strains and the strains appear to stall in the progression from meiosis I to meiosis II.


cis- and trans-acting suppressors of a translation initiation defect at the cyc1 locus of Saccharomyces cerevisiae. The cyc1-362 mutant of Saccharomyces cerevisiae is deficient in iso-1-cytochrome c as a consequence of an aberrant ATG codon that initiates a short open reading frame (uORF) in the cyc1 transcribed leader region. We have isolated and characterized functional revertants of cyc1-362 in an effort to define cis- and trans-acting factors that can suppress the effect of the uORF. Genetic and DNA sequence analyses have defined three classes of revertants: (i) those that acquired point mutations in the upstream ATG (uATG), restoring iso-1-cytochrome c to its normal level; (ii) substitution of the normal A residue at position -1 relative to the uATG by either C or T, enhancing iso-1-cytochrome c production from approximately 2% to 6% (C) or 10% (T) of normal, indicating that the nucleotide immediately preceding the initiator codon can affect the efficiency of AUG start codon recognition and that purines are preferred over pyrimidines at this site; and (iii) extragenic suppressors that enhance iso-1-cytochrome c expression to 10-40% of normal while retaining the uATG. These suppressors are represented by five different genes, designated sua1-sua4 and sua6. In contrast to the previously described sua7 and sua8 suppressors, they do not compensate for the uATG by affecting cyc1 transcription start site selection. Potential suppressor mechanisms are discussed.


DNA position-specific repression of transcription by a Drosophila zinc finger protein. Expression of the yellow (y) gene of Drosophila melanogaster is controlled by a series of tissue-specific transcriptional enhancers located in the 5' region and intron of the gene. Insertion of the gypsy retrotransposon in the y2 allele at -700 bp from the start of transcription results in a spatially restricted phenotype: Mutant tissues are those in which yellow expression is controlled by enhancers located upstream from the insertion site, but all other structures whose enhancers are downstream of the insertion site are normally pigmented. This observation can be reproduced by inserting just a 430-bp fragment containing the suppressor of Hairy-wing [su(Hw)]-binding region of gypsy into the same position where this element is inserted in y2, suggesting that the su(Hw)-binding region is sufficient to confer the mutant phenotype. Insertion of this sequence into various positions in the y gene gives rise to phenotypes that can be rationalized assuming that the presence of the su(Hw) protein inhibits the action of those tissue-specific enhancers that are located more distally from the su(Hw)-binding region with respect to the promoter. These results are discussed in light of current models that explain long-range effects of enhancers on gene expression.


HAP1 positive control mutants specific for one of two binding sites. The expression of the yeast CYC1 and CYC7 genes is controlled by the HAP1 activator. A GAL4-like zinc finger (residues 1-148) specifies binding to the dissimilar sites UAS1 (of CYC1) and CYC7, and an acidic domain (residues 1307-1483) is essential for activation of transcription. To analyze how HAP1 binds to UAS1 and CYC7, we performed saturation mutagenesis of the DNA-binding domain and recovered mutants with altered activity. Class 1 mutants had a reduced activity at both UAS1 and CYC7, and class 2 mutants selectively eliminated activity at CYC7. Surprisingly, several mutants of both classes exhibited wild-type DNA binding, indicating that they were specifically defective in activation. These positive control (PC) mutants alter residues that bracket the zinc finger. We explain these mutants in a model involving cofactor proteins that bind UAS1 and CYC7 along with HAP1. The existence of PC mutants that only affect activity at CYC7 raises the possibility that different cofactors may exist for UAS1 and CYC7.


The construction of Streptomyces cyaneus genomic libraries in Escherichia coli is dependent upon the use of Mcr-deficient strains. Streptomyces cyaneus genomic DNA ligated into either lambda phage or plasmid vectors was very inefficiently cloned into standard Escherichia coli host strains. However, the same material could be efficiently cloned using Mcr-deficient E. coli strains. These results suggest that the S. cyaneus genome contains 5-methylcytosine residues, some of which occur within the recognition sequences of the E. coli Mcr restriction system.


A new family of polymorphic metallothionein-encoding genes MTH1 (CUP1) and MTH2 in Saccharomyces cerevisiae. By pulsed-field gel electrophoresis of chromosomal DNA and hybridization with a cloned MTH1 (CUP1) gene, we determined the locations of metallothionein-encoding gene sequences on chromosomes in monosporic cultures of 76 natural strains of Saccharomyces cerevisiae. Most of the strains (68) exhibited a previously known location for the MTH sequence on chromosome (chr.) VIII. Seven strains (resistant or sensitive to Cu2+) showed a MTH sequence in a new locus, MTH2, on chr. XVI. One strain carried an MTH locus on both chromosomes VIII and XVI. Restriction fragment and Southern blot analyses showed that the two MTH loci were very closely related. The strains displayed heterogeneity in the size and structure of their MTH2 locus. The length of the repeat unit of MTH2 varied: a 1.9-kb or 1.7-kb unit was found, instead of the 2.0-kb unit of the MTH1 locus. The most resistant strain (resistant to 1.2 mM CuSO4) contained a 0.9-kb repeat unit in addition to those of 1.9 kb and 1.7 kb. All three sensitive (to over 0.3 mM CuSO4) strains with an mth2 locus had a repeat unit of 1.9 kb or 1.7 kb, suggesting the presence of at least two copies of the MTH2 gene, with one always being in the junction area outside of the repeat unit. A monogenic tetrad segregation of 2:2 was usually found in crosses of resistant MTH2 and sensitive mth2 strains. Hybrids between strains with different MTH loci in all combinations showed low ascospore viability, suggesting that the complete lack of an MTH locus may lead to the death of segregants on YPD medium. The MTH1 and MTH2 loci were exchangeable. Strains with a high level of Cu2+ resistance were also resistant to Cd2+. However, these two properties did not cosegregate in heterozygotic hybrids.


Mammalian expression vectors with modulatable promoters and two multiple cloning sites. To facilitate the use of a wide range of selectable markers in transfection studies with human cells, in conjunction with the use of modulatable promoters for regulated expression of the genes of interest, we constructed two pUC19-based mammalian expression vectors, each containing two lacZ alpha-based multiple cloning sites (MCS). Selectable markers can be inserted into the MCS derived from pUC19, and the recombinants can be screened by lacZ complementation. The genes of interest can be inserted into the second MCS. The new MCS contains an amber stop codon in-frame with translation of the LacZ alpha-peptide. The presence of insert in the second MCS can also be screened on XGal plates, but in an Escherichia coli host containing an amber suppressor gene. Expression of the genes of interest can be modulated through transcription from the promoter of the mouse metallothionein-I-encoding gene or the long terminal repeat of the mouse mammary tumor virus. These vectors, as well as several of the intermediate plasmids described in this report, can be used to clone any two genetic elements into a single plasmid.


Sequences of the bovine herpesvirus 1 homologue of herpes simplex virus type-1 alpha-trans-inducing factor (UL48). A virion protein of herpes simplex virus type-1, called Vmw65, alpha TIF or VP16, interacts with cellular transcription factors to transactivate immediate early viral genes. We have cloned and determined the nucleotide sequence of the gene encoding the homologous protein in bovine herpesvirus 1 (BHV-1). The amino acid sequence of the BHV-1 protein is similar to that of alpha TIF, except in the C-terminal one-third of the protein. Since the ability of alpha TIF to activate transcription is dependent on this region, our results suggest that the BHV-1 homologue either does not act as a transactivator or activates genes by a different mechanism.


Cloning and integration of DNA fragments in human cells via the inverted terminal repeats of the adeno-associated virus 2 genome. In current systems for molecular cloning of eukaryotic genes, bacterial cells are routinely utilized as intermediate hosts. We investigated the possibility of using a viral system for cloning DNA fragments independent of bacterial cell usage. In this report, we provide an alternative approach for molecular cloning of DNA fragments in eukaryotic cells by utilizing the inverted terminal repeats (ITRs) of the genome of a nonpathogenic human parvovirus, the adeno-associated virus 2 (AAV). We constructed a series of chimeric linear duplex DNA molecules, ranging in length from 1.8 to 7.2 kb, containing the cruciform structures of AAV-ITRs at both ends. These 'no-end' (NE) DNA structures, when transfected into adenovirus-infected human cells in the presence of AAV replication proteins (Rep), underwent DNA replication. Furthermore, in the presence of AAV capsid proteins (Cap), all replicated DNA molecules of less than 5.0 kb were packaged into mature, biologically active AAV progeny virions. When a chimeric NE DNA (NE-neo) containing a gene (neo) encoding resistance to neomycin was transfected into human cells, neoR clones could be readily isolated in the presence of G418 (Geneticin). Southern-blot analysis of genomic DNA of several independently isolated neoR clones suggested stable integration of the NE-neo DNA into the host chromosomal DNA. AAV-ITRs, therefore, offer an alternative system for molecular cloning, as well as packaging of DNA fragments in mammalian cells independent of bacterial cell usage.


Tissue-specific expression and chromosome assignment of genes specifying two isoforms of subunit VIIa of human cytochrome c oxidase. Subunit VIIa of mammalian cytochrome c oxidase (COX; EC 1.9.3.1) exists in at least two isoforms, one present in all tissue types ('liver' isoform; COX VIIa-L) and the other specific for cardiac and skeletal muscle (COX VIIa-M). We have isolated a full-length cDNA encoding human COX VIIa-M. The deduced polypeptide represents the human ortholog of COX VIIa-M, as it shares 78% identity with bovine COX VIIa-M, but only 63% identity with human COX VIIa-L. Northern-blot analysis of primate tissues demonstrated that COXVIIa-M mRNA is present only in muscle tissues; in contrast, the COXVIIa-L mRNA is present in both muscle and nonmuscle tissues. Southern-blot hybridization of human-rodent cell hybrid genomic DNA indicates that the COXVIIa-M gene maps to a single locus on chromosome 19, designated COX7AM. In contrast, COXVIIa-L cDNA probes hybridized to fragments from two COX7AL loci, on chromosomes 4 and 14.


Differential expression of genes specifying two isoforms of subunit VIa of human cytochrome c oxidase. Subunit VIa of mammalian cytochrome c oxidase (COX; EC 1.9.3.1) exists in two isoforms, one present ubiquitously ('liver' isoform; COX VIa-L) and the other present only in cardiac and skeletal muscle (COX VIa-M). We have now isolated a full-length cDNA specifying human COX VIa-M. The deduced mature COX VIa-M polypeptide is 62% identical to the human COX VIa-L isoform, but is approximately 80% identical to the bovine and rat COX VIa-M isoforms, suggesting that the two COX VIa isoform-encoding genes arose prior to the mammalian radiation. Transcriptional analysis showed a tissue-specific pattern: whereas COXVIa-L is transcribed ubiquitously, COXVIa-M is transcribed only in heart and skeletal muscle. The cDNA specifying COX VIa-M is a prime candidate for use in investigations of Mendelian-inherited COX deficiencies with primary involvement of muscle.


Mutational analysis of the Escherichia coli serB promoter region reveals transcriptional linkage to a downstream gene. Genes encoding proteins with unrelated functions can be cotranscribed, and this may be used by cells to coordinate different metabolic pathways during growth. We describe a gene, designated sms, which is downstream from the serine biosynthetic gene serB in Escherichia coli but does not appear to be involved in amino acid (aa) biosynthesis. The sms gene is 1380 bp long. The Sms product migrates at 55 kDa on sodium dodecyl sulfate(SDS)-polyacrylamide gels and has a M(r) of 49472 (460 aa residues) calculated from the nucleotide sequence. The deduced Sms aa sequence shares regions of similarity with two ATP-dependent proteases, Lon and RecA, and contains two motifs: a C-x(2)-C-x(n)-C-x(2)-C motif, which is found in some nucleic acid binding proteins, and an ATP/GTP binding site motif. Insertional inactivation of sms led to increased sensitivity to the alkylating agent methylmethane sulfonate, but not to a requirement for serine or other metabolites. Several promoter mutations were isolated and characterized, which suggest that serB has a typical promoter recognized by sigma 70. After the serB coding sequence there is a 48-bp region with no obvious promoter sequence preceding the sms translation start codon. Analyses using sms'-lacZ fusions cloned downstream from wild-type and mutant serB promoters showed that sms is cotranscribed with serB.


Novel streptococcal-integration shuttle vectors for gene cloning and inactivation. Seven new streptococcal integration shuttle vectors have been constructed which contain different antibiotic-resistance-encoding genes capable of expression in both Streptococcus sp. and Escherichia coli. These plasmids can replicate in E. coli, but not in streptococci because of the absence of a streptococcal origin of replication. The size, antibiotic resistance, and number of unique restriction sites available for cloning for each plasmid are as follows: pSF141 (7.6 kb, CmR and KmR, 7 sites), pSF143 (5.7 kb, TcR, 6 sites), pSF148 (7.3 kb, CmR and SpR, 7 sites), pDL285 (3.4 kb, KmR, 3 sites), pDL286 (3.1 kb, SpR, 4 sites), pSF151 (3.5 kb, KmR, 10 sites), pSF152 (3.2 kb, SpR, 9 sites). If these plasmids carry a fragment of streptococcal DNA they can specifically integrate into the chromosome via Campbell-like, homologous recombination. Therefore, they should be useful for gene inactivation, cloning, chromosomal walking, or linkage analysis in streptococci. The availability of these integration plasmids resistant to different antibiotics, along with the previously described plasmid, pVA891 (ErR), should also allow the construction of mutants possessing multiple insertionally inactivated genes useful for a variety of genetic studies.


Role of the histone-like proteins OsmZ and HU in homologous recombination. The HU protein of Escherichia coli has been implicated in various site-specific recombination reactions. Moreover, recent data suggest that HU may also participate in homologous recombination. In particular, it has been shown that P1 transduction is inhibited in the absence of HU [Kano and Imamoto, Gene 89 (1990) 133-137]. In contrast, we found that transductional recombination and conjugational recombination were almost normal in hupA hupB mutants. However, it appeared that the recombination proficiency of hupA hupB mutant bacteria was reduced tenfold in an intrachromosomal recombination assay. Moreover, we found that intrachromosomal recombination was reduced tenfold in a gyrB226 strain and by more than 100-fold in an osmZ205 strain. The gyrB226 mutation affects the DNA gyrase activity, while mutations in osmZ are highly pleiotropic, affecting the expression of a variety of genes and increasing the frequency of site-specific inversion events. Since it has been shown that the hupA hupB mutations, like the gyrB226 mutation, decrease the level of DNA supercoiling, whereas the osmZ205 mutation increases the level of DNA supercoiling, it appears that the histone-like proteins HU and OsmZ may play a key role in intrachromosomal recombination by affecting the DNA topology.


Sequence of ISRm4 from Rhizobium meliloti strain GR4. ISRm4, an IS-like sequence structurally similar to Pseudomonas cepacia insertion element IS402, was identified by sequence analysis. This 933-bp element carries 17-bp putative terminal inverted repeats with five mismatches and a putative direct target duplication of 3 bp.


Analysis of the genes and gene products of Xanthomonas transposable elements ISXc5 and ISXc4. A series of deletion mutants have been constructed for the gene analyses of transposable elements, ISXc5 and ISXc4, derived from Xanthomonas. At least two element-encoded polypeptides of 48 kDa and 40 kDa have been identified in the minicell-producing Escherichia coli strain, TC410. A study of the element transposition and cointegrate resolution revealed that the 48-kDa and 40-kDa polypeptide are both involved in translocation of the elements, the 48-kDa product being involved in transposition of the elements and the 40-kDa product being involved in cointegrate resolution.


A selective lambda phage cloning vector with automatic excision of the insert in a plasmid. A bacteriophage lambda cloning vehicle has been constructed for the generation of cDNA libraries. The vector has the following properties. (1) It has a unique BamHI site engineered into the lambda gam gene. Segments of DNA can be cloned into this site and clones with an insert can be selected by their ability to grow on an Escherichia coli host lysogenic for phage P2 (Spi- phenotype). (2) When the recombinant phage infects a Cre-producing E. coli strain, a site-specific recombination event results in the excision of a plasmid replicon with the cloned insert. (3) Single-stranded DNAs can be recovered by growing helper M13 phages on bacteria harboring such plasmids. The vector, lambda MGU2, has been used to construct a nematode (Caenorhabditis elegans) cDNA library.


Replication of bovine papillomavirus vectors in murine cells. Varying capacities for autonomous replication have been obtained with bovine papillomavirus type 1 (BPV-1)-based expression vectors in mouse C127 cells. Both integration of the vector DNA into the genome of the host cell and replication as monomeric extrachromosomal elements have been observed. In this study, we have examined what features of BPV-1 vectors influence their replication potential. Transfection of the entire BPV-1 genome into C127 cells resulted in the replication of extrachromosomal monomeric BPV-1 elements. The same result was obtained when a plasmid sequence was inserted into the BPV-1 DNA. However, introduction of foreign, transcriptionally active units resulted in chromosomal integration of the expression vectors. This result was obtained with clones isolated by co-transfection followed by neomycin selection, as well as with clones isolated from neoplastic foci. Supertransfection of a BPV-1-based expression vector into cells harbouring unintegrated replicating BPV-1 genomes resulted in integration of the vector DNA, whereas replication of the resident BPV-1 genomes was unaffected. Extrachromosomal replication of such a vector was achieved when the enhancer and promoter region of the foreign gene were deleted.


Characterization of a LINE retroposon dispersed in the genome of three non-sibling Aedes mosquito species. A family of long interspersed repetitive elements (LINEs) dispersed in the genome of Aedes mosquitoes is described. Basically, full-length copies of the element designated Juan-A are dispersed in the genome of A. aegypti, but some elements are truncated or deleted. Complete Juan-A elements are 4.7 kb long, and their overall genetic organization is similar to that of LINEs from other species in which this class of nonviral retrotransposons has been described. Juan-A elements are terminated at the 3' end by an adenosine(A)-rich sequence and are flanked by target-site duplications. They display two long open reading frames potentially encoding two polypeptides. The first one contains Cys-rich motifs typical of nucleic-acid-binding proteins, while the other shows homology to the reverse transcriptases. These features are characteristic of LINE retroposons and indicate that Juan-A elements can be transposed by reverse transcription of an RNA intermediate. Furthermore, Juan-A retroposons display significant homologies with the Drosophila LINEs Jockey and F, suggesting that all these elements have arisen from a common precursor. The full-length Juan-A copies which are amplified in the genomes of various strains belonging to the three non-sibling species, A. aegypti, A. albopictus and A. polynesiensis, form an internally homogeneous family. These data are interpreted to indicate that active Juan-A retroposons underwent a recent amplification in the strains analyzed. Furthermore, they suggest that these elements have spread by horizontal transfer between the three non-sibling species.


Construction of a new universal vector for insertional mutagenesis by homologous recombination. We describe here the construction of a vector (pSSC-9) which can be used for the insertional mutagenesis of any gene for which genomic sequences have been cloned. This vector contains a neomycin-resistance-encoding gene (neoR) which is driven by a modified thymidine kinase (tk) promoter for positive selection. Flanking neoR are two tk genes driven by their own promoters for negative selection of nonhomologous insertions. The neoR and tk cassettes are separated by four unique cloning sites on the right-hand side of the neoR cassette and three unique sites on the left-hand side. The vector also includes two SfiI sites, one on each side of the tk cassettes, for the excision of the cloned genomic DNA fragments along with the selectable markers. Electroporation of pSSC-9 into mouse embryonic stem (ES) cells and cultured diploid mouse adrenal Y-1 cells conferred resistance to G418 and sensitivity to ganciclovir in both cell lines. These results illustrate the expression of the positive and negative selectable markers in two different cell lines and thus suggest that the vector could be used in ES cells, as well as in cultured somatic cells.


Sequence of a rat cDNA encoding the ERK1-MAP kinase. Mitogen-activated protein (MAP) kinases are cytoplasmic and/or nuclear protein kinases which are activated by one or several signal transduction pathways from the cell surface into the nucleus. Their activity is regulated by phosphorylation on Tyr as well as on Ser/Thr residues. A cDNA encoding the rat ERK1 member of the MAP kinase family was isolated and sequenced. The longest cDNA consisted of 1875 nucleotides and coded for a polypeptide of 380 amino acids with a predicted M(r) of 42987.


Sequence comparisons for a hepatitis C virus genome RNA isolated from a patient with liver cirrhosis. The nucleotide (nt) sequence was determined for an isolate of hepatitis C virus (HCV) derived from cirrhotic tissue of a patient with hepatocellular carcinoma. The 9408-nt sequence (EMBL Acq. No. X61596) showed homology of 90.7-91.4% on the nt level, as compared to two Japanese isolates from patients with a high titer of serum transaminase, 78.4-78.8% to those obtained in the United States, and 65.0% to that from an asymptomatic Japanese carrier. The phylogenetic tree of the six isolates classified them into three groups.


Dual mode cytotoxicities of reactive oxygens on L5178Y radiosensitive mutant M10. Cytotoxic effects of O2- and H2O2 on mammalian cells were investigated in comparison with the relative sensitivity of mouse L5178Y cells and its radiosensitive mutant M10. Both O2- and H2O2 exhibited two different modes of cytotoxic actions depending on their exposure rates: At a high exposure rate (4.3 nmol of O2-/mL/min), M10 was more sensitive to O2- than L5178Y normal type cell, in agreement with the case of X-rays; while at a low exposure rate (five times less than the high exposure rate), M10 became more resistant than L5178Y. Similar results were obtained with H2O2. Reactive species responsible for these two different cytotoxic actions were examined with special reference to the metal-catalyzed Haber-Weiss reaction, by using a metal chelator, 1,10-phenanthroline, and an .OH scavenger, dimethylsulfoxide (DMSO). Significant protection by 1,10-phenanthroline was observed, which indicates the presence of a metal-dependent process in both cytotoxic actions. DMSO showed a marked protective effect, except for the case of M10 exposed at the low exposure rate, in which DMSO showed no protection. The resistance of M10 to O2- and H2O2 observed at the low exposure rate suggests the possibility that reactive species other than .OH are involved in the cytotoxicity.


Formation of aminoxyl radicals in the reaction between penicillins and hydrogen peroxide. Aminoxyl radicals are formed in high yield in the reaction between penicillins and hydrogen peroxide in water solutions in the pH range between 7 and 8. The nine-line EPR spectrum, 3 x 3 (1:2:1), indicated an interaction of the unpaired electron with one 14N nucleus (aN = 1.44 mT) and two equivalent hydrogen nuclei (aH = 2.00 mT). The reaction involves an oxidative cleavage of the beta-lactam ring of the penicillins with the formation of a cyclic aminoxyl radical, in which the thiazolidine ring carries the nitroxide group (= N-O.). It is suggested that the reaction with the formation of aminoxyl radicals can also take place in vivo in the deactivation of penicillins by metabolically formed hydrogen peroxide.


Modulation of human upper intestinal nutrient transit by a beta adrenoreceptor mediated pathway. To explore the role played by beta adrenoreceptor mediated pathways on human upper gut function a series of studies were conducted into the effects of beta adrenoreceptor agonists and antagonists on orocaecal and duodenocaecal transit and on antral and duodenal motor activity. Under control conditions orocaecal transit was consistent within individuals (mean coefficient of variation (18.0%) but varied widely between individuals (median transit 63 minutes, range 33-164). Prior administration of the non-selective beta adrenergic antagonist propranolol consistently hastened orocaecal transit (median transit 51:25-93, v control p < 0.005). The selective beta-1 antagonist, atenolol, also hastened transit (median transit 50:35-93 minutes, v control p < 0.01). The magnitude of an individual's response to beta blockade correlated closely with the orocaecal transit (Tau = 0.54, p < 0.01). Duodenocaecal transit was also hastened by propranolol from control values of 66:45-107 minutes to 50:16-62 minutes, p < 0.025). In contrast neither duodenal nor antral motility were consistently altered by beta blockade. The beta adrenoreceptor agonist, isoprenaline, delayed both orocaecal transient (97:55-178 minutes, v control p < 0.005) and also duodenocaecal transit (160:45-215 minutes, v 73:40-133) (p < 0.025). Isoprenaline also reduced antral motility by an effect which appeared to occur predominantly through a reduction in contraction amplitude (from a median amplitude of 27:5.39 mm Hg to 14:3-24 mm Hg, p < 0.03) rather than an effect on the interval between contractions. No effect on either amplitude or frequency of duodenal motor activity was observed. A beta adrenoreceptor mediated pathway thus appears to exert a biologically relevant effect on gut function not only under conditions of sympathetic stimulation, but also at rest when a basal beta adrenergic tone appears to influence the speed of nutrient transit through the human upper gut.


Effect of the dietary fibre content of lifelong diet on colonic cellular proliferation in the rat. The effect of the fibre content of lifelong (18 months) diets on proximal and distal colonic cellular proliferation and short chain fatty acid (SCFA) content was investigated in 40 rats. Rats were fed a low fibre diet (17 g/kg non-starch polysaccharides NSP) or the stock diet (133 g/kg NSP). The higher fibre fed rats had increased caecal and colonic total contents (p < 0.001) and SCFAs than the low fibre fed rats (caecal SCFAs: higher fibre rats 96.4 (6.8) mumol/g wet weight v low fibre 22.7 (3.0): p < 0.001, colonic SCFAs: higher fibre 52.3 (3.1) mumol/g wet weight v low fibre 6.9 (2.2) mumol/g wet weight: p < 0.001). Cellular proliferation was increased in the proximal colon (bromodeoxyuridine labelling index, higher fibre 9.3 v low fibre 8.4 p < 0.05; flow cytometry, % cells in S phase higher fibre diet 7.9 v low fibre 6.9; p < 0.01) and there was a shift of proliferating cells to a higher region in each crypt. There was no significant difference in the percentage of cells in S phase in the distal colon of rats in both diet groups. The proliferative zone, however, was expanded in the distal colon of the higher fibre diet fed rats. This study indicates that long term higher fibre intake in rats is associated with a modest increase in cellular proliferation in the proximal colon but not the distal colon.


Gastrointestinal symptoms in patients infected with human immunodeficiency virus: relevance of infective agents isolated from gastrointestinal tract. The correlation of gastrointestinal symptoms and infections in 186 consecutive patients with human immunodeficiency virus (HIV) infection undergoing diagnostic endoscopy (oesophagogastroduodenoscopy, n = 124; colonoscopy, n = 37; both, n = 25) was investigated. Biopsy and stool samples were examined for infective agents. Only weight loss (p = 0.003) and dysphagia (p = 0.027) were more common in patients at stage CDC IV compared with earlier stages. In three of 27 patients at stage II/III and in 93 of 159 patients at stage IV an infective agent was identified in stool or gastrointestinal biopsy specimen (p < 0.001). Cytomegalovirus (n = 35), Candida sp (n = 28), M avium complex (n = 10), and Cryptosporidium (eight) were the most frequent agents detected. At stage IV, diarrhoea was more frequent in infected compared with non-infected patients (p = 0.006); however, an infective agent was also found in 39 of 82 patients at stage IV without diarrhoea. The frequency of gastrointestinal symptoms was not consistently increased in patients harbouring specific infective agents compared with non-infected patients. Our findings indicate that the pathogenic relevance of a gastrointestinal infection in HIV infected patients has to be verified and indirectly support the existence of an HIV associated enteropathy.


Feeding nursing home residents with Alzheimer's disease. The purpose of this exploratory, descriptive study was to identify and categorize the behaviors of caregivers and residents that elicit, sustain, or extinguish feeding. Although preliminary analysis yielded initial categories, mutually exclusive categories were difficult to distinguish. Relationships among identified cues and specific feeding behaviors are being examined in further analysis. The relationship between specific feeding behaviors and the environmental context of feeding is also being examined. The goals of studying feeding behaviors in nursing home residents with Alzheimer's disease are to enhance mealtime for both residents and caregivers and to encourage functional feeding behavior as much as possible. Identification of behaviors that elicit, sustain, and extinguish functional feeding can facilitate development of behavioral and environmental nursing interventions to promote functional feeding.


Tuberculosis surveillance program: a nursing home experience. Tuberculosis surveillance efforts in the nursing home for elderly persons need organization into an effective program. The ability to tally results of testing gives reliable parameters for comparison and may alert practitioners to early active disease presentation and transmission within the setting. Such efforts based on current research and recommendations ensure provision of prudent and useful standards of elderly care. Our experience during a 10-year period shows that tuberculosis may be an active problem for nursing home populations.


Humor: no geriatric nurse should be without it. Humor is an approach caregivers can use to assist the elderly in facing some of the disappointments of old age. The ability to laugh at oneself, at others, and at situations stimulates trust, commitment, and a positive sense of working together that is therapeutic to both staff and patients. Humor offers a tool for patients, families, and staff who deal with the elderly for stress management, coping, and to facilitate implementation of the nursing process. When humor is used it is important for the nurse to bear in mind that for patients who are cognitively impaired attempts at humor may be misunderstood. Sarcastic humor or humor that does not show respect for individuals should never be used. It is essential that when humor is used a careful assessment for appropriateness be made of the situation. An evaluation must also be made of the effect of the humor on the other persons in the interaction so a positive interpersonal relationship is facilitated.


Lectin histochemistry of sex cord-stromal tumors and small cell carcinoma of the ovaries. Binding sites of peanut agglutinin (PNA), Ulex europaeus (UEA-1), concanavalin A (Con A), and wheat germ agglutinin (WGA) were localized in 10 granulosa cell tumors, 10 Sertoli-Leydig cell tumors, 4 theca cell tumors, and 5 small cell carcinomas. Con A and WGA reacted positively with the majority of the studied neoplasms. PNA and UEA-1 were persistently negative in the sex cord-stromal tumors (SCST) but showed focal positivity in small cell carcinomas. Negative reactions of SCST with PNA and UEA-1 may serve for differentiation between them and common epithelial tumors that are usually PNA and UEA-1 positive.


Safe withdrawal from acute alcohol abuse in the aged. It is imperative to consider chronic and acute alcoholism as possibly underlying problems that aged persons may exhibit. The potential exacerbation of organ dysfunctions, the complexity of medication regimens, and negative interactions of these are all more likely to create problems in the treatment of the elderly alcoholic patient. The nurse, the social worker, the nutritionist, the physician, and the pharmacologist must work as a team to treat these patients properly. The nurse focuses on maintaining patient safety and function, continually assessing, observing, and reassessing. On the basis of these data the pharmacologist collaborates with the physician to manage medications and titrate dosages properly. Malnutrition is a prevalent problem among the aged, particularly older alcoholic men who live alone. The nutritionist must assess dietary deficiencies and recommend therapeutic vitamins, minerals, and foods as needed. It is imperative to evaluate serum levels of folic acid, cyanocobalamin, thiamine, magnesium, and to monitor periodically potassium. Before discharge the social worker assists the patient in beginning an appropriate alcoholic treatment plan. The situation of the acutely ill alcoholic patient requires ongoing and precise collaboration of the team to achieve acceptable outcomes.


[Molecular cardiopharmacology of selective inhibitors of cyclic nucleotide phosphodiesterase isozymes]. The existence of multiple isozymes of cyclic nucleotide phosphodiesterase (PDE) in many tissues including the heart has been demonstrated. Five isozyme families, each composed of several subtypes and having different tissue and subcellular distributions, have been characterized. Selective inhibitors of PDE III (cGMP-inhibited PDE) elevates the cAMP level which mediates positive inotropic actions with compartmentation of cAMP related to cardiac cell particulate structures. Both cardiac cytosolic and particulate PDE III were potently and selectively inhibited by the new cardiotonic agents competitively with respect to cAMP, except for vesnarinone. There might be at least two subtypes of PDE III, and vesnarinone may be a selective subtype inhibitor of PDE III in human heart. It was also reported that vesnarinone was beneficial in treating patients with congestive heart failure. Moreover, selective inhibitors of PDE III with ancillary properties such as calcium sensitization may prove to be more useful drugs for the treatment of heart failure.


[Formation and removal of active oxygen species and lipid peroxides in biological systems]. The mechanisms of formation and removal of active oxygen species and lipid peroxides in biological systems have been briefly reviewed. Cytotoxic active oxygen species can be classified into two types: (a) radical species such as O2-. (superoxide) and HO. (hydroxyl radical) and (b) non-radical species such as H2O2 (hydrogen peroxide) and 1O2 (singlet oxygen). The direct or indirect attack of active oxygen species on polyunsaturated fatty acids, essential constituents of biological membranes, has been shown to result in the formation of a number of peroxidative lipid breakdown-products: LOOH (lipid hydroperoxide), LOO. (lipid peroxyl radical) and LO. (lipid alkoxyl radical). The lipid peroxide decomposition is probably dependent on the presence of ferric-ferrous ions. These processes are called lipid peroxidation reactions. In recent years, there has been a renewed interest in the role played by lipid peroxidation in many disease states. The multiple lines of defense against toxic oxygen intermediates consist of enzymatic systems, glutathione peroxidase, catalase and superoxide dismutase, and furthermore involves antioxidant capacities such as those of vitamin E and vitamin C. In biological systems, there are naturally occurring lipid-soluble (vitamin E and ubiquinone) and water-soluble (vitamin C, reduced glutathione and uric acid) antioxidants. Therefore, so long as homeostasis is maintained between the rate of radical generation and the rate of radical dissipation, the cellular generation of radicals may not be harmful. In contrast, this balance can be disturbed if cellular defenses are decreased or if there is a significant increase in the flux of radical generation. Once lipid peroxidation is initiated, the reactive intermediate formed induces cell damage.(ABSTRACT TRUNCATED AT 250 WORDS)


Hepatotoxicity and hepatocarcinogenicity in rats fed squid with or without exogenous nitrite. The popular seafood squid contains high levels of naturally occurring amines such as dimethylamine (DMA) trimethylamine and trimethylamine-N-oxide (TMAO). The hepatotoxicity and hepatocarcinogenicity of squid with or without exogenous nitrite were investigated in rats. Acute necrosis including polymorphogenic neutrophil infiltration, haemorrhage and cholangiofibrosis were observed in the livers of most rats fed squid. Hepatocellular carcinoma (HCC) was induced in two out of 12 rats (16%) by feeding 10% squid in Purina rat chow for 10 months. The incidence of HCC was increased to four out of 10 rats (33%) when 0.3% NaNO2 was added to the above diet. At the end of the experiment a marked elevation of serum gamma-glutamate transferase was observed in treated groups, but no significant changes in the activities of serum glutamic-oxaloacetic transaminase and glutamic-pyruvic transaminase were detected. Vitamin C (0.3%) gave partial protection against hepatic damage. The concentration of DMA in squid is estimated to be 0.19%; this concentration did not induce HCC under the experimental conditions used. Therefore it is suggested that another major naturally occurring amine in squid, TMAO, could be one of the important factors involved in the induction of hepatotoxicity and hepatocarcinogenicity in rats.


A case report of mixed tumor arising in the vagina. We reported a case of mixed tumor arising in the vagina of a 32-year-old Japanese woman. The tumor was a finger-like polyp protruded from the lower portion of the vagina. The histologic feature of the tumor resembled that of pleomorphic adenoma of the salivary gland, consisting of stromal-type cells embedded in the myxomatous stroma, although there was no cartilagenous component. Immunohistochemical study also suggested resemblance to pleomorphic adenoma of salivary gland, being positively stained with antibodies against cytokeratin, epithelial membrane antigen, vimentin and muscle actin, but negatively with an antibody against S-100 protein. The histogenesis of the tumor is not clear, but it may be originated from myoepithelial cells.


Hypothalamopituitary deficiency and precocious puberty following hyperhydration in diabetic ketoacidosis. We report on a 5-year-old child who survived an intracerebral crisis, following ketoacidosis-revealing diabetes (DKA), with visual impairment due to a vascular occipital lesion. Two and 4 months after the initial episode, a unique hypothalamopituitary disorder consisting in GH, ACTH, TSH deficiencies and central precocious puberty, was detected. Cranial magnetic resonance images showed no visible lesion in the hypothalamopituitary region. The most likely hypothesis is the ischemia of hypothalamopituitary and occipital regions following possible cerebral edema after hyperhydration. She survived with low visual acuteness and received a combined replacement therapy for the neuroendocrinological deficiencies. This case emphasizes that the rehydration at the initial period of DKA is critical, especially when risk factors for cerebral edema are present (young age, marked hyponatremia). The neuroendocrinological consequences of acute cerebral edema are rare, but physicians must be attentive in survivors of these accidents.


Bilateral and simultaneous inferior petrosal sinus sampling in the early diagnosis of an ACTH-producing pituitary microadenoma and its detection by magnetic resonance one year later. This study describes a case of pituitary-dependent Cushing's syndrome where standard biochemical and radiological techniques were discordant in localizing the origin of the autonomous adrenocorticotropic hormone (ACTH) hypersecretion in the pituitary. Hormonal evaluation suggested a pituitary genesis for the disease, but both sellar computed tomography and cranial magnetic resonance (MR) were unable to give clear-cut evidence for a pituitary neoplasm. Simultaneous and bilateral inferior petrosal sinus sampling (SBIPS) correctly identified the left side of the pituitary gland as the source of autonomous ACTH production. One year later, the shaded signs of a pituitary lesion in the left side of the gland were seen with MR imaging, and a 0.5-cm in height adenoma was surgically removed. At the 2-year follow-up the patient's symptoms had completely disappeared, and her menses were restored. In this case, SBIPS correctly diagnosed the presence of an ACTH-secreting pituitary microadenoma one year before shaded signs of the pituitary lesion appeared with MR imaging. This is a clear-cut demonstration of the accuracy of the SBIPS technique in localizing small pituitary lesions.


Lack of control of liver gluconeogenesis in cholestatic rats with reduced portal blood flow. Previous studies indicated a role for ischemia in the metabolic changes induced by cholestasis. Liver pyruvate kinase is a key enzyme for the concurrent control of glycolysis and gluconeogenesis. In this experiment the control of pyruvate kinase activity was investigated in cholestatic rats. Pyruvate kinase kinetics changed from a sigmoidal type in sham-operated rats to a hyperbolic type in obstructed rats. The change in the enzymatic kinetics paralleled the reduction in the portal blood flow, which reached 50% of the control value 7 days after obstruction. Dibutyryl cyclic AMP (5 mumol/kg body wt) plus theophylline 0.1 mmol/L failed to inactivate the enzyme when injected into the portal veins of rats whose livers were obstructed 7 days before. Both the kinetics changes and the lack of phosphorylation control are compatible with ischemia.


Treatment of children with chronic hepatitis C with recombinant interferon-alpha: a pilot study. Twelve children with chronic non-A, non-B hepatitis were entered in a pilot trial of recombinant interferon-alpha. Although all the children had hepatitis C virus RNA in serum, only five had antibodies against this virus. Children received 3 MU/m2 body surface area interferon-alpha 3 times/wk for 6 mo; they were followed for 24 mo, including the therapy period. One child was dropped from the study, so the results are from the 11 children who completed the study. At the end of the therapy period, 36% of the children had normal ALT levels; this percentage increased to 90% at mo 15 of follow-up. Thereafter, relapse occurred in five children; thus ALT normalization was observed in 5 of 11 children at the 24th month. Moreover, two different ALT patterns were found: HCV antibody-negative children had significant peaks of ALT levels with respect to the basal samples (p less than 0.05) until the third month of the therapy; these levels later decreased. In contrast, HCV antibody-positive children had slight fluctuations of ALT until normal levels were reached. At the end of treatment, three children had HCV RNA; one demonstrated a rebound in ALT levels. Finally, histological activity had decreased significantly in the second liver biopsy specimen in all children. In summary, interferon treatment in children with chronic hepatitis C may be helpful, although these results should be confirmed in controlled trials.


Postresection recurrence of hepatocellular carcinoma treated by arterial embolization: analysis of prognostic factors. Of 270 consecutive patients with hepatocellular carcinoma who underwent surgery, 50 who had recurrence and were subsequently treated with transcatheter arterial embolization were analyzed. The longest interval between surgery and recurrence in the 50 patients who underwent transcatheter arterial embolization was 7 yr. Recurrence was initially found in the remnant liver in all patients but one; extrahepatic metastases were detected in 13 patients (26%) during follow-up. A "multiple" type was the most common (64%) hepatic recurrence pattern on angiography, followed by the "solitary" (16%) and "tumor thrombus" (12%) patterns. Hepatic recurrence was most frequently found in the ipsilateral lobe (48%) relative to the site of the primary hepatocellular carcinoma. Multivariate analysis of the factors affecting survival after transcatheter arterial embolization indicated that recurrence pattern (p = 0.025) and distant metastases (p = 0.011) were significant. Of 13 patients with distant metastases, 11 had the "multiple" pattern of hepatic recurrence. Survival rates for all 50 patients after initial surgery and after transcatheter arterial embolization were 90% and 64%, respectively, at 1 yr; 52% and 24%, respectively, at 3 yr; and 27% and 5%, respectively, at 5 yr. On analysis of survival rates after transcatheter arterial embolization in 37 patients with recurrence only in the liver and of the response of recurrent hepatocellular carcinoma to transcatheter arterial embolization, a significant difference was noted between those with "partial response" and "progressive disease" (p less than 0.05) and between those with "no change" and "progressive disease" (p less than 0.05).


Elevated fibrin-related and fibrinogen-related antigens in patients with liver disease. Patients with liver disease have a variety of coagulation abnormalities. These derangements are of uncertain origin and do not always correlate with disease severity or activity. We have measured the levels and proportions of the total fibrin-related and fibrinogen-related antigens, the principal fibrin (D-dimer) and fibrinogen (D-monomer) degradation fragments and intermediates of fibrin formation (fibrin monomers) in patients with a variety of acute and chronic liver diseases in whom all known other precipitating causes of disseminated intravascular coagulation had been excluded. Fibrin-related and fibrinogen-related antigens were extracted from serum using antihuman fibrinogen-IgG covalently bound to activated amino-phenylthioether paper disks and were subjected to 4% to 11% sodium dodecyl sulfate-polyacrylamide gel electrophoresis under nonreducing conditions. Fibrin-related and fibrinogen-related antigen proportions were determined by densitometry, and their levels were measured by radioimmunoassay. Levels of total fibrin-related and fibrinogen-related antigens (and D-dimer) were significantly elevated (p less than 0.01) in patients with cirrhosis (121 to 641 ng/ml) and hepatocellular carcinoma (416 to 8,786 ng/ml) when compared with patients with acute viral hepatitis (84 to 322 ng/ml) and control subjects (38 to 186 ng/ml). In addition, D-monomer levels were elevated. These findings strongly suggest that disseminated intravascular coagulation is a component of the coagulopathy of certain liver diseases. Because fibrin-related and fibrinogen-related antigens have anticoagulant, vasoactive and immunosuppressive properties, their elevated presence may be biologically significant in these patients.


Macroregenerative nodules and hepatocellular carcinoma in forty-four sequential adult liver explants with cirrhosis. Macroregenerative nodules, also called nodules of adenomatous hyperplasia, have been well documented in Japan. Extensive studies support the hypothesis that in the Japanese population these lesions represent a possible pathway for hepatocarcinogenesis. However, reporting of these lesions in non-Japanese populations has so far been rare. We examined 44 sequential cirrhotic hepatectomy specimens from adult patients who underwent orthotopic liver transplantation at our institution. All livers were serially sectioned every 0.5 cm. Macroregenerative nodules were defined as regenerative nodules at least 1 cm in diameter. Forty-eight macroregenerative nodules were found in 11 livers (25% of livers). The antecedent diseases in these livers included hepatitis C (3), alcoholism (2), primary biliary cirrhosis (2) (one with iron overload), cryptogenic cirrhosis (2), hepatitis B (1) and alpha 1-antitrypsin deficiency (1). The macroregenerative nodules often differed from the surrounding nodular parenchyma in color, texture or the degree to which they bulged beyond the cut liver surface. Three livers contained grossly apparent hepatocellular carcinomas. Microscopically, macroregenerative nodules could be classified as those with (type 2) and without (type 1) dysplasia. Four livers had type 1 lesions, two had type 2 lesions and five had lesions of both types. We found 36 type 1 lesions in all and 12 type 2 lesions, 3 containing foci of microscopic carcinoma. All hepatocellular carcinomas arose in livers containing macroregenerative nodules (either type). Liver cell dysplasia, large-cell or small-cell, was observed in cirrhotic nodules of 27 livers. Microscopic or macroscopic hepatocellular carcinoma occurred in three livers with large-cell but not small-cell dysplasia and in one liver without dysplasia.(ABSTRACT TRUNCATED AT 250 WORDS)


Evidence for normal nitric oxide-mediated vasodilator tone in conscious rats with cirrhosis. Because it has been hypothesized that the hyperkinetic circulation in portal hypertension is the result of increased synthesis of nitric oxide, we compared the hemodynamic effects of nitric oxide synthesis--specific agonist (L-arginine) and antagonist between normal and cirrhotic conscious rats. The dose-response curves showed that L-arginine significantly decreased arterial pressure and increased heart rate. These changes started at the 200 mg/kg dose and were similar in both groups of rats. In both groups of rats NG-monomethyl-L-arginine (25 mg/kg) significantly decreased cardiac output by 35%. In cirrhotic rats, NG-monomethyl-L-arginine decreased portal pressure from 15.3 +/- 0.9 mm Hg to 13.6 +/- 0.7 mm Hg and portal tributary blood flow from 7.8 +/- 0.7 ml.min-1.100 gm-1 to 5.9 +/- 0.7 ml.min-1.100 gm-1; it significantly increased portal territory vascular resistance from 950 +/- 108 dyn.sec.cm-5.100 gm-1 x 10(3) to 1,579 +/- 258 dyn.sec.cm-5.100 gm-1 x 10(3). In normal rats, portal tributary blood flow decreased similarly, by 27%, and portal territory vascular resistance increased by 55%. In neither group was hepatic arterial blood flow altered. Before and after NG-monomethyl-L-arginine administration, arterial cyclic GMP concentrations were not significantly different between normal and cirrhotic rats. In conclusion, this study shows evidence of a normal role for nitric oxide-mediated vasodilatation in rats with cirrhosis and that inhibition of nitric oxide synthesis reduces portal hypertension. These results did not support the hypothesis that nitric oxide synthesis is increased in cirrhosis.


Histological features and interphase nucleolar organizer regions in hyperplastic, dysplastic and neoplastic epithelium of intrahepatic bile ducts in hepatolithiasis. Neoplastic transformation occurs in the intrahepatic biliary tree in hepatolithiasis. The present study aimed to clarify the neoplastic processes by correlating the histological features of the bile duct lesions with counts of interphase argyrophilic nucleolar organizer regions (AgNORs), which reflect cell proliferative activity. We studied 55 cases of hepatolithiasis and 25 normal autopsy livers. The biliary epithelial lesions in hepatolithiasis were divisible into hyperplasia, dysplasia and neoplasia. These lesions were found in bile ducts containing calculi. All cases of hepatolithiasis showed a varied degree of hyperplasia. Additionally, eight cases showed dysplasia, five non-invasive intraductal adenocarcinoma and 10 invasive adenocarcinoma. Cases of non-invasive and invasive carcinoma frequently harboured areas of dysplasia, and areas of dysplasia and non-invasive carcinoma, respectively. The mean and standard deviation of the number of interphase AgNORs in the normal and abnormal biliary epithelium showed a step-wise increase in the following order: normal (1.32 +/- 0.36), hyperplasia (1.52 +/- 0.37), dysplasia (2.28 +/- 0.56), non-invasive carcinoma (3.23 +/- 1.00), and invasive carcinoma (3.72 +/- 0.77). These histological and cell kinetic observations suggest that, in hepatolithiasis, carcinogenesis in bile duct epithelial cells progresses in a multi-step manner, through hyperplasia, dysplasia, non-invasive adenocarcinoma and invasive adenocarcinoma.


The predictive value of human papilloma virus (HPV) typing in the prognosis of bronchial squamous cell papillomas. Five solitary squamous papillomas of bronchus with variable degrees of dysplasia, one combined with a laryngeal papilloma and with a neuroendocrine carcinoma in the contralateral lung, and five papillomas combined with invasive squamous cell carcinomas were investigated for their expression of human papilloma virus DNA by in situ hybridization. Benign squamous cell papillomas showed an association with papilloma virus type 11 and rarely type 6, whereas types 16 or 18, sometimes in combination with types 31/33/35 were found in papillomas associated with carcinomas. In one patient a benign papilloma containing human papilloma virus type 18 and 31/33/35-positive preceded a recurrence combined with carcinoma by 2 years; this recurrent papilloma and the carcinoma were also positive for human papilloma virus 18 DNA. We suggest that human papilloma virus typing should be performed in every squamous cell papilloma of bronchus. Patients with papillomas exhibiting human papilloma virus 16 or 18 positivity are at high risk for the development of squamous cell carcinoma. Furthermore, virus typing may be of prognostic importance in relation to the development of squamous carcinoma.


Clear cell sarcoma of kidney: a clinicopathological study of eight cases from Malaysia. Eight cases of clear cell sarcoma of kidney were seen in the Department of Pathology, University Hospital, Kuala Lumpur, Malaysia over the 16-year period from 1973 to 1989. Five of the patients were males. Six patients were Malay, one Chinese and one Indian. The patients' ages ranged from 8 months to 3 years. Clear cell sarcoma was the original diagnosis in two patients while six were diagnosed as blastemal-predominant Wilms' tumours at presentation. Metastases developed in five patients. Metastatic sites included the thoracic vertebra, skull, orbit, humerus, radius, ulna, shoulder, lung and liver. The prolonged survival, of 9 years and 9 months, seen in one patient despite omission of Adriamycin (doxorubicin) from the chemotherapeutic protocol is highlighted. We also emphasise the histological factors which are of help in differentiating clear cell sarcoma from Wilms' tumour.


Small cell (neuroendocrine) carcinoma of the vagina. We report a case of vaginal small cell (neuroendocrine) carcinoma. Immunostaining for neuron-specific enolase, PGP 9.5, chromogranin, synaptophysin, Leu 7 and cytokeratin was positive. Neurosecretory granules were found by electronmicroscopy. There was local recurrence and regional lymph node metastases. The patient survived for 10 months following local surgical therapy only.


Outcomes and costs of two VA inpatient treatment programs for older alcoholic patients. One hundred thirty-seven older alcoholic patients were randomly assigned to two different inpatient treatment programs at a Veterans Affairs medical center and followed for one year after discharge. The older alcoholic rehabilitation (OAR) program was operated by a tolerant staff that specialized in treating elderly alcoholics. Treatment included reminiscence therapy with goals of developing patient self-esteem and peer relationships. The traditional care program emphasized confrontation to focus on patients' past failures and present conflicts. Patient care costs were slightly lower (2.5 percent lower) in the OAR program than in the more traditional program, and OAR patients were 2.1 times more likely to report abstinence at one year. Response to the OAR program was best for patients over 60 years of age.


Age-related psychiatric comorbidities and level of functioning in alcoholic veterans seeking outpatient treatment. The relationship of age and of level of adaptive functioning to comorbidity of mental disorders among alcoholics was studied in a survey of all alcoholics seeking outpatient mental health treatment in the Veterans Affairs mental health care system during a one-month period in 1986 (N = 22,463). More than half of the alcoholic outpatients had one or more comorbid psychiatric diagnoses. Rates for comorbid substance abuse disorders, posttraumatic stress disorder, schizophrenia, and personality disorders peaked in younger alcoholics and then decreased with age. Age-related increases were observed for major depression, anxiety disorders, and organic brain syndrome or dementia. DSM-III axis V ratings of poor to grossly impaired functioning were consistent across age groups. More than half of alcoholics with a comorbid psychiatric disorder were rated as severely impaired, compared with less than a third of those with no comorbid mental disorder.


Alu insertion polymorphism: a new type of marker for human population studies. A PCR-based method was used to screen 462 individuals from Japan, Papua New Guinea, Indonesia, and Australia for an Alu family insertion polymorphism. The frequency of this Alu insertion shows significant heterogeneity among island subgroups of the Indonesian sample and between the Japanese-Indonesian populations and the Australian-New Guinean populations. The simple, rapid PCR-based screening technique and the significant frequency differences among populations demonstrate that Alu insertion polymorphisms are potentially valuable markers for studies of the evolutionary history and migration patterns of modern humans.


[Preliminary use of recombinant glycoprotein 52kd as an antigen in the diagnosis of human cytomegalovirus infection]. Interest in the human cytomegalovirus (HCMV) mainly derives from its associations with congenital malformations, mental retardation, and severe or fatal infections in immunosuppressed individuals such as transplant patients, tumor and AIDS patients. It is evidenced that there has been a need for a rapid and sensitive methods to detect an ongoing acute infection. The recent studies showed that high titers of antibody to the glycoprotein 52kd are present in sera of patients undergoing acute HCMV infection. However, purification of individual glycoprotein from HCMV-infected cells is a daunting prospect. HCMV glycoprotein 52 kd expressed via recombinant DNA techniques are a promising approach to solve this problem. In order to evaluate the diagnostic value of the recombinant glycoprotein 52 kd antigenic code region for HCMV infection, we have used the polymerase chain reaction (PCR) and recombinant DNA techniques to construct successfully the high-level expression plasmid pHCMV containing the HCMV GP-52 kd antigenic code region, with the predicted protein at levels up to 20% in total bacterial protein. The expressed protein was purified from SDS-PAGE, used as an antigen in Western-blot, and reacted with 12 cases of the positive sera, 4 cases of the negative sera, following by reaction with HRP-labelled horse IgG antibody against human. The results indicated that the approach we are using to detect antibody to HCMV acute infection are as sensitive as general serological methods such as ELISA, with the advantages of easy preparation of antigen with high quantity, and clinical practicability.


[The mechanism of inhibition of calmodulin-dependent cyclic nucleotide phosphodiesterase by dihydropyridine calcium antagonists]. Kinetic studies of the monoclonal antibody-purified 60 kd calmodulin dependent cyclic nucleotide phosphodiesterase (PDE) from bovine brain indicate that the dihydropyridine calcium antagonists, nicardipine and felodipine are partially competitive inhibitors of the enzyme. This was supported by the facts that the inhibition approached a finite level as the concentrations of drug reached saturation and the inhibition showed non-linearity characteristics in the Dixon plot. Furthermore, at the constant concentration of felodipine which brings about close to maximal inhibition, increasing concentrations of nicardipine can alleviate enzyme inhibition to approach the level of maximal nicardipine inhibition, suggesting that the calmodulin-dependent PDE may contain specific dihydropyridine binding site distinct from the active site.


[Experimental study of use of shuttle vector to develop a system of detecting mutagenesis in mammalian cells]. The paper presents a series of experimental studies of the use of shuttle vector to develop a short-time mutagenesis system of detecting gene mutation in the mammalian cell at DNA level. The system is based on the finding that in the EB virus the shuttle vector pMCi5 carries LacI gene. The LacI is used as a target of induced mutagenesis in mouse cells. Use pMCi5 plasmid to transform the 3T3 mouse cell, expose it to a chemical mutagen, ethyl methanesulfonate (EMS); transfer the treated plasmid DNA to E. coli strain MC1061F' 150Kan; and then plate the transformation on a special culture medium containing X-gal for rapid detection and analysis of LacI mutation. The spontaneous mutant frequency of the system is less than 1.21 x 10(-4). After the treatment with 300 micrograms/ml EMS, the induced mutant frequency increases to 3.8 x 10(-3). No gross alteration has been discovered in seven LacI mutants at DNA level. No point mutation has occurred in the EcoRI site by analysis with restriction enzyme EcoRI.


[Antisteatotic effects of four kinds of dietary fibers in rats fed on high cholesterol diet: a preliminary morphometric analysis]. The hypolipidaemic and antisteatotic effects of the Konnyaku Powder (KP) have been reported before. In order to evaluate further the antisteatotic role played by KP, the effects of KP on the levels of liver lipid and on the hepatic histopathology and morphometry in comparison with those of pectin, algin and agar were studied. Sixty Wistar strain rats were divided into 6 groups: a normal diet group, a high cholesterol diet group (HC), and 4 test groups, in which the animals were fed on a diet similar to that of the high cholesterol diet group with addition of KP, pectin, algin or agar at a dosage of 5%, respectively. All the animals were killed at the end of the diet treatment for 9 weeks. The results showed that relative liver weights were lower in four experimental groups than in the HC group. The levels of total and free cholesterol, and triglyceride in the liver were lower in KP group than in HC group and in the other three experimental groups. Hepatic histopathology and morphometric examination indicated that antisteatotic effects of KP appeared to be much more significant than those of the other fibers.


[Effects of aluminum, fluorine and their mixtures on the enzyme activities of jejunal mucus membrane in the perfusion of small intestine of rats in vivo]. Buffer (control), F as NaF (100 mmol/L), Al as AlCl3 (32 and 170 mmol/L) and the two mixtures of Al and F (Al:F = 0.3:0.7 and 0.7:0.3) in Tris-buffered media of pH 7.0 were respectively perfused through the small intestine of rats over a period of 40 min. The jejunum was taken out from each animal immediately. Some of its parts were for HE staining; and the others for enzyme histochemical staining. Fifteen enzymes were observed. The results showed that (1) Fluorine chiefly inhibited the metal-enzymes and metal-activated enzymes; aluminum mainly depressed the activities of enzymes in relation to the cell energy metabolism, such as hydrogenases and oxidase. (2) The decreased activities of enzymes induced by Al or F increased significantly after mixing with each other, indicating that the simultaneous existence of Al and F decreased their ionic concentrations, so that strong antagonistic effects on the inhibition of enzymes were noticed since F or Al inhibited enzyme activities by their ions. (3) A high dose of Al intensively inhibited the enzymes related to the cell energy metabolism, which explained why less quantity of Al was absorbed in animals with high dose of Al than that of low dose Al seen in the absorption experiment. (4) The enzyme activities in the animals given high dose of Al mixed with F apparently were elevated in comparison with that in the animals given a pure high dose of Al which was the result of antagonistic effects of F on the inhibition of enzyme activities by Al, therefore the quantity of the absorbed Al was high.


Primitive neuroectodermal tumors of the uterus: a report of four cases. Four cases of primitive neuroectodermal tumor (PNET) of the uterine corpus are reported, bringing the total number of reported PNETs in this site to seven. The four women were in their seventh decade of life and presented with abnormal vaginal bleeding and, in two cases, an enlarged uterus. The patients underwent total or subtotal abdominal hysterectomy and bilateral salpingo-oophorectomy and, in one patient, pelvic lymphadenectomy. Three patients received postoperative radiation therapy, chemotherapy, or both. Gross examination revealed fleshy polypoid masses filling the endometrial cavity and, in two cases, deeply invading the myometrium. Histologic, immunohistochemical, and, in two cases, ultrastructural examination revealed typical PNETs that exhibited variable degrees of neural, glial, ependymal, and medulloepithelial differentiation. Two PNETs were admixed with other neoplasms: in one case a grade I endometrial adenocarcinoma and in the other a low-grade endometrial stromal sarcoma. The prognosis of the tumors was related to their stage: two patients with stage I tumor were alive with no evidence of disease at 5 and 6 years, whereas two patients with stage III or IV tumor died of tumor at 6 and 12 months. Although it has been suggested that uterine PNETs may be derived from displaced germ cells or implanted fetal tissues, evidence provided by this study, including the advanced ages of the patients and an admixture with neoplasms of unquestioned müllerian origin, suggests a müllerian origin for these tumors in at least some cases.


Prevalence of genital human papillomavirus infection in Wellington women. To determine prevalence of human papillomavirus (HPV) in Wellington women, to identify risk factors for HPV infection, to correlate presence of HPV with cervical cytology, and to identify characteristics of women infected with HPV but with normal cytology. Demographic, social, personal and clinical data were collected by a confidential self-administered coded questionnaire. The presence of DNA from HPV types 6/11, 16 + 18 and 31 + 33 in cervical scrapes was determined by dot-blot DNA hybridisation. All data were correlated with cervical cytology results. Two thousand and twenty one women attending family planning clinics in the Wellington region participated in the study. The mean age of participants was 26 years, 33.3% currently smoked, 72.3% used hormonal contraceptives, 31.4% were married, and 91.4% were of European origin. We found 10.9% of the study group infected with HPV. HPV types 16 and/or 18 predominated, being detected in 71.5% of HPV-positive women either alone or with other types. Of those infected 26.2% had multiple infections. Dysplasia (n = 87) or atypia (n = 84) were observed in 26.7% of infected women (n = 221) and 6.25% of uninfected women (n = 1792). Over 8% of women with normal smears were HPV positive, and types 16/18 were most common in these women. Women with cervical dysplasia or atypia were six times more likely to have HPV infection than other women. The main risk factor for HPV infection, particularly with types 16 and/or 18, was multiple (> 5) sexual partners in the last year independent of other variables. Multivariate analysis of data showed no independent association between HPV infection and ethnicity, educational background, smoking history, marital status, contraceptive use, age at first sexual intercourse, or number of lifetime sexual partners.


Pefloxacin and ciprofloxacin in the treatment of uncomplicated gonococcal urethritis in males [corrected]. To study the effectiveness of single-dose pefloxacin and ciprofloxacin in the treatment of uncomplicated gonococcal urethritis in males. Department of STD Control, Kelantan Road, Singapore. 160 male patients with uncomplicated gonococcal urethritis were assigned alternately to receive single oral doses of either pefloxacin 800 mg or ciprofloxacin 250 mg. Of the pefloxacin group 98.5% (65/66 patients) and of the ciprofloxacin group 98.6% (74/75 patients) were cured of gonorrhoea. The rates of post-gonococcal urethritis were 64.3% and 67.3% in the pefloxacin and ciprofloxacin groups, respectively. Both drugs were well tolerated and reported side-effects were minor and transient. There was a high incidence of penicillinase-producing gonococci (32.3%) and tetracycline resistant isolates with MIC > or = 2 mg/l (99.3%). High level tetracycline resistance (MIC > or = 16 mg/l) was found in 7.4% of isolates. The drugs in the dosages studied may be recommended for first-line treatment of uncomplicated gonococcal urethritis in males in Singapore. However, the emergence of bacterial resistance to the fluoroquinolones in the literature calls for vigilance in the monitoring of antimicrobial susceptibility [corrected].


Macrophages in host defence--an overview. The importance of macrophages in host defence is well documented. They are distributed in various tissues where they perform functions in normal steady state as well as in diseased condition. Macrophages secrete a number of enzymes, plasma proteins, complement and coagulation factors which regulate the effector functions of the macrophages. Exposure of macrophages to pathogens results in further metabolic changes which activate the former to secrete oxygen metabolites leading to their augmented microbicidal activity. Macrophages respond to the external stimuli by expressing a large repertoire of surface receptors which play an important role in the activation, recognition and endocytosis of foreign microorganisms. A large number of intracellular pathogens are harboured in the macrophages which can reside and replicate in them. A variety of strategem has been employed to target drugs to vacuolar apparatus of the macrophages in order to combat intracellular pathogens. This review covers some of these aspects particularly in relation to hose defence and methods by which therapeutic agents could be specifically delivered to macrophages.


Functions of cytochrome c in regulation of electron transfer and protein folding. Cytochrome c, a "mobile electron carrier" of the mitochondrial respiratory chain, also occurs in detectable amounts in the cytosol, and can receive electrons from cytochromes present in endoplasmic reticulum and plasma membranes as well as from superoxide and ascorbate. The pigment was found to dissociate from mitochondrial membranes in liver and kidney when rats were subjected to heat exposure and starvation, respectively. Treating cytochrome c with hydroxylamine gives a partially deaminated product with altered redox properties; decreased stimulation of respiration by deficient mitochondria, increased reduction by superoxide, and complete loss of reducibility by plasma membranes. Mitochondria isolated from brown adipose tissue of cold-exposed rats are found to be sub-saturated with cytochrome c. The ability of cytochrome c to reactivate reduced ribonuclease is now reinterpreted as a molecular chaperone role for the hemoprotein.


Mixed tumour of salivary gland type of the male breast. Benign breast tumours with a mixed cartilaginous and epithelial component are distinctly rare as evident from the literature. A case of Mixed Tumour of the breast presenting pre-operatively as a hard mass in a 65 year old male is reported. Histologically, it was composed of a mixture of benign cartilage, myoepithelial cells, tubules and a myxoid stroma in fat. A brief review of cartilage bearing lesions and mixed tumour in the mammary region is discussed.


In vitro effects of interleukin-4 on interferon-gamma-induced macrophage activation. Interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) have been shown to be secreted by distinct T-helper cell subsets which have different roles in the determination of host resistance to infection. We studied the activity of these two cytokines on effector mechanisms of mouse macrophages. In vitro cultured bone marrow-derived macrophages from C57BL/6 mice were treated with IFN-gamma, IL-4, or a combination of both cytokines and the ability to secrete superoxide or nitrite or to restrict growth of Mycobacterium avium and Toxoplasma gondii was then evaluated. We found that IL-4 could inhibit the priming of macrophages for enhanced superoxide production induced by IFN-gamma although IL-4 when used alone did have some enhancing effect of its own. This effect of IL-4 on IFN-gamma-primed superoxide production was dose dependent and could be observed even if the treatment by IL-4 was done 24 hr after treatment by IFN-gamma. IL-4 did not, however, influence the enhanced production of nitrogen reactive intermediates, the induction of bacteriostatic activity against M. avium, or the restriction of T. gondii by IFN-gamma-treated macrophages, and did not have any effect of its own regarding these latter functions.