Laboratory evaluation of a filter for the control of cross-infection during pulmonary function testing.

Nosocomial transmission of respiratory pathogens is a possible complication of lung function testing. The use of a filter, placed between the patient and the spirometer equipment may be one way of preventing such nosocomial spread. This paper reports the laboratory evaluation of the efficiency of such a filter at removing bacteria from exhaled breath. Volunteers exhaled 100 normal breaths or four forced exhalations through a filter on to a blood agar plate. Bacterial counts on the agar plate were compared with recoverable bacterial counts from the filter. Total challenge from the forced exhalations ranged from 161-84,200 colony forming units. The calculated efficiency of 10 volunteers was 99.9%. For the normal breaths the challenge ranged from 0-262,000 colony forming units. There was no growth on any of the blood-agar plates. These filters appear to be highly efficient at removing exhaled bacteria.


Epidemiology of Pseudomonas aeruginosa in an intensive care unit using selective decontamination of the digestive tract.

Selective decontamination of the digestive tract (SDD) aims to reduce the rate of nosocomial infections in critical care patients. Pseudomonas spp. are common nosocomial pathogens and in this study isolates collected from patients and the environment during an SDD trial were examined. The study enrolled 161 SDD cases and 170 controls. Pseudomonads were isolated from 27% of SDD patients and 30% of controls. SDD partially suppressed colonization in the 'gastro-respiratory' mucosae but not in the rectum. A total of 108 isolates of pseudomonads were recovered from the environment. Resistance in rectal isolates was minimal but isolates from 'gastro-respiratory' sites showed increasing aminoglycoside resistance. Eighty-six per cent of aminoglycoside-resistant isolates from both patient groups and environment were pyocine type 1x. Episodes of infection were reduced in the SDD patients (6) compared with the controls (16), aminoglycoside-resistant strains being associated with zero episodes in SDD patients but with five in the control group.


The housefly (Musca domestica) as a carrier of pathogenic microorganisms in a hospital environment.

Houseflies have long been regarded as potential carriers of microorganisms. Since pathogenic microorganisms are widespread in the hospital environment, there is abundant opportunity for flies to become contaminated and, in turn, to contaminate the patient environment. In the present study, an attempt was made to isolate and identify pathogenic bacteria, fungi and parasites from the housefly Musca domestica collected in the surgical ward of the All India Institute of Medical Sciences Hospital and also in a remote residential area located 5 km from the hospital. A total of 113 flies were collected: 65 from a surgical ward (test) and 48 from a residential area for comparison. Ten genera of bacteria were isolated from the test group of flies compared with nine from the control group. In primary isolations, it was observed that the load of bacteria carried by the test group of flies was significantly more (P less than 0.001) than for the control flies. Pseudomonas aeruginosa, Enterococcus faecalis and viridans streptococci were isolated only from the test flies. The isolation rate of Staphylococcus aureus was significantly higher (P less than 0.001) in test houseflies than in the control houseflies. There was no significant difference in isolation of parasitic ova and cysts from test and control houseflies. Candida spp. were isolated in almost equal numbers from both groups of houseflies, yet none of these was Candida albicans. Houseflies therefore may act as vectors of potentially pathogenic bacteria in a hospital environment.


The use of the Body Mass Index in studies of abdominal wound infection.

Abdominal wound infection rates were significantly related to obesity, as measured by Body Mass Index, in two large clinical trials of prophylactic antibiotics in at-risk abdominal surgery. Obesity is a risk factor that should be taken into account in any study in which wound infection is an outcome measure.


A novel methodology for the establishment of neuroblastoma cell lines from metastatic marrow. Expression of surface markers, neurofilaments, MDR-1 and myc proteins.

A methodology for rapid isolation of neuroblastoma (NB) cells from marrow with metastatic NB cells was developed using a cocktail of five antibodies and magnetic microspheres coated with secondary antibodies. Cells bound to microspheres were released by brief exposure to chymopapain, followed by repeated culture of released cells in serum supplemented DMEM medium and selection for adherent cells. Using this methodology, over 35 primary cell lines were obtained free of contaminating normal cells. Detailed analyses of over 14 cell lines revealed gross differences in cell phenotype, size, morphology development of neurite processes, and doubling time (40h-80 h). All cell lines expressed the 145 kDa neurofilament (NF) and a few expressed the 200 kDa NF, with very little or no expression of the 68 kDa NF. DNA analyses revealed 80% near diploid cell lines. High expression of the MDR-1 protein was detected in 6/22 of the cell lines tested.


Simultaneous assay for oxidative metabolism and adhesion of human neutrophils: evidence for correlations and dissociations of the two responses.

An assay method for the simultaneous evaluation of the oxidative metabolism and adherence of human neutrophils is described, together with certain specific applications. Incubations were performed in serum-coated microtiter plates, where oxidative metabolism was measured as O2- release and, after washing out the nonadherent cells, the adhesion was measured as activity of acid phosphatase. Three agonists tested in this system--opsonized zymosan, concanavalin A, and N-formyl-methionyl-leucyl-phenylalanine--induced both activation of O2- release and cell adhesion, but the two functions had time course and dose dependence patterns that varied depending on the stimulant. Particularly with concanavalin A, O2- release and adhesion response were markedly dissociated; this lectin at low doses increased neutrophil adherence without triggering any O2- production, whereas at high doses it increased both O2- production and adherence. Anti-integrin monoclonal antibodies did not affect adhesion induced by low-dose concanavalin A but inhibited the adhesion induced by the other tested agonists. Adhesion and O2- production were also found to be differentially affected by the NADPH oxidase inhibitor diphenylene iodonium, the sulfhydryl reagent N-ethylmaleimide and the A2 agonist adenosine, indicating that these neutrophil responses have various transductional pathways that also depend on the type of stimulus.


Monocyte adherence to fibronectin: role of CD11/CD18 integrins and relationship to other monocyte functions.

Adherence of monocytes to extracellular matrix components is critical for their accumulation at sites of infection. To gain insight into the factors that regulate monocyte recruitment, we have studied monocyte adherence with regard to the regulatory effects of bacterial lipopolysaccharide (LPS) and the mechanisms involved; moreover, we have contrasted the phenotypes of adherent and nonadherent cells. Our results show that only a minor subpopulation of monocytes (20-25%) adhere spontaneously to fibronectin and that LPS stimulated a threefold increase in the proportion of adherent cells. Basal adherence and LPS-stimulated adherence of monocytes to fibronectin were substantially mediated by CD11/CD18 integrins. Further studies revealed that spontaneously adherent monocytes were 14-fold more actively phagocytic, released 1.6-fold more superoxide anion, and contained 20-fold more peroxidase activity than nonadherent cells, whereas LPS-adherent cells had an intermediate phenotype. These results indicate that LPS may enhance the accumulation of monocytes with an antimicrobial phenotype and thereby promote resolution of tissue infection.


The glnA gene of the extremely thermophilic eubacterium Thermotoga maritima: cloning, primary structure, and expression in Escherichia coli.

The structural gene (glnA) encoding the glutamine synthetase (GS) of the extremely thermophilic eubacterium Thermotoga maritima has been cloned on a 6.0 kb HindIII DNA fragment. Sequencing of the region containing the glnA gene (1444 bp) showed an ORF encoding a polypeptide (439 residues) with an estimated mass of 50,088 Da, which shared significant homology with the GSI sequences of other Bacteria (Escherichia coli, Bacillus subtilis) and Archaea (Pyrococcus woesei, Sulfolobus solfataricus). The T. maritima glnA gene was expressed in E. coli, as shown by the ability to complement a glnA lesion in the glutamine-auxotrophic strain ET8051. The recombinant GS has been partially characterized with respect to the temperature dependence of enzyme activity, molecular mass and mode of regulation. The molecular mass of the Thermotoga GS (590,000 Da), estimated by gel filtration, was compatible with a dodecameric composition for the holoenzyme, as expected for a glutamine synthetase of the GSI type. Comparison of the amino acid sequence of T. maritima GS with those from thermophilic and mesophilic micro-organisms failed to detect any obvious features directly related to thermal stability.


Application of screening and confirmatory assays for anti-HTLV-I/II in U.K. populations.

During an epidemiological study of a low risk U.K. population diverse screening and confirmatory assays for the detection of anti-HTLV-I/II were assessed. Sera from 2,900 antenatal patients were tested for anti-HTLV-I/II by gelatin particle agglutination assay (GPA). All reactive sera, and 133 randomly selected unreactive sera, were further tested by Abbott and DuPont ELISAs, "in house" competitive and IgG capture radioimmunoassays (RIAs), and Western blot (WB). Sera which reacted with any HTLV-I proteins by WB were tested by radio immunoprecipitation assays (RIPA). The two ELISAs detected all the GPA reactive specimens that were subsequently confirmed as anti-HTLV-I/II positive. Confirmation of positive screening results required the use of both WB and RIPA. Serological diagnosis of HTLV infection involves access to specialised assays that are not commercially available.


Gerstmann-Sträussler-Scheinker disease in an Alsatian family: clinical and genetic studies.

The clinical progression of Gerstmann-Sträussler-Scheinker disease in a family of Alsatian origin is reported. The age of onset and the duration of evolution were variable. The clinical picture became more complex over the generations: in the first generations, isolated dementia and in later generations a triad of pyramidal, pseudobulbar syndromes and dementia associated with spinal cord and cerebellar features. Prion gene analysis showed that four surviving patients carry double missense changes at codons 117 and 129, identical to those found in one case at necropsy and 10 other healthy members of the family. The missense changes were not found in 100 controls. No member of the family had modification of condons 102, 178, or 200. The lod score suggests linkage between the missense change at codon 117 and Gerstmann-Sträussler-Scheinker disease in this family.


Laparoscopy for the impalpable testes: experience with 53 testes.

Impalpable testes constitute approximately 20% of most series of undescended testes. From January 1986 to March 1991, we performed laparoscopies on 53 patients with impalpable testes. Thirty-two of them were found to have normal vasa and vessels entering each internal ring on the side in question. Of these, 14 were found to have "vanishing testes" at exploration, 12 others underwent successful orchiopexy, and the remaining 6 had excisional biopsies of fibrotic testicular remnants. Five patients had no visible vessels and a sixth had a blind-ending vas and vessels adjacent to the internal ring; in these cases no further investigations were deemed necessary. Fifteen patients were found to have abdominal testes and underwent high testicular vessel ligation and division at the time of the laparoscopy; 14 of them have undergone staged orchiopexy 6 months after laparoscopy and one is scheduled for this procedure. A 3-month follow-up of those who had orchiopexy showed excellent results in 10 patients and poor results in 3, all of whom had small testes that were unimproved or worse following vessel ligation. Four boys were spared operations as a result of findings at laparoscopy. Early in the series there was one failed laparoscopy, but it was successfully completed later. the procedure, but it was successfully treated with antibiotics. There were no other complications. Laparoscopy is a safe procedure that allows accurate diagnosis and may prevent additional intervention in the treatment of the absent testes. It facilitates the locating of the impalpable testis and the planning and timing of subsequent orchiopexy. We believe that laparoscopy is the preferred procedure in the management of impalpable testes.


Short-acting beta-adrenergic blockade and the fetus. A case report.

An infant was born to a woman who received intravenous esmolol for intrapartum supraventricular tachyarrhythmia. Despite the very short acting and cardio-selective beta-1 adrenergic blockade induced by that agent, neonatal effects can occur up to 48 hours after delivery.


Prevalence of substance use among US physicians.

To estimate the prevalence of substance use among US physicians. A mailed, anonymous, self-report survey that assessed use of 13 substances and permitted comparison with results of the National Household Survey on Drug Abuse. Rates of physician substance use were weighted to provide national prevalence estimates. A national sample of 9600 physicians, stratified by specialty and career stage, and randomly selected from the American Medical Association master file. The response rate after three mailings was 59%. Demographic characteristics of respondents closely reflected those of the US physician population. Subjects' self-reported use of 13 substances in their lifetime, the past year, and the past month; reasons for use; self-admitted substance abuse or dependence; and whether treatment was received. For controlled prescription substances, respondents were asked to report only use "not prescribed by another physician for a legitimate medical or psychiatric condition." Physicians were less likely to have used cigarettes and illicit substances, such as marijuana, cocaine, and heroin, in the past year than their age and gender counterparts in the National Household Survey on Drug Abuse. They were more likely to have used alcohol and two types of prescription medications--minor opiates and benzodiazepine tranquilizers. Prescription substances were used primarily for self-treatment, whereas illicit substances and alcohol were used primarily for recreation. Current daily use of illicit or controlled substances was rare. Although physicians were as likely to have experimented with illicit substances in their lifetime as their age and gender peers in society, they were far less likely to be current users of illicit substances. The higher prevalence of alcohol use among respondents may be more a characteristic of their socioeconomic class than of their profession. A unique concern for physicians, however, is their high rate of self-treatment with controlled medications--a practice that could increase their risk of drug abuse or dependence. Uniform national guidelines are needed to sensitize medical students and physicians to the dangers of self-treatment with controlled prescription substances.


Urinary aflatoxin biomarkers and risk of hepatocellular carcinoma.

Aflatoxins have long been suspected to be human hepatic carcinogens but no direct study was feasible until assays to measure individual aflatoxin exposure became available. We have used assays for urinary aflatoxin B1, its metabolites AFP1 and AFM1, and DNA-adducts (AFB1-N7-Gua) to assess the relation between aflatoxin exposure and liver cancer, as part of an ongoing prospective study of 18,244 middle-aged men in Shanghai, People's Republic of China. After 35,299 person-years of follow-up, 22 cases of liver cancer had been identified. For each case, 5 or 10 controls were randomly selected from cohort members without liver cancer on the date the disorder was diagnosed in the case and matched to within 1 year for age, within 1 month for sample collection, and for neighbourhood of residence. Subjects with liver cancer were more likely than were controls to have detectable concentrations of any of the aflatoxin metabolites (relative risk 2.4, 95% confidence interval 1.0-5.9). The highest relative risk was for aflatoxin P1 (6.2, 1.8-21.5). In an analysis adjusting for the effects of hepatitis B surface antigen seropositivity, level of education, cigarette smoking, and alcohol consumption, the relative risk for the presence of aflatoxin metabolites was 3.8 (1.2-12.2). There was a strong interaction between serological markers of chronic hepatitis B infection and aflatoxin exposure in liver-cancer risk. Reduction of aflatoxin exposure may be a useful intermediate goal in prevention of liver cancer, since the benefits of wide-scale hepatitis B vaccination will not be apparent for many years.


Prophylactic ciprofloxacin for catheter-associated urinary-tract infection.

Patients receiving antibiotics during bladder drainage have a lower incidence of urinary-tract infections compared with similar patients not on antibiotics. However, antibiotic prophylaxis in patients with a urinary catheter is opposed because of the fear of inducing resistant bacterial strains. We have done a double-blind, placebo-controlled trial of prophylactic ciprofloxacin in selected groups of surgical patients who had postoperative bladder drainage scheduled to last for 3 to 14 days. Patients were randomly assigned to receive placebo (n = 61), 250 mg ciprofloxacin per day (n = 59), or 500 mg ciprofloxacin twice daily (n = 64) from postoperative day 2 until catheter removal. 75% of placebo patients were bacteriuric at catheter removal compared with 16% of ciprofloxacin-treated patients (relative risk [RR] [95% CI] 4.7 [3.0-7.4]). The prevalence of pyuria among placebo patients increased from 11% to 42% while the catheter was in place; by contrast, the rate of pyuria was 11% or less in patients receiving ciprofloxacin (RR 4.0 [2.1-7.3]). 20% of placebo patients had symptomatic urinary-tract infections, including 3 with septicaemia, compared with 5% of the ciprofloxacin groups (RR 4.0 [1.6-10.2]). Bacteria isolated from urines of placebo patients at catheter removal were mostly species of enterobacteriaceae (37%), staphylococci (26%), and Enterococcus faecalis (20%), whereas species isolated from urines of ciprofloxacin patients were virtually all gram-positive. Ciprofloxacin-resistant mutants of normally sensitive gram-negative bacteria were not observed. Ciprofloxacin prophylaxis is effective and safe in the prevention of catheter-associated urinary tract infection and related morbidity in selected groups of patients requiring 3 to 14 days of bladder drainage.


Hepatitis B virus and apparent fulminant non-A, non-B hepatitis.

While there is evidence that hepatitis C virus (HCV) does not cause fulminant non-A, non-B hepatitis, the causal agent remains unknown. To evaluate the role of hepatitis B virus (HBV) in this disease, we used a two-step polymerase chain reaction (PCR) to amplify the surface and core regions of HBV DNA in serum and liver samples taken prospectively from twenty-six patients (mean age 36 years, range 1 to 64) with acute hepatic failure undergoing liver transplantation. HBV DNA was absent from the serum of all patients before transplantation. Seventeen patients were diagnosed as having non-A, non-B hepatitis because they lacked serological evidence of hepatitis A virus or HBV infection. Liver samples were taken from twelve of these patients, and six samples were positive for HBV DNA. By contrast HBV DNA was not detected in liver from three patients with acute liver failure caused by hepatitis A or toxins. HCV RNA was not found in pretransplant samples by PCR. Four of the six patients with detectable HBV DNA in liver and presumptive non-A, non-B hepatitis had detectable HBV DNA in serum after transplantation. One additional patient who did not donate pretransplant liver had HBV DNA in a post-transplant serum sample. Thus, HBV DNA was present before or after transplantation in seven of seventeen patients with apparent non-A, non-B hepatitis. Three of five patients with detectable post-transplant serum HBV DNA were serologically positive for HBV surface antigen. These findings indicate that HBV may be a common cause of fulminant hepatic failure in patients lacking serological evidence of HBV infection.


Subcurative chemotherapy and fatal post-treatment reactive encephalopathies in African trypanosomiasis.

The treatment of late-stage African sleeping sickness in man is often complicated by a post-treatment reactive encephalopathy. The bases of this pathological reaction was investigated in a mouse model of African trypanosomiasis. Subcurative treatment with diminazene aceturate, which did not clear parasites from the central nervous system, resulted in a post-treatment meningoencephalitis similar to that seen in man. By contrast, a curative regimen of melaminylthioarsenite and 5-nitroimidazole, which cleared parasites from the central nervous system, did not cause any pathological reaction in the mice. This result indicates that subcurative treatment leads to the development of the post-treatment encephalopathy. Evidence that this may also be the case in man was provided by the detection of trypanosome DNA with the polymerase chain reaction in the brains of 9 patients who had died as the result of a post-treatment reaction. Our findings suggest that more aggressive treatment regimens, which ensure the elimination of trypanosomes from the central nervous system, may prevent post-treatment reactions in patients.


Human papillomavirus type 16 in cervical smears as predictor of high-grade cervical intraepithelial neoplasia [corrected].

The management of women with mild to moderately dyskaryotic cervical smears would benefit from a non-invasive test that predicts which women have high-grade cervical intraepithelial neoplasia. Detection of human papillomavirus type 16 (HPV16) DNA in cervical smears may be such a test. With the polymerase chain reaction (PCR), we estimated the amount of HPV16 DNA in cervical smears from 85 women referred for colposcopy because of abnormal cytology. An intermediate or high amount of HPV16 DNA predicted the presence of high-grade cervical intraepithelial neoplasia in a subsequent biopsy in almost 90% of patients irrespective of the cytological grade of the referral smear. This technique may allow early identification of those women with low-grade cytological abnormalities who have high-grade underlying cervical disease.


Controlled trial of endoscopic injection treatment for bleeding from peptic ulcers with visible vessels.

Controlled trials have shown that bleeding peptic ulcers can be successfully treated by endoscopy and injection of adrenaline, with or without sclerosant. However, these trials have been done in major research centres, and endoscopic treatment of upper gastrointestinal bleeding has not yet become routine in general hospitals. We have done a prospective, randomised, controlled trial of injection treatment for bleeding peptic ulcers in a district general hospital. Between April, 1989, and June, 1991, all patients with acute upper gastrointestinal bleeding (n = 555) underwent endoscopy by an experienced endoscopist within 24 h of admission. 98 patients were found to have an ulcer with a visible vessel, of whom 93 were randomised to injection (n = 48) or standard treatment alone (n = 45). Injection treatment consisted of 1-2 ml of 1 in 10,000 adrenaline injected at four to six sites around the ulcer. Adrenaline and 5% ethanolamine oleate (1-2 ml) were then injected directly into the vessel. The medical team managing the patient was unaware of the endoscopic treatment given. The two groups were similar for age, initial haemoglobin concentration, shock, and ulcer site. Rebleeding (injected 8 [16.7%] vs control 21 [46.7%], p = 0.011) was significantly reduced in treated patients. The treated group also had lower mortality (4 [8.3%] vs 9 [20%]), requirement for surgery (4 [8.3%] vs 8 [17.8%]), and mean blood-transfusion requirement (5 vs 7.5 units). Endoscopic injection treatment in our patients significantly reduced rebleeding rate and may have other benefits. This cheap and widely applicable treatment can be used routinely in the management of patients with bleeding peptic ulcers who are at high risk of rebleeding.


Clinical study of the lactational amenorrhoea method for family planning.

The effect of breastfeeding on fertility is well known; however, its use as a method of family planning was, until recently, untested. In 1988, the Bellagio Consensus Conference proposed guidelines that became the basis for a method of family planning called the lactational amenorrhoea method (LAM). The principle of LAM is that a woman who continues to fully or nearly fully breastfeed her infant and who remains amenorrhoeic during the first 6 months postpartum is protected from pregnancy during that time. We have assessed this method in the context of a breastfeeding support intervention study of 422 middle-class women in urban Santiago, Chile. The cumulative 6-month life-table pregnancy rate was 0.45% among women who relied on LAM as their only family planning method (1 woman pregnant in month 6). The findings indicate that LAM, with its high acceptance and efficacy, is a viable method of family planning and can safely serve as an introductory method for breastfeeding women.


Cutaneous malignant melanoma, Scotland, 1979-89. The Scottish Melanoma Group.

The Scottish Melanoma Group (SMG) was established in 1979 to assess mortality from and incidence, features, pathological data, and management of cutaneous malignant melanoma in Scotland. Incidence during the first five years and five-year survival have already been reported. We now have data about incidence and mortality over eleven years in relation to anatomical site and pathological types. From 1979 to 1989, 1354 male and 2459 female patients with primary cutaneous malignant melanomas were first diagnosed in Scottish residents. The incidence rate per 100,000 population per year has increased from 3.4 in 1979 to 7.1 in 1989 for men, and from 6.6 to 10.4 for women. The overall increase over eleven years is 82% (7.4% per year). The greatest rates of increase are seen in lesions of the superficial spreading histogenetic type, arising on the female leg and the male trunk. Following public education programmes started in 1985, the proportion of all melanomas less than 1.5 mm thick has shown a sustained and significant increase. Mortality data for 1661 patients for whom a minimum of five-year follow-up is available shows five-year survival of 71.6% overall (77.6% for women, 58.7% for men). The survival advantage for women persists when appropriate statistical adjustment is made for thickness, ulceration, and histogenetic type. These data are useful in designing public education programmes aimed at both primary and secondary prevention of melanoma and in auditing changes in trends that might result from such education.


Embryonal germ-layer antigens: target for autoimmunity.

Histopathological analysis of some systemic autoimmune diseases and syndromes led us to the conclusion that the common feature of the organs involved might be their embryonal origin. We suggest that organs derived from the same germ layer express common germ-layer-specific antigens. Such antigens could serve as target antigens for the autoimmune response.


Retroperitoneal germ cell neoplasm: MR and CT.

A case of germ cell neoplasm in an undescended retroperitoneal testicle is reported. CT revealed a large mass most consistent with a chronic hematoma. MRI demonstrated findings typical for neoplasm, and this was confirmed on biopsy.


Risk factors for hypertension in Kimberley aborigines.

To determine physical, biochemical and lifestyle factors associated with high blood pressure among Aborigines in the Kimberley region. Blood pressure and electrocardiographic (ECG) abnormalities in an age and sex stratified random sample of the Aboriginal population were related to other observations and measurements made at the same time specifically for the purpose of these comparisons. A field study in which subjects were interviewed and measurements made mostly in community clinics. All 249 men and 241 women from the prevalence study were included although only complete data sets for the various comparisons were analysed. A sample of venous blood was obtained in addition to physical measurements and information at interview. Statistical analysis of the relationships between blood pressure or hypertension and alcohol consumption, plasma gamma-glutamyl transpeptidase (GGT) activity, use of tobacco, body mass index (BMI, kg/m2) and non-fasted plasma cholesterol level. Hypertension was defined as systolic blood pressure of 160 mmHg or greater or diastolic blood pressure of 95 mmHg or greater. High blood pressure in Aboriginal men below 30 years was associated both with current drinking status and with circulating GGT level. There was a positive association of diastolic hypertension with consumption of alcohol in middle aged men (30 to 49 years) and in older women. Drinking was highly prevalent among men, especially below 30 years, but was less prevalent among women. Both systolic and diastolic blood pressure were positively related to BMI across the population but obesity (BMI greater than or equal to 30 kg/m2) was highly prevalent only among middle-aged women. Both systolic and diastolic blood pressure were positively and strongly related to plasma cholesterol level independently of the latter's relationship to age and BMI. The high prevalence of drinking among Aboriginal men and of obesity among Aboriginal women involves a risk of hypertension. The association between plasma cholesterol and blood pressure in Aboriginal men and women may be relevant to the demonstrated link between systolic hypertension and ischaemic heart disease.


Recommendations for prophylaxis against Pneumocystis carinii pneumonia for adults and adolescents infected with human immunodeficiency virus.

In 1989, the United States Public Health Service convened a Task Force of experts to consider the expanding knowledge base about prevention of Pneumocystis carinii pneumonia (PCP) among adults and adolescents (greater than or equal to 13 years of age) with human immunodeficiency virus (HIV) infection. This Task Force concluded that the morbidity, mortality, and cost due to PCP could be substantially reduced by appropriate use of antipneumocystis prophylaxis in subgroups of HIV-infected patients known to be at high risk, and developed recommendations for the administration of prophylactic regimens (1). The recommendations state that CD4+ T-lymphocyte counts should be monitored prospectively at 3- to 6-month intervals and prophylaxis should be instituted when patients become immunologically susceptible to PCP. Susceptibility was defined by a CD4+ T-lymphocyte count less than 200 cells/microliters or less than 20% of total circulating lymphocytes, or the occurrence of a previous episode of PCP. The goal of this approach was to reduce the frequency both of initial episodes of PCP (primary prophylaxis) and of relapses or recurrences (secondary prophylaxis). Either oral trimethoprim-sulfamethoxazole (TMP-SMX) or aerosol pentamidine was recommended for prophylaxis, but because direct comparative data were lacking, neither regimen was endorsed as "preferred." Since the recommendations were issued in 1989, additional information has become available about the efficacy and safety of aerosol pentamidine and oral TMP-SMX. A trial sponsored by the National Institute of Allergy and Infectious Diseases AIDS Clinical Trials Group compared these two regimens in a prospective randomized study; in August 1991, this study was terminated by an independent data and safety monitoring board because statistically significantly fewer recurrences of PCP were observed in the oral TMP-SMX group than in the aerosol pentamidine group (2). On the basis of this finding and other studies assessing PCP prophylaxis, the Task Force was reconvened on October 5, 1991. This report contains the revised recommendations issued by the Task Force.


[Encephalitis. Recent clinical, diagnostic and therapeutic aspects].

The subjects of this paper are mainly the encephalitis of viral origin and in addition 3 types of non-viral encephalitis of practical importance. A review is given on the current diagnostic possibilities including; clinical criteria; examination of the cerebrospinal fluid; neuroradiology and; etiological investigation by means of direct identification of the microbes in the cerebrospinal fluid or by measuring intrathecally produced antibodies. After some general statements concerning treatment the specifically treatable encephalitis caused by Herpes simplex virus, Mycoplasma pneumoniae, Borrelia burgdorferi and Toxoplasma gondii are discussed in more detail.


Antidiabetogenic effect of glucagon-like peptide-1 (7-36)amide in normal subjects and patients with diabetes mellitus.

Glucagon-like peptide-1 (7-36) amide (glucagon-like insulinotropic peptide, or GLIP) is a gastrointestinal peptide that potentiates the release of insulin in physiologic concentrations. Its effects in patients with diabetes mellitus are not known. We compared the effect of an infusion of GLIP that raised plasma concentrations of GLIP twofold with the effect of an infusion of saline, on the meal-related release of insulin, glucagon, and somatostatin in eight normal subjects, nine obese patients with non-insulin-dependent diabetes mellitus (NIDDM), and eight patients with insulin-dependent diabetes mellitus (IDDM). The blood glucose concentrations in the patients with diabetes were controlled by a closed-loop insulin-infusion system (artificial pancreas) during the infusion of each agent, allowing measurement of the meal-related requirement for exogenous insulin. In the patients with IDDM, normoglycemic-clamp studies were performed during the infusions of GLIP and saline to determine the effect of GLIP on insulin sensitivity. In the normal subjects, the infusion of GLIP significantly lowered the meal-related increases in the blood glucose concentration (P less than 0.01) and the plasma concentrations of insulin and glucagon (P less than 0.05 for both comparisons). The insulinogenic index (the ratio of insulin to glucose) increased almost 10-fold, indicating that GLIP had an insulinotropic effect. In the patients with NIDDM, the infusion of GLIP reduced the mean (+/- SE) calculated isoglycemic meal-related requirement for insulin from 17.4 +/- 2.8 to 2.0 +/- 0.5 U (P less than 0.001), so that the integrated area under the curve for plasma free insulin was decreased (P less than 0.05) in spite of the stimulation of insulin release. In the patients with IDDM, the GLIP infusion decreased the calculated isoglycemic meal-related insulin requirement from 9.4 +/- 1.5 to 4.7 +/- 1.4 U. The peptide decreased glucagon and somatostatin release in both groups of patients. In the normoglycemic-clamp studies in the patients with IDDM, the GLIP infusion significantly increased glucose utilization (saline vs. GLIP, 7.2 +/- 0.5 vs. 8.6 +/- 0.4 mg per kilogram of body weight per minute; P less than 0.01). GLIP has an antidiabetogenic effect, and it may therefore be useful in the treatment of patients with NIDDM:


Usefulness of the assessment of urinary enzymes and microproteins in monitoring ciclosporin nephrotoxicity.

The clinical usefulness of serial assays of urinary N-acetyl-beta-D- glucosaminidase (NAG), gamma-glutamyltransferase (GGT) and beta 2-microglobulin (beta 2M) were tested to evaluate and follow up the nephrotoxicity resulting from the prolonged administration of ciclosporin (CS). Three groups of patients were studied for 18 months: group A: functioning renal transplant patients (n = 13) on maintenance therapy from 12-31 months with CS and prednisone; group B: functioning renal transplant patients (n = 11) treated with prednisone and azathioprine; group C: patients (n = 10) affected by autoimmune steroid-unsensitive uveitis, free from previous renal disorder and treated with CS (for 8-16 months) at progressively decreasing doses. In groups A and B, the urinary enzymes and beta 2M underwent overlapping increases, so that these parameters cannot be considered reliable indices of CS-induced nephrotoxicity. This is due to the fact that transplanted kidneys are already altered by concomitant or preexisting affections, or by persistent immunologic injury. Conversely, in patients with uveitis, the serial assays of such urinary parameters prove to be quite reliable to evidence clinically yet unrecognizable kidney involvement due to prolonged CS administration. High enzymuria has been shown to be an earlier marker of nephrotoxicity only in nephropathy-free patients; on the other hand, the regression of elevated beta 2Muria into normal ranges indicates complete tubule cell recovery.


Dopa accumulates in the hypothalamic-hypophysial portal vessels and is taken into the anterior pituitary of NSD-1015-treated rodents.

DOPA was measured in the anterior pituitary and hypothalamic-hypophysial portal blood after treatment with NSD-1015, a DOPA decarboxylase inhibitor. NSD-1015 caused DOPA to accumulate in the anterior pituitary of mice and rats, and increased DOPA in the hypothalamic-hypophysial portal blood of rat. Serum prolactin was also increased. Interruption of the anterior pituitary blood supply from the hypothalamic-hypophysial system by cannulation of the entire pituitary stalk eliminated the NSD-1015-induced DOPA accumulation in the rat pituitary. We conclude that DOPA can be taken into the anterior pituitary from the portal blood of NSD-1015-treated rodents and that the anterior pituitary lacks tyrosine hydroxylase activity in both mice and rats.


Central mechanisms subserving the impaired growth hormone secretion induced by persistent blockade of NMDA receptors in immature male rats.

Recently, we have reported in immature female rats that short-term blockade of glutamate receptors of the N-methyl-D-aspartic acid (NMDA) subtype by the noncompetitive antagonist MK-801 induced a reduction of growth rate, basal and stimulated growth hormone (GH) release and plasma somatomedin C levels. In the present study, we investigated in immature male rats the mechanism(s) through which agonists and antagonists of glutamate receptors affect GH secretion. In 21-day-old male rats, administration of MK-801 (0.2 mg/kg i.p.b.i.d.) for 10 days induced a significant impairment of growth rate, which was unrelated to a significant reduction of food intake. GH secretion from anterior pituitary fragments of MK-801-treated rats was not significantly reduced under basal conditions but was significantly less under stimulation by 40 mM K+. Incubation of dispersed pituitary cells of 31-day-old rats with N-methyl-aspartic acid (1 and 100 microM), alone or associated with MK-801 (1 microM) did not change GH secretion. Semi quantitative densitometric analysis of hypothalami of MK-801-treated rats evidenced a clearcut decrease in the intensity of GHRH-like immuno-reactivity (LI) staining in the median eminence (ME), whereas no difference was observed in the ME-somatostatin (SS)-LI. Finally, GHRH mRNA but not SS-mRNA, evaluated by slot-blot hybridization, was reduced in the hypothalamus of MK-801-treated rats. These and our previous data would demonstrate that NMDA glutamate receptors play an important role in the neuroendocrine control of GH secretion in the rat, and suggest an action mediated by GHRH-secreting neurons.


The effect of benzodiazepines on the fetus and the newborn.

The effect of the maternal use of benzodiazepines (BZD) on the fetus and the newborn infant has been studied in a representative series of 17 newborn infants (BZD group). The pregnancy and the perinatal period were characterized by 20 items. On the 2nd day of life, a neurologic investigation was performed and comprised a total of 38 items, subgrouped into items of reflexes/reactions, tonus, and other symptoms and signs. An optimum finding for each item was selected. The results were compared with a group of 21 newborns fetally exposed to psychotropic drugs other than BZD (drug group) and a reference group of 29 newborns with no known fetal exposure to drugs. Infants in the BZD group had a lower birth weight for birth length, as compared to both the drug group and the reference group. Significant differences in frequency of pre- and perinatal complications and in neuro-behavior between the BZD group and the reference group were found in all groups of items. We conclude that the use of BZD during pregnancy is associated with impaired intrauterine growth and an increased frequency of pre- and perinatal events. It affects the newborn infant neurologically mainly in the form of intoxication and withdrawal symptoms.


Mononeuritis multiplex in a child with cutaneous polyarteritis.

A 14-year-old boy presented with a febrile illness associated with arthritis. Shortly later he developed mononeuritis multiplex. After certain typical skin lesions had developed after two months, the diagnosis cutaneous polyarteritis could be made. The diagnostic features of this benign disease, which may involve peripheral nerves, are discussed.


Prion protein mutation at codon 102 in an Italian family with Gerstmann-Sträussler-Scheinker syndrome.

We present the first family from Italy with the Gerstmann-Sträussler-Scheinker syndrome (GSS) and a substitution of leucine for proline at codon 102 of the prion protein gene. This mutation is associated with the ataxic form of GSS in a number of reported families. The clinical presentation of our family includes amyotrophic changes in some affected family members in addition to ataxia.


Discordance of the T-cell receptor alpha-chain gene in familial multiple sclerosis.

We studied restriction fragment length polymorphisms of the T-cell receptor alpha-chain (TCR alpha) gene in DNA obtained from 99 individuals of 14 multiplex families with multiple sclerosis (MS). Thirty-four family members had definite MS and two had probable MS. Six normal family members had abnormal cranial MRIs. Linkage analysis utilized constructed haplotypes of EcoRV, Sst I, and Taq I polymorphisms. With penetrance values from 0.1 to 0.7, and scoring the normal individuals with abnormal MRIs as either unknown or affected, LOD scores were between -3.16 and -7.95 for the autosomal dominant model. For the autosomal recessive model with a penetrance range from 0.1 to 1, the LOD scores ranged from -6.77 to -23.08. These findings do not support a direct role of TCR alpha in the inheritance of MS.


[Takayasu disease in the genesis of cerebrovascular insufficiency. Description and considerations of a clinical case].

The authors in explaining their limited clinical experience relative to one single case, describe the peculiarity of Takayasu's disease and its rare incidence in western populations. The case under observation because of its peculiar rarity had been underestimated and treated in a completely inappropriate manner. Important instrument in its diagnosis is the High Resolution echography which in determining the diagnosis favors and speeds up the diagnostic therapeutic iter.


Cloning and characterization of the mouse neu promoter.

We have isolated a genomic clone containing the mouse neu gene. The 5' end of the mouse neu gene was localized by Southern analysis, subcloned and characterized. DNA sequence analysis revealed that the promoter region is 67% G+C-rich and lacks a TATA box, although a CAAT box is present. By sequence comparison, we identified several consensus recognition sequences for general transcription factors such as Sp1, E4TF1, AP2, OTF-1 and GCF, as well as recognition sequences for RVF, E1A and GTG, which have recently been identified in the rat neu promoter. Functional promoter activity was demonstrated by the ability of the promoter to drive transcription of the bacterial chloramphenicol acetyltransferase gene. Using a series of 5'-end deletion mutants, we have identified multiple positive and negative cis-acting elements that regulate mouse neu gene transcription.


Neisseria gonorrhoeae PilC expression provides a selective mechanism for structural diversity of pili.

Pili of Neisseria gonorrhoeae undergo both phase and structural variation. Phase variation of gonococcal pili can be caused by a RecA-independent on/off switch in PilC, a protein involved in pilus biogenesis. We show here that spontaneous nonpiliated PilC- derivatives as well as PilC- insertional mutants have also acquired sequence alterations in pilE relative to the pilE gene of the piliated MS11mk(P+)-u parent, so that the pilin produced is processed to soluble S-pilin and can be released into the medium. It is proposed that pilin alterations are selected for in PilC- bacteria if the parental nonassembled pilin is toxic to the cells--i.e., is not degradable to S-pilin at rates sufficient to allow viability of the cells. Toxicity is indicated by the extreme instability of certain unassembled pilin sequences and by the low frequency of nonpiliated, pilin+, PilC- variants that emerge from piliated recA- cells. The presence of a point mutation changing leucine-39 to phenylalanine at the cleavage site for S-pilin in one nonpiliated, PilC-, recA- variant relative to its piliated parent is a further argument for a selective mechanism of structural diversity of the gonococcal pilin.


RHS2, a POU domain-containing gene, and its expression in developing and adult rat.

Gene expression within the central nervous system is regulated by complex interactions of DNA-binding proteins, among which are the POU domain-containing proteins, which are distantly related to homeobox proteins. These POU domain-containing proteins have been implicated in control of transcription and replication within the central nervous system. We used degenerate primers with the PCR to isolate another POU domain-containing cDNA, RHS2, from hypothalamic RNA. Isolation of a putative full-length cDNA was accomplished by using serial dilutions of a hypothalamic cDNA library grown on solid medium. This member of the class III POU family is expressed in rats from embryonic day 11.5 into adulthood, being especially prominent in the brain. We performed double-labeling hybridization histochemistry and determined that RHS2 is coexpressed with a variety of neuropeptides in medium-sized neurons in the caudate putamen and with dynorphin in the paraventricular and supraoptic nuclei of the hypothalamus. Expression of RHS2 in the caudate putamen was increased by elimination of its nigrostriatal dopaminergic innervation.


P-type voltage-dependent calcium channel mediates presynaptic calcium influx and transmitter release in mammalian synapses.

We have studied the effect of the purified toxin from the funnel-web spider venom (FTX) and its synthetic analog (sFTX) on transmitter release and presynaptic currents at the mouse neuromuscular junction. FTX specifically blocks the omega-conotoxin- and dihydropyridine-insensitive P-type voltage-dependent Ca2+ channel (VDCC) in cerebellar Purkinje cells. Mammalian neuromuscular transmission, which is insensitive to N- or L-type Ca2+ channel blockers, was effectively abolished by FTX and sFTX. These substances blocked the muscle contraction and the neurotransmitter release evoked by nerve stimulation. Moreover, presynaptic Ca2+ currents recorded extracellularly from the interior of the perineural sheaths of nerves innervating the mouse levator auris muscle were specifically blocked by both natural toxin and synthetic analogue. In a parallel set of experiments, K(+)-induced Ca45 uptake by brain synaptosomes was also shown to be blocked or greatly diminished by FTX and sFTX. These results indicate that the predominant VDCC in the motor nerve terminals, and possibly in a significant percentage of brain synapses, is the P-type channel.


Identification of a mammalian 10-kDa heat shock protein, a mitochondrial chaperonin 10 homologue essential for assisted folding of trimeric ornithine transcarbamoylase in vitro.

We have identified a 10-kDa stress-inducible mitochondrial protein. The protein is synthesized at elevated rates in cultured rat hepatoma cells challenged with heat shock or amino acid analogues and, therefore, designated heat shock protein 10 (Hsp10). Hsp10 was purified to homogeneity from rat liver and found to exhibit a native molecular mass of 65 kDa, as opposed to a monomeric molecular mass of 10,813.4 +/- 0.41 Da. The amino acid sequence of rat Hsp10 disclosed extensive sequence similarity with bacterial chaperonin (Cpn) 10. Rat Hsp10 and Escherichia coli Cpn60 were used to reconstitute functional trimeric rat ornithine transcarbamoylase from a chemically denatured state with high efficiency. This process depended completely upon rat Hsp10 and was abolished in the presence of a nonhydrolyzable ATP analogue. We conclude that Hsp10 is a eukaryotic Cpn10 homologue and, therefore, together with Cpn60 essential for mitochondrial protein biogenesis. The Cpn-mediated protein-folding apparatus, thus, exhibits a high degree of conservation between prokaryotes and mitochondria of higher eukaryotes.


mRNA coding for neurotransmitter receptors in a human astrocytoma.

Electrophysiological techniques and Xenopus oocytes were used to study the expression of neurotransmitter receptors encoded by mRNAs isolated from three human glioma cell lines. Oocytes injected with mRNAs from two glioblastoma cell lines did not show electrical responses to the various neurotransmitters tested. In contrast, oocytes injected with mRNA from an astrocytoma cell line (R-111) acquired acetylcholine and glutamate receptors as well as a small number of N-methyl-D-aspartate (NMDA) receptors. Acetylcholine elicited oscillatory Cl- currents that were abolished by muscarinic antagonists. The muscarinic receptors are coupled to the inositol phosphate-Ca2+ receptor-channel coupling system. Glutamate and its analogs kainate, quisqualate, and alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid induced smooth currents. The non-NMDA responses were potently blocked by 6,7-dinitroquinoxaline-2,3 dione. Our results show that human astrocytoma cells contain mRNAs coding for functional acetylcholine and glutamate receptors that have properties similar to those of neurons. In contrast, human glioblastoma cells lacked those mRNAs. These differences might be useful for the development of new diagnostic and therapeutic procedures.


Genetic transfer of non-P-glycoprotein-mediated multidrug resistance (MDR) in somatic cell fusion: dissection of a compound MDR phenotype.

A non-P-glycoprotein-mediated mechanism of multidrug resistance (non-Pgp MDR) has been identified in doxorubicin-selected sublines of the human non-small cell lung carcinoma cell line SW-1573. These sublines are cross-resistant to daunorubicin, VP16-213, Vinca alkaloids, colchicine, gramicidin D, and 4'-(9-acridinylamino)methanesulfon-m-anisidide (m-AMSA). They accumulate less drug than the parental cells and their resistance is not due to the MDR1-encoded P-glycoprotein, as the resistant cell lines have lost the low amount of MDR1 mRNA detectable in parental cells. Here we show that the resistant cell lines also contain less topoisomerase II mRNA and enzyme activity than the parental cells. This might contribute to the resistance of these lines to drugs interacting with topoisomerase II, such as doxorubicin, daunorubicin, and VP16-213, but cannot account for the resistance to the other drugs. We have tested whether all properties of the non-Pgp MDR cell lines cosegregate in somatic cell fusions between lethally gamma-irradiated, resistant donor cells and drug-sensitive acceptor cells. Whereas a MDR phenotype with reduced drug accumulation and the loss of MDR1 P-glycoprotein mRNA were cotransferred to the acceptor cells, the decrease in topoisomerase II gene expression was not. We conclude that the MDR phenotype, the reduced drug accumulation, and the loss of MDR1 P-glycoprotein mRNA are genetically linked. They might be due to a single dominant mutation, which does not cause the alteration in topoisomerase II.


[Lymphocytic alveolitis in the early stages of HIV infection: correlation with biological and prognostic factors].

Broncho-alveolar lavage was performed to assess the degree of pulmonary lymphocytic alveolitis in 32 asymptomatic patients who were infected with the Human Immunodeficiency Virus (VIH). The patients were stages II and III of the CDC classification and the aim of the study was to determine the frequency, nature and prognostic role of the findings. 62.5% of the subjects (20/32) presented with a lymphocytic alveolitis which consisted predominantly of CD8 lymphocyte (64.3 +/- 3.5%), in the absence of an opportunistic infection or broncho-pulmonary tumours. Two sub-populations of alveolar CD8 were shown at comparable levels, a) sub-population CD8+D44+ (22.1 +/- 5%), in whom we showed the possession of cytotoxic activity in particular specific for VIH; b) sub-population CD8+CD57+ (19.6 +/- 3%) which we have shown to be capable in vitro of inhibiting the effector phase of cytotoxic activity of CD8+D44+ alveolar cells specific for VIH. In this group of 32 patients the occurrence of an alveolitis was not correlated with the usual prognostic factors of infection by VIH measured simultaneously with broncho-alveolar lavage (the level of CD4+ blood lymphocytes, and the beta 2-plasma microglobulins and the presence of p24 antigenaemia). In addition the level of CD4 lymphocytes supperior to 400/mm3 and of beta 2-microglobulins less then 3 mg/l whether a lymphocytic alveolitis was there or not confirmed the relatively poorly developed state of the VIH infection in these asymptomatic patients. Also the occurrence of a lymphocytic alveolitis did not seem to be linked to progression of the disease in the group of patients studied.(ABSTRACT TRUNCATED AT 250 WORDS)


[A case of congenital long QT syndrome associated with T wave alternans].

A case was presented in which a rare T wave alternans occurred in association with congenital long QT syndrome. A 71-year-old woman, who had experienced several syncopal attacks per year for the previous forty years, was admitted for further evaluation of the syncope. She had a family history of sudden death (sister) and QT prolongation (son). Electrocardiogram showed a corrected QT interval of 0.68 seconds. Treadmill exercise-tolerance test revealed both T wave alternans immediately after exercise and torsades de pointes 150 seconds after exercise. The syncope was induced by the mental excitation. A prolonged corrected QT interval reduced from 0.70 seconds to 0.58 seconds by the correction of her serum potassium and magnesium. The effect of propranolol, verapamil, phenytoin or mexiletine on T wave alternans and ventricular arrhythmia was evaluated by the treadmill exercise-tolerance test. The treatment with propranolol was most effective.


[New virological data: replication and persistence of the human immunodeficiency virus].

Human immunodeficiency virus behaves in vitro as an acutely replicating, cytopathic retrovirus. However, HIV infection in vivo is persistent and slowly progressive. Many virological properties of HIV could be involved in establishing the persistence of infection: the complex regulation of HIV replication, particularly of HIV gene expression; the requirement of cell activation for several steps of HIV replication; the role of HIV accessory genes; the great genetic variability of HIVs, allowing their adaptation to host responses in vivo.


HIV1 quantitation in infected patients: a comparison of cell viraemia, plasma viraemia and R-HEV.

Anti-HIV1 trials require sensitive, reproducible and reliable parameters which measure virus load and virus production. The aim of the present study of 14 patients at different CDC stages was to compare results provided by three techniques which evaluate, respectively, cell-associated HIV1 infectivity (cell viraemia), plasma-associated HIV1 infectivity (plasma viraemia) and radiation-resistant expression ex vivo (R-HEV). For each patient, TCD4 lymphocytes were numbered and an HIV1 antigenaemia determination was carried out. We found no correlation between cellular viraemia and R-HEV; on the contrary, data concerning plasma viraemia and R-HEV were significantly correlated. While cellular viraemia and R-HEV were not correlated with TCD4 levels, plasma viraemia exhibited a significant negative correlation with TCD4 counts. Despite theoretical and practical differences, plasma viraemia and R-HEV may measure events in the viral replication cycle which bear a close resemblance; however, as designed, R-HEV seems to be far less discriminative than plasma viraemia.


[Asthma therapy: are bronchodilators obsolete?].

Short acting beta 2-agonists, such as salbutamol, fenoterol and terbutaline, are the most potent bronchodilating agents in the treatment of acute asthmatic attacks. In chronic asthma, however, they must be viewed as purely symptomatic treatment compared to antiinflammatory agents (inhaled corticosteroids, cromoglycate and nedocromil sodium), prophylactic agents which can also decrease airway hyperresponsiveness and thereby stabilize the disease. Modern asthma treatment are based on the present understanding of asthma pathophysiology and take into account the new therapeutic options with high dose inhaled steroids and long acting beta 2-agonists. Patients with only occasional asthma symptoms should receive inhaled short acting beta 2-agonists as needed. These agents continue to be central to the treatment of any asthmatic attack. If asthma symptoms occur more than 4-6 times a week a regular inhalative antiinflammatory regimen should be initiated (e.g. inhalative corticosteroids b.i.d.). In cases with frequent symptoms or persisting airway obstruction, regularly administered inhalative bronchodilators (beta 2-agonists, possibly long acting) should be added to high dose inhaled steroids. Treatment of severe acute asthma should always include systemic glucocorticosteroids (e.g. prednisone 30-60 mg/day). Special emphasis should be placed on self-management plans, encouraging patients to monitor and treat their disease themselves in close cooperation with the physician.


Balancing selection at allozyme loci in oysters: implications from nuclear RFLPs.

Population genetic analyses that depend on the assumption of neutrality for allozyme markers are used widely. Restriction fragment length polymorphisms in nuclear DNA of the American oyster evidence a pronounced population subdivision concordant with mitochondrial DNA. This finding contrasts with a geographic uniformity in allozyme frequencies previously thought to reflect high gene flow mediated by the pelagic gametes and larvae. The discordance likely is due to selection on protein electrophoretic characters that balances allozyme frequencies in the face of severe constraints to gene flow. These results raise a cautionary note for studies that rely on assumptions of neutrality for allozyme markers.


Competition for overlapping sites in the regulatory region of the Drosophila gene Krüppel.

A 730-base pair element regulates expression of the Drosophila gap gene Krüppel (Kr) in response to the fly anterior morphogen bicoid (bcd). Two hormone receptor-like proteins, encoded by the genes knirps (kni) and tailless (tll), bind specifically to the element. In vitro, kni protein competes with the homeodomain-containing bcd protein in binding to a 16-base pair target sequence. In vivo experiments suggest that both kni and tll act as competitive repressors of bcd-mediated activation of Kr. These results suggest a mechanism by which developmental genes can be regulated in response to an activating morphogen gradient antagonized by repressors.


Use of a dihydrogen osmium complex as a versatile 1H NMR recognition probe.

A new recognition probe for biomolecules, [en2Os(eta 2-H2)]2+ (1; en, ethylenediamine), is reported. In aqueous solution, 1 binds readily to a variety of biomolecules, including nucleotides, RNA, amino acids, peptides, and phospholipids. In each case, binding leads to a characteristic proton nuclear magnetic resonance (1H NMR) for the dihydrogen that appears in a spectral window in the range delta = 0 to -20 parts per million, and as well to characteristic values of the coupling JHD and of the relaxation time T1. Small structural differences in molecules such as DGMP (2'-deoxyguanosine 5'-monophosphate) and IMP (inosine 5'-monophosphate) or Asp and Glu can readily be distinguished, such as when 1 binds to the N-7 position of the nucleobase of DGMP or IMP and when 1 binds to the carboxylate of Asp or Glu. Upon one-electron oxidation of the metal center, diamagnetic 1 is converted to a paramagnetic probe.


Functional complementation of yeast ste6 by a mammalian multidrug resistance mdr gene.

Multidrug resistance in mammalian tumor cells is associated with the overexpression of mdr genes encoding P-glycoproteins, which function as drug efflux pumps. A yeast homolog of mdr, STE6, mediates export of a-factor mating peptide. Yeast MATa cells carrying a ste6 deletion produce no extracellular a-factor and therefore are defective in mating. Expression of a complementary DNA for the mouse mdr3 gene in a yeast ste6 deletion strain restored ability to export a-factor and to mate. A mutation (a serine to phenylalanine substitution at amino acid 939) known to affect the activity of the mdr3 gene product abolished its ability to complement the yeast ste6 deletion. Thus, functions of P-glycoproteins in normal mammalian cells may include the transmembrane export of endogenous peptides.


Magnetic resonance imaging of tendon and ligament abnormalities: Part II. Pelvis and lower extremities.

Magnetic resonance imaging (MRI) has provided an ideal means, unmatched by other preexisting modalities, of examining musculoskeletal abnormalities, particularly those involving tendons and ligaments in the lower extremities. Lack of motion artifact, convenience of application of surface coil, and absence of overlying structures have made the lower extremities ideally suited to MRI. In addition, the abundance of adjacent adipose tissue provides a superb contrast background. Although evaluation of trauma remains the most common reason for MRI examination, many other conditions may also affect tendons and ligaments. As in other soft-tissue, chondral, and osteochondral lesions, MRI provides exquisite details of abnormalities in these structures. Part II of this review systematically reviews the abnormalities of tendons and ligaments in the pelvis and lower extremities.


Dissection of the uncinate process during pancreatoduodenectomy.

An alternative approach to the dissection of the uncinate process during pancreatoduodenectomy is described. This involves mobilisation of the midgut superiorly and bringing the uncinate process into an antero-inferior position within the operative field. Access to, and visualisation of, the posterior aspect of the superior mesenteric vessels is considerably improved. This approach may avoid many of the potential problems associated with freeing of the uncinate process during pancreatic resection.


Laparoscopy for the non-palpable testis--look before you cut!

Before definitive surgery laparoscopies were performed to define the position of 24 non-palpable testes in 21 patients. The anatomical site of the testes was confirmed by surgery in all cases. Laparoscopy is helpful in planning the definitive procedure in these cases.


The influence of antihypertensive agents on circadian rhythms of blood pressure and heart rate in patients with essential hypertension.

The effects of once-daily administration of calcium (Ca) channel blockers, beta-blockers and and angiotensin-converting enzyme (ACE) inhibitors on circadian rhythms of blood pressure (BP) and heart rate (HR) were studied using the cosinor method. Sixty-two recruited patients with essential hypertension (WHO stage I or II) were divided into three groups based on the class of administered drugs. In the Ca channel blocker group (n = 37, age 54 +/- 9.0 years), 18 patients were given YM 730 at a mean dose of 11 +/- 4.0 mg/day (mean +/- S.D.), 8 were given nitrendipine (11 +/- 6.7 mg/day), and 11 were given nisoldipine (8 +/- 6.4 mg/day). In the beta-blocker group (n = 15, age 42 +/- 13.5 years), 13 patients were given atenolol (44 +/- 11.0 mg/day), 1 was given nadolol (30 mg/day), and 1 was given sustained-release propranolol (60 mg/day). In the ACE inhibitor group (n = 10, age 56 +/- 8.7 years), 7 patients were given enalapril (6 +/- 2.8 mg/day), and 3 were given lisinopril (20 mg/day). Ambulatory BP monitoring (ABPM) was performed before and during treatment. Mean arterial pressure (MAP) and HR were monitored under ambulatory conditions every five minutes for 24 hr with a finger volume oscillometric device. In all three groups, the mesor of MAP decreased significantly, while the amplitude and acrophase did not change during treatment. beta-Blockers reduced the amplitude as well as the mesor of HR. Ca channel blockers increased the amplitude of HR without influencing the mesor. ACE inhibitors had no effect on the circadian rhythm parameters of HR. These results suggest that Ca channel blockers, beta-blockers and ACE inhibitors lowered BP throughout the day without changing the circadian BP rhythm. However, the three drug classes may have different influences on the autonomic nervous system that regulates circadian cardiac rhythm.


Exogenous Thy-1.1 expression as a marker for thymocyte maturation in transgenic mice.

We established a line of transgenic mice carrying the exogenous mouse Thy-1.1 gene (8.2 Kb Eco RI genomic DNA fragment). In these mice, Thy-1.1 was expressed on thymocytes but not on peripheral T cells, presumably due to the lack of cis-acting element(s) on the microinjected genomic DNA. Even in the thymus, however, while most of the CD3/TCR- thymocytes were positive for the transgenic Thy-1.1 gene expression, the CD4+ or CD8+ single positive (SP) thymocytes were composed of two groups, one Thy-1.1+ and one Thy-1.1-, suggesting that the former belongs to cells at premature stages of terminal T cell differentiation. There was no difference in the amount of CD3/TCR complexes expressed on two such SP thymocyte subsets. In the double negative (DN) thymocytes, all the CD3/TCR+ cells were Thy-1.1-. These results suggest that the maturational process of CD3+ DN thymocytes differs from that of SP thymocytes. The unique distribution of Thy-1.1+ population among CD3+ thymocytes suggests that the transgenic Thy-1.1 gene expression can serve as a useful marker to examine terminal maturation processes of CD3+ thymocytes.


Monoclonal antibodies against human T cell adhesion molecules--modulation of immune function in nonhuman primates.

The cytotoxic T cell is thought to be a primary effector of allograft rejection. In vitro studies have demonstrated that the interaction between cytotoxic T cells and target cells involves cell surface adhesion molecules that result in conjugate formation, with subsequent antigen recognition, T cell activation, and target cell lysis. Experiments have also demonstrated the ability of monoclonal antibodies with specificity for two human T cell adhesion molecules, lymphocyte function associated (LFA) antigen-1 (LFA-1, CD11a, alpha-chain/CD18, beta-chain) and LFA-2 (CD2), to inhibit conjugate formation in vitro. Studies in a nonhuman primate model were undertaken to determine whether the in vivo administration of monoclonal antibodies with specificity for the alpha chain of LFA-1 (CD11a) or with specificity for CD2 could modulate in vivo T cell function. Cynomolgus monkeys (Macaca fascicularis) received 10 daily intravenous infusions of either anti-CD11a, anti-CD2 or both anti-CD11a and anti-CD2 monoclonal antibodies. Antibody administration was well tolerated and resulted in high levels of circulating murine monoclonal antibody in the peripheral circulation. Nearly all the animals generated antimurine antibodies that were specific for both idiotypic and nonidiotypic determinants of the infused mouse protein. Circulating lymphocytes and T cells were not depleted by treatment with anti-CD11a or anti-CD2 mAbs; in fact, treatment with the combination of anti-CD11a plus anti-CD2 or anti-CD11a alone led to increased numbers of circulating lymphocytes and T cells. Modulation of the LFA-1 molecule on circulating T cells occurred as a result of treatment with anti-CD11a (or the combination of anti-CD11a plus anti-CD2), whereas treatment with anti-CD2 (or anti-CD11a plus anti-CD2) did not result in modulation of the CD2 antigen despite detectable levels of circulating anti-CD2 mAb. In vivo T cell function was assessed by placement of skin allografts. As compared with treatment with saline or a control mAb, allograft survival was significantly prolonged in animals treated with anti-CD11a or combination treatment but not in animals receiving anti-CD2 alone. We conclude that the in vivo administration of anti-LFA-1 mAb may be useful for the blockade of effector T cell activity during allograft rejection, that saturation of antigen and antigen modulation may be important for efficacy of such antibody effects in vivo, and that monoclonal antibodies with specificity for functionally important T cell surface molecules may alter T cell function in vivo without lymphocyte depletion.


Stimulation of CD4+ T lymphocytes by allogeneic MHC peptides presented on autologous antigen-presenting cells. Evidence of the indirect pathway of allorecognition in some strain combinations.

A preliminary analysis of the alloantibody response to free, unconjugated class I and class II MHC peptides in several rat and mouse strains was performed, to screen for an effective interaction between the allogeneic MHC peptides and recipient MHC molecules. The PVG rat strain was noted to produce very strong, MHC-restricted, primary and secondary responses to a synthetic peptide derived from the alpha helical region of the alpha 2 domain of an RT1.C/E class I MHC molecule of the DA strain. In vitro proliferation studies demonstrated that CD4+ but not CD8+ T cells of the PVG strain responded in a recipient APC-dependent manner to the peptide, whereas the BN strain (which showed no antibody response to this peptide) gave no T cell proliferation. Immunization of PVG rats with the peptide did not influence the rejection of DA skin allografts. The relevance of these studies to the possible mechanisms of allograft rejection by an indirect pathway are discussed.


Cell kinetic analyses and expression of carcinoembryonic antigen, carbohydrate antigen 19-9 and DU-PAN-2 in hyperplastic, pre-neoplastic and neoplastic lesions of intrahepatic bile ducts in livers with hepatoliths.

We evaluated cell proliferative activity and expression of carcinoembryonic antigen (CEA), carbohydrate antigen 19-9 (CA 19-9) and DU-PAN-2 in various bile duct lesions in livers with hepatoliths, using histochemical and immunohistochemical methods. Histologically, the bile duct lesions were divisible into hyperplasia, dysplasia, adenocarcinoma in situ and invasive adenocarcinoma. All cases showed mucosal hyperplasia in stone-bearing bile ducts. Livers with invasive adenocarcinoma frequently contained adenocarcinoma in situ and dysplasia, and livers with adenocarcinoma in situ occasionally harboured dysplasia. Proliferating cell nuclear antigen (PCNA) labelling index was low in hyperplasia (mean +/- SD = 20.5 +/- 8.7%), intermediate in dysplasia (35.4 +/- 15.9%), and high in adenocarcinoma in situ (46.4 +/- 9.3%). The mean number of argyrophilic nucleolar organizer regions (AgNORs) was low in hyperplasia (1.52), intermediate in dysplasia (2.26) and high in adenocarcinoma in situ (2.69). There was a significant positive correlation between PCNA labelling index and AgNORs count. CEA was expressed on invasive adenocarcinoma cells and adenocarcinoma in situ cells in most cases and on dysplastic cells in about a half, while CEA was never present in hyperplastic epithelia. Expression of CA 19-9 was low in adenocarcinoma, intermediate in dysplasia and rather high in hyperplasia. There was no significant difference in DU-PAN-2 expression among these bile duct lesions. These data suggest that cell replicative activity is low in hyperplasia, intermediate in dysplasia and high in adenocarcinoma in situ, and that CEA appears in the following order: dysplasia, adenocarcinoma in situ, invasive adenocarcinoma. We suggest that carcinogenesis in biliary epithelial in livers with stones is a multi-step process through hyperplasia, dysplasia and adenocarcinoma in situ to invasive adenocarcinoma.


Immunohistochemical studies on oncogene products (EGF-R, c-erbB-2) and growth factors (EGF, TGF-alpha) in human breast cancer: their relationship to oestrogen receptor status, histological grade, mitotic index and nodal status.

In this investigation, 83 human mammary carcinomas were examined for the expression of oestrogen receptor (ER), epidermal growth factor receptor (EGF-R), epidermal growth factor (EGF), transforming growth factor alpha (TGF-alpha), c-erbB-2, histological grade, mitotic index and nodal status, all of which are reportedly prognostically significant factors (Bloom and Richardson 1957; Baak et al. 1985; Wright et al. 1989). ER expression was biochemically recognized in 43.4% of mammary carcinomas, and EGF-R, EGF, TGF-alpha and c-erbB-2 were histochemically recognized in 25.3, 14.5, 27.7 and 18.0% of mammary carcinomas examined respectively, using conventional sections of buffered formalin-fixed, paraffin-embedded tissue and monoclonal or polyclonal antibodies. There were significant relationships between negative ER and positive EGF-R or TGF-alpha; positive EGF-R and TGF-alpha; positive EGF-R and c-erbB-2; and positive c-erbB-2 and TGF-alpha. The single changes which were the negative ER and the positive c-erbB-2 correlated with histological grade and mitotic index. Co-expression of EGF-R and TGF-alpha correlated with positive nodal status. Therefore, the present investigation indicates that the negative ER, single expression of c-erbB-2 and co-expression of EGF-R and TGF-alpha are important markers which contribute indirectly to prognosis, which reconfirms previous findings on the former two while adding the new finding that immunohistochemical demonstration of expression of EGF-R and TGF-alpha may provide useful information for selecting the appropriate treatment.


Establishment and characterisation of two cell lines with different grade of differentiation derived from one primary human pancreatic adenocarcinoma.

From a liver metastasis of a human pancreatic adenocarcinoma, we have established cell lines for studying the cell biology of this tumor. We obtained two cell lines with different morphological, chromosomal and functional properties. One of them, named PaTu 8988s, revealed a solid growth in nude mouse xenografts with cells exhibiting only occasional polar organisation of the cytoplasm. In general, no apical or basolateral plasma membrane domains could be distinguished and the sparse organelles were randomly distributed throughout the cytoplasm. Secretory products, such as mucin, were weakly stained histochemically or were completely absent. Transglutaminase (TGase) activity used as a marker for cellular differentiation was low in these cells. The other cell line, named PaTu 8988t, grew tumors composed of tubular structures when injected subcutaneously into nude mice. Cells were polarized with distinct apical and basolateral plasma membranes and the cytoplasmatic organelles were arranged with the nucleus in the lower part of the cell, while the apical cytoplasm contained the Golgi complex and numerous secretion granules. A high content of mucin was stained histochemically and transglutaminase activity was ten times higher than in PaTu 8988s. Comparing the chromosome number per metaphase plate, both cell lines showed a major peak, with 45-55 chromosomes per metaphase plate in PaTu 8988s and about 110-120 chromosomes per metaphase plate in PaTu 8988t. When the two cell lines were injected intravenously into the tail vein of nude mice, only PaTu 8988s developed metastases localized exclusively in the lung, whereas PaTu 8988t produced no metastases in any organ. We conclude, that two cell lines exhibiting different grades of differentiation as well as a different potency to metastasize can be established from the same primary tumor, and that these cell lines represent a suitable model for further study of the cell biology of human pancreatic adenocarcinoma.


Autometallographic detection of mercury in rat spinal cord after treatment with organic mercury.

Autometallography was used to localize mercury in rat spinal cord after intraperitoneal administration of methylmercuric chloride (200 micrograms CH3HgCl daily). The technique permits small amounts of mercury sulfides and mercury selenides to be visualized by silver-enhancement. Mercury deposits were observed by light microscopy only in neurons. In all of the spinal cord segments selected (first cervical segment, C1; fifth cervical segment, C5; sixth thoracic segment, T6; and first lumbar segment, L1) the mercury was observed with cumulative dosages of 6000 micrograms CH3HgCl and greater. Laminae VII, VIII, and IX contained the majority of stained neurons, whereas laminae IV, V, VI, and X had a relatively lower density of mercury-containing neurons. Stained neurons were confined to specific cell groups, such as Clarke's column, nucleus intermedio-lateralis, nucleus cervicalis centralis, and nucleus dorsomedialis. At the ultrastructural level, mercury deposits were restricted to lysosomes of neurons and occasional accumulations in the lysosomes of ependymal cells.


Distribution of retinol-binding protein in the human digestive tract.

By employing polyclonal antibodies for retinol-binding protein (RBP), its distribution in the human pancreas and digestive tract mucosa was compared with those of transthyretin (TTR) and various peptide hormones. The materials used included surgically removed pancreas, esophagus, stomach, small and large intestines. Paraffin sections were stained by the indirect immunoenzyme method. The results indicate that RBP-containing cells are found in the pancreas and the gastrointestinal mucosa, but most frequently in the gastric antrum and duodenum. In the pancreas, RBP-containing cells are found in the islets and among acinar and ductal epithelial cells, and consistently stain for chromogranin A. RBP-containing cells in the gastrointestinal mucosa showed typical features of endocrine cells and also stained for chromogranin A. The distribution of TTR in these tissue sites resembled that of RBP, but the immunoreactive intensities of both peptides altered independently. Comparison of the distribution of RBP, TTR, and various gastrointestinal peptide hormones revealed that the distribution of RBP coincided with none of the other peptides, although some of the RBP-containing cells stained for most of the peptides examined and vice versa. These results suggest that RBP may be a consistent component of gastrointestinal endocrine cells.


Effects of X-irradiation and adriamycin on quiescent and proliferating cells of the seminal vesicle in the castrated mouse.

The sensitivity of resting and proliferating cells of the seminal vesicle to X-irradiation and adriamycin has been investigated. Stimulation with testosterone propionate (250 micrograms/day) was started 11 days after castration in BALB/c mice. X-rays (2.5-7.5 Gy total body irradiation) and intraperitoneal injections of adriamycin (4-16 mg/kg body weight) were administered at various times before or after induction of proliferation by testosterone injection. The DNA contents and the weights of the seminal vesicles were determined at 4 days after the start of stimulation. A Do for X-rays of about 10 Gy was found for the seminal vesicle epithelium. For both X-irradiation and adriamycin no significant differences in sensitivity were observed between quiescent (Go) and proliferating (G1; S) seminal vesicle cells.


The distribution of alpha 1-antichymotrypsin and amyloid production in the brain in Alzheimer's disease.

In this immunohistopathological study alpha 1-antichymotrypsin, which is barely demonstrable in the normal brain, was found in amyloid fibrils, endothelial cells and the cytoplasm of astroglial cells in brains from patients with Alzheimer's disease. Amyloid precursors stained with methenamine silver were arrayed mainly along the membranes, and amyloid fibrils, which stained densely with anti-alpha 1-antichymotrypsin, were in direct contact with the fibrous structures connecting with the membranes of vascular feet or astrocytic processes. From the above findings, alpha 1-antichymotrypsin seems to play a role in the production of amyloid fibrils in Alzheimer's disease.


The role of macrophages in the uptake of endotoxin by the mouse liver.

The purpose of this investigation was to analyse the macrophage subpopulations involved in the uptake of endotoxin in the liver. The results show that in normal B10.D2 mice the liver macrophages constitute a heterogeneous population of cells which, depending on their state of differentiation, are distinguished by their differential distribution in the liver acinus and by their ability to phagocytose latex. Following the intravenous administration of endotoxin (lipopolysaccharide = LPS) from Salmonella abortus equi, endotoxin-carrying non-parenchymal cells of the liver (NPLC) were investigated immunohistochemically (in situ) and immunocytochemically (after isolation) between 1 h and 14 days after the injection. The endotoxin content of the blood and of isolated NPLC was also determined, using radioactivity labeled LPS. Following LPS injection, the total number of macrophages in the liver increased, reaching a maximum after 3 days. There was a striking increase in the ratio of mature to immature macrophages. After day 3, the number of macrophages decreased again, returning to the pre-injection values by day 14. 1 h after the administration of LPS, 41% of the isolated NPLC were already endotoxin-positive, a percentage which remained constant until the 3rd day. Thereafter, the number of LPS-bearing cells increased to a maximum of about 52% on the 5th day. This increase mostly involved macrophages which had taken up endotoxin. Concurrent with these changes there was a threefold increase in radioactivity-labeled LPS from the 7th h to the 5th day after injection.(ABSTRACT TRUNCATED AT 250 WORDS)


Ultrastructural study of the glomerular slit diaphragm in fresh unfixed kidneys by a quick-freezing method.

In normal kidneys fixed by perfusion with tannic acid and glutaraldehyde, glomerular slit diaphragms have been reported to consist of highly ordered and isoporous substructures with a zipper-like configuration. We have re-evaluated the ultrastructure of fixed or unfixed glomeruli using quick-freezing and deep-etching (QF-DE) and freeze-substitution (QF-FS) methods. In the fixed slit diaphragms, zipper-like substructures were often observed by the QF-DE method. In contrast, in fresh unfixed glomeruli the slit diaphragms mainly consisted of non-porous substructures. The slit diaphragms were more widely opened in the fixed glomeruli, as examined by the QF-FS method. These results suggest that the foot processes shrink during tissue preparation by conventional methods with chemical fixatives, and that the broadening of slit diaphragms and zipper-like substructures are formed by the pulling apart of adjacent foot processes due to shrinkage.


Influence of Parkinson's disease on oral health.

Thirty patients with Parkinson's disease were investigated with regard to their oral health. They had significantly more teeth and less caries than a control group of corresponding age. However, the salivary secretion rate was significantly lower with advancing parkinsonian symptoms. It is concluded that not only motor impairment but also autonomic dysfunction, as an expression of a more advanced neuron degeneration, could be of importance when the possibility of maintaining a good oral health in parkinsonian patients is considered.


Treatment of bile duct stones by laser lithotripsy: results in 12 patients.

We used a pulsed tunable dye laser (operating at 60 mJ per pulse, 504-nm wavelength) to fragment large (0.8-4.5 cm) stones retained in the hepatic ducts or common bile duct in 12 patients after cholecystectomy. Attempts to extract stones via a T-tube or endoscope had been unsuccessful in all patients. In nine of 12 patients, all stone fragments were successfully eliminated during the initial treatment. In one patient, fragmentation occurred but debris remained, requiring endoscopic stenting. Pseudomonas sepsis developed in this patient 30 days after the procedure and was treated by extraction of the stone fragments. Fragments remaining after lithotripsy were cleared at the same sitting by using saline flushing or endoscopic or percutaneous basket extraction. In two of 12 patients, the treatment was unsuccessful because of laser malfunction. The treatment was performed without complications, except for clinically insignificant hyperamylasemia, which occurred in two patients. Our experience suggests that laser lithotripsy offers a safe alternative for nonsurgical treatment of large retained biliary stones for patients in whom traditional treatments have failed.


Restoration of stable metabolic conditions during islet suppression in dogs.

These studies were undertaken to examine the stability of metabolic conditions during islet suppression with fixed-rate insulin and glucagon replacement. Somatostatin was infused peripherally at 0.8 microgram.min-1.kg-1, insulin was infused intraportally at 200 microU.min-1.kg-1, and glucagon was infused intraportally at 0, 0.6, 1, 2, 5, or 20 ng.min-1.kg-1 in conscious overnight-fasted dogs. [3-3H]glucose was infused for measurement of glucose kinetics. During infusion, plasma insulin was 7.2 +/- 0.4 microU/ml. Plasma glucagon rose linearly with glucagon dose, achieving basal levels at 2 ng.min-1.kg-1 infusion (164 +/- 18 vs. basal = 182 +/- 57 pg/ml). Plasma glucose and hepatic glucose output (HGO) decreased from basal at doses 0, 0.6, and 1 ng.min-1.kg-1, increased from basal at doses 5 and 20 ng.min-1.kg-1, and remained close to basal at dose 2 ng.min-1.kg-1 (92 +/- 20 vs. basal = 99 +/- 3 mg/dl and 2.4 +/- 0.2 vs. basal = 2.7 +/- 0.2 mg.min-1.kg-1 for glucose and HGO, respectively; P greater than 0.47). Glucose clearance and blood lactate were also closely matched to basal at dose 2 ng.min-1.kg-1. Coefficients of variation during 2 ng.min-1.kg-1 glucagon infusion (last hour) were 3.4, 4.6, 4.9, and 4.7% for glucose, HGO, clearance, and lactate, respectively. These findings indicate that fasting metabolic conditions, as inferred from blood glucose, lactate, insulin, and glucagon levels, and the rates of glucose production and uptake can be recreated in toto during fixed-rate islet hormone replacement.


Hormones increase oxygen uptake in periportal and pericentral regions of the liver lobule.

The effect of several hormones known to alter intracellular free Ca2+ on rates of O2 uptake in periportal and pericentral regions of the liver lobule was studied in the perfused liver. Regional O2 uptake was measured by stopping the flow and monitoring the decrease in O2 concentration. When perfusion was in the anterograde direction, basal rates of O2 uptake were two to three times higher in periportal than in pericentral regions, and phosphorylase alpha activity, which increases as a function of intracellular free Ca2+ levels, was higher in periportal regions. In contrast, when perfusion was in the retrograde direction, rates of O2 uptake were two to three times greater in pericentral regions. Infusion of epinephrine (0.1 microM) or angiotensin II (5 nM) increased the rate of O2 uptake nearly exclusively in downstream areas of the lobule where O2 tension was low. When perfusions were in the anterograde direction, epinephrine increased phosphorylase alpha activity significantly only in pericentral regions. Stimulation of O2 uptake by epinephrine was blocked by the alpha-adrenergic receptor blocker phentolamine (1 microM) but not by the beta-receptor blocker propranolol. Thus hormones that increase intracellular calcium stimulate O2 uptake predominantly in regions of the liver lobule where O2 tension is lowest, supporting the hypothesis that oxygen tension regulates O2 uptake in the liver via mechanisms involving intracellular free Ca2+.


Pertussis toxin-sensitive and pertussis toxin-insensitive inhibition of parietal cell response to GLP-1 and histamine.

We have recently shown that in rat parietal cells the glucagon-like peptide 1 (GLP-1) variants 7-36 amide, 1-37, and 1-36 amide stimulate H+ production as indirectly measured by [14C]aminopyrine (AP) accumulation. This response to the GLP-1 peptides was intracellularly mediated by activation of adenylate cyclase and by adenosine 3',5'-cyclic monophosphate (cAMP) as second messenger. In the present study, we compared prostaglandin (PG)E2, somatostatin, and the protein kinase A antagonist Rp-adenosine-3',5'-monophosphorothioate (Rp-cAMPS) with respect to their inhibitory effects on parietal cell function induced by GLP-1 or histamine. PGE2 and somatostatin noncompetitively inhibited AP accumulation and cAMP production in response to the GLP-1 variants and histamine (IC50): [mean inhibitory concn 5 x 10(-9) M PGE2; 3 x 10(-7) somatostatin]; at their maximal concentrations PGE2 (10(-7) M) and somatostatin (10(-6) M) caused 85 and 65% inhibition, respectively. Treatment with pertussis toxin (PT; 250 ng/ml; 4 h) reversed the inhibitory effect of PGE2 and somatostatin on AP accumulation and cAMP production. At 2 x 10(-3) M (IC50: 3 x 10(-4) M) Rp-cAMPS completely inhibited AP accumulation induced by the GLP-1 variants or histamine; this effect was insensitive to PT. Specificity of Rp-cAMPs as protein kinase A inhibitor is suggested by inhibition of AP accumulation in response to Sp-cAMPS and N6,O2-dibutyryl adenosine 3',5'-cyclic phosphate sodium, and forskolin, activators of protein kinase A and adenylate cyclase, respectively. We conclude that the parietal cell responses to GLP-1 and histamine are inhibited by identical mechanisms. Effects of PGE2 and somatostatin are mediated by the PT-sensitive subunit of adenylate cyclase Gi, whereas Rp-cAMPS interferes with cAMP-dependent mechanisms that are insensitive to PT.


Peptone stimulates gastrin secretion from the stomach by activating bombesin/GRP and cholinergic neurons.

The mechanism by which partly digested protein (peptone) stimulates gastrin secretion was examined in isolated antral tissues with intact intramural innervation. In the isolated vascularly perfused rat stomach, luminal perfusion with 0.5% peptone increased gastrin (62 +/- 14 pg/min; P less than 0.01) and decreased somatostatin (74 +/- 19; P less than 0.01) secretion. The axonal blocker tetrodotoxin (TTX) abolished the gastrin and somatostatin responses indicating that the responses were neurally mediated. Atropine partly inhibited the gastrin response (50%) and converted the somatostatin response to an increase above basal level. The selective bombesin/gastrin-releasing peptide (GRP) antagonist [Leu13-psi(CH2NH)-Leu14]-bombesin partly inhibited the gastrin response (65%) and caused a further decrease in somatostatin secretion. A combination of atropine and the bombesin/GRP antagonist, like TTX, abolished the gastrin and somatostatin responses. The pattern of response to peptone in superfused antral segments was identical to that in the vascularly perfused stomach. In fundic segments that do not secrete gastrin, the somatostatin response to peptone alone and with various antagonists was identical to that in antral segments. The results indicate that peptone stimulates gastrin secretion by activating stimulatory cholinergic and bombesin/GRP neurons. Cholinergic neurons stimulate gastrin directly as well as indirectly by eliminating the inhibitory paracrine influence of somatostatin.


Nitric oxide as a putative nonadrenergic noncholinergic inhibitory transmitter in the canine pylorus in vivo.

Antropyloroduodenal motility was recorded in seven anesthetized dogs to assess the role of nitric oxide and L-arginine metabolites in nonadrenergic noncholinergic (NANC) mediation of pyloric relaxation. Pyloric activity induced by duodenal field stimulation was inhibited by antral field stimulation and electrical vagal stimulation. Intra-arterial NG-L-arginine-methyl-ester (L-NAME) reduced the inhibition from antral or vagal stimulation (P less than 0.05). Intravenous infusion of L-NAME also blocked the inhibitory effect of vagal and antral stimulation but left the tetrodotoxin-insensitive action of intra-arterial vasoactive intestinal peptide (VIP) and sodium nitroprusside unchanged. L-Arginine reversed the effect of L-NAME whereas D-arginine did not. L-NAME enhanced pyloric contractions to intra-arterial acetylcholine. The NANC inhibition of the substance P-stimulated pyloric response in vitro was blocked by L-NAME and reversed by addition of L-arginine. Sodium nitroprusside was effective as a relaxant in vitro but VIP was not. These data suggest that metabolites of L-arginine mediate neural inhibition of canine pyloric motor activity.


Calcitonin gene-related peptide: mechanisms of modulation of antral endocrine cells and cholinergic neurons.

Actions of human calcitonin-gene related peptide (hCGRP) on acetylcholine (ACh) discharge and gastrin and somatostatin release from rat antral mucosal-submucosal fragments were examined in both dynamic perifusion experiments and short-term static incubation studies. The principal findings of the dynamic perifusion experiments were that hCGRP exerted a dual or biphasic effect on ACh discharge and gastrin release. Initial exposure of antral tissues to hCGRP (1 x 10(-8) M) resulted in stimulation of both ACh and gastrin release that was of brief duration. Continued hCGRP perifusion caused subsequent inhibition of ACh and gastrin release that was substantially greater in duration and magnitude than the initial stimulatory responses. Static incubation studies indicated that hCGRP (10(-10) to 10(-7) M) stimulated somatostatin and inhibited gastrin release in a dose-dependent manner. Inhibition of gastrin and ACh release by hCGRP appeared to be an indirect effect that was mediated by somatostatin as suggested by studies with pertussis toxin (200 ng/ml). Furthermore, studies with atropine (1 x 10(-6) M) and tetrodotoxin (1 x 10(-6) M) indicated that CGRP-induced stimulation of somatostatin release and inhibition of ACh discharge occurred independent of muscarinic receptor activation and nerve excitation. In conclusion, results of these studies indicate that CGRP is capable of exerting both stimulatory and inhibitory effects on ACh release from mucosal-submucosal neurons and gastrin release from antral mucosal G cells in in vitro studies. These data suggest that the inhibitory effects of CGRP on cholinergic discharge and gastrin release are due to the paracrine effects of somatostatin released from antral D cells by direct action of CGRP.


Role of gastrin, histamine, and acetylcholine in the gastric phase of acid secretion in anesthetized rats.

To determine the relative contributions of gastrin, histamine, and cholinergic stimulation to the gastric phase of acid secretion, peptone-stimulated acid output was measured in urethan-anesthetized pylorus-ligated rats after intravenous administration of gastrin monoclonal antibody, cimetidine, and atropine. Intragastric peptone stimulated acid secretion four-fold over basal, which was associated with a significant increase in plasma gastrin levels. Gastrin immunoneutralization and simultaneous H2- and muscarinic-receptor blockade demonstrated that approximately 40% of peptone-stimulated acid output as attributed to endogenous gastrin through a histamine-dependent pathway, whereas 20% of acid output was accounted for by a cholinergic component. Another 10% of titratable acid was omeprazole-insensitive and presumably due to intragastric digestion of peptone. Therefore, approximately 30% residual acid output in response to peptone could not be accounted for by known acid stimulatory mechanisms. In rats given somatostatin monoclonal antibody to block the tonic inhibitory effect of endogenous somatostatin, residual acid output was a similar fraction of meal-stimulated acid output. In contrast, gastric distension induced by intragastric instillation of saline stimulated acid secretion to 1.5-fold over basal. Although 60% of distension-induced acid secretion could be inhibited by either H2 blockade or gastrin immunoneutralization, acid output returned to basal levels after simultaneous muscarinic blockade. These results indicate that gastrin, through a histaminergic pathway, is the principal mediator of meal-stimulated acid secretion in anesthetized rats. Approximately 30% of acid output was due to other unidentified mechanisms, such as chemical secretagogues, a direct effect of amino acids, or novel peptides.