Chronic cocaine alters brain mu opioid receptors. The possibility that dopamine may modulate the expression of opioid receptors was investigated by determining the effects of chronic cocaine administration on the density of mu opioid receptors. Quantitative in vitro autoradiography with the highly selective mu opioid ligand [3H]DAMGO was used to measure and localize changes in mu opioid receptors in the brains of rats administered cocaine or saline three times daily for 14 days. Significant increases in [3H]DAMGO binding were measured in areas of the cingulate cortex, nucleus accumbens, caudate putamen, and basolateral amygdaloid nucleus of the cocaine-treated animals. These results demonstrate that mu opioid receptors undergo upregulation in response to chronic cocaine exposure and suggest that dopamine activity can regulate the expression of mu opioid receptors.


Modulation of nerve membrane sodium channel activation by deltamethrin. Deltamethrin is a highly potent pyrethroid insecticide that causes hypersensitivity, choreoathetosis, tremors, and paralysis in mammals. It is known to modify the sodium channel in such a way as to prolong the tail current associated with step repolarization following a depolarizing pulse. Using the axial-wire voltage-clamp technique with the giant axon of the squid Loligo pealei, we have demonstrated that deltamethrin also greatly slows the opening of the sodium channel. This was first observed as a decrease, by as much as 80%, in the peak sodium current flowing during a short, 10 ms depolarization. Current flowing through these slowly opening deltamethrin modified sodium channels was observed during the first depolarizing pulse after deltamethrin exposure and developed with a time constant of 320 ms. This supports the idea that deltamethrin can modify sodium channels when they are in the closed or resting state. Further, evidence of this hypothesis was provided by experiments using 0.1 and 10 microM deltamethrin and measuring the tail current amplitude after depolarizing pulses of varying duration (1-1200 ms). The mean time constant for the increase in tail current amplitude was almost concentration independent; 253 ms at 0.1 microM and 193 ms at 10 microM. We conclude that deltamethrin modifies the activation kinetics of sodium channels in such a way as to slow opening and that this modification occurs predominantly when channels are in the closed or resting state.


Central tachykinin injection potently suppresses the need-free salt intake of the female rat. Repeated sodium depletions produce a persistent, enhanced need-free salt intake in the rat, particularly in the female. The neurochemical mechanisms underlying the phenomenon are still unknown. The present studies evaluated the effect on the enhanced need-free salt intake of the female rat (1) of pharmacological interference with the natriorexigenic hormones angiotensin II and aldosterone and (2) of the central injection of the tachykinin peptides, which are endowed with antinatriorexic activity. The need-free salt intake of the female rat is not modified by treatment with the angiotensin-converting enzyme inhibitor captopril or by the aldosterone receptor antagonist RU-28318. On the other hand, the behavior is highly sensitive to the inhibitory effect of central tachykinins, suggesting the possibility that need-free salt intake might be linked to modification (down-regulation) of the inhibitory tachykininergic system.


Ageing: a normal degenerative process--with or without regular exercise. The purpose of this symposium was to review the physical, metabolic, psychological, and physiological manifestations of the ageing process. A great deal of research has been carried out in the past 10 years with elderly subjects. Investigations are currently expanding to use invasive methodologies previously applied to healthy young subjects. A most comprehensive account of exercise and the elderly is given by Stamford (1988), with insight into the physical capabilities of some 10% of the Canadian population. Ageing affects the total body and its metabolism including hemodynamics, enzyme activities, strength, muscle contraction, and incidence of disease. Equally important, ageing affects the Canadian economy, geographic demographics of retirees, and ethical consideration of the prolongation of life without quality. The symposium papers analyze the ageing process in humans and animal species from biochemical (Kaldor), physiological (Paterson), mechanical (Vandervoort), and psychological (Brown) viewpoints.


Spectrophotometric and kinetic studies on normal and thyrotoxic cardiac myosins. Kinetic and microcalorimetric methods were used to study the interaction of thyrotoxic (V1) and normal adult cardiac myosin (V3) with ATP. It was shown that the overall enthalpy and entropy change was larger in the interaction of ATP with the thyrotoxic than with the normal myosin. There is evidence that the observed enthalpy changes were generated by protein conformational changes connected to the sequential destabilization and restabilization of the prevalent, energetically favored conformation of the myosin molecule. The V1-ATP interaction created more entropy than the V3-ATP interaction, and also the thyrotoxic isomyosin required more activation energy than the normal protein to attain the activated state. All of these results indicated that the catalytic activity of the V1 myosin was thermodynamically less efficient than that of the V3 isoenzyme.


Effects of ageing on the cardiorespiratory system. Ageing is characterized by a loss of functional capacity. A linear loss from age 30 to 65 years has been described for various measures. After age 65 years, evidence also suggests an accelerated decline in some measures, particularly those of muscle size and function. Nevertheless, up to 85 years, the rate of decline of VO2max across ages appears to remain rather constant. As well, the central cardiovascular capacity, despite some suggested age effects, would appear to be adequate to support the smaller muscle mass. Thus, despite the losses in absolute exercise capacity with age, the ability to sustain a relatively high intensity of aerobic exercise appears preserved. It has also been shown that cardiorespiratory training of older men and women is effective in increasing VO2max and results in the usual changes during submaximal exercise. These benefits of cardiorespiratory training are substantial in that the increases observed compensate for the losses with ageing described over the period of a decade or more.


Effects of ageing on human neuromuscular function: implications for exercise. This review focuses on the age related declines in muscle mass and strength, including a discussion of the potential for improvement of neuromuscular function following exercise training programmes. In the literature, limb muscles have been compared between groups of men and women throughout the adult age range, showing that decreases in voluntary strength do not become apparent until after the age of about 60. Rate of decline then amounts to 10 to 15% per decade, stemming from age related decreases in the amount of excitable muscle tissue. However, high-resistance exercise training programmes are effective in improving both muscle size and voluntary strength, even in very old and frail men and women. These improvements may yield significant gains in the performance of sports and the activities of daily living, such as walking ability. Maintenance exercise programmes must also be advocated to avoid rapid detraining effects seen in elderly people who become sedentary.


Physical activity, ageing, and psychological well-being: an overview of the research. Research has shown that physical activity leads to greater psychological well-being for many young and middle-aged individuals. This paper reviews studies to determine whether there is also evidence for an association between physical activity and psychological well-being across the life span, or in the elderly. Several methodological problems related to exercise, ageing, and psychological well-being are explored. Although an association between exercise and mental well-being in the elderly is noted, there is little experimental evidence to support a conclusion that physical activity is causally related to enhanced psychological well-being in the elderly. Methodological problems, a paucity of well-controlled studies, and equivocal findings regarding the topic make it premature to conclude that physical activity leads to enhanced mental health in elderly persons.


Fitness testing and counselling in health promotion. For the past 15 years the University of Ottawa has conducted on-site fitness assessments of over 5,000 federal public servants. The testing sessions and accompanying counselling session are conducted within a framework of health promotion to encourage managers to adopt a healthy lifestyle. The data collected on this population are quite unique since the managers represent a cross-section from across Canada, and it is an important source of information regarding associations among fitness, lifestyle, and health characteristics. The assessment includes a lifestyle and stress questionnaire, a 12-hour fasting lipid profile, determination of resting and exercise heart rate and blood pressure, body composition, upper body strength and muscular endurance, flexibility, pulmonary function, and aerobic power (Canadian Aerobic Fitness Test). Results are presented in a computerized format and interpreted during the course of a debriefing session; an exercise prescription is also provided. The sessions foster awareness, influence attitudes, and identify health behaviour alternatives. Not only can testing be used as a diagnostic and intervention procedure but it also serves as an excellent education and motivational tool that could be integrated in a routine medical examination.


Development and validation of a fitness screening protocol for firefighter applicants. It is imperative that fitness screening protocols for firefighter applicants embody the specific physical requirements of fighting fires. Based on the physiological characterization of experienced firefighters performing essential tasks, a test battery was developed that includes both job related performance tests and a combination of health related and performance related laboratory tests of physical fitness. Fifty-three firefighters with an average of 5.4 years of experience completed the battery and provided Likert scale comparisons of the tests with actual firefighting operations for criterion validation. "Acceptable" through "maximum" times were established for the job related performance tests, and "minimum" through "optimum" standards were developed for the physical fitness tests. Guidelines for the medical screening of firefighter applicants are also described. In addition, an overall scoring procedure was formulated for the ranking of acceptable applicants.


Characterization of the physical demands of firefighting. To characterize the physical demands associated with on-the-job use of current firefighting equipment and the performance of essential firefighting operations, an initial task analysis of all firefighting operations was followed by an in-depth physical and physiological characterization of those tasks deemed to be physically demanding. The most commonly encountered applications of strength and endurance were lifting and carrying objects (up to 80 lbs), pulling objects (up to 135 lbs), and working with objects in front of the body (up to 125 lbs). The most demanding firefighting operations required a mean VO2 of 41.5 ml/kg.min-1 with peak lactate concentrations of 6 to 13.2 mM. Ninety percent of the demanding firefighting operations that were studied required a mean VO2 of 23 ml/kg.min-1. These aerobic energy requirements corresponded to 85 and 50% VO2max, respectively. Therefore a minimum VO2max standard for firefighter applicants of 45 ml/kg.min-1 is recommended.


Development of physical fitness standards for Canadian Armed Forces younger personnel. The purpose of this study was to develop minimum physical fitness standards based on common task criteria for personnel younger than 35 years of age in the Canadian Armed Forces. A random sample of 66 men and 144 women performed the Exercise Prescription (EXPRES) test and five physically demanding tasks that simulated common military tasks. Common cutting scores were selected as the point at which 75% of the total weighted sample passed each task. Since there were significant differences between the sexes in task performance and technique execution, the groups were analyzed separately. Results indicated a range in variance of 14 to 48% between military task performance and physical fitness test score, thus suggesting that fitness measures are a poor predictor of task performance. Because of the low variance observed the passing group of each task was treated as a representative sample of subjects whose fitness profiles were indicative of those able to meet task criteria. The 5th percentile fitness scores of the passing group were proposed as the minimum fitness standard. These conditions resulted in fitness standards that were physically more demanding for women than for men.


Development of fitness screening protocols for physically demanding occupations. The fitness screening of applications to physically demand occupations has become increasingly complex with the advent of Equal Opportunity legislation and Affirmative Action initiatives. A comprehensive study was conducted of the physical demands and associated physiological responses of workers in strenuous jobs at a large multifaceted natural gas company. Based on these findings, an applicant fitness screening protocol was developed and validated. The protocol was implemented for an 18-month period, after which the results were examined with regard to gender differences in success rates and other influencing factors. Because of ongoing changes in attitudes and practices in the workplace, fitness screening for applicants was interrupted and a review was initiated of all strenuous jobs with the aim of reducing their physical demands.


Police Officer's Physical Abilities Test compared to measures of physical fitness. This study compared the Police Officer's Physical Abilities Test (POPAT) with selected field and laboratory tests of physical fitness. Ninety-eight volunteer police officers (73 men, 25 women) completed all aspects of the testing. Fifty-five (55)% of the total group passed the POPAT by bettering the 4 min 15 sec "cut" point. Only 16% of the women and 68% of the men passed the overall test. Laboratory tests revealed a rather unfit sample of subjects (mean VO2 max = 42.6 ml.kg.min-1; % body fat = 22.9). Stepwise multiple regression indicated that 55% of the variance on the run component of the test was accounted for by maximal aerobic power and anaerobic capacity. The fight component of POPAT did not correlate highly with standard field tests of strength. Pass/fail aspects of the test were not clearly delineated by selected lab and field tests. It was concluded that POPAT, being a valid, task-specific, job related test, consists of motor abilities and technique as much as generalized fitness parameters.


Periodisation of training stress--a review. Athletic performance improves as the athlete adapts to progressively increasing training loads. Empirical observations and studies investigating fluctuations in performance indicate that this adaptation occurs during periods of reduced training, termed regeneration periods. Thus it is essential that adequate regeneration time be included in training programmes so that adaptation can be achieved. In order to induce adaptation, heavy periods of training are used to provide a stimulus for adaptive processes to become functional. The literature and anecdotal accounts suggest that the cycling of light, medium, and heavy periods of training is an optimal method for combining the heavy periods of training with the periods of light training needed to allow adaptation and supercompensation.


Periodisation and the prevention of overtraining. It may be essential for the athlete to train in cycles in order to induce optimal improvements and prevent overtraining. Without sufficient recovery time, adaptation may not occur and the athlete may develop the symptoms of overtraining due to continuous and/or excessive exposure to training stress. Training in cycles provides guidelines for the times in the training programme when regeneration should be complete, and therefore the times when the athlete can be screened for overtraining without confusing the fatigue of overload training with that of overtraining. A periodised training structure provides guidelines for conducting research into the mechanisms of training adaptation and overtraining.


Transforming growth factor-alpha in human hepatocellular carcinoma and coexpression with hepatitis B surface antigen in adjacent liver. Hepatitis B virus (HBV) infection is closely associated with the development of hepatocellular carcinoma (HCC) in many patients, but the mechanisms by which HBV contributes to HCC are not known. Transforming growth factor-alpha (TGF-alpha), a regulator of growth and regeneration in rat liver that can be found in high levels in some human cancers, theoretically could play such an intermediate role in the development of HCC. The expression of TGF-alpha and its relation to the HBV antigens were evaluated in human HCC and adjacent nontumorous livers from 33 patients from the United States and China using immunoperoxidase staining of paraffin-embedded sections. TGF-alpha was detected in HCC from 27 of 33 (82%) patients; the frequencies were similar in patients from the United States and China. TGF-alpha was detected in HCC more frequently in patients whose adjacent nontumorous livers had detectable hepatitis B surface antigen (HBsAg) and/or hepatitis B core antigen (HBcAg) than in those whose adjacent livers lacked HBsAg and HBcAg. Detection of TGF-alpha was not affected by tumor size, histologic type, or grade. TGF-alpha was detected in adjacent nontumorous livers from 31 of 33 patients (94%). Coexpression at a high intensity of TGF-alpha and HBsAg in the same hepatocytes could be demonstrated by specific staining of consecutively cut sections for 17 of 33 patients (52%). TGF-alpha is expressed at a high level in 82% of human HCC. Localization of HBsAg within the same hepatocytes as TGF-alpha suggests a possible interaction between HBV and TGF-alpha during hepatocarcinogenesis in humans. Stimulation of TGF-alpha expression could be part of a chain of events by which HBV contributes to the development of HCC in some patients.


Expression of HLA-DR in intrahepatic cholangiocarcinoma. Expression of HLA-DR was investigated immunohistochemically in 20 cases of intrahepatic cholangiocarcinoma, as well as 9 normal livers that had been obtained at autopsy. Five of the nine normal livers had peribiliary glands that showed HLA-DR. Positive staining for HLA-DR on tumor cells was observed in 6 of 13 cases of cholangiocarcinoma of the hepatic hilus and in only 1 of 7 cases of peripheral cholangiocellular carcinoma. In cholangiocarcinomas of the hepatic hilus, the 5-year survival rate for tumors that had positive staining for HLA-DR was better than that for tumors that had negative staining. Based on these results, the presence or absence of HLA-DR on tumor cells could have a pathologic significance for intrahepatic cholangiocarcinoma.


Flow cytometric analysis of nuclear DNA content of duct cell carcinoma of the pancreas. This study was designed to evaluate the efficacy of nuclear DNA content analysis in determining the prognosis of carcinoma of the pancreas. Resected and paraffin-embedded specimens from 72 patients with duct cell carcinoma of the pancreas were examined, and flow cytometry was used to explore the relationship between DNA ploidy and TNM classification or histologic grade. DNA aneuploidy was found histologically in 42.9%, 56.8%, and 71.4% of Grade 1, 2, and 3 tumors, respectively. DNA ploidy showed a statistically significant correlation with T category and retroperitoneal invasion. The cumulative survival rate of patients with retroperitoneal invasion was shorter than that of those without retroperitoneal invasion. Among the patients with retroperitoneal invasion, those with DNA aneuploidy had a significantly shorter survival time than did those with DNA diploidy. The DNA ploidy pattern, in combination with the presence or absence of retroperitoneal invasion, appears to be useful in predicting the prognosis for duct cell adenocarcinoma of the pancreas.


Ultrasonography and ultrasonographically guided fine-needle aspiration biopsy of impalpable cervical lymph nodes in patients with non-small cell lung cancer. Neck ultrasonography (US) and ultrasonographically guided fine-needle aspiration (UGFNA) cytologic examination were used in the staging work-up of 51 consecutive patients with non-small cell lung cancer (NSCLC) with clinically impalpable cervical lymphadenopathy. All patients had chest radiography, fiberoptic bronchoscopic examination, computed tomography (CT) scans of the chest and abdomen, bone scan, and US of the abdomen and the neck. Six patients (12%) were found to have cervical lymph nodes by neck US. The cytologic examination of the lymph node aspirates obtained from UGFNA confirmed the metastasis. The clinical stage thus was changed from Stage IIIa to Stage IIIb in one patient. In the other five patients, UGFNA of cervical lymph nodes prevented more invasive diagnostic procedures. This preliminary study shows that neck US and UGFNA of lymph nodes may be helpful in the pretreatment evaluation of patients with NSCLC with impalpable cervical lymph nodes. However, additional study is needed to confirm this result.


Unusual relapse of hepatocellular carcinoma. The authors report a patient with iatrogenic dissemination of hepatocellular carcinoma (HCC). A 65-year-old Caucasian man was found to have a moderately well-differentiated HCC diagnosed by laparoscopy and biopsy; the patient had atypical left liver lobe resection. Thirty-three months after definitive surgery a double relapse was found at the site of the previous laparoscopy and at the surgical scar; no other metastases were found. Surgical procedure for removal of these lesions was performed, and the patient received complementary radiation therapy. At 30 months of follow-up, the patient is alive and disease free. The risk of neoplastic seeding through biopsy and improved safety in surgical techniques justify the omission of diagnostic biopsy in patients who have surgical procedures.


The morphologic transition in hepatocellular carcinoma. A comparison of the individual histologic features disclosed by ultrasound-guided fine-needle biopsy with those of autopsy. In hepatocellular carcinoma (HCC), it has been strongly suggested that as the tumor increases in size, foci of less-differentiated malignant tissues arise in the well-differentiated tumor and increase until they replace the well-differentiated tumor tissues. It has also been suggested that tumor growth is attributed to such dedifferentiation. In the current study, the individual histologic features of ultrasound-guided fine-needle biopsy specimens, taken from 12 small HCC in an early stage, were compared with those of autopsy to confirm the dedifferentiation of HCC. In all 12 cases, autopsy was performed 6 months or more after the initial biopsy. Dedifferentiation was demonstrated in 9 of 12 cases (75.0%). Most of the biopsy specimens from minute HCC were well-differentiated. All tumors that had been well-differentiated when evaluated by biopsy were found to have become moderately differentiated at autopsy. This comparative study confirmed the dedifferentiation of HCC with tumor growth.


Small cell carcinoma of the ampulla of vater. Primary small cell carcinoma of the periampullary region is rare. Only four patients have been reported previously. The authors examined a patient with ampullary small cell carcinoma using a range of immunocytochemical stains. This tumor shows morphologic and neuroendocrine features similar to its pulmonary and extrapulmonary counterparts.


Small cell undifferentiated carcinoma of the pancreas. Report of a patient with tumor marker studies. Small cell undifferentiated carcinoma of the pancreas is a rare type of pancreatic neoplasm. The authors report the clinical and pathologic aspects of a patient with this malignant lesion and an extensive serologic and immunohistochemical survey of potential ectopically produced hormones or tumor markers. Neuron-specific enolase (NSE) emerged as a tumor marker. NSE could be valuable in the diagnosis and management of other patients with this rare disease.


Immunohistochemical study of the human 17 beta-hydroxysteroid dehydrogenase and steroid receptors in endometrial adenocarcinoma. 17 beta-Hydroxysteroid dehydrogenase (17HSD) and estrogen (ER) and progestin (PR) receptors were analyzed immunohistochemically in tissue specimens of 66 patients with endometrial adenocarcinoma. Plasma steroid concentrations were correlated to immunohistochemical data. 17HSD was detected in 48% of the specimens and was stained in the cytoplasm of epithelial cells. The tissues were characterized by a heterogeneous staining pattern for 17HSD. In some patients, intensively stained epithelial cell clusters were seen, indicating that local factors were responsible for the expression of the protein. Poorly differentiated adenocarcinoma specimens tended to have no 17HSD more frequently than did well or moderately differentiated tissues. ER and PR were detectable in 24% and 28% of patients, respectively, and were localized in the nuclei of epithelial and stromal cells. There was a significant correlation between 17HSD and PR staining and an inverse correlation between plasma progesterone concentrations and 17HSD staining. This is contrary to the data obtained with normal endometrium. The main reason for this inverse relation between endometrial 17HSD staining and plasma progesterone concentrations was that, in some postmenopausal patients with low plasma progesterone concentrations, intense staining for 17HSD was detectable in the endometrial carcinoma specimens. This indicates a major difference in the regulation of 17HSD expression in endometrial adenocarcinomas, compared with normal tissues of premenopausal women.


Primary intracranial germ cell tumor. Pathologic study of 51 patients. Fifty-one primary intracranial germ cell tumors (GCT), including germinoma, teratoma, endodermal sinus tumor, choriocarcinoma and mixed GCT, were studied. The incidence of GCT in the surgically removed intracranial neoplasms was 11.1% for pediatric patients and 0.6% for adult patients. The age/sex of the patients and the location of the tumors were analyzed. Morphologic findings of these tumors were identical to that of their gonadal counterparts. Immunohistochemical studies showed that alpha-fetoprotein (alpha-AFP), human chorionic gonadotropin (HCG), and placental alkaline phosphatase (PLAP) were helpful, whereas carcinoembryonic antigen (CEA) and cytokeratin (CKER) were of little help in determining the diagnosis. Serum tumor markers, alpha-AFP and HCG, were helpful in recognizing GCT producing them. However, they could not be used for specific diagnosis because different tumors could have similar serum levels. Histopathologic study was handicapped by the small size of most specimens (which usually could not include all of the components if the tumor was a mixed GCT), but it was the only means for specific diagnosis.


Epstein-Barr virus in acquired immune deficiency syndrome (AIDS) and non-AIDS primary central nervous system lymphoma. Primary central nervous system lymphoma (PCNSL) still occurs mainly in patients who are immunosuppressed, but its incidence is rising dramatically among immunocompetent individuals. The Epstein-Barr virus (EBV) has been detected by in situ hybridization in PCNSL tumor tissue from patients who are immunodeficient, but not from patients who are immunocompetent. Using the more sensitive polymerase chain reaction (PCR) technique, the authors analyzed PCNSL tissue from 13 patients with acquired immune deficiency syndrome (AIDS) and 13 patients who were immunocompetent for the presence of EBV genome. DNA was extracted from paraffin-embedded biopsy or autopsy specimens. PCR was run using primers for EBV (from the first internal repeat segment of the EBV genome), and identical samples were run simultaneously with primers against actin or the p53 gene as controls to establish the presence of DNA in the sample. Reaction products were also Southern blotted to confirm EBV specificity. EBV was detected in the tumor tissue of 11 of 13 patients (85%) with AIDS and of 7 of 13 patients (54%) who did not have AIDS. There was a history of illness that might suggest or predispose to immune compromise in 5 of 13 patients without AIDS; however, prior illness did not predict EBV-positive tumors. Although mechanisms remain to be clarified, EBV was present in a high percentage of patients with AIDS-related PCNSL and non-AIDS-related PCNSL:


Use of proton nuclear magnetic resonance spectroscopy of plasma in screening for malignant disease. In 1986, narrow plasma proton nuclear magnetic resonance (NMR) methyl and methylene line widths were reported to be associated with malignant disease, but more recent studies have not confirmed this relationship. The authors analyzed 106 plasma samples from healthy control subjects and patients with cirrhosis, hepatocellular carcinoma, metastatic liver tumors, other untreated cancers, and hyperlipidemia. NMR spectroscopy was performed using a proton NMR spectrometer operating at 399.65 MHz. A significant difference was found between the mean line widths of the plasma methyl resonances in control subjects and those in patients with cancer or hyperlipidemia. However, no significant difference was found between the mean plasma methylene line widths in control subjects and patients with cancer. Plasma samples from patients with liver disease or hyperlipidemia showed a characteristic methylene spectral pattern. The methylene pattern could be separated into three types: type A had a small peak on the right shoulder of the main peak; type B was a sharp single peak; and type C was a broad single peak. All control subjects had type A pattern; patients with liver disease had type C pattern; and patients who had hyperlipidemia had type B pattern, and hyperlipidemia may affect methyl and methylene line widths in NMR spectra. Because the methyl and methylene levels and their average line widths correlated inversely with triglyceride levels, considering the spectral patterns that indicate hyperlipidemia should decrease false-positive results and make the methyl line width useful for cancer screening.


Neuroblastoma. Effect of genetic factors on prognosis and treatment. BACKGROUND AND METHODS. Genetic analysis of tumor tissue has provided considerable insight into mechanisms of malignant transformation and progression. Neuroblastomas have been studied by cytogenetics, flow cytometry, and molecular genetic techniques, and these studies have identified several specific abnormalities that allow subclassification of these tumors into genetic/clinical subtypes. RESULTS AND DISCUSSION. Four genetic abnormalities have been identified that are characteristic of certain neuroblastomas. These include: (1) loss of heterozygosity (LOH) for the short arm of chromosome 1, including band 1p36; (2) amplification of the N-myc protooncogene; (3) hyperdiploidy, or near triploidy; and (4) defects in expression or function of the nerve growth factor receptor (NGFR). Abnormalities of the NGFR are found in virtually all neuroblastoma cell lines, and some primary tumors. The latter have not been studied extensively. Hyperdiploidy is associated with lower stages of disease and with a favorable outcome in infants. LOH for chromomors. The latter have not been studied extensively. Hyperdiploidy is associated with lower stages of disease and with a favorable outcome in infants. LOH for chromosome 1, band p36, and N-myc amplification are more common in patients older than 1 year of age with advanced stages of disease. The latter two genetic abnormalities may be related, and LOH for 1p36 may precede the development of amplification. When these abnormalities are combined with assessment of DNA content, three distinct genetic subsets of neuroblastomas can be identified. The first is characterized by a hyperdiploid or near-triploid modal karyotype, with few if any cytogenetic rearrangements. These patients generally are younger than 1 year of age with localized disease and a good prognosis. The second has a near-diploid karyotype, with no consistent abnormality identified currently. These patients generally are older with more advanced stages of disease that progress slowly and are often fatal. The third group has a near-diploid or tetraploid karyotype, with deletions or LOH for 1p36, amplification of N-myc, or both. These patients generally are older with advanced stages of disease that rapidly are progressive. Thus, genetic analysis of neuroblastoma cells provides information that has prognostic significance and can direct a more appropriate choice of treatment.


Population genetics of colonic cancer.

Population genetics of colonic cancer. There are several well known but rare syndromes of inherited colonic cancer. Genetic epidemiologic studies also have demonstrated that relatives of individuals with colonic cancer in general exhibit an excess risk for this malignancy. This report reviews the literature pertinent to genetic and familial risk for colonic cancer with emphasis on the recent work that suggests that inherited susceptibility to colonic neoplasms is common. The adenomatous polyposis syndromes are rare inherited colonic cancer conditions caused by a mutant gene which recently has been characterized. Hereditary nonpolyposis colorectal cancer is likewise inherited and may account for up to 5% of cases. The molecular genetics of this disease remain to be clarified. The majority of colonic cancer cases are considered sporadic but are known to often cluster in families. Recent work suggests that inherited susceptibility may be the basis of this familial occurrence. Screening strategies based on inherited and familial risk are suggested. Knowledge of the familial and inherited risk for colonic cancer is leading to a better understanding of this disease and is suggesting more directed preventive strategies.


Accumulation of genetic defects during astrocytoma progression. The development of human cancer generally is thought to entail a series of events that cause a progressively more malignant phenotype. This hypothesis predicts that tumor cells of the ultimate stage will carry each of the events; cells of the penultimate stage will carry each of the events minus the last one; and so on. Therefore, a dissection of the pathway from a normal cell to a fully malignant tumor may be viewed as the unraveling of a nested set of aberrations. In experiments designed to elucidate these events, genotypic combinations were compared at genomic loci defined by restriction endonuclease recognition-site variations in normal and tumor tissues from patients with various forms and stages of cancer. The first step, inherited predisposition, is described best for retinoblastomas in which a recessive mutation of a locus residing in the 13q14 region of the genome is unmasked by aberrant, but specific, mitotic chromosomal segregation. A similar mechanism involving the distal short arm of chromosome 17 is apparent in astrocytic tumors, and the event is shared by cells in each malignant stage. This is distinct from a loss of heterozygosity for loci on chromosome 10, which is restricted to the ultimate stage, glioblastoma multiforme. In addition, this approach has been extended to include a wide variety of human cancers, and generally it is applicable. These results identify a genetic approach to defining degrees of tumor progression and a means for determining the genomic locations of genes involved in the pathway as a prelude to their molecular isolation and characterization. They provide a molecular genetic-based oncology with clinical utility in differential pathologic findings, in disease groupings for therapeutic purposes, and in prenatal identification of latent disease carriers.


Viral role in cervical and liver cancer. Hepatitis B virus (HBV) and certain human papillomaviruses (HPV) have been suspected of playing a role in human malignancy. The long latency period (decades) and the fact that only a small proportion of infected individuals subsequently have cancer suggests that these viruses probably contribute in an indirect manner to the development of cancer. Analysis of viral gene function at the molecular level supports this conclusion. More studies are needed to understand the subtle interactions between host cell and viral functions and the genetic and environmental factors that may contribute to malignancy.


The influence of TNF on the membrane fluidity of tumor cells. Possible TNF (tumor necrosis factor) effects on the membrane fluidity of tumor cells were investigated. Viable tumor cells, TNF sensitive, were obtained from the ascitic form of the SA-1 tumor bearing mice. The influence of in vitro and in vivo treatment of cells with the TNF analog was investigated by EPR (electron paramagnetic resonance). SA-1 cells were spin labeled with the methylester of 5-doxylpalmitate, which primarily dissolves in the membranes. The maximal hyperfine splitting was determined and the empirical correlation time calculated. The results show that TNF significantly decreases the correlation time, i.e. it increases the fluidity of SA-1 cell membranes. Such alteration could contribute to the cytotoxicity of TNF.


Polyclonal and monoclonal antibodies monospecific to MMTV LTR orf protein produced in E. coli. Monoclonal and polyclonal antibodies specific to an open reading frame of the mouse mammary tumor virus long terminal repeat were generated using an open reading frame-beta-galactosidase fusion protein produced in E. coli. Both antibodies reacted with the open reading frame-beta-galactosidase fusion protein but not with beta-galactosidase alone using an immunoblotting technique. It is concluded that these antibodies were specific for the protein encoded by the open reading frame of the mouse mammary tumor virus long terminal repeat.


Antitumoral activity of bombesin analogues on small cell lung cancer xenografts: relationship with bombesin receptor expression. Gastrin releasing peptide (GRP), the human homologue of bombesin (BN), is an autocrine growth factor for small cell lung cancer (SCLC) cells. The synthetic octapeptides [D-cpa1-beta-Leu8-des-Met9]litorin (BIM 26182) and [D-Phe6-Leu13-CH2NH-Cpa14]bombesin(6-14)NH2 (BIM 26189) are potent GRP/BN antagonists of the proliferation of 3T3 and rat pancreas cells. The effect of these analogues on the proliferation of four SCLC cell lines (SCLC 6, SCLC 41, SCLC 75, SCLC 74R) was tested in vitro and in vivo. Two of these SCLC lines (SCLC 41M and SCLC 75) had receptors for BN/GRP and expressed the prepro-GRP mRNA. BIM 26182 and BIM 26189 inhibited [3H]thymidine incorporation into the DNA of SCLC 41 cells, stimulated [3H]thymidine incorporation in SCLC 6, and had no effect on the two other cell lines. The SCLC implanted s.c. in nude mice were treated with either BIM 26182 or BIM 26189. BIM 26182 and BIM 26189 injected at the doses of 50 micrograms twice a day (s.c.) around the tumor for 10 to 21 days delayed the growth of SCLC 41 and of SCLC 75. The maximal effect was observed during the treatment period, after which the tumors regrew, suggesting a cytostatic effect of these peptides. No inhibitory effect of the peptides on SCLC 74R or SCLC 6 growth was observed. These data suggest that GRP antagonists are able to inhibit the in vitro and in vivo growth of BN/GRP receptors-positive SCLC.


Suppression of melanoma cell motility factor receptor expression by retinoic acid. beta-All-trans-retinoic acid (RA) has been shown to inhibit the growth, enhance the differentiation, and suppress the transformed and metastatic properties of certain human and murine melanoma cells. This study examined the effect of RA on the level of a cell surface receptor (M(r) 78,000) (gp78) for an autocrine motility factor, which has been implicated in invasion and metastasis. Treatment of murine melanoma cell lines S91-C2, B16-F1, and K1735-P with RA (10 microM) for 5 days decreased the level of gp78 by 37, 72, and 92%, respectively, as revealed by immunoblotting with monoclonal antibodies raised against gp78. In contrast, RA had only a limited effect on gp78 levels in melanoma cell clones or variant cell lines that are resistant to the growth-inhibitory effects of RA (S91-C154, B16-F10, and K1735-Cl19). Further studies with K1735-P, the most sensitive cell line with respect to modulation of gp78, showed that the decrease in gp78 level required at least 1 microM RA and 4 to 5 days of treatment. The binding of anti-gp78 antibodies to the surface of intact RA-treated cells and to intracellular gp78 in permeabilized cells was also lower than in untreated cells. Furthermore, RA treatment decreased the induction of cell motility, on colloidal gold-coated glass coverslips, by anti-gp78 antibodies, which mimic the effect of autocrine motility factor. The RA-induced decrease in antibody-enhanced cell motility was similar to the time- and RA concentration-dependent decrease in the amount of gp78, suggesting that the two events are related. These results raise the possibility that the previously reported suppression by RA of tumor cell invasion and metastasis may be related, at least in part, to suppression of cell motility resulting from the decreased level of the autocrine motility factor receptor.


Expression of matrix metalloproteinase genes in transformed rat cell lines of high and low metastatic potential. The enzymes that comprise the family of matrix metalloproteinases (MMPs) share the capacity to degrade extracellular matrix components. A large body of evidence indicates that certain members of this metalloproteinase gene family play critical roles in determining the malignant phenotype of solid tumors. We previously have derived transformed cell lines with vastly different metastatic potentials by transfecting different combinations of oncogenes into primary rat embryo cells. Conditioned medium from those cell lines was assayed by Western blot analysis for the production of four separate matrix metalloproteinases to see whether a correlation could be found between protease expression and the metastatic phenotype. The transformed rat embryo cell lines with high metastatic potential were found to produce high levels of the stromelysin 1 (MMP-3) and stromelysin 2 (MMP-10) proteases, while the nonmetastatic lines produced low or undetectable levels of these two enzymes. No correlation was seen between the metastatic phenotype of the cell lines and the level of expression of two other matrix metalloproteinases, the M(r) 72,000 type IV collagenase (MMP-2) and the M(r) 92,000 gelatinase (MMP-9). These data suggest that the differential regulation of the stromelysin proteases may contribute to the difference seen in the metastatic potential of these cell lines.


gamma-Interferon increases metaiodobenzylguanidine incorporation and retention in human neuroblastoma cells. Iodine-labeled m-iodobenzylguanidine (MIBG) is a widely used radiopharmaceutical for both diagnosis and biologically targeted radiotherapy of neuroblastoma. However, resistance to the radiotherapeutic effects of MIBG is often encountered, mainly due to lack of MIBG accumulation by neoplastic cells. We have investigated whether the induction of neuroblastoma cell differentiation modifies MIBG incorporation and retention. LAN-5 cells were selected, due to their moderate ability to take up MIBG. Treatment of these cells with gamma-interferon (IFN-gamma) resulted in morphological changes accompanied by a significant increase in overall cell-associated MIBG. Desimipramine, but not reserpine, easily depleted IFN-gamma-treated LAN-5 cells of their MIBG content. This suggests that the mechanism involved is an uptake enhancement rather than an improved storage ability. Indeed, IFN-gamma induces de nov synthesis of MIBG receptor-transporters, as demonstrated by polymerase chain reaction amplification and semiquantitative analysis. Our results suggest that pretreating neuroblastoma patients with IFN-gamma before MIBG administration may enhance the efficacy of both biologically targeted radioimaging and therapy of this tumor.


Antiproliferative effects of the somatostatin analogue octreotide (SMS 201-995) on ZR-75-1 human breast cancer cells in vivo and in vitro. The somatostatin analogue octreotide (SMS 201-995) inhibits secretion and growth of certain tumor cells, and current efforts are directed toward the elucidation of its mode of antiproliferative action. In this study, the effect of octreotide on the growth of ZR-75-1 human breast cancer cells has been characterized in immunodeficient nude mice and in cell culture. These results have been related to the expression of somatostatin receptors in vivo and in vitro. Continuous infusion of 10 micrograms/kg/h of octreotide yielded plasma levels of 5.7 ng/ml and elicited highly significant growth inhibitory effects on solid ZR-75-1 breast tumors in nude mice. After 2 and 4 weeks of treatment, tumor volumes in the octreotide group were 39.1 and 36.7% of those of control animals treated with vehicle, respectively. Autoradiographic studies demonstrated that 8 of 12 ZR-75-1 tumors studied were somatostatin receptor positive. When ZR-75-1 tumor cells were exposed in vitro to nanomolar concentrations of octreotide, a dose-dependent inhibition of cell growth was observed in the presence of 5% fetal calf serum or under serum-free conditions using epidermal growth factor, insulin-like growth factor type I, or insulin as growth stimulus. In parallel receptor-binding experiments, ZR-75-1 cells were shown to express specific high-affinity somatostatin receptors (Kd value = 0.9 nM, Bmax = 6000 sites/cell). From these experiments, we conclude that octreotide is a powerful inhibitor of ZR-75-1 tumor cell growth in nude mice and in culture. This inhibitory action of octreotide and the presence of somatostatin receptors on ZR-75-1 tumor cells in vitro and in vivo suggest a direct, somatostatin receptor-mediated effect of octreotide.


Oncogenic human papillomavirus type 16 is associated with squamous cell cancer of the male urethra. Human papillomaviruses (HPV), especially genotypes 16 and 18, are probable effectors of human urogenital malignancies. Although the male urethra is a proposed reservoir of HPV transmission, the association between HPV and squamous cell carcinoma (SCC) of the male urethra has not been studied. The highly sensitive technique of polymerase chain reaction with type-specific HPV 16 and 18 primers and general primers, including nine other genotypes was used to survey a series of SCC of the male urethra for the prevalence of an association with HPV. Archival surgical specimens from 14 patients were analyzed, and primary, recurrent, and metastatic lesions from 4 (29%) patients contained HPV 16 DNA. No other HPV genotype (6b, 11, 13, 18, 30, 31, 33, 35, 45, 51) was detected. Complete concordance for the presence of HPV in primary and recurrent or metastatic disease was demonstrated. These findings strongly suggest that HPV type 16 is associated with a substantial subset of SCCs of the male urethra. Analysis of clinical data revealed that HPV-positive tumors had a significant predilection for location in the pendulous urethra versus the bulbar urethra. Survival data analysis showed that the presence of HPV more closely correlated with prolonged survival than did tumor location. The presence or absence of HPV 16 DNA defines two subsets of SCC of the male urethra which differ in the site of occurrence and, possibly, progression.


A novel gene (HIP) activated in human primary liver cancer. Differential screening of a human hepatocellular carcinoma complementary DNA library using subtracted probes allowed us to identify a novel gene named HIP whose expression at the transcriptional level was elevated in liver tumors. The protein potentially encoded by the complementary DNA showed 68.5% identity with the bovine pancreatic thread protein and 49% identity with the human reg protein, which has been proposed as a pancreatic islet cell regenerating factor and is identical to the pancreatic stone or pancreatic thread protein. Sequence analysis suggests that the bovine pancreatic thread protein encoding gene is, in fact, the bovine homologue of the HIP gene. Furthermore, data base searches revealed a significant similarity of the HIP and pancreatic stone protein/pancreatic thread protein/reg sequences with the C-type lectin superfamily. The HIP sequence, like pancreatic stone protein/pancreatic thread protein/reg protein, consists of a single carbohydrate recognition domain linked to a signal peptide which would be involved in secretion of the protein. HIP mRNA was expressed at a high level in the tumors of seven of 29 hepatocellular carcinomas. In contrast, HIP mRNA was not detected in nontumorous adjacent areas or in normal adult and fetal liver, suggesting that HIP could be involved in liver cell proliferation or differentiation. HIP mRNA expression is tissue specific, since it is present in the normal small intestine and pancreas, while it could not be evidenced in colon, brain, kidney, or lung. In summary, our results show the existence of a novel family within the superfamily of C-type lectin which may be involved in liver, pancreatic, and intestinal cell proliferation or differentiation.


Release of ATP from human erythrocytes in response to a brief period of hypoxia and hypercapnia. The aims were, first, to detect and quantify the release of ATP from human erythrocytes in response to a brief exposure to a hypoxic/hypercapnic environment, similar to that found in vigorously exercising skeletal or cardiac muscle; and second, to explore the mechanism of ATP release in response to hypoxia. Washed human erythrocytes suspended in Krebs-Henseleit solution were exposed for 50 s to an atmosphere of approximately 8.0 kPa PCO2 and 2.7 kPa PO2; ATP released into the suspension was assayed using the firefly luminescence technique. Samples of human blood were obtained by venepuncture of the median cubital vein from male and female volunteers ranging in age from 21 to 74 years. Anticoagulation was with EDTA. A background of 0.49 x 10(6) (SEM 0.037 x 10(6)) ATP molecules.cell-1 was attributed to spontaneous haemolysis of 1% of the cell population, as estimated by levels of haemoglobin measured in the suspension fluid. When the erythrocytes were exposed to the hypoxic/hypercapnic gas mixture at 37 degrees C the ATP concentration in the suspension fluid rose to 2.67 x 10(6) (0.27 x 10(6)) molecules.cell-1. An efflux rate of 276(37) molecules.mu-2.s-1 was calculated. The hypoxia induced ATP release was blocked in three different ways: first, by application of 50 microM of the specific band 3 anion channel blocking agents niflumic acid (a translocation inhibitor), DIDS (a transport site inhibitor), or dipyridamole (a channel blocker); secondly, by replacement of extracellular chloride and bicarbonate with the impermeant anion methanesulphonate; and thirdly, by application of 5 nM of the nucleoside transport blocker nitrobenzylthioinosine. None of these blocking techniques affected the background levels of ATP attributed to haemolysis. A situation of hypoxia/hypercapnia, such as would be found in exercising muscle, induces release of ATP from the erythrocyte via the plasma membrane protein moiety known as band 4.5 (a nucleoside transporter) and electrical balance across the erythrocyte membrane is maintained by the simultaneous influx of extracellular chloride and/or bicarbonate via the plasma membrane protein known as band 3 (anion channel). The circulation of erythrocytes into a region of hypoxia in vivo could promote an increase in local blood flow through release of endothelium dependent relaxing factor in response to released ATP.


Benzo[e]pyrene elicits changes in the biochemical activities and chromatographic behavior of murine hepatic cytochromes P-450 that are distinct from those induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin. The objective of this study was to examine the potential for a specific ligand of carcinogen binding protein (CBP) to induce changes in the overall character of hepatic microsomal cytochromes P-450 (P450) and to compare potential changes with those induced by an Ah receptor ligand. Benzo[e]pyrene (BeP) was previously shown to bind CBP with high affinity and Ah receptor with low affinity. In contrast, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) binds Ah receptor avidly and CBP weakly. Hepatic microsomes were prepared from C57BL/6J (B6) and DBA/2J (D2) mice treated with corn oil, BeP or TCDD. Relative to corn oil controls, pretreatment of B6 mice with BeP or TCDD increased the nmol P450/mg microsomal protein content 26 and 28%, respectively. In D2 mice, nmol P450/mg microsomal protein was increased 23% in the BeP pretreatment, while TCDD pretreatment had no effect relative to the corn oil controls. For the O-alkyl ethers of resorufin, rates of metabolism (per nmol P450) were affected differently in B6 and D2 by BeP pretreatment. Pentoxyresorufin O-dealkylase activity was reduced to 44% of control activity in B6 mice and increased 39% relative to controls in D2 mice. BeP pretreatment had no effect on ethoxyresorufin O-dealkylase activity in B6 mice, while this activity was decreased to 58% of controls in D2 mice. Additionally, benzyloxyresorufin O-dealkylase activity was reduced to 65% of control levels in B6 mice and not affected in D2 mice. Methoxyresorufin O-dealkylase activity was reduced in both strains to an average of 55% of control values. As expected, TCDD pretreatment resulted in increases of all O-dealkylations measured in both strains of mouse. For both inbred strains of mouse, anion exchange chromatography revealed a P450 peak associated with BeP pretreatment that was not present in chromatograms generated with corn oil or TCDD pretreatments. Results of enzyme linked immunosorbant assays also indicated that the pattern of P450 isoenzyme expression associated with BeP pretreatment was distinct from that associated with TCDD pretreatment. Overall, these data show that treatment with a specific ligand of CBP induces changes the biochemical activities and chromatographic behavior of P450 isozymes in murine hepatic microsomes. Moreover, they indicate that changes in P450 occurring after treatment with a CBP ligand are distinct from those changes that are associated with treatment with an Ah receptor ligand (TCDD). Differences between B6 and D2 strains suggest that the hepatic P450 changes occurring in response to pretreatment with a CBP ligand may be influenced by the presence of Ah receptor.


Reductase and oxidase activity of rat liver cytochrome P450 with 2,3,5,6-tetramethylbenzoquinone as substrate. The main objective of the present study was to investigate the proposed role of cytochrome P450 in the reductive metabolism of quinones as well as in the formation of reduced oxygen species in liver microsomes from phenobarbital (PB-microsomes) and beta-naphthoflavone (beta NF-microsomes) pretreated rats. In the present study, 2,3,5,6-tetramethylbenzoquinone (TMQ) was chosen as a model quinone. Anaerobic one-electron reduction of TMQ by PB-microsomes showed relatively strong electron spin resonance (ESR) signals of the oxygen-centered semiquinone free radical (TMSQ), whereas these signals were hardly detectable with beta NF-microsomes. Under aerobic conditions TMSQ formation was diminished and concomitant reduction of molecular oxygen occurred in PB-microsomes. Interestingly, TMQ-induced superoxide anion radicals, measured by ESR (using the spin trap 5,5'-dimethyl-1-pyrroline-N-oxide), and hydrogen peroxide generation was found to occur with beta NF-microsomes as well. Furthermore, SK&F 525-A (a type I ligand inhibitor of cytochrome P450) inhibited TMQ-induced hydrogen peroxide formation in both PB- and beta NF-microsomes. However, metyrapone and imidazole (type II ligand inhibitors of cytochrome P450) inhibited molecular oxygen reduction in beta NF-microsomes and not in PB-microsomes. The present study indicates that cytochrome P450-mediated one-electron reduction of TMQ to TMSQ and subsequent redox cycling of TMSQ with molecular oxygen constitutes the major source for superoxide anion radical and hydrogen peroxide generation in PB-microsomes (i.e. from the reductase activity of cytochrome P450). However, most of the superoxide anion radical formed upon aerobic incubation of TMQ with beta NF-microsomes originates directly from the dioxyanion-ferri-cytochrome P450 complex (i.e. from the oxidase activity of cytochrome P450). In conclusion, both the one-electron reduction of TMQ and molecular oxygen were found to be cytochrome P450 dependent. Apparently, both the reductase and oxidase activities of cytochrome P450 may be involved in the reductive cytotoxicity of chemotherapeutic agents containing the quinoid moiety.


Cholesterol increases the L-type voltage-sensitive calcium channel current in arterial smooth muscle cells. To determine whether membrane free cholesterol affects calcium currents in vascular smooth muscle cells, whole-cell patch clamp recordings were made before and after cholesterol enrichment of cells by exposure to cholesterol-rich liposomes. Exposure to cholesterol-rich liposomes resulted in a gradual increase in the L-type current over 20 hours and a plateau (73 +/- 7% increase over basal) between 20 and 32 hours. This effect was associated with a rightward shift in the inactivation potential and a decrease in the sensitivity to (-)-PN-202-791, a dihydropyridine antagonist. There was no change in the maximum L-type current stimulated by (+)-PN-202-791, a dihydropyridine agonist. Liposome exposure caused a small, transient increase in the T-type current (peak effect, 20 minutes). We conclude that membrane cholesterol has important effects on the L-type calcium current in vascular smooth muscle cells, which is most likely due to an alteration in channel functional state rather than an increase in channel expression.


Captopril enhances intracellular calcium handling and beta-adrenergic responsiveness of myocardium from rats with postinfarction failure. To examine the cellular mechanisms of contractile dysfunction in postinfarction heart failure, we studied the effects of beta-adrenergic receptor stimulation on contractile function and Ca2+i handling of noninfarcted papillary muscles from sham-operated (n = 17) and infarcted (n = 17) rats. Ca2+i transients measured with the bioluminescent protein aequorin and parameters of isometric contraction were recorded during graded isoproterenol stimulation. Developed tension and peak rate of tension rise were depressed (p less than 0.05) in muscles from infarcted rats at physiological and maximally stimulating [Ca2+]oS. The time to peak tension was prolonged in the muscles from the infarcted rats, corresponding with prolongation of the time to peak Ca2+i. In muscles from sham-operated rats, isoproterenol increased both the amplitude of the Ca2+i transient and the peak rate of tension rise. In contrast, the inotropic response to isoproterenol was severely blunted in the muscles from infarcted rats despite a large increase in the amplitude of the Ca2+i transient. Isoproterenol abbreviated the time course of the isometric twitch and the Ca2+i transient in both groups. These findings suggest that postinfarction heart failure may be related in part to decreased force-generating capacity of the myofilaments. Treatment with captopril for 5 weeks, beginning 1 week after infarction (n = 14), resulted in reduction of left ventricular filling pressures and partial normalization of myocardial contractility and Ca2+i handling. In addition, compared with muscles from untreated infarcted rats, muscles from the captopril-treated rats exhibited improved contractile responses to increasing [Ca2+]o or isoproterenol. The inotropic response to isoproterenol in muscles from all three groups of rats had a significant negative correlation (r = -0.64, p less than 0.0001) with left ventricular end-diastolic pressure measured in vivo. Thus, the defect in excitation-contraction coupling in rats with postinfarction heart failure may be partially normalized by chronic load reduction with an angiotensin converting enzyme inhibitor.


Membrane-bound nucleoside diphosphate kinase activity in atrial cells of frog, guinea pig, and human. Muscarinic K+ channels in inside-out patches of atrial cells from guinea pig or rabbit can be activated by Mg(2+)-ATP in the absence of acetylcholine and GTP or GDP. The ATP-dependent activation involves a phosphorylation and is postulated to be due to the association of a membrane-bound nucleoside diphosphate kinase (NDPK) with the G protein GK: direct phosphorylation of the GK-bound GDP into GTP, catalyzed by NDPK, would result in activation of the G protein and, hence, activation of the channels. The aim of this study was to identify the presence of NDPK activity in atrial membranes by investigating the phosphate transfer between tritium-labeled nucleotides. We show that frog, guinea pig, and human atrial membranes contain a substantial NDPK activity since they catalyze the conversion from [3H]GDP+nucleoside triphosphate (NTP or NTP gamma S) to [3H]GTP (or [3H]GTP gamma S), from [3H]ADP+NTP to [3H]ATP, and from [3H]GTP+nucleoside diphosphate (NDP) to [3H]GDP. The phosphate transfer rates for the [3H]GDP+ATP to [3H]GTP conversion are 1.8, 0.5, and 2.4 mumol inorganic phosphate formation/mg per 10 minutes at 37 degrees C in frog, guinea pig, and human, respectively. The order of substrate efficiency for different NTPs was ATP greater than ITP approximately equal to GTP greater than UTP greater than CTP, which parallels the efficiency of these nucleotides in their activation of the muscarinic K+ channels. Addition of other nucleotides blocked the transphosphorylation reaction, indicating that the NTP-NDP conversion mechanism is aspecific, as is expected for an NDPK-catalyzed reaction. In conclusion, the data support the concept of NDPK involvement in the atrial muscarinic signal transduction cascade.


Influence of vascular smooth muscle heterogeneity on angiotensin converting enzyme activity in chicken embryonic aorta and in endothelial cells in culture. The smooth muscle of the abdominal region of the chicken aorta derives from locally recruited mesenchyme (mesenchymal smooth muscle), whereas that of the thoracic region derives from the neural crest (ectomesenchymal smooth muscle). We hypothesized that this smooth muscle heterogeneity might affect important enzymatic functions of the vessel wall. Therefore, we measured angiotensin converting enzyme (ACE) activity in homogenates of chicken thoracic and abdominal aorta at different embryonic stages (days 10, 14, and 18 of gestation). ACE activity increased in both regions over the time of gestation (p less than 0.001 in both cases); the increase was steeper and ACE activity was higher in thoracic than in abdominal segments (p less than 0.001). Km values were similar (approximately 7 microM) at all times and between the two segments, whereas changes in Vmax values closely paralleled those in enzyme activity, indicating gestation-dependent increases in the amount of enzyme. Neural crest ablation at an early developmental stage resulted in an increase of ACE activity in thoracic homogenates (p less than 0.001), predictably leaving that in abdominal homogenates unaffected. Bovine pulmonary artery endothelial cell monolayers exposed to media conditioned with cultured mesenchymal or ectomesenchymal smooth muscle cells exhibited elevated ACE activity (46% and 83%, respectively, relative to control medium, with p less than 0.01 in both cases; p less than 0.05 between the two media). Increases in endothelial cell ACE activity corresponded to proportional increases in ACE protein determined by enzyme-linked immunosorbent assay (r = 0.99) and were interpreted as indicative of enhanced enzyme synthesis subsequent to exposure of endothelial cells to smooth muscle-conditioned media.(ABSTRACT TRUNCATED AT 250 WORDS)


Endothelin increases myofilament Ca2+ sensitivity in alpha-toxin-permeabilized rabbit mesenteric artery. This study was designed to investigate the mechanism of endothelin-1 (ET-1) contractions in Staphylococcus alpha-toxin-permeabilized vascular smooth muscle. Rabbit small mesenteric arteries permeabilized with alpha-toxin were mounted for isometric or isotonic force recording or were processed for determination of myosin light chain (MLC) phosphorylation levels. Addition of 100 nM ET-1 plus 10 microM GTP significantly enhanced myofilament Ca2+ sensitivity as compared with the addition of Ca2+ alone (EC50, 0.47 microM Ca2+ for Ca2+ alone and 0.13 microM Ca2+ for ET-1 plus (GTP). This enhanced sensitivity was reversed by GDP beta S. ET-1-induced contractions were relaxed at a constant [Ca2+] by the addition of 30 microM cAMP or cGMP, demonstrating a direct effect of the cyclic nucleotides on contractile regulation. Inhibition of protein kinase C activity by 100 nM staurosporine relaxed ET-1 plus GTP-induced contractions, and pretreatment with 40 microM chelerythrine inhibited the ET-1 plus GTP increase in force. At 0.32 microM Ca2+, steady-state levels of shortening velocity were not increased by ET-1 plus GTP, although steady-state levels of MLC phosphorylation were significantly enhanced. The ET-1-induced increase in MLC phosphorylation was not altered by changes in [Ca2+], whereas the shortening velocity was Ca2+ dependent, suggesting that the increase MLC phosphorylation level may be the result of protein kinase C, rather than MLC kinase, activation. These results are consistent with the hypothesis that ET-1 increases myofilament Ca2+ sensitivity by a G protein-dependent pathway and subsequent activation of protein kinase C.(ABSTRACT TRUNCATED AT 250 WORDS)


Nitric oxide is a mediator of hypoxic coronary vasodilatation. Relation to adenosine and cyclooxygenase-derived metabolites. Hypoxia is a potent coronary-vasodilating signal; its mechanisms are still controversial. We have assessed the possible role of nitric oxide (NO) in hypoxic coronary vasodilatation (HCVD) in isolated guinea pig hearts perfused at constant pressure. HCVD was elicited by a 1-minute 100% N2 exposure; coronary flow doubled within 1 minute of hypoxia (early phase) and returned to baseline within 40 seconds after reoxygenation (late phase). The early phase of HCVD was associated with a rapid approximately eightfold increase in cGMP overflow, an indication of NO release. The specific NO synthase inhibitor N omega-methyl-L-arginine (NMA, 0.1-1 mM) antagonized HCVD and the associated increase in cGMP spillover (maximum inhibition, approximately 65%); excess arginine (1.2 mM) prevented both effects. The late phase of HCVD was associated with an increase in adenosine overflow and was attenuated by the adenosine receptor antagonist BW A1433 (1 microM; maximum inhibition, approximately 45%). Indomethacin (10 microM) inhibited HCVD in spontaneously beating hearts by approximately 35% but had no effect in hearts paced at faster rates. NMA and BW A1433 were more effective in combination than alone (maximum inhibition, approximately 72%). However, irrespective of the concentrations used, there was no synergism among the anti-HCVD effects of NMA, BW A1433, and indomethacin, nor was HCVD completely inhibited by the antagonists, whether alone or in combination. Our findings indicate that NO is an important mediator of the early phase of HCVD, whereas additional mechanisms and/or factors, including adenosine and vasodilatatory prostaglandins, contribute to the late phase.


Congenital hypertrophy of retinal pigment epithelium in patients with colonic polyps associated with cancer family syndrome. Congenital hypertrophy of retinal pigment epithelium (CHRPE) has been shown to be a frequent extracolonic manifestation of adenomatous polyposis coli (APC). The presence of CHRPE in patients with adenomatous polyps from families with cancer family syndrome suggests possible involvement of the APC gene locus in syndromes associated with less florid polyp formation than seen in APC. It also emphasises that caution must be exercised in using the presence of CHRPE clinically as a marker for APC in isolated at-risk individuals.


Potentiation of a weakly active ricin A chain immunotoxin recognizing the neural cell adhesion molecule. A ricin A chain immunotoxin, SEN36-ricin A chain, directed against the neural cell adhesion molecule (N-CAM) had no selective cytotoxic activity against three different small cell lung cancer (SCLC) cell lines in tissue culture despite expression of the target antigen on more than 98% of cells in each line detected by indirect immunofluorescence. Treatment of the SW2 SCLC cell line with suramin and interferons alpha and gamma increased the level of N-CAM expression only slightly and had no significant effect on the cytotoxic activity of the SEN36 immunotoxin. In the presence of the carboxylic ionophore monensin at a concentration of 0.1 microM, the toxicity of SEN36-ricin A chain to the SW2 cell line was enhanced by 12,000-fold. In contrast, lysosomotropic amines showed little or no potentiation of activity, suggesting that lysosomal degradation was not the major factor limiting the action of the anti-N-CAM immunotoxin. The findings of this study indicate that ricin A chain immunotoxins directed against N-CAM on SCLC are unlikely to have sufficient activity to be useful therapeutic agents in the absence of potentiating agents such as monensin, which can interfere with the normal intracellular pathways of antigen routing.


Elevated serum levels of soluble tumour necrosis factor receptors (sTNF-R) in patients with HIV infection. Serum levels of the soluble form of tumour necrosis factor receptor type II (p75) (sTNF-R) were determined in HIV-infected individuals and risk groups and were then correlated with the course of infection and prognosis. sTNF-R levels were determined by an ELISA with MoAbs and polyclonal antibodies to urine-derived sTNF-R proteins. The mean +/- s.e. levels of sTNF-R in the sera of 49 HIV+ male homosexuals, 34 HIV- male homosexuals and 44 matched controls were 6.1 +/- 0.3 ng/ml, 4.4 +/- 0.3 ng/ml and 3.4 +/- 0.2 ng/ml, respectively. All these values were significantly different between each of the groups (P less than 0.001-0.05). Sequential studies of sTNF-R revealed higher levels following seroconversion in 5/8 individuals, remained persistently high during the asymptomatic phase of the infection and became even more elevated in some ARC and AIDS patients. At the same time TNF-alpha was undetectable in sera obtained from HIV+ male homosexuals and from healthy controls. This was independent of stage of HIV infection, serum sTNF-R level and type of ELISA kit used. These findings suggest that TNF-alpha/TNF-R system is turned on before and during HIV infection and raise the possibility that sTNF-R, the natural inhibitor of TNF, may be of importance in determining the course and probably prognosis of the disease.


Mononuclear cells from HIV-infected patients produce factors which enhance functional activity of polymorphonuclear neutrophils from healthy subjects. The influence of mononuclear cell supernatants (MNCS) from nine healthy donors and 35 HIV-infected patients (17 with lymphoadenopathy syndrome (LAS), 15 with ARC and three with AIDS) on functional activity of polymorphonuclear neutrophils (PMN) from healthy donors was investigated. MNC after short-term cultivation (24 h) produced factors which enhanced chemiluminescence (CL) and chemotaxis of PMN. This augmentation did not depend on stimulation of MNC by mitogens (lipopolysaccharide Escherichia coli (LPS) and concanavalin A (Con A)) or on activation of PMN by FMLP. After 48 h of cultivation only MNC stimulated by LPS produced these factors. MNCS from HIV-infected patients provoked a more pronounced augmentation of PMN CL compared with MNCS from healthy subjects. This enhancement was observed in patients at all stages of infection, but was more pronounced in patients with LAS. MNCS impact on PMN CL was not connected with proliferative activity of MNC but was correlated with the level of CD4 cells. It was shown that removal of adherent cells from MNC fraction resulted in decreased MNCS impact. Treatment of MNCS by antibody to IL-1 beta, IL-8, interferon-alpha (IFN-alpha) and tumour necrosis factor-alpha (TNF-alpha) did not decrease MNCS impact on PMN CL.


Impaired proliferative responses of peripheral blood B cells from splenectomized subjects to phorbol ester and ionophore. The responses of peripheral blood B cells to mitogenic stimulation were examined in 12 splenectomized subjects without residual splenic function, as determined by pitted erythrocyte counts. These were compared to a group of healthy controls matched for age and sex. Polyclonal anti-immunoglobulin evoked a normal transient elevation in intracellular free Ca2+ in splenectomized subjects, thereby suggesting that the early events of the signal transduction pathway are not impaired. However, mitogenic stimulation by pre-treatment with phorbol ester and culture in presence of a calcium ionophore (Ionomycin) resulted in reduced uptake of 3H-thymidine and subsequent proliferation. Nevertheless, entry into the mitotic cycle, as assessed by expression of Ki67, was slightly, but not significantly impaired. Unlike in normal controls, where up to 7% of freshly-isolated B cells were Ki67+, almost no Ki67+ peripheral B cells were observed in splenectomized subjects. The data are consistent with the hypothesis that peripheral B cells in splenectomized subjects are in a reduced state of activation compared with normal controls and require additional growth factor stimulation before they can undergo mitosis.


Regional cellular responses to intraperitoneal infection. Leucocytes in different body compartments were examined in a murine intra-abdominal abscess model: (i) peripheral blood; (ii) peritoneal exudate; (iii) abscess wall; and (iv) abscess pus, with respect to their phagocytic ability, CR3 expression and Ia antigen expression (murine MHC antigen). The ability of serum or supernatant in each compartment to opsonize Staphylococcus aureus was also determined. Mice responded to caecal ligation and puncture (CLP) with systemic monocytosis, an increased amount of opsonins in peritoneal fluid and an increased phagocytosis in pus leucocytes. This increased phagocytic ability and CR3 expression both subsequently decreased. Monocyte/macrophage Ia expression was reduced in all measured compartments over time. In contrast, sham-operated animals, without an intra-abdominal abscess, responded to operation with an even greater increase in peritoneal exudate phagocytic ability and in monocyte/macrophage Ia expression in all compartments which was sustained beyond 1 week. Decreased peritoneal exudate cell and peripheral blood Ia was still present after 28 days in CLP animals, compared with both sham and normal animals (P less than 0.05, P less than 0.02, respectively). For all parameters, changes observed in peripheral blood did not reflect those present near the site of infection. Proper understanding of local or regional infection must take into account and ultimately alter these generally unappreciated changes in and about the actual site of infection.


A recombinant topoisomerase I ELISA: screening for IgG, IgM and IgA anti-topo I autoantibodies in human sera. An ELISA for the detection of anti-topoisomerase I autoantibodies in sera from patients with suspected or manifest rheumatic diseases is described. The antigen source used in this assay consists of a recombinant protein containing the last 695 C-terminal amino acid residues of human topoisomerase I (topo I). The sensitivity of the assay was 61%, while the specificity was more than 98%. Using this ELISA, 47 sera from scleroderma patients and immunopositive for anti-topo I antibodies, were screened for the presence of the isotypes IgG, IgA and IgM to topo I. Our finding that relatively high levels of IgA antibodies to topo I are present in most of the sera tested is consistent with the results of Hildebrandt et al. [1]. In addition, it is demonstrated that the IgG and IgA antibodies in a serum may recognize different epitope regions on the topo I polypeptide.


Signal transduction pathways leading to the production of IL-8 by human monocytes are differentially regulated by dexamethasone. Previous studies have shown that IL-8 gene expression is enhanced by various stimuli, which induce different signal transduction pathways. A lipopolysaccharide (LPS)-induced pathway has been reported to be inhibited by glucocorticoids in monocytes. We have now examined the effect of dexamethasone on the LPS-induced and other signal transduction pathways leading to the production of IL-8 by human monocytes. Dexamethasone inhibited the production of IL-8 stimulated with a cyclic adenosine monophosphate analog or LPS. In contrast, dexamethasone had no significant effect on a phorbol ester (PMA)-stimulated IL-8 production. These results suggest that the signal transduction pathways leading to the production of IL-8 by human monocytes are differentially regulated by dexamethasone.


Inhibitory effect of neuropeptide Y on stimulated renin secretion of awake rats. 1. (1-36)-NPY is a vasoconstrictor peptide widely distributed in sympathetic nerve terminals. This peptide exerts an inhibitory action on renin release induced by various stimuli. Post-synaptic neuropeptide Y (NPY) receptors show a high affinity for (1-36)-NPY as well as for the agonist (Pro34)-NPY, while presynaptic receptors bind preferentially (13-36)-NPY. 2. This study was undertaken to assess whether the NPY induced renin suppression in awake normotensive rats infused with the beta-adrenoceptor stimulant isoproterenol is mediated by activation of pre- or post-synaptic receptors. 3. Non-pressor doses of (1-36)-NPY and (Pro34)-NPY markedly attenuated the renin secretion triggered by isoproterenol whereas (13-36)-NPY had no effect. This suggests that the effect of NPY on renin release is due to the stimulation of post-synaptic receptors. However it remains unknown whether NPY acts directly on juxtaglomerular cells or indirectly by modifying intraglomerular haemodynamics.


Effective anticoagulation by a low molecular weight heparin (Fragmin) in hemodialysis with a highly permeable polysulfone membrane. The efficacy and kinetics of a low molecular weight heparin fragment (LMWH-fragment, Fragmin) were studied during one hemodialysis session with a highly permeable polysulfone membrane and compared to a second dialysis session using a conventional cuprophane membrane. All patients received 5000 U of Fragmin given as an injection into the arterial line at start of dialysis. The anticoagulative efficacy was evaluated by measuring plasma fibrinopeptide A concentrations. LMWH-fragment concentrations in plasma and ultrafiltrate were determined by an amidolytic activity assay and by a radioimmunoassay using a monoclonal antibody. During hemodialysis with cuprophane and polysulfone membranes the fibrinopeptide A concentrations were low indicating adequate anticoagulation. LMWH concentrations in plasma did not differ in the two membranes at any time. The LMWH-fragment in the ultrafiltrate could neither be detected with the amidolytic assay nor with the radioimmunoassay. We conclude that a single injection of Fragmin effectively prevents clotting during hemodialysis with a highly polysulfone membrane. No significant amounts of the anticoagulant are lost over the dialysis membrane.


Limits of adaptation to a diet low in protein in normal man: urea kinetics. 1. Urea kinetics were measured using prime/intermittent oral doses of [15N15N]urea in six healthy men taking diets adequate in energy and containing either 74 or 30 g of protein/day. 2. On 74 g of protein/day, urea production (199 mg of N day-1 kg-1) was 121% of intake, with 60% of the urea produced being excreted in the urine and 40% being salvaged in the colon; 69% of the salvaged nitrogen was retained in the metabolic nitrogen pool. 3. Nitrogen balance was not maintained on 30 g of protein/day. There was a significant decrease in the urea production rate (123 mg of N day-1 kg-1) and 54% of production was excreted in urine, with 46% being salvaged. 4. The pattern of urea production and salvaging on 30 g of protein/day was different to that seen in an earlier study on 35 g of protein/day, with a significant decrease in both production (71%) and salvaging (50%). 5. These data reinforce the conclusions drawn from an earlier study, that the salvaging of urea nitrogen by the colon is an integral part of the process of adaptation to low protein diets. The salvage system appears to fail on an intake of 30 g of protein/day and nitrogen is no longer conserved in sufficient amounts for balance to be maintained. 6. The changes seen in urea kinetics reinforce the conclusion based upon nitrogen balance that the minimum physiological requirement for protein in normal adult man lies between 30 and 35 g of protein/day.


Comparisons between basal metabolic rate and diet-induced thermogenesis in different types of chronic obstructive pulmonary disease. 1. Some patients with the emphysematous type of tobacco-related chronic obstructive pulmonary disease are hypermetabolic. Since the likely mechanism is the increased work of breathing, other groups of patients with chronic obstructive pulmonary disease should be similar. We have now measured basal metabolic rate and diet-induced thermogenesis in six patients with chronic obstructive pulmonary disease with an arterial partial pressure of CO2 of less than 5 kPa (emphysematous), nine patients with chronic obstructive pulmonary disease with an arterial partial pressure of CO2 of greater than 6 kPa (bronchitic), eight patients with chronic obstructive pulmonary disease due to chronic asthma and seven control subjects. Diet-induced thermogenesis was measured for 4h after a meal of 87% carbohydrate, 11% protein and 2% fat as energy, with a total energy content of 40% of basal metabolic rate. 2. There was no difference between measured and predicted basal metabolic rate in the control (5541 +/- 272 versus 5881 +/- 245 kJ/24h) or emphysematous (5552 +/- 370 versus 6239 +/- 197 kJ/24h) groups, but measured basal metabolic rate was significantly higher than predicted in the bronchitic (6126 +/- 387 versus 5405 +/- 250 kJ/24h) and asthmatic (6293 +/- 197 versus 5701 + 245, mean +/- SEM, P less than 0.01) groups. All the control subjects had measured basal metabolic rates within 10% of predicted, whereas two out of six emphysematous patients, four out of nine bronchitic patients and five out of eight asthmatic patients were hypermetabolic. The contributions of fat, carbohydrate and protein oxidation rates to the overall basal metabolic rate were similar between groups. 3. Diet-induced thermogenesis was similar between groups.(ABSTRACT TRUNCATED AT 250 WORDS)


Cutaneous heat loss shortly after burn injury in children. 1. Total heat loss and its components have been studied in cool (20 degrees C) and warm (30 degrees C) environments in 30 healthy children and 21 children who had been burned (10-17% body surface area) 0.5-29 h previously. 2. In healthy naked children at 20 degrees C, the partition of total heat loss was: radiation, 64%; convection, 32%; evaporation, 4%. On transfer to the warm, total heat loss was reduced by approximately 50%, with disproportionate reductions in the contributions from radiation and convection being offset, to some extent, by an increase in evaporative heat loss. 3. In patients during the first 5.5h after injury, the magnitude and pattern of heat loss at 20 degrees C and 30 degrees C were similar to those in control subjects and were unaffected by bandaging. 4. Ten to twenty-nine hours after injury, when the patients were bandaged and body temperature and heat content were significantly higher than in control subjects, radiant and convective heat losses were increased, but as evaporative heat loss tended to be reduced; total heat loss in the warm was unchanged. However, at this time at 20 degrees C, total heat loss was reduced compared with healthy children at the same ambient temperature. 5. The findings of unchanged or reduced total heat loss and reduced evaporative heat loss in injured patients are interpreted as inappropriate responses to an increased body temperature and heat content in children after burn injury.


Hypertension in Dahl salt-sensitive rats: biochemical and immunohistochemical studies. 1. The renin-angiotensin and kinin-kallikrein systems of Dahl salt-sensitive and salt-resistant rats fed diets with different salt contents were analysed using biochemical and immunocytochemical techniques. 2. Blood pressure increased by 45% in salt-sensitive rats only, after 4 weeks on a high-salt diet. The plasma renin activity and plasma angiotensin II concentration remained at the same levels in salt-sensitive rats on the high-salt diet as on the normal salt diet, whereas the plasma renin activity and plasma angiotensin II concentration of salt-resistant rats fed the high-salt diet were lower. The plasma renin activity and the plasma angiotensin II concentration were elevated in both salt-resistant and salt-sensitive rats fed the salt-deficient diet but were much more elevated in salt-resistant than in salt-sensitive rats. 3. The kidney immunocytochemical data paralleled the data on plasma parameters. Salt-sensitive rats had fewer renin positive juxtaglomerular apparatuses than salt-resistant rats on the normal diet, and the increase on the sodium-deficient diet was also smaller in salt-sensitive rats. Salt-sensitive rats fed the high-salt diet and the standard diet had almost no angiotensin II immunoreactivity compared with the salt-resistant rats on the same diets. 4. The total renal kallikrein content of salt-sensitive rats was lower than that of salt-resistant rats on all three diets, as was the amount of kallikrein excreted in the urine on the standard and the high-salt diets. The differences resulted from a reduction in active kallikrein. The increase in kallikrein in salt-sensitive and salt-resistant rats on the salt-deficient diet was not significantly different. 5. There were similar changes in immunopositive kallikrein in the kidneys of salt-sensitive and salt-resistant rats with diet, with a large increase in kallikrein biosynthesis on the low-salt diet. The plasma concentration of high-molecular-mass kininogen was not significantly different in salt-sensitive and salt-resistant rats, but there was a significant increase in T-kininogen in salt-sensitive rats fed the high-salt diet. 6. In conclusion, the absence of decreases in the plasma renin activity and the plasma angiotensin II concentration in salt-sensitive rats fed the high-salt diet might partially explain the increase in blood pressure.


Influence of dietary protein on glomerular angiotensin II-receptor binding in normotensive and spontaneously hypertensive rats. 1. The binding of angiotensin II to glomerular receptors was studied in spontaneously hypertensive and normotensive Wistar-Kyoto rats in response to 7, 16 and 32% isocaloric, isonatraemic protein diets. 2. Increased dietary protein elevated the systemic angiotensin II levels of both spontaneously hypertensive and Wistar-Kyoto rats [F0.05(2,26) = 4.758, P less than 0.05; n = 36], and this was not associated with changes in either systemic blood pressure or cortical renin activity. 3. Furthermore, no significant changes in the affinity or density of angiotensin II receptors were associated with changes of dietary protein intake in either strain. 4. These results indicate a dissociation between the system renin-angiotensin system and the tissue renin-angiotensin system in response to protein intake.


Measurement of plasma volume in pregnancy. 1. Determination of the plasma volume in pregnant women is a useful research tool and may become an important clinical measurement. We used three methods to determine plasma volume using Evans Blue dye: (1) the 'usual' method, measuring serum absorbance at a wavelength of 610 nm, (2) a two-wavelength method, and (3) precipitation of non-albumin proteins by the addition of polyethyleneglycol before measuring serum absorbance at a wave-length of 620 nm. These were each compared with the standard 125I-human serum albumin method in 20 non-pregnant subjects. Subsequently, the polyethylene glycol method was considered the standard and the three Evans Blue dye methods were compared in 20 pregnant women. 2. In non-pregnant subjects mean plasma volumes did not differ significantly according to the method used. However, the limits of agreement with 125I-human serum albumin method were closest for the polyethyleneglycol method, for both clear and turbid sera. 3. In pregnant women, mean plasma volume values did not differ according to the Evans Blue dye method used, but the limits of agreement were significantly closer with the two-wavelength method than with the 'usual' method (P less than 0.05) largely owing to the effects of turbid sera. 4. These studies demonstrate that considerable error may occur when the Evans Blue dye concentration is determined in turbid sera by the 'usual' method. This can be overcome by the use of the two-wavelength method or the polyethyleneglycol method. The most accurate results will be obtained if the latter method is employed routinely to determine plasma volume in pregnant women.


Atrial natriuretic peptide inhibits fluid intake in hyperosmolar subjects. 1. The effect of atrial natriuretic peptide on osmotically stimulated thirst appreciation and consequent fluid intake was investigated in healthy man. 2. Six seated male subjects were studied on two occasions: synthetic alpha-human atrial natriuretic peptide (99-126) (2 pmol min-1 kg-1) or placebo (saline, 150 mmol/l NaCl) was infused intravenously for 105 min; 30 min after the start of atrial natriuretic peptide/placebo infusion, hypertonic saline (855 mmol/l NaCl) was infused (0.06 ml min-1 kg-1) for 60 min. Subjects were then allowed free access to water for the next 2 h; infusion of atrial natriuretic peptide/placebo continued for the first 15 min of the drinking period. 3. The plasma atrial natriuretic peptide concentration did not alter significantly during infusion of hypertonic saline and placebo; it rose to a steady state of 12.7 +/- 1.1 pmol/l (mean +/- SEM) during the infusion of atrial natriuretic peptide and hypertonic saline, and remained at this level during the first 15 min of the drinking period. During infusion of hypertonic saline and atrial natriuretic peptide or placebo, similar increases in plasma osmolality (P less than 0.001) and plasma vasopressin concentration (P less than 0.005) occurred. During infusion of hypertonic saline and atrial natriuretic peptide or placebo, thirst increased significantly over the time course of both studies (P less than 0.01), but the effect of atrial natriuretic peptide infusion compared with placebo infusion was to significantly decrease thirst at 60 min. 4. Drinking rapidly abolished thirst and vasopressin secretion before changes in plasma osmolality occurred.(ABSTRACT TRUNCATED AT 250 WORDS)


Systemic and splanchnic haemodynamic effects of pentifylline in rats with portal hypertension. 1. The systemic and splanchnic haemodynamic effects of pentifylline (40 mg/kg body weight intravenously) were assessed in rats with portal hypertension associated either with CCl4-induced cirrhosis (n = 13) or portal vein ligation (n = 13). 2. Heparinized catheters were placed into the portal vein, inferior vena cava, aorta and left ventricle with exits from the neck. Haemodynamic studies were performed 4 h after consciousness was regained. Cardiac output and regional blood flows were measured using radiolabelled microspheres and the reference sample method in seven rats in each group; portal-systemic shunting was measured using microsphere injection in the ileo-colic vein in six rats in each group. 3. Forty-five minutes after injection, pentifylline had no effect on mean arterial pressure, cardiac output, peripheral resistance, portal venous flow, hepatic artery flow or portal-systemic shunting in either group of rats with portal hypertension. The drug lowered portal pressure (-18%) in cirrhotic rats, but not in portal-vein-ligated rats. 4. These data demonstrate that pentifylline lowers portal pressure in cirrhotic rats without affecting portal venous flow and portal-systemic shunting; this effect is possibly mediated by changes in intrahepatic resistance related to the effects of pentifylline on blood viscosity and/or on intrahepatic vasomotor tone.


Intrasplenic blood cell kinetics in man before and after brief maximal exercise. 1. After a 4 min period of maximal exercise in 10 normal subjects (14 studies), there was a consistent decrease in total blood volume and a consistent increase in erythrocyte indices, which were maximal immediately after exercise. Peripheral platelet and leucocyte counts increased, but did not reach maximal values until 5-10 min after the end of exercise. 2. The distributions of 99mTc-labelled erythrocytes (five studies), 111In-labelled platelets (five studies) and 111In-labelled granulocytes (four studies) were monitored with a gamma-camera immediately after injection and before and after maximal exercise performed 60 min after injection. 3. Labelled erythrocytes equilibrated rapidly between the spleen and circulating blood after injection, whereas labelled platelets and granulocytes equilibrated more slowly. After exercise, each cell type was released from the spleen with a time course that was the reciprocal of the time course of the corresponding cell count in peripheral blood. Thus, whereas the radioactivity of 99mTc-labelled erythrocytes in the spleen, which fell to 0.46 (SD 0.09) of the pre-exercise value, increased towards its baseline value as soon as exercise was completed, the radioactivities of 111In-labelled platelets and 111In-labelled granulocytes decreased, to respective minimum values of 0.61 (0.09) and 0.63 (0.09) of the pre-exercise levels, 5-10 min after the end of exercise. The exercise-induced changes in lung radioactivity for each cell type, and their time courses, broadly reflected those in the corresponding cell counts in peripheral blood. Liver radioactivity tended to decrease for each cell type.(ABSTRACT TRUNCATED AT 250 WORDS)


Measurement of pelvic blood flow changes in response to posture in normal subjects and in women with pelvic pain owing to congestion by using a thermal technique. 1. A method is described for studying pelvic blood flow in women by the indirect method of measuring vaginal temperature changes in response to a posture change. Ten women with chronic pelvic pain and venous congestion and 10 normal subjects were observed during posture changes over a 2 h period. 2. Vaginal temperature minus axillary temperature rose after subjects changed from the supine to the seated position, indicating a fall in the rate of pelvic blood flow. 3. A significantly greater variance in the rate of change in vaginal temperature minus axillary temperature was found in patients with pelvic venous congestion compared with control subjects (P less than 0.005). 4. The findings are consistent with a disorder of blood flow regulation in women with pelvic pain owing to congestion.


Differential responses in superior mesenteric artery blood flow may explain the variant pressor responses to clonidine in two groups with sympathetic denervation. 1. Measurement of superior mesenteric artery blood flow along with systemic and regional haemodynamic changes in blood pressure, heart rate, cardiac index, forearm blood flow, digital skin blood flow and index finger temperature were made before and after administration of clonidine (2 micrograms/kg body weight intravenously) in 10 patients with multiple-system atrophy, 10 patients with pure autonomic failure and 15 age-matched healthy control subjects. 2. After clonidine, blood pressure fell in patients with multiple-system atrophy and control subjects but not in patients with pure autonomic failure. 3. Resting superior mesenteric artery blood flow was similar in patients with multiple-system atrophy and control subjects, but was higher in patients with pure autonomic failure. The fall in blood pressure after clonidine was accompanied by active dilatation of the superior mesenteric artery in patients with multiple-system atrophy and control subjects. This did not occur in patients with pure autonomic failure. 4. After clonidine, there was a fall in cardiac index in patients with multiple-system atrophy. 5. After clonidine, changes in other haemodynamic parameters were not significant in any group, except for a fall in forearm blood flow and a rise in index finger temperature in control subjects. 6. We conclude that after clonidine there are differential superior mesenteric artery blood flow responses in the two groups with autonomic failure (multiple-system atrophy and pure autonomic failure). These may relate to differences in the site of the sympathetic lesion, which is considered to be mainly central in multiple-system atrophy but peripheral in pure autonomic failure.(ABSTRACT TRUNCATED AT 250 WORDS)