Exact stratification of person-years.

Exact stratification of person-years. This paper describes methods for the exact assignment of person-years of follow-up to strata of time-dependent factors. It reviews methods applicable to variables that directly measure the passing of time and that increase in step with follow-up (monotonic time factors), and it extends their use to nonmonotonic time factors. When the same categorization scheme is used for all time factors (for example, 5-year categories), the follow-up period consists of segments whose length and position in time are predictable. A new exact method presented here uses this property to improve the efficiency of computations. The new method is generally faster than a method previously described. Finally, the paper outlines a method of exact person-year stratification that is applicable to all time-dependent factors.


Confidence limit analyses should replace power calculations in the interpretation of epidemiologic studies. Frequently, after an epidemiologic study is completed, statistical power to detect a relative risk of interest is recalculated using data obtained during the course of the study. A negative study may then be dismissed on the grounds that its power was too low. However, post hoc power calculations ignore the actual relative estimate and its variance, which are by then known. We present evidence that post-study power calculations have little value and should be replaced by a more informative method using the upper (1 - alpha)% confidence limit of the point estimate that touches the value of the relative risk of interest.


Confidence interval estimation of interaction. Relative excess risk due to interaction, the proportion of disease among those with both exposures that is attributable to their interaction, and the synergy index have been proposed as measures of interaction in epidemiologic studies. This paper presents the methodology for obtaining confidence interval estimates of these indices utilizing routinely available output from multiple logistic regression software.


The effects of nondifferential confounder misclassification in ecologic studies. In ecologic studies, covariate levels of groups are often quantified as the prevalence of a dichotomous covariate. We show that, under certain conditions, nondifferential misclassification of such a binary covariate does not reduce the ability to control confounding by the covariate in ecologic studies. Thus, any remaining exposure-disease association in an adjusted ecologic analysis cannot be ascribed to incomplete control for confounding due to nondifferential misclassification of the dichotomy under those conditions, although residual confounding by the underlying covariate may still be present. This point is illustrated by ecologic analyses of the association between population density and mortality from lung cancer in women in 30 administrative districts of the Federal Republic of Germany, in which control for cigarette smoking is required.


Risk factors for cardiovascular disease predict the development of diabetes among Utah families. In 1989, we sent a medical follow-up questionnaire to 2,728 members of 98 Utah families originally screened from 1980 to 1983 in the Cardiovascular Genetics Research Clinic. The response rate was 69.9%. Of 1,134 nondiabetic individuals initially age 18 or older who returned the questionnaire, 10 were found to be newly diagnosed with diabetes. The incidence of diabetes was higher among individuals who were found at baseline to have central obesity, lipid abnormalities, especially increased triglyceride levels, and hypertension. Family histories of coronary heart disease and diabetes were not related to the development of diabetes. Our findings that cardiovascular disease risk factors predict the development of diabetes in this relatively young, Caucasian population are consistent with the results of studies from several different populations.


Cancer epidemiology in the former Soviet Union. Cancer epidemiology in the former USSR is predominantly descriptive and depends heavily on cancer registration. Cancer epidemiologists have spent most of the last 35 years "correcting" the serious inconsistencies in reported incidence data, as official cancer statistics are notoriously incomplete and inaccurate. Professional standards of Soviet cancer epidemiologists reflect the prevailing conditions they have worked in, notably, severe censorship, bans on publishing, lack of computers for compilation of data or analysis, loose recordkeeping practices in institutions, restricted access to scientific literature, and limited opportunities for training in biostatistics and epidemiology. In the eyes of a new generation of young scientists, modern epidemiology is not an attractive discipline. Despite having one of the largest and most diverse populations in the world, the scientific productivity of ex-Soviet cancer epidemiologists is small. The increasing number of publications from the former USSR in international journals and of ongoing projects is an encouraging sign that cancer epidemiology in the republics that comprised the Soviet Union may be emerging from its prolonged infancy. Prospects depend on the ability of researchers to weather current economic and political disturbances.


[Cisapride and sucralfate in dyspepsia associated with duodenogastric reflux gastritis]. The present investigation was undertaken with the aim of evaluating the clinical efficacy on dyspeptic symptoms associated with duodenogastric reflux gastritis of two drugs belonging to two different groups: a prokinetic (cisapride) and a cytoprotective agent (sucralfate). A total of 18 patients with duodenogastric reflux gastritis diagnosed on the basis of symptoms, endoscopy and histology were studied. Nine were given 30 mg of cisapride/daily and 9 4 g of sucralfate/daily for two months according to a randomization list. Pyrosis, epigastric pain, sense of epigastric repletion and foul-tasting mouth were considered on a scale from 0 to 4 attributed by the patient. The total score of dyspeptic symptoms significantly decreased only after cisapride (p less than 0.05). Considering each symptom alone, neither cisapride or sucralfate were able to significantly improve them. Cisapride seems to the better than sucralfate in improving dyspeptic symptoms associated with duodeno-gastric reflux gastritis.


[Effects of the administration of magnesium hydroxide on gastric acidity in health volunteers]. Antacids are often the first therapeutic approach in patients with pyrosis. We carried-out a study on 8 healthy volunteers who underwent a 24 hour gastric pH-metry to assess the real efficacy of magnesium hydroxide administration on gastric acidity. Magnesium hydroxide was alternatively administered at different dosages (400 mg or 800 mg) in the population studied. Our results showed a mild and non reproducible response to lower dose, but, on the contrary, the 800 mg dose always induced an immediate, effective and prolonged antacid action, reaching a maximum pH value of 5 and lasting up to 40 minutes. Our study confirms, using a modern and reliable technique as the 24-hour gastric pH-metry, the antacid activity of magnesium hydroxide.


Historical trends in road accident types, deaths and casualties in Western Australia. Road accident casualties are major consumers of health service resources in Australia, using inpatient care, accident and emergency treatment and other facilities. We analysed hospitalisations resulting from road accidents in Western Australia from 1968 to 1987 to examine trends in accident types, deaths and casualties. Data from the Western Australian Health Department's Hospital Morbidity System were used. Although hospitalisations from road accidents generally decreased, they were still higher than the Australian average in 1985. The rate of decline in hospitalisations was similar to that for fatalities and was not generally related to age or sex. There was some evidence of a more rapid decline in the rate of severe injuries than in minor ones. Further steps need to be taken to reduce the number of casualties from road accidents, as road accidents represent a major public health problem in Western Australia.


Identifying hazards and risk opportunity in child farm injury. Although children are overrepresented in farm injury, little is known about the environmental hazards and risk behaviours associated with injury, or about how to identify these factors at a local level. This study addresses the measurement of these hazards and hazardous behaviours. The study was conducted in 1990 at Gloucester, New South Wales, a small dairy, beef and hobby farm community. After some formative research and local consultation, a checklist survey was constructed and sent to 120 farm families with school-age children. Families were sent the checklist forms again two weeks later to assess test-retest reliability, which was found to be acceptable among the 38 per cent who responded on both occasions. The findings on the prevalence of environmental hazards and risk behaviours from the 84 per cent of respondents were useful to refine the existing injury information available from local hospital morbidity figures, which had identified injuries related to riding (horses, bicycles and motorcycles) and to machinery and drowning as major rural injury issues. In particular the importance of bicycle riding and horse-related injury were confirmed. The survey importantly identified some previously undetected issues, most notably the danger to children's hearing. The prevalence data were used to identify targets for the development of local health promotion initiatives, leading the local farm safety action group to select horses, helmets and hearing as issues for preventive action. Findings from the method indicated the importance of local information, involving farmers in constructing the checklist, and feeding back results to the community.


Dogma disputed: potential endemic heterosexual transmission of human immunodeficiency virus in Australia. The concept of tertiary sexual transmission of human immunodeficiency virus (HIV) has been central to government efforts to communicate notions of risk to heterosexuals in Australia. Data on heterosexually transmitted acquired immune deficiency syndrome (AIDS) and HIV for Australia are reviewed with emphasis given to the probability of misclassification bias in the heterosexually acquired and 'other/undetermined' categories. Tertiary cases are almost certainly rare in Australia, with little evidence of any increase in their incidence since the first cases were recorded. Three factors (low probability of exposure, the infectivity of HIV and a comparatively low rate of sexual partner change) make it improbable that Australian heterosexuals with no risk factors will experience endemic HIV infection, with a caveat to this conclusion lying in the potential of Australian sex tourism to Southeast Asia for introducing HIV into the Australian heterosexual population. Four hegemonic factors which have acted to suppress any serious debate of the notion that HIV in Australia is unlikely to become endemic among heterosexuals are discussed: the political 'democratization' of risk inspired by concerns that gay men should not be further vilified as a victim group; the preventive imperative; a reluctance among health educators to question the very foundations of the message they are employed to deliver; and a reluctance to curtail 'Trojan horse' benefits to sexually transmissible disease prevention engendered by HIV education promoting safe sex messages.


Discourse analysis: a new methodology for understanding the ideologies of health and illness. Discourse analysis is an interdisciplinary field of inquiry which has been little employed by public health practitioners. The methodology involves a focus upon the sociocultural and political context in which text and talk occur. Discourse analysis is, above all, concerned with a critical analysis of the use of language and the reproduction of dominant ideologies (belief systems) in discourse (defined here as a group of ideas or patterned way of thinking which can both be identified in textual and verbal communications and located in wider social structures). Discourse analysis adds a linguistic approach to an understanding of the relationship between language and ideology, exploring the way in which theories of reality and relations of power are encoded in such aspects as the syntax, style and rhetorical devices used in texts. This paper argues that discourse analysis is pertinent to the concerns of public health, for it has the potential to lay bare the ideological dimension of such phenomena as lay health beliefs, the doctor-patient relationship, and the dissemination of health information in the entertainment mass media. This dimension is often neglected by public health research. The method of discourse analysis is explained, and examples of its use in the area of public health given.


Cultural identification in aboriginal and Torres Strait Islander AIDS education. The emergence of the disease AIDS in the early 1980s has resulted in a unique response. Medical, sociocultural, political, sexual, moral and racial issues have all been raised. This paper examines the way in which participation of Aboriginal and Torres Strait Islander people has resulted in the culturally appropriate and distinctive approaches evident in health education materials produced in Aboriginal and Torres Strait Islander communities. Specific cultural issues relevant to AIDS education are considered, including the use of visual and narrative communication for AIDS education; the significance of the specific concepts related to communication on sexual issues; perceptions of AIDS as alien and genocidal; the use of the Dreaming in AIDS educational resources; and implications for AIDS education.


[Enteral nutrition in the elderly].

[Enteral nutrition in the elderly]. Elderly is particularly at risk of malnutrition: he is not able to feed himself adequately, it is then important to attain correct intakes using also artificial enteral nutritional techniques (nasogastric tube, gastrostomy, etc.). These techniques may lead to complications (ab ingestis pneumonia, metabolic complications, alvus disorders): the use of artificially nutrition in the elderly must be carefully evaluated. 257 patients (M = 180, F = 77) aged 65 or more, mainly affected by neoplastic diseases (n 195) and by neurological and vascular diseases (n 62). The feeding route were evaluated in this study: 74% by nasogastric tube, 13% by gastrostomy, 11% by jejunostomy. In a group of 55 patients similar concerning clinical and nutritional conditions we evaluated at the beginning of enteral feeding and four months later, caloric/protein intake, body weight and plasmatic albumin. In patients fed by nasogastric tube a mean intake of 1300 +/- 365 Kcal n.p./die, with a protein rate of 58.5 +/- 16.9 g/die was attained; by gastrostomy 1450 +/- 324 Kcal n.p./die and 65.5 +/- 16 g/die; by jejunostomy 1219 +/- 398 Kcal n.p./die and 53.3 +/- 21 g/die. The compliance to enteral nutrition was well in 37% of patients night administration was performed. Clinical complications: nausea and vomiting were observed in 9 patients with nasogastric tube, in 1 patient with gastrostomy and in 3 patients with jejunostomy; diarrhea has been noticed in 6 patients with nasogastric tube and in 1 patient with jejunostomy. Mechanical complications; nasogastric tube (n 189): 35 displacements, 7 breakages, 4 obstructions; pharyngostomy (n 6): 2 displacements and 1 obstruction; gastrostomy (n 33): 3 displacements; jejunostomy (n 29): 2 misplacements.(ABSTRACT TRUNCATED AT 250 WORDS)


The national cot death prevention program in New Zealand. A case-control study examining the risk factors for sudden infant death syndrome (SIDS) in New Zealand identified three risk factors that are potentially amenable to modification: prone sleeping position of the infant, maternal smoking and lack of breastfeeding. In total these three risk factors may account for 79 per cent of deaths from SIDS in New Zealand. We describe the planning and implementation of the cot death prevention program, which has involved a wide range of groups and different strategies. The outcome of the prevention program is being evaluated.


Evaluation of a heart disease survey and education program for general practitioners. In 1989 we mailed a questionnaire to all 461 general practitioners (GPs) identified as currently practising in the Eastern Metropolitan Health Region of Sydney. This was the first phase of a program to assess, amplify and reassess GPs' knowledge of the risk factors for heart disease and measure their attitudes and beliefs about their role in the prevention of cardiovascular disease. The second phase was an education program designed to meet the needs identified by the first questionnaire. Phase three was a postintervention questionnaire. Fifty-six per cent (260/461) responded to the first questionnaire. This follow-up group were mailed the second questionnaire, to which 52 per cent (135/260) responded. Thirty per cent of the original sample (139/461) attended the education program and 30 per cent (79/260) of the follow-up group did so. At baseline, the respondents' level of risk factor knowledge was good, but after the education program there was still a large gap between what they said they knew and the amount of advice they said they would give to patients. The only significant increase in the amount of advice after the intervention was to 'control blood pressure'. This applied whether the GP had participated in the intervention or not. When GPs were asked how often in the last month they had actually given advice to reduce cardiovascular disease risk, program attenders reported offering it more frequently than nonattenders. We also attempted to determine whether any particular demographic characteristics could predict respondents to the questionnaires and/or the educational program.


[Chief cell mass in gastric ulcer: cyto-secretory correlations]. The aim of this experience has been to evaluate the chief cell mass and serum pepsinogen I in gastric ulcer patients. Comparisons were also made with parietal cell mass and acid secretion. Chief cell mass and serum pepsinogen I are not only influenced by the localization of ulcer but, also, by the histological condition of fundic mucosa. In fact, the behaviour of serum pepsinogen I and chief cell mass in type I gastric ulcer is the same observed in case of fundic chronic gastritis without gastric ulcer. In case of gastric ulcer type I with superficial fundic gastritis it emerges normozymogenism with hyperpepsinogenemia++ I, with preatrophic fundic gastritis hypozymogenism with normopepsinogenemia, with atrophic fundic gastritis hypozymogenism with hypopepsinogenemia I. In type II and III gastric ulcer the chief cell mass and serum pepsinogen I behaviour as they do in duodenal ulcer with hyperpepsinogenemia although hypozymogenism is present.


Food in low-income families.

Food in low-income families. This descriptive study used both quantitative and qualitative research methods to examine the food and nutrient intake, food purchasing patterns and budgeting strategies of 29 sole-parent low-income families with dependent children living in Corio Shire, Victoria, in 1989-90. Expenditure on food and nonalcoholic beverages when compared with the average for all Australian households showed that the study families allocated a greater proportion, but similar amounts of money to cereals, dairy products, fruit and vegetables and miscellaneous foods, and less to meat, nonalcoholic beverages and food eaten away from home. Despite large differences in the amount of money spent in the first and second weeks of a social security payment period, the nutrient density of the parents' diets was maintained at a similar level in both weeks. The only exception was vitamin C, for which the median intake was significantly lower in the second week, consistent with reduced purchases of fruit and vegetables in the same period. In interviews with 13 of the parents a variety of purchasing and budgeting strategies were described, indicating common concerns with making the most of their limited resources. The data from this study support the notion that low-income families give priority to food purchases above other expenses such as recreation. They also support the view that low-income families are concerned about health and nutrition and manage to consume nutritionally sound diets under difficult circumstances. Nutrition education programs could support them in this endeavour.


Preventive health behaviours among parents of infants aged four months. Six preventive health behaviours have been frequently identified as having the potential to reduce mortality and morbidity during infancy: breast-feeding until the age of six months; no solid food until after four months of age; immunisation against whooping cough, diphtheria, poliomyelitis and tetanus; the use of a baby capsule to restrain the infant when travelling in a motor vehicle; regular attendance at a health care provider for preventive health checks; and no maternal smoking. This study surveyed 191 primiparous women four months after the birth of their babies to explore the proportion of parents who perform the recommended preventive health behaviours and the association among the behaviours. Thirty-nine per cent of the women reported that they were no longer breast-feeding by the time their infant was four months old; 35 per cent had introduced solids before 16 weeks of age; 35 per cent did not always use a baby capsule when travelling with their baby in their car, 25 per cent did not regularly attend the early childhood health centres and 22 per cent smoked. Forty-eight per cent of the sample were performing four or fewer of the six preventive health behaviours and 21 per cent were performing three or fewer. The relationship between performing each preventive health behaviour and a range of demographic variables was investigated. A logistic regression indicated that performing three or fewer of the health behaviours was associated with lower levels of education, having public health insurance and being born in a country other than Australia.


Coffee shops and clinics: the give and take of doing HIV/AIDS research with injecting drug users. We discuss recruiting and interviewing injecting drug users and using research as health promotion in the context of collecting information related to human immunodeficiency virus (HIV) or acquired immune deficiency syndrome (AIDS) from a convenience sample of 200 injecting drug users, half in treatment and half not, in 1989 and 1990 in Perth, Western Australia. A variety of recruiting methods were used including advertising, referral by agency staff, 'snowballing' and approaches to personal contacts and others known to inject by the interviewer. Snowballing and personal contacts were the most successful means of recruiting those not in treatment, while advertising was comparatively unsuccessful with this group because of the importance of establishing the credibility of the study and the interviewer among injecting drug users before they will volunteer to be involved. The promotion of behavioural risk reduction among respondents during the interview is detailed. We argue that the traditionally rigid separation between research and intervention is inappropriate in the HIV/AIDS context. When lives are potentially at stake, any contact with injecting drug users, especially those not in treatment (where may receive HIV/AIDS education), must be used as an HIV/AIDS prevention opportunity, and the interview is an ideal opportunity. The employment of research as community intervention is also discussed.


Self-help smoking cessation materials.

Self-help smoking cessation materials. Smoking-cessation campaigns and services use printed self-help materials as a major medium for assisting smokers to quit. Such materials are associated with abstinence rates of up to 20% at 12-month follow-ups. Adherence to the specific content of self-instructional programs can be poor, but higher levels of reported adherence may be associated with a greater likelihood of cessation. Personalized cessation materials do not enhance cessation rates, but social support and minimal levels of personal contact do. Research is needed to identify which specific components of these programs are effective, how adherence may be promoted and how materials may be used in conjunction with other interventions--particularly cost-effective forms of counselling and social support.


Sunglasses and clothing--an unhealthy correlation? Increasing awareness of health effects of solar ultraviolet radiation has focused attention on protection measures, including sunglasses. Sunglasses, or fashion glasses with tinted lenses, may be associated with some risks, and quality is not evident from casual inspection. This study investigates one possible 'hazard' of sunglasses--that they may induce behavioural changes which reduce the wearer's tendency to use adequate sun protection for the rest of the body. Findings consistent with such a proposition were shown in a photographic survey of 373 people in the summer of 1991 in Perth, Western Australia. This revealed maladaptive associations between the wearing of sunglasses and a high degree of leg or arm exposure as recorded by length of sleeves or trouser legs. This was significant for the arms and the legs in males (P less than 0.05), but approached significance only for the arms in females (P less than 0.10). There was no significant association between degrees of arm and leg exposure in either sex. The health implications of these maladaptive behaviours, and some advantages and disadvantages of the survey technique, are discussed. The results suggest that promotion of sun-protective behaviour should encourage the use of both sunglasses and protective clothing, as subjects' behaviours with respect to these do not appear to be correlated in a rational manner. If public awareness of links between skin cancer and eye disease could be increased, promotion of sunglasses might lead to a reduction in the incidence of skin cancers in general, not just those in the periorbital area.


The family v. the family court: sterilisation issues. Parents as guardians of minor children have the right and duty to give and withhold consent to medical treatment when the treatment is neither routine nor urgent. Parental authority, however, is not absolute and dwindles as the child gradually matures. In general, teenagers can give consent to medical treatment if they understand the nature and consequences of the proposed treatment. The diminution of parental authority is based on the premise that the child will eventually become autonomous. In cases where a sterilisation or hysterectomy procedure is being considered for a severely intellectually disabled teenager the question of consent is most contentious. Should this power belong to parents or the state? This paper examines some recent Family Court cases concerning this issue and also addresses questions about human rights, medical autonomy and the role of the Family Court. Finally, a proposal for an alternative means of decision-making in these cases is briefly outlined.


Otitis externa induced with Malassezia pachydermatis in dogs and the efficacy of pimaricin. Eight beagles were experimentally inoculated intraotally with Malassezia pachydermatis to induce acute otitis externa. Three or 4 days after the inoculation, the animals showed the symptoms of otitis externa. All ear canals were erythematous and the dogs were shaking their heads. A large number of M. pachydermatis was noticed in exudate taken from every ear canal. Clinical signs of otitis externa were reduced after treatment with 0.1 ml (per canal) of 1% pimaricin suspension twice a day for 3 days. The amount of exudate decreased gradually and 12 of the 16 ear swabs examined, thereafter, were found to be negative for M. pachydermatis within 10 days. No side effects were observed in all the treated cases. These results suggested that M. pachydermatis could induce the canine otitis externa, and that pimaricin is effective agent for M. pachydermatis infection in ear canals.


A balanced anesthesia with a combination of xylazine, ketamine and butorphanol and its antagonism by yohimbine in pigs. The effects of intramuscular injections of xylazine (2 mg/kg)-ketamine (15 mg/kg) [X-K15], and xylazine (2 mg/kg)-ketamine (5 mg/kg)-butorphanol (0.22 mg/kg) [X-K5-B] were compared in atropinized (0.05 mg/kg) miniature pigs (pigs). Both combinations induced the anesthesia for more than 1 hr, however X-K5-B induced the more potent and well balanced anesthesia as compared with X-K15, although the amount of ketamine was reduced to one third. The duration of loss of pedal reflex, an indicator of surgical anesthesia, in X-K5-B (62 +/- 13 min) was significantly (P less than 0.05) longer than in X-K15 (28 +/- 19 min). In addition, X-K5-B was accompanied by loss of laryngeal reflex in all pigs. Recovery from anesthesia in X-K5-B was much smoother than in X-K15, and the administration of yohimbine (0.05 mg/kg) could rapidly and smoothly reverse the anesthesia induced by X-K5-B, although it was accompanied by a transient fall in blood pressure and tachycardia. The combination of xylazine, ketamine and butorphanol appears to be a relatively safe and widely available anesthesia for the period of one hour in pigs.


Evaluation of plasma erythropoietin levels in normal adult dogs by in vivo bioassay using concentrated plasma. Measurement of plasma erythropoietin level in normal dogs by in vivo bioassay has been considered to be impossible so far. In the present study, we successfully determined it by using concentrated plasma 60 times which allowed the lower limit to 2.7 mU/ml. This normal plasma erythropoietin level was the first to be determined as an in vivo bioactivity and was 9.14 +/- 7.81 mU/ml in 75 normal adult dogs. This value was sufficiently reliable in terms of accuracy of determination and considered to be meaningful as the low level in vivo bioactivity that hasn't been known to date. Furthermore, erythropoietin levels in normal plasma were within a certain lower range and showed neither difference in plasma erythropoietin level between males and females or among breeds nor correlation between erythropoietin and hemoglobin level.


Role of adherent spleen cells in the induction of cytotoxic activity by Toxoplasma lysate antigen. In order to identify mechanisms responsible for the anti-tumor effects of Toxoplasma lysate antigen (TLA), we used an in vitro 51Cr release assay to study the functional properties of plastic-adherent cells during induction of splenic cytotoxic activity by TLA. Cytotoxic activity of non-adherent cells was measured in all experiments after a 6 days incubation. Induction of cytotoxic non-adherent cells by TLA required the presence of plastic-adherent spleen cells. In contrast, rhIL-2 alone was able to induce transformation of cytotoxic non-adherent cells from non-adherent spleen cells. Contact between adherent and non-adherent spleen cells was necessary for successful induction of cytotoxic non-adherent cells by TLA. Treatment of spleen cells with anti-macrophage serum prevented induction of cytotoxic activity by TLA. Biologically active IL-2 was not detected in culture supernatants of spleen cells exposed to TLA. These findings suggest that contact between TLA-sensitized non-adherent cells and macrophages is necessary for induction of cytotoxic cells in the presence of TLA. This contact, however, is not necessary for generation of IL-2-induced killer cells.


Effect of hyperlipidemia on cardiovascular responses to adrenergic stimulation in piglets. This study was designed to assess the effect of hyperlipidemia on cardiovascular responses to adrenergic stimulation in a porcine model. Four-week-old piglets (n = 10) were divided into two groups; one fed a control diet and the other was fed an atherogenic diet for 8 weeks. Cardiovascular responses were evaluated to both norepinephrine (NE; 0.5 and 2.5 micrograms/kg) and isoproterenol (ISO; 0.1 and 0.5 microgram/kg) from simultaneous recordings of femoral arterial pressure, heart rate, left intraventricular pressure and left intraventricular dP/dt. It was found that no significant difference in the baseline values of cardiovascular function was observed between the control and hyperlipidemic groups. However, in the hyperlipidemic group as compared with the control group: 1) arterial blood pressure responses to NE were significantly increased (P less than 0.05), 2) cardiac contractile responses to NE and ISO were significantly potentiated (P less than 0.05), and 3) reflex bradycardia in response to increasing arterial blood pressure did not occur. These findings indicate that hyperlipidemia can potentiate the cardiovascular responses to adrenergic stimulation, whereas reflex cardiovascular regulation is somewhat altered by hyperlipidemia. Conceivably, these observations may have relevance to the possible role of the mechanism of the interaction of hyperlipidemia and hypertension in atherogenesis.


Sedative effects of medetomidine in pigs. Sedative effects of medetomidine, a potent selective and specific alpha 2-adrenoceptor agonist, were evaluated in pigs using 5 different doses (30, 50, 80, 100 and 150 micrograms/kg of body weight) and compared with those of xylazine (2 mg/kg). Atropine (25 micrograms/kg) was mixed with both drugs to prevent severe bradycardia. All drugs were administered intramuscularly. Medetomidine at a dosage of 30 micrograms/kg produced more potent sedation than xylazine. The depth of sedation induced by medetomidine was dose dependent within the range from 30 to 80 micrograms/kg. At 100 or 150 micrograms/kg, the depth of sedation was mostly the similar level to that at 80 micrograms/kg but the duration was prolonged. The degree of muscle relaxation produced by medetomidine also seemed to be dose dependent from 30 to 80 micrograms/kg and was stronger than that produced by xylazine. An increase in the duration of muscle relaxation was dose dependent up to 150 micrograms/kg. No analgesic effect was produced by xylazine, however moderate analgesia was obtained by medetomidine. There were no marked changes in heart rate and respiratory rate during the observation period in pigs of any groups, however mild hypothermia after the administration of both drugs was observed. From these results, medetomidine has a significant and dose-dependent sedative effects which are much more potent than that of xylazine, and a combination of 80 micrograms/kg of medetomidine and 25 micrograms/kg of atropine is suitable for sedation with lateral recumbency and moderate muscle relaxation without notable side effects in pigs.


Serological survey of Rhodococcus equi infection in horses in Hokkaido. Serological survey of Rhodococcus equi infection in horses in Hokkaido was performed using ELISA. Of 2,879 horse sera, 318 (11.0%) gave antibody-positive (OD greater than or equal to 0.3) reactions. The antibody-positive rate of female was significantly higher (p less than 0.01) than that of male, and no statistical difference between Anglo-Arab and thoroughbred was detected in the antibody-positive rate. The maximum antibody-positive rate (27.1%) was shown at 14 years of age. The antibody-positive rates on the 160 farms were found to vary widely from 0 to 78.9%. A significant difference (p less than 0.01) in the antibody-positive rate was detected among horse farms. It was elucidated that 100 (62.5%) out of 160 horse farms had an antibody-positive horse. These results indicate that R. equi was widespread on horse farms, and the level of environmental contamination with R. equi differed among horse farms.


Effect of soft x-ray irradiation on immunological functions in mice. Effect of soft x-ray irradiation on immunological functions in mice was investigated. Soft x-ray irradiation with 100R or more induced a significant reduction in the number of plaque-forming cells (PFC). The reduction in the number of PFC depended on the irradiation doses. Irradiation with 600R or more showed a significant reduction in the delayed reaction of footpad swelling. However, soft x-ray irradiation with doses ranging from 100R to 1000R did not exert significant influence on the K values of carbon clearance test. Irradiation with 100R or more of soft x-ray showed a remarkable reduction of response to concanavalinA (ConA) or lipopolysaccharide (LPS) in spleen cells, and the response to ConA was lower than that to LPS. These results suggest that in the soft x-ray-irradiated mice, antibody-producing ability, delayed type hypersensitivity reaction and mitogenic activity are sensitive to soft x-ray irradiation and furthermore, T cell is more sensitive than B cell, but phagocytic activity of reticulo-endothelial system (RES) is resistant to soft x-ray irradiation.


Immunohistochemical studies on canine cerebral amyloid angiopathy and senile plaques. Amyloid protein was isolated from the cerebral meninges of 4 aged dogs with cerebral amyloid angiopathy. By immunoblot analysis, antiserum against synthetic oligo-peptide consisting of 1-28 amino acid of amyloid beta protein recognized prominent wide band ranging from 14 to 18 kilodalton (kd). When amyloid samples were solubilized by formic acid, the antiserum recognized lower molecular weight band ranging from 3 to 4 kd. Immunohistochemical studies on cerebral amyloid angiopathy and senile plaques were performed in 17 aged dogs. Anti-amyloid beta protein serum labeled amyloid deposits in cerebral vessel walls and senile plaques. Compact deposits of beta protein were detected in primitive or classical plaques. After using formic acid pretreatment, diffuse deposits of beta protein in the neuropil representing diffuse plaques were detectable. Classical and primitive plaques reacted with antiserum against glial fibrillary acidic protein, while not with antisera against alpha 1-antichymotrypsin, IgG and IgM. Amyloid deposits in the intestines of aged dogs examined, did not react with anti-amyloid beta protein serum.


Effects of atherosclerosis on mean and daily variation of arterial pressure in conscious WHHL rabbits. Effects of atherosclerosis on the mean value and daily variation of arterial pressure were studied in 12 Watanabe-heritable hyperlipidemic (WHHL) rabbits aged 12 to 35 months and 25 normal Japanese white rabbits aged 6 to 30 months. A pressure catheter was inserted through the left subclavian artery under pentobarbital anesthesia. A few days after the catheterization, the mean arterial pressure (MAP) of the rabbits, which were active and in a good state of appetite, was recorded by an analogue-to-digital converter every second for about 6 hrs and stored in a computer. The mean (M) and standard deviation (SD) in the WHHL rabbit, calculated from each successive MAP record, ranged widely from 85.8 to 131.4 mmHg and 5.6 to 12.6 mmHg, respectively. There was no significant correlation between M and SD in the WHHL rabbit. M and variance (V) of MAP in the WHHL rabbit were significantly higher than those in the normal rabbit. M did not show any significant change with increasing ages, whereas SD increased significantly with aging in the WHHL rabbit. Concentrations of serum total cholesterol and triglyceride in the WHHL rabbit were 475 and 328 mg/dl, which were about nine and seven times as high as those in the normal rabbit, respectively. Macroscopic and histopathological examinations of the aorta revealed development and spread of sclerotic lesions with aging in the WHHL rabbit. We can conclude that development of atherosclerosis with aging in the WHHL rabbit causes malfunction of the baroreceptors, which contributes to hypertension and lability of arterial pressure.


Biological and biophysical characteristics of phages isolated from Clostridium botulinum type C and D strains, and physicochemical properties of the phage DNAs. Toxin-converting phages CE beta and d-16 phi and non-converting phages CE gamma and d-1', isolated from toxigenic strains C-468 and D-CB16 of Clostridium botulinum types C and D, respectively, were characterized biologically and biophysically. DNAs isolated from these four phages were studied physicochemically. Phages CE beta, d-16 phi, CE gamma and d-1' were adsorbed to their susceptible cells at the constant rates of 1.1 x 10(-8), 3.3 x 10(-8), 1.4 x 10(-8) and 1.4 x 10(-8) ml/min, and grew after latent periods of 35, 45, 35 and 35 min at the burst sizes of 38, 85, 35 and 35, respectively. Converting phages were more susceptible than non-converting phages to physical and chemical treatments such as temperature, pH, time, UV-irradiation and organic solvents. GC% of phage DNAs were determined by melting temperature, buoyant density and HPLC analysis to be 26, 26, 29 and 29% for CE beta, d-16 phi, CE gamma and d-1' phage DNAs, respectively. The restriction digestion profiles of phage DNAs with seven endonucleases were compared by agarose gel electrophoresis, revealing that the two converting phage DNAs were similar in regard to five enzyme digestion profiles, but different in the other two enzymatic profiles. Non-converting phage DNAs produced the same restriction digestion profiles with all seven endonucleases. The molecular sizes determined from the size of restriction enzyme digestion fragments were about 110 kb for converting phage CE beta and d-16 phi DNAs and 65 kb for non-converting phage CE gamma and d-1' DNAs. Dot blot hybridization experiments revealed that DNA homology between the converting phages CE beta and d-16 phi was 50-75%, while that between non-converting phages CE gamma and d-1' was about 100%. Converting phage and non-converting phage DNAs did not hybridize at all.


Pathological studies on local tissue reactions in guinea pigs and rats caused by four different adjuvants. We investigated pathological changes at the injection site in guinea pigs and rats for 16 weeks following a single intramuscular injection of one of the following oil adjuvant emulsions; oil adjuvant ISA-70, Freund's incomplete adjuvant, Freund's complete adjuvant, and aluminium phosphate gel. In the animals injected with ISA-70 emulsion prepared by manual shaking, grossly, there was partial thickening of subcutaneous tissue, discoloration of inter-muscular connective tissue, and swelling of the inguinal lymph nodes at 2 and 4 weeks post injection (PI). Histopathologically, ISA-70 injected sites revealed acute inflammatory changes at 72 hrs PI, and peak reactions consisting of macrophage accumulation around oil cysts and fibrosis were observed at 4 weeks PI. These changes were less severe and of shorter duration than those in the other three adjuvants. Guinea pigs and rats injected with materials containing inactivated Newcastle disease virus (NDV) antigen similarly showed an infiltration of plasma cells and lymphocytes in addition to the changes described above. ISA-70 containing NDV antigen induced similar hemagglutination-inhibition titer to that induced by Freund's incomplete adjuvant.


Isolation and identification of intestinal bacteria from Japanese tree frog (Hlya japonica) with the special reference to anaerobic bacteria. The bacteria in the large intestines of eight Japanese tree frogs (Hlya japonica) were enumerated by using an anaerobic culture system. The microorganisms at approximately 3.1 x 10(9) bacteria per g (wet weight) of intestinal contents were present in the intestine of all the frogs tested. No difference of the total bacteria in the frog intestine was observed between two different incubation-temperatures (room temperature and 37 degrees C). Eleven genera and 16 species were isolated from the frog intestine. In most frogs, Bacteroides (B.) caccae and B. vulgatus were detected as the predominant organisms. Escherichia coli was also present in greater numbers in the intestine. Other bacteria isolated at high dilutions were strict anaerobes, including Fusobacterium and Clostridium. Enterococcus faecalis was frequently isolated from the frog intestine. However, four genera of Bifidobacterium, Eubacterium, Peptostreptococcus, and Lactobacillus were not isolated from the frog intestine.


Impact of the advances in age on the gastrointestinal microflora of beagle dogs. The gastrointestinal microflora of male beagle dogs in two different age groups; I) less than month 12 of age, and II) more than year 11 of age, was compared. No detectable difference occurred on the microflora of stomach, duodenum, jejunum, and ileum in both dogs. Large bowel (cecum, colon and rectum) microflora in both dogs yielded the different microbial populations. In all regions of large bowel, the levels of bacteroides, eubacteria, peptostreptococci, bifidobacteria, lactobacilli, and staphylococci in the elderly dogs were lower than those in the younger animals, whereas the numbers of Clostridium perfringens and streptococci in the elderly animals were higher than those in the youngers. The high incidence of lecithinase-negative clostridia was observed with advances in age, but not that of spiral shaped rods. The result of this study shows that the advances in age of beagle dogs yield some changes in the microbial population of large bowel in the animals.


Characteristics of ventricular activation and recovery patterns in the rat. Bipolar or unipolar epicardial and intraseptum electrograms were recorded from the in situ rat hearts with spontaneous sinus rhythm or myocardial electrical stimulation of 400 bpm in order to investigate the sequence and duration of cardiac activation in the rat. The ventricular epicardial isochronous maps in the rat showed the activation sequence comparable to those in humans and dogs, although the activation time (6 to 13 msec) of epicardium was approximately one-third of those of such species. The delay of activation between septum and ventricular surface in the rat was 5 to 8 msec, which was also shorter as compared with those in humans and dogs. However, the conduction velocity of the ventricular epicardium in the rat was approximately 40 cm/sec, mostly equal to that of humans. These findings might be accounted for by the possible presence of the bundle branch-purkinje system, at least functionally, similar to humans. The local QT interval in the septum were relatively longer (69 to 123 msec), compared with that of the ventricular surface (50 to 98 msec), and such durations of ventricular epicardium and septum were considerably less than those of dogs and humans. These results are consistent with the configuration of QRS-T on the limb lead ECG.


Isolation of secretory IgA from feline bile and bile IgA levels in growing cats. Secretory IgA was isolated from feline bile, using ammonium sulphate precipitation, gel-filtration chromatography and affinity chromatography. It formed a precipitating line between anti-cat whole serum or anti-cat bile serum by immunoelectrophoresis and double diffusion. It consisted of three subunits that molecular weight were 80 kd, 62 kd and 27 kd by SDS-PAGE analysis. The 80 kd protein was equivalent to secretory component of human and other animals. Immunoglobulin levels were observed on bile collected continuously by gallbladder cannulation in growing cats. Although immunoglobulins were not detected in bile of feline fetus, after birth its levels increased gradually. Biliary IgA levels reached the adult levels earlier than that of their serum.


Effects of estrous cycle, estrogen and progesterone administration on the antidromic and orthodromic reaction threshold of medial preoptic neurons in the rat hypothalamus. In the present study, attempts were made to evaluate the effects of the estrous cycle and the administration of estradiol (Ovx-E) or progesterone (Ovx-P) after ovariectomy on the threshold of antidromic (AD) and orthodromic (OD) responses of neurons in the medial preoptic area (POA) induced by electrical stimulation of the median eminence-arcuate nucleus (ARC) of the hypothalamus. Single units were evaluated with extracellular recording. The threshold of AD and OD response in POA neurons declined during estrus and proestrus of the estrous cycle and in the Ovx-E group, but increased in the Ovx-P group and during diestrus. The number of neurons with the inhibitory OD response increased during proestrus and in the Ovx-E group, but decreased during diestrus. The neurons with the excitatory OD response increased in the Ovx-P group and during diestrus, and decreased during proestrus. The threshold of the inhibitory and excitatory OD response decreased during proestrus, estrus and in the Ovx-E group, but increased in the Ovx-P group. It is apparent from these studies that estrogen appears to activate the neurons of the POA receiving inhibitory synaptic input via the axonal collateral branches of the ARC neurons, and that progesterone seems to activate the neurons of the POA receiving excitatory synaptic input.


Analysis of swinepox virus antigens using monoclonal antibodies. Seventeen monoclonal antibodies (MAbs) against swinepox virus (SPV) were produced and characterized. These MAbs were classified into eight groups (A through H) on the basis of the molecular weight of the polypeptides which they recognized and the staining patterns of antigens in SPV-infected cells by the indirect immunofluorescent (IF) technique. The MAbs belonging to groups A, B, C and G recognized late antigens in cytoplasmic inclusion bodies with molecular weights of 97 kD, 65 kD, 48 kD and 15 kD, respectively. The MAbs belonging to groups D and H respectively recognized 35 kD and 12 kD late antigens, which first appeared in cytoplasmic inclusion bodies and spread to the cytoplasms and surface membranes of the infected cells. The MAb of group F recognized an 18 kD late antigen with granular distribution in the cytoplasm. The MAbs of group E recognized a 32 kD early antigen. Although all the MAbs belonging to the six groups (A, D through H) were specific for SPV, some of those belonging to groups B and C showed cross-reactivity with members of the other genera of poxviridae. An MAb in group B, SP14, cross-reacted with orf and rabbit fibroma viruses. Two MAbs in group C, SP24 and SP32, cross-reacted with vaccinia, cowpox, ectromelia, and rabbit fibroma viruses. These findings indicate that at least two SPV antigens contain cross-reactive epitopes with different genera of poxviridae.


Relationship between pulmonary arterial pressure and lesions in the pulmonary arteries and parenchyma, and cardiac valves in canine dirofilariasis. The relationship between pulmonary hypertension and lesions was examined in 41 dogs infested naturally with heartworms, which consisted of 28 cases with pulmonary heartworm disease and 13 cases with caval syndrome. Pulmonary arterial pressure (PAP) was measured before and 1 or 7 days after heartworm removal with a flexible alligator forceps. In these dogs, lesions were examined after the last measurement of PAP. The mean PAP was 28.2 +/- 16.0 mmHg (10.9 to 81.4 mmHg in range) at post-removal phase. Pulmonary arterial intimal lesions, pulmonary thromboemboli, pneumonic lesions, tricuspid valvular lesions and mitral valvular lesions were macroscopically recognized in 95, 59, 39, 54 and 56% of cases, respectively. These lesions were classified by severity and the relationship with PAP was examined by the multiple correlation analysis. The multiple coefficient correlation was found the highest between PAP and thromboemboli, followed by mitral valvular lesion, tricuspid valvular lesion, and pneumonic lesion. There was no significant correlation between PAP and intimal lesions. The coefficient of determination showed the highest value in thromboemboli when one variable was used, and increased only very slightly when a variable of thromboemboli was added to those of other lesions. The cases with high PAP had fresh thromboemboli in large pulmonary arteries. From these evidences, it was concluded that thromboemboli following natural death of heartworm was the most important factor causing an increase in PAP and developing clinical signs in canine heartworm disease.


Cardiopulmonary function in dogs with serious chronic heartworm disease. Cardiopulmonary function was examined in 18 dogs with serious chronic heartworm disease showing ascites, subcutaneous edema, prostration, weakness, jaundice and so on. After surgical heartworm removal from the pulmonary arteries, 10 dogs recovered (surviving group), and 8 dogs died or were euthanatized because of poor prognosis (nonsurviving group). The number of live heartworms residing in the pulmonary arteries of the surviving group tended to be larger than that in the nonsurviving group. At necropsy, severe pulmonary arterial lesions such as thromboembolism including dead heartworms, proliferative and villous lesions and intimal hyperplasia were noticed in all dogs examined, and tended to be severer in the nonsurviving group. Heartworm-coiling around the tricuspid valve chord was found in 1 dog of the surviving group and 4 dogs of the nonsurviving group. Before heartworm removal, there was no significant difference in the mean pulmonary arterial pressure (MPAP) between the surviving and nonsurviving group. Right atrial pressure (v-wave) was higher, and the cardiac index (CI) was lower in the nonsurviving group. Arterial oxygen tension was lower in the surviving group than in the heartworm-free group, and it was lower in the nonsurviving group than in the surviving group. Carbon dioxide tension was lower in the surviving group than in the heartworm-free group. Bicarbonate concentration (HCO3-) was lower both in the surviving and nonsurviving groups than in the heartworm-free group. One week after heartworm removal, MPAP decreased (P less than 0.05), and CI and HCO3- tended to increase in the surviving group.


[Inhibition of enzymatic activity of alpha-thrombin by low molecular weight synthetic inhibitor]. The effect of the organophosphoric inhibitor, SA-152, on the fibrinogen-coagulating and TAME-esterase activity of bovine alpha-thrombin was studied. The irreversible inhibition constants (k11 = 1.1 x 10(4) M-1.min-1,Ki = 0.7 x 10(-4) M, k2 = 0.8 min-1 towards the coagulating activity and kII = 0.7 x 10(4) M-1.min-1, Ki = 0.3 x 10(-4) M, k2 = 0.2 min-1 towards the esterase activity) were determined. The SA-152 inactivated alpha-thrombin was dialyzed and incubated with 0.5 M and 2.5 M NaCl and 10 mM TAME. There was no reconstitution of activity of the SA-152 modified alpha-thrombin after dialysis and treatment with high concentrations of NaCl and TAME. Heparin interactions with the anion-binding site of the high molecular weight recognition center in the alpha-thrombin molecule did not significantly influence the values of the kinetic constants for the enzyme inhibition by SA-152. This finding is consistent with the hypothesis on the irreversible binding of SA-152 in the active center of the enzyme.


[Isolation and properties of three forms of phosphorylase and phosphorylase kinase from human skeletal muscle]. Three forms of phosphorylase (I, II and III), two of which (I and II) were active in the presence of AMP and one (III) was active without AMP, were isolated from human skeletal muscles. The pI values for phosphorylases b(I) and b(II) were found to be identical (5.8-5.9). During chromatofocusing a low molecular weight protein (M(r) = 20-21 kDa, pI 4.8) was separated from phosphorylase b(II). This process was accompanied by an increase of the enzyme specific activity followed by its decline. During reconstitution of the complex the activity of phosphorylase b(II) returned to the initial level. Upon phosphorylation the amount of 32P incorporated into phosphorylase b(II) was 2 times as low as compared with rabbit phosphorylase b and human phosphorylase b(I). It may be supposed that in the human phosphorylase b(II) molecule one of the two subunits undergoes phosphorylation in vivo. This form of the enzyme is characterized by a greater affinity for glycogen and a lower sensitivity to allosteric effectors (AMP, glucose-6-phosphate, caffeine) compared with phosphorylase b(I). Thus, among the three phosphorylase forms obtained in this study, form b(II) is the most unusual one, since it is partly phosphorylated by phosphorylase kinase to form a complex with a low molecular weight protein which stabilizes its activity. A partially purified preparation of phosphorylase kinase was isolated from human skeletal muscles. The enzyme activity necessitates Ca2+ (c0.5 = 0.63 microM). At pH 6.8 the enzyme is activated by calmodulin (c0.5 = 15 microM). The enzyme activity ratio at pH 6.8/8.2 is equal to 0.18.


[Comparative kinetic studies of the primary specificity of bovine and salmon trypsin]. A comparative kinetic analysis of Pacific salmon and bovine trypsins revealed that the former hydrolyzes p-nitroanilide-N,L-benzoyl-D,L-arginine (BApNA) with a far greater efficiency in comparison with bovine trypsin due to the decrease in Km. The inhibition constants for the BApNA hydrolysis by bovine and salmon trypsin with glycine, beta-alanine, L-lysine, L-arginine and benzamidine were determined. With an increase in the length of the hydrocarbon chain in the inhibitor molecule (i.e., in the order of glycine-beta-alanine-L-lysine) the inhibiting effect increased both with salmon and bovine trypsins. The Ki values for benzamidine and L-arginine appeared to be by one order of magnitude higher with salmon trypsin than with bovine trypsin. L-arginine was a much more effective inhibitor compared to L-lysine when both salmon and bovine trypsins were used.


[Substrate specificity of collagenolytic proteases from the hepatopancreas of the of the Kamchatka crab]. The substrate specificity of two isozymes of collagenolytic protease of the crab (Paralithodes camtschatica) was studied. It was found that both proteases can effectively hydrolyze type I and III collagens, as well as gelatin, the set of products yielded by enzymatic hydrolysis being different for isozymes A and C. Hydrolysis of some well-known peptides revealed that isozyme A predominantly cleaves the peptide bonds containing arginine and lysine residues, whereas isozyme C predominantly hydrolyzes bonds containing hydrophobic amino acids. The catalytic constants for the hydrolysis of several low molecular weight substrates in the presence of P. camtschatica proteases were determined, which allowed to attribute isozyme A to trypsin-like, and isozyme C to chymotrypsin-like proteinases. The peptide substrates of collagenase, Pz-Pro-Leu-Gly-Pro-D-Arg and Z-Gly-Pro-Ala-Gly-Pro-Ala are not hydrolyzed isozymes of crab collagenolytic protease.


[The role of terminal DNA groups in the activation of (ADP-ribose)polymerase]. Some peculiarities of activation of (ADP-ribose) polymerase by DNA fragments were studied. DNA fragments were produced by the digestion of calf thymus DNA by micrococcal nuclease and with a subsequent enzymatic modification of their end groups by nuclease S1, polynucleotide kinase of phage T4 and alkaline phosphatase. The dependence of the activating effect of DNA on the chemical structure of its end groups was established. It was shown that the terminal phosphate groups are involved in the formation of a catalytically active complex of (ADP-ribose) polymerase with DNA.


[Interaction of human thrombin I fragment, prethrombin I, and alpha-thrombin with tissue thromboplastin]. The binding of 125I-labeled prothrombin fragment I. prethrombin I and alpha-thrombin to native and papain-treated tissue thromboplastin in the presence of CaCl2 of EDTA was studied. The experimental curves plotted in the Scatchard coordinates testify to the presence in thromboplastin of two types of fragment I binding sites: those with a high (Kd = 7.6 x 10(-6) M) and moderate (Kd = 1.3 x 10(-8) M) binding affinity. The parameters of fragment I binding and their changes reproduced, for the most part, the mode of prothrombin binding observed in previous studies. The experimental results provide indirect evidence in favour of a hydrophobic role of Ca(2+)-dependent binding of prothrombin fragment I to thromboplastin. The binding of prethrombin I was nonspecific and Ca(2+)-independent, whereas alpha-thrombin showed a relatively high level of nonspecific electrostatic binding which was competitively inhibited by Ca2+. Thromboplastin proteins interacted (both directly and in a Ca(2+)-independent fashion) with all the prothrombin derivatives under study.


[Inhibition of pyruvate kinase of bovine adrenal cortex by ATP: the role of magnesium]. The effect of ATP on bovine adrenal cortex pyruvate kinase has been studied. ATP is a competitive inhibitor of the enzyme, the Ki being 3.2 mM. Based on the efficiency of tryptophan fluorescence quenching, it was concluded that the magnesium complex of ATP is a true enzyme inhibitor. The role of Mg2+ in the inhibition process consists in the formation of a bridge between the enzyme and ATP. The ATP-dependent mechanism of pyruvate kinase inhibition is a potential physiological regulator of the enzyme determining the lower threshold of its sensitivity in vivo.


[Gangliosides and antibodies to gangliosides in blood serum]. The concentration and composition of gangliosides from normal and pathological blood serum of animals and man are reviewed. Data concerning the elevation of the ganglioside content in the serum under malignization are summarized. The appearance of ganglioside-specific antibodies in some pathological states is described. The possible influence of changes in the serum ganglioside content and composition on immunity is discussed.


[Mechanism of inhibiting the cytochrome P-450-dependent monooxygenase system in liver with fluorocarbons]. The inducer of the liver monooxygenase system perfluorodecalin added to microsomes as a submicron emulsion forms an enzyme-substrate complex with cytochrome P-450. The K(app) values for the perfluorodecalin binding to cytochrome P-450 in microsomes isolated from the livers of control and phenobarbital-treated rats are 5 x 10(-5) M and 2.3 x 10(-6) M, respectively. Perfluorodecalin competitively inhibits the binding of substrates to cytochrome P-450 and decreases the rates of monooxygenase reactions. Perfluorodecalin extrusion from the active center of cytochrome P-450 occurs when an excess of perfluorocarbons non-interacting with cytochrome P-450 is added to microsomes. There is a significant vagueness in the rates of various monooxygenase reactions because of simultaneous induction and inhibition of monooxygenase enzymes after perfluorodecalin administration to rats. The data obtained are consistent with the hypothesis that constitutive forms of cytochrome P-450 are primary receptors for xenobiotic-inducers of phenobarbital-type cytochrome P-450 isoforms.


[Purification and properties of GTP-cyclohydrolase from Bacillus subtilis]. Highly purified GTP-cyclohydrolase was obtained by fractionation of cell extracts with ammonium sulfate, ion-exchange and hydrophobic chromatography. The N-terminal amino acid sequence and amino acid composition of the protein were determined. According to SDS-PAGE data, the molecular weight of the enzyme is 45 kDa. The active enzyme has several isoforms separable by native electrophoresis. The maximal enzyme activity is determined at 1.5 mM Mn2+; 70% of enzymatic activity is detected with Mg2+. The enzyme is inhibited by heavy metal ions and chelators and is inactive in the absence of thiol-reducing agents. The enzyme activity is detected in a broad range of pH with a maximum at pH 8.2. The pyrimidine product of the GTP-cyclohydrolase reaction. 2.5-diamino-6-hydroxy-4-ribosylaminopyrimidine-5'-phosphate was purified and identified. Another product of this reaction is pyrophosphate.


[Rat liver antioxidant defense mechanism to the action of aromatic amines]. The effects of diaminobiphenyl, biphenylamine and tetraaminobiphenyl on lipid peroxidation and antioxidant protective mechanisms in the subcellular fractions of rat liver have been studied. It was found that activation of lipid peroxidation plays a crucial role in the manifestation of hepatotoxic activities of diaminobiphenyl and biphenylamine, this effect being due to the decrease of the protective activity of the antioxidant system during intoxication by these compounds. Tetraaminobiphenyl does not influence the rate of lipid peroxidation. It is concluded that structural differences determine the differences in the mechanisms of adaptation of the antioxidant system to the effect of aromatic amines.


[The binding of SSB-proteins with DNA in chromatin of Ehrlich ascites carcinoma cells]. Using UV-induced cross-linking between proteins and DNA, the contacts between single-stranded DNA-binding proteins (SSB proteins) and chromatin DNA have been demonstrated. Ehrlich ascites tumour DNA was labeled in vivo by inoculation of tumour-bearing mice with 3H-thymidine. The cells were irradiated with the UV light dose of 3000 J/m2, destroyed in a Triton X-100-containing hypotonic medium, and separated by centrifugation into the extrachromatin fraction and chromatin. Chromatin DNA was digested with DNAase 1, and the chromatin proteins were extracted with 2 M NaCl-polyethyleneglycol. SSB proteins from the extrachromatin fraction and chromatin were purified. Only SSB proteins from UV-irradiated cell chromatin appeared to possess a high specific radioactivity which exceeded 7.5-fold that of non-irradiated cells. There were no differences between chromatin SSB proteins in control and irradiated cells as could be evidenced from SDS electrophoresis data. It is assumed that in irradiated cells SSB proteins of DNA-digested chromatin are covalently cross-linked with DNA fragments.


[Modification of a disulfide in the hinge region of rabbit IgG and study of the interaction of polyvalent ferritin antigen, protein A, and anti-IgG]. Since immunoglobulins are used in a vast variety of immunoassays, the problem of obtaining antibodies with enhanced antigen-binding activity is of great importance. In order to discriminate between putative approaches to activating antibody modification, some functional characteristics of rabbit IgG modified at the hinge disulfide by three reagents: iodoacetamide, N-ethylmaleimide, and 2.2'-dipyridyl disulfide, have been studied. As can be judged from gel-permeation chromatography data, the molecular sizes of modified rabbit IgG were slightly increased in comparison with the native protein. Using enzyme immunoassay, it was shown that modification by each of the above reagents results in the same degree of activation of the antibody binding to the protein polyvalent antigen-human ferritin, due to the increase in segmental flexibility, i.e., Fab motion around the Fo fragment of IgG. The type of concentration dependencies of antigen binding suggest that another determinant stimulating the antigen binding in addition to the increase in segmental flexibility, can be attributed to intra- or interdomain flexibility of domains constituting the Fab fragments. Using protein A and anti-IgG as conformational probes for the antibody Fo fragment, the conformation and conformational dynamics of both CD2 domain epitopes and the switch region between the CD2 and CH3 domains have been shown to be essentially unaffected by modification.


[Analysis of the quaternary structure of secreted repressible acid phosphatase from the yeast Saccharomyces cerevisiae]. The structural organization of extracellular repressible acid phosphatase from S. cerevisiae has been studied. The existence of multiple acid phosphatase forms with isoelectric points at pH 4.1-4.8 has been confirmed by isoelectrofocusing. The molecular masses of three acid phosphatase isoforms (56, 57-59, and 60 kDa) obtained after enzymatic deglycosylation correlate with the data obtained previously during the analysis of translation products in cell-free systems. Electron microscopic studies revealed that the acid phosphatase molecule has a square shape and is made up of four identical subunits with molecular masses of about 125 kDa.


[Polyfunctional enzymes].

[Polyfunctional enzymes]. The properties of enzymes possessing several autonomous functions distributed among the domains of the polypeptide chain are considered. Three types of such proteins have been characterized: (i) bifunctional enzymes catalyzing consecutive reactions of metabolism: (ii) enzymes capable of catalyzing different reactions that are not consecutive and, (iii) enzymes catalyzing oppositely directed reactions. Possible advantages of various reactions catalyzed by the same protein are discussed.


[Successes in studying the physiological activity of terpenoids and steroids]. Data concerning the structure and function of prenyl proteins and peptides of fungi and animals are reviewed. There exist proteins that are posttranslationally modified by thioether-linked farnesyl- or geranylgeranyl groups; the modification affects the cysteine residue near the C-terminus. Prenylation increases hydrophobicity and is expected to promote protein binding to membrane lipids or other hydrophobic proteins. Isopentenyl-adenylated proteins mediate isoprenoid control over DNA synthesis and are involved in regulation of cell proliferation in animals. Sterols of fungi and plants play a role in regulation of the membrane structure as well as in proliferation. The significance biologically active terpenoids for biotechnology, medicine, and agriculture is discussed.


[Structure-functional properties of eukaryotic aminoacyl-tRNA synthetase]. Data on structural and functional peculiarities of eukaryotic aminoacyl-tRNA synthetases (structure, supermolecular organization, and localization in eukaryotic cell) are reviewed. The functional significance of aminoacyl-tRNA synthetase association with high molecular weight complexes and other cellular components is discussed.


[Interaction of lactate dehydrogenase with structural cell components: possible physiological significance]. Reversible binding of cytoplasmic enzymes to structural elements of the cell is one of mechanisms of in vitro regulation of enzyme properties. The results on lactate dehydrogenase interactions with myofibrillar proteins and membranes and the changes in enzyme properties induced by these interactions (modification of kinetic parameters and stability) are analyzed. A hypothesis is proposed concerning the functional role of reversible lactate dehydrogenase interactions with structural components of the cell during glycolysis activation.


[Nuclear proteoglycans from mouse liver cells: isolation and identification]. Proteoglycans (PG) have been isolated from mouse liver nuclei and identified. Nuclear PG are represented by various classes: i) PG containing dermatan sulfate (DS) chains; ii) PG containing heparan sulfate (HS) chains and, apparently, iii) mixed PG whose protein core contains both DS and HS chains.


[Protein synthesis during hyperthermia in ground squirrels]. The intensity of [14C]leucine incorporation into heart, liver, brain, muscle, and blood plasma protein in gophers under deep artificial hypothermia has been studied. It was shown that the intensity of protein synthesis decreased sharply under these conditions. Insignificant incorporation of [14C]leucine into proteins was observed during the first two hours after the onset of hypothermia and then ceased completely.


[Lipid biosynthesis and metabolism of native and acetylated low density lipoproteins in macrophages stimulated by zymosan in vivo and in vitro]. The effects of zymosan on lipid metabolism in mouse peritoneal macrophages (MPM) in vitro and in vivo were studied with special reference to the following parameters: i) 14C-oleate incorporation into cholesteryl esters (CE), triglycerides (TG), and phospholipids (PL) in MPM incubated with low density lipoproteins (LDL) and acetylated LDL; ii) cholesteryl-14C-oleate-acetyl LDL uptake and 125I-acetyl LDL degradation; iii) oxidative modification of LDL. Zymosan administered to mice caused significant stimulation of 14C-oleate incorporation into CE, TG, and PL with no effect on 3H-cholesterol (Ch) incorporation into CE or 3H-glycerol incorporation into TG and PL in MPM. The 14C-oleate incorporation into cellular lipids was unaffected by 18-hour incubation of MPM with zymosan (100-500 micrograms/ml) but increased after incubation of unstimulated MPM with blood serum and peritoneal fluid harvested harvested from zymosan-treated mice. One possible explanation of this phenomenon is oleyl-CoA formation induction in cytokine-stimulated MPM in vivo. Zymosan decreased the Ch-14C-oleate-acetyl LDL uptake, 125I-acetyl LDL degradation, and Ch esterification in the presence of acetyl LDL in MPM both in vitro and in vivo. An increase in Ch esterification after incubation of MPM with zymosan for 6-18 hours in the presence of LDL was accompanied by an increase in lipid peroxidation of LDL and its electrophoretic mobility. The data obtained suggest that the macrophage acetyl LDL receptor pathway may be inhibited by zymosan and that cytokines released from zymosan-stimulated cells may influence the generation of foam cells.


[Nonphysiological redox-agents are reduced at the binding center of NADP(H) glutathione reductase]. Studies of the acceptor reductase reaction of yeast glutathione reductase (EC 1.6.4.2) revealed that the competitive inhibitors for NADPH, 2',5'-ADP and Br- decrease the rate constants for the enzyme oxidation by ferricyanide, phenanthrene quinone, and juglone. A similar effect is observed when NADH which does not bind to the reduced enzyme is used as substrate. These observations support the hypothesis that non-physiological redox agents are reduced at the NADP(H)-binding center of glutathione reductase and that NADP(H) binding stimulates the reaction by displacing tyrosine-197 which protects FAD from the solvent.


[Selective solubilization and biochemical analysis of R. prowazekii outer membrane proteins]. Solubilization of proteins from total membranes (a mixture of cytoplasmic and outer membranes) of Rickettsia prowazekii, a typical gram-negative bacterium, was studied using three different detergents. It was shown that isolated outer membranes and sarkosyl-insoluble material contain major polypeptides of 134, 31, 29.5 and 25 kDa as well as minor polypeptides of 78, 60, 42, and 17 kDa, while the total membranes--the same plus a great number of additional minor proteins. The material solubilized by octyl glucoside in the presence of MgCl2 contains exclusively major proteins (134, 31, 29.5, and 25 kDa). No differential solubilization takes place upon membrane treatment with octyl glucoside in the absence of Mg2+ or with Triton X-100. Rickettsial proteins are insensitive to trypsin in both whole cells and total membranes, unless the latter are presolubilized with octyl glucoside. Proteinase K degrades all of the total membrane proteins but only the 134 kDa polypeptide of whole cells. Upon immunoblotting predominantly the major outer membrane proteins (134, 31, and 20.5 kDa) and, to a lesser extent, the minor proteins (60, 42, and 17 kDa) interact with human convalescent serum.


[Serine proteinase from the archaebacterium Halobacterium mediterranei--an analog of eubacterium subtilisin]. A homogeneous serine proteinase was isolated from cultural filtrates of the extreme halophilic bacteria Halobacterium mediterranei 1538 using affinity chromatography on bacitracin-Sepharose, ultrafiltration and gel filtration on Sephadex G-75, with a 48% yield and 260-fold purification. The enzyme was completely inactivated by specific inhibitors of serine proteinases, PMSF and DFP, as well as by Hg2+ and PCMB. The enzyme activity was strongly dependent of NaCl concentration, the enzyme being inactivated below 0.75 M NaCl. Inactivation of the enzyme was also seen in the presence of 2-7% organic solvents. The pH optimum for Glp-Ala-Ala-Leu-pNA hydrolysis is 8.0-8.5; Km is 0.14 mM, kcat is 36.9 s-1. The stability optimum lies at pH 5.5-8.0, temperature optimum is at 55 degrees C. The enzyme molecular weight is 41,000 Da; pI is 7.5. The substrate specificity of the enzyme is comparable to that of secretory subtilisins; the extent of protein substrate hydrolysis is similar to that of proteinase K. The N-terminal sequence of Halobacterium mediterranei serine proteinase, Asp-Thr-Ala-Asn-Asp-Pro-Lys-Tyr-Gly-Ser-Gln-Tyr-Ala-Pro-Gln-Lys-Val-Asn- Ala- Asp-, reveals a 50% homology with the aminoterminal sequence of Thermoactinomyces vulgaris serine proteinase. Hence, the serine proteinase secreted by halophilic bacteria may be considered as a structural and functional analog of eubacterial enzymes.


[Thyroid hormone conjugates with rhodamine B as fluorescent ligands of human plasma transport proteins]. Conjugates of thyroxine (T4) and triiodothyronine (T3) with rhodamine B in which the hormone and the fluorescent dye are linked via a thiourea bond have been synthesized. These conjugates possess an ability to inhibit in a competitive manner the binding of [125I]T4 to three protein preparations: T4-binding globulin (TBG), apolipoprotein A-I (ApoA-I), and high density lipoprotein particles (ApoA-I-HDL) isolated from human serum by T4-Sepharose 4B chromatography and further purified. The following values of association constants have been estimated: for the T4 derivative-3 x 10(7) M-1 (TBG), 4.1 x 10(5) M-1 (ApoA-I), and 4.2 x 10(5) M-1 (ApoA-I-HDL); for the T3 derivative-1.6 x 10(7) M-1 (TBG), 5.3 x 10(5) M-1 (ApoA-I), and 5.4 x 10(5) M-1 (ApoA-I-HDL). The binding of rhodamine B-labeled thyroid hormones to TBG or ApoA-I do not alter significantly the parameters of rhodamine B chromophore absorption and fluorescence. The interaction of the conjugates with ApoI-HDL leads to a significant enhancement of the absorption intensity and a 3 nm blue shift in the absorption maximum as well as to a 1.5-fold increase in the fluorescence band amplitude at 586 nm. Biological and fluorescent properties of T4 and T3 derivatives suggest that these compounds may be a useful tool in fluorescence studies of plasma binding protein-driven transport of thyroid hormones in model biological systems.


Cell proliferation in the small intestine and colon of intravenously fed rats: effects of urogastrone-epidermal growth factor. There is marked intestinal hypoplasia in the intestine of intravenously fed (TPN) rats. Recombinant urogastrone-epidermal growth factor (URO-EGF) reversed these changes by significantly increasing the length of the intestinal crypts. Crypt diameter, however, was not affected to the same extent. Few differences in labelling indices were seen between the orally fed and TPN groups, however, this was the consequence of the concomitant changes in crypt population. The number of mitoses and labelled cells per crypt, and thus the crypt cell production rates, were significantly decreased in the TPN group when compared to the orally fed. URO-EGF significantly increased both proliferative indices and the number of dividing cells per crypt. Crypt cell production in the small intestine was restored to those levels seen in the orally fed rats, moreover, labelling per crypt in the colon was increased to more than twice that of orally fed rats. The location of the mean labelling position and the half maximum labelling position followed the changes in crypt length in the small intestine, but to a lesser extent; thus the growth fraction was significantly increased in the TPN rats in comparison with the orally fed and the URO-EGF treated groups. Similar changes in these positions were seen in the colon, but the growth fraction was much reduced in the URO-EGF treated rats, as a consequence of the large increase in crypt length without a concomitant alteration in label distribution.


Altered proliferative kinetics in PHA-activated human T-lymphocytes treated with the anti-HLA class I monoclonal antibody 01.65. Anti-HLA class I monoclonal antibody (mAb) 01.65 inhibited phytohaemagglutinin (PHA)-induced human lymphocyte proliferation. The inhibitory effect was inversely correlated to the strength of the proliferative response. It was increased when lymphocytes were stimulated with suboptimal doses of PHA but it disappeared with supraoptimal doses. Proliferation inhibition was achieved by prolonging the cell cycle time and by slowing down its recruitment rate. The former effect was not restricted to the G1-phase but also included the S phase. These results support the idea that HLA class I molecules are important in the PHA-induced proliferation of human T-lymphocytes.


Proliferative changes in two murine tumours in response to single or fractionated doses of X-rays. Studies were carried out to investigate proliferative changes in two murine experimental tumours in response to radiation. Results were generated using bromodeoxyuridine labelling and flow cytometry. This study demonstrates the possible ambiguity of previous studies using tritiated thymidine in which inability to discriminate normal and tumour cell components in murine tumours may lead to different values for cell kinetic parameters. In particular, the sarcoma F appeared to have a growth fraction of 0.62 when all cells were considered; in reality the growth fraction of the tumour cells only (based on DNA content discrimination) was close to unity. Radiation, administered either as single or fractionated doses, caused little change in the proliferative characteristics of the sarcoma F tumour but had profound effects on the adenocarcinoma Rhodesia tumour. Major changes were the accumulation of cells in G2 for several days after the end of the radiation treatment in both tumours and a dramatic drop in labelling index of the Rhodesia tumour. In neither tumour was there any evidence to suggest an increase in tumour cell proliferation during or after the irradiations. The diploid cells within the sarcoma F tumour showed an initial depression of labelling index followed by a rapid increase overshooting the control labelling index at higher radiation doses. Much of the effects could be attributed to cell cycle delays.


Cell cycle analysis of asexual stages of erythrocytic malaria parasites. Intra-erythrocytic Plasmodium species can be stained with the DNA binding dye, Hoechst 33342, and the distribution of DNA content determined for parasite populations by flow cytometric measurement of fluorescence. Analysis of this distribution will determine the parasitaemia (percentage of erythrocytes infected), and the percentages of trophozoite infected red blood cells, polyparasitized (trophozoite) red blood cells, and schizont/segmenter infected red blood cells. This analysis is based on the hypothesis that the asexual parasites cycle with single G1 period, and effectively, a single S phase with no significant G2/M period except at schizogony when the genome DNA content is equivalent to 8 N or higher, dependent on the species. Data are presented to support this model.


Growth fraction measured using the comet assay. Growth fraction, an important determinant of tumour response to therapy, was measured using a novel assay in WiDr human colon carcinoma cells grown as monolayers, spheroids, or xenografts. The assay is based on the fact that the anticancer agent etoposide produces DNA strand breaks in proliferating but not non-proliferating cells. Strand breaks were detected in individual cells using the alkaline 'comet' assay, and growth fraction was defined as the fraction of cells containing elevated numbers of DNA strand breaks. The specificity of the method for detecting proliferating cells was verified directly by allowing cells to incorporate bromodeoxyuridine (BrdUrd) into DNA, followed by exposure to etoposide and treatment of the comets with anti-BrdUrd antibodies. All cells stained with anti-BrdUrd antibodies were also damaged by etoposide. Similarly, growth fraction measured using Ki-67 correlated well with the new assay. The accuracy, speed and convenience of the comet assay for measuring growth fraction suggest that it may be useful for predicting response of human cancers to therapy.


Systemic administration of recombinant interleukin-6 in mice induces proliferation of lymphoid cells in vivo. Interleukin-6 (IL-6) is a pleiotropic cytokine and has been shown to support the growth of T and B lymphocytes in the presence of mitogens in vitro. IL-6 can induce human natural killer (NK) and interleukin-2 (IL-2)-induced lymphokine-activated killer cell (LAK) activity in vitro. It can also mediate antitumor effects in various murine models. In order to understand the mechanism of in vivo action, we have investigated the proliferation of lymphoid cells in vivo and the effects on NK, and LAK cell activities in response to IL-6 administration in mice. In vivo proliferation was measured by labeling the DNA of dividing cells with [125I]iodo-2'-deoxyuridine. C57BL/6 mice were injected ip with either IL-6 or HBSS control two times a day for 3 days and in vivo proliferation was measured. For comparative purpose IL-2 was administered and in vivo effects were analyzed. IL-6 caused significant proliferation of cells mainly in the spleen, while, IL-2 caused proliferation in the lungs, liver, spleen, and kidneys. Pretreatment irradiation (500 rad) of mice abrogated the IL-6-induced proliferation indicating radiosensitive cells are involved. Furthermore, in vivo proliferation was not observed in young nude mice treated with IL-6. To investigate whether the proliferating cells were cytotoxic, we tested for LAK (vs. fresh MCA-102 tumor targets) and NK (vs. Yac-1 tumor targets) activities in the organs of mice treated with IL-6 or IL-2 by 4 h 51Cr-release assay. IL-2 administration induced the generation of LAK activity and increased NK cytotoxicity in various organs, but IL-6 had no effect.(ABSTRACT TRUNCATED AT 250 WORDS)


Employment of antipeptide antisera to distinguish two closely related cytokines induced in human T cells. AT464 and AT744 are two cytokines encoded by mitogen-induced genes from human T lymphocytes. These proteins belong to a family of structurally related chemotactic proteins associated with inflammation. By expression of their full length cDNAs in COS cells and in baculovirus-infected Sf9 cells, we found that pAT464 and pAT744 cDNAs represent secreted polypeptides of a molecular mass of about 8,000 and 11,000 Da, respectively. Biochemical characterization of these proteins has been persued through polyclonal antisera, which were derived to synthetic peptides. Using these sera for Western blotting analyses the recombinant AT464 and AT744 proteins could be detected as secreted products from transfected COS cells and from baculovirus-infected Sf9 cells. Similarly the native proteins could be detected in the supernatants of activated human peripheral blood T cells. The recombinant and the T cell-secreted AT464 and AT744 proteins appear to be identical as judged by their mobility by SDS-PAGE and by their reactivity with the rabbit polyclonal antisera.


Adhesion molecules on MHC-nonrestricted lymphocytes: high density expression and role in oncolysis. We have shown that interleukin-2 (IL-2) -activated adherent lymphocytes (A-LAK) display superior oncolytic activity against acute myelogenous leukemia (AML) blasts when compared to conventionally prepared lymphocytes with lymphokine-activated killing (LAK) activity. The A-LAK activity was generated promptly and from donors whose lymphocytes did not display any LAK activity. In comparison to LAK, a higher percentage of A-LAK expressed the CD25 and HLA class II (HLA-DR) activation-associated structures and high density of HLA class I antigens. Most striking, however, was the observation that lymphocytes from A-LAK cultures consistently contained a high density of CD2, CD11a, and CD18 adhesion molecules, as indicated cytometrically by their "bright" fluorescence intensity. Three color flow cytometric analysis indicated that virtually all CD56+,CD3- natural killer (NK) and CD56+,CD3+ T cells in unstimulated, LAK and A-LAK populations displayed this "bright" phenotype, while most CD56-,CD3+ T cells (with the exception of the small proportion found in A-LAK) were of the "dim" phenotype. The A-LAK cultures also contained a higher percentage of lymphocytes expressing CD11b (CR3 receptor) and CD54 (ICAM-1) antigens. The CD11a, CD18, and partially CD2 molecules were important in the A-LAK cytolytic mechanism against AML, since blocking of these structures with monoclonal antibodies (MAb) significantly decreased the antileukemia effect. Additionally, the ability of A-LAK to adhere to plastic was most strongly inhibited by anti-CD11a MAb and less, but significantly, by MAb against CD2, CD18, and CD56 molecules.(ABSTRACT TRUNCATED AT 250 WORDS)


Interleukin-6 production by human melanoma cell lines. We examined the expression of interleukin-6 (IL-6) by 12 established human melanoma cell lines. Two constitutively produced low levels of IL-6 protein, as measured by enzyme-linked immunosorbent assay. Cells from these two lines, as well as those from two non-IL-6-producing cell lines, contained IL-6-specific mRNA as demonstrated by Northern hybridization. Treatment of the two IL-6-producing melanoma cell lines with interleukin-1 beta, tumor necrosis factor-alpha, or phorbol myristate acetate caused a marked increase in IL-6 production. These induction signals failed to stimulate IL-6 production in the nonproducing cells, even those that expressed IL-6 mRNA. IL-6 did not appear to act as an autocrine growth factor since the addition of exogenous human recombinant IL-6 or polyclonal anti-IL-6 antibody did not alter cellular proliferation. The production of this multifunctional cytokine by tumors may play a role in tumor-host interactions and this should be recognized in the design of biologic therapy trials.


Regulation of freshly isolated, IL-4-producing T cells: IFN-gamma-sensitive and -resistant cells. Cytokine responsiveness of interleukin-4 (IL-4)-producing T cells (IL-4p) during primary in vitro stimulation was investigated. Freshly isolated T cells were stimulated with anti-CD3 epsilon antibodies in the presence of macrophages. Using limiting dilution analysis, we found that IL-4p were not detected when endogenous IL-2 activity was blocked with anti-IL-2 antibodies. These data support previous observations that IL-4p require IL-2 for induction, and further indicate the presence of macrophages was not sufficient to overcome the requirement for IL-2. The effects of IFN-gamma on IL-4p were studied in this system. Addition of exogenous IFN-gamma decreased the frequency of IL-4p but did not completely inhibit induction of these cells. Furthermore, blocking endogenous IFN-gamma during stimulation resulted in a 2- to 4-fold increase in the frequency of IL-4p detected. These data demonstrate the novel finding that both IFN-gamma-resistant and IFN-gamma-sensitive IL-4p were present in freshly isolated T cell populations. Thus, during early in vitro culture, there were at least two types of IL-4p, both of which require IL-2. These data indirectly support a current model of Th ontogeny as defined by cytokine production patterns, and extend the model to include cytokine responsiveness.


Use of biodegradable plates and screws in a rabbit model. During the last decade, rigid internal fixation with miniplates and screws has gained widespread acceptance in the correction of both congenital and acquired craniomaxillofacial deformities. Recent studies have proposed that the currently employed metallic plates and screws may require removal because of potential facial growth restriction in growing children. Others have reported bone resorption under the plate due to stress shielding, infection, extrusion, and palpability in regions where there is minimal tissue coverage. Because these implants are radiopaque, they generate significant that interfere with radiological studies and with radiation therapy in patients undergoing treatment for malignancies. There is no question that the use of a biodegradable plating system would eliminate each of these potential or real problems, because stability is necessary only for a reasonably short period until the fracture segments have become united. We report the initial phase of a long-term study examining various materials that will be available for fabrication of a biodegradable plate and screw system. We evaluated a commercially developed biodegradable plate and screw system to treat zygomatic arch fractures in a rabbit model. Fractures were surgically created at the midpoint of each zygomatic arch. The experimental animals were then divided into three equal groups. Fractures in the first group were permitted to heal without any form of stabilization. In the second group, segments were secured with standard titanium plates and screws. Biodegradable plates and screws were employed for stabilization in the experimental group. Animals were then killed, and radiographs were obtained at 2, 4, 6, and 8 weeks.(ABSTRACT TRUNCATED AT 250 WORDS)


Effects of nerve growth factor on craniofacial onlay bone graft survival: preliminary findings. A study was undertaken to evaluate the effects of nerve growth factor (NGF), which had previously demonstrated bone formation around mandibular nerve regenerates, on resorption of onlay bone grafts to the rabbit skull. Iliac grafts were onlayed on the parietal bone bilaterally and immobilized with wire ligatures in 15 animals. NGF and a control solution were introduced into the grafts through subcutaneous osmotic pumps. After 60 days, graft survival was compared by weights, histology, and fluorochrome labels. NGF-treated grafts were characterized by an average of 88.4% weight retention, preservation of the outer cortical plate with firm fixation to the underlying calvarium, and fluorescent labels localized to the inner cortex. Conversely, control grafts had an average weight retention of 42.8% (p < 0.01), nearly complete outer cortical plate loss with increased mobility of the residual inner cortex, and a lack of fluorescence in any part of the grafts. Despite the incongruous name, NGF exhibited a beneficial effect on onlay bone graft volume maintenance in this study. Although the mechanism and the long-term effects are unknown, inhibition of graft resorption rather than enhancement of bone formation is suggested.


Clinical experiences with patients with facial bone deformities associated with hemifacial microsomia. Eighteen adult patients with hemifacial microsomia were treated with a combination of skeletal and augmentation surgery. Three typical cases are presented. In principle, skeletal and augmentation surgery have recently been performed in combination in a single stage. Groin flaps and scapular or scapular ostocutaneous flaps have mainly been employed for augmentation surgery.


Maternal discourse features used with language-normal preschoolers with facial deformities. Discourse features of 10 mothers of language-normal preschool children with repaired cleft palates were examined. Mothers had been provided training in infant stimulation. Fifty-six percent of the communication initiations within the mother-child dyads were made by the mothers. That level of responsibility for continuity of communication resembles the percentage of discourse initiation reported for mothers of language-normal children without orofacial clefts. All categories of utterance types were used by the subjects; a predominance was found among attention-getters and indirect directives. Distribution of discourse features appears to differ from that reported for mothers of both language-normal and language-impaired children who do not have orofacial clefts.


Surgical correction of unilateral lambdoid synostosis: occipital rotation flap. Virchow (1851) is credited with initially describing calvarial development following premature closure of individual sutures. He noted that resultant compensatory growth occurred perpendicular to the fused suture, resulting in asymmetrical calvarial development. Lambdoid synostosis, whether unilateral, bilateral, or associated with other cranial fusions, is reasonably uncommon. Previous authors report an incidence of less than 10% of all reported cases of premature fusion of the calvarial or skull base sutures. We report 27 patients (20 boys, 7 girls) ranging in age from 3 months to 8 years with unilateral lambdoid craniosynostosis followed over a three-year period. In 19 patients, calvarial reshaping was performed by repositioning a parietooccipital bone flap stabilized with lag-screw fixation to provide an increased radius of curvature to the affected area and to reestablish the resultant craniectomy between the parietooccipital junction and into the posterior fossa retromastoid occipital bone as an appropriate site for growth. Indications for one-stage calvarial reshaping included untreated lambdoid stenosis in older children or a failed simple lambdoid synostectomy after approximately 18 months. Each child experienced significant improvement in calvarial shape and ipsilateral ear position. Although we had no operative complications, two children underwent a second outpatient procedure for removal of hardware palpated by their parents.


The course of the inferior alveolar neurovascular canal in relation to sliding genioplasty. The anterior course of the inferior alveolar neurovascular canal was determined in 52 hemimandibles using high-resolution radiographs. Significant variability was found in its course. It was noted that if the osteotomies for sliding genioplasty were performed at least 6 mm below the inferior border of the mental foramen, injury to the mental nerve would be reduced.


Comparison of inorganic bovine bone mineral particles with porous hydroxyapatite granules and cranial bone dust in the reconstruction of full-thickness skull defect. Twenty adult rabbits were used to evaluate the biocompatibility and osteoconductivity of Bio-Oss, an inorganic bovine bone mineral, in the reconstruction of full-thickness skull defects. Skull defects were treated with either autogenous bone dust, porous hydroxyapatite granules, Bio-Oss particles, or were left untreated as controls. Histological examination of decalcified sections showed incorporation of Bio-Oss into the host tissue without a significant inflammatory reaction. Measurement of the profile area occupied by the bone revealed that Bio-Oss, hydroxyapatite, and the control had the same amount of bone ingrowth, whereas autogenous bone dust produced a greater amount of bone (p < 0.01). We conclude that Bio-Oss, like porous hydroxyapatite, has sufficient osteoconductive properties and can also be used as a bone substitute material.