Particles resembling C-type oncornaviruses in normal rat prostate. A large number of particles resembling oncornaviruses were observed in the ventral lobe of a normal rat prostate. The particles were seen budding into the intercellular spaces of the epithelial lumen cells. They exhibited features morphologically similar to immature C-type particles, with an electron-lucent center and a diameter of approximately 100 nm. Neither intracellular A-type particles nor mature C-type particles were observed. This is the first report of particles resembling oncornaviruses in normal rat prostatic tissue.


Effect of submaximal isometric exercise on catecholamine, cAMP and lactate concentrations in the coronary circulation of man, following atropine and oxprenolol. The effects of atropine and oxprenolol on changes occurring in total catecholamine, cyclic AMP (cAMP) and lactate concentrations in arterial and coronary sinus blood, during submaximal isometric exercise, were studied in 10 patients. Static one-third-maximal handgrip exercise, sustained for 5 minutes, did not produce an increase in either arterial or coronary sinus plasma catecholamine concentrations (measured at rest and during the last minute of exercise) and was not influenced by atropine and oxprenolol. Myocardial lactate production did not occur. Coronary sinus cAMP concentrations fell during isometric exercise from 11.53 +- 0.93 to 9.42 +/- 0.81 nmol/l (+/ SEM), and following autonomic blockade from 12.46 +/- 1.12 TO 9.6 +/- 0.87 nmol/l but rose on subsequent isometric exercise to 11.27 +/- 0.8 nmol/l (p less than 0.05). Although this latter increase could still be due to beta-adrenergic stimulation, the absence of any change in catecholamine concentrations in the presence of beta-blockade suggests that other factors may have been responsible.


Effects of ACTH and 3',5'-cyclic purine nucleotides on the morphology and metabolism of normal adult human adrenocortical cells in primary tissue culture. Stereological studies showed that treatment of normal adult human adrenocortical cells in primary culture with ACTH or cyclic-AMP for 2 days results in similar increases in the volume of cells, of the mitochondrial and "membrane space" compartments and of the surface area of the smooth endoplasmic reticulum and mitochondrial cristae, and decrease in the lipid content of the cells. These changes were more marked after 8 days of treatment. Treatment for 2 days with cyclic-GMP had no striking effects on cell ultrastructure, whereas an 8-day treatment led to ultrastructural changes similar to those obtained after 2 days of ACTH- or cyclic-AMP-treatment. A discrete population of untreated cortical cells maintained a slow proliferation that was not effected by exposure to cyclic-GMP, but was significantly increased in cultures treated with ACTH or cyclic-AMP. Radioimmunological studies showed that untreated cortical cells kept secreting progesterone and cortisol and that ACTH, but neither cyclic nucleotide, increased the secretion rate per cell of both hormones. These results assign a major role to cyclic-AMP and a minor one to cyclic-GMP in the mediation of the differentiation-promoting and trophic effects, but not in the steroidogenic effects of ACTH on the human adrenal cortex.


Pesticide-induced changes in hepatic microsomal enzyme systems: further studies on the effects of 1,1,-di(p-chlorophenyl)-2-chloroethylene (DDMU) in the Japanese quail. Changes in the liver resulting from the low level dietary administration of 1,1-di(p-chlorophenyl)-2-chloroethylene (DDMU),p,p'-DDT, o,p'-DDT, p,p'-DDD and p,p'-DDE to Japanese Quail have been monitored. DDMU was exceptional in causing substantial increases in relative liver wt. and hepatic glucose-6-phosphatase after feeding at 100 ppm for 28 days. The time course of liver enzyme induction by DDMU has also been studied in Japanese Quail after periods of dietary administration ranging from 1--28 days with particular reference to changes in hepatic cytochrome P-450 and relative liver wt. Structural changes in the liver have been followed by reference to protein and lipid components. The hepatic response to DDMU appears to be biphasic. Initially there are substantial increases in hepatic cytochrome P-450 and relative liver wt., but the latter is largely due to accumulation of triglycerides. After approximately 20 days the level of hepatic cytochrome P-450 remain at a high 'plateau' level. This secondary phase of liver induction probably involves cell proliferation. It is concluded that DDMU causes major changes in the avian liver and either directly or through a metabolite causes pronounced microsomal enzyme induction.


[Agenesis of the femoral vein in the framework of vascular dysplasias of the lower limbs. Clinical case]. Venous dilatations of the lower limbs, especially if associated with others abnormalities, may reflect pathogenetic mechanisms far more complex and diverse than plain insufficiency of the sephena, all the way to congenital vascular anomalies of all possible types and still dubious classification. In such conditions a prerequisite of prime importance is the complete exploration of the whole arterial and venous vasculature of the limb, first of all in order to avoid rash therapeutic decisions, and then to help clarify the diagnosis and select the most appropriate treatment from among the numerous technics available.


Glucose-6-phosphatase activity in liver and blood platelets of two patients with glycogen storage disease type I. Glucose-6-phosphatase (G-6-Pase) activity in liver and blood platelets of two patients with glycogen storage disease (GSD) type I is described. Both patients had a reduced activity of G-6-Pase in liver. The km value for glucose 6-phosphate (G-6-P) of residual activity in liver of both patients was similar to that of control liver. We could not demonstrate any reduced activity of platelet G-6-Pase in the patients. Platelet G-6-Pase with our assay method seems to represent a nonspecific phosphatase activity. Our observation suggests that it is necessary to examine platelet G-6-Pase of many other patients with GSD type I to confirm that G-6-Pase deficiency can be diagnosed by enzyme assay performed on blood platelets.


Triglyceride lipase activity in postheparin plasma and plasma lipoproteins in liver disease. Hepatic lipase activity and lipoprotein lipase activity were studied in postheparin plasma from 14 patients with various liver disorders. Plasma lecithin: cholesterol acyltransferase (LCAT) activity and lipoprotein composition and structure were also estimated. Five patients had lower hepatic lipase activity than the lowest control value, and in three of these no hepatic lipase activity was detected. Lipoprotein lipase was low in 5 patients, but in only one of them was hepatic lipase activity also low. Hepatic lipase was not significantly correlated to the concentration of plasma triglycerides, either in controls or in patients, whereas lipoprotein lipase was negatively correlated with plasma triglycerides both in controls and patients. Lipoprotein lipase and LCAT activity, but not hepatic lipase, was negatively correlated to the triglyceride content of the low density lipoproteins (density 1.019-1.063 g/ml) from the patients. No specific lipid or lipoprotein pattern was found in plasma from the patients with a low or without any hepatic lipase activity. The results suggest an important role of lipoprotein lipase and LCAT, for the increased content of triglycerides in the low density lipoproteins in patients with liver disease. The role of hepatic lipase remains unclear.


An evaluation of quantitative agarose-gel electrophoresis of serum lipoproteins. The method of Hatch et al. (Hatch, F.T., Lindgren, F.T., Adamson, G.L., Jensen, L.C. and Wong, A.W. (1973) J. Lab. Clin. Med. 81, 946-960) of quantitative determination of serum lipoproteins has been compared with preparative ultracentrifugation and chemical analysis of the lipoprotein fractions and a good concordance was demonstrated. The replacement of the coefficients proposed by Hatch et al. by the ones derived from the present study does not bring about relevant changes of the results when the harmonic mean of two calibration factors, derived from serum cholesterol and triglycerides, is used, thus indicating that this method of calculation minimizes the effect of variation of the chemical composition of lipoproteins. The method is sufficiently reliable and reproducible for practical pruposes and is recommended as the standard method for the clinical laboratory.


The quantitative determination of apolipoprotein A-I (apo-lp-Gln I) in human serum by radial immunodiffusion assay (RID). A radial immunodiffusion (RID) assay was developed to determine the concentration of the major apolipoprotein (apolipoprotein A-I, apo-lp-Gln I) of human high density lipoproteins (HDL): (1) In a random population sample of apparently healthy persons (104 women, 95 men) aged 40-49 years, serum levels of apo A-I were determined. For both women and men a value of 137 +/- 23 mg/dl was found. (2) Values of 139 +/- 21 mg/dl were found in a group of 31 women taking oral contraceptives, i.e. an insignificant increase in apo A-I level. (3) In two volunteers, apo A-I levels showed no significant variation in a 24-h period.


Effects of dietary saturated and polyunsaturated fat on the metabolism of apolipoproteins A-I and B. Study of a patient with type IIb hyperlipoproteinaemia. The effects of dietary saturated and polyunsaturated fat on the metabolism of apolipoprotein A-I (apoA-I) and apolipoprotein B (apoB) were studied in a patient with type IIb hyperlipoproteinaemia. On the saturated fat diet, the rate of synthesis of very low density lipoprotein apoprotein B (VLDL-apoB) was approximately twice normal, accounting for the increased plasma VLDL pool in this subject. However, 54% of the synthesized VLDL-apoB was catabolized by a pathway independent of low density lipoproteins (LDL). The metabolic conversion rate of VLDL-apoB to LDL-apoB was normal in this subject and his expanded plasma LDL-apoB pool resulted, not from increased input of the apoprotein from VLDL, but from a decrease in its fractional clearance rate. On the polyunsaturated diet, there was a significant fall in the plasma cholesterol and triglyceride concentrations and a change in the fatty acid composition of all plasma lipoprotein fractions. These changes were accompanied by a decrease in the plasma concentrations of apoA-I and apoB which resulted from a reduction of apoprotein synthetic rate.


[Stability of lipid and protein composition of very low-density lipoproteins during the storage after freezing of whole serum and isolated lipoproteins (author's transl)]. The lipid and protein composition of lipoprotein is now regarded with great interest for the identification of hyperlipoproteinemias. Because this relatively time-consuming study cannot be envisaged currently in clinical services, it seems useful to analyse a storage procedure which does not affect the composition of very-low density lipoproteins, whose proportion is very important among subjects with type IV hyperlipoproteinemia. Two studies were considered, one using whole sera stored at -20 degrees C and one using very low-density lipoproteins held in the same conditions. The lipid composition was determined for triglycerides and cholesterol concentrations of total proteins, proteins soluble in tetramethylurea (apoprotein C, "arginine-rich fraction", and apoprotein D), and peptides CII, CIII1 and CIII2 of apoprotein C. For both procedures considered, no significant changes were seen in lipid and protein composition for periods as long as 8 weeks.


On the "rapid beta" (Rbeta) lipoprotein variant of human serum. By visual inspection of agarose gel electrophoretograms of lipoproteins, a beta lipoprotein with increased mobility, here named "rapid beta" lipoprotein, has been observed. By objective measurements of the migration distances of the lipoprotein bands in 542 electrophoretograms by means of an exact ruler, this visual impression was objectively confirmed. In samples, classified by inspection as "rapid beta", the beta lipoprotein was found to have migrated approx. 20% further than in control samples. The "rapid beta" variant was initially detected in samples from grossly obese patients where it occurs frequently, but does not seem to be specific for any clinical abnormality.


High-density lipoprotein cholesterol, monthly variation and association with cardiovascular risk factors in 1000 forty-year-old Dutch citizens. High density lipoprotein cholesterol (HDL chol) levels were measured in about 1000 forty-year-old residents of the town of Leiden, The Netherlands. Mean levels (+/- S.D.) in women were 48.0 +/- 11.0 mg/dl; in men 42.5 +/- 10.7 mg/dl. In March mean levels in women were about 5 mg/dl lower than in June (P less than 0.001); in men there was a similar difference between mean levels in April and June (P less than 0.002). HDL chol levels were negatively related to smoking habits, relative body weight, and in women to use of oral contraceptives. Weak positive and negative correlations between HDL chol and serum cholesterol, respectively, were found in subgroups with high and low HDL chol levels, respectively.


A simple spectrophotometric method for estimation of plasma angiotensin I converting enzyme activity. The procedure described is rapid, fairly simple, and inexpensive. The method may easily be used even in a small clinical laboratory. This method, based on a specific reaction which gives a product, hippuric acid azlactone, which absorbs in the visible region, avoids the interference of reagents and solvents encountered in the UV spectrophotometric assay, in which hippuric acid is determined directly.


Evaluation of an automated immunoassay procedure for apo-LDL for phenotyping type II hyperlipoproteinemia. Apo-low density lipoproteins were determined by an automated immunoassay procedure on serum samples from 88 normolipidemic individuals and 84 hyperlipoproteinemic subjects, to establish whether this method was useful in the routine detection of type II hyperlipoproteinemia. The results obtained were compared with the cholesterol levels of the same specimens. In subjects with type II hyperlipoproteinemia, the apo low density lipoprotein levels, as well as the ratio of low density lipoprotein cholesterol/apo-low density lipoprotein were higher, as expected, than in normals or in subjects with other types of hyperlipoproteinemia. However, there was considerable overlap in individual values of both these parameters, between patients with type II hyperlipoproteinemia and normals or subjects with other types of hyperlipoproteinemia, suggesting that apo low density lipoprotein levels alone were not sufficiently discriminatory for the laboratory determination of type II hyperlipoproteinemia.


Discrete separation of HDL2 from HDL3 of human serum by means of polyacrylamide gel. A relatively short, simple procedure is presented to separate serum high-density lipoproteins discretely into the two main classes, those with densities between 1.063 and 1.125 g/ml (HDL2) and those with densities between 1.125 and 1.210 g/ml (HDL3). A 3.5% polyacrylamide gel in 10 cm glass tubes and the use of Tris/glycine buffer, pH 8.4, will accomplish this separation. The components can be identified in several different ways, examples of which are given. This procedure will give rapid and reliable estimations of both HDL2 and HDL3, and can be used to relate their levels and proportional amounts to incidence or risk of atherosclerosis, coronary-artery disease and possibly cancer.


Cushing's syndrome, nodular adrenal hyperplasia and virilizing carcinoma. A 48-year-old hypertensive diabetic woman rapidly became virilized. Urine 17-oxo-and oxogenic steroids and plasma testosterone, androstenedione, DHEA, DHEA-sulphate and androstenediol were greatly elevated. Plasma cortisol was constantly high and was not suppressed by dexamethasone. Circulating immunoreactive ACTH was consistently detectable at 18-24 ng/l. A 450 g carcinoma arising from a nodular hyperplastic right adrenal gland was resected. Production by the tumour of 17a-hydroxypregnenolone, 17a-hydroxyprogesterone and five C-19 steroids, but very little prenenolone, progesterone or cortisol, was shown by blood sampling, tumour culture and dramatic falls after operation. The plasma cortisol fell to half, with no diurnal variation, consistent with persistent Cushing's syndrome, and the plasma ACTH rose to 55 ng/l. She died 3 months later from a myocardial infarction. Autopsy revealed a pituitary basophil adenoma at a site where radiologically there had been an indentation in the fossa floor for at least 7 years. The left adrenal gland showed nodular hyperplasia. Therefore we conclude that mild pituitary-dependent Cushing's syndrome may have been present for many years before development of a virilizing carcinoma. This case demonstrates that adrenal carcinoma in man can sometimes develop as a consequence of nodular adrenal hyperplasia which may in turn be due to long-standing trophic hyper-stimulation.


In vitro studies of the interaction of isolated Lp(a) lipoprotein and other serum lipoproteins with glycosaminoglycans. The interaction of isolated Lp(a) lipoprotein or other lipoprotein classes with different glycosaminoglycans (GAG) bound to activated Sepharose was studied. In contrast to LDL, the Lp(a) lipoprotein did not bind to the GAG tested if sodium was used as a buffer cation. In the presence of Ca++, however, even the Lp(a) lipoprotein was bound to GAG. This type of binding, probably mediated by divalent cation bridges, is apparently not a simple function of the GAG used. Addition of GAG in solution revealed that this binding may be the only one existing under physiological conditions, and it appears possible that the Lp(a) lipoprotein is bound more firmly to GAG than is LDL under such conditions.


Inter observer agreement in the assessment of the motor response of the Glasgow 'coma' scale. The motor response of the  Glasgow 'coma' scale  is readily learned by nurses and doctors. It can be used as a reliable indicator of changes in the state of a comatose patient, as there is a high agreement rate between different observers.


Gliomas: new ways of treatment?

Gliomas: new ways of treatment? Evidence starts to appear that immune reactions may be important in the development and growth of gliomas. The purpose of this article is to review this evidence and to mention its implications for the future.


Craniosynostosis: a new operative technique. In the treatment of craniosynostosis several methods have been proposed to prevent craniectomies from early reclosure. New bone is formed at the edge of the craniectomy, but particularly at the outer surface of the dura. The author has developed a method to separate the edges from each other and to stop osteogenesis inside the craniectomy. The outer layer of the dura is dissected free from the inner layer folded over the edge of the groove and sutured to the outer periosteum. The operative technique and the results in 40 patients with a follow up of 17 years are described.


Kinking of the carotid artery: indications for surgery and surgical procedure. In seven patients surgical correction of kinking with stenosis of the extracranial part of the carotid artery was performed. The indications for surgical treatment of carotid kinking are discussed in detail. The following criteria for surgical correction are applied: 1. symptoms of cerebral ischemia must be present; 2. position or motion of head and neck can provoke symptoms; 3. definite stenosis of the corresponding carotid artery at the level of kinking; 4. no other causes of cerebral ischemia; 5. no irreversible brain infarction. A simple and effective surgical procedure as a routine method for correction of carotid kinking is described. Only in special circumstances a more extensive open arterial reconstruction might be necessary.


Methodology for analyzing clinical neurological data: ACTH in multiple sclerosis. From 1965 to 1968 a multicenter (USA) clinical trial was conducted to compare ACTH with placebo for the treatment of multiple sclerosis patients in a stage of acute exacerbation. Of seven evaluation systems which were used, one was a quantitative neurological examination. In this report, the quantitative neurological data have been re-evaluated using a transformation of the data which express patient scores in terms of a percentage of age- and sex-matched normal function followed by a reduction of the data into composite neurological functions vs time. The results support the positive findings of previous reports. In addition, the methodology helps to simplify the therapists' task of interpreting results by enabling him to determine how near normal function the patient came following the treatment trial.


Intestinal cholecalciferol absorption in the elderly and in younger adults. 1. A method for assessing cholecalciferol absorption in man is described. 2. The intestinal absorption of [3H]cholecalciferol was studied in 20 female geriatric patients, most of whom were vitamin D-depleted. 3. The plasma [3H]cholecalciferol response after oral ingestion was significantly lower than that of a group of younger female subjects. 4. The plasma response of labelled polar metabolites of cholecalciferol was also lower in the geriatric than in the younger group, suggesting that increased removal of label by conversion into more polar metabolites could not account for the reduced plasma [3H]cholecalciferol response. 5. There was no evidence that alteration in gastrointestinal motility could account for the different rate of appearance of the labelled vitamin in the plasma in the two groups. 6. It is suggested that there is a defect in intestinal absorption of cholecalciferol in the elderly.


Biochemical and ultrastructural analysis of SV40 chromatin. SV40 chromatin can be isolated in two forms: At moderate ionic strength (mu = 0.1-0.3) it contains histone H1 in addition to the four nucleosomal histones and has a highly condensed appearance in the electron microscope, being composed of a few closely connected large spheres [190 A (160, 220) diameter]. At high ionic strength (mu = 0.6-0.8) or after prolonged exposure to very low ionic strengths (mu less than 0.02), the compact form unfolds and the chromatin shows a typical nucleosomal morphology. Native SV40 DNA-protein complexes contain a median number of 24 nucleosomes. The number of superhelical turns does not differ in DNA obtained from the compact and the unfolded forms of chromatin. DNA-relaxing enzyme is found associated with SV40 chromatin and is capable of acting both on extraneously added circular DNA and on its own DNA in the nucleoprotein complex. Purified DNA-relaxing enzyme forms transiently nicked DNA intermediates where the enzyme can be found covalently attached to the site of the nick in the DNA. Transcriptionally active SV40 complexes undergo the same ionic-strength-dependent structural transition as that of bulk SV40 chromatin and may therefore also have a compact configuration at physiological salt concentrations.


A rapid method for isolation of the plasma membrane of the myxamoebae and swarm cells of the myxomycete Didymium iridis. A method for the isolation of the plasma membranes of the acellular slime mould, Didymium iridis in the myxamoebae and swarm cell stages was developed using a modified dextranpolyethylene glycol aqueous two-phase polymer system. It was found to be far superior to the widely accepted technique of density gradient centrifugation concerning this cellular system. Chemical and enzymatic assays performed on the plasma membranes and other cell fractions as well as microscopic examination were used as a basis for positive identification and assessment of purity. Results of the chemical and enzymatic assays indicate that plasma membranes are recovered with high yield and purity using the modified two-phase polymer technique. The method is both rapid and effective and can be performed using low-speed centrifugation.


The diagnostic value of lysosomal enzyme patterns in bronchial aspirates of patients with suspected bronchial carcinoma. The measurement of the activity of acid hydrolases and of alkaline phosphatase in bronchial aspirates obtained through bronchoscopic procedures from a series of 300 patients forms the basis for a screening program to diagnose bronchial malignant neoplasms more effectively. We define such a screening test as one permitting rapid measurements indicative of pathologic abnormalities and producing a preliminary diagnosis which, if in error, yields preferably a false-positive result. Using this approach, we demonstrated that an elevation of the activity of alkaline phosphatase or cathepsin D predicts a 50 percent likelihood of cancer, but elevation of both the concentrations of alkaline phosphatase and cathepsin D has an additive prediction of 71 percent. Data obtained in this study showed that the presence of a pulmonary tumor can cause increased levels of alkaline phosphatase or cathepsin D (or both) in bronchial aspirates before the presently accepted methods yield a diagnostic result. Furthermore, those patients with an elevated activity of alkaline phosphatase or cathepsin D (or both) but with no histologically demonstrable pulmonary carcinoma can be reexamined intermittently.


Ultrastructure of somatotroph cells at different times in a circadian period, in mice bearing a slow growing transplanted hepatoma. An electronmicroscopic study of STH cells from mice bearing transplanted hepatomas was performed at different time points, in a circadian period. Normal mice served as controls. The STH cells of control animals showed circadian variations inplasmic reticulum and lysosomes, which suggest increased secretion of growth hormone along the light peroid. In mice bearing hepatomas the morphologic aspect of these organelles would also indicate an increase of STH elaboration at 1200 and 1600. These changes were more remarkable than in controls and mainly consisted of hypertrophy of the Golgi complex, extended and compact endoplasmic reticulum and presence of large amounts of lysosomes. Besides, another peak of secretion seems to be present at 0000, considering the dilatation of the Golgi complex and endoplasmic reticulum cisternae. Some findings are coincident with observations made by others in the hypophysis of animals bearing transplantable tumors, where circadian periodicity was not studied. The STH cytological circadian variations could be correlated with other variations could be correlated with other variables, such as STH values in plasma and hypophysis, and DNA synthesis and mitotic activity of SS1-H hepatoma.


Corticosteroids: clinical pharmacology and therapeutic use. The widespread use of corticosteroids in clinical practice emphasises the need for a thorough understanding of their metabolic effects. In general, the actions of corticosteroids on carbohydrate, protein, and lipid metabolism result in increased hepatic capacity for gluconeogenesis and enhanced catabolic actions upon muscle, skin, lymphoid, adipose and connective tissues. Because of the morbidity associated with steroid therapy, the clinician must carefully consider in each case the gains that can reasonably be expected from corticosteroid therapy versus the inevitable undesirable side effects of prolonged therapy. Thus, it is important to remember that the enhanced anti-inflammatory activity of the various synthetic analogues of cortisol is not dissociated from the expected catabolic actions of glucocorticoid hormones. Replacement therapy with physiological doses of cortisol in primary or secondary adrenal insufficiency is intended to simulate the normal daily secretion of cortisol. Short term, high dose suppressive glucocorticoid therapy is indicated in the treatment of medical emergencies such as necrotising vasculitis, status asthmaticus and anaphylactic shock. With improvement of the underlying disorder, the steroid dosage can be rapidly tapered and then discontinued over a 2 to 3 day period. Long term, high dose suppressive therapy is often commonly used to treat certain diseases (see sections 4.7.2 and 4.7.3). In this setting, suppression of the hypothalamic-pituitary-adrenal axis may persist for as long as 9 to 12 months following steroid withdrawal if steroid doses are administered in the supraphysiological range for longer than 2 weeks. In general, higher doses, longer duration of usage, and frequent daily administration are all correlated with the severity of pituitary ACTH suppression. When steroid therapy is to be withdrawn, gradual tapering of the dosage is necessary; the steroid dosage should also be given as a single morning dose if possible. Rapid or total withdrawal of the steroid therapy may be associated with exacerbation of the underlying disease or with a steroid withdrawal syndrome. An additional important point to remember in any withdrawal programme is that the steroid dosage should be appropriately increased for an exacerbation of the underlying disease or for intercurrent major stress. Alternate day therapy is recommended as a steroid maintenance programme for patients requiring high dose glucocorticoid therapy over a prolonged period of time. Thus, it is usually employed to maintain a therapeutic benefit which had previously been extablished by daily steroid treatment. Complications resulting from corticosteroid therapy include: (1) proximal muscle weakness; (2) osteopenia; (3) unmasking of latent diabetes mellitus; (4) sodium retention and/or elevation of mean arterial blood pressure; (5) adverse psychiatric reactions; (6) development of glaucoma; and (7) reactivation of latent infections (such as tuberculosis).


Field concentrations and persistence of polybrominated biphenyls in soils and solubility of PBB in natural waters. Soil samples were collected from 28 fields which had received manure from Michigan's most highly contaminated dairy herds. The number of fields in each concentration range of PBB in soil were: 2, not detectable; 15, 0.0 to 8.0 ppb; 6, 14-102 ppb, and 5, 153 to 371 ppb. Plant tissue sampled from the 10 most highly contaminated fields contained no detectable PBB. No evidence of significant degradation of PBB was noted after 1 year incubation in soil. When 14C hexabromobiphenyl and heptabromobiphenyl isomers were incubated in soil less than 0.2% of the 14C was volatilized. Also gas chromatographic analysis of soil extracts showed no difference in recovery of the six major PBB isomers between sterilized and nonsterilized soil. Analysis of these extracts by thin layer chromatography and autoradiography showed no 14C-PBB intermediates. Photodegradation products of the major hexa- and heptabromobiphenyl isomers showed more but still minor (approximately 3%) biodegradation in soil. Much of the photodegradation products appeared bound to soil, since these products could not be extracted from soil. Photodegradation does not appear to be a significant fate of PBB in manures spread on fields since no change was noted in the relative concentrations of isomers in soil samples from our field survey. Studies with distilled, tap, river, and soil waters showed that PBB solubility was markedly influenced by water composition.


Comparison of findings among residents on Michigan dairy farms and consumers of produce purchased from these farms. Consumers who had purchased farm products from both quarantined and nonquarantined farms were examined during the cross-sectional clinical survey of 1,029 Michigan residents. Since PBB had inadvertently contaminated cattle and other farm animals, ingestion of meat, milk, eggs and other farm products was thought to have possibly resulted in significant PBB body burdens in some consumers. Findings were considered in comparison with those made among farm residents. Prevalence of symptoms in consumers of farm products from quarantined farms (CQ) was similar to that found in farmers on quarantined farms (FQ); the prevalence was lower in consumers of products from nonquarantined farms (CNQ). Liver function abnormalities were found with similar prevalence in dairy farmers and consumers. Distribution, mean and median values of PBB serum levels in consumers were found to be similar to those of dairy farmers. These results indicate that significant body burdens of PBB had been accumulated by some consumers of farm products in Michigan and that prevalence of symptoms and liver function abnormalities resembled those found among dairy farm residents.


Effects of PBB on cattle. II. Gross pathology and histopathology. Toxicosis was induced in pregnant Holstein heifers by feeding FireMaster BP-6 (polybrominated biphenyls) in daily oral doses of 25 g/head/day for 33--60 days. The individual heifers were dosed until each became moribund (days 33, 36, 39, 40, 41, or 66), at which time they were necropsied. Gross findings included dehydration, subcutaneous emphysema and hemorrhage, atrophy of the thymus, fetal death with concomitant necrosis of cotyledons, thickened wall of the gallbladder, inspissated bile, edema of abomasal folds, mucoid enteritis, linear hemorrhage and edema of the rectal mucosa, and secondary pneumonia. The livers were enlarged approximately 40%. Kidneys were approximately double the normal size and were pale tan to grey in color. The perirenal lymph nodes were enlarged and edematous. Microscopic changes were the most marked in the kidneys, gallbladder and eyelid. Extreme dilatation of collecting ducts and convoluted tubules with epithelial degenerative changes of cloudy swelling, hydropic degeneration and separation from the basement membrane were principal changes in the kidney. Hyperkeratosis with accumulations of keratin in hair follicles of the epidermis and squamous metaplasia with keratin cysts in the tarsal glands were characteristic findings in sections of eyelids. Moderate to marked hyperplasia and cystic dilatation of the mucous glands in the lamina propria were common changes in the gallbladder. Foci of fatty degeneration and glycogen depletion were observed in liver sections. Necropsy of heifers immediately after 60 days exposure to 0.25 and 250 mg/head/day of PBB showed no gross or histopathological signs indicating toxicosis. Following parturition, at approximately 220 days after the PBB doses, heifers from the 0.25 and 250 mg/head/day groups and their calves were necropsied and displayed no signs of toxicosis.


Effects of PBBs on cattle. III. Target organ modification as shown by renal function and liver biochemistry. Efforts were made to more clearly delineate target organs and mechanisms of toxicity for PBBs in cattle. Methods were developed to obtain sequential liver biopsies on bovine heifers which yield 0.5 to 1.0 g of tissue. PBB was fed at a dose of 250 mg/head/day to Holstein heifers for 202 days. This dose produced no clinical signs of toxicity in any of the heifers, yet this produced tissue PBB concentration of greater than 100 times the FDA tolerance in body fat of 0.3 ppm. Liver biopsies (0.5-1.0 g each) were taken at days 0, 90, and 180. The liver tissue was homogenized and microsomes were prepared. Dithionite difference spectra were determined on the carbon monoxide treated microsome suspension and the cytochrome P-450 content determined. Also, the 100,000g supernatant was saved for ornithine decarboxylase analysis as a measure of hepatocyte proliferative activity. Results of the cytochrome P-450 analysis showed a significant (p < 0.05) two-fold elevation (per gram of wet liver) by day 90 and remained significantly (p < 0.05) elevated on day 180. The cytochrome P-450 values of control animals not receiving PBBs showed no such increase with time. The biopsy procedure appeared not to adversely affect the liver cytochrome P-450 concentration in the control heifers. These results show that PBBs at a dose of 250 mg/day induced the drug metabolism system of the liver, of which the cytochrome P-450 is a part, indicating that the liver is a potential target organ for PBBs. However, this has not been shown to cause clear signs of hepatotoxicity in the cow as determined from histopathology or serum enzyme analyses. The observed elevation of gross liver weights of the PBB-treated animals might be an expected consequence of the cytochrome P-450 induction. In contrast to rodents, the kidney has been identified by histopathology as a target organ for PBB toxicity in cattle. However, renal function studies with (131)I-sodium-iodohippurate and (125)I-sodium iodothalamate in PBB treated cows indicated that PBB toxicity to the kidney did not affect glomerular filtration rate or effective renal plasma flow even though nephrotoxic effects were produced. From these studies, both liver (as expected) and kidney (unexpected) were affected by PBBs. For liver this did not result in hepatotoxicity while for kidney nephrotoxicity was produced but could not be mechanistically explained.


Cytogenetic and teratogenic test of polybrominated biphenyls in rodents. Previously we reported negative terata and c-mitosis synergism of FireMaster (polybrominated biphenyls) with colchicine in subacutely treated rats. Now we report absence of chromosome aberrations from FireMaster and absence of c-mitosis synergism of FireMaster and colchicine in male mice. For the study of chromosome aberrations groups of three mice received 0, 50, or 500 mg/kg FireMaster or 4.5 mg/kg triethylenemelamine (TEM) dissolved in dimethyl sulfoxide through a single stomach gavage administration. Five hours before killing the animals were injected with 5 mg colchicine/kg. Groups of 3 mice from each treatment killed 12, 24, and 48 hr after treatment. From the bone marrow of each of 36 mice 100 metaphases were scored for gaps, chromatid and chromosome breaks, rearrangements and pulverized chromosomes. Only TEM induced chromosome damage. For detection of synergism between FireMaster and colchicine, slides prepared for chromosome analysis were also scored for metaphase and mitotic indeces. Control mice for detection of synergism were treated as for the chromosome study but were not injected with colchicine. Approximately 1000 cells were scored from each of 72 animals for determination of metaphase and mitotic indeces. FireMaster did not show c-mitosis synergism with colchicine in mice. Treatment with FireMaster did not cause visually recognizable toxicity.


Polybrominated biphenyl (PBB) in the growing pig diet. Twelve pigs which averaged 13.7 kg were randomly allotted from litters to a corn-soybean meal grower diet containing 0, 20, or 200 ppm of polybrominated biphenyls (PPB). During a 16-week growth trial, average daily gain (kg), average daily feed (kg) and feed/gain for pigs on diets containing 0, 20, or 200 ppm of PBB, respectively, were 0.82, 2.45, 2.99; 0.67, 1.88, 2.79; 0.45, 1.23, 2.70. Mean daily gain differences between all lots were highly significant (p < 0.01). Blood from each pig was withdrawn biweekly through the first 8 weeks of the trial and at 4 week intervals thereafter. Hemoglobin and hematocrit differed significantly only at the 6 weeks bleeding, being reduced in pigs receiving 200 ppm of PBB. Erythrocyte reduced glutathione concentration and glutathione peroxidase activity were not significantly influenced by level of dietary PBB. Serum lactic dehydrogenase activity was significantly higher in control pigs than in either PBB supplemented lots at 16 weeks. There was no significant influence of PBB upon serum glutamic oxaloacetic transaminase, serum alkaline phosphatase or serum creatine phosphokinase. Based on these enzyme assays, PBB produced no evidence of significant necrosis of liver, myocardium, or skeletal muscle. There was no consistent effect of dietary PBB upon total serum protein concentration or electrophoretic profile. Pigs on either level of PBB did not have overt clinical signs of toxicity during the 16-week test period with the exception of a dermatosis on the ventral surface of two of the pigs receiving 200 ppm of PBB. There was a marked increase in liver weight of pigs receiving either level of dietary PBB. Heart, kidney, and adrenals of pigs receiving either level of dietary PBB were heavier as a percent of body weight than that of control pigs. Fat retention of PBB and urinary and fecal PBB excretion were significantly affected by dietary PBB level. Grossly, the glandular portion of the stomach appeared somewhat hyperplastic in pigs on 200 ppm of PBB. Two pigs which had received 200 ppm of PBB were placed on the control diet and over the next 14 weeks normal growth rate occurred. One of these pigs was killed and organ weights were normal. The other pig, a gilt, came into estrus. She was bred and conceived. At the end of gestation, four pigs were born. Three survived and grew normally; the one death at birth examined at gross necropsy did not reveal changes in organ size or other tissue alterations.


Effect of diet on the hepatotoxicity of polybrominated biphenyls (FireMaster PB-6). The consumption of diets formulated with Cruciferae vegetables, e.g., cauliflower, cabbage, and Brussels sprouts, has been shown to result in a stimulation of the intestinal and hepatic microsomal enzyme systems in rats. This study was designed to determine if this increase in intestinal and hepatic microsomal enzyme activity affected the hepatic response to polybrominated biphenyls (PBB). After three weeks of consuming either a semipurified or 25% cauliflower leaf-supplemented diet (CLD), male Sprague-Dawley rats were maintained for an additional 20 days on their respective diets containing either 0, 1, or 50 ppm PBB. A significant decrease in body weights, but not feed efficiency, was observed over all levels of PBB in animals consuming CLD compared to semipurified diets; consumption of up to 50 ppm of PBB had no effect on body weights with either diet. Relative liver weights (RLW), hepatic aryl hydrocarbon hydroxylase (AHH), N- and O-demethylase, as well as intestinal AHH were all increased in CLD-consuming animals before the addition of PBB. While PBB supplementation alone resulted in increased RLW, hepatic AHH, N- and O-demethylase, microsomal protein, and cytochrome P-450, rats consuming cauliflower diets + PBB had even higher RLW and N- and O-demethylase activity and microsomal protein concentrations. Hepatic PBB residue and total hepatic lipids were significantly reduced in CLD groups receiving 50 ppm PBB. These results suggest that the antitoxic effects of certain vegetables are related to more rapid metabolism and excretion of xenobiotic compounds.


Responses of nonhuman primates to a polybrominated biphenyl mixture. In a series of experiments, rhesus monkeys have been given in their diets 0.3, 1.5, and 25 ppm of a commercial polybrominated biphenyl (PBB) (as FireMaster FF-1). The seven adult female monkeys receiving 0.3 ppm PBB have been on the treatment regime for 15 months and have consumed over 22 mg of PBB. During the initial 6 months of exposure, they lost weight and 2 of the animals develop sterile abscesses. At 6 months, 4 of the 7 animals had flattened and lengthened serum progesterone peaks. This change was correlated with an increase in length of their menstrual cycles. After 6 months of PBB exposure, the animals were bred. Two of the 7 animals showed excessive and prolonged implantation bleeding. Two abortions and 5 live births were recorded. All of the experimental infants were smaller than the controls at birth. The animals receiving a diet containing 1.5 ppm PBB for 36 weeks (total intake 70 mg) have shown a moderate weight loss and decrease in serum cholesterol. Similar changes have also been recorded in the group given the 25 ppm PBB diet for 14 weeks (approximately 500 mg total intake). In addition, these animals have also developed a hyperplastic gastritis.


FireMaster BP-6: fractionation, metabolic and enzyme induction studies. FireMaster BP-6 is a commercial polybrominated biphenyl (PBB) preparation containing a complex mixture of isomers with the major component being identified as 2,2',4,4',5,5'-hexabromobiphenyl. Column chromatographic techniques have been developed in which the crude FireMaster is separated into three fractions, F1, F2, and F3, in increasing order of polarity. F1 consists of highly purified 2,2',4,4',5,5'-hexabromobiphenyl (94%) whereas F2-F3 contain less of this isomer and correspondingly more of the other bromobiphenyl components. Previously we have shown that crude FireMaster BP-6 is metabolized in mammals to give hydroxylated degradation products and the metabolism of F1, not unexpectedly, gives comparable results. It is well known that PBBs are effective inducers of diverse microsomal enzymes including including the aryl hydrocarbon hydroxylase (AHH) system. The effects of FireMaster BP-6 and F1-F3 as AHH inducers have been investigated by using the following approach: the substrates used to monitor AHH activity are model halogenated aromatic compounds; the levels of metabolites and metabolite conjugates formed have been quantitated for control and induced enzymes; the levels of macromolecular adducts have also been quantitated for the inducers. This approach thus not only measures the rate of increase of detoxification products (metabolites and metabolite conjugates) but also monitors the macromolecule adduct formation which represents a toxification route. The effects of the PBBs as AHH inducers will be discussed in terms of the above approach.


Effects of polybrominated biphenyls on kidney function and activity of renal microsomal enzymes. Polybrominated biphenyls (PBBs) cause hepatic microsomal enzyme stimulation and histopathological alterations in several organs, including kidney. Concern about effects of PBBs on the health of newborns has increased after the discovery of PBBs in milk of nursing mothers. Therefore, it was of interest to investigate the effects of PBBs on kidney function and the activity of renal microsomal enzymes in adult and immature animals. Seven and eleven day old pups were treated with a single IP injection of either peanut oil or 150 mg/kg PBBs (FireMaster BP-6) in peanut oil. Adult virgin rats were fed diet containing 0 or 100 ppm PBBs for 30 or 90 days. Treatment with PBBs only retarded weight gain after 90 days exposure. Kidney-to-body weight ratio was not altered by PBBs. Arylhydrocarbon hydroxylase activity was increased while epoxide hydratase activity was decreased (adults) or not affected (immature rats) in kidney following treatment with PBBs. Administration of PBBs had no effect on blood urea nitrogen, the clearance of inulin, p-aminohippurate (PAH), or fractional sodium excretion. Similarly, the in vitro accumulation of PAH and N-methylnicotinamide (NMN) by thin renal cortical slices and ammoniagenesis and gluconeogenesis in renal cortical slices were not affected by PBBs. In conclusion, treatment with PBBs resulted in modification of the activity of renal microsomal enzyme activities but had no detectable effect on renal function.


Induction of drug metabolizing enzymes in polybrominated biphenyl-fed lactating rats and their pups. Polybrominated biphenyls (PBBs) cause a mixed-type (phenobarbital- plus 3-methylcholanthrene-like) induction of liver microsomal drug metabolizing enzymes in rats. However, 2,2',4,4',5,5'-hexabromobiphenyl and 2,2',3,4,4',5,5'-heptabromobiphenyl, which together comprise less than 80% of PBBs (FireMaster), were shown to be strictly phenobarbital-type inducers. Other components (unidentified) must therefore cause the 3-methylcholanthrene-like effects. The potential for PBBs to exert effects on neonates through milk was examined. Lactating rats were fed 0, 0.1, 1.0, or 10 ppm FireMaster for the 18 days following delivery, at which time mothers and most pups were sacrificed. Pups nursing from mothers fed 10 ppm PBBs showed significant increases in liver weights and microsomal protein, and both mothers and pups had increased cytochrome P-450, aminopyrine demethylation, benzo[a]pyrene hydroxylation, and UDP-glucuronyltransferase. Pups nursing from rats fed 1.0 ppm had increases in microsomal protein, cytochrome P-450, aminopyrine demethylation, and benzo[a]pyrene hydroxylation, while their mothers were unaffected. Several pups from the 0, 0.1, and 1.0 ppm groups were maintained on their mother's diets, raised, and allowed to mate. Their pups showed much the same responses to PBBs as did the original group of pups. The effects on both generations of adult female rats were also comparable. PBBs cause a mixed-type induction in both lactating rats and their nursing pups; PBB components responsible for both aspects of this induction must be transmitted through milk. Nursing rats are approximately tenfold more sensitive to the effects of PBBs in their mother's diets than are the dams. The approximate no-effect level for microsomal induction in nursing rats is 0.1 ppm PBBs in the diet of the adult.


Toxicity of PBBs with special reference to porphyrinogenic action and spectral interaction with hepatic cytochrome P-450. Some of the polyhalogenated aromatic compounds (PHAs) which are able to produce porphyria are presently known as environmental contaminants. Chronic exposure to PHAs causes hepatic porphyria in different species. Qualitatively PBBs act comparable to PHAs. An increase in accumulation of porphyrins caused by PHAs is not simply related to an increase of delta-ALAS activity in liver. Heme cannot exert a feedback when porphyria develops. Induction of P-450 mediated drug enzymes is needed. The PHAs interact with P-450 in vitro. The PHAs are converted into a reactive intermediate, not a known metabolite, which depletes liver GSH and then becomes reactive to tissue structures. Mitochondria are damaged; fluorescence of porphyrins is detected in the region of central veins where degenerative change in hepatocytes is most marked. A possible pathological change in the cell membrane permeability is assumed too. In this porphyric stage uropophyrinogen decarboxylase (urogen decarboxylase) is inhibited. The proportion of steroid hormones product by ovaries and testes compared to each other are possibly involved in sensitivity to porphyrinogenic compounds.


Relation of DDE and PBB serum levels in farm residents, consumers, and Michigan Chemical Corporation employees. Results of serum PBB determinations on 524 Michigan diary farm residents and consumers of products from the farms, 55 chemical workers, and 56 Wisconsin farm residents are reported. Mean and median values were highest for the chemical workers, followed by consumers from and residents of quarantined and nonquarantined farms. Serum DDE was higher among chemical workers, but was similar for all other groups. Statistical analysis of serum of PPB and serum DDE levels was done with respect to quarantine status, age, sex, and obesity. The most significant correlate with PBB was quarantine status. Serum DDE, age, sex, or obesity were not consistently correlated with serum PBB. For DDE, age was invariably the most significant correlate. Both serum PBB and DDE were higher in males than females in husband-wife pairs in most cases, although the differences occurred less frequently among older age groups. These results support the hypothesis that PBB exposure was a recent interim exposure whereas DDE exposure has been cumulative throughout a person's lifetime. Higher PBB and DDE mean concentrations in serum of Michigan Chemical workers suggests an occupational exposure to these chemicals.


A simple technique for fat biopsy of PBB-exposed individuals. A simple nonsurgical technique of obtaining fat samples by aspiration from the gluteal prominence was developed by Hirsh in 1960 and has been in use in our Nutrition Clinic at the Mount Sinai Hospital for several years. We have modified it for field use and the analysis of fat-soluble hydrocarbon residues. All the materials which will contain the fat sample to be analyzed are washed with acetone and pesticide residue-free hexane, and a 15 gauge needle and 33 cc syringe are sterilized. Aspiration of fat from the lateral gluteal prominence is accomplished under local xylocaine anesthetic. The anesthetic also serves as the vehicle into which the fat is broken by the shearing action of the 15 gauge needle. Fat particles are sucked into the syringe by a constant vacuum kept on the syringe during lateral movement of the needle under (and parallel to) the skin within the gluteal fat pad; 200--500 ml of fat can be obtained for hydrocarbon residue analysis. The only complications have been some mild hematomas at the site of the aspiration. The method avoids surgical biopsy and sutures and takes about 7--8 min.


Investigation of the health status of Michigan chemical corporation employees. Clinical findings are reported for a group of 55 employees of the Michigan Chemical Corporation which manufactured FireMaster BP-6 from 1970 to 1974, in addition to a variety of other halogenated fire retardant chemicals. The results are compared with those from a group of male farm residents and consumers from Michigan examined at the same time. An increased prevalence of chest and skin symptoms was observed, compared with farmers. Skin symptoms were more prevalent among former PBB production personnel. Musculosketal symptoms were less prevalent among these workers than among farmers. Serum PBB concentrations are signicantly higher than among farmers. Blood chemistry results were similar for workers and farmers. However, both groups exhibited a significantly higher prevalence of elevated liver function tests (SGOT, SPGT) than a control population of nonexposed farmers. Both farmers and chemical workers showed an association of elevated CEA with serum PBB greater than 10 ppb.


Polybrominated biphenyl (PBB) contamination of the Pine River, Gratiot, and Midland Counties, Michigan. Michigan Chemical Corporation, St. Louis, Michigan manufactured PBB from 1970 until November 20, 1974. Studies in 1974 showed significant quantities of PBB in effluent discharged from the facility and in water, fish, ducks, and sediments from the Pine River. Fish uptake rates and bioconcentration factors were estimated. Followup surveys over the three year period since the termination of PBB production indicate a decline in PBB loadings to the river but no significant corresponding decline of PBB levels in sediments, fish and duck tissue. A Michigan Department of Public Health warning against consumption of Pine River fish from St. Louis downstream to its confluence with Chippewa River remains in effect.


Carcinoembryonic antigen (CEA) plasma levels in Michigan and Wisconsin dairy farmers. Carcionogenic embryonic antigen (CEA) titers were determined for 611 Michigan farmers exposed to PBB and a control nonexposed population of 139 Wisconsin farmers. The overall prevalence of elevated CEA titers was slightly higher in the Michigan study group, but the difference was not statistically significant. Smoking and/or significant past or present conditions (inflammatory bowel disease, ulcers, polyps, liver disease, chronic lung disease, or malignancies) were found to result in higher prevalence of elevated CEA titers, in both the Michigan study group and the Wisconsin control group, thus confirming previous reports. In addition, serum PBB concentrations appeared to be positively correlated with CEA titers. The possibility that the effect of PBB concentrations appeared to be positively correlated with CEA titers. The possibility that the effect of PBB may be additive to that of other factors which are known to result in an increased prevalence of elevated CEA titers is discussed.


Comparative neurobehavioral study of a polybrominated biphenyl-exposed population in Michigan and a nonexposed group in Wisconsin. An analysis of findings regarding the prevalence and time course of symptoms and the results of neurobehavioral testing among Michigan and Wisconsin dairy farmers, is reported. Reviewed are: (1) differences in the prevalence of neurological symptoms at the time of examination; (2) differences in the incidence and time course of symptoms for the period 1972--1976; (3) differences among populations and subgroups (sex and age) regarding performance test scores; (4) correlations between performance test scores and neurological symptoms; and (5) correlations between serum PBB levels as indicators of exposure and performance tests and neurological symptoms.


PBB homologs in sera of Michigan dairy farmers and Michigan chemical workers. In November, 1976 the Environmental Sciences Laboratory of the Mount Sinai School of Medicine, examined 1029 persons in Michigan for the potential health effects of exposure to PBB. Included in this group were 55 persons from Michigan Chemical Corporation and 237 farming families. Blood serum was analyzed for PBB. FireMaster FF-1 contains several bromobiphenyl components, the major constituent being approximately 60% 2,4,4,2'4'4'-hexabromobiphenyl. Other PBB homologs identifiable as peaks by gas chromatography--mass spectrometry include two pentabromobiphenyl peaks, three additional hexabromobiphenyl peaks, and two heptabromobiphenyl peaks. The relative concentrations, with respect to the major hexabromobiphenyl peak, of these minor constituents (1--20%) of PBB were different for persons from Michigan Chemical Corporation and for farmers. Penta-, hexa-, and hepta-bromobiphenyl components in serum samples analyzed from farming families from Michigan Chemical Corporation employees were compared with relative concentrations of these homologs in FireMaster mixture and in tissue and blood samples from rats fed FireMaster FF-1. Varying concentrations of these PBB components are attributed to different routes of exposure and the relative ease of metabolism and excretion of one pentabromobiphenyl component.


Unanticipated prevalence of symptoms among dairy farmers in Michigan and Wisconsin. Adverse human health effects resulting from exposure to polychlorinated biphenyls (PBBs) were unknown until the accidental contamination of Michigan dairy farm animal feed in 1973-1974. Human exposure resulted from the consumption of contaminated meat, milk, dairy products, and eggs. In November 1976, the Environmental Sciences Laboratory conducted comprehensive examinations of 933 farmers and residents in Mighigan who were likely to have consumed farm products contaminated with PBB. A comparison group of 229 Wisconsin dairy farmers were examined in March 1977 and the same scientific and medical staffs that conducted the Michigan survey were responsible and the same procedures used. A complete history of symptomatology by organ system, including year of first onset, duration, frequency, and severity of each symptom was obtained by a physician on all adults examined. Statistical analysis of the prevalence of symptoms at the time of examination or during the preceding year in the Michigan and Wisconsin populations studied found the Michigan group to have a significantly higher prevalence of skin, neurological and musculoskeletal symptoms. The increase was seen among the younger age groups 16-35 and 36-55. Michigan females had a higher prevalence of neurological symptoms than the Michigan males. The existing differences could not be explained without considering an etiologic role for exposure to PBB.


Effects of polybrominated biphenyls (PBB) on immune response in rodents. Studies were performed to investigate the effects of FireMaster FF-1, a chemical fire retardant consisting of a mixture of polybrominated biphenyls (PBB), on immune functions in mice and rats. Animals received 22 daily treatments of 0.03, 0.3, 3.0, or 30 mg PBB/kg body weight in a period covering 30 days. PBB exposure severely depressed cell mediated immunity in both mice and rats at the higher dosage levels as indicated by depressed responsiveness of splenic lymphocytes to mitogenic stimulation by polyclonal T-cell activators. Additionally, humoral immunity was depressed in mice at the 30.0 ppm dosage level. Assays for humoral immune functions included antibody production, serum immunoglobulin levels, and mitogenic stimulation of splenic lymphocytes to a polyclonal B-cell activator. These studies indicate that PBB exposure can lead to suppression of both humoral and particularly cell-mediated immune responses.


Effect of polybrominated biphenyls on hepatic excretory function in rats and mice. The purpose of this investigation was to determine the influence of polybrominated biphenyls (PBBs) on hepatic excretory function in developing and adult rats and mice. Prenatal or postnatal dietary exposure to PBBs (50 ppm in diet of pregnant or lactating mother or in diet of rat weanlings) resulted in elevated liver weight in developing rats. In 15-day-old rats that had been treated with PBBs increased liver weight correlated to enhanced ouabain transport from plasma into bile. Liver weight was also elevated in 21, 35, and 49-day-old rats exposed to PBBs, but this effect was not associated with stimulation of ouabain transport in these animals. However, adult rats fed 100 ppm PBBs for two weeks had significantly lower plasma concentrations of sulfobromophthalein (BSP) and increased biliary excretion of BSP, when compared to controls. PBBs-fed adult rats also excreted a greater percentage conjugated BSP (BSP-GSH) into bile. Two week dietary treatment of 100, 150, and 200 ppm PBBs resulted in enhanced initial disappearance of indocyanine green (ICG) from plasma of adult mice. However, dietary doses of 100 and 200 ppm PBBs to adult mice was not associated with enhanced capacity for ouabain excretion. In contrast, treatment with PBBs through the mother's diet (50 ppm) resulted in an almost twofold increase in cumulative ouabain excretion in 15-day-old mice. The results suggest that PBBs stimulate hepatic drug elimination in rats and mice, but the magnitude of the effect is dependent on age and transported compound.


Potentiation of hepatic and renal toxicity of various compounds by prior exposure to polybrominated biphenyls. Mice ingesting a standard rodent diet supplemented with polybrominated biphenyls (PBBs) were more susceptible to chlorinated hydrocarbon solvent-induced renal and hepatic damage, as well as the lethal effects of CHCl3 and CCl4, than were mice consuming control diet. As little as 0.025 ml/kg CHCl3 caused a significant increase in serum glutamic oxaloacetic transaminase (SGOT) and blood urea nitrogen (BUN) and a significant decrease in renal cortical slices accumulation of p-aminohippurate (PAH) in PBB-pretreated but not control mice. SGOT and serum glutamic pyruvate transaminase (SGPT) were greater in PBB-pretreated mice than in control mice after 0.125 and 0.005 ml/kg CCl4, respectively. Renal cortical PAH accumulation was greatly reduced in PBB-pretreated but not control mice aftter 0.125 ml/kg CCl4. The solvent-induced decrease in PAH accumulation was also greater in PBB-pretreated mice than in control mice following administration of 1.0 ml/kg trichloroethylene (TRI) and 0.15 ml/kg 1,1,2-trichloroethane (TCE).


PBB fed to immature chickens: its effect on organ weights and function and on the cardiovascular system. In a series of polybrominated biphenyl (PBB) feeding trials in White Leghorn cockerel chicks, the effects of PBB on various physiological parameters and organ weights were determined. These measurements included: growth, thyroid function, ECG, cardiac output, blood pressure, hematocrit, erythropoietin levels, and spleen, bursa of Fabricius, thyroid, testes, comb, liver weights. In addition, tissues were histologically examined. PBB, in the commercial grade form of hexabromobiphenyl, was administered continuously in the diet at levels ranging from 50 to 250 ppm. Pair-feeding studies were conducted to ascertain the effects of the drug per se since preliminary trials indicated that PBB administration resulted in decreased feed intake. Chronic administration resulted in depressed body weight as a result of decreased feed intake; decreased comb, testes, spleen, and bursa weights; increased liver and thyroid weight; hydropericardium and ascites; decreased hematological values due to depressed plasma erythropietin levels; decreased heart rate, packed cell volume, hemoglobin and cardiac output; decreased voltage amplitude of the ECG and a shift in the mean electrical axis. No significant differences were found in blood pressure (mean arterial and pulse pressure), stroke volume, or respiratory rate.


Behavioral and neurological toxicity of polybrominated biphenyls in rats and mice. Male, albino rats of the F-344/N strain and mice of the B6C3F1 strain were dosed by gavage, 5 days per week for a toal of 22 doses with 0.03--30 mg/kg of FireMaster FF-1, 0.168-16.8 Mg/kg of 2,4,5,2',4',5'-hexabromobiphenyl (HBB), or corn oil vehicle. A battery of tests was administered at the end of repeated dosing (30 day examination) and 30 days after dosing ceased (60 day test). FF-1 and, to a much lesser extent, HBB decreased body weight and performance on a variety of tests designed to detect neuromuscular dysfunction. Included in these tests were activity in the open field, forelimb grip strength, and muscular reflexes. Visual placement responses were also decreased in some animals, while hypothermia was observed in others. Emotionally, as measured by the number of defecations and urinations in the open field, was not affected by exposure to either compound. At the end of 30 day test, mice were less affected by exposure to these polybrominated biphenyls (PBBs) than rats; rats tended to worsen during the 30 days of no dosing, while mice tended to improve. These experiments indicate that oral dosing with levels of PBBs below those required to produce signs of acute toxicity produced behavioral or neurological toxicity when given repeatedly.


Persistent liver lesions in rats after a single oral dose of polybrominated biphenyls (firemaster FF-1) and concomitant PBB tissue levels. In a preliminary study, 12 male and 12 female weanling Sherman strain rats were given a single dose of 1000 mg polybrominated biphenyls (PBBs) FireMaster FF1 Lot 7042 kg/body weight as a 5% solution in corn oil. Three male and three female weanling rats were given corn oil. One day after dosing PBB blood levels ranged from 78 to 162 ppm and 42 days later they ranged from 1.1 to 2.99 ppm. The liver was the only organ with pathological changes. In a long-term recovery study groups of 20 male and female rats, 2 months old, were given 0 or 1000 mg PBBs/kg body weight as a single dose in peanut oil. Five rats per group killed 2, 6, 10, and 14 months after dosing had pronounced liver pathology, including hepatic porphyria in the female rats and neoplastic nodules also mainly in female rats. Chemical analyses of blood, liver, and adipose tissue for PPBs 10 and 14 months after dosing gave the following mean results. Blood levels in females were 2.9 and 2.92 ppm, respectively, and males 0.94 and 1.34 ppm, respectively. Adipose tissue levels in females were 1202 and 783 ppm and in males 713 and 866 ppm, respectively. The liver levels in females were 37 and 22 ppm and in males 60 and 63 ppm, respectively.


Environmental considerations for the disposal of PBB-contaminated animals and wastes. Accidental contamination of livestock feed in 1973 by polybrominated biphenyls (PBB) led to the destruction of over 30,000 animals in Michigan. Animal carcasses of mostly dairy cattle along with some beef cattle, hogs, sheep and rabbits destroyed under the Federal Food and Drug Administration guidelines were disposed on the land at an environmentally safe site in Kalkaska County, Michigan. The geology and hydrology of the disposal site on state-owned land is considered favorable for the disposal of contaminated carcasses and to prevent any migration of PBBs into ground and surface waters of the area. Materials underneath the site are predominantly sand with layers of silts and clays of glacial origin. The vertical isolation from the surface to the water table is over 90 ft, and the horizontal isolation to the privately owned properties and surface water bodies is well over 1.5 mile in all directions. The site design provides necessary safeguards for minimizing surface water infiltration into disposal trenches and maximizing the protection to the environment. A series of water wells in the direction of flow are established for monitoring groundwater quality for years to come. A 40-acre Gratiot County landfill located near St. Louis, Michigan, has received 269,000 lb of wastes containing 60 to 70% PBBs between 1971 and 1973. PBB wastes are intermixed with general refuse at various depths predominantly in the eastern half of the landfill. Phase I of the hydrogeological investigation shows that the landfill is situated immediately above the groundwater aquifer and a divide. Recently drilled test wells show traces of PBBs in the aquifer in all directions. Additional studies are planned in the near future for corrective measures and monitoring.


EM changes and other toxic effects of firemaster BP-6 (polybrominated biphenyls) in the mouse. Groups of Swiss ICR mice were fed 1000 ppm polybrominated biphenyls (FireMaster BP-6) in rodent chow for 4, 8, 11, and 14 days. Control groups were fed standard rodent chow without FireMaster BP-6. Animals were killed at the end of each feeding period and the livers examined by electron microscopy. EM changes noted were progressive increase in size of hepatocytes, a decrease in rough endoplasmic reticulum, a marked increase in smooth endoplasmic reticulum, mitochondrial degeneration, increased lysosomes, and a decrease in glycogen. In addition, there was increasing proliferation of microvilli in bile canaliculi with increasing feeding times. A group of mice fed 1000 ppm FireMaster BP-6 in rodent chow for 11 days had livers with a mean of 13.93% of total body weight as compared with 6.49% for the control group (P=0.02). Tissue distribution following ingestion of 100 ppm FireMaster BP-6 for 14 days was studied. Twelve weeks post-feeding, the tissue concentrations of hexabromobiphenyl in order of highest concentration to lowest were as follows: perithymic fat, perirenal fat, adrenal glands, thymus gland, liver and stomach.


PBB fed to adult female chickens: its effect on egg production, reproduction, viability of offspring, and residues in tissues and eggs. Two experiments with light breed laying chickens fed polybrominated biphenyl (PBB) as FireMaster FF-1. The first involved feeding PBB at dietary levels of 0.2, 1, 5, 25, 125, 625, and 3125 ppm, the second involved levels of 30, 45, 60, 90, and 120. Each group had 24 hens, and each experiment had a control group of 24 hens. PBB diets were fed for 5 weeks. Feed intake, production, reproduction, tissue residues and viability of offspring were monitored during that time and a subsequent 8 weeks. Production, hatchability, and viability of offspring were significantly affected by feeding PBB at 45 ppm. Marked inanition occurred at levels of 625 and 3125 ppm, and there was some loss of feed intake at 125 ppm. There was a return to normal production and hatchability in 3 to 4 weeks after PBB withdrawal of diets with levels of 125 ppm or less. Dose--response lines are presented for PBB in muscle, liver, kidney, adipose tissue, and eggs. Withdrawal curves for PBB from these tissues are also given.


Pediatric health aspects of PBBs.

Pediatric health aspects of PBBs. Many Michigan farm children are viewed as having suffered a deterioration in health coincident with contamination of dairy cattle by PBBs. Among Wisconsin dairy farm children such a deterioration was a rare event. There is a suggestion that Michigan farm children who had multiple symptoms during the 1973-76 period are getting better. A discrete syndrome of ill health has not been identified among Michigan farm children. The symptoms complained of indicate a disruption of various physiological functions, including neurobehavioral, gastrointestinal and immunological. Children from quarantined farms do not have a higher prevalence of multiple symptoms than those from nonquarantined farms.


Embryotoxic effects of polybrominated biphenyls (PBB) in rats. Pregnant Sprague Dawley rats were given 0, 0.25, 0.5, 1, 5, and 10 mg of a commercial polybrominated biphenyl, FireMaster BP-6 (PBB), in olive oil by gavage each day from days 7 through 15 of pregnancy. Laboratory chow and water were given ad libitum. Treatment with PBB had no significant effect on body weight gain, food and water consumption, and urine production. The mothers were killed on day 20, and the only significant effect observed was an increased liver weight of those given 1, 5, and 10 mg PBB. Spleen, kidney, ovarian, gravid uterine, and perirenal fat pad weights were similar to those of control mothers. PBB had no significant effect on number of live/dead fetuses, crown-rump length or fetal weight. No grossly malformed fetuses were observed in PBB-treated mothers. The effects of PBB transfer from mothers to nursing pups was studied by reciprocal exchange of pups between control mothers and mothers treated with 10 mg PBB. When the pups were 21 days old, they were weaned and fed control chow. The following four combinations of prenatal-postnatal exposure were studied: control-control (C:C); control-PBB (C:PBB); PBB-control (PBB:C); and PBB-PBB. Although the birth weights of pups from PBB-treated mothers were similar to those of the controls, body weights of 60-day-old males exposed prenatally and postnatally (PBB:C, C:PBB, and PBB:PBB) were less (p less than 0.05) than those of the controls (C:C). The weights of the perirenal fat pads of male and female pups exposed to PBB were less (p less than 0.05) than the control. Liver weights, on a body weight basis, were higher in male and female pups exposed to PBB. Vaginal openings were delayed; the percentages of 36-day-old pups with open vaginas were 50 (C:C), 38 (PBB:C), 28 (C:PBB), and 30 (PBB:PBB).


Characteristics of cytochrome P-450 and mixed function oxidase enzymes following treatment with PBBs. The mixture of PBBs in FireMaster BP-6 has been demonstrated to constitute potent inducers of hepatic and extrahepatic mixed function oxidase (MFO) enzymes. Chronic dietary administration of PBBs to mature female rats results in a "mixed" pattern of induction, with increases in both cytochrome P-450 and P1-450 associated enzymes. Acute administration of PBBs (150 mg/kg IP) to mature female rats resulted in a time-dependent induction of MFO activities; the P-450-dependent enzymes were simulated early (24-48 hr after administration) while the P1-450 dependent enzymes reached maximal activities at later time points. However, studies of the kinetics and patterns of inhibition of the induced enzymes along with gel electrophoresis studies of the microsomal proteins indicate that PBBs may induce different proteins from those induced by the classical P-450 and P1-450 inducers, phenobarbital and 3-methylcholanthrene. In addition, the pattern of enzyme induction caused by PBBs in developing rats differs from that in adults, in that the P1-450-associated enzymes are stimulated prior to the P-450-associated enzymes. The overall pattern of enzyme induction in extrahepatic tissue differs from that seen in the liver and sex differences in enzyme induction have also been demonstrated. As modifications of MFO activity may alter the toxicity of chemicals, these findings suggest that the toxicity of chemicals may be altered in animals exposed to PBBs and that these toxicities may exhibit age, sex, and organ specificities different from those seen in control animals.


Induction of monooxygenation in rainbow trout by polybrominated biphenyls: a comparative study. Two commercial polychlorinated biphenyl mixtures (Aroclor 1254 and Aroclor 1242) and one polybrominated biphenyl mixture (FireMaster BP-6) were examined for their abilities to induce hepatic microsomal monooxygenation in rainbow trout (Salmo gairdneri). Pretreatment of rainbow trout with Aroclors 1254 and 1242 (150 mg/kg IP) resulted in an approximate 10-fold induction of arylhydrocarbon (benzo[a]pyrene) hydroxylation, ethoxycoumarin-O-deethylation and ethoxyresorufin-O-deethylation within 7 days after injection. These enzyme activities remained elevated above control values for at least 2-3 weeks. Administration of FireMaster BP-6 (150 mg/kg IP) also resulted in an induction of several monooxygenase activities. Arylhydrocarbon (benzo[a]pyrene) hydroxylation, ethoxycoumarin-O-deethylation and ethoxyresorufin-O-deethylation were increased by 6-, 3,- and 25-fold, respectively. Only the latter two activities remained elevated two weeks post-injection. Ethylmorphine-N-demethylation was unaffected by the polyhalogenated biphenyls. Significant increases in P-450 hemoprotein were not observed after pretreatment with any of the polyhalogenated biphenyls studied.


Family clustering of PBB and DDE values among Michigan dairy farmers. Family clustering of varying levels of serum PBB and DDE was evaluated for 62 Michigan families by using the mean and standard deviation for family units, ranked percentile comparison of family members, and correlation by linear regression of family members. The results indicate that levels of serum PBBs cluster within family units and for children within families. Serum DDE clustering within families occurs only for children. These findings are consistent with recent, interim PBB exposure, perhaps from a common dietary source for families. Serum DDE represents a lifetime, low-level exposure to DDT-DDE which has been both less prolonged and less intense for children. Thus similar levels, or clustered serum DDE, was observed for children within families.


Toxicity of firemaster FF-1 and 2,2',4,4',5,5'-hexabromobiphenyl in cultures of C3H/10T 1/2 mammalian fibroblasts. A procedure for purifying 2,2', 4,4',5,5'-hexabromobiphenyl (HBB) from FireMaster FF-1 in gram amounts by crystallization is presented. Following purification, the structural assignment of HBB was made by using proton and carbon-13 nuclear magnetic resonance (NMR) spectroscopy, elemental analysis, and mass spectroscopy (MS). The growth of C3H/10T 1/2 cells treated with 5, 37, 75, and 150 microgram of HBB and FF-1 per milliliter of medium was measured at 4, 8, and 13 days following treatment. FF-1 was more toxic at 37 and 75 microgram/ml at both 4 and 8 days, but the same at 13 days. At 150 microgram/ml cell growth was completely inhibited by both compounds. Growth of cells was stimulated at 5 microgram/ml, by HBB at 4 and 8 days, and FF-1 at 8 and 12 days. HCB was compared with HBB and FF-1 for cell growth toxicity at 37 and 75 microgram/ml. At 75 microgram/ml, HCB was more toxic than HBB and FF-1 during the entire time period. At 37 microgram/ml, HCB was more toxic than HBB and FF-1 at 4 and 8 days. Cells seeded at high densities and treated with HBB for three days lost the high degree of postconfluence inhibition of cell division observed in control cultures. Cells treated with FF-1 for three days did not adhere well to the plastic growth surface. Ultrastructural features of the HBB- and FF-1-treated cells included decreased surface villi and increased lysosomes relative to the control cells.


Effect of polybrominated biphenyls on adenylate cyclase activity in rat lung alveoli. The in vitro effect of polybrominated biphenyls (FireMaster BP-6) on adenylate cyclase [ATP pyrophosphate lyase (cyclizing), EC 4.6.1.1] activity in plasma membranes of rat lung alveoli was determined. Two fractions of plasma membranes, PM-I (approximate d = 1.13 g/cm3) and PM-II (approximate d = 1.16 g/cm3) were isolated under hypotonic homogenization and fractionation conditions (1mM NaHCO3, 5mM dithiothreitol, pH 7.5) by using differential centrifugation and nonlinear sucrose gradient centrifugation techniques. Polybrominated biphenyls (10 microgram/ml) dissolved in ethyl acetate (0.25%) stimulated basal adenylate cyclase activity of plasma membranes by 2.1 to 2.5-fold. The implication of this observation is discussed.


Liver function tests among Michigan and Wisconsin dairy farmers. Serum activity of SGOT, SGPT, LDH, and alkaline phosphatase was measured in 614 Michigan adults exposed to PBB and 141 Wisconsin adults not so exposed. The Michigan group had higher prevalence of abnormal SGOT (p less than 0.005) and SGPT (p less than 0.005). A clear sex difference was observed. Michigan men had a higher prevalence of abnormal SGPT (p less than 0.005) and LDH (p less than 0.005) than Michigan women, and a higher prevalence than Wisconsin men of abnormal SGOT (p less than 0.005) and SGPT (p less than 0.01). These differences could not be ascribed to differing patterns of alcohol consumption, laboratory error, or choice of criteria for normality/abnormality. Seven Michigan subgroups were defined on the basis of the criteria by which they had been selected to participate. The two subgroups who were essentially self-invited did not differ from the remaining five randomly selected subgroups combined in prevalence of these abnormal liver function tests. Based on 364 serum PBB analyses thus far analyzed of the 614 Michigan participants, no obvious relationship between serum PBB values and liver function tests was observed. However, this is a tentative conclusion that will be further evaluated when remaining serum PBB analyses are completed. The greater prevalence of abnormal SGPT and SGOT among Michigan dairy farm residents compared to the Wisconsin dairy farm residents is tentatively ascribed to the former group's exposure to PBB.


Polybrominated biphenyl toxicosis in rats fed an iodine-deficient, iodine-adequate, or iodine-excess diet. Young male Sprague-Dawley rats were fed 0, 1, 10, or 100 ppm of polybrominated biphenyls (PBB) in iodine-deficient, iodine-adequate (0.2 ppm), or iodine-excess (1000 ppm) diets. Six rats in each of the 12 groups were killed at 30 days and the remaining six in each group at 60 days. Growth rates were similar in all rats fed diets containing 0, 1, or 10 ppm PBB but were slower from 30 to 60 days in rats given 100 ppm PBB. Results of routine hematologic examinations and urinalyses were essentially normal. Although liver weights were substantially increased by PBB, the smallest increases were in rats fed an iodine-deficient diet. Thyroid weights were increased by iodine deficiency and by 10 and 100 ppm PBB. Electropherograms of serum proteins, serum lipoproteins, and LDH isozymes at 60 days from rats given PBB indicated hepatic alterations, but changes were least dramatic in rats fed an iodine-deficient diet plus PBB and most severe in rats fed iodine-excess diets plus PBB. Hepatic lesions were basically similar to those previously described except that bile duct proliferation was seen at 60 days only in rats fed an iodine-deficient diet and 100 ppm PBB. Histologic changes in thyroid glands were associated with iodine deficiency and with PBB. The iodine-excess diet plus 100 ppm PBB induced squamous metaplasia of respiratory bronchiolar epithelium. These results indicate interrelationships between PBB and iodine which may affect the toxicosis caused by PBB.


Formation of brominated dibenzofurans from pyrolysis of the polybrominated biphenyl fire retardant, firemaster FF-1. The polybrominated biphenyl (PBB) fire retardant, FireMaster FF-1, was pyrolyzed for 20 min at 380-400 degrees C in open glass tubes and in tubes sealed after nitrogen flushing. The pyrolyzed residue was extracted with benzene, and extracts were cleaned up on columns of graphite (Carbopack A) and alumina. Analysis was carried out by low resolution direct probe mass spectrometry (MS). Spectra from extracts of the open tube pyrolyzed material had a series of ions characteristic of tetra- and pentabrominated dibenzofurans as evidenced by comparison with spectra from 2,3,7,8-tetrabromodibenzofuran (TBDF). Confirmatory evidence for the brominated dibenzofurans was obtained by high resolution MS dual ion analysis of certain fragment and molecular ions. Recovery values of TBDF through the cleanup procedure averaged 50% and, using this recovery value and TBDF as an external standard, dual ion analyses indicated that 40 ppm tetra- and 4 ppm pentabrominanted dibenzofuran were produced based on the PBB level used in the pyrolysis experiments. Additional analysis of the open tube pyrolyzed material by gas chromatography/mass spectrometry provided evidence that there was one tetrabromodibenzofuran compound with a retention time equal to that of TBDF. Trace levels (less than 1 ppm) of the molecular ion of tetrabrominated dibenzofuran were found after analysis by low resolution MS of the PBB pyrolyzed under nitrogen in sealed tubes. The experimental evidence is consistent with a mechanism for brominated dibenzofuran formation involving attack of oxygen on PBB compounds.


Assessment of the hazards of polybrominated biphenyls. During their peak use period, PBBs represented under 1% of the total sales of fire retardant chemicals, and very probably would have escaped intensive study if they had not been mixed accidentally with animal feed preparations. Instead, international attention was drawn to PBBs by the state-supervised killing of over 35,000 cattle which had been contaminated with PBBs. Interestingly, low doses of PBBs exert a broad spectrum of toxicological, pharmacological, and biochemical effects despite low acute toxicity. These effects and the intensive bioaccumulation of PBBs derive from their structure and their consequent resistance of biotransformation and high solubility in fat. In rodents, PBBs are teratogenic, immunosuppressive, and potentially carcinogenic. In bovine, rodent, and avian species, PBBs reduce feed intake and induce mixed function oxidases of liver microsomes. The latter effect may be responsible for steroid level changes which underline hormonal toxicities observed in cows, mink, rats, and chickens. The effects of PBBs on humans are controversial, but data suggestive of immunological, skin, and liver disorders continue to accumulate. Concern about the clinical effects of PBBs is heightened by the knowledge that these compounds readily enter the fetus by crossing the placental barrier and can be transferred to newborn children after extensive passage into breast milk.


Effects of processing and cooking on PBB residues. To study the effect of processing on polybrominated biphenyl (PBB) levels, milk from four dairy herds containing less than 0.3 ppm (fat basis) of physiologically incorporated PBBs was presented individually into cream, skim milk, butter, and stirred curd cheese. Pasteurized and freeze-dried whole milk, skim milk, and cream, spray-dried whole milk and skim milk, and condensed whole milk were also made. PBBs were concentrated in the high-fat products. Spray-drying reduced PBBs in whole milk and skim milk while pasteurization, freeze-drying, aging of cheese, and condensation were not effective. To study the effect of cooking on PBB levels, thigh meat, thigh skin, drumstick and breast (with skin) from half of chickens fed PBBs were analyzed raw, and pieces from the other halves were analysed following separate pressure cooking. The level of PBBs expressed as parts per million on a solids basis was lower in the cooked sample than in the corresponding raw piece and part of the PBBs lost were found in the drip. Recoveries of PBBs in cooked tissue and broth ranged from 68.1% in the thigh skin to 84.6% in the drumstick, with approximately two-thirds of the recovered PBBs found in the cooked meat itself. Therefore, pressure cooking resulted in a loss ranging from 36% for the drumstick to 53% for the thigh skin.


Distribution and kinetics of PBB residues in cattle. Cows fed a constant amount of polybrominated biphenyl (PBB) reached a steady-state concentration in milk fat within 30 days. This concentration was approximately four times the concentration in the total diet. When feeding of PBB was stopped, the concentration in milk was adequately described as a sum of two first-order elimination rates. Biological half-life in environmentally contaminated cows, studied for 6 months about a year after contamination, was 60 days. The stage of lactation affected the rate of elimination, and in some concentrations increased shortly after calving. Residues were distributed in body tissues proportionally to concentration of fat in the tissues. Liver and brain were exceptions. Concentration in liver fat was generally higher than other tissues and possibly related to the treatment of some cows with phenobarbital. Residues in brain fat were significantly lower than all other tissues. The ratio of the concentrations in milk fat to concentration of residues in the blood of calves and fat of fetal tissues to the concentration in the corresponding tissue in the dams was 0.36:1. It was estimated that people consuming milk from the highly contaminated Michigan cows could have received PBB doses as great as 10 g from this source alone.


Distribution of polybrominated biphenyls after dietary exposure in pregnant and lactating rats and their offspring. Female rats were fed PBBs in the diet (50 ppm) from day 8 of gestation to day 21 of gestation, from day 1 postpartum to day 14 postpartum or from day 8 of gestation through day 14 postpartum. Levels of PBBs were measured in various tissues. Small concentrations of PBBs (less than 5 microgram/g) were found in the brain, heart, lung, liver, small intestine, placenta, and gravid uterus. Larger concentrations (less than 30 microgram/g) were found in kidneys, the nongravid uterus, skin, mammary tissue, and fat. Lactation did not significantly alter the concentrations of PBBs found in tissues other than mammary tissue. Offspring were subjected to several exposure regimens by cross-fostering. Concentrations of PBBs in the neonatal livers were higher than in the adults nursing them. Transfer of PBBs via the milk appears to be much more important to appearance of PBBs in newborns than does placental transfer.


Effects of PBBs on cattle. IV. Distribution and clearance of components of firemaster BP-6. Sixty dairy animals were utilized in seven experiments to determine aspects of the distribution and clearance of FireMaster BP-6. Experimental protocols of various studies provided daily exposures from 0.25 to 25,000 mg, exposures for 1 to 202 days, and total study periods from 10 to 1100 days. Necropsy of 28 animals provided information on residue concentrations in 35 tissues, and the excretion in milk was determined in 15 animals. These studies showed that the major brominated biphenyls of this commercial mixture were absorbed from the gastrointestinal tract and appeared in the blood plasma within 4 hr. With continued exposure to the residue plasma concentrations reached a steady state by 15 days. Free PBB was not detectable in urine. During PBB feeding feces was the major route of excretion, representing approximately 50% of the amount fed to animals not displaying signs of toxicosis. Following a withdrawal of PBB, fecal concentrations declined to 1 to 2% of concentrations during dosing, yet, feces remained the major excretory route in nonlactating animals. In contrast, in post-exposure lactating animals milk fat became an important excretory route removing three-times the quantity of residue cleared in feces. Following parturition, concentrations of PBB in milk fat declined approximately twofold in 6 days. Thereafter, the residue concentration in milk fat was approximately 0.4 that in depot fats. PBB had a predilection for lipid tissues with similar concentrations in various depot fats. Concentrations of the residue were notably low in tissues of the nervous system despite the high content of lipid material. Liver contained residue concentrations that were disproportionately high when compared to the lipid content of the organ. Calves born to PBB-exposed cows had similar distribution of residues in body tissues although concentrations were less than those of the dam.


Immunological studies in cattle exposed to polybrominated biphenyls. The intactness of the immune system in cattle exposed to polybrominated biphenyls (PBBs) has been investigated by using several immunoassays. Eighty-seven animals have been studied, 35 control animals (not exposed to PBBs) and 52 animals exposed to PBBs (0.02-30 ppm/g fat equivalent). The immunoassays included a complete blood count, identification of peripheral blood T and B lymphocyte subpopulations, serum immunoglobulin levels (IgG, IgM, and IgA), the in vitro response to lymphocytes to phytolectins (PHA, Con A, PWM), the antibody response to Keyhole limpet hemocyanin (KLH), the cell-mediated response to PPD, and determination of autoantibodies and/or immunosuppressive serum factors. For control and PBB-exposed cattle, there was no statistical difference between the number of circulating erythrocytes or leukocytes, the hematocrit, or hemoglobin content; the percentage or number of T and B lymphocytes; the isotope incorporation index (DNA synthesis) of lymphocytes in response to mitogens; the concentrations of serum immunoglobulins IgG, IgM, or IgA; the mean peak titer to KLH; or in vivo or in vitro immune response to PPD.Additional evaluation of cattle with tissue levels of PBB greater than 3 ppm/g tissue for hematological and immunological parameters revealed no statistical difference from control animals. Other experiments were performed to evaluate serum from cattle exposed to PBBs for autoantibodies to smooth muscle, mitochondrial or nuclear antigens. No evidence for autoantibodies was observed. Further studies were done to examine the cytotoxic and/or immunosuppressive activity of sera from PBB-exposed animals. In these studies, the blastogenic response of lymphocytes from control cattle and humans were evaluated in the presence and absence of serum from animals exposed to PBBs (> 3 ppm/g tissue). No evidence for either a cytotoxic or an immunosuppressive influence of such sera was demonstrable. Our studies indicate that PBB, at the levels studied, does not alter or interfere with lymphocyte surface antigens, the complex nuclear and cytoplasmic events required for mitosis and cell division, or the biological events required for antibody formation and cell-mediated immune reactions. Further, PBB exposure at the levels studied does not predispose cattle to autoantibody production or leucotoxic serum factors.


Effects of PBBs on cattle. I. Clinical evaluations and clinical chemistry. Toxicosis was induced in pregnant heifers by feeding 25,000 mg/head/day of FireMaster BP-6, a commercial blend of polybrominated biphenyls (PBB). The PBB feeding decreased dry matter intake approximately 50% by 4 days exposure. Emaciated animals became anorexic a few days prior to death at 33 to 66 days. Weight losses of heifers average 80 kg. Other clinical signs observed were dehydration, diarrhea, excessive salivation and lacrimation, fetal death, abortion, and general depression as evidenced by depressed heart and respiratory rates. Clinical signs were apparent after 10 days exposure and progressively intensified along with loss of condition until death. Clinicopathologic changes included significantly increased serum glutamic-oxaloacetic transaminase and decreased serum calcium by 30 days exposure. Lactate dehydrogenase, urea nitrogen, and bilirubin were elevated, and serum albumin decreased by 36 to 40 days. Principal urine changes were decreased specific gravity and moderate proteinuria. Pregnant heifers fed 0.25 or 250 mg/head/day for 60 days and nonpregnant heifers fed 250 mg/head/day for 180 days displayed neither clinical signs nor clinicopathologic changes indicating adverse effects from PBB exposure. Post-exposure, all heifers exposed to PBB for 60 days calved normally with zero calf mortality and were successfully rebred. Milk production was not different from control animals. Birth weights of calves from dams exposed to 250 mg PBB/head/day were significantly greater than calves of dams exposed to 0 mg or 0.25 mg/head/day. PBB exposure of dams produced no detrimental effects on calves as indicated by clinical signs, clinicopathologic changes, or performance.


Absence of plant uptake and translocation of polybrominated biphenyls (PBBs). Studies of polybrominated biphenyl (PBB) uptake by plants have been conducted in hydroponic solutions and in greenhouse experiments with soil. Autoradiograms of corn and soybean seedlings grown in hydroponic solutions showed no translocation of 14C-PBB from 14C-PBB-treated solutions to plant tops or within the leaf from 14C-PBB-treated spots on the upper leaf surface. A significant portion of the 14C-PBB associated with the roots was removed when the roots were dipped in acetone. Three root crops (radishes, carrots, and onions) were grown in two soils, each treated with a mixture of FireMaster BP-6 (PBB) and 14C-PBB to achieve final concentrations of 100 ppm and 100 ppb. All roots showed more PBB when grown in the soil with the lower clay and organic matter content than they did when grown in the soil with more clay and organic matter. In the latter soil (clay loam) no PBB was detected in any roots from the 100 ppb treatment. More PBB was associated with roots of carrot than of radish or onion. Corn leaf whorls containing dust from a PBB contamination soil and washed radishes from a heavily contaminated garden showed no PBB.