Enzyme-linked immunosorbent choriomammotropin assay.

We have devloped an enzyme-linked immunosorbent assay for determining choriomammotropin (human placental lactogen) in serum. Unlabeled hormone competes with choriomammotropin-beta-galactosidase conjugate for antibody bound to polystyrene tubes. The entire assay can be performed in 2.5 h with good precision. The coefficient of variation for one sample with a mean concentration of 5.6 mg/L, assayed 10 times on the same day, was 5.7%. The coefficient of variation for nine samples (3.5 to 9.0 mg/L) assayed on five different days was 7.9%. Forty-eight clinical samples were assayed (y) and compared with results obtained by radial immunodiffusion (x). The resulting regression equation was: y = 1.05x + 0.78; r = 0.91.


The influence of citrate and phosphate on the Mancini single radial immunodiffusion technique and suggested improvements for the determination of urinary albumin.

When adapting the immediate bromcresol green method for urinary albumin determination a correlation study with the Mancini single radial immunodiffusion (SRID) method was performed. This study showed large disparities between the two methods, SRID giving the higher results. The unsuitability of the currently used SRID methods is demonstrated and improvements to the method are suggested. Known amounts of albumin were added to urine samples as well as to a "synthetic urine", both giving falsely elevated results with the SRID method. On investigating the different components of the "synthetic urine", it was found that the disparities were due to the influence of citrate and phosphate. On addition of citric acid or phosphate to the dilution buffer and/or the gel buffer, the results of the SRID method agreed with those of other methods and with the expected values. The findings presented in this paper can probably be extended to other immunological methods too since it seems to be the antigen-antibody reaction which is affected.


Changes in serum lysosomal hydrolases in marasmus.

The activities of several lysosomal hydrolases including beta-glucuronidase, acid phosphatase and hexosaminidase were compared in serum from 19 well-nourished subjects and 13 children (age 5--24 months) who were suffering from marasmus. The marasmic children exhibited growth retardation and muscle wastage but had normal serum protein values and absence of psychomotor retardation or oedema. Significant changes were observed in serum beta-glucuronidase and acid phosphatase activities. Compared to the control group, serum beta-glucuronidase (determined at pH 4.5 using the fluorogenic substrate, 4-methylumbelliferyl-beta-D-glucuronide) was 2.3-fold higher (p less than 0.001) in the marasmic children. In contrast, serum acid phosphatase values were approximately 50% lower (p less than 0.01) in the marasmic population. Serum hexosaminidase values in the two groups under study were not significantly different. Determination of the beta-glucuronidase to acid phosphatase ratio permitted effective discrimination (p less than 0.001) of serum from normal and protein-calorie malnourished children. The finding that the elevated value of the beta-glucuronidase : acid phosphatase ratio (0.64--1.37) decreased to within the normal range of values (0.10--0.43) after nutritional rehabilitation of several marasmic cases indicates that the determination of serum lysosomal hydrolases using fluorogenic substrates might provide a rapid and sensitive quantitative method for objectively evaluating the status of protein-calorie malnourished children and their responsiveness to nutritional therapy.


ABO and Rh phenotyping of foetal blood obtained by foetoscopy.

Blood samples taken from the foetus by foetoscopy early in the 2nd trimester of pregnancy can be used for reliable blood grouping. However, the sampling technique is not perfect. In 5 out of 15 pregnant women, no foetal blood was obtained, although between 1 and 10 samples were taken on each occasion. One-third of 56 samples from the 10 women contained foetal blood. Contamination of samples with amniotic fluid varied, but did not interfere with the blood grouping. ABO and Rh typing of the foetus was possible even if the samples were contaminated with large amounts of maternal blood. In nine cases, blood grouping of the foetus was performed on red cells from foetoscopy samples. Control blood samples from the foetus following abortion were obtained in seven of these cases, and the foetoscopic results were confirmed.


Genetic structure of the Greek gypsies.

Data are presented on several polymorphic genetic markers in 200 Greek gypsies. Polymorphic loci studied were: the ABO, MN, Rhesus, Kell and Duffy blood groups, hemoglobin, and ceruloplasmin. A survey for congenital malformations and hereditary diseases was also carried out on this group. The ABO, Rhesus, MN and Duffy system frequencies varied significantly from the figures obtained for the Greek population. However, there is a characteristic similarity between various gypsy groups studied in other nations and the distribution of polymorphic traits in the Punjab region of India. Cystic fibrosis, renal tubular acidosis, 21-hydroxylase deficiency, Hoty-Oram syndrome and homozygous beta-thalassemia were diagnosed within the gypsy group studied.


Reduced red blood cell carbonic anhydrase activity in a patient with nephrolithiasis.

Low levels of red blood cell (RBC) carbonic anhydrase (CA) activity in hemoglobin-free hemolyzate (HFH) were found in 2 children with nephrolithiasis when compared to age-matched controls. The lowest levels were consistently found over a two year period in a 6 1/2 year old boy (U.P.) whose renal calculi contained uric acid, ammonia, calcium carbonate and oxalate. His RBC CA values ranged from 1.4-2.7 U/g Hb compared with levels of 3.9-5.3 U/g Hb in control subjects. Statistical comparisons of the mean values for U.P., 2.06 U/g Hb, and his age-matched control subjects, 4.46 U/g Hb, revealed a significant difference (P = less than 0.001). Similar reductions in RBC CA activity were found in his father--2.0 and 2.1 U/g Hb compared with 3.3 and 3.9 U/g Hb in adult controls. HFH CA activity was not decreased in the mother or sister. Polyacrylamide gel electrophoresis of HFH from 4 of the children and the father of U.P. was abnormal. However, this abnormal electrophoretic pattern could only be demonstrated when the gel was run for 120 minutes and not when it was run for 80 or 160 minutes. We have identified a patient and his father with low levels of RBC CA activity.


The contraction induced by ethylenediaminetetracetic acid-4 sodium (EDTA-4Na) in guinea-pig isolated vas deferens kept in isotonic sucrose medium.

1. Ethylenediaminetetracetic acid--4 sodium (EDTA--4Na) induced a contraction in guinea-pig isolated vas deferens kept in isotonic sucrose medium containing 0.07% Nahco3 and 0.1% glucose after preincubation in Locke's solution. The maximum contraction was 82.1 +/- 6.7% of that induced by added KCl. 2. EDTA--Ca--2Na (10(-7) to 10(-3) g/ml) induced no contraction in preparations kept in the sucrose medium. 3. The EDTA--4Na-induced contraction was also obtained when sucrose in the sucrose medium was replaced by glucose but not when the preparation was kept in Locke's solution. 4. The EDTA--4Na-induced contraction was decreased by reduction of the CaCl2 of KCl concentration of the Locke's solution in which the preparation had been preincubated before placing in the sucrose medium. 5. The EDTA--4Na-induced contraction was suppressed with increasing concentrations of NaCl used to partially replace sucrose in the sucrose medium. 6. When the preparation was kept in EDTA-free sucrose medium after pretreatment with 10(-4) g/ml of EDTA--4Na in the sucrose medium for 20 min, a contraction was induced by added CaCl2. The maximum contraction induced by CaCl2 was 30.1 +/- 11.8% of that of the EDTA--4Na-induced contraction, when CaCl2 was added at 20 min after the exchange of medium.


Heterotopic ossification in the lumbar region. Report of a case.

Delayed appearance of heterotopic ossification in the lumbar region occurred in a 34-year-old man with a high thoracic paraplegia. The occurrence of lumbar heterotopic ossification seems not to have been previously reported in the literature.


Cardiovascular and respiratory effects of carotid body stimulation in the monkey.

1. The carotid bodies were stimulated in the anaesthetized pig-tailed macaque monkey (M. nemestrina) using (i) brief injections of cyanide or CO2-equilibrated bicarbonate solution into a common carotid artery, and (ii) longer perfusion with hypoxic hypercapnic blood in vascularly isolated chemoreceptor preparations. 2. In spontaneously breathing animals brief stimuli (thirty-one tests, seven monkeys) consistently increased pulmonary ventilation (by 97 +/- 10% of control), slowed the heart rate (the pulse interval increasing by 36 +/- 7.5%), and increased femoral vascular resistance (by 44 +/- 7%). 3. More sustained chemoreceptor stimulation with asphyxial blood (nineteen tests, five monkeys) increased ventilation by 187 +/- 23%, but transient bradycardia occurred in only eight of nineteen tests and was followed by tachycardia; in the remaining tests, only tachycardia occurred. After 20--40s, the pulse interval was 5.8 +/- 0.9% below the control level. Femoral vascular resistance either increased (five tests, two animals) or decreased (six tests, two animals). 4. Evidence is presented that in the monkey the autonomic effects of chemoreceptor stimulation are influenced by the level of respiratory activity with bradycardia and vasoconstriction occurring when the level is low, and tachycardia and vasodilatation when it is high. 5. The interaction of autonomic responses resulting from carotid body stimulation and from mechanisms initiated by the concomitant hyperventilation are qualitatively similar in the monkey and in subprimate species, although there may be quantitative differences such as would account for the species differences to distrubances produced, for instance, by arterial hypoxia.


Physeal injuries of the ankle in children: classification.

Seventy-one cases of ankle fractures in children were studied. An analysis of the fracture lines involving the distal tibial and fibular epiphyses were done. A new classification is proposed based on the position of the foot at the moment of trauma and the direction of the abnormal force. Four mechanisms were considered, grades were described for each mechanism and each grade always occurred in the same order. In almost half of the cases, the supination-inversion mechanism was responsible for the injury. A correlation with the physeal injury classification of Salter and Harris is imperative, since complications such as varus deformity of the ankle are common when a Salter-Harris III or IV injury is present. The proposed classification has an important practical use. It facilitates closed reduction, because once the mechanism of injury is known, reduction is obtained by reversing the direction of abnormal force and immobilizing the foot in the opposite position given by the mechanism.


Surface replacement arthroplasty of the hip.

The principle of hip joint resurfacing is replacement of diseased joint surfaces and simultaneous restoration of the normal anatomy and biomechanical function to the maximal degree possible. This concept offers several theoretical advantages over conventional total hip joint replacement and the clinical results in this series of 426 cases appears to confirm the value of both the method and the concept. Successful joint resurfacing surgery with attention to detail. Most problems can be anticipated and handled appropriately. Complications are few. The operation should only be done in cases of severe hip disability, when the patient's level of suffering demands operative intervention and when the only reasonable alternatives are fusion, total joint replacement or head and neck resection. It is an operation designed and recommended as an "in-between" procedure to gain time against the progressive disease. Resurfacing should not be performed if conservative measures or classic hip osteotomies offer significant benefit. The principal advantages of this procedure relate directly to the prosthetic design. Only the joint surfaces are removed during surgery, most of the normal bone is preserved, the medullary canal is not opened, and the implants utilized are of small volume. As a result the risk of infection is low compared to other implant arthroplasty techniques and clinical statistics confirm this anticipated advantage. The operation is designed to interfere minimally with the normal joint mechanics so it is also anticipated that prosthesis longevity will be greater than when rigid stem prostheses are placed in elastic bone. As yet follow-up is too short to make valid judgments on this point. The technique is applicable to younger patients, however, because if it should, in time, fail and other surgical treatment becomes necessary the original alternatives of total hip replacement, arthrodesis, or head and neck resection remain available. Relief of pain is predictable and almost all patients have experienced significant improvement in function. The procedure has a broader indication in cases of prior bone or joint infection and is definitely a preferable procedure in young individuals with severe hip disability.


Nutrition of flexor tendons in monkeys.

The hydrogen washout technique was used to investigate the role of synovial diffusion versus vascular perfusion in the nutrition of monkey flexor tendons within the digital sheath. There was no significant difference in the uptake and washout of hydrogen tracer by tendons in contact with synovium but detached from the surrounding vasculature, compared to control tendons. However, there was insignificant uptake of tracer by tendons with intact vasculature, but separated from synovium. Synovial diffusion is a primary nutrient pathway of monkey flexor tendons within the digital sheath.


THARIES surface replacements: a review of the first 100 cases.

The first 100 cases of the 135 THARIES surface replacement procedures with 4--32 months follow-up, are evaluated in terms of clinical results, radiographic information and complications. The short but detailed follow-up suggests that this procedure was an excellent alternative for the younger and more active population. The overall hip ratings (pain, walking and function) and range of motion are comparable to that of stem-type total hip replacements. When the THARIES and conventional hip results are further compared in 34 patients matched by sex and age for 3 major etiological groups (osteoarthritis, osteonecrosis and congenital hip dysplasia), the interim results are essentially comparable for both types. Three cases in the first 100 have required revision. One patient (no. 1) with osteoporosis, chondrolysis and arthrofibrosis following slipped capital femoral epiphysis had loose femoral and acetabular components 24 months postoperatively. He was revised to a T-28 hip replacement. The polyethylene socket in another patient (no. 4), the first dysplastic hip in this series, was 20 nm uncovered superiorly, became loose and was revised 9 months postoperatively. Now 15 months postoperative with a more medial THARIES acetabulum, the patient continues to have a good result. Another patient (no. 12) with bilateral dysplastic hips became progressively more disabled due to heterotopic bone, which was then excised 18 months postoperatively. Radiographic studies of the THARIES sockets demonstrate radiolucent zones at the cement-bone interfaces of the acetabulum in 88 cases, partial in 51 and complete in 37. Three hips were currently considered to have evidence of progressive socket loosening but are active and asymptomatic. There have been no femoral neck fractures in this series which we attribute to the custom fitting ability inherent in the range of components, the reaming protocol and the various remodelling guides. There have been no prosthetic breakages, subluxations, dislocations or sepsis. The complications observed in this series are minor and comparable to that of many other total hip arthroplasty operations. Nerve palsy and trochanteric separation have not been major problems although one existing peroneal nerve dysfunction and one trochanteric migration emphasize the need to minimize these complications by careful handling of the operative leg, and an accurate trochanteric reattachment technique.


Endogenous prostaglandin synthesis inhibitor in the renal cortex. Effects on production of prostacyclin by renal blood vessels.

1. The capacity of various tissues of the porcine kidney to convert [1-14C]arachidonic acid into radiolabelled prostaglandins was studied. 2. Only after removal from the cortical matrix, were renal blood vessels able to convert arachidonic acid into prostaglandins (primarily prostacyclin). In contrast, convoluted tubules showed a low capacity to metabolize arachidonic acid. 3. The failure to demonstrate prostaglandin synthesis by renal cortical slices is related to the presence of an inhibitor of cyclo-oxygenase. Thus the addition of renal cortical incubate to isolated vascular tissues and ram seminal vesicles inhibited their ability to synthesize prostaglandins. 4. Slices of renal medulla metabolized arachidonic acid primarily to prostaglandin F2alpha; lesser amounts of prostaglandin E2 and prostacyclin were generated. 5. The large capacity of the renal vasculature to generate prostacyclin is consistent with an important role for this prostaglandin in regulation of renin release and renal haemodynamics.


Evaluation of polymer flock and metal alloy intra-tubal device in pigtail monkeys.

Two intratubal devices, one covered with a flock made from ethylene vinyl acetate and the other constructed of titanium-aluminum-vanadium alloy with an etched surface were evaluated after being placed in the Fallopian tubes of pigtail monkeys. In some instances, the devices were medicated with 10% silver nitrate or 50% quinacrine hydrochloride. The microflock device anchored in the tube mechanically, but there is no evidence that either the polymeric or metal alloy device formed an actual attachment with Fallopian tube epithelium. It is suggested that a mechanical design approach to ITDs might be more productive than one which assumes a tissue-device bond.


In vivo evaluation of cholesterol as a matrix for the controlled-release of progesterone.

Twelve adult, female baboons were used to test the rate and duration of progesterone (P4) release from cholesterol (C) pellets implanted subcutaneously. Four different preparations were tested (3 baboons per group). Each group was treated with pellets containing a different sized distribution and P4-cholesterol ratio (P4:C). Group 1 = 0.25--0.5 mm, 59% P4:41% C; Group 2 = 0.25--0.5 mm, 65% P4:35% C; Group 3 = 0.5 -1.0 mm, 59% P4:41% C; and Group 4 = 0.5 -1.0 mm, 65% P4:35% C. All baboons received the identical quantity of P4 (500 mg). Following treatment, blood samples were obtained at selected intervals, and the levels of progesterone were determined by radioimmunoassay. Serum-progesterone clearance curves were parallel for each experimental group. The rate of release of progesterone calculated from the serum clearance curves for groups 1--4 were: 42; 55; 29; and 38 mg P4/week. The mean duration of release for experimental groups 1--4 were 12, 9, 17, and 13 weeks respectively. The continuous progesterone treatment inhibited normal baboon ovarian function during the period of progesterone release.


[Studies on staphylococcal enterotoxin B. 1. Quantitative assay by capillary tube single diffusion test (author's transl)].

A method for rapidly identifying and quantitating small amount of staphylococcal enterotoxin B was developed on capillary tube single diffusion test. The high sensitivity of method has the advantages to save antiserum and incubation period. Some factors such as concentration of antigen and antiserum, incubation temperature, buffer systems, and position of capillary tube all might affect the test accuracy. If all those affecting factors were concerned, the toxin would probably be standardized. The most accessible quantitative estimation was accomplished by means of (1) plotting a precipitation curve from the different concentrations of standardized staphylococcal enterotoxin B; (2) the sample to be test was diluted by serial two-fold method with the same buffer system; (3) select suitable 3--5 dilutions to perform capillary tube diffusion test, taking the values of precipitation bands as abcissa and sample dilutions as longitudinal axis, rendered the precipitation curve of sample exquisitely paralled to that of the standard; (4) calibrate the figure values of sample exactly on the precipitation curve, and reading the toxin content of specimen to be tested from standard curve.


Mixed acid-base abnormalities in diabetes.

This study is a description of a patient who exhibited diabetic ketosis associated with an alkalosis rather than acidosis and a review of eight previously reported cases. Precipitating factors for this syndrome are severe vomiting with loss of hydrogen, potassium, and chloride ions, and dehydration. The ingestion of alkali may also result in this mixed acid-base disturbance. Treatment consists primarily of replacement of potassium and chloride. All reported patients had received large doses of insulin for initial therapy; however, limited insulin (20 U) therapy in this patient almost completely reversed the metabolic abnormality with 12 hours.


Toxicity tests on ciliates--a short review.

A variety of methods has been proposed to study the toxicity of chemicals or polluted waters on ciliates as representative test organisms of the microfauna of aquatic ecosystems. These methods are based on morphological, ultrastructural, ethological, or metabolic criteria. The techniques proposed are mostly species dependent and so are their applications and restrictions. In view of a future standardization of protozoan toxicity tests a selection has been made by the authors on the basis of the applicability of the tests for routine analysis. The methods withheld are briefly described and comments are made on their advantages and limitations. The equipment involved, the operational complexity, and the accuracy of the results and their interpretation are listed in a synoptic table.


Endocrine effects of chlorinated hydrocarbons in rhesus monkeys.

After the rhesus monkey was demonstrated to be a suitable model for man in both metabolic and endocrinological studies, effects of hexachlorobenzene (HCB) and polychlorinated biphenyls (PCB) on the pattern of sexual hormones in cycling female rhesus monkeys were investigated. After confirmed ovulation, four adult female rhesus monkeys were treated during the following cycle with 4 mg/kg/day of HCB, and four other monkeys were treated with the same dose of Clophen A 30. Ovulation was blocked in three PCB-treated and one HCB-treated monkeys. Whereas the levels of luteinizing hormone and follicle-stimulating hormone did not seem to be changed directly by the treatment, low estrogen levels were found during the anovulatory cycles. Studies with PCB- and HCB-treated superovulated rats indicated interaction of the chemicals with ovarian steroidogenesis. Altered hepatic steroid metabolism may also cause low estrogen levels in treated animals.


Environmental chemical-induced immune dysfunction.

Antibody formation, endotoxin sensitivity, and resistance to a challenge malarial infection were evaluated in mice fed a diet containing polychlorinated biphenyl (PCB) (Aroclor 1242) or hexachlorobenzene (HCB). Antibody synthesis to the antigen sheep RBC (SRBC) was significantly depressed in the PCB- and HCB-treated (167 ppm) animals as evidenced by the fact that control mice elicited an approximate twofold increase in antibody formation over the chemical-treated mice. Serum IgA concentrations in the PCB- and HCB-treated mice were consistently 40--80 mg/dl lower than control values. Gram-negative endotoxin (Salmonella typhosa) sensitivity in PCB- and HCB-treated mice was increased 5.2- and 32-fold, respectively, following the dietary administration of 167 ppm of Aroclor 1242 or HCB for 6 weeks. An endotoxin hypersusceptibility was also noted at 3 weeks after dietary administration. Decreased resistance to a malaria challenge was also demonstrated in the xenobiotic-treated mice. A 20% decrease in mean survival time of mice fed Aroclor 1242 for 3 to 6 weeks and inoculated with Plasmodium berghei (NYU-2) was observed. Infected mice which had received HCB for 3 or 6 weeks manifested reductions in mean survival time of 24 and 31%, respectively. The data indicated that environmental chemical contaminants impair host resistance and, since no concomitant histopathological alterations were observed in the treated mice, the evaluation of immune parameters may possibly be a sensitive indicator of toxicity.


Prolonged psychic epileptic seizures: a study of the absence status.

Long-lived epileptic seizures associated with spike-and-wave complexes are presently considered to be the absence status, i.e., the generalized nonconvulsive status. EEG radiotelemetry allowed us to record three prolonged seizures of 3 epileptic patients. Clinical manifestations included selective rather than global impairment of higher cortical functions. Clinical impairment appeared only when patients were in a state of activity and if those altered functions were used. EEG abnormalities were diffuse, but among them spike-and-wave complexes were never diffuse. It was impossible to establish close electroclinical correlation. However, the clinical and electrical evolution was roughly isomorphic, i.e., cyclic. Major clinical manifestations were associated with spikes rather than with slow waves. Lastly, patients showed common ictal psychopathological symptoms. The problem of classifying such seizures in either the generalized or partial status is discussed. The role of selective impairment of mental functions in psychopathological symptoms is also dealth with.


Chronic epileptic foci in monkeys: correlation between seizure frequency and proportion of pacemaker epileptic neurons.

A total of 1,802 neurons from 15 alert, undrugged Macaca mulatta monkeys were studied. Thirteen monkeys had chronic epilepsy induced by subpial alumina injections in precentral cortex. Precentral neurons were judged epileptic by the magnitude and variability of the percentage of interspike intervals less than 5 msec during periods when the monkeys were awake. This method of quantifying epileptic single neuron activity appears highly reliable in distinguishing epileptic neurons from precentral neurons in either normal cortex, cortex contralateral to, or within the focus. For the 13 epileptic monkeys, the relative proportion of strongly epileptic neurons found within foci was logarithmically correlated with the mean number of daily seizures. Because of the similarity between the physiology of the alumina focus in monkeys and epileptic foci in humans, these data imply that the severity of focal human epilepsy is a function of epileptic neuronal mass.


Neostigmine activated epileptiform discharge in the amygdala: electrographic-behavioral correlations.

This investigation was carried out to test the hypothesis that amygdaloid epileptiform activity is due to cholinergic hyperactivity. It was designed to study the underlying physiopathology of, and to act as an experimental model for, psychomotor epilepsy. Neostigmine was injected intracerebrally into the amygdala of the cebus monkey with chronically implanted "chemitrodes" fitted with EEG recording electrodes. The injections were made in the basal amygdaloid nucleus which normally shows very high acetylcholinesterase (AChE) enzymatic activity in histochemical preparations. Neostigmine injection resulted in very high amplitude spike activity in the amygdala only. Other brain areas, including the neighboring temporal cortex, did not show any marked EEG changes. In the first day or two, these EEG changes were associated with myoclonus localized in the ipsilateral muscles of facial expression and also associated with masticatory seizures. Subsequently the animal became aggressive and remained so several months after the injection of neostigmine. The EEG changes continued for approximately 6 weeks. Intramuscular injections of atropine diminished the amplitude of the epileptiform EEG discharges and modified slightly the animal's behavior.


Studies on the structure and I-blood-group activity of poly(glycosyl)ceramides.

Employing a modified technique of acetolysis, which allows almost a complete recovery of constituent sugars from poly(glycosyl)ceramides, the glycolipids were found to contain an excess of N-acetylglucosamine over galactose. On the basis of Smith degradation, methylation study, chromium trioxide degradation and the structures of oligosaccharides released from the glycolipids by partial acid hydrolysis, the presence of two types of sugar sequences has been established in poly(glycosyl)ceramides: a) Galbeta1 leads to 4GlcNAcbeta1 leads to 6Gal3 comes from R1 b) Galbeta1 leads to 4GlcNAcbeta1 leads to 4GlcNAc1 leads to R2. The repeating unit of poly(glycosyl)ceramides seems to be the GlcNAcbeta1 leads to 3Gal sequence. The specificity of one anti-I serum (Woj) is directed against the non-reducing ending of the first kind of chain. Three other anti-I sera reacted with inner portions of the oligosaccharide chains of the glycolipids.


Translation of MS2 RNA in vitro in the absence of initiation factor IF-3.

An Escherichia coli cell-free translational system, deprived of initiation factor IF-3, has been used to study the role of the factor in protein synthesis. In this system, 30-S ribosomal subunits are preincubated together with MS2 phage RNA in a small volume in the presence of 10 mM Mg(Ac)2; the missing components required for protein synthesis are then added and assembly of elongating ribosomes is allowed to occur. This stepwise assembly process permits formation of functional complexes which can carry out protein synthesis in the complete absence of IF-3. The translational products, obtained in the absence of IF-3, have been analysed and shown to be similar to those synthesized in the presence of the factor. The main product observed is the phage coat protein.


Plant microbody proteins. Purification and glycoprotein nature of glyoxysomal isocitrate lyase from cucumber cotyledons.

1. Isocitrate lyase from cotyledons of cucumber seedlings (Cucumis sativus) has been purified 100-fold. Two methods of preparing the soluble glyoxylate cycle enzyme are described: an elaborated method which used crude extracts of cucumber cotyledons, and another procedure which started with purified glyoxysomes from 4-day-old cotyledons and included a separation of glyoxysomal matrix enzymes by zonal centrifugation. The product behaved as a single species when tested by (a) polyacrylamide gel electrophoresis in the presence of dodecyl sulfate, (b) zonal centrifugation, and (c) double immunodiffusion against rabbit antibody to isocitrate lyase. 2. Isocitrate lyase of cucumber glyoxysomes exhibited a molecular weight of 255,000 and was composed of four apparently identical subunits of Mr 64,000. An isoelectric point of 5.9 was determined. 3. It was shown that isocitrate lyase is a glycoprotein, (a) by Schiff stain on polyacrylamide gels, (b) by periodate oxidation of the enzyme, subsequent reduction with NaB[3H]4 and electrophoretic analysis of the labelled glycoprotein, and (c) by incorporation of [3H]glucosamine in vivo into a protein which could be precipitated with antibodies to isocitrate lyase and revealed a 64,000-Mr band upon electrophoresis.


Pituitary-testicular function in patients with chronic alcoholism.

Testis and liver histology, and pituitary-testicular function were studied in thirty chronic alcoholics. Severe reduction of spermatogenesis was found in 30% and in these patients serum follicle stimulating hormone and prolactin concentrations were significantly higher. There was no correlation between abnormalities in the liver and testis. The serum testosterone was normal in most cases. Sexual dysfunction and testicular atrophy occurred in more than half of the patients and were not related to liver disease. Testicular disorder in chronic alcoholism may be independent of liver disease.


Sex steroids and sex-hormone binding globulin in males with chronic alcoholism.

Thirty men with chronic alcoholism were studied. Biopsies of the liver and testis were performed in all. Serum concentrations of total and non-protein bound (free) testosterone and oestradiol, dihydrotestosterone and sex-hormone binding globulin (SHBG) were determined. Testosterone and dihydrotestosterone concentrations were normal in most patients, whereas oestradiol and free oestradiol were above normal in approximately 50% of the patients. None of the hormones measured differed significantly between patients with and without cirrhosis. SGBG was significantly higher in men with severely reduced spermatogenesis compared to those with intact germinal epithelium, but there was no difference between men with and without cirrhosis. No relation could be demonstrated between clinical signs of hypogonadism and any of the hormones measured. The results support the view that hormonal and sexual disturbances may occur in chronic alcoholism independent of the presence of liver disease.


Influence of chenodeoxycholic acid on the kinetics of endogenous triglyceride transport in man.

Plasma lipids and triglycerides kinetics were studied in ten subjects before and after 6-8 weeks treatment with 1 g/day chenodeoxycholic acid for radiolucent gallstones. Plasma triglyceride concentration fell by 20% and phospholipid concentration rose by 5% on average; there was no change in cholesterol or free fatty acids. Body weight remained constant. Triglyceride kinetics, studied by a precursor-labelling technique, revealed a reduction both of triglyceride plasma pool and turnover rate, accompanied by a decline of more than 35% both of plasma triglyceride precursor pool and of incorporation of plasma free fatty acids into newly synthesized plasma triglycerides. Fractional turnover rates in both pools remained unaltered. The appearance-time for newly synthesized plasma triglycerides after injection of label did not change. These results indicate that chenodeoxycholic acid lowers plasma triglyceride by reducing its synthesis, thereby precluding alternative, and possibly undesired, modes of action, such as impaired secretion or increased peripheral catabolism of endogenous triglyceride. Several biochemical mechanisms may be responsible for this effect, among which stimulation of phospholipid synthesis by bile acids could play a role.


Hepatic and extrahepatic triglyceride lipase activity in uraemic patients on chronic haemodialysis.

In eleven patients with chronic renal insufficiency treated by intermittent haemodialysis and in ten normal subjects, hepatic and extrahepatic triglyceride lipase activity of post heparin plasma was selectively measured, utilizing the different sensitivity of both enzymes to inhibition by protamine sulphate. In uraemic patients, hepatic triglyceride lipase activity was significantly decreased and extrahepatic triglyceride lipase activity was normal when compared with the control group. The uraemic subjects showed a moderate hypetriglyceridaemia; their serum cholesterol level, however, was normal. The high triglyceride concentration was due to an increase of very low density lipoproteins and low density lipoproteins of the density between 1.006 and 1.019 g/ml (LDL1). The concentration of low density lipoproteins of the density between 1.019 and 1.063 g/ml (LDL2) was decreased. LDL2 were relatively rich in triglycerides when compared with LDL2 from the control group.


Interactions of calcium, magnesium and atropine on exocrine pancreatic secretion in man.

The effects of the intravenous administration of atropine or magnesium on pancreatic secretion which has been stimulated by secretin and induced hypercalcaemia have been studied in man. In the presence of secretin (0.5 CU/kg.h) the infusion of Ca2+ (0.3 mmol/kg.105 min) resulted in an increase in secretion of enzymes by 100-200%, and in that of Ca2+ and Mg2+ by 50-100% without affecting fluid and bicarbonate secretion. The additional injection of atropine (0.5 mg i.v. and 0.5 mg s.c.) were followed by a prompt fall in enzymes but not in Ca2+ and Mg2+ to the secretin-stimulated values. The additional infusion of Mg2+ (0.12 mmol/kg.45 min) to the Ca2+-infusion did not alter the secretion of enzymes, Ca2+ or Mg2+ compared with the calcium infusion alone. It is suggested that the hypercalcaemic stimulus depends on an intact innervation of the acinar cells. In these experiments the secretion of Ca2+ and Mg2+ seem to originate mainly from extracellular fluxes.


Effects of increased left ventricular work, hypoxia, or coronary ligation on hearts from rats at high altitude.

Isolated perfused hearts from rats acclimatized to Jungfraujoch (altitude 3454 m) were less resistant to the effects of high levels of heart work or mild hypoxia than hearts from litter mates reared at Geneva (340 m). However, in response to coronary artery ligation hearts from acclimatized rats were better able to maintain tissue contents of adenosine triphosphate, phosphocreatine and glycogen. A projected explanation for the increased biochemical resistance to ischaemia was a relative increase in the coronary flow rate. The data support the hypothesis that exposure to high altitude helps to protect the heart against coronary occlusion by an increased resistance developing as a result of increased sensitivity to high levels of heart work and hypoxia.


The galactose elimination capacity as a quantitative measure of liver function in acute carbon tetrachloride intoxication of rats.

In rats given rising single doses of carbon tetrachloride (CCl4) intragastrically the relation between dose and mortality, between time after injection and the quantitative liver function measured by the galactose elimination capacity (GEC), and between the dose and the GEC, was examined. The change in hepatic contents of galactose metabolites after CCl4 was measured. There was a linear relation between the dose and mortality. No rat died later than 36 h after injection. Following injection of a dose lethal to 15% of rats the GEC fell to 40% of control after 36 h and was normalized after 72 h. There was a dose dependent decrease in the GEC with rising doses given 36 h earlier up to a dose lethal to 15%. Galactose metabolites other than UDP-galactose, which was decreased, were not affected by CCl4, suggesting a general enzyme depression. The results are compatible with the concept of proportionality between the GEC and 'the functioning liver mass' and indicate that the GEC presents prognostically valuable information during acute hepatic insufficiency.


Human colonic adenylate cyclase: effects of bile acids.

Three different bile acids--deoxycholic acid, chenodeoxycholic acid and ursodeoxycholic acid--were tested for their capacity to stimulate the adenylate cyclase in human colonic mucosa. This enzyme system was found to be sensitive towards vasoactive intestinal polypeptide and prostaglandin E2. These three bile acids were ineffective in activating the human cyclase system over a wide concentration range tested. Concentrations above 1 X 10(-5) mmol/l induced a dose-dependent inhibition of basal enzyme activity. These results suggest that bile-acid induced diarrhoea is not associated with activation of the membrane-bound adenylate cyclase system at least in man.


Determination of microsomal UDP-glucuronyltransferase in needle-biopsy specimens of human liver.

Sensitive, reliable and convenient assays are described for the study of human liver microsomal UDP-glucuronyltransferase. Using 14C-1-naphthol as substrate about 2 mg of a liver biopsy specimen and for 14C-morphine about 20 mg of tissue will suffice for enzyme estimation. Lack of inhibition of 1-naphthol glucuronidation by morphine suggests that the substrates are glucuronidated by different forms of the enzyme. Enzyme levels in the native and activated state were studied in biopsies from patients grouped according to histopathological and clinical criteria. The enzyme assays may help to characterize UDP-glucuronyltransferases in human tissues and their induction by drugs and environmental chemicals, as well as their alteration in various diseases.


Selective participation of immunoglobulin V region and major histocompatibility complex products in antigen binding by T cells.

Antigen-binding inhibition studies using microscopic autoradiography were performed on T or B cell-enriched lymphocyte populations. Antibodies specific for the "framework" of immunoglobulin heavy or light chain variable domains (VH or VL), or anti-H-2 and anti-Ia antisera were used. T cell subclasses were separated with anti-Lyt antisera and complement. It was found that antigen-binding T cells of different subclasses can be inhibited selectively with only one of the two anti-V region antibodies. Antigen binding to Lyt-1+ cells was inhibited by anti-VH, while Lyt-2+,3+ cells were inhibited by anti-VL specifically. Anti-Ia antisera inhibited unprimed Lyt-1+ antigen-binding cells, whereas anti-H-2K or anti-H-2D anti-sera inhibited unprimed Lyt-2+,3+ antigen-binding cells, and both classes of immune T antigen-binding cells.


B lymphocytes contain three species of mu chains.

The murine B cell line cloned from a single cell, 38C-13, synthesizes three species of mu chains, that of cell surface membrane IgM (m-mu), that of secreted IgM (s-mu) and that of intracellular IgM (i-mu). They differ in their mobility on sodium dodecyl sulfate polyacrylamide gel electrophoresis. Sequence analysis of the different mu chains suggests that they are identical in the N-terminal as well as in their C-terminal positions. The ratio between incorporated radioactive monosaccharides to radioactive amino acids into the three different mu chains was higher in s-mu than in m-mu, but nevertheless m-mu migrated more slowly than s-mu on sodium dodecyl sulfate polyacrylamide gel electrophoresis. However, since this ratio may also be influenced by the rate of synthesis, it may not represent a real molar ratio of carbohydrate to protein. Studies with normal spleen cells clearly indicated the presence of the same three types of mu chains.


Guinea pig immunoglobulin light chain isotypes. I. Separation of kappa and lambda chains and the identification of three isotypes of the lambda chain constant homology region.

The separation of intact kappa chain and two fragments comprising lambda (lambda) chain from immunoglobulin light chain pools isolated from strain 13 guinea pigs is achieved by cyanogen bromide digestion and gel filtration before and after reductive clevage of disulfide bonds. The smaller lambda chain fragment derives from the original carboxyl-terminus of lambda chain. The partial sequence of component tryptic and thermolytic peptides of this thirty-nine residue fragment allowed its complete sequence to be deduced. Three isotypic forms of this lambda chain constant region fragment, distinguished by four residue positions showing alternative amino acids, are found expressed by guinea pigs. Each of these three isotypes is more homologous to the other two than to lambda chains of any other species. The distribution of amino acid substitutions differentiating these isotypes further supports the view that lambda chain isotypes arose in guinea pigs, and in several other species, independently by gene duplication.


Guinea pig immunoglobulin light chain isotypes. II. The preferential expression of lambda chain-bearing anti-phenyltrimethylammonium antibodies is associated with restricted variable region expression, but not with any alteration in the proportion of lambda chain constant region isotypes, relative to normal immunoglobulins.

The inbred strain 13 guinea pig expresses at least three isotypes of immunoglobulin (Ig) lambda chain. The proportion of these isotypes in normal Ig can be estimated by isolating a constant (C) region cyanogen bromide peptide from lambda chains and quantitatively analyzing its component tryptic peptides. Anti-phenyltrimethylammonium antibody, when analyzed in this way, was found to contain the same proportion of each lambda chain C region isotype as did normal Ig. The specific antibody light (L) chain pools were found to contain predominantly lambda chains, although these are the minority type in guinea pig normal Ig L chain pools. Moreover, amino-terminal, 5-pyrrolidone-2-carboxylic acid-containing peptides isolated from antibody lambda chains show no demonstrable sequence heterogeneity, while their homologs isolated from normal lambda chains are markedly heterogeneous. That a restriction in heterogeneity of variable region framework residue positions is not associated with any alteration in the proportion of C region isotypes in these antibody lambda chains, implies that translocation of all guinea pig lambda chain variable region genes to the isotypic C region genes occurs in a random way.


Phencyclidine-induced stereotyped behavior in monkeys: antagonism by pimozide.

The psychotomimetic substance, phencyclidine (PCP), was administered chronically to selected members of a primate social colony. PCP induced stereotyped behavior in all treated monkeys at each of three test doses. Furthermore, there was a significant increase in the amount of stereotypy seen during the second and third weeks of treatment compared to the first week. The dopamine receptor blocking agent pimozide antagonized PCP stereotypy. The results are discussed in terms of dopamine mediation of these responses.


Histamine-induced release of thyrotropin releasing hormone from hypothalamic slices.

In vitro release of TRH from mediobasal hypothalamic (MBH) slices was increased in the presence of 56 mM K+ in a Ca2+-dependent manner. Addition of histamine (HA) induced a concentration-dependent release of TRH; concentrations of 3 x 10(-7) M and 10(-6) M elicited maximal and maximal stimulation respectively. Cimetidine 10(-6) M, a potent antagonist of HA H2-receptors, inhibited the effect of HA. The present results suggest that HA can trigger the release of TRH from neurosecretory nerve endings and that MBH slices are a useful preparation for studying neurohormone release.


A study of the changes in motor behaviour caused by TRH on intracerebral injection.

20 microgram TRH injected bilaterally into the caudate-putamen, tuberculum olfactorium, nucleus accumbens, amygdala, lateral ventricles, midbrain or cerebral cortex failed to induce any increase in locomotor activity (measured using photocells), although other behavioural changes were observed after each injection, and included body shakes, limb tremor, repetitive head and limb movements, biting, scratching and an alert appearance. These behavioural changes could result in positive readings from equipment used to measure locomotor activity, but careful investigations focussing on the nucleus accumbens used photocell boxes, activity wheels and Animex recorders to emphasise the inability of intracerebral TRH (10--40 microgram) to enhance locomotor activity. Intraaccumbens TRH also failed to enhance amphetamine hyperactivity or reduce the motor depression caused by haloperidol and analeptic drugs. The data do not support a central locomotor stimulant action of TRH.


Anterior pituitary adenylate cyclase: stimulation by dopamine and other monoamines.

Adenylate cyclase activity of homogenates of anterior pituitary from rat, rabbit and monkey was stimulated by dopamine, other amine neurotransmitters and Gpp(NH)p. The concentration of dopamine for half-maximal stimulation of activity was 2--5 micrometer. Studies with selective agonists and antagonists indicated the presence of dopamine receptors and beta-adrenergic receptors coupled to adenylate cyclase in anterior pituitary. In addition, histamine stimulated activity in rabbit and monkey, but not in rat, anterior pituitary.


Characteristics of interhemispheric impulse conduction between prelunate gyri of the rhesus monkey.

Cells of origin of the corpus callosum (callosal efferent neurons) in prelunate gyrus (area OA) of the rhesus monkey were studied using electrophysiological techniques. Monkeys were chronically prepared and callosal efferent neurons were identified by their antidromic activation following electrical stimulation of the contralateral prelunate gyrus and/or the splenium of the corpus callosum. Interhemispheric antidromic latencies ranged from 2.6--18.0 ms (median = 7.0 ms) while the conduction velocity along the length of the axon ranged from 2.8 to 22.5 M/s (median = 7.4 M/s) while the conduction velocity along the length of the axon ranged from 2.8 to 22.5 M/s (median = 7.4 M/s. Following the relative refractory period of a single prior impulse, all but one of 61 callosal efferent neurons studied showed a supernormal period of increased axonal conduction velocity and excitability. Following several prior impulses, the supernormal period was followed by a subnormal period of decreased axonal conduction velocity and excitability, which, depending on the number of prior impulses, lasted from several hundred ms to nearly 2 min.


The retino-geniculo-cortical pathway in Callithrix. I. Intraspecific variations in the lamination pattern of the lateral geniculate nucleus.

Injections of tritiated tracer precursors into one eye revealed a highly variable lamination pattern of the LGN in the marmoset Callithrix. In one specimen, only four cell layers were apparent with some indication of further differentiation. In a second specimen, six almost complete relay layers were found in the occipital portion of the LGN. Three other animals showed stages between these two extremes. The lamination pattern of Callithrix thun represents an intermediate stage between a four-layered LGN suggested as the basic primate pattern, and the advanced six-layered LGN of most other anthropoid monkeys. In addition, bilateral retinal terminations were found consistently in layer 0, and contralateral terminations in a narrow band between the magnocellular and parvocellular portions of the nucleus, and within the interlaminar zone which separates the two magnocellular layers.


Spinal branching of pyramidal tract neurons in the monkey.

The branching pattern of individual pyramidal tract (PT) neurons of the monkey motor cortex was studied by activating these neurons antidromically from within the cervical motor nuclei and also from other regions of the spinal cord. 1. Fifty-four neurons were activated from motor nuclei in the cervical cord. Twenty-eight of these were activated from one segment and six (11%) were activated from motor nuclei of different segments. The remaining 20 neurons were activated from motor nuclei and also from unspecified region(s) of the gray matter. 2. Another 156 neurons were activated from unspecified regions(s) of cervical gray matter which could have been motor nuclei or outside the nuclei, and 64 of these were activated from more than one segment. 3. The branching patterns of PT neurons sending axons directly to motor nuclei innervating distal forelimb muscles suggested that they branch less than the rest of PT neurons.


Projection of individual pyramidal tract neurons to lumbar motor nuclei of the monkey.

The projection of individual pyramidal tract (PT) neurons from the hindlimb area in the precentral gyrus of the cerebral cortex to the lumbar spinal cord was studied in the monkey by systematically searching for sites within identified regions of the spinal gray from which the PT neurons could be antidromically activated by local stimulation. All investigated neurons belonged to the fast conducting fraction of PT neurons. The following results were obtained. 1. Each PT neuron could be activated from more than one region of the spinal gray matter, including identified spinal motor nuclei and areas dorsomedial to these nuclei, but do not the intermediate nucleus or regions dorsal to it. "Passage areas" and "termination areas" were defined. 2. Half of the PT neurons with termination areas within motor nuclei had these areas in more than one nucleus. There were thus strong suggestions for synaptic contacts of some PT neurons with motoneurons of more than one muscle. 3. Four groups of three or four neurons were recorded simultaneously by the same cortical electrode. Comparisons of passage and termination areas within groups revealed both similarities and differences in projections of neighboring neurons. Every neuron was activated from some region(s) where others of the group were not. Common passage areas, or passage and termination areas, for two or three neurons of a group within at least one motor nucleus were found for all groups. Termination areas in the same motor nucleus have been found for the majority of the neurons of only one group. These common projection areas are compatible with, but not prove, that group of adjacent PT neurons has common target cells in the spinal cord.


Compartmentalization of prostaglandins and prostacyclin within the kidney: implications for renal function.

When renal function is compromised, the circulation to the kidney is sustained by a major prostaglandin component, withdrawal of which results in significant hemodynamic effects, particularly reduction in blood flow to the inner cortex and medulla. Prostaglandins modulate the effects of vasoactive hormones by attenuating the renal actions of the renin-angiotensin system and contributing to and, perhaps, mediating some of those of the kallikreinkinin system. In addition, a prostaglandin mechanism, presumably located in the renal arterioles, participates in the regulation of renin release. Although cyclooxygenase is present in several renal tissues, the major products of arachidonic acid metabolism may be tissue specific and, consequently, their effects may be primarily restricted to one compartment, e.g., the proposed interaction of prostacyclin and renin within the vascular pole of the glomerulus; and PGE2/PGF2a with the kallikrein-kinin system within the urinary compartment. The former is related to the regulation of renin release and renal vascular resistance and the latter to the excretion of water and perhaps salt.


Calcium as a second messenger in the stimulation of luteinizing hormone secretion.

In cells dissociated from porcine anterior pituitary glands and maintained in culture for 48 h the specific secretagogue luteinizing hormone-releasing hormone (LH-RH) induces a biphasic pattern of luteinizing hormone (LH) release. A biphasic pattern of release is also induced by 57 X 10(-3) M K+ and the ionophore A-23187. By reducing the availability of Ca2+, either by omission from the medium, chelation or interfering with Ca2+ transport across the plasma membrane, it is shown that LH release stimulated by LH-RH is much less dependent upon the availability of extracellular Ca2+ than that stimulated by either high K+ or A-23187. Nevertheless, by using a lanthanum displacement protocol to follow the influx of 45Ca2+ it is shown that LH-RH stimulation does induce an influx of extracellular Ca2+. Parallel experiments in which the stimulated 45Ca2+ efflux from preloaded cells is followed confirm the influx data but suggest, in addition, that when the influx of extracellular Ca2+ is inhibited, the peptide is able to mobilize Ca2+ from an intracellular location. It is thus concluded that while LH release can be initiated by an increase in the intracellular level of Ca2+, and although LH-RH stimulation does increase the permeability of the plasma membrane to Ca2+, the stimulation of LH release by LH-RH is not dependent upon extracellular Ca2+.


A new look at the assessment of mental development in young children recovering from severe malnutrition.

Previous studies of malnourished children in hospital have shown that their developmental quotients (DQS) improve as their nutritional status improves, but they have not taken into account the possible effects of being in hospital and of test practice. The present study assessed development of malnourished and adequately nourished children in hospital and found that mean DQS of each group rose to a similar extent during recovery from illness. It is concluded that DQS on admission to hospital were lowered by the children's discomfort and unhappiness at being in hospital, and/or by illness, and that rises in DQ shown by children recovering from malnutrition are not necessarily attributable to improving nutritional status.


Cerebrovascular changes in neurofibromatosis.

Vascular changes in neurofibromatosis are most commonly described in the renal arteries. In the present study, two children with neurofibromatosis and cerebral vascular occlusive changes demonstrated by cerebral angiography are reported. Although focal neurological findings in children with neurofibromatosis are often due to tumours, the sudden development of neurological symptoms in such cases should alert paediatricians to the possibility of cerebral vascular disease.


Alrestatin: gastric acid antisecretory-antiulcer activity in the rat.

Alrestatin sodium (AY-22,284-A) inhibited gastric acid secretion and decreased the volume of gastric juice produced in the pylorus-ligated rat when administered intraperitoneally or perorally. Pentagastrin-induced gastric acid output in the unanesthetized rat was antagonized. The pyloric ligation-induced ulcer formation in the rat was inhibited. Alrestatin sodium did not exhibit an anticholinergic profile. The drug did not block the norepinephrine neuronal uptake mechanism in rat brain or heart; it did not alter the endogenous norepinephrine concentration in the heart and decreased endogeneous brain norepinephrine concentration. Alrestatin sodium is an effective inhibitor of gastric acid secretion and ulcer formation in the rat.


Effect of starvation on small intestinal enzyme activity in germ-free rats.

Starvation overnight and starvation for 48 h reduced the weight and the protein content of mucosal scrapings, but only minimally reduced the DNA content of the mucosal scrapings. The activity of sucrase and maltase was reduced by both periods of starvation. The activity of lactase and of acid and alkaline phosphatase, however, was less subject to starvation. There were striking differences in the response to starvation between the proximal, mid and distal third of the small intestine. The importance of the proper reference system was discussed.


Plasma arginine vasopressin in diabetic ketoacidosis.

Plasma Arginine Vasopressin (AVP) was measured serially in 10 patients during the first 24 hours of treatment of diabetic ketoacidosis. AVP was elevated in all cases initially, ranging from 4.0--122.0 pmol/l, the basal level in healthy hydrated subjects being 1.57 +/- 0.59 pmol/l (+/- 1 SD). The levels fell progressively during the course of treatment. While there was no evidence that the massive increases in AVP noted in this study were associated with water overload it remains to be determined whether the high levels observed have any significant metabolic or haemodynamic role in this disorder.


Atypical esophageal diverticula associated with progressive systemic sclerosis.

Five cases of unusual esophageal diverticula associated with progressive systemic sclerosis (scleroderma) or collagen vascular disease are presented. These wide-mouthed saccular diverticula were infrequently seen in a group of PSS patients with the typical motility disturbance of esophageal involvement and are reminiscent of the diverticula of the colon involved by PSS.


Modern operative fluorocholangiography: utopia or overlooked entity?

A system of image amplification with television fluoroscopy and 100 X 100 mm cut film technique is presented for use in the operating room. Biliary surgery has benefited from this equipment as operative cholangiography has been improved and unnecessary extension of operating time avoided. Applications of the equipment for other procedures are discussed.


Association of congenital cystic dilatation of the common bile duct and congenital diverticulum of the hepatic duct with concomitant ascariasis.

A case with association of congenital cystic dilatation of the common bile duct and congenital diverticulum of the hepatic duct is reported. The etiology and classification are briefly discussed and a radiological approach is recommended.


Effect or oral administration of propylene glycol on the induction of enzymes and proteins in microsomes and cytosol of the rat liver.

Male Wistar rats were divided into 4 groups, receiving water, propylene glycol (PG), sodium phenobarbital (PB) and PB + PG, respectively. After oral administration of these materials for 7 days, various enzymes in hepatic microsomes and the binding capacity of Y and Z fractions in the cytosol were assayed. As compared with the water group, the aniline hydroxylase activity, the cytochrome b5 content and the amount of Z fraction increased in the PG group. As compared with the PB group, the microsomal protein increased but the activities of paranitrophenol-UDP-glucuronyltransferase (pNP-GT) and aminopyrine demethylase (AD) decreased in the PB + PG group. When the PB + PG group and the PG group were compared with the PB group and the water group respectively, there was no parallel change. Even though PG added to hepatic microsomes of untreated rats in vitro, there was no change in the difference spectrum produced by binding of cytochrome p-450 and aniline and also in the activities of AH and pNP-GT. Based on these observations, it can be concluded that PG affected the in vitro assay data of drug-metabolizing enzyme without changing the cytochrome P-450 content by its certain in vivo action rather than by its direct interaction with microsomes, and modified the effect of PB on the microsomal enzymes and the cytosol proteins. Therefore, it is not appropriate to investigate the effect of the drugs on enzymes and proteins of the liver cell by the use of PG as a solvent.


A receptor for polymerized human and chimpanzee albumins on hepatitis B virus particles co-occurring with HBeAg.

A receptor for polymerized human serum albumin was demonstrated on Dane particles as well as on 20-nm hepatitis B surface antigen particles, isolated from asymptomatic carriers of hepatitis B virus who were positive for HBeAg. In contrast, such receptor was not born by 20-nm hepatitis B surface antigen particles obtained from carriers positive for antibody to HBeAg. Hepatitis B surface antigen particles with the receptor were heavier than those without, and when treated with pronase, they became lighter and lost the receptor. The receptor is responsible for the agglutination of erythrocytes coated with polymerized human serum albumin by the serum of patients with Type B hepatitis and asymptomatic carriers, which have been attributed to autoantibodies directed to denatured albumin molecules. When albumin fractions of chimpanzees were polymerized with glutaraldehyde, they also bound with the receptor on hepatitis B surface antigen. Polymerized albumin fractions of all the other experimental animals without susceptibility to hepatitis B virus, however, failed to bind with the receptor. These results seem to suggest a possible role of the receptor on Dane particles (presently accepted hepatitis B virions) for polymerized albumin molecules in infecting hepatocytes both in humans and chimpanzees.


Chronic persistent hepatitis and unconjugated hyperbilirubinemia.

The authors studied 12 patients with chronic persistent hepatitis and persistent or intermittent mild unconjugated hyperbilirubinemia. Maximum serum total bilirubin concentration ranged from 2.1 to 3.6 mg/dl. Hemolysis was not evident. Hepatic bilirubin UDP-glucuronyltransferase activity assayed in each patient ranged from 0.16 to 0.39 U (mean +/- SEM = 0.27 +/- 0.02) compared to 0.68-1.99 (1.35 +/- 0.08) in 23 normals, 0.78-2.28 (1.41 +/- 0.05) in 53 patients with acute hepatitis, 0.34-1.74 (0.81 +/- 0.09) in 16 patients with anicteric chronic persistent hepatitis, and 0-0.62 (0.24 +/- 0.03) in 33 patients with Gilbert's syndrome. The mean UDP-glucuronyltransferase activity was significantly lower in anicteric chronic persistent hepatitis compared to normals, but higher than in Gilbert's syndrome. The incidence of unconjugated hyperbilirubinemia among first degree relatives was 0:32 in icteric chronic persistent hepatitis compared to 24:85 (28%) in Gilbert's syndrome. These results show that the likely cause for the unconjugated hyperbilirubinemia associated with chronic persistent hepatitis is an acquired depression of hepatic bilirubin UDP-glucuronyltransferase activity. The data suggest that the enzyme defect is related to chronic persistent hepatitis.


Detection of carcinogen-induced stimulation of cytochrome P-448-associated enzymes by 14CO2 breath analysis studies using dimethylaminoazobenzene.

Rats pretreated with three agents known to stimulate cytochrome P-448-associated enzymes, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 3-methylcholanthrene (3MC), and Aroclor 1254 were studied with a 14CO2 breath analysis technique after administration of [14C-dimethyl]aminoazobenzene (DMAB). The half-life of breath 14CO2 after [14C]DMAB administration was significantly decreased in the TCDD-, 3MC-, and Aroclor 1254-treated rats compared with controls. In vitro studies indicated that DMAB N-demethylase was increased by these three agents. Phenobarbital, an inducer of cytochrome P-450-associated enzymes, had no effect on 14CO2 half-life or on DMAB N-demethylase. Studies with [14C]aminopyrine, a cytochrome P-450 substrate, showed that TCDD and 3MC had no effect on 14CO2 half-life or aminopyrine N-demethylase; Aroclor 1254 and phenobarbital decreased 14CO2 half-life and stimulated aminopyrine N-demethylase. The data suggest that a [14C]DMAB breath analysis technique may be useful for in vivo studies of inducers of cytochrome P-448.


Dynamics of correlated genetic systems. IV. Multilocus effects of ethanol stress environments.

Four replicate populations of Drosophila melanogaster, two reared on medium supplemented with ethanol and two reared on standard medium, were electrophoretically monitored for 28 generations. During the first 12 generations, allelic, genotypic and gametic frequencies were determined for eight polymorphic enzymes: GOT, alpha-GPDH, MDH, ADH, TO, E6, Ec and ODH. Samples from generation 18 and 28 were electrophoretically typed for ADH and alpha-GPDH. In addition, samples from generation 27 were analyzed for the presence of inversion heterozygotes. The experimental results showed rapid gene-frequency divergence between control and treatment populations at the Adh locus in a direction consistent with the activity hierarchy of Adh genotypes. Gene-frequency divergence between control and treatment populations also occurred at the alpha-Gpdh locus, although the agreement among replicates appeared to have broken down by generation 28. No differential gene-frequency change occurred at any of the six remaining marker loci. Furthermore, values of linkage disequilibria among all linked pairs of genes were initially small and remained small throughout the course of the experiment. Taking these facts into account, it is argued that the gene-frequency response observed at ADH is most probably caused by selection at the Adh locus. The gene frequency response at alpha-Gpdh can also be be accounted for in terms of the effect of ethanol on energy metabolism, although other explanations cannot be excluded.


Rabbit variable kappa light chain regions: subgroups contain polypeptides encoded by multiple genes.

Two identical light chain variable regions were identified in anti-streptococcal Group A-variant antibodies elicited in litter-mate rabbits by hyperimmunization with vaccine. In addition, one rabbit produced two additional clonally restricted antibodies to this polysaccharide antigen. The partial amino acid sequence of the light chain of one of these antibodies was identical with the dominant antibody light chain sequence, while the light chain of the other antibody, also partially established, showed significant variations in the framework-associated regions with identical CDRI and II. Since all of these light chains were from a small subset of rabbit kappa light chain pools (b4 allotype) the data suggest, together with other light chains reported in the literature, that more than one copy of variable region genes are present in the germ-line per subgroup. Furthermore, framework associated amino acid substitutions are not random; this suggests the existence of some "ordered" mechanism for linked amino acid substitutions (presumably recombination). Furthermore one light chain can pair with more than one heavy chain to yield functional antibodies.


The complete amino acid sequence of both subunits of C-phycocyanin from the cyanobacterium Mastigocladus laminosus.

The amino acid sequences of both subunits of the C-phycocyanin from the thermophilic cyanobacterium Mastigocladus laminosus have been determined. The alpha-chain consists of 162 amino acid residues and has a molecular weight of 18000, whereas the beta-chain consists of 172 residues and has a molecular weight of 19400. For the first three quarters of their length the polypeptide chains are 31% homologous, whereas there is no significant homology in the final quarter up to the C-terminus. This could mean that the introduction of an additional chromophore binding site in the last quarter of the beta-chain during evolution was achieved via a large number of point mutations or by exchange of the whole C-terminal part in an ancestral gene.


A hospital must study cost, codes, alternatives to decide on reuse or new building.

To cope with the impact of codes, design, and attitudes in a renovation project, a hospital may have to make compromises and consider many alternatives to its plan.


Medical center applies financial strategies to renovation project.

A Chicago-based medical center abandons the classical approach to preconstruction planning to save both time and money.


Could two-level process eliminate costly planning problems?

Lack of concurrent review and incompatibility with cost containment objectives are serious problems with the present certificate-of-need approval process.


To reuse or reconstruct: that is the question.

With cost containment being a vital concern, health care facilities must carefully assess differences  in the cost between new construction and renovation.


Prenatal diagnosis of GM1-gangliosidosis: biochemical manifestations in fetal tissues.

A prenatal diagnosis of GM1-gangliosidosis was made in a pregnancy at risk, on the basis of a deficiency of beta-galactosidase activity demonstrated in cultured amniotic fluid cells. Biochemical analyses were performed in the aborted fetus. GM1-ganglioside beta-galactosidase activity was reduced to 1% of the control value in both the brain and liver of the affected fetus. Lamellar bodies suggestive of membranous cytoplasmic bodies were found in cells of basal ganglions, while the accumulation of GM1-ganglioside in the brain was not remarkable.


[Routine test for sensitivity (author's transl)].

The agar diffusion test is a simple but in most instances reliable method for the determination of the sensitivity of pathogenic bacteria, if it is performed under controlled standardised conditions and evaluated by defined criteria. The accuracy of the test results can be achieved by a permanent control of the test with control strains during routine testing.


[Relationship between MIC and Zone Size (author's transl)].

The one antibiotic-one disk agar diffusion test for susceptibility testing of clinical isolates, though technically similar to that used for determining antibiotic concentrations in body fluids, lacks the exactness of the latter. In correlating zone diameters to MICs, stochastic linearity should be evaluated besides the regression lines and the correlation coefficient.


[A method for testing susceptibility of Haemophilus influenzae to co-trimoxazole (author's transl)].

Various methods for determining the activity of co-trimoxazole against haemophilus influenzae have been compared. The results were cortically influenced by antagonistic substances in the medium. The content of p-aminobenzoeacid and thymidine in the medium should be as low as possible. The antagonistic influences are compensated by lysed horseblood, which contains thymidine-phosphorylase. Haeminchloride and nicotinamide adenine dinucleotide (NAD) are better growth-factors than other poorly defined preparations (Supplement C, Fil-des-Enrichment, Iso-VitaleX). For the determination of the minimal inhibition concentration (MIC) we propose the microtiter technique. Likewise a modified agar diffusion test can be recommended for screening purposes. Of the 143 haemophilus influenzae strains tested against co-trimoxazole 134 (94%) were sensitive.


[Susceptibility testing with the autobac system (author's transl)].

The Autobac I provides an automated method for the rapid measurement of antimicrobial susceptibility of fast growing pathogenic microorganisms. To check instrument accuracy, 477 cultures were tested by the Bauer-Kirby method for comparison. According to the DIN limits, the results were comparable in 81.8%, whereas within the NCCLS limits the agreement was 86.5%. The discrepancy was found to be due to incorrect disc mass and different limits of inhibitory zone diameters. Another reason for disagreement seemed to be the incubation time, since its influence on the results varied when different bacterium/antibiotic combinations were used.


Fracture of an ossified Achilles tendon.

A case of rupture of an ossified Achilles tendon is described. It is suggested that the management of this condition is surgical. Previously undescribed histological appearances demonstrate osteogenic potential. The significance of subcutaneous Achilles tenotomy is discussed.


Immunological method to differentiate between antigens of tubercle bacilli, other mycobacterial species, and non-acid-fast bacteria.

Sera from rabbits immunized with sonicates of Mycobacterium bovis BCG were passed through an immunoadsorbent made of a soluble BCG extract to make partially purified antibodies to BCG. These antibodies were in turn used to prepare an immunoadsorbent through which the BCG extract was passed. The partially purified antigenic material was radiolabeled and subjected to electrophoresis in acrylamide gels. One of the radiolabeled fractions isolated (BCG-C) was found to bind to antibodies to BCG and H37Rv, but not to antibodies in sera from rabbits immunized with other mycobacterial species or Nocardia asteroides. The reaction between BCG-C and the partially purified antibodies to BCG was inhibited by small amounts of different BCG antigens. Cultures obtained from 25 patients with tuberculous diseases, other bacterial cultures, and various bacterial extracts were tested for their capacity to inhibit this reaction. Each of 13 mycobacteria identified as M. tuberculosis inhibited this reaction. Equivalent numbers of 12 strains of mycobacteria other than M. tuberculosis and high concentrations of other bacterial extracts did not inhibit, indicating that determinants of BCG present in M. tuberculosis were not detected in the other mycobacteria or in non-acid-fast bacteria. The use of sequential purification procedures could be of potential clinical value in quickly differentiating between M. tuberculosis and a variety of other mycobacteria.


Suppression of humoral responses during Trypanosoma cruzi infections in mice.

C57BL/6 mice exhibit low parasitemias and often survive Trypanosoma cruzi infections, whereas C3H(He) mice die during the acute phase with relatively high parasitemias. The present study showed that both strains of mice develop nonspecific immunosuppression to challenge with sheep erythrocytes during the course of infection. Several major differences in immunosuppression-related phenomena between the two strains of mice were determined, yet there is no apparent relationship between immunosuppression and resistance to T. cruzi. Both the number of direct plaque-forming cells and the titer of 2-mercaptoethanol-sensitive agglutinating antibody were significantly lower on day 11 for C57BL/6 mice and day 9 for C3H(He) mice. The number of indirect plaque-forming cells and the titer of mercaptoethanol-resistant agglutinating antibody were reduced by day 36 of infection in C57BL/6 mice and 13 days postinfection in C3H(He) mice. In both strains the degree of humoral response suppression of mice increased concomitant with the period of infection, but was not correlated with the changes in spleen cell numbers. Preliminary experiments designed to explore the mechanism underlying the induction and maintenance of immunosuppression in this hostparasite model disclosed the presence of suppressor substance in the serum of T. cruzi-infected mice. The passive transfer of serum from infected mice to syngeneic recipients elicited a state of immunosuppression to sheep erythrocytes, but did not diminish anti-erythrocyte activity in allogeneic recipients. The induction of immunosuppression in normal mice was further found to be dependent on the interval between serum transfer and challenge with antigen. No quantitative differences existed between the magnitude of suppressed humoral responses in mice infected for varying lengths of time and recipients of serum collected from similarly infected mice.


K-1 antigen of Escherichia coli: epidemiology and serum sensitivity of pathogenic strains.

K-1 Escherichia coli are far more frequent in neonatal sepsis (36% of E. coli sepsis) and meningitis (80% of E. coli meningitis) than would be expected by the frequency of K-1 E. coli colonization in neonates (11 to 25%). There is no apparent parallel in cases of sepsis in adults. To study further this apparent age-related difference in virulence, E. coli K-1 clinical isolates were tested for their sensitivity to sera. Strains isolated from cases of neonatal meningitis were more sensitive to serum bactericidal activity than those from cases of neonatal or adult sepsis or adult meningitis (P < 0.01). Serum sensitivity did not appear to be determined by K or O antigens. Four isolates sensitive to serum bactericidal activity obtained from neonatal cerebrospinal fluid were killed by adult serum chelated with 0.05 M Mg(2+) ethyleneglycol-bis (beta-aminoethyl ether)-N,N-tetraacetic acid (EGTA), suggesting that the alternative pathway was activated. Although untreated neonatal sera killed these strains as well as adult sera did, EGTA-treated neonatal sera were less effective than EGTA-treated adult sera. This suggests that the alternative pathway function was not activated in neonatal sera. The bactericidal defect of neonatal EGTA-treated serum was partially corrected by addition of either A or B hyperimmune equine meningococcal antiserum.


Nonhuman primate model for the study of respiratory Klebsiella pneumoniae infection.

Squirrel monkeys were inoculated by the intratracheal inoculation of 700 Klebsiella pneumoniae organisms and developed lobar pneumonia in about 24 h. Characteristic clinical findings were fever, anorexia, and coughing. Laboratory findings included leukocytosis or leukopenia (with the latter more prominent in ultimately fatal infections), bacteremia, and shedding of bacteria into the pharynx. Infected monkeys showed increased plasma lysozyme activity as well as increased plasma ceruloplasmin, haptoglobin and alpha1-antitrypsin. The mortality rate was 60%, and the mean time of death was 50.5 h. Pathologically, the disease spread by means of Kohn's pores and other pathways that generally did not involve airways as a means of dissemination until about 30 h. Squirrel monkeys seem to be better models for human respiratory K. pneumoniae infection than rats or mice.


Effects of temperature on the stability of Rickettsia tsutsugamushi and gamma-irradiated scrub typhus immunogens.

Unirradiated Rickettsia tsutsugamushi and a component of gamma-irradiated Karp immunogen required for homologous immunity were more stable than the immunogen component that elicited heterologous (Kato strain) protection.


Gamma-irradiated scrub typhus immunogens: development and duration of immunity.

The development and duration of immunity to lethal scrub typhus infection was studied in BALB/c mice vaccinated with gamma-irradiated Rickettsia tsutsugamushi, strain Karp. One intraperitoneal injection containing approximately 10(8) 50% mouse lethal doses (MLD(50)) of irradiated organisms elicited an immune response protective against challenge with 10(5) MLD(50) of viable Karp. The same mass of immunogen given in three injections at 5-day intervals increased homologous (Karp strain) protection 25-fold and heterologous (Kato strain) protection 60-fold. Further temporal expansion of the immunization regimen did not increase protection. Subcutaneous vaccination provided significant, but lower, levels of protection than were achieved by intraperitoneal immunization, but the levels of cell-transferable immunity elicited by the two routes were approximately the same. Immunologically specific protection after intraperitoneal vaccination developed rapidly enough to provide resistance against simultaneous challenge with 200 MLD(50) of Karp. Homologous immunity was protective against a 10(6)-MLD(50) challenge 7 days after completion of the three-injection regimen, remained at that level for 3 months, dropped to 10(4) MLD(50) by 9 months, and was effective against a 50-MLD(50) Karp challenge at 12 months. Protection against heterologous challenge was first observed on day 17 and peaked on day 38, when the mice resisted a 10(5)-MLD(50) Kato challenge. Thereafter, heterologous protection waned rapidly and was not significant at 6 months.


Studies on gonococcus infection. XVI. Purification of Neisseria gonorrhoeae immunoglobulin A1 protease.

A protease which cleaves human immunoglobulin A1 (IgA1) has been purified from broth cultures of Neisseria gonorrhoeae. This IgA1 protease is produced by pilated and nonpilated gonococci throughout their growth cycles. A combination of ammonium sulfate precipitation, column chromatography, and either isoelectric focusing or affinity chromatography was utilized to obtain an enzyme preparation that showed approximately 3,800-fold purification and exhibited two bands (65,000 and 70,000 daltons) by analytical polyacrylamide electrophoresis in the presence of sodium dodecyl sulfate and reducing conditions. IgA1 protease activity is dependent on divalent cations and is heat labile. Detection and quantitation of IgA protease activity utilized an assay in which [125I]IgA1 is incubated with protease preparations and the cleavage products are analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.


Gonococcal color and opacity variants: virulence for chicken embryos.

Previous studies have noted that the prevalence of certain gonococcal colony types is influenced by the physiological state of the human host. Consequently eight different gonococcal strains were passed on clear typing medium, and opaque (Op) and transparent (Tr) variants of heavily pilated (P++) bacteria were selected. P++Op and P++Tr were injected into 11-day-old chicken embryos, and the resulting mortality was determined. In every case the Tr variants were more lethal to the embryos than were the Op variants (P = 0.01). P++Tr were stable in vivo, but chicken embryos injected with P++Op strains developed bacteremia with P++Tr organisms. However, chicken blood was not bactericidal for either colonial variant. These results indicate that transparent pilated bacteria may be more virulent than opaque pilated bacteria and that in vivo selection of transparent forms occurs.


Brucella antigen preparations for in vitro lymphocyte immunostimulation assays in bovine brucellosis.

Three Brucella antigen preparations, Brucella abortus soluble antigen, B. abortus strain 45/20 enriched protein antigen, and B. melitensis enriched protein antigen, were compared in terms of their ability to induce specific in vitro lymphocyte immunostimulation responses. Lymphocytes were prepared from peripheral blood of cattle with different exposure experiences to B. abortus organisms. Lymphocytes were processed by the Ficoll-diatrizoate technique, and results were assayed for [3H]tymidine incorporation into DNA by liquid scintillation spectrometry. The three Brucella antigen preparations were compared both at the optimal concentrations of protein and on an equal-dry-weight basis. The results were evaluated in terms of specific lymphocyte immunostimulation responses induced by each preparation and the degree of correlation with infection. B. abortus soluble antigen-induced lymphocyte immunostimulation response correlated best with infection status followed by B. abortus 45/20 and B. melitensis enriched protein antigens. The implications of these findings are discussed and a hypothesis is proposed.


Antibody-dependent cell-mediated cytotoxicity in cattle: activity against 51Cr-labeled chicken erythrocytes coated with protozoal antigens.

Bovine mononuclear cells in the presence of bovine anti-chicken erythrocyte sera at high dilutions induce release of chromium-51 from labeled chicken erythrocytes. Bovine effector cells are capable of recognizing both bovine immunoglobulin G(1) and bovine immunoglobulin G(2); in contrast, human effector cells only recognize immunoglobulin G(1). Effector cell activity of bovine mononuclear cells is equally distributed between peripheral blood and spleen. As in other species, thymus and lymph node cells exert no antibody-dependent effect, although some direct cytotoxicity by lymph node cells may be observed. Antibody-dependent cell-mediated cytotoxicity against a bovine cell line can also be detected. By using a tannic acid technique, it was found that chicken erythrocytes coated with Theileria parva piroplasm antigen or with Trypanosoma rhodesiense variant-specific coat antigen form suitable targets for bovine antibody-dependent cell-mediated cytotoxicity assays. By using such targets, a moderate degree of direct cytotoxicity by bovine mononuclear cells, in the absence of antibody, is always observed; this may be reduced by choosing optimal conditions of tannic acid treatment and antigen sensitization and by the use of short incubation periods for the cytotoxicity assay. Observations have been made on the variant specificity, time course of appearance, and association with immunoglobulin G(1) of the antibody activity responsible for cell-dependent cytotoxicity against chicken erythrocytes coated with T. rhodesiense antigens. The potential usefulness of this technique in the analysis of protective immune responses against protozoal infections is discussed.


Experimental urinary tract infection with Pseudomonas aeruginosa in mice.

Urinary tract infection with Pseudomonas aeruginosa was induced in mice by transurethral inoculation of the organism into the bladder, followed by urethral obstruction for 6 h. The infection was mostly localized in the urinary organs. P. aeruginosa P9 was selected as the challenge organism from 10 laboratory strains of P. aeruginosa. After the inoculation of 10(7) colony-forming units of P. aeruginosa P9, transient bacteremia was observed in some of the mice from 6 h to 1 day after the inoculation. The number of organisms in the bladder tissue gradually decreased, whereas that in the kidneys increased to levels of 10(6) to 10(7) colony-forming units in 3 days, and these levels remained up to 2 weeks after the inoculation. The organisms gradually disappeared thereafter, and spontaneous recovery took place. The organisms could be recovered from the kidneys of 95% of the mice, and the gross lesions in the kidneys were observed in 77% of the mice 1 week after inoculation. The method developed here is simple and may be useful in the study of urinary tract infections due to P. aeruginosa and other species of bacteria.20


R-factor inheritance and plasmid content in mucoid Pseudomonas aeruginosa.

Eighteen strains of alginate-producing mucoid Pseudomonas aeruginosa were evaluated with respect to plasmid content and the ability to maintain well-characterized R plasmids. The spontaneous loss of alginate production in these strains varied from 0.01 to 0.7% and was not significantly increased by plasmid curing regimens. Examination of cleared lysates of these strains and their isogenic nonmucoid derivatives by agarose gel electrophoresis failed to reveal plasmid DNA. R-plasmid (P-incompatibility-group) transfer to mucoid P. aeruginosa was unaffected by the presence of the alginate capsule. Maintenance and expression of such plasmids in the mucoid strains were confirmed by agarose gel electrophoresis and by verification of plasmid-linked drug resistance and pilus-specific bacteriophage sensitivity. These studies demonstrate that alginate production does not appear to be plasmid linked and that mucoid P. aeruginosa are capable of receiving and donating certain drug resistance plasmids. Since some of the plasmids used here have been shown to mobilize chromosomal DNA, strains constructed in this study should afford the means for exploring the genetic basis of the mucoid phenotype.


Contact sensitivity responses in mice infected with Trypanosoma cruzi.

Mechanisms of depression of contact sensitivity responses in C57BL/10 mice infected with Trypanosoma cruzi were studied. Cellular involvement during sensitization with oxazolone was investigated in mice acutely infected with T. cruzi. Contact sensitivity was not expressed in mice during the latter stages of the acute infection. Spleen cells from sensitized, infected mice which were unable to respond to oxazolone could confer contact sensitivity upon normal syngenic mice as effectively as spleen cells from uninfected, sensitized donors. The ability of mice infected with T. cruzi to respond to an eliciting dose of oxazolone was significantly improved when macrophages from normal syngenic donors were administered to them at the time of skin test. When either normal or infected mice were used as recipients of lymphocytes from sensitized donors, the normal mice responded significantly better than did infected mice after administration of an eliciting dose of oxazolone. An increase in pyroninophilic cells was observed in draining lymph nodes after application of a sensitizing dose of oxaxolone to the ears of either normal or acutely infected mice. These results indicate that suppression of contact sensitivity during acute T. cruzi infection is directed toward the efferent arm rather than the afferent arm of the response.


Immunity to antigenically related salmonellae: effects of humoral factors on the bactericidal activity of normal and immune peritoneal exudate cells.

Immunity against Salmonella enteritidis and Listeria monocytogenes was studied by measuring in vitro the bactericidal activity of peritoneal exudate cells (PEC) of control (normal PEC) and S. typhi Ty2-immune (immune PEC) mice. Specific immune serum, anti-S. tyhphi Ty2, heat inactivated at 56 degrees C for 30 min, significantly inhibited the growth of S. enteritidis only with immune PEC. These opsonic factors had no effect upon the activity of normal PEC. That such inhibition could not be demonstrated in Listeria experiments, either with immune or normal PEC, suggests that S. enteritidis was specifically recognized, in vitro, by the thermostable opsonin anti-S. typhi Ty2 and that macrophages from immune PEC were more efficient in inhibiting bacterial growth than those from normal PEC. Thus, the interaction between macrophages and the microorganism seems to play an essential role in cell-mediated as well as humoral immunity.


Heat-labile B-cell mitogen obtained from Listeria monocytogenes.

A water-soluble extract of Listeria monocytogenes strain 10403 acts as a mitogen on cultured mouse spleen lymphocytes. This mitogen induced a response six to nine times that of controls, as measured by [3H]thymidine incorporation. The mitogen extract was derived from washed bacterial cells which were mechanically disrupted with a French press. The extract was centrifuged at 105,000 X g and filtered through a 0.22-micrometer filter. Similar levels of lymphocyte stimulation were observed in lymphocyte cultures prepared from spleens of nude mice, indicating the effect of this mitogen on B-cells. The mitogenic property of this extract was destroyed by heating to 56 degrees C. This heat treatment does not destroy the antigens in the extract, which stimulate spleen cell cultures obtained from specifically immune mice. Similarly prepared extracts from Staphylococcus epidermidis and Salmonella typhimurium did not show similar levels of mitogenic activity. The mitogenic property of the L. monocytogenes extract was present in two strains of Listeria tested and was not associated with mouse virulence.


Purification of heat-labile enterotoxin from four Escherichia coli strains by affinity immunoadsorbent: evidence for similar subunit structure.

A single-step method for the purification of heat-labile enterotoxin of Escherichia coli is described. The method involves an affinity immunoadsorbent made with antiserum to cholera toxin. Crude toxin preparations of three human and one porcine enterotoxinogenic strains of E. coli were purified on this immunoadsorbent, and the elution products suggest that the toxin molecule is composed of subunits. One kind of subunit shared by these four strains showed similar mobility of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, close antigenic relationship, and an antigen in common with cholera enterotoxin.


Biological activities of pyochelins: iron-chelating agents of Pseudomonas aeruginosa.

Strains of Pseudomonas aeruginosa able to grow readily in serum (serum resistant) produce siderophores in large quantity, enabling them to extract iron from transferrins. The term pyochelin has been proposed for this group of compounds. Pyochelin extractable with ethyl acetate and designated pyochelin A appears to be a mixture of catechols and other phenolates. The structures of water-soluble siderophores, designated pyochelin B, have not been determined. Pyochelins enabled growth in serum of strains of serum-sensitive P. aeruginosa and other gram-negative bacilli. Serum-resistant strains of P. aeruginosa tended to be more virulent than equally toxigenic strains of the serum-sensitive group. However, incorporation of pyochelins into the inocula of serum-sensitive strains could reduce, rather than enhance, their virulence. Utilization of pyochelins by serum-sensitive strains of P. aeruginosa rendered some of these organisms resistant to pyocins which were otherwise lethal to them.


Isolation and characterization of a high-molecular-weight polysaccharide from the slime of Pseudomonas aeruginosa.

A procedure is described for isolating a high-molecular-weight polysaccharide (PS) from the slime of Pseudomonas aeruginosa immunotype 1. The resultant material, obtained from the void volume of a Sephadex G-100 column, was composed of carbohydrate and water. No lipopolysaccharide (LPS), 2-keto-3-deoxyoctonoate, heptose, phosphate, or protein was detectable, and nucleic acid contamination was generally below 1%. The carbohydrate composition of the PS was glucose, rhamnose, galactose, arabinose, and mannose. PS had a molecular weight of between 100,000 and 350,000 and did not disaggregate when chromatographed in the presence of sodium deoxycholate. An antigen immunologically indistinguishable from PS could be obtained from LPS by either acetic acid hydrolysis and column chromatography or by allowing solutions of LPS to stand at room temperature for 3 days. Some of this LPS-associated polysaccharide eluted as the void volume of a G-100 column but differed from PS by its lack of galactose and arabinose. LPS also contained an immunodeterminant not shared with PS that was detected by its stability to dilute alkali treatment (0.1 N NaOH, 37 degrees C, 2 h). PS was destroyed by alkali treatment. PS appeared to represent a form of LPS polysaccharide side chain that contains galactose and arabinose and is of a high molecular weight.


Protective immunity induced in mice by immunization with high-molecular-weight polysaccharide from Pseudomonas aeruginosa.

A high-molecular-weight alkali-labile polysaccharide (PS) isolated from the slime of immunotype 1 Pseudomonas aeruginosa was tested for its ability to protect mice from lethal challenge with the live, homologous organism. Intraperitoneal (i.p.) injection of 10 to 25 microgram of the PS protected 60 to 70% of the mice against challenge with up to 50 50% lethal dose units. Although single immunization of mice with up to 250 microgram of PS effected protective levels of only 70%, two successive immunizations provided 100% protection. Subcutaneous and intravenous immunization with PS also provided protection to i.p. challenges with immunotype 1 P. aeruginosa, but not to i.p. challenge with immunotype 4 P. aeruginosa. Although lipopolysaccharide (LPS) was found to be more immunogenic than PS in out studies, contamination of the alkali-labile PS with LPS did not account for the protection seen. Alkali treatment (0.1 N NaOH, 37 degrees C, 2 h) of the PS destroyed its protective effectiveness, while similarly treated LPS retained its capacity for inducing immunity in mice. Adsorption and passive protection studies with sera raised to either PS or a mixture of PS and LPS indicated that antibody directed to the alkali-labile PS antigen was capable of contributing to the protection of mice against challenge with P. aeruginosa.


Soluble antigens of virulent and attenuated biotypes of Brucella abortus.

Several methods were used to characterize three Brucella abortus biotypes (1, 5, and 7), including the attenuated vaccine strain S-19. Chemical analysis did not reveal remarkable differences among these strains, and only minor differences were noted in elution patterns of soluble extracts subjected to column chromatography. Qualitative and quantitative differences in extract components were demonstrated, however, by polyacrylamide gel isoelectric focusing. A distinctive difference was the presence of components in extracts from one or more of the virulent biotypes that were absent in similar preparations from the attenuated strain. In addition, one component common to all virulent strains was absent in strain S-19. Results of immunodiffusion experiments employing adsorbed and unadsorbed antisera also suggested that the quantity, quality, and surface distribution of various cellular antigens differed among the biotypes studied.