Effect of reducing agents on oxidation-reduction potential and the outgrowth of Clostridium botulinum type E spores.

Oxidation-reduction potential (Eh) levels were measured and standardized to pH (Eh7) for Trypticase soy broth containing various concentrations of reducing agents. Prereduced Trypticase soy broth with no added reducing agents exhibited a potential of -141 mV. Ascorbic acid at 0.2 to 0.005% and sodium thioglycolate at concentrations below 0.05% produced an Eh7 higher than the prereduced Trypticase soy broth containing no added reducing agents. The addition of cysteine hydrochloride,2-mercaptoethanol, and sodium formaldehyde sulfoxylate to prereduced Trypticase soy broth resulted in a reduction of Eh7 compared to the system without added reducing agents. The order of relative reducing intensity (from highest to lowest) for the reducing agents when comparing molar concentration was: sodium formaldehyde sulfoxylate,2-mercaptoethanol, cysteine hydrochloride, sodium thioglycolate, and ascorbic acid. Optimal growth of the test organism occurred at low Eh7 and low concentration of the reducing agents. A direct correlation existed between growth of the test organism and -Eh7 x -log concentration of the reducing agent.


Sodium nitrite and sorbic acid effects on Clostridium botulinum spore germination and total microbial growth in chicken frankfurter emulsions during temperature abuse.

Samples of (i) a control or of (ii) sodium nitrite-containing or (iii) sorbic acid-containing, mechanically deboned chicken meat frankfurter-type emulsions inoculated with Clostridium botulinum spores, or a combination of ii and iii, were temperature abuse at 27 degrees C. Spore germination and total microbial growth were followed and examined at specified times and until toxic samples were detected. The spores germinated within 3 days in both control and nitrite (20, 40 and 156 micrograms/g) treatments. Sorbic acid (0.2%) alone or in combination with nitrite (20, 40, and 156 micrograms/g) significantly (P less than 0.05) inhibited spore germinations. No significant germination was recorded until toxic samples were detected. A much longer incubation period was necessary for toxin to be formed in nitrite-sorbic acid combination treatments as contrasted with controls or nitrite and sorbic acid used individually. Total growth was not affected by the presence of nitrite, whereas sorbic acid appeared to depress it. Possible mechanisms explaining the effects of nitrite and sorbic acid on spore germination and growth are postulated.


Mutagenic activity of Fusarium moniliforme isolates in the Salmonella typhimurium assay.

A total of 33 isolates of Fusarium moniliforme from several food or feed crops were grown on sterile cracked corn, and chloroform-isopropanol extracts were assayed for mutagenic activity in the Salmonella typhimurium-microsome system by using tester strain TA98 or TA100 or both. Extracts of 21 (64%) of the isolates assayed against TA100 were mutagenic. Activities of seven of these extracts were increased markedly with incorporation of the liver homogenate (S-9) into the assay. Seven (33%) of the isolates assayed against TA98 were weakly active, with the liver homogenate having little effect on reversion rates.


Comparison of bacterial populations of the pig cecum and colon based upon enumeration with specific energy sources.

Concentrations of bacteria in the ceca and colons of pigs were measured by determinations of colony counts on rumen fluid-based media in anaerobic roll tubes. With our most complete medium (medium CCA), the mean colony count of cecal samples from 20 pigs was 2.37 X 10(10) +/- 1.0 X 10(10) (+/- standard deviation)/g (wet weight). The mean number of bacteria attached to or associated with cecal epithelial tissues from three pigs on medium CCA was 2.67 X 10(7) +/- 0.81 X 10(7)/cm2 of tissue. The proportions of gut bacterial populations able to use various energy substrates were estimated on the basis of relative colony counts. The following substrates are listed in descending order of their capacity to support growth of cecal bacteria: glucose, starch, cellobiose, xylose, Trypticase, gastric mucin from swine, mannitol, glycerol, and lactate. The effect of diet upon this distribution was not examined. The relative proportions of bacteria from a given population that were able to grow on various selective media were used as population profiles. Comparisons of populations in this way indicated that differences could be detected between (i) populations from the cecum of littermate pigs, (ii) populations from the cecum and colon of the same pig, and (iii) populations in the lumen of the cecum as compared with populations associated with cecal mucosa.


Propagation of ribonucleic acid coliphages in gnotobiotic mice.

To clarify the propagation cycle of bacteriophages in their natural habitats, we tested whether animals could support ribonucleic acid (RNA) phage propagation in their intestines, using germfree mice as the test animal. Propagation of four different antigenic types of RNA phages was tested. No detectable propagation or colonization of RNA phages was observed either in germfree mice or in gnotobiotic mice infected with the F- strain of Escherichia coli. Propagation or colonization was observed when RNA phages were orally introduced into gnotobiotic mice harboring the F+ or F' strain of E. coli. These results were consistent with data for in vitro propagation experiments. Fecal titers of phages were monitored over 24 to 98 days and were found to vary from 10(5) to 10(11) plaque-forming units per g of feces. Streptomycin administration gradually led to the disappearance of bacteria and, concomitantly, the RNA phages. Phages recovered from gnotobiotic mice feces included some of novel antigenic types. The bacterial isolates recovered from gnotobiotic mice harboring F+ bacteria included the original F+ strain, strains which had become F-, and some which had become inefficient hosts for the propagation of RNA phages.


Enzyme-linked immunosorbent assay for detection of Clostridium botulinum type E toxin.

The enzyme-linked immunosorbent assay using the "double-sandwich" technique was utilized to determine Clostridium botulinum type E toxin. With this technique, about 80 mouse intraperitoneal 50% lethal doses of toxin could be detected. Cross-reaction was hardly observed with C. botulinum type A and B toxins. No cross-reaction was observed with culture supernatants of C. botulinum type C or other Clostridium strains. In all probability this was due to the high specificity of the antiserum prepared aginst the toxic component of type E toxin.


New prodigiosin-like pigment from Alteromonas rubra.

The red prodigiosin-like pigment from Alteromonas rubra was shown to be a mixture of prodigiosin (pigment 1) and a new cyclic isomer (pigment 2). The new structure was elucidated by mass and nuclear magnetic resonance spectra. Careful examinations of the prodigiosins produced by Serratia marcescens, Vibrio psychoerythrus, and an unidentified red bacterium (LL-100-6) failed to disclose any of the new pigment, pigment 2.


Isolation of Legionella pneumophila from nonepidemic-related aquatic habitats.

Continuous centrifugation of large volumes of water from natural southeastern lakes allowed quantitative detection of Legionella pneumophila by direct immunofluorescent staining. Positive samples were injected intraperitoneally into guinea pigs, and the L. pneumophila were isolated and identified by their morphological, cultural, physiological, and serological characteristics.


Bullous dermatosis and systemic lupus erythematosus in a 15-year-old boy.

A 15-year-old boy had a bullous eruption suggestive of bullous pemphigoid and established systemic lupus erythematosus (SLE). Direct immunofluorescence studies of the bullae and adjacent skin revealed the linear deposition of IgG and complement localized to the basement membrane zone. Indirect immunofluorescence examination of the serum failed to reveal circulating basement membrane zone antibodies. The differential diagnosis of the bullous eruption is reviewed, and the problem of diagnosis in cases of coexistent bullous disease and SLE is discussed.


Physiological responses of the newborn infant to resuscitation.

Lung ventilation, endotracheal, and intraoesophageal pressures were measured during standard resuscitation of 20 asphyxiated babies born by caesarean section. The most common response to resuscitation was the production of a large positive intraoesphageal pressure. An opening pressure greater than 2.0 kPa was required to expand the lungs of most of these babies. The need for resuscitation was associated with prolonged maternal anaesthesia before delivery.


Comparative trial of manual and mechanical percussion technique with gravity-assisted bronchial drainage in patients with cystic fibrosis.

Chest physiotherapy still remains one of the most important aspects in the treatment of chest complications of cystic fibrosis. A mechanical device that allows the patient with cystic fibrosis to do his own chest physiotherapy will be of great benefit if it is as effective as manual percussion. 14 patients with cystic fibrosis using mechanical and manual percussion physiotherapy were studied by measuring sputum volumes, and FEV and FVC. Results with mechanical percussor were as good as with the manual percussor and, therefore, it would be reasonable for the older patient to use the former on his own.


Metabolic inhibition of plaque-forming cells: comparison of human rheumatoid-factor-producing cells with mouse anti-sheep erythrocyte-producing cells.

While rheumatoid-factor-producing haemolytic plaque-forming cells (RF--PFC) of the human peripheral blood were easily inhibited by cycloheximide, mouse spleen cells immune to sheep red cells (anti-SRC PFC) were inhibited only after prolonged preincubation in the drug. The RF--PFC were easily inhibited by propranolol, while the anti-SRC PFC were not at all inhibited. Vinblastine inhibited both systems equally. These differences are taken to suggest that the RF--PFC have very little preformed antibody in them and therefore depend upon active protein synthesis for their demonstration. In contrast, anti-SRC PFC, which may be predominantly mature plasma cells, generally need no new protein synthesis for their demonstration because of increased quantities of preformed antibody. A possible mechanism is that RF--PFC may represent primarily RF-specific B cells, the RF of which is released by surface immunoglobulin shedding and therefore susceptible to membrane stabilising agents such as propranolol.


The antiperinuclear factor. 1. The diagnostic significance of the antiperinuclear factor for rheumatoid arthritis.

In 1964 Nienhuis and Mandema reported the presence of antibodies against cytoplasmic granules in buccal mucosal cells in the serum of 50% of patients with rheumatoid arthritis (RA). Although they reported a good specificity for RA of these so-called antiperinuclear antibodies (APF), their results never threatened the monopoly of the rheumatoid factor as a serological tool for the diagnosis of RA. A re-evaluation with improved immunofluorescence methods showed a frequency of the APF of 78% in 103 patients with RA. The latex test and the Waaler-Rose test were positive in only 70% and 58% respectively of these patients. Only 15% of the RA patients were negative for all 3 tests. Thus, 40% of patients who were seronegative by the traditional methods gave a positive result on performance of the APF test. The high sensitivity of the APF test was combined with a good specificity, for the frequency in patients with other autoimmune diseases or degenerative joint disease and in healthy subjects was low. For the serodiagnosis of RA it seems best to combine the use of the APF test with one for rheumatoid factor.


Patterns of induced variation in the morphology, hyaluronic acid secretion, and lysosomal enzyme activity of cultured human synovial cells.

In contrast with newly isolated cells or early primary cultures, synovial cell lines in standardised growth conditions assume a rather uniform fibroblast-like appearance. However, 2 distinct variations in the cytological pattern can be induced at this stage. The first is characterised primarily by increased numbers of small phase-dense organelles that show the distinctive fluorescence of lysosomes after supravital staining, and are interspersed with vacuoles. The associated functional changes include increased enzyme activity and decreased net synthesis of hyaluronic acid. This variation can be induced by exposure to indigestible neutral sugars, adenosine, or its 5' nucleotides. The second variation consists of a striking reorganisation of cytoplasm by condensation into dense ridges or a dendritic network of processes. It is accompanied by increased hyaluronic acid secretion and is induced by agents that enhance intracellular activity of cyclic adenosine monophosphate, such as dibutyryl cyclic adenosine monophosphate and cholera enterotoxin. It appears possible to direct differentiation in synovial cell lines to correspond at least in part with the presumed functions of the different cell types in the parent tissue. The 2 patterns may be useful markers to correlate with other aspects of synovial cell function in vitro.


Effects of carrageenin on human synovial cells in vitro: morphology, hyaluronic acid production, growth, and the lysosomal system.

Some in-vitro effects of the arthritogenic polysaccharide carrageenin were studied on cells from human synovium. Synovial cells were isolated from intact human knee joints, and cell lines were developed by passaging with trypsin. Carrageenin was ingested by the cells but did not significantly affect cell growth, numbers of lysosomes, intracellular lysosomal enzyme activity (N-acetyl-beta-D-glucosaminidase), or release of lysosomal enzyme from cells. Carrageenin produced a reduction in net hyaluronic acid synthesis. It also induced a striking morphological change in a high proportion of synovial cells, characterised by increased spreading over the culture surface and apparent condensation of the cytoplasm into a pattern of ridges. Nonrheumatoid and rheumatoid synovial cells behaved similarly to one another.


[Congenital rubella. Present knowledge and perspectives (author's transl)].

This paper deals with features and problems of congenital rubella and perspectives in the prevention of this infection. We discussed the factors determining whether or not the maternal infection will involve the fetus, the pathogenesis and the clinical aspects of congenital rubella, the epidemiological importance of infected infants who may be contagious for long periods of time, the problems encountered in diagnosis. The efficacy of the active immunization programs, the data available in the literature on inadvertent vaccination of pregnant women, the restriction in the prophylactic use of immunoglobulins are also reported. The great importance of the premarital serological test for women is stressed.


[Candida infections in humans (author's transl)].

A complete review of the candidiasis dealing with taxonomy and morphology of the fungus and with epidemiology, experimental pathogenicity, immunology, mode of transmission, pathologic features, clinical aspects, diagnosis and treatment of the disease.


[R factors influence on motility of "E. coli" and "S. typhimurium" (author's transl)].

Influence on the motility of E. coli and S. typhimurium strains by several R factors has been studied; both classes of inhibiting and stimulating R factors have been detected; the plasmidic effect on the motility can be inverted by transfer to another carrier strain.


Management of perforating colon trauma: randomization between primary closure and exteriorization.

During a 44 month trial, 268 patients with wounds of the colon were entered into a prospective, randomized, nonblinded study. Consideration for primary closure demanded that: preoperative shock was never profound, blood loss was less than 20% of estimated normal volume, no more than two intra-abdominal organ systems had been injured, fecal contamination was minimal, operation was begun within eight hours, and wounds of colon and abdominal wall were never so destructive as to require resection. Once such criteria had been satisfied, colon wound management was dictated by last digit in the randomly assigned hospital number; odd indicated primary closure; even, exteriorization of the wound or primary closure with protection by a proximal vent. Results obtained in 139 determinant patients eligible for randomization revealed that primary closure (67 patients) had a lower infection rate of the incision (48% vs S7%, p > 0.05) and a still lower infection rate for the abdomen proper (15% vs 29%, p < 0.05) on comparison to the 72 patients with a randomized colostomy. Morbidity otherwise for the randomized colostomy was tenfold greater than if a primary closure had been performed. Average postoperative stay was six days longer (p < 0.01) if a colostomy had been created, exclusive of subsequent hospitalization for colostomy closure; while the total extra cost for management of the colon wound by colostomy was approximately $2,700.00. Although immediate mortalities were identical, one late death occurred following colostomy closure. These data not only confirm the safety of primary closure for colon wounds in selected cases, but also indicate that such should become the preferred method of treatment whenever specific criteria have been met.


[Serological diagnosis of hydatid disease with indirect immunoperoxidase and immunofluorescence methods (author's transl)].

Indirect immunoperoxidase (IP) method is compared with indirect immunofluorescence (IFI) in 44 cases of active hydatidosis established at operation (30 cysts of the liver, 9 cysts of the lung and 5 extra-hepatic and extra-pulmonary cysts). The sensitivities of the IP and IFI tests were 88.6% and 63.6%, respectively. The IP test is easy, specific and very sensitive.


Induction of unresponsiveness to major transplantable organs in adult mammals: a recapitulation of ontogeny by irradiation and bone marrow replacement.

Transplantation of renal allografts obtained from prospectively selected genotypically DLA-identical donors into supralethally irradiated dogs reconstituted with their own stored bone marrow has produced a state of unresponsiveness to these kidneys in the recipients. Eleven of 18 kidneys transplanted at 12 hours after marrow replacement currently survive with normal function and maintain life in the recipients for 757, 800, 825, 978, 1062, 1092, 1136, 1282, 1373, 1380, and 1381 days, respectively. Similar results occurred in eight of 13 allografts transplanted at 28 hours after marrow replacement, which currently survive for 349, 363, 377, 407,436,470, 485, and 513 days, respectively, and in eight of 13 kidneys grafted at 36 hours after marrow replacement, which are surviving for 197, 247, 298, 324, 337, 396, 443, and 472 days, respectively. Achievement of optimal results is dependent on the specific timing and sequence of each procedure. Only four of 16 recipients of kidneys transplanted at the time of marrow replacement were unresponsive to their allografts. Similarly, only five of 19 recipients of kidneys placed in irradiated dogs at 40 hours before marrow replacement accepted such allografts. When kidney transplants were placed into the recipients 20 hours before removal of marrow, irradiation, and reconstitution with stored marrow, only three of 21 dogs became unresponsive to such ailografts. In five of 12 instances, the recipients were also unresponsive to skin allografts obtained from their respective kidney donors. Such skin grafts currently survive for 606, 673, 687, 701, and 708 days, respectively. The remaining seven skin grafts were rejected at 28, 39,42, 84, 90, 92, and 115 days, respectively. Second- and third-set skin grafts from the same kidney donor were rejected by six of these dogs at 19, 20, 21, 29, 29, and 30 days, and at 21, 22, 23, 24, 27, and 27 days, respectively. Rejection of these skin grafts had no detectable effect on the function and survival of kidney allografts from the same source. Seven of eight skin grafts obtained from other DLA-identical donors were rejected at 13,14,16,25,28,38, and 84 days, respectively; one allograft continues to survive for 708 days. Eleven DLA-incompatible skin allografts placed on the recipients at the same time were rejected within 11-20 days. Supralethal total body irradiation and bone marrow replacement can establish in the adult canine host a privileged phase of immunological reactivity during which exposure to alloantigens produces specific long-term unresponsiveness rather than sensitization. The use of stored autologous rather than allogeneic bone marrow for reconstitution of the irradiated recipient eliminates the hazards of GVH complication usually associated with this procedure. This consideration and the apparent capacity of the tolerant host to maintain a long-term state of unresponsiveness without any further immunosuppressive therapy point to the potential relevance of the results to human transplantation.


Thoracic duct fistula and renal transplantation.

Thoracic duct drainage (TDD) was established for 21-115 days in 40 kidney recipients with an average removal per patient day of 4.7 1 lymph and 1.88 billion cells. Cellular and humoral immunity were depressed. TDD and immunosuppressive drugs were started at transplantation in 35 recipients of cross-match negative grafts. Although the results were better than in precedent non-TDD controls, eight patients rejected their grafts before a full TDD effect, and three of the eight developed predominantly anti-B lymphocyte cytotoxic antibodies which were probably responsible for positive cross-matches with their next donors. With continuing TDD, all eight patients had good initial function after early retransplantation. In five more "nontransplantable" patients with performed cytotoxic antibodies, TDD was started 30-56 days before transplantation. In these five pretreated patients, antibodies persisted with positive antidonor cross-matches. Hyperacute rejection occurred repeatedly in two patients with high anti-T (and anti-B) titers, but was surmounted in three patients with lower titers. From the clinical and immunologic data, we have concluded that TDD should be used for pretreatment of all cases with or without prior antibodies, and have suggested an adjustable management plan that takes into account new developments in antibody monitoring.


[Comparative study of the different media of enrichment and isolation for the identification of enteric pathogens (author's transl)].

In the present study, were compared several media in use in diagnostic laboratory for the isolation of enteric pathogens. The AA. have remarked the efficiency of Bacto-Selenite F and of Bacto-TT Broth Base Hajna, for the enrichment; it is suggested the use of Bacto Agar and of Bacto-Hektoen Enteric Agar for the isolation.


Ten year experience with renal transplantation in juvenile onset diabetics.

Between 1968 and 1978, 305 juvenile onset diabetic patients with uremia and 462 nondiabetic uremic patients of similar age received primary renal allografts at the University of Minnesota. Two hundred eight of the diabetic patients are alive and 190 have functioning renal grafts three months to ten years after transplantation. Cumulative patient survival rates at two years for diabetic recipients of kidneys from HLA identical siblings, other related and cadaver donors are 90, 73 and 68%, respectively, the corresponding graft functional survival rates are 90, 67 and 55%. For nondiabetic patients receiving kidneys from the same donor categories the corresponding patient survival rates are 97, 86 and 75%, while the graft functional survival rates are 94, 77 and 64%. The differences in patient and graft survival between diabetic and nondiabetic recipients are statistically significant only for the patients receiving grafts from HLA-nonidentical related donors. For all recipients under the age of 30, there are no statistically significant differences in patient and graft survival. Regardless of the age of the patient or the source of the kidney, the survival of diabetic patients treated with transplantation at our institution is better than the use of chronic hemodialysis, alone. Technical complications do not occur more frequently in diabetic transplant recipients. Cardiovascular disease is responsible for most of the late deaths in these diabetic patients. Amputations of digits or extremities have been required in 15% of the diabetic patients. On the positive side, the vision of 88% of these recipients remained stable or had improved visual acuity, and 82% of the diabetic patients were actively rehabilitated after transplantation. Kidney transplantation is the treatment of choice for end-stage renal failure in diabetic patients, just as it is for most uremic patients.


Alimentary tract complications after renal transplantation.

A computer analysis of post renal transplantation gastrointestinal problems was performed to identify important associated clinical factors. Thirty-seven per cent of all transplant recipients developed one or more significant problems. Hemorrhage, nondiverticular intestinal perforation, and esophagitis occurred most frequently in hospitalized patients. Pancreatitis, diverticulitis, and gastroduodenal perforation occurred characteristically in long-term survivors with well functioning allografts. Eleven of 32 HLA identical recipients treated with maintenance corticosteroids during stable kidney function developed gastrointestinal disease while only one of 13 HLA identical recipients not given maintenance steroids developed a problem, which strongly suggests a causal role for steroids in the development of late complications. The association of preexisting peptic ulcer and diverticular disease with hemorrhage and perforation supports previous recommendations that documented peptic ulcer disease or diverticulitis should be corrected surgically prior to transplantation.


Obliteration of esophageal varices by PTP: a follow-up of 43 patients.

The percutaneous transhepatic portal vein catheterization (PTP) with selective obliteration of the coronary vein and/or the short gastric veins in treating bleeding esophageal varices was introduced in 1974. In order to prevent recanalization of the vessels Bucrylate (isobutyl-2-cyano-acrylate) has been used in 43 patients 55 times during a period of 34 months (October 1975 to July 1978). The obliterative treatment was followed by rebleeding in 35% of the cases and continued bleeding occurred in two patients. Fourteen patients were treated on 16 occasions during acute bleedings, and five of these (36%) died within two months from a portal vein thrombosis caused by the obliterative procedure. Because of these findings PTP with obliteration of the veins feeding the esophageal varices is not recommended as an elective way of treatment. It should only be used in the acute bleeding patient when transesophageal sclerosering therapy, continuous vasopressin infusion and balloon tamponade have failed. Fifty-six per cent of the patients acutely treated stopped bleeding for more than one week, thus avoiding an emergency shunt or devascularization operation which are associated with a high mortality rate.


Effect of diazepam on serum testosterone and the ventral prostate gland in male rats.

A study was initiated to investigate the effect of diazepam on serum testosterone and testosterone-sensitive tissues in male rats. Diazepam was administered in a dose of 50 mg/kg body weight daily for 10 days. When compared with an equally matched group of control male rats, this dose of diazepam was associated with a significant reduction in both the weight of the ventral prostrate of treated rats and the serum testosterone levels. These changes were not associated with alterations in serum LH and FSH and the hypothalamic luteinizing hormone-releasing hormone content. Since diazepam does not interfere with the radioimmunoassay of testosterone, it is suggested that the above observations were biologically induced via direct suppression of the interstitial cells of the testis.


Effect of a 5'-substituted adenosine analogue (744c96) on insulin and glucagon release in isolated rat islets of Langerhans.

The effect of adenosine-5'-(N-ethylcarboxamide) (744-96), a highly vasoactive adenosine analogue, was investigated on insulin and glucagon release from isolated rat islets of Langerhans. The compound inhibited glucose-induced insulin release in a dose-dependent manner (concentration range 10(-8) to 10(-5 M). No effect of 744-96 was observed on basal or on arginine-induced glucagon release. By contrast, 744-96 (5 microgram/kg i.v.) produced a significant increase in plasma glucagon and glucose levels in normal as well as in reserpinized rats. The observed failure to promote glucagon release from isolated islets may be attributable to the isolation procedure, to a release of glucagon in vivo from non-pancreatic stores, or to a non-specific effect of 744-96 on glucagon secretion.


The role of prostaglandins in platelet-vessel wall interaction.

Flurbiprofen, a cyclooxygenase inhibitor, and tranylcypromine, a PGI2 synthetase inhibitor, in topical superfusion over an arterial segment respectively decrease and increase ADP-induced local white platelet thrombus formation. Superfusion of the arterial segment with PGI2 demonstrates a dose-response related inhibition of ADP-induced thrombosis.


Facilitation of bicuculline- and picrotoxin-induced seizures by sodium valproate in rats.

Paralysed rats anaesthetized with urethane or halothane were injected with bicuculline or picrotoxin at doses which induced seizure activity in the EEG. Sodium valproate (50--1200 mg/kg) or saline was injected i.p. and its effect on the amplitude and duration of the seizures was measured. The seizures induced by either convulsant were increased significantly by doses of valproate greater than or equal to 200 mg/kg. The sites of action of the drugs in the brain, and the possible transmitter mechanisms involved, are discussed.


Renal transplantation in the amyloidosis of familial Mediterranean fever. Experience in ten cases.

Ten patients with familial Mediterranean fever (FMF) and histologically confirmed amyloidosis received cadaver kidney transplants for treatment of terminal renal disease. Colchicine, 1 mg daily, was included in the routine postoperative regimen from 1974 for amyloidotic patients. Graft and patient survival were compared with ten nonamyloidotic recipients of renal grafts matched for age, sex, type of allograft, and HLA compatibility. In the FMF group, five of ten grafts have survived from 20 to 64 months; in the control group, six of ten. While only recipients with functioning grafts survived in the FMF group, patient survival in the control group is eight of ten after one year. In all five FMF survivors, graft function is satisfactory, proteinuria is absent, and blood creatinine levels are normal. Amyloid involvement of an allograft was documented 16 months after transplantation in the only patient whose maintenance colchicine dosage had been reduced to 0.5 mg daily.


Covalent linkage of lipoprotein to peptidoglycan is not essential for outer membrane stability in Proteus mirabilis.

Isolated rigis layers from Proteus mirabilis harvested at different growth phases were degraded by endo-N-acetylmuramidase from Chalaropsis B, and the degradation products were investigated. The results show the complete absence of covalently linked lipoprotein in exponential-phase cultures. Stationary cells, however, possess covalently linked lipoprotein in amounts similar to those found in Escherichia coli or Salmonella typhimurium during all growth phases. The overall peptidoglycan structure did not change during transition from logarithmic to stationary growth. Implications of these findings for the organization of the outer membrane are discussed.


Relaxed mutants of Serratia marcescens SM-6. Biochemical traits and relevance of the rel+ allele for the formation of exoenzymes.

Serratia marcescens SM-6 when starved for a required amino acid stops synthesizing protein and RNA and accumulates two nucleotides which co-chromatograph with ppGpp and pppGpp. These features are characteristic of bacterial strains with stringent RNA control (rel+). Two independent mutants were isolated which resemble relaxed (relA) mutants of Escherichia coli; they continue to synthesize RNA and accumulate neither ppGpp nor pppGpp when deprived of the required amino acid. The extracellular enzyme activities (nuclease, protease, lipase) of the relaxed mutants are about the same as those of the parental stringent strain when studied under standard growth conditions. Exoenzyme-deficient (nuc;prt) and exoenzyme-hyperproducing (nucsu) mutants were isolated from both stringent and relaxed strains of S. marcencens SM-6 and no change of the cellular ability to form ppGpp and pppGpp could be observed. From these results it appears that the formation of exoenzymes of S. marcescens SM-6 is independent of stringent/relaxed RNA control.


[The adolescent confronted with hemodialysis and transplantation, psychological problems and developmental factors].

A study has been undertaken of the psychological problems of more than 250 adolescents and young adults with chronic renal failure in a dialysis and transplantation programme. The centre provided treatment and helped the patients to adapt to their illness. As the illness is of indefinite duration observations could be made of the course and the outcome. The individuals reaction to different forms of treatment, the possible final outcome and the institution were studied.


Serendipity in biological psychiatry--a myth?

It is often stated that major biological treatments in psychiatry were discovered by accident or serendipity. Tracing the history of the concept of serendipity, we find that serendipity has been subjected to greatly divergent interpretations. According to the current usage, it is a discovery in which chance was a necessary and/or sufficient condition. With this definition, none of the discoveries of major biological treatments in psychiatry can be labeled serendipitous. The necessary factors common to these discoveries were creative minds that were variably influenced by the zeitgeist and that were persistent in their search for answers. Another important prerequisite was the availability of crucial basic knowledge of many related sciences. We conclude that chance cannot substitute for long-term research and that the latter is the most likely way to lead to valuable discoveries.


Exploratory psychotherapy in the treatment of psoriasis. Twelve hundred years ago.

Twelve hundred years ago, Jasaliq, a Persian physician, treated a patient who was suffering from psoriasis. Although psoriasis was not clearly differentiated from leprosy in the West until the 1840s, it was clinically described and differentiated as a separate entity in Persia in the eighth century AD. In this case history reported in one of the oldest Persian texts, Chahar Maqala, written in 1155 AD, the physician formulated a dynamic relationship between the outbreak of psoriasis in his patient and the existence of severe interpersonal conflicts. In treating his patient, he skillfully used therapeutic modalities that are now prevalent in exploratory and psychodynamic psychotherapy. The techniques of developing a therapeutic alliance, therapeutic confrontation, clarification, dynamic interpretation, and exploration of intrapsychic and interpersonal conflicts are in accord with our present concepts of exploratory and dynamic psychotherapy.


Plasmodium berghei--infected mice. Focal glomerulonephritis in hyperimmune state.

Twenty-one male Swiss albino mice were injected intraperitoneally with 2 x 10(5) Plasmodium berghei berghei-infected cells. Twenty-four mice were used as a noninfected control group. On the tenth day after infection, three mice were killed and the remainder were treated with chloroquine phosphate (0.025% solution) given orally. The dosage was subsequently adjusted to keep the parasitemia below 5%. Eighteen of the controls received the same chloroquine dosage. Diffuse proliferative glomerulonephritis was seen from day 10 of infection up to two months after treatment. Granular deposits of immune complex were seen in all glomeruli. At the end of third month and thereafter, all experimental mice were in a hyperimmune state, and the kidneys showed the characteristic of focal glomerulonephritis resembling some types of human focal glomerulonephritis.


Electron-dense subepithelial glomerular deposits in Henoch-Schönlein purpura syndrome.

A case of Henoch-Schönlein purpura (HSP) with renal lesions is presented. The clinical, laboratory, and morphologic findings revealed features commonly described in HSP. A renal biopsy revealed diffuse, generalized, proliferative glomerulonephritis. The unusual findings in this case were numerous electron-dense subepithelial deposits ("humps") and a few small intramembranous deposits in the glomerular capillaries without deposits in the mesangium, mimicking those of acute poststreptococcal glomerulonephritis. The electron-dense deposits were well correlated with deposits of IgG, IgA, C3, C4, C1q, and fibrin-fibrinogen demonstrated by immunofluorescent microscopy.


Primary excision and grafting of burns located over an open fracture.

Early excision of the burn, immediate autografting, reduction of the fracture, and stabilization with a plaster-of-paris cast is offered as an alternative method for managing burns with associated long-bone fractures. The opportunity to excise and immediately autograft a burn wound requires rethinking of the entire problem of burns associated with a fracture.


A clinical and laboratory evaluation of a polyurethane foam: a new donor site dressing.

A polyurethane foam (Lyofoam) has been reported to accelerate epithelization of a wound. The purpose of this study was to evaluate its efficacy as a donor-site dressing for thermally injured patients. Thus, partial-thickness injuries were made in ten pigs and covered with Lyofoam, Xeroform, Telfa, Scarlet Red, and fine-mesh gauze. Gross and histologic examinations failed to show accelerated healing under the Lyofoam dressing but did show that Scarlet Red covered donor sites healed the fastest. On clinical evaluation, nine patients only showed that Lyofoam separated earlier from the underlying wound but there was no evidence to suggest that the wound was more mature than that covered with fine-mesh gauze.


Lung expansion, airway pressure transmission, and positive end-expiratory pressure.

Transmission of airway pressure to the intrapleural space and change in functional residual capacity by positive end-expiratory pressure (PEEP) were measured in ten anesthetized swine. Measurements and calculations were performed with varying lung and chest wall compliances. When both compliances were normal, approximately half of the applied airway pressure was transmitted. Aspiration of hydrochloric acid reduced lung compliance approximately fourfold and decreased airway pressure transmission. Increased thoracic compliance also reduced airway pressure transmission. When acid aspiration reduced lung compliance and sternotomy simultaneously increased thoracic compliance, pressure transmission was maximally reduced. Decreases in either thoracic or lung compliance reduced the volume-expanding effects of PEEP. Positive end-expiratory pressure was least effective when thoracic and lung compliances were reduced simultaneously. Careful assessment of both lung and thoracic compliances may be helpful in treating patients requiring elevated airway pressure.


Attempted enhancement in patients undergoing renal allotransplantation: lack of long-term sensitization.

It seems timely and pertinent to review the long-term follow-up of four patients who were deliberately exposed before transplant to donor antigens in an "enhancement" protocol. Not only were there no adverse effects observed, but three of the four patients never had an identifiable rejection episode. One of the successful allograft recipients had cytotoxic antibodies to his donor's cells. These appear to have been cold, T- and B-cell-reactive antibodies of doubtful significance. Both long-term surviving recipients showed weakness in degree of mitogenesis in mixed culture of donor and recipient peripheral blood mononuclear cells, which does not appear to be related to a macrophage defect but may be related to a lack of active rosette-forming T cells.


Urinary tract infections in kidney transplant recipients.

In 65 kidney transplant recipients who were followed up for a mean period of 14.7 months, the incidence of urinary tract infection (UTI), and how the incidence was affected by length of graft survival, age, HLA-A and HLA-B matches, complications, duration of Foley catheter use, and other aspects, were examined. The total incidence of infection included an unexpectedly high rate of late infections. The incidence was found to be statistically increased with nephrectomy, splenectomy, recatheterization, and age older than 40 years. There was no correlation noted with graft source, antigen match, graft loss, or previous history of UTI. A group of patients with persistent UTI was noted and an inability to suppress UTI with long-term therapy with antibiotics was found. The asymptomatic nature of most of the UTIs confirmed the need for frequent periodic cultures of urine in the immunosuppressed patient.


Basic requirements of pancreatic mass for transplantation.

In canine pancreas autografts, more than 20% (30% to 40%) of the pancreatic mass (body and distal head or proximal tail) is required for the establishment of a satisfactory endocrine response. Transplantation of 20% of the pancreatic mass did not result in an adequate restoration of the endocrine function. We believe that this study gives a practical answer to the question of how much pancreatic mass is necessary for the establishment of normal functional response after transplantation. This autotransplantation model precludes the determination of the content and volume of transplanted pancreatic islets. Thus, our model indicates the requirements of the pancreatic mass of the specific anatomic areas that were transplanted, but does not indicate the actual amount of transplanted islets.


The carcinogenicity of lead.

The potential for lead to cause neoplasm in animals and man is reviewed. A multitude of studies indicate that principally renal tumours may be produced by various forms of inorganic lead in small rodents. No human study either of an epidemiological form or of a case report in industrial, agricultural or community medicine has proven that lead may cause cancer in man.


IgM fluorescence antibodies in sera of pregnant women exposed to rubella.

A high per cent of false positive results for rubella virus IgM antibodies determined by immune fluorescence were detected in sera of pregnant women. After absorption with Staphylococcus suspension, Cowan 1 strain, and aggregates of human immunoglobulins, the false positive reactions due to IgM antiimmunoglobulin were eliminated.


[Origin of fibroblasts and macrophages of skin wound granulation tissue].

By the method of thymidine autoradiography and fluorescing antibodies, the origin of cells participating in healing the experimentally produced wounds was investigated. 3H-thymidine was administered a day before the production of the wounds. 14C-thymidine was injected impulsively 1 h before the material was fixed 1,3 and 8 days after the trauma. The analysis of the autographs demonstrated that during wound healing the cell-precursors of macrophages and fibroblasts migrate from beyond the limits of the connective tissue. By means of the fluorescing antibodies in mouse radiation xenogenic chimaeras it was demonstrated that macrophages and fibroblasts participating in wound healing take their origin from the donor's elements, that is, are of bone marrow origin.


[Stereologic analysis of the thyroid gland (review of methods)].

The most important stereological methods applied for quantitative morphologic investigation of the thyroid gland are considered: estimation of relative volumes of the organ's components, of follicular surface, of follicular surface/volume ratio, of number of cells and follicles per volume unit and in the whole organ, their distribution according to their size. The main formulae are presented and the most effective ways in application of stereological of stereological methods for investigation of normal and pathological gland in men and animals are described.


[Endothelial barrier, foam cells and lipoprotein metabolism in arterial walls (based on experimental arteriosclerosis in rabbits)].

The features of impaired permeability of endothelium and increased pinocytosis of endothelial cells in developing experimental atherosclerosis in rabbits are described on the basis of electron microscopy findings. Foam cells of various degrees of vacuolation were recovered from the tissue of atherosclerotic aorta by enzymatic treatment. The stages of smooth muscle cell transformation into foam cells were followed. An analysis of morphological changes occurring in foam cells in response to lipids is given. Interactions of intracellular lipid vacuoles with lysosomes are analysed. Possible causes and modes of impairment of the structure and function of aorta cells in experimental atherosclerosis are discussed.


The pregnant epileptic: a review and recommendations.

The major concerns in the pregnant epileptic patient are loss of seizure control and the teratogenic effects of antiepileptic drugs on the fetus. Loss of seizure control is usually caused by a progressive decline of antiepileptic plasma levels throughout pregnancy. This decline can be prevented by monthly dose adjustments based on plasma level determinations. Although infant malformations are a more prevalent outcome of the pregnancies of epileptics than of nonepileptics, the role of antiepileptic drugs in teratogenicity is not fully established. Only trimethadione has been convincingly linked to fetal malformation. Recommendations for the management of epilepsy in pregnancy are made.


Meiges disease: a clinical form of facial convulsion, bilateral and medial.

In 1910 the French neurologist Henry Meige described in detail a disorder characterized chiefly by symmetric dystonic spasms of the facial muscles, which he called "spasm facial median." Cases with this disorder are rare, and frequently misdiagnosed and inappropriately treated. We report here a translation of Meige's original description of "spasm facial median." We hope that this translation will make it easier to recognize and diagnose this disorder and that it will stimulate greater interest in this unusual syndrome.


Disappearance of Kayser-Fleischer rings after liver transplantation.

Kayser-Fleischer rings were observed in a 14-year-old boy with progressive liver disease. Laboratory data supported a diagnosis of hepatolenticular degeneration (Wilson's disease). Therapy with chelating agents was initiated, but discontinued because of side reactions. Successful liver homotransplantation was performed. The Kayser-Fleischer rings gradually faded and had completely disappeared by six years postoperatively.


Interferometric visual acuity testing in anterior segment disease.

Preoperative and posteroperative visual acuities for Landolt ring targets and for a grating target of comparable retinal illuminance produced by two-beam interference were assessed in patients with anterior segment disease. Good preoperative interferometric acuities (6/12 or better) were found in over half of the cases in which Landolt acuities were 6/90 or worse. In general, good preoperative interferometric acuities were predictive of good postoperative Landolt acuities. However, poor interferometric acuities in patients with opacified media did not preclude good Landolt acuities postoperatively. We conclude that interferometric acuity testing can provide valuable information about potential macular resolution capacity in some cases. Both the strengths and weaknesses of interferometric acuity testing must be appreciated for successful application.


Suspending sutures in blepharoplasty.

Lateral canthopexy using permanent suspending sutures to shorten the lateral canthal tendon helps prevent scleral show, increased sloping of the upper lid, and inferomedial displacement of the lateral canthus following lower lid blepharoplasty. While these complications usually are prevented by flap suspension techniques, in which more skin is excised above and lateral to the commissure than below it, sutures running from the inferomedial part of the lateral canthal tendon to its superolateral aspect and to periosteum just inside the orbit still further reduce the tendency toward the complications mentioned. We report technical aspects and results. After several years of clinical experience, we have the impression that the technique is a useful adjunct, even though it may produce more swelling and inflammation and a longer recuperative period.


Implants of Supramid, Proplast, Plasti-Pore, and Silastic.

Four types of synthetic materials were implanted in 54 rabbits: Supramid, Proplast, Plasti-Pore, and Silastic. Each material was implanted into the auricle and concomitantly into the subdermis of the face. Postoperative assessment was every one to three days, while killing was done at six weeks, six months, and one year postoperatively for microscopic evaluation. Supramid mesh contains black pigment, which became incorporated within the histiocytes and giant cells around the implanted mesh by six weeks; at one year, the soft tissues were pigmented microscopically. Proplast was not extruded despite an intense and persistent histiocytic and giant-cell reaction; its black color constituted a major drawback for implantation beneath thin skin. The Plasti-Pore used in these studies was rigid and somewhat difficult to sculpture, and it induced a persistent but minor cellular reaction. Silastic was easily displaced.


The psychological treatment of cancer: the patient's confusion of the time for living with the time for dying.

It has been shown that it is possible to influence cancer growth by a form of intensive meditation, although it is not yet established whether it can be influenced to the point of cure. In working with these patients it has been observed that the course of the illness has often been influenced by the patient's confusion of the biologically appropriate time for living and the time for dying. Without recourse to any formal psychotherapy, the family physician aware of this reaction may be able to enhance the immune defences and increase the quality of life of such patients.


Relationship between thermogenic drinking and plasma renin activity in the rat.

The spontaneous water intake of rats increases when they are transferred abruptly from a cold (5 degrees C) to a neutral (25 degrees C) environment. This has been termed thermogenic drinking. Treatment of cold-acclimated rats with SQ 14,225, an angiotensin converting enzyme inhibitor, at 10-50 mg/kg of body weight prior to removal from cold, inhibited the thermogenic drinking response in a dose-dependent manner. Plasma for determination of plasma renin activity (PRA) was obtained by cardiac puncture from methoxyflurane anesthetized rats maintained chronically at both 25 degrees and 5 degrees C. In addition, plasma was obtained from cold-acclimated rats 15 min after removal from 5 degrees to 25 degrees C. PRA values were 2.2 +/- 0.4 (S.E.) ng/ml/h for control rats; 1.9 +/- 0.8 ng/ml/h for cold-acclimated rats and 8.5 +/- 1.7 ng/ml/h for cold-acclimated rats removed from cold for 15 min. Thus, PRA was significantly increased in rats removed acutely from cold. These data suggest that thermogenic drinking may be mediated by the renin-angiotensin system.


Evaluation of several methods of detecting salmonellae in groups of chickens.

Chickens exposed experimentally to Salmonella agona, S. blockley, S. enteritidis, S. infantis, and S. typhimurium were tested serologically by microagglutination and microantiglobulin procedures and culturally by cloacal swabs and environmental samples. Chicks infected at one day old yielded the highest level of positive cloacal swab cultures, whereas exposure of older birds gave maximal serologic titers. Recovery of salmonella from environmental samples is dependent on a number of factors, including excretion rate by the population and survival rate of salmonellae in the environment. Serologic titers persisted after salmonellae could no longer be isolated from cloacal swabs or environmental samples. The possibility of false-positive serologic reactions was explored in two groups of chickens, one exposed to other Enterobacteriaceae, and one composed of eight specific-pathogen-free flocks. Occasional serologic microantiglobulin titers were observed only in the latter group. Extensive culturing of the environment from these flocks had negative results.


Prevalence of certain antibodies to selected disease-causing agents in wild turkeys in Texas.

In Texas in 1976 and 1977, Rio Grande turkeys (Meleagris gallopavo intermedia) from seven counties and Eastern turkeys (M. g. silvestris) from one county were tested for antibodies to selected poultry pathogens. Standardized serological tests disclosed reactors to Salmonella pullorum (2.4%), S. typhimurium (2.3%), and Mycoplasma gallisepticum (4.8%). There were no reactors to Newcastle disease virus or Chlamydia psittaci. Prevalence of M. gallisepticum antibody in wild turkeys was significantly higher for counties with commercial turkey operations than for counties lacking domestic turkeys, whereas the incidence of S. pullorum and S. typhimurium did not differ significantly.


Adenosine triphosphate consumption by bacterial arginyl-transfer ribonucleic acid synthetases.

ATP consumption by arginyl-tRNA synthetases from Escherichia coli and Bacillus stearothermophilus has been investigated by the firefly luciferin--luciferase assay. Arginyl-tRNA synthetase from E. coli utilizes ATP only for aminocylation of tRNA with a 1:1 stoicheiometry. In contrast, we have shown an adenosine triphosphatase activity of arginyl-tRNA synthetase from B. stearothermophilus in the absence of tRNAArg. Dowex chromatography revealed the formation of ADP by the thermophile enzyme; under aminoacylation conditions, AMP was also formed in amounts stoicheiometric with arginyl-tRNA formation.


Thiamin biosynthesis in Saccharomyces cerevisiae. Origin of carbon-2 of the thiazole moiety.

Radioactivity from [2-14C]glycine enters C-2 of the thiazole moiety of thiamin and no other site, in Saccharomyces cerevisiae (strains A.T.C.C. 24903 and 39916, H.J. Bunker). Radioactivity from L-[Me-14C]methionine or from DL-[2-14C]tyrosine does not enter thiamin.


Metabolism and insulin-releasing capabilities of glucosamine and N-acetylglucosamine in isolated rat islets.

The ability of glucosamine and N-acetylglucosamine to stimulate insulin secretion from perifused rat islets and the suitability of these hexoses to be metabolized in a static incubation was studied under various conditions. N-Acetylglucosamine alone stimulated insulin release with a threshold of 10 mM, with half-maximal effect at approx. 16 mM, and maximally at 20 mM. With higher concentrations stimulation was slightly diminished. Release caused by 20 mM-N-acetylglucosamine was unaffected by 30 mM-mannoheptulose, but was blocked by 2-deoxyglucose or iodoacetate (1 mM). At moderate concentrations, (2.75--20 mM), the metabolism of N-acetyl[1-3H]glucosamine was similar to that of [1-3H]glucose and secretion rates paralleled the corresponding rates of metabolism with these hexoses. Glucosamine (27.5 mM) alone weakly stimulated insulin secretion, which was unaltered by 30 mM-mannoheptulose but blocked by 2-deoxyglucose or iodoacetate. A lower rate of [1-3H]glucosamine metabolism appeared to account for its weaker stimulatory efficacy. Insulin release caused by 27.5 mM-glucosamine or 27.5 mM-N-acetylglucosamine in the presence of basal (2.75 mM) glucose was accurately predicted based on the summed metabolic rates of these compounds. The data strengthen the theory proposing that metabolites or cofactors generated during metabolism are essential for triggering insulin secretion.


Studies on the inhibition of protein synthesis by selenodiglutathione.

Amino acid incorporation in a cell-free system derived from rat liver has previously been found to be inhibited by GSSeSG (selenodiglutathione). In the present experiments the effect of GSSeSG on protein synthesis in 3T3-f cells, on growth and protein synthesis in Escherichia coli, and on amino acid incorporation in a cell-free system derived from E. coli, was studied. GSSeSG inhibits the incorporation of [3H]leucine into protein by 3T3-f cells. This inhibition cannot be reversed by removing GSSeSG and is correlated with the uptake of GSSeSG. Sodium selenite (Na2SeO3) and oxidized glutathione had no inhibitory effect in this system. [3H]Uridine or [3H]thymidine incorporation into RNA or DNA was not inhibited, indicating that the primary action of GSSeSG was on protein synthesis. GSSeSG did not influence the growth of E. coli in a synthetic medium, although enhanced amino acid incorporation was observed. In the cell-free system derived from E. coli, amino acid incorporation was not changed by GSSeSG, indicating that elongation factor G, in contrast to elongation factor 2 of mammalian cell systems, is not blocked by GSSeSG.


Defective calcium handling and insulin release in islets from diabetic Chinese hamsters.

In pancreatic islets from normal Chinese hamsters preloaded with 45Ca2+, glucose-induced biphasic insulin release was associated with increased 45Ca2+ efflux; islets from diabetic hamsters showed decreased insulin release and no increase in 45Ca2+ efflux. The lack of stimulated 45Ca2+ efflux persisted even when glucose-induced insulin release was potentiated by 3-isobutyl-1-methylxanthine. Since glucose-stimulated 45Ca2+ uptake by diabetic islets was not impaired, a defect in intracellular Ca2+ handling may be involved in the defective insulin release of the diabetic Chinese hamster.


Localization of the heparin-releasable lipase in situ in the rat liver.

Immunofluorescence and immuno-electron microscopy were used for the localization of the heparin-releasable lipase in situ in the rat liver. The lipase is located exclusively on the liver endothelial cells. No labelling could be detected on the parenchymal of Kupffer cells, or in the livers of heparin-pretreated animals. The physiological significance of the endothelial localization of the hepatic lipase is discussed.


[Effect of non-steroidal anti-inflammatory drugs on plasma protein binding of corticosterones (author's transl)].

A series of non-steroid antiinflammatory drugs (NSAD) as well as substances with different spectra of pharmacological action were investigated with regard to the influence on binding of corticosterone to plasma protein in rats after repeated oral administration. At pharmacologically active doses, NSAD cause a time and dose dependent increase in rat plasma of the corticosterone fraction which can be ultra-filtrated. The effect on corticosterone binding to plasma protein largely parallels the antiinflammatory efficacy of the NSAD. This does not occur by a mere displacement of corticosterone from the plasma protein binding, but is apparently the consequence of a qualitative and/or quantitative change of the binding proteins. Organic acids, cytostatic or other drugs which strongly bind to plasma protein do not show any corresponding influence on the corticosterone binding to plasma protein under comparable conditions. The increase of the fraction of corticosterone which can be ultra-filtrated effected by the NSAD accelerates the excretion of corticosterone into the bile of rats. The phenomenon of decrease in binding of corticosteroids to plasma protein is discussed with respect to its importance for the mode of action of the NSAD.


[Methods of determination and theoretical principles of the serum protein binding of drugs (author's transl)].

A survey is given of the methods applied in the determination of protein binding (electrophoresis, ultrafiltration, gel-filtration, ultracentrifugation, dialysis and microbiological plate test). Basing on our own investigations as well as on papers by other authors the possibilities of errors and limitations of the methods are discussed. The protein binding is reversible and can be explained by the law of mass action. By the representation of the test results according to Scatchard plot it is possible to ascertain the number of the binding groups of the albumin molecule and the equilibrium constant of the reaction. Based on these results the affinity (free reaction, energy) of the protein binding can be calculated. The protein binding can be described especially in therapeutical concentration rather well by the Freundlich isothermal curve. The binding powers which cause the protein binding are discussed. The examination of various substance groups, such as sulphonamides, antibiotics and steroid hormones has shown that the hydrophobic binding plays the main role in the binding of drugs to albumin. As an example of an ionogenic binding the binding or aminoglycosides to alubmin is discussed.


[Optically active barbiturates. Synthesis, configuration and pharmacological effects (author's transl)].

Optically active N-alkylated barbiturates are synthesized from disubstituted cyanoacetates. The configuration of the synthesized compounds is determined by chemical and chiroptical procedures and by X-ray analysis. In animals the barbiturates show a different anesthetic activity, in some cases the CNS activity is opposed, one enantiomer is anesthetically active, the other is a convulsive agent. In their pharmacokinetic behaviour the enantiomers show remarkable differences.


Bactericidal activity of cefazedone.

Distinct bactericidal activity of (6R,7R)-7-(2-[3,5-dichloro-4-oxo-1(4H)-pyridyl]-acetamido)-3-([(5-methyl-1,3,4-thiadiazol-2-yl)-thio]methyl)-8-oxo-5-thia-1-aza-bicyclo[4,2,0]oct-2-ene-2-carboxylic acid (cefazedone, Refosporen) could be demonstrated in 2 Staphylococcus aureus and 3 Escherichia coli strains. The effect was less in 1 strain of Proteus mirabilis. Whereas the activity in serum and bile was higher than in nutrient broth, the bactericidal activity in urine was less pronounced. The combination of cefazedone and gentamicin proved to be bactericidal in low concentrations.


Investigations on the effectiveness of cefazedone in experimental E. coli pyelonephritis.

The therapeutic effectiveness of (6R,7R)-7-(2-[3,5-dichloro-4-oxo-1(4H)-pyridyl]-acetamido)-3-([(5-methyl-1,3,4-thiadiazol-2-yl)-thio]methyl)-8-oxo-5-thia-1-azabicyclo[4,2,0]oct-2-ene-2-carboxylic acid (cefazedone, Refosporen), cephazolin, cephacetrile and cephalothin was compared in the test model of estradiol-induced E. coli pyelonephritis in the rat. Cefazedone and cephazolin were similar in effect. The relations between results of treatment and microbiological and pharmacokinetic characteristics of the cephalosporins tested are discussed.


Antibiotic activity of cefazedone in experimental pyelonephritis.

The new cephalosporin derivative (6R,7R)-7-(2-[3,5-dichloro-4-oxo-1(4H)-pyridyl]-acetamido)-3-([(5-methyl-1,3,4-thiadiazol-2-yl)-thio]methyl)-8-oxo-5-thia-1-azabicyclo[4,2,0]oct-2-ene-2-carboxylic acid (cefazedone, Refosporen) has been tested for antibacterial activity in the infection and therapy model of acute E. coli pyelonephritis in rats. The reference substance was cephalothin. The tests showed a superiority of cefazedone which, however, could not be clearly confirmed by the significance test. Owing to its favourable pharmacokinetics in combination with good antibacterial activity, cefazedone can be added to the cephalosporins considered for use in clinical practice, especially so as the statistics for 1977 show that the causative agents of pyelonephritis which these drugs can often combat effectively are present in the majority of the urine strains: E. coli accounted for 45%, enterococci for 18%, and Proteus for 15% out of a total number of 6100 urine strains.


New aspects of cell membrane radiobiology and their impact on radiation protection.

Recent developments in the field of the membrane biology of eukaryotic cells have revived relevant radiobiological studies. The supramolecular structures of membranes built up by the various chemical components that ensure certain functions and connections of the cells seem to react sensitively to ionizing radiations. Data concerning those radiation-induced alterations that reflect changes of such features and functions as surface charges, membrane-bound enzyme activities and receptor functions are reviewed. As the new concepts arising from the results obtained so far may have consequences in the theory and practice of radiation protection, a few areas are mentioned where the impact of the relevant accumulated knowledge might be expected to result in practical applications.


Evaluation of microhaemagglutination assay to determine treponemal antibodies in CSF.

The microhaemagglutination assay for Treponema pallidum antibodies (MHA:TP-CSF), the Veneral Disease Research Laboratory (VDRL-CSF), and the fluorescent treponemal antibody (FTA-CSF) tests were used to detect treponemal antibodies in 3157 samples of cerebrospinal fluid from patients with syphilis, with neurological, metabolic, or immunological disorders, and with undetermined diagnoses. An agreement of 99.5% between the results of the MHA:TP-CSF and FTA-CSF tests was obtained for all the CSF samples whereas that of only 95.5% was obtained for samples from the known case of syphilis. The MHA:TP-CSF test is simple, economical, rapid, and specific but not as sensitive as the FTA-CSF test.


False-positive test results for syphilis in relatives of a patient with systemic lupus erythematosus.

In a family of nine members, two had systemic lupus erythematosus and seven positive serological test results for syphilis. None of the affected subjects had a history or physical signs of syphilis, but two had positive results to the Treponema pallidum immobilisation test. The explanation for these findings is not known, but possibily they were all false-positive reactions.


[Research on surface antigens with immunofluorescence on enterobiopsies of patients with hepatitis B].

ABsAg surface antigen was detected, on enterobiopsies at duodenal level by means of Crosby capsule, in 12 subjects with various types of hepatitis B and in 2 control subjects, by means of immunofluorescence. The antigen was present in the duodenal mucosa of a certain number of subjects with hepatitis: this case-list included subjects with a tendency to relapse or with a poor chronic course notwithstanding the treatment. Some interpretative hypotheses are set forth.


Cultures from thymus and liver cells and blood lymphocytes for chromosome analysis in human abortions.

The authors propose a rapid method for chromosome studies in human abortions as a supplementary or alternative method to embryonic tissue cultures. Cultures were set up from fetal lymphocytes and thymus and liver cells. 72 hours was the longest time necessary to obtain suitable chromosome preparations. A very high rate of success was obtained in the 28 cases studied.


[Mother and child sharing the same room and breast feeding].

The author analyzes the existing relationship among breast feeding and socioeconomical level and degree of instruction of the mother. He also describes the causes for the interruption of breast feeding in a maternity where rooming-in existed. He stressed the importance of the motivation to the mother for breast feeding and the independence between levels of instruction and frequency of breast feeding.


[Levamisol in the treatment of recurrent urinary tract infections].

Levamisole, a widely used antihelminthic drug, possesses immunotropic properties. It has been proved that children with recurrent infections of the respiratory tract who were given the drug, were capable of preventing the repetition of the infections. Twenty-seven (27) children with recurrent urinary tract infections without structural malformations were studied and in all of them, a diminished cellular immunity was detected (blastic transformation of lymphocytes, rosettes of T lymphocytes and the skin tests). The drug was administered in doses of 2.5 mg/k/day, twice a week, for eight months. During the period of study, in 21 children repetition of the infections were not detected and in 13, a significant improvement of the cellular immunity was achieved. In some children it was possible to prove a rise of the IgA secretary in the urine.


A comparison of the effects of labetalol, bendrofluazide and their combination in hypertension.

1 Bendrofluazide (10 mg/day) or labetalol (300 mg and 600 mg/day) produced significant reductions in lying, standing, and post-exercise blood pressure in ten hypertensives. 2 Active treatments were approximately equivalent in anti-hypertensive effect. However, comparing lying and standing determinations, labetalol (600 mg/day) produced significantly greater additional postural falls in systolic blood pressure than during placebo or bendrofluazide treatment. 3 Systolic blood pressure rose after exercise during placebo or bendrofluazide treatment. However, on labetalol, mean changes in systolic blood pressure after exercise were negative and significantly different from those seen on placebo. 4 Greater reductions in lying and post-exercise systolic blood pressure were produced by combination treatment than by either individual drug. Additional postural and exercise-related falls in systolic blood pressure tended to be smaller with combination treatment than during treatment with labetalol alone. 5 Labetalol significantly reduced lying, standing, or post-exercise heart rate by comparison with placebo or bendrofluazide. 6 Labetalol significantly reduced lying, standing, or post-exercise heart rate by comparison with placebo or bendrofluazide. 6 Labetalol moderately reduced plasma renin activity, whereas bendrofluazide caused marked elevation. The effect of bendrofluazide predominated during combination treatment.


Clinical evaluation of a new antihypertensive vasodilating agent PR--G 138 Cl.

1 The cardiovascular effects of a new substance (PR--G 138) with vasodilating action were analysed in 12 patients with moderately severe essential hypertension on a 60 mEq sodium diet in a metabolic ward. To prevent tachycardia, propranolol 40 mg four times daily was given during the control period until blood pressure (BP) was stabilized, and continued throughout the study. 2 The compound was effective in every patient except one, who also was resistant to hydrallazine and diazoxide. Mean arterial pressure was lowered from a mean control value of 121 +/- 11 supine and 118 +/- 13 standing to 98 +/- 18 and 95+/- 15 mm Hg (P less than 0.001) respectively after a single oral dosage of 5 to 15 mg PR--G 138. The effect was maximal after 1--2 h and lasted up to 6 h. 3 With adequate dosage, there was no orthostatic reaction. Pulse rate and plasma renin activity did not rise during PR--G 138 treatment, and cardiac output increased only slightly, doubtlessly as a result of the propranolol therapy. In most patients 5 mg of the drug was sufficient to cause a drop of BP to normal levels. Exercise tolerance (bicycle ergometry) was constant or improved during drug action. One patient complained of headache, but no other side effects were seen provided that propranolol was taken when the drug was given. During 3 months treatments on an out-patient basis the effect was sustained in four of eight patients. No toxic effects have been noticed.


A single dose comparison of piretanide and bumetanide in congestive cardiac failure.

1 This study has compared the diuresis produced by a single oral administration of 6 mg piretanide, 9 mg piretanide and 1 mg bumetanide in a group of nine patients with cardiac failure using a balanced randomized design. 2 The natriuresis and kaliuresis produced in the first 6 h after administration of piretanide 9 mg and bumetanide 1 mg were similar. Piretanide 6 mg produced a lesser response. 3 There was evidence of sodium and water conservation following the diuresis for up to 48 h with all three treatments. 4 The patterns of urate and calcium excretion were similar for the two diuretics.


A human testicular teratoma serially transplanted in immune-deprived mice.

Serial transplantation of an HCG-producing human testicular teratoma in immune-deprived mice is described. The xenografted tumour was compared to the tumour of origin in histology, immunohistochemistry (using an immune peroxidase technique to localize HCG) autoradiography, marker production and growth rate. It is concluded that the xenograft retained the characteristics of the original tumour with the exception of a reduction in HCG-producing elements at transplantation beyond 5 serial passages.


Amyloid in basal cell carcinomas.

Stromal or intratumour deposits of amyloid were found in thirty of forty-six randomly selected basal cell carcinomas. Amyloid was found less often in histologically aggressive tumours than in other sub-types. Immunoglobulins, predominantly IgM, were demonstrated in all but one of these thirty cases, corresponding to the distribution of the amyloid deposits. Electron microscopy revealed 'early' amyloid in one case which showed numerous stromal IgM bodies on immunofluorescence but no evidence of amyloid by routine histochemical methods.


Identification of tertiary base pair resonances in the nuclear magnetic resonance spectra of transfer ribonucleic acid.

The low-field hydrogen-bond ring NH proton nuclear magnetic resonance (NMR) spectra of several transfer ribonucleic acids (tRNAs) related to yeast tRNAPhe have been examined in detail. Several resonances are sensitive to magnesium ion and temperature, suggesting that they are derived from tertiary base pairs. These same resonances cannot be attributed to cloverleaf base pairs as shown by experimental assignment and ring current shift calculation of the secondary base pair resonances. The crystal structure of yeast tRNAPhe reveals at least six tertiary base pairs involving ring NH hydrogen bonds, which we conclude are responsible for the extra resonances observed in the low-field NMR spectrum. In several tRNAs with the same tertiary folding potential and dihydrouridine helix sequence as yeast tRNAPhe, the extra resonances from tertiary base pairs are observed at the same position in the spectrum.


Nuclear magnetic resonance studies on transfer ribonucleic acid: assignment of AU tertiary resonances.

The hydrogen-bonded ring NH nuclear magnetic resonance (NMR) spectra of several transfer ribonucleic acid (RNA) species have been examined with particular emphasis on the extreme low-field portion. Betwen --13.8 and --15 ppm there are two extra resonances which are not derived from cloverleaf base pairs. A combined approach involving undermodified tRNAs, chemical modification, and hairpin fragment studies has assigned the T54--A58 resonance at --14.3 ppm in yeast tRNAPhe and Escherichia coli tRNA1 Val., the U8--A14 resonance has been assigned at --14.3 ppm, and the s4U8--A14 resonance in bacterial tRNAs has been assigned at --14.9 ppm. The T54--A58 resonance shifts between --14.3. and --13.8 ppm depending on the surrounding nucleotide sequence in the ribothymidine loop.


Paramagnetic ion effects on the nuclear magnetic resonance spectrum of transfer ribonucleic acid: assignment of the 15--48 tertiary resonance.

We have studied the effects of Co2+ and Mn2+ ions on the low-field nuclear magnetic resonance (NMR) spectra of pure class 1 transfer ribonucleic acid (tRNA) species. With 1.2 mM tRNA in the presence of 15 mM MgCl2 discrete paramagnetic effects were observed for Co2+ at concentrations in the range 0.02--0.1 mM and for Mn2+ in the range 0.002--0.01 mM, indicating fast exchange of these cations with tRNA. Both of these cations paramagnetically relax the s4U8--A14 resonance as well as other resonances from proximal base pairs. The Co2+ site appears to be the same site on G15 which was observed crystallographically [Jack, A., Ladner, J. E., Rhodes, D., Brown, R. S., & Klug, A. (1977) J. Mol. Biol. 111, 315-328]; the initially occupied tight Mn2+ site is the cation site involving the phosphate of U8. There are three base pairs within 10 A of both sites, namely, G15--C48, A14--s4U8, and C13--G22; this has led to the assignment of the G15--C48 and C13--G22 resonances in the NMR spectrum [Jack, A., Ladner, J. E., Rhodes, D., Brown, R. S., & Klug, A. (1977) J. Mol. Biol. 111, 315--328; Holbrook, S. R., Sussman, J. L., Warrant, R. W., Church, G. M., & Kim, Sung-Hou (1977) Nucleic Acids Res. 4, 2811--2820; Quigley, G. J., Teeter, M. M., & Rich, A. (1978) Proc. Natl. Acad. Sci. U.S.A. 75, 64--68].


Nuclear magnetic resonance studies on the tertiary folding of transfer ribonucleic acid: assignment of the 7-methylguanosine resonance.

Analysis of the low-field nuclear magnetic resonance (NMR) spectra of several class 1 D4V5 transfer ribonucleic acid (tRNA) species containing 7-methylguanosine in their variable loops reveals a set of six to seven tertiary base pair resonances, one of which is always located at ca. --13.4 ppm. Other tRNA species which do not contain 7-methyl-guanosine do not contain the tertiary resonance at --13.4 ppm. Chemical removal of 7-methylguanosine from several tRNAs containing the same dihydrouridine (DHU) helix sequence as yeast tRNAPhe results in the loss of the --13.4-ppm tertiary resonance. In the initiator methionine tRNA, which contains a different DHU helix sequence, the 7-methylguanosine hydrogen bond has been assigned at --14.55 ppm by chemical removal of this residue. In these experiments the aromatic C8H proton of 7-methylguanosine was also assigned (--9.1 ppm). The unexpectedly low-field position of the 7-methylguanosine resonance is explained by the deshielding effect of the delocalized positive charge in this nucleoside.


In vivo studies on the incorporation of microinjected acidic proteins of the large ribosomal subunit from Escherichia coli and artermia salina into oocyte ribosomes from Xenopus laevis.

Tritium-labeled acidic proteins from the large ribosomal subunit of Artermia salina or Escherichia coli were microinjected into the cytoplasm of stage IV/V oocytes from Xenopus laevis. eL12 from the large ribosomal subunit of A. salina but not L7/L12 or L7/L12--L10 from E. coli is specifically incorporated into 60S ribosomal subunits of oocytes. This incorporation is not significantly inhibited by actinomycin D. Incorporation of eL12 into the 60S subunits occurs in enucleated oocytes, suggesting that active ribosomal ribonucleic acid synthesis and ribosome assembly as well are not prerequired for this reaction. Apparently the incorporation proceeds via an exchange reaction between a free cytoplasmic pool of eL12 and ribosomal eL12.


Peptidyl transfer ribonucleic acid hydrolase activity of proteinase k.

Proteinase k, a seryl-protease obtained from Tritirachium album, is able to specifically hydrolyze N-blocked aminoacyl transfer ribonucleic acids (tRNAs). The blocked amino acid is released, and the tRNA molecule remains able to be recharged by its cogante amino acid. Aminoacyl-tRNAs are highly resistant to hydrolysis by the protease. This activity is not due to contamination of the protease preparation. A commercial protease from Streptomyces griseus displayed a similar activity, while trypsin, chymotrypsin, and papain unspecifically hydrolyzed all charged tRNAs tested. The characteristics of the hydrolysis performed by proteinase k closely resemble the peptidyl-tRNA hydrolase activity described in different cells as a scavenger for the peptidyl-tRNA that eventually falls from the polysomes. Out results warn about a hasty identification of any N-blocked aminoacyl-tRNA hydrolase activity in the cytoplasm as an independent peptidyl-tRNA hydrolase.


A single amino acid substitution in a histidine-transport protein drastically alters its mobility in sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Mutation hisJ5625 has altered the histidine-binding protein J of Salmonella typhimurium such that histidine transport is impaired, even though binding of histidine by the J protein is unimpaired [Kustu, S.G., & Ames, G.F. (1974) J. Biol. Chem. 249, 6976--6983]. We have determined by protein analytical methods that the only effect of this mutation has been the substitution of a cysteine residue for an arginine at a site in the interior of the polypeptide chain. This arginine residue is therefore potentially essential for the transport function of the protein. The mutant protein migrates in sodium dodecyl sulfate-polyacrylamide gel electrophoresis more slowly than the wild type protein, as if its molecular weight were greater by as much as 2000. Since this behavior is apparently due to a single amino acid replacement, a molecular weight difference even between two closely related proteins should not be inferred solely on the basis of sodium dodecyl sulfate gel electrophoresis.


Thallous ion is accumulated by potassium transport systems in Escherichia coli.

The accumulation of 204T1+ by Escherichia coli occurs primarily via either of two K+ transport systems called Kdp and TrkA. T1+ influx is inhibited and T1+ efflux is stimulated by the addition of K+ to the assay medium. Mutants defective in both the Kdp and TrkA systems accumulate little T1+. Uptake of triphenylmethylphosphonium, a lipid-soluble cation whose distribution is widely used to estimate the membrane electrical potential in bacteria, occurs to about the same extent in mutants that accumulate little T1+ as in strains that accumulate T1+ to high levels. These findings indicate that T1+ may be useful as a probe of bacterial K+ transport systems but is not a reliable indicator of the membrane electrical potential in E. coli.


Isolation and characterization of fourteen ribosomal proteins from small subunits of yeast.

A method for preparation of a large amount of ribosomal subunits from Saccharomyces cerevisiae by a Ti-15 zonal rotor is described. The proteins of the small subunits (ca. 50 000 A260 units) were separated into 22 fractions by chromatography on carboxymethylcellulose columns. Fourteen proteins were then purified from the ten chromatographic fractions by filtration through Sephadex G-100 or Sephacryl S--200. The isolated proteins are YP 6, YP 7, YP 9, YP 12, YP 14', YP 14'', YP 28, YP 38, YP 45, YP 50, YP 52, YP 58, YP 63, and YP 70. The molecular weight and amino acid compositions of these proteins are presented.


Internal motions in myosin.

High-resolution proton nuclear magnetic resonance (1H NMR) measurements were made on myosin, heavy meromyosin (HMM), myosin subfragment 1 (S1), light meromyosin (LMM), and actin. A strong signal from amino acid side chains undergoing motions too fast to be accounted for by simple rotations of groups on a rigid backbone was obtained from myosin. Comparison of myosin, HMM, S1, and LMM showed that the mobile region is located almost entirely in S1 and accounts for approximately 22% of its structure. Adenosine triphosphate (ATP) and ATP analogues had no measurable effect on the S1 spectrum. Actin, on the other hand, quenched the internal motions of S1. When S1 was titrated with actin, an association was obtained which was in agreement with other measured values. The actin effect was reversed by adding magnesium pyrophosphate (MgPPi) or adenyl-5'-yl imidophosphate (MgAMPPNP). Quantitative treatment of the broad signals from myosin and its subfragments substantiated the existence of two flexible regions in myosin. The highly mobile portion of myosin may be located in the "swivel" between S1 and the rest of myosin or in the actin binding site or in both. These possibilites are discussed, and a new possible mechanism for muscle cross bridge elasticity is proposed.


Covalent cross-linking of transfer ribonucleic acid to the ribosomal P site. Mechanism and site of reaction in transfer ribonucleic acid.

The covalent cross-linking of unmodified Escherichia coli N-acetylvalyl-tRNA to the 16S RNA of Escherichia coli ribosomes upon near-UV irradiation previously reported by us [Schwartz, I., & Ofengand, J. (1978) Biochemistry 17, 2524--2530] has been studied further. Up to 70% of the unmodified tRNA, nonenzymatically bound to tight-couple ribosomes at 7 mM Mg2+, could be cross-linked by 310--335-nm light. Covalent attachment was solely to the 16S RNA. It was dependent upon both irradiation and the presence of mRNA but was unaffected by the presence or absence of 4-thiouridine in the tRNA. The kinetics of cross-linking showed single-hit behavior. Twofold more cross-linking was obtained w-th tight-couple ribosomes than with salt-washed particles. Puromycin treatment after irradiation released the bound N-acetyl[3H]valine, demonstrating that the tRNA was covalently bound at the P site and that irradiation and covalent linking did not affect the peptidyl transferase reaction. Cross-linking was unaffected by the presence of O2, argon, ascorbate (1 mM), or mercaptoethanol (10 mM). Prephotolysis of a mixture of tRNA and ribosomes in the absence of puly(U2,G) did not block subsequent cross-linking in its presence nor did it generate any long-lived chemically reactive species. There was a strong tRNA specificity. E. coli tRNA1Val and tRNA1Ser and Bacillus subtilis tRNAVal and tRNAThr could be cross-linked, but E. coli tRNA2Val, 5-fluorouracil-substituted tRNA1Val, tRNAPhe, or tRNAFMet could not. By sequence comparison of the reactive and nonreactive tRNAs, the site of attachment in the tRNA was deduced to be the 5'-anticodon base, cmo5U, or ,o5U in all of the reactive tRNAs. The attachment site in 16S RNA is described in the accompanying paper [Zimmerman, R. A., Gates, S. M., Schwartz, I., & Ofengand, J. (1979) Biochemistry (following paper in this issue)]. The link between tRNA and 16S RNA is either direct or involves mRNA bases at most two nucleotides apart since use of the trinucleotide GpUpU in place of poly(U2,G) to direct the binding and cross-linking of N-acetylvalyl-tRNA to the P site did not affect either the rate or yield of cross-linking. Both B. subtilis tRNAVal (mo5U) and E. coli tRNA1Val (cmo5U) gave the same rate and yield of cross-linking when directed by the trinucleotide GpUpU. Therefore, the presence of the charged carboxyl group in the cmo5U-containing tRNA apparently does not markedly perturb the orientation of this base with respect to its reaction partner in the 16S RNA. The cross-linking of AcVal-tRNA only takes place from the P site. At 75 mM KCl and 75 mM NH4Cl, less than 0.4% cross-linking was found at the A site, while 55.5% was obtained at the P site. However, when the salt concentration was lowered to 50 mM NH4Cl, 5% cross-linking to the A site was detected, compared to 49% at the P site. Thus, a simple change in the ionic strength of the incubation mixture was able to alter the affinity labeling pattern of the ribosome.


Circular dichroism anisotrophy of DNA with different modifications at N7 of guanine.

The complexex DNA-Ag1+, DNA-Cu1+, protonated DNA and DNA methylated at N7 of guanine were oriented by pumping the solutions through a multicapillary cell in the direction of a light beam. The CD components along the DNA axis, delta epsilon parallel, and normal to it, 2 delta epsilon perpendicular, were calculated from the CD spectra of the oriented samples by the method of Chung and Holzwarth, (1975) J. Mol. Biol. 92, 449--466. It was shown that in most cases, except that of the protonated DNA, the degree of orientation was only slightly less than that for pure DNA. This demonstrated the absence of aggregation and of appreciable denaturation. In all cases the modifications of DNA give rise to a negative component 2 delta epsilon perpendicular, whose magnitude increased as the extent of modification increased. From both the CD spectra of non-oriented samples and the absorption spectra, an inference is drawn that Ag1+ and Cu1+ are attached to the same site as CH3 groups i.e., to the N7 atom of guanine. Proton transfer along the H-bond from the N1 atom of G to the N3 atom of the complementary cytosine is suggested to be a result of the modifications, although the case of H+-DNA may differ from the others. Based on the CD spectra for the anisotropic components, delta epsilon parallel and 2 delta epsilon perpendicular, it is proposed that ligand binding is accompanied by winding of the DNA helix.


Optimization of conditions for the in vitro formation of hybrid DNA molecules by DNA ligase.

Conditions for the ligation with T4 induced DNA ligase of two DNA molecules via their complementary sticky ends have been established which lead preferentially to the formation of hybrid molecules. This is demonstrated with two combinations of parent molecules varying greatly in their relative molecular weights. In one case the intact hybrid molecule could be directly isolated. In addition a DNA dependent quantitative electrophoretic assay for DNA ligase activity is described which does not need a radioactively labeled substrate. The ligation procedure has been shown to be useful in molecular cloning experiments.


Action of thiazolidine-2-carboxylic acid, a proline analog, on protein synthesizing systems.

Thiazolidine-2-carboxylic acid, or beta-thiaproline, is a proline analog in which the beta methylene group of proline is substituted by a sulfur atom. It has been deomonstrated that beta-thiaproline is activated and transferred to tRNAPro by Escherichia coli and rat liver aminoacyl-tRNA synthetases, and inhibits proline incorporation into polypeptides in protein synthesizing systems from E. coli, rat liver or rabbit reticulocytes. In mammalian systems beta-thiaproline inhibits also leucine incorporation; in rabbit reticulocyte lysate it inhibits ribosome run-off. Both these effects may be explained by the fact that beta-thiaproline once incorporated into the growing polypeptide chain impairs its further elongation, as shown by experiments made with puromycin. All tests were performed in comparison with thiazolidine-4-carboxylic acid, or gamma-thiaproline, another proline analog having the gamma methylene group substituted by a sulfur atom; it was shown that in all the reactions studied both compounds act as competitive inhibitors of proline. Some differences in the effects of the two analogs have been evidenced: in almost all the reactions and mainly in the whole protein synthesizing systems, beta-thiaproline shows an higher inhibitory activity.


Enzyme-enzyme interaction and the biosynthesis of aromatic amino acids in Escherichia coli.

The technique of affinity chromatography has been used to demonstrate that enzymes involved in the biosynthesis of tyrosine and phenylalanine in Escherichia coli undergo reversible interactions. Thus it has been shown that the aromatic amino acid aminotransferase (aromatic-amino-acid: 2-oxoglutarate amino-transferase, EC 2.6.1.57) reacts specifically with chorismate mutaseprephenate dehydrogenase (chorismate pyruvate mutase, EC 5.4.99.5 and prephenate: NAD+ oxidoreductase (decarboxylating), EC 1.3.1.12) in the absence of reactants and with chorimate mutase-prephenatedehydratase (prephenate hydro-lyase (decarboxylating), EC 4.2.1.51) in the presence of phyenylpyruvate. Tyrosine causes dissociation of the aminotransferase: mutasedehydrogenase complex while dissociation of the aminotransferase-mutasedehydratase complex occurs on omission of phenylpyruvate. Only the active form of chorismate mutase-prephenate dehydrogenase participates in complex formation.


Isolation and characterization of porcine beta-casein.

Porcine beta-casein was isolated by chromatography on DEAE-cellulose. The protein had a molecular weight of 24 900 as determined by gel filtration on Sephadex G-100 in guanidine-HCl. Its amino acid composition differed from bovine beta-casein especia-ly in respect to serine, alanine and leucine. In common with bovine beta-casein the N-terminal amino acid was arginine; the C-terminal was either alanine or valine, while the C-terminal of bovine beta-casein is valine. At any temperature porcine beta-casein was more sensitive to Ca2+ than bovine beta-casein, while at a fixed Ca2+ concentration porcine beta-casein aggregated at a lower temperature than bovine beta-casein. Porcine beta-casein was susceptible to hydrolysis by calf chymosin but the proteolytic specificity differed from that of calf chymosin on bovine beta-casein.


Quaternary structure of DNA-dependent RNA polymerase from Escherichia coli. Measurement of distances by fluorescence energy transfer.

Distances between the subunits in Escherichia coli RNA polymerase (core and holo enzyme) and the rifamycin binding site have been determined using the nonradiative energy transfer technique. The appropriate donor and acceptor labels have been chosen in order to optimize the spectral overlap and maximize the energy transfer. Spacer linked derivatives of rifamycin SV possessing nitrobenzo-oxadiazole groups (energy acceptor) were synthesized for this purpose. The donor label, acetylaminoethylaminonaphthalene sulfonate, was introduced into the intact enzyme, and the subunits were separated. Enzyme molecules selectively labelled on one kind of subunit were produced by mixed reconstitution techniques employing labelled and non labelled subunits. The labelled beta'-subunit could not be prepared in sufficient amounts. Energy transfer distances between the enzyme-bound rifamycin derivative and the subunits were determined to be approximately 5.9 nm for sigma, 7.2 nm for alpha 2 and 6.1 nm for beta.


Shape of proteins S3 and S17 from the small subunit of Escherichia coli ribosome.

The shapes of proteins S3 and S17 purified from the 30 S subunit of Escherichia coli A19 were studied by hydrodynamic methods. The proteins have s020,w values of 2.1 +/- 0.1 S and 1.2 +/- 0.1 S and D020,w values of 7.4 +/- 0.5 . 10(-7) cm2/s and 11.4 +/- 0.6 . 10(-7) cm2/s. The respective molecular weights determined by sedimentation equilibrium are 25 800 +/- 500 and 9900 +/- 300. The intrinsic viscosity values for the two proteins are 5.8 +/- 0.3 ml/g and 4.2 +/- 0.2 ml/g. From these hydrodynamic parameters a slightly elongated shape for S3 and a globular shape for S17 have been concluded.


Stimulation of dehydrogenase synthesis by cadmium in a cadmium-resistant strain of Saccharomyces cerevisiae.

The activity of dehydrogenase in Saccharomyces cerevisiae was estimated by reduction of 2,3,5-triphenyltetrazolium chloride. By the adaptation of yeast to cadmium, the high activity of dehydrogenase was observed. Furthermore, the activity of dehydrogenase in Cd-resistant cells was increased by growing in medium containing CdSO4. However, the activity of dehydrogenase was inhibited by the addition of CdSO4 to the reaction mixture. The activity of dehydrogenase in Cd-sensitive cells was increased slightly by incubation with low concentrations of CdSO4. High activity of dehydrogenase in Cd-resistant cells was completely negated by the addition of cycloheximide to the incubation medium. The increase of dehydrogenase activity is due partly to de novo synthesis of protein.


The primary structure of ribosomal protein S7 from E. coli strains K and B.

Ribosomal proteins S7 from 30S subunits of Escherichia coli strains K and B differ extensively in their aminoacid compositions. The experimental details which led to the determination of the complete primary structures of proteins S7K and S7B are presented. Protein S7K consists of a single polypeptide chain of 177 aminoacids giving a calculated molecular weight of 19, 732, whereas protein S7B has 153 residues which amount to a molecular weight of 17,131. Aminoacid sequences were determined by a combination of automated Edman degradation of the intact proteins in a modified Beckman sequenator and sequencing of peptides obtained by digestion with trypsin. Staphylococcus aureus protease, thermolysin and pepsin, either by solid-phase Edman degradation or by dansyl-Edman degradation. Additional information about the primary structure was derived from peptides resulting from chemical cleavages of the protein by 2-(2-nitrophenyl-sulphenyl)-3-methyl 3' bromoindolenine at its tryptophanyl bonds and by cyanogen bromide at its methionyl bonds leading to large fragments. The mutational event occurring between S7B and S7K was characterized. Protein S7K contains an additional sequence of 24 aminoacids at its C-terminal end. The aminoacid sequence of both proteins S7K and S7B was compared to the published sequences of the other ribosomal proteins of Escherichia coli and predictions for the secondary structure of these proteins were made.


[Dehydrogenases reducing NAD+ in the presence of D (-) 3- phosphoglycerate in mycobacteria (BCG, H37Ra, M. phlei)].

Referring to the elution volume on a Sephadex G-150 column only one specific peak is obtained, the same for the BCG, H37Ra and Mycobacterium phlei strains grown on Sauton synthetic medium. Some properties of these partially purified dehydrogenases are studied (conservation and dialysis in media of different salt concentrations, equilibrium constant, Km, heat stability). All enzyme preparations from tubercle bacilli (BCG, H37Ra) are readily inactivated by heat and are very unstable in solutions of low ionic strength. In contrast, under the same experimental conditions, all enzyme samples from M. phlei are, comparatively, much more stable towards these factors [heat, salt (potassium phosphate, NaCl) concentration].