Coordinate control of collagen synthesis and cell growth in chick embryo fibroblasts and the effect of viral transformation on collagen synthesis. Using collagenase digestion as an assay for collagen in partially synchronized secondary cultures of chick embryo fibroblasts, we find that the rate of collagen synthesis remains at a constant fraction of overall protein synthesis (5%) regardless of the growth rate of the cells even when the rate of protein synthesis is accelerated 5-fold by adding serum and altering the pH of the culture medium. However, in cells oncogenically transformed by Rous sarcoma virus, the relative rate of collagen synthesis was decreased by 50% 24 hours after infection and was 10% of the initial rate after 5 days. This selective decrease in rate of collagen synthesis could be reversed in cells infected with an RSV temperature-sensitive transformation-defective mutant at the non-permissive temperature, indicating that the decrease in the rate of collagen synthesis was not merely the result of viral infection but was a direct consequence of oncogenic transformation.


Effects of sugars on melanogenesis in cultured melanoma cells. A permanent cell line C2M of mouse melanoma B16 was highly melanized in a modified Eagle's MEM supplemented with 10% calf serum, when the medium contained 1 mM galactose and 10 mM pyruvate instead of 5.5 mM glucose. The activity of the key anzyme for melanogenesis, tyrosinase (EC 1.14.18.1), of living cells cultured in the galactose-pyruvate medium was consistently 27 times higher than that of cells in normal MEM. This high level of tyrosinase activity was maintained in the stationary phase, in contrast to the activity of cells in normal medium, which decreased sharply in the stationary phase. It seems likely that tyrosinase activity is suppressed by the presence of glucose rather than stimulated by galactose. This modified medium should be useful obtaining a high level of tyrosinase activity in living cells in culture and in cell-free extracts.


The self-assembly of synthetic filaments of myosin isolated from Chaos carolinensis and Amoeba proteus. Synthetic myosin thick filaments were formed from preparations of electrophoretically homogeneous myosin isolated from Chaos carolinensis and Amoeba proteus when dialysed to physiological ionic strength and pH. Myosin dialysed directly against low ionic strength buffers formed native-like thick filaments in the presence and absence of exogenous divalent cations. The average dimensions of the synthetic filaments grown under these conditions were 455 nm long and 16 nm wide with a distinct bare central zone 174 nm long. Myosin predialysed against EGTA-EDTA solutions at high ionic strength and then dialysed to low ionic strength formed native-like filaments only in the presence of 1mM Mg2+. 1 mM Ca2+ could not be substituted for Mg2+ under these conditions to achieve native-like filaments. Filaments grown from predialysed myosin in the absence of Mg2+ resembled EGTA-dissociated myosin filaments observed in EGTA-treated cytoplasm and were highly branched, poorly formed filaments lacking a distinct bare central zone. The average dimensions of the filaments grown from predialysed myosin in the absence of Mg2+ were 328 nm long, 13 nm wide with a bare central zone 111 nm long. Under the conditions tested, myosin isolated from these amoebae did not demonstrate a divalent cation requirement for thick filament formation. The results obtained with myosin isolated from the 2 organisms were identical.


[Identification and quantitation of impurities from benorilate (Salipran) by high-performance liquid chromatography (author's transl)]. High-performance liquid chromatography is used for identification and quantitation of impurities which may be encountered in a new antalgic, benorilate (or Salipran), an ester of aspirin with paracetamol. Gradient elution is carried out using a stationary phase consisting of porous 10-micron silica beads bonded to alkylnitrile (Micropak CN), and a mixture of hexane-methylenechloride-methanol-acetic acid with varying methanol percentage as mobile phase. The following impurities were separated from benorilate: acetylsalicylic anhydride, aspirin, acetylsalicylsalicylic acid, salophene, amino-4-phenylacetoxy-2-benzoate, paracetamol, p-aminophenol. The repeatability of the quantitative analysis is good with a standard variation of 0.54% for benorilate (7 injections). Detection is by UV absorption at 254 nm, and detectability is between 2-10(-9) moles for p-aminophenol and 4-10(-11) moles for salophene.


Purification of alpha-L-fucosidase from various sources by affinity chromatography. An affinity column for alpha-L-fucosidases was constructed by linking p-amino-phenyl 1-thio-alpha-L-fucopyranoside to Sepharose 4B through linkers of succinyl 3,3'-diamino-dipropylamine. Excellent purification of alpha-L-fucosidase from rat epididymis, Clostridium perfringens and Limulus polyphemus (horse shoecrab) could be effected inone step with good yield. An affinity column purification step can be introduced at any point in published purification procedures. The purified enzyme is essentially free of other glycosidases and proteolytic enzymes. The column material is stable and can be reused for at least two years.


Nickel gas chromatographic columns: an alternative to glass for biological samples. Nickel tubing may be substituted for glass in the fabrication of gas chromatographic columns for use with samples of biological interest. Comparisons of separations of mixtures of steroids, narcotic alkaloids, phenothiazines, and amphetamines on stainless stell, glass, and nickel packed columns showed little or no observable sample decomposition on glass or nickel as contrasted to complete loss of certain compounds on stainless steel. The nickel columns are easily prepared, durable, economical, and not subject  to breakage.


Analysis of water soluble vitamins by high pressure liquid chromatography. Resolution of the water-soluble vitamins--pyridoxin, riboflavin, niacinamide, vitamin B12, thiamin, ascorbic acid, niacin and folic acid--by high pressure liquid chromatography was examined on two bonded-phase columns, muBondapak C18 and muBondapak NH2. The effect on the retention times of individual vitamins and the separation of a multivitamin sample was determined using varying proportions of water/methanol as the eluting solvent and by addition of various salts, buffer solutions and PIC reagents to the water/methanol. Each vitamin was able to be eluted satisfactorily from muBondapak C18. It was found that seven vitamins could be resolved from a multivitamin mixture in a single analysis in several solvent systems with the total time for the analyses being always less than 40 min. With muBondapak NH2, all the vitamins except folic acid were eluted and six vitamins could be resolved from a mixture in a single analysis. The speed of analysis was greater with muBondapak NH2 with all compounds eluted in 15 min and the peaks were sharper. The order of elution was essentially the reverse of that obtained with muBondapak C18.


Isolation of an obligately anaerobic Streptococcus pneumoniae from blood culture. An obligately anaerobic strain of Streptococcus pneumoniae was isolated from blood culture in a 14-month-old child with an upper respiratory tract infection.


Bicarbonate transport by rabbit cortical collecting tubules. Effect of acid and alkali loads in vivo on transport in vitro. Rabbit cortical collecting tubules were perfused in vitro to investigate the control of bicarbonate transport. Bicarbonate was measured by microcalorimetry as total CO2. The perfusate and bath were identical solutions containing 25 mM bicarbonate at pH 7.4. The mean pH of the urine in the bladders of untreated rabbits at the time they were killed was 7.4. Their individual tubules, studied in vitro, either absorbed or secreted bicarbonate, and, combining the results, there was on the average no significant net transport. When the rabbits were treated with NH4Cl the day before the experiment, their urine was acidic and their tubules studied in vitro absorbed bicarbonate (i.e., there was net lumen-to-bath transport). In contrast, when the rabbits were treated with NaHCO3, their urine was significantly more alkaline, and their tubules studied in vitro generally secreted bicarbonate (i.e., net bath-to-lumen transport). Thus, the direction of bicarbonate transport by cortical collecting tubules studied under standard conditions in vitro correlated with the urine pH and was determined by the preceding treatment of the animals in vivo with acidifying or alkalinizing salts. These results demonstrate a previously unrecognized mechanism which contributes to the control of urinary bicarbonate excretion.


Relationship between phosphaluria and acute hypercapnia in the rat. Standard clearance studies were performed in mechanically ventilated intact and acutely thyroparathyroidectomized (TPTX) rats to document and characterize the effect of hypercapnia (HC) on urinary phosphorus excretion (U(P)V). HC as compared to normocapnia (NC) was associated with an increase in U(P)V in intact (62.5 vs. 7.93 mug/min) and TPTX (30.5 vs. 0.59 mug/min) rats, an increase in filtered load of phosphorus in intact (218 vs. 191 mug/min) and TPTX (243 vs. 146 mug/min) rats, an increase in blood bicarbonate concentration in intact (27.8 vs. 26.0 meq/liter) and TPTX (24.5 vs. 22.3 meq/liter) animals, and a decrease in blood pH in intact (7.15 vs. 7.42) and TPTX (7.07 vs. 7.39) rats. Additional TPTX rats with NC and HC were studied during phosphorus infusion at a comparable filtered load of phosphorus (NC = 307 mug/min and HC = 328 mug/min). U(P)V was 18.5 mug/min in NC and 85.2 mug/min in HC animals. Intact NC animals infused with NaHCO(3) achieved a blood bicarbonate of 45.9 meq/liter compared to 26.0 meq/liter in intact NC NaCl-infused rats. U(P)V was 10.0 mug/min in the NaHCO(3) and 7.93 mug/min in NaCl-infused animals. In intact HC animals infused with NaHCO(3), blood pH was 7.36 compared to 7.42 in NC intact NaCl-infused animals. U(P)V was 83.2 mug/min in the HC bicarbonate-infused and 7.93 mug/min in the NC NaCl-infused rats. These experiments demonstrate that elevated blood carbon dioxide tension per se increases U(P)V. Increases in filtered load of phosphorus and blood bicarbonate which are associated with HC contribute to the phosphaturia as does parathyroid hormone. The phosphaturia is not dependent upon reduction of extracellular pH.


Interactions among heparin, cold-insoluble globulin, and fibrinogen in formation of the heparin-precipitable fraction of plasma. Fibrinogen and the cold-insoluble globulin of plasma (CIg) are the main protein components of the heparin-precipitable fraction of normal plasma. The interactions among these proteins and heparin were examined. Heparin formed a cold-precipitable complex with purified CIg or with mixtures of CIg and fibrinogen but not with purified fibrinogen alone. Cryoprecipitation was augmented by addition of Ca(++) or by selection of optimal heparin levels; it was reduced or even abolished by raising the ionic strength or pH or both, or by raising the heparin concentration above that for maximum precipitation of CIg. Fibrinogen reduced the threshold for heparin-induced CIg cryoprecipitation and, by coprecipitating with heparin and CIg, increased the amount of precipitate that formed. In contrast to the heparin-precipitable fraction of normal plasma which contained both fibrinogen and CIg, that from a patient with congenital afibrinogenemia contained CIg but lacked fibrinogen. Normal plasma depleted of CIg by immunoabsorption failed to form a heparin-induced cryoprecipitate. Thus, CIg is essential for heparin-induced cryoprecipitation to occur. Fibrinogen, as assessed by chromatographic experiments with heparin-Sepharose columns, had a considerably lower binding affinity for heparin than did CIg, suggesting that it participates in precipitate formation mainly, if not entirely, by virtue of its affinity for CIg. The region of the fibrinogen molecule accounting for its precipitation with CIg appears to be localized in the carboxy-terminal segment of the Aalpha-chain; fibrinogen subfractions lacking this region failed to augment cryoprecipitation of heparin-CIg mixtures and, even though such species were present in normal plasma, they failed to coprecipitate in the heparin-induced complex.


Effects of hypermetabolism on ventilation and chemosensitivity. Muscular exercise is associated with hypermetabolism and increased hypoxic ventilatory response (HVR). In order to dissociate mechanical and metabolic factors, the effect of hypermetabolism on hypoxic ventilatory response was evaluated at rest. Carbohydrate and protein feeding increases metabolic rate, and their effects on chemosensitivity, ventilation, and blood pH were evaluated in six normal subjects 2 h and 3 h after calorically equal test meals (1,000 cal). After carbohydrate, base-line oxygen consumption (Vo(2)) increased from 237+/-11.3 ml/min (SEM) to 302+/-19.4 (P < 0.001) and 303+/-18.5 (P < 0.001) at 2 h and 3 h, respectively. Hypoxic ventilatory response, measured as shape parameter A, increased from a control of 144+/-11.8 to 330+/-61.0 (P < 0.01) at 2 h and 286+/-57.0 (P < 0.05) at 3 h. These changes were associated with a mild metabolic acidosis as pH decreased from a control of 7.402+/-0.004 to 7.371+/-0.009 (P < 0.005) at 2 h and 7.377+/-0.008 (P < 0.005) at 3 h. After protein, Vo(2) increased from 241+/-6.7 to 265+/-6.2 (P < 0.02) and 270+/-5.4 (P < 0.001), an overall increase less than that which occurred after carbohydrate (P < 0.01). Hypoxic ventilatory response increased from 105+/-14.5 to 198+/-24.3 (P < 0.02) at 2 h and 219+/-17.3 (P < 0.01) at 3 h, which was not different from the increase with carbohydrate. After protein, no acidosis occurred. Thus, after protein, HVR increased despite the absence of a systemic acidosis. We conclude that both carbohydrate and protein feedings are associated with resting hypermetabolism and increased HVR compared with the fasting state. For both meals, increased metabolic rate was correlated with increased hypoxic response.


The influence of thyroid hormones on in vitro erythropoiesis. Mediation by a receptor with beta adrenergic properties. The erythropoietic effect of various thyroid hormones has been studied using erythroid colony formation by canine marrow cells. Although erythropoietin was required for colony growth, physiologic levels of thyroid hormones significantly enhanced colony numbers. The order of potency of the thyroid compounds in their in vitro erythropoietic effect parallels their known calorigenic potency in vivo, suggesting that the in vitro effect is physiologically relevant. A series of studies linked the mechanism of thyroid action to adrenergic receptors on responsive cells. Propranolol, a global beta-blocker, inhibited thyroid hormone-responsive erythroid colonies. When adrenergic antagonists having different blocking characteristics were added to culture, the thyroid hormone effect was blocked by those compounds having beta(2)-subspecificity. Velocity sedimentation analysis showed that the peak of colony-forming cells which respond to thyroid hormone and the adrenergic agonist, isoproterenol, sedimented at an identical rate (7.54 mm/h), which is slower than the major peak of colony-forming cells responding to erythropoietin alone (8.62 mm/h). These results demonstrate thyroid hormonal enhancement of in vitro erythroid colony growth which appears mediated by a receptor with beta(2)-adrenergic properties. The data suggest that changes in hormone-target cell interaction may occur during states of abnormal thyroid function.


Judgment of emotion among chronic schizophrenics. This study investigated accuracy of judgment as to posed facial expressions and nonverbal scenes of various emotions. Ss were 16 male and 16 female chronic schizophrenics and a normal control group of equal size and sex composition. The results revealed that normal Ss were significantly more accurate than schizophrenics in identifying emotions from both posed photographs of the face and nonverbal videotape scenes. S sex was not found to affect differentially the schizophrenic or normal Ss' response accuracy to both the photographs and videotapes. Further, it was observed that both groups' accuracy improved when given multiple-choice alternatives to select from as contrasted to their open-ended free responses; this was especially true for the schizophrenic group. Teaching patients to identify and practice expressing discrete emotions was suggested.


The Brief Outpatient Psychopathology Scale (BOPS). The pertinence of a rating scale capable of characterizing presenting psychopathology and measuring degrees of change for psychoneurotic patients treated in an outpatient setting is described. Data for 328 patients who represent 1191 rating profiles were used to develop the factor structure for the New Physician's Rating List, a rating scale completed by health professionals after Patient interviews. Findings were used to develop the new Brief Outpatient Psychopathology Scale. A proposed factor structure is discussed. The structures of both scales include an Anxiety Syndrome composed of Anxiety and Psychomotor Activation Factors. The scales also include Depression and Somatization factors.


Patients' expectancies and improvement in treatment: the shape of the link. It has been hypothesized (a) that patients' expectancies for therapeutic gain are linked to the clinical improvement that the patients realize in treatment; and (b) that patients' expectancies may play a causative role in such improvement. The first hypothesis has received empirical support, but the second has not. This study tested the causativeinterpretation of patients' expectancies and a second interpretation, which states that patients' expectancies predict, but do not cause clinical improvement. The shape of thelink between expectancy and improvement for hospitalized schizophrenics was explored. Based on motivation research it was reasoned that a curvilinear relationship between expectancy and improvement would support a causative interpretation and that a linear relationship would support a predictive interpretation of the nature of expectancy. Multiple regression analyses found a linear relationship between expectancies and objective measures of improvement for the patients, but no evidence of a curvilinear relationship between these measures. The results were interpreted as supporting a predictive interpretation of the expectancies of hospitalized schizophrenic patients.


Comparison of bumetanide and hydrochlorothiazide on renal potassium and hydrogen ion excretion. The purpose of this study was to compare the renal electrolyte excretion pattern of bumetanide with that of hydrochlorothiazide in dogs anesthetized with pentobarbital. In bumetanide-treated animals, mean sodium excretion rose to 12 per cent of the filtered load, while hydrochlorothiazide increased sodium excretion to 4 per cent of the filtered load. After bumetanide, urine pH fell from 6.1 to 5.1 and net hydrogenion excretion increased significantly. After hydrochlorothiazide, urinary pH went from 6.4 to 7.4, and there was no change in net hydrogen ion excretion. Potassium excretion rose to 106+/-22 muEq/min with bumetanide and to 99+/-17 muEq/min with hydrochlorothiazide. These changes in electrolyte excretion occurred despite lack of changes in arterial blood gases, arterial blood pressure, and glomerular filtration rate. In addition, bumetanide did not exert an inhibitory effect on potassium excretion under conditions of potassium loading. It is concluded that bumetanide produces a higher urinary Na+:K+ ratio with a lower pH than hydrochlorothiazide and that renal potassium ion excretion in response to sulfamoyl diuretics is not solely dependent on the rate of sodium excretion.


Twin crossover relative potency analgesic assays in man. II. Morphine vs. 8-methoxycyclazocine. Using the twin crossover, balanced incomplete block design described in the previous paper, a double-blind determination of the relative analgesic potency of graded intramuscular doses of Win 20,836 (8-methoxycyclazocine) and morphine was carried out in patients with postoperative pain. Although no preliminary data at all on the human analgesic activity of Win 20,836 were available, the sequential decision-making process designed to choose the doses of the test medication most closely equianalgesic with the standard functioned efficiently to establish doses of Win 20,836 that had analgesic activity. Unfortunately, the occurrence of psychotomimetic side effects prevented the administration of doses of Win 20,836 equieffective with the morphine standard, and this necessitated substantal extrapolation of the dose-response curve of the test drug to arrive at a relative potency estimate. However, our relative potency estimate, which indicated that Win 20,836 is three to six times as potent as morphine, was dependable enough to predict with reasonable certainty that doses of Win 20,836 equieffective to the usual doses of morphine would produce an unacceptable level of psychotomimetic side effects. Clinical investigation of the drug was therefore terminated.


Reactivity and aggression in the rat: induction by alpha-adrenergic blocking agents injected ventral to anterior septum but not into lateral septum. Intracranial injections were made bilaterally through permanently implanted cannulas ending in the lateral septum or in the region ventral to the anterior septum. The rats were first screened with injections of a local anesthetic, lidocaine, which blocks both synaptic and axonal conduction. Those animals that showed an increase in reactivity and aggression were then injected with a synaptic transmitter blocking agent. The results showed that transmitter blocking agents reproduced the effect of the local anesthetic only in the region ventral to the anterior septum and that alpha-adrenergic (phentolamine, tolazoline), but not beta-adrenergic (propranolol, hydralazine), cholinergic (atropine, hyocine), or dopaminergic (haloperidol) blocking agents were effective. These results suggest that synapses in the forebrain system controlling reactivity and aggression are alpha-asrenergic and are located in the region ventral to the anterior septum just lateral to the diagonal band of Broca. The septum itself may be involved only to the extent that it is traversed by fibers of passage.


Clinical pharmacokinetics of lorazepam. III. Intravenous injection. Preliminary results. Four healthy male volunteers received 5 mg lorazepam as a single intravenous injection. Concentrations of lorazepam and its glucuronide metabolite were determined in multiple venous blood samples drawn during the 48 hours after dosing and in all urine collected during 96 hours after the dose. Mean pharmacokinetic parameters for lorazepam were: apparent elimination half-life, 13.2 hours; volume of distribution, 0.84 liter/kg; total clearance, 55.3 ml/min. Lorazepam glucuronide, the major metabolic product of lorazepam, promptly appeared in blood, reached peak levels within 6 hours of the dose, then declined in parallel with the parent compound. A mean of 69 per cent of the dose was recovered in urine as lorazepam glucuronide.


Twin crossover relative potency analgesic assays in man. I. Morphine vs. morphine. Although the four-point relative potency assay using crossover design has proven a powerful technique for the clinical evaluation of analgesics in patients with chronic pain, excessive dropouts have made this design impractical in postoperative pain. In a relative potency assay comparing single graded intramuscular doses of morphine standard and morphine test in postoperative patients, we have managed to circumvent this difficulty while preserving many of the advantages of a complete crossover by using the "twin-crossover" balanced incomplete block design, which requires that each subject receive only two of the four possible treatments. The "twin crossover" design, coupled with a sequential decision-making process that expedites choosing the doses of the test medication which are most closely equianalgesic with the standard, yielded excellent analgesic assay sensitivity and made efficient use of our population of postoperative patients.


Evaluation of butaclamol in chronic schizophrenic patients. In a double-blind, placebo-controlled study, an attempt was made to evaluate butaclamol in chronic schizophrenic patients using chlorpromazine (CPZ) as the standard comparative drug. With doses up to 50 mg/day, butaclamol was shown to have significant antipsychotic activity comparable to CPZ but with a much higher incidence of extrapyramidal signs. A more reasonable maintenance dose may be in the range of 5 to 20 mg/day. Rebound insomnia was noted again with butaclamol, which warrants further study.


Metabolic and cardiovascular effects of carbuterol and metaproterenol. Metabolic and cardiovascular responses to selective beta-adrenergic bronchodilators, carbuterol and metaproterenol, were studied during an asymptomatic period in 8 male subjects with bronchial asthma diagnosed as mile to moderate. On separate days each individual received either placebo, carbuterol 2 mg, carbuterol 4 mg, or metaproterenol 20 mg orally in a double-blind fashion. Subsequently, metabolic and cardiovascular responses were measured periodically for 5 hr. Carbuterol 2 mg was indistinguishable from placebo except for small elevations of glucose at 3 and 4 hr. Carbuterol 4 mg produced significant increases in glucose, insulin, lactate, and free fatty acids as well as in pulse rate and arterial pulse pressure. Metaproterenol produced increases only in plasma glucose and insulin. The majority of patients reported drug-related side effects which were all mild, after taking either carbuterol 4 mg or metaproterenol 20 mg. Fewer subjective side effects were noted with carbuterol 2 mg. These findings indicate that a 2-mg dose of carbuterol can be administered to typical asthmatic subjects without significant subjective or objective side effects. The larger dose (4 mg) may be accompanied by a greater frequency of side effects.


Clinical pharmacokinetics of lorazepam. IV. Long-term oral administration. Fifteen healthy male volunteers received long-term daily treatment with oral lorazepam at doses as high as 10 mg per day for a period of 26 weeks. Steady-state plasma concentrations of lorazepam and its glucuronide metabolite were measured in all subjects at least every two weeks. At daily doses of 6 mg per day, the mean steady-state lorazepam level was 88 ng/ml and that of lorazepam glucuronide was 170 ng/ml. Mean levels among seven subjects who received 10 mg per day were 164 and 266 ng/ml, respectively. Lorazepam concentrations fluctuated from week to week despite constant dosage, but there was no evidence of systematic variation. Mean steady-state lorazepam levels were highly correlated with daily dose in mg/kg, but were not related to age. Lorazepam was not detected in any plasma samples drawn one week after discontinuation of treatment.


The dispersal of cells from human gynecologic specimens: chemical agents. An investigation of nonenzymatic chemical solutions for the dispersal of gynecologic cells in suspension was undertaken using known general information on the composition of cell surfaces and cell adhesive properties. Studies on human cells cultured in vitro showed that thiol reducing agents release microextension-mediated contacts. Dispersing solutions containing lithium diiodosalicylate separated cells, presumably by extracting glycoproteins. Solutions without this agent but containing tetramethylurea and mixed sugars similar to those found on the cell surface separated cells by interacting with hydrogen bonds and other noncovalent polysaccharide interactions thought to hold cells together. Solutions containing ethylenediaminetetraacetate, dithiothreitol, tetramethylurea, mixed sugars, mixed amines and inorganic ions are suggested as mildly-acting dispersal solutions on the basis of evaluationtions by phase contrast microscopy, Papanicolaou staining and particle volume analysis.


The antibody response to the T1699 murine adenocarcinoma: antibody class and subclass heterogeneity detected in serum and in situ. We have used indirect immunofluorescence to study antibody responses directed against membrane antigens expressed on in vitro and in vivo T1699 mammary adenocarcinoma cells. IgG1, IgG2a, IgG2b, IgG3, IgA, and IgM antibodies were present in the serum of DBA/2 mice bearing T1699 tumors; IgG2a and IgG2b antibodies were readily detected on the cells in situ. Lesser amounts of the other classes and subclasses could be detected by indirect immunofluorescence measurements on in vivo tumor cells and with low pH eluates of in vivo cells tested on the in vitro line of T1699. The antigenic determinants on in situ tumor cells are not saturated with antibody as these cells demonstrated enhanced fluorescence of all immunoglobulin classes and subclasses when treated with autologous serum. Experiments with thymus-depleted mice indicated that immunoglobulin production was strongly dependent on thymus-derived cells for all immunoglobulin classes and subclasses except IgG2b. Our studies suggest that IgG2a may be active in the macrophage-mediated cytotoxic reaction and IgG2b in the immediate hypersensitivity reaction to T1699 cells. These results provide further evidence for an active role of tumor-specific antibody in the host defense to the T1699 adenocarcinoma in situ.


Sensitivity of amplifier T cells involved in the antibody response to type III pneumococcal polysaccharide to anti-lymphocyte serum. Amplifier T cells responsible for enhancement of the antibody response to type III pneumococcal polysaccharide have been shown to be resistant to the effects of antilymphocyte serum (ALS) given at the time of immunization, a treatment that eliminates suppressor T cell activity. The resistance of amplifier T cells to ALS can be attributed to the fact that their activity develops after that of suppressor T cells. ALS given 1 or 2 days after immunization does abrogate amplifier T cell activity, independent of the mode by which that activity is elicited. The data emphasize the importance of kinetic considerations in understanding the effects produced by immunologically active agents such as ALS.


Effect of concanavalin A on lymphocyte interactions involved in the antibody response to type III pneumococcal polysaccharide. II. Ability of suppressor T cells to act on both B cells and amplified T cells to limit the magnitude of the antibody response. When administered 2 days after immunization with 0.5 microgram Type III pneumococcal polysaccharide (SSS-III), the T lymphocyte mitogen concanavalin A (Con A) stimulates a 2.6-to 7-fold enhancement of the plaque-forming cells (PFC) response to SSS-III in vivo. This enhancement requires the presence of amplified T cells, which act by driving PFC or their precursors to extra rounds of proliferation. The extra proliferation that can be stimulated by Con A is not seen in the normal primary response to SSS-III; but treatment with anti-lymphocyte serum (ALS) to remove suppressor T cells will permit the additional proliferation to occur. This indicates that in the primary response to SSS-III, suppressor T cells act on amplifier T cells to limit the magnitude of the antibody response. Only suppression of B cells can account for the further suppression induced by Con A given at the time of immunization or by low-dose paralysis of the SSS-III response. The relatively late development of amplified activity compared to suppressor activity appears to account for the absence of amplifier activity after primary immunization with SSS-III. It is apparent that one can explain the regulatory effects observed during the development of an immune response to SSS-III only by considering both T cell- B cell and T cell- T cell interactions, together with the temporal relationships involved in those interactions.


The presence of immunoregulatory cells in chicken thymus: function in B and T cell responses. The presence of immunoregulatory cells in chicken thymus was studied by using several different systems. Chickens injected with large numbers of syngeneic thymocytes were tested for their ability to produce antibody to heterologous red cells. Similar chickens were studied for their ability to reject allogeneic skin grafts. In separate studies, mixtures of thymocytes with spleen cells or with peripheral blood leukocytes were assayed for their ability to respond to PHA or to produce a graft-vs-host reaction in embryonic chicks. These studies indicated that immunoregulatory cells exist in chicken thymus, which displays both helper and suppressor activity. The suppressor cells were more prevalent or more easily detectable in young birds and in chickens with intact bursas. The helper function of thymocytes was seen to better advantage with cells derived from older animals and from bursectomized donors.


Immune responses during measles infection in immunosuppressed Rhesus monkeys. Rhesus monkeys immunosuppressed with horse anti-human thymocyte gamma-globulin (ATG) were infected with measles and simultaneously inoculated with sheep erythrocytes (SRBC), a thymus-dependent antigen, and with pneumococcal polysaccaride type III (SSS-III), a thymus-independent antigen. ATG treatment alone suppressed SRBC antibody production, had no effect on SSS-III antibody production, and effectively eliminated circulating T cells compared to nonsuppressed monkeys. ATG treatment of measles-infected monkeys resulted in delayed virus clearance and delayed antibody production compared to nonsuppressed infected monkeys. After cessation of ATG treatment, measles antibodies and T cells reached normal levels, and measles virus was eliminated. Thus, immune clearance of measles virus is T cell-dependent, but the relative roles of cellular- and humoral-mediated immunity in vivo could not be clearly separated. Also, measles infection was associated with a decreased T cell mitogen responsiveness of circulating lymphocytes but not of lymph node lymphocytes, suggesting an altered circulating pattern of the cells responsible for delayed hypersensitivity. Also, measles infection had no effect on T-dependent antibody production to SRBC.


Cross-protective immunity to Gram-negative bacilli: studies with core glycolipid of Salmonella minnesota and antigens of Streptococcus pneumoniae. Two immunoprophylactic approaches to the control of infections caused by gramnegative bacilli were evaluated by study of experimental infections in animals. The core glycolipid antigen derived from the Re mutant of Salmonella minnesota R595 is shared by virtually all enteric bacteria, and immunization with this endotoxin protects against the hemodynamic sequelae of bacterial infection and pyrexia without enhancing intravascular clearance of bacteria. The degree of protection afforded by active and passive immunization with core glycolipid was significantly less than that conferred by type-specific immunization. Escherichia coli and Klebsiella pneumoniae share capsular antigens with some strains of Streptococcus pneumoniae; by the mechanism of enhanced opsonization, antibodies to S. pneumoniae may cross-protect against infection with E. coli or K. pneumoniae.


Comparison otitis media due to types 3 and 23 Streptococcus pneumoniae in the Chinchilla model. Only a few pneumococcal serotypes are responsible for the majority of cases of acute otitis media due to Streptococcus pneumoniae in children. The immunopathogenesis of otitis media cause by two serotypes, type 3 and 23, was studied in chinchillas. Chinchillas with type 23 pneumococcal otitis media were capable of clearing the infection over a six-week period without treatment, whereas the type 3 infection persisted throughout the six-week study. In addition, bacteremia or meningitis occurred more frequently and earlier with type 3 pneumococcal otitis media than with type 23. The resolution of the type 23 pneumococcal infection paralleled the development of type-specific antibody, as measured by radioimmunoassay, whereas type 3 infection was associated with a fall in serum antibody to low levels. Since the pathogenesis of pneumococcal otitis media in the chinchilla differs between pneumococcal serotypes, it may be important to correlate pneumococcal serotypes with sequealae and recurrence of otitis media in children.


Maturation of regulatory factors influencing magnitude of antibody response to capsular polysaccharide of type III Streptococcus pneumoniae. Mice of different ages were evaluated for their ability to give a plaque-forming cell response to the capsular polysaccharide of Streptococcus pneumoniae (SSS-III). The response of amplifier and suppressor thymus-derived (T-) cells was also evaluated. The responses to an optimally immunogenic dose of SSS-III for two-and three-week-old mice were only 7% and 14%, respectively, of that produced by adult mice; values comparable to those of adult mice were attained by four weeks of age. Activity of amplifier T-cells, which was minimal at two to four weeks of age, matured slowly and did not reach a maximum until eight to 10 weeks of age. However, activity of suppressor T-cells was found to be fully developed as early as two weeks of age. These findings indicate that the inhibitory effects of suppressor T-cells are predominant in young mice and that such cells may play an active role in determining the ease with which immunological unresponsiveness is induced in neonates.


Temperature-sensitive mutants of type I Streptococcus pneumoniae: preparation, characterization, and evidence for attenuation and immunogenicity. Thirteen temperature-sensitive (ts) mutants of type I Streptococcus pneumoniae were selected after exposure of virulent wild-type (ts+) organisms to nitrosoguanidine. Each mutant resembled the ts+ parent in properties of alpha-hemolysis, bile solubility, optochin sensitivity, antibiotic sensitivity, and serotype. Unlike the ts+ parent, however, each ts mutant was restricted in its capacity to form colonies on blood agar at 38 C. With the exception of two mutants, there was a correlation between the degree of temperature-sensitivity of a mutant and its genetic stability. When inoculated intraperitoneally into mice, 11 of 13 mutants were attenuated and induced homologous resistance. Three mutants (ts 1, ts 3, and ts 4) were also studied in hamsters and were found to be attenuated and immunogenic after intraperitoneal injection. Study of the behavior of mutants ts 1, ts 3, and ts 4 in the blood of hamsters suggested that attenuation may be related, in part, to decreased growth and survival of ts organisms at body temperature. Mutants ts 1 and ts 4 were completely attenuated for hamsters when administered intranasally and induced significant resistance to subsequent challenge with wild-type organisms by the same route. Local administration of ts mutants of type 1 S. pneumoniae to hamsters may provide a model for evaluating the potential of live vaccines in the prevention of disease due to bacterial respiratory tract pathogens.


Adenyl cyclase in Bordetella pertussis vaccines. Brodetella pertussis organisms have a large amount of extracytoplasmic adenyl cyclase, part of which is found in the supernatant culture medium during exponential growth. The enzyme differs from previously studied bacterial adenyl cyclases in biochemical characteristics as well as in location. Several commercial pertussis vaccines were found to contain adenyl cyclase activity; this activity is probably due to the extracytoplasmic enzyme associated with and released from the whole cells in the vaccine.


Prevention of pneumococcal infection by immunization with capsular polysaccharides of Streptococcus pneumoniae: current status of polyvalent vaccines. Because of the continuing morbidity and mortality resulting from pneumococcal infection, a program was instituted to redevelop polyvalent vaccines consisting of capsular polysaccharides of Streptococcus pneumoniae. Vaccines containing 50 microgram each of the capsular polysaccharides of as many as 13 pneumococcal types have been shown to be safe, antigenic, and 78.5% effective in the prevention of type-specific putative pneumococcal pneumonia and of type-specific pneumococcal bacteremia in adults. In a population in which pneumococcal pneumonia predominated, the total incidence of radiologically confirmed pneumonia, irrespective of cause, was reduced by 54.3% by use of a tridecavalent vaccine. The efficacy of vaccine in the prevention of infection during the first two years of life is under investigation. The vaccine is recommended for those at high risk of pneumococcal infection or of a fatal outcome from such illness.


Platelet-aggregating factor and the aggregation of fixed washed platelets. Platelet-aggregating factor (PAF) was removed from bovine plasma by human platelets fixed with 2% formaldehyde. The degree of adsorption was directly related to the platelet concentration and the length of incubation. Fixed washed platelets (FWP) aggregated with bovine plasma could be deaggregated by 1M KCl, Evans blue, and 8M urea but not by beta-galactosidase. Incubation with 1M KCl eluted some but not all of the PAF, as the deaggregated platelets spontaneously aggregated upon removal of the deaggregating conditions. Also, fixed platelets adsorbed PAF even in the presence of 1M salt or after treatment with Evans blue. Platelet aggregation was not affected by thrombin (20 micron/ml) but was abolished by trypsin at concentrations as low as 4 X 10(-1) microgram/ml. The data suggest that deaggregation is not the result of elution of the loosely bound aggregating factor from the platelet surface, but rather the disruption of noncovalent interplatelet bridging between one or more PAF molecules bound to a specific receptor.


A comparison of lipoprotein lipase activity and adipocyte differentiation in growing male rats. During adipose tissue development changes in lipoprotein lipase activity per adipocyte precede significant changes in fat cell size. Lipoprotein lipase activity per adipocyte increases fourfold from the second to seventh postnatal week. Furthermore, when isolated adipocytes and stromal--vascular cells are prepared by collagenase digestion of adipose tissue, there is a progressive shift in enzyme activity during development from the stromal-vascular compartment to the adipocyte fraction. The data support the concept that during normal development a "bed" of preadipocytes is synthesized during the suckling period. The data further suggest a regulatory role for lipoprotein lipase in the control of "lipid-filling" during early postnatal development.


Blood respiratory properties of rainbow trout (Salmo gairdneri) kept in water of high CO2 tension. 1. Blood O2 transport and acid-base balance were studied at 20 degrees C in rainbow trout (Salmo gairdneri) which had been kept in water of high CO2 content (15 mmHg) for at least a week. Also the blood gas chemistry of fish rapidly entering or leaving the hypercapnic environment was studied. 2. Fish entering high CO2 water suffered a sharp decrease in blood pH which significantly reduced O2 transport by the blood, but after a few hours considerable compensation was achieved. 3. After at least a week in high CO2 water, trout showed elevated plasma bicarbonate and PCO2 levels, and a decrease in plasma chloride, while pH was about 0 - 1 pH unit below the level for control fish. Oxygen transport by the blood was marginally reduced. 4. Hypercapnic fish rapidly entering fresh water showed a sharp increase in blood pH and a decrease in blood PO2. These parameters regained normal values after a few hours but plasma bicarbonate and chloride levels took much longer to regain control concentrations. 5. Acid-base balance in hypercapnic fish is discussed with particular reference to the role of the branchial ion exchanges.


pH and haemoglobin oxygen affinity in blood from the Antarctic cod Dissostichus mawsoni. Blood pH in the antarctic cod (Dissostichus mawsoni) and in two Trematomus species, occlrring at --1-9 degrees C, is extremely high (approximately 8-2 to 8-3). This supports and extends Rahn's (1966) model for the temperature-pH relationship in cold-blooded vertebrates. The blood of D. mawsoni shows a low oxygen affinity (P50 approximately equal to 14-5 mmHg at pH 8-16 and -1-9 degrees C). Despite normal in vitro temperature and pH sensitivities, blood P50 increases only slightly when live fish are temperature-stressed (+ 4-0 degrees C), or become acidotic as a result of agitational stress (blood pH 7-71), primarily as a result of compensatory decreases in blood ATP levels. Oxygen-binding properties of 'stripped' (cofactor-free) solutions of D. mawsoni haemoglobin were measured in attempts to elucidate the molecular mechanisms involved in the function of the pigment.


The effect of progressive hypoxia on respiration in the toad Bufo marinus. 1. As Bufo marinus became progressively hypoxic over a period of 90 min, there was a rise in arterial pH, presumably brought about by hyperventilation. The alkalosis gradually disappeared when oxygen levels became very low. It is suggested that this is the result of a respiratory or a metabolic pH adjustment, or both. 2. Hypoxic animals developed a characteristic breathing pattern in which discrete periods of lung ventilations alternated with buccal oscillations or respiratory pauses. 3. A pronounced bradycardia was associated with the concomitant decline of inspired and arterial PO2. 4. Although respiratory rates were greater than normal resting values in the initial stages of post-hypoxia, the pre-exposure breathing pattern was quickly restored. Following recovery from bradycardia (60 min), the breathing rates, arterial blood gases and pHa returned to normal within 30 min.


Suppression of in vitro cytotoxic response by macrophages due to induced arginase. Arginine was found to be completely depleted from cell-free supernates of mixed leukocyte cultures suppressed by the addition of excess macrophages. Partial reversal of macrophage-mediated suppression was accomplished by daily addition of a cocktail containing arginine and nonamino acid nutrients. Complete reversal of the suppression was accomplished by the addition of arginine and glucose to the medium and the nonadherent cells after their separation from the adherent macrophages. A marked increase in the enzyme arginase was found in macrophages that had been cultured 24 h in vitro in Eagle's minimum essential medium plus 10% fetal calf serum, in peritoneal cells activated by prior injection of thioglycollate, and in one spleen activated by a graft vs. host reaction.


Elastase secretion by peritoneal exudative and alveolar macrophages. Mouse alveolar macrophages (AM) cultured in the absence of serum secrete an elastolytic enzyme. The elastase from AM resembles the previously described elastase from peritoneal macrophages (PM) in pH optimum and inhibition profile. The macrophage enzymes do not appear to be stored, and with periodic changes in the culture medium, accumulate extracellularly for up to 10 days. Resident PM produce barely detectable levels of extracellular elastase unless given a phagocytic load. Thioglycollate-stimulated peritoneal exudative macrophages (PEM), however, secret easily detectable levels of elastase, which can be further increased with a phagocytic load. Without any additional stimulation, AM secret an elastolytic activity comparable to that of the PEM receiving a phagocytic load, but unlike PM they do not increase elastase secretion after phagocytosis.


Influence of splenectomy on first set rejection reactions of C57BL/6 females to male skin isografts. The tempo with which C57BL/6 females reject male skin isografts is determined in part by the immunogenicity of the H-Y antigen and in part by the capacity of the host to respond immunologically. Our studies indicate that the spleen plays an important role in determining the briskness of the rejection process in that splenectomy 7-30 days before grafting with male skin significantly shortens the survival time. The results of reconstitution experiments suggests that a population of cells is present in spleens of normal, but not specifically sensitized, females which can restore the conventional first set reaction in splenectomized females. It is inferred that this resident population normally operates in spleen-intact females to delay the development of specific effector responses. Lymphoid cells from H-Y antigen-sensitized, splenectomized females failed to evoke graft-vs.-host responses in males whereas similar cells from females with spleens intact did possess graft-vs.-host potential.


[Critical observations on the use of phenolphthalein monophosphate as a substrate for the determination of prostatic acid phosphatase (author's transl)]. At pH 10, which is in the alkaline measurement range, phenolphthalein is bound by albumin. The resonance system of this acid-base indicator cannot therefore be completely formed, and the colour intensity is correspondingly lower. Albumin also binds phenolphthalein monophosphate at pH 4-8, in the pH range of the enzymic reaction. Depending on the relative concentrations, the substrate concentration is therefore either suboptimal (manifested as a lower catalytic concentration of phosphatase), or the substrate excess is bound by albumin, so that the substrate inhibition of the phosphatase is decreased or abolished. Binding of phenolphthalein and its monophosphate to albumin is partly prevented by the addition of alcohol. These results show that the determination of prostate phosphatase with the substrate phenolphthalein monophosphate is markedly influenced by various factors. The interpretation of the observed colour intensity is therefore difficult, and the calculation of the catalytic concentration is frequently subject to error.


Relations between ameboid movement and membrane-controlled electrical currents. We have studied the pattern of electrical currents through amebas (mainly Chaos chaos) with an ultrasensitive extracellular vibrating probe. Amebas drive both steady currents and current pulses through themselves. Relatively steady current with an average surface density of 0.1-0.2 muA/cm2 enters the rear quarter of an ameba and leaves its pseudopods. Streaming reversals are preceded by changes in this current pattern and the region with the largest new inward current becomes the new tail. Ion substitution studies suggest that some of the steady inward current is carried by calcium ions. Characteristic stimulated pulses of current sometimes follow the close approach of the vibrating probe to the side of an advancing pseudopod. Such a pulse enters the cytoplasm through a small patch of membrane near the probe (and seems to leave through the adjacent membrane), is usually followed by hyaline cap and then by pseudopod initiation, is calcium dependent, lasts about 5-10 s, and has a peak density of about 0.4 muA/cm2. Spontaneous pulses of similar shape and duration may enter or leave any part of an animal. They are much less localized, tend to have higher peak densities, and occur in physiological salt solutions at about 0.2-4 times per minute. Retraction of a pseudopod is always accompanied or preceded by a spontaneous pulse which leaves its sides.


Temperature-dependent changes of chloride transport kinetics in human red cells. Chloride self-exchange in human red cells was studied between 0 degrees C and 38 degrees C. At higher temperatures the flow-tube method was used. Although the general features of chloride transport at 0 degrees C and 38 degrees C are similar, the following differences were found: (a) the maximum pH of chloride self-exchange flux was lowered 0.6 pH unit from 7.8 to 7.2 when temperature was increased from 0 degrees C to 38 degrees C; (b)the apparent half-saturation constant increased from 28 mM at 0 degrees C to 65 mM at 38 degrees C; (c) chloride transport at body temperature is slower than predicted by other investigators by extrapolation from low-temperature results. Chloride transport increased only 200 times when temperature was raised from 0 degrees C to 38 degrees C, because the apparent activation energy decreased from 30 kcal mol(-1) to 20 kcal mol(-1) above a temperature of 15 degrees C; (d) a study of temperature dependence of the slower bromide self-exchange showed that a similar change of activation energy occurred around 25 degrees C. Both in the case of Cl(-) (15 degrees C) and in the case of Br(-) (25 degrees C), critical temperature was reached when the anion self-exchange had a turnover number of about 4x10(9) ions cell (-1)s(-1); (e) inhibition of chloride transport by DIDS (4,4'- diisothiocyano-stilbene-2,2'-disulfonate)revealed that the deflection persisted at 15 degrees C at partial inhibition (66 percent) presumably because DIDS inactivated 66 percent of the transport sites. It is suggested that a less temperature- dependent step of anion exchange becomes rate limiting at the temperature where a critical turnover number is reached.


Preferential hemolysis of postnatal calf red cells induced by internal alkalinization. Red blood cells from neonatal calves, but not from adult cows, rapidly hemolyze in buffered 300 mM solutions of a variety of nonelectrolytes and amino acids. Of these compounds, sucrose is chosen to elucidate the mechanism by which this preferential hemolysis takes place. As in other mammalian red cells, both calf and cow cells are found to be impermeable to sucrose and, in an isosmolar sucrose solution, to undergo volume shrinkage caused by the net loss of chloride ions with concomitant increase in intracellular pH. To test the potential role of intracellular pH change associated with chloride loss in promoting hemolysis, intracellular pH was altered by: (a) a direct addition of fixed acid or base to sucrose solution; (b) the removal of dissolved CO(2) from sucrose solution; and (c) the addition of cells to isotonic NaHCO(3) solution in the absence of sucrose. In all cases, only calf and not cow cells underwent hemolysis. Moreover, 4-acetamido-4'-isothiocyano-2,2'-stilbene disulfonic acid, a potent anion transport inhibitor, completely protected calf cells from hemolysis and caused a nearly total inhibition of both chloride loss and intracellular alkalinization. Furthermore, the hemolytic process is closely related to the integrity of a membrane protein, the band 3 protein, which can be cleaved to varying degrees by the combined treatment of pronase and lipase. Hemolysis is progressively inhibited as the band 3 protein undergoes proteolysis, until a total inhibition of hemolysis takes place when almost all of the band 3 protein is digested into smaller protein components with a mol wt of 65,000 and 35,000 daltons. These results suggest that the intracellular alkalinization process leading to a structural instability of the membrane band 3 protein is responsible for this calf cell hemolysis.


Influence of solute and hydrogen ion concentration on the water relations of some xerophilic fungi. Germination and growth of six xerophilic fungi, Aspergillus flavus, Aspergillus ochraceus, Eurotium chevalieri, Chrysosphorium fastidium, Wallemia sebi and Xeromyces bisporus were examined on media of a wide range of water activities (aw). The influence of three solutes, NaCl, glycerol and a glucose/fructose mixture, was studied at pH 4-0 and pH 6-5 using a plate-slide technique. Germination times and growth rates were affected by solute type, but the influence of pH was less marked. Except for Wallemia sebi, the fungi grew most strongly on glucose/fructose and were partially or completely inhibited by NaCl. The results showed that a universal isolation medium for xerophilic fungi could be based on glycerol or glucose/fructose but not on NaCl as aw-limiting solute.


Effect of dietary threonine supplementation on tyrosine toxicity in the rat. The objective of this study was to determine the effect of threonine supplementation on tyrosine metabolism in rats fed a low protein diet with excess tyrosine. The growth retardation and the development of eye and paw lesions that occur in rats ingesting a basal plus 3% or 5% L-tyrosine diet could be alleviated partially by the addition of 0.5% or 1.0% L-threonine to the diet. An increased blood tyrosine level in rats fed excess tyrosine was also lowered by threonine supplementation. In tyrotoxic conditions, the activities of liver tyrosine transaminase (EC 2.6.1.5) and threonine dehydratase (EC 4.2.9.16) were elevated, but p-hydroxyphenyl pyruvic acid oxidase (EC 1.13.11.27) which is also intimately associated with tyrosine toxicity was found to be inactivated. Furthermore, biosynthesis of ascorbic acid in liver was significantly lowered in this condition. However, addition of L-threonine in the diet, not only could cure the signs developed due to excess tyrosine, but also could affect the levels of enzymes studied.


Decreased opsonization for Streptococcus pneumoniae in sickle cell disease: studies on selected complement components and immunoglobulins. Opsonic activity for Streptococcus pneumoniae in the sera of patients with sickle cell disease was reduced in comparison to the opsonic activity of sera from age-matched normal children. No difference in opsonic activity for Escherichi coli was observed in the sera from patients or normals. Total hemolytic complement, conversion of C3 by inulin and cobra venom factor, and levels of C3, factor B, properdin, C3b inactivator, and immunoglobulins G, A, and M were normal in patients' sera. The opsonic abnormality for S. pneumoniae was attributed to a deficiency of serum proteins rather than to an inhibitor of opsonic function. The data suggest that decreased opsonization was not associated with a deficiency of those complement components or immunoglobulins measured in this study.


Methylmalonic aciduria: a variant form of methylmalonyl coenzyme A apomutase deficiency. Sisters aged 3 and 4 1/2 years, respectively, who had grown and developed normally were found to have methylmalonic aciduria. The elder had had only one previous illness, at 18 months, and was thriving at six years without treatment; she was excreting up to 2 gm methylmalonic acid per day. The younger sister died during her third episode of illness, at 3 years of age. Studies of cultured skin fibroblasts showed deficient methylmalonyl coenzyme A mutase apoenzyme activity; cobalamin metabolism was normal. There was a moderate response in cultured cells to large amounts of added hydroxycobalamin; but treatment of the patient with high doses of intramuscular vitamin B12 for ten days failed to lower the urinary excretion of methylmalonic acid.


Bilateral retractile testes--subsequent effects on fertility. A group of 43 adults who as children had been diagnosed as having bilateral retractile testes were traced. Seventy-four per cent of the married patients had children. The testicular volume was normal in all the 19 patients examined.


Fetal and maternal blood glucose, insulin and acid base observations following maternal glucose infusion. The aim of the present investigation was to examine the fetal and maternal blood glucose and insulin response following glucose infusion to the mother. The studies were performed on 11 primigravid patients with a gestational age of 38-40 weeks during the first stage of labor. Glucose was given intravenously by a bolus injection of 330 mg/kg body weight, followed by a glucose infusion of 27.5 mg/kg/min for 60 min. Glucose concentration, immuno-reactive insulin (IRI), pH and base excess of the maternal and fetal blood were measured before and during maternal glucose load. Maternal blood glucose rose within 10 min. up to 280.0 mg% (SD 25.9). This level could be fairly maintained throughout the experiment. The maternal glucose was after 60 min. infusion 326.5 mg% (SD 46.9). Fetal glucose concentration rose continuously from 65.8 mg% (SD 5.8) at control to 249.2 mg% (SD 23.3) after 60 min. The increase of maternal and fetal glucose was associated with an elevation of immuno-reactive insulin (IRI). The maternal insulin was 24.0 micronU/ml (SD 8.0). It was scattered over a wide range (55.4 micronU/ml-217.1 micronU/ml) after 60 min. glucose infusion. The fetal insulin was 17.0 micronU/ml (SD 5.2) at control and rose by 86.5% (SD 80.5) after 60 min. glucose load. One case of a mother with a subclinical diabetes mellitus deviated where the fetal insulin rose from 26.0 micronU/ml at control to 215.6 micronU/ml after 60 min. infusion. The increase of insulin per glucose rise was correlated to fetal body weight. During glucose infusion to the mother of both, fetal and maternal, acid base parameters remained unchanged. From these observations it may be concluded that in the human fetus insulin secretion following a single glucose load is generally low, however, it increases in cases where the maternal insulin response to glucose load is abnormal. This might be related to a chronic stimulation by glucose of the fetal pancreatic islet cells in poorly controlled diabetic and possibly prediabetic patients.


The effect of temperature on the tensile strength and disintegration of paracetamol and oxytetracycline tablets. The tensile strengths and disintegration times of paracetamol and oxytetracycline tablets at room temperature are higher when they have been prepared at high temperatures, e.g. 85 degrees, than at room temperature or below, e.g.--20 degrees. The activation energies of the two materials, 3 and 1 k cal mol-1 (13 and 4kJmol-1) respectively, were derived from plots of log tensile strength and log disintegration time versus the reciprocal of the absolute temperature. The results have been explained in terms of sintering theory and the formation of welded bonds between particles.


Binder distribution onto binary mixtures of glass spheres. Glass spheres (0-8 and 0-4 cm diam.) in differing weight ratios were treated in a rotating pan with four pharmaceutical binder fluids. Binder uptake for a sphere of 1 g was greater onto the smaller spheres irrespective of binder or wt ratio used. There were differences in uniformity of uptake, the solution binders (PVP and gelatin) being more evenly distributed than the mucilaginous binders (starch and methylcellulose). Binder distribution on the basis of unit surface area showed less obvious differences between the sphere sizes. Possible consequences of such binder distribution through powder aggregates are discussed.


Permeation of oestrone, oestradiol, oestriol and dexamethasone across cellulose acetate membrane. The permeation across cellulose acetate of three oestrogens, differing only in the number of hydroxyl groups attached to the nucleus, and a 'standard' steroid, dexamethasone, was investigated using the lag-time method for calculating diffusion parameters, between 10 and 40 degrees. Diffusion coefficients for the similarly-sized oestrogens were relatively insensitive to marked changes in polarity, but increased permeation was correlated with increased partition coefficients, decreased polarity and fewer hydroxyl groups on the nucleus. Permeation increased with temperature and energies of activation were calculated from Arrhenius-type plots. Ep values ranged from 4-84 k cal mol-1 (20 kJ mol-1) for the least polar steroid (oestrone) to 6-91 k cal mol-1 (29 kJ mol-1) for the most polar steroid (oestriol). The results implied that steroid diffusion occurred through aqueous membrane channels, but that it was impeded to various extents by both obstruction and polar interaction effects.


Effect of a second solubilizate on the partition coefficient of drugs in micellar solution and their permeation rate across an artificial membrane. The distribution of a solubilizate between micelles and the aqueous phase does not obey a simple partition law when a second solubilizate species is present. Alterations of the apparent partition coefficient cannot be explained in terms of a simple displacement mechanism, following the interaction of both solubilizates with the same site of the micelle. A non linear increase in solubilizate association to micelles following an increase in surfactant concentration is observed in the presence of a second solubilizate. A depression in the cloud point temperature follows the addition of a second species and is such that cannot be interpreted as a simple additive effect. Alteration of the apparent partition coefficient in a miscellar solution has an effect on the permeation rate of the solubilizate across an artificial membrane. Biopharmaceutical implications are discussed.


The release of plasticizer from polyvinyl chloride haemodialysis tubing. The extent of migration of plasticizer from haemodialysis lines into saline and plasma has been examined and a number of different types of tubing have been compared. A model system designed to simulate in vivo haemodialysis was evolved. The method uses a continuous cyclohexane extraction in order to mimic uptake of plasticizer by body tissues. Although not an ideal system, the results obtained are indicative of the extent to which the plasticizers are leached.


The effect of tris buffers on rat liver mitochondrial monoamine oxidase. The effect of tris buffers upon the monoamine oxidase (MAO) activity in rat liver mitochondria has been investigated. Tris buffer was shown to inhibit MAO in a non-competitive manner with a Ki of 15-25 mM. Tyramine, 5-hydroxytryptamine and beta-phenethylamine but not benzylamine oxidations were all inhibited by tris buffer. All inhibitions, except that of 5-HT, were completely reversible. It is suggested that these effects are produced by conformational changes in the structure of the MAO. The significance of these results is discussed with respect to the use of tris buffers in the extraction and estimation of the activity of MAO.


Metabolic oxidation of aralkyl oximes to nitro compounds by fortified 9000g liver supernatants from various species. Incubation of 'amphetamine oxime' (IIa, anti-benzyl methyl ketoxime) with fortified rabbit liver 9000 g supernatants gave the nitro compound (Ie) and the beta-hydroxylated oxime (IIc) in addition to the previously reported ketone (IIb) and alcohol (Ic) metabolites. Formation of the products was cofactor dependent. The nitro compound was also formed using mouse, hamster and guinea-pig 9000 g liver supernatants and to a minor extent by rat liver. The oximes of 2-phenethylamine (IIe) and norfenfluramine (IIg) were also metabolized to the corresponding nitro compounds, ketones and alcohols with rabbit 9000 g liver supernatants; however, no nitro compound (IIIb) was detected after the incubation of 'mexiletine oxime' (IVa). The metabolic products were identified and characterized by g.l.c., t.l.c. and g.l.c. linked mass spectrometry by comparison with synthetic materials.


Dopamine receptors in rat striatum and nucleus accumbens; conformational studies using rigid analogues of dopamine. A study was made of the actions of dopamine and of some 2-amino-1,2,3,4-tetrahydronaphthalenes on dopamine-sensitive adenylate cyclase in homogenates of rat striatum and nucleus accumbens. The compounds were also tested for their ability to stimulate motor activity following bilateral injection into the nucleus accumbens of conscious rats. The most active compounds on adenylate cyclase from both striatum and nucleus accumbens were dopamine and 2-amino-6,7-dihydroxy-1,2,3,4-tetrahydronaphthalene(6,7-diOHATN). The 5,6-dihydroxy analogue (5,6-diOHATN) was 50 times less active than 6,7-diOHATN in striatal homogenates and 350 times less active in homogenates of nucleus accumbens. All dihydroxy compounds tested were active in causing stimulation of motor activity, the most active compounds being 6,7-and5,6diOHATN. Both dimethoxy derivatives tested were inactive on the adenylate cyclase and as locomotor stimulants.


The effects of diethylcarbamazine citrate on smooth muscle. The effect of the anthelmintic drug diethylcarbamazine citrate (DECC) was examined on the guinea-pig isolated ileum, rabbit duodenum, chick oesophagus, rat portal vein and pig coronary artery. DECC contracted all the gastrointestinal smooth muscle preparations. The contractions were antagonized by hexamethonium and atropine but they were not affected by mepyramine or methysergide in concentrations that abolished, or markedly reduced, responses to histamine and 5-hydroxytryptamine. DECC inhibited the responses of the guinea-pig ileum to other spasmogens, namely, acetylcholine, histamine and nicotine. Physostigmine markedly potentiated the responses of the chick oesophagus and the rabbit duodenum to DECC. DECC relaxed the potassium chloride-induced contractions of the pig coronary artery strips; these relaxations were not modified by propranolol or calcium chloride. There was no evidence that DECC released histamine from skin or muscle.


Interpretation of dissolution rate data from in vitro testing of compressed tablets. To find if theoretically and experimentally a relation existed between the dissolution rate theory of Kitazawa, Johno & others (1975) and that of Wagner (1969), a study was undertaken with uncoated caffeine, aspirin and proxyphylline tablets using two dissolution methods. Although the original treatment for surface area of drug available for dissolution was quite different between the two dissolution theories, the dissolution rate constants obtained were in fair agreement. Hence it might not be always necessary to take into consideration changes in the surface area as a function of dissolution rate, and the 1n W infinity/(W infinity) versus time plot devised by Kitazawa & others might be a useful and simple means of obtaining the dissolution rate constant of an active ingredient from a dosage form such as compressed tablet.


The extraction and analysis of benzodiazepines in tissues by enzymic digestion and high-performance liquid chromatography. A simple, rapid and sensitive method is described for the analysis of unchanged benzodiazepine drugs and their metabolites, in human and animal tissues, using the proteolytic enzyme subtilisin Carlsberg followed by reverse-phase liquid chromatography. The enzymic digestion yields far higher recoveries than conventional extraction methods, while the liquid chromatogrpahic analysis allows the rapid separation and detection of nanogram quantities of these drugs without the need for elaborate "clean-up" procedures.


Biochemical differentiation of amphetamine vs methylphenidate and nomifensine in rats. Amphetamine-like stimulants were divided into two groups, one in which the stereotyped behaviour was not antagonized by reserpine [(+)-amphetamine, (-)-amphetamine, methamphetamine, phenmetrazine and phenethylamine] and another group in which the behavioural effects were blocked by reserpine (methylphenidate, nomifensine, pipradrol and amfonelic acid (NCA; Win 25978)). Both groups increased homovanillic acid (HVA) in whole brain 2 h after administration. The 'methylphenidate group' also increased brain 3,4-dihydroxyphenylacetic acid (DOPAC) in naive rats; whereas the '(+)-amphetamine group' decreased DOPAC in naive rats, as well as in reserpinized rats, alpha-methyl-p-tyrosine-treated rats and after acute hemisection. The reserpine antagonism of the 'methylphenidate group'-induced stereotyped behaviour was partially reversed by type A monoamine oxidase inhibition. The '(+)-amphetamine group'-induced stereotyped behaviour was not blocked by short time pretreatment with alpha-methyltyrosine, only by longer pretreatment intervals. The mechanisms by which the two groups are differentiated biochemically is discussed with special attention to possible intra-neuronal inhibition of dopamine oxidation by the '(+)-amphetamine group'.


Neurochemical correlates of ethanol withdrawal: alterations in serotoninergic function. Cessation of chronic ethanol administration, and elimination of ethanol from the body, results in a withdrawal syndrome in mice characterized by behavioural symptoms and hypothermia. During withdrawal, the accumulation rate of [14C] 5-hydroxytryptamine (5-HT) from [14C]tryptophan, was significantly lower in the brainstem of the ethanol-withdrawn animals than in controls. A similar pattern was seen in forebrain. When the rate of 5-HT accumulation was determined using pargyline, no differences occurred between control and ethanol-treated animals. The endogenous concentrations of tryptophan in plasma, and tryptophan, 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) in brain were the same in ethanol-treated and control animals. It is suggested that the changes in accumulation of 14C-5-HT and 14C-5-HIAA in ethanol-withdrawn animals reflected alterations in electrical activity of serotoninergic neurons during withdrawal.


Identification of di- and tri-substituted hydroxy and ketone metabolites of delta1-tetrahydrocannabinol in mouse liver. In vivo liver metabolites of delta1-tetrahydrocannabinol (delta1-THC) were examined with a gas chromatograph--mass spectrometer--computer system as trimethylsilyl (TMS), [2H9]TMS and methyloxime-TMS derivatives. In addition to the reported monohydroxy, acid, and hydroxyacid metabolites, the following multiply substituted metabolites were identified: 2'',7-, 3'', 7-, and 6beta,7-dihydroxy-delta1-THC; 2'',6alpha,7-, and 3'',6alpha,7-trihydroxy-delta1-THC; 2''-, 3''-, and 7-hydroxy-6-oxo-delta1-THC, and 2'',7- and 3'',7-dihydroxy-6-oxo-delta1-THC. The ketones and hydroxyacids were reduced to common alcohols with lithium aluminium deuteride and the number of deuterium atoms in the product was used to distinguish the metabolic alcohols from those produced by reduction.


Neutral in vivo metabolites of cannabinol isolated from rat faeces. The in vivo transformation of cannabinol (CBN) in the rat has been studied. Unchanged CBN and nine neutral mono-oxygenated and dioxygenated CBN metabolites have been identified. In the mono-oxygenated series the metabolites occurred in decreasing order of prominence as follows: 7-hydroxy-CBN, 4''-hydroxy-CBN, 1''-hydroxy-CBN, 2''-hydroxy-CBN, 3''-hydroxy-CBN, 5''-hydroxy-CBN and CBN-7-al. In the dihydroxylated metabolite series only 1'',7-dihydroxy-CBN and 4'',7-dihydroxy-CBN were found with the former as the more prominent metabolite.


Cannabinoid patterns in seedlings of Cannabis sativa L. and their use in the determination of chemical race. We have examined the cannabinoid contents of seedlings from twelve strains of Cannabis of known chemical race. Fourteen days after emergence of the shoot those of the tetrahydrocannabinol (THC) type could be distinguished from those of the cannabidiol (CBD) type. The true leaves of the THC type contained a relatively high content of THC and also contained cannabichromene (CBC), sometimes as the major cannabinoid, whereas the CBD type had much lower amounts of THC and no CBC; CBD being the major component. A strain from China corresponded to the THC type but showed some unusual features. Seeds purchased at seven outlets in Britain were also examined. Only two batches germinated by these proved to be of a THC type and resembled the Chinese strain.


Degradation of carmustine in aqueous media. The degradation rate of carmustine was investigated in buffered aqueous media at several pH values. The buffering agents studied were those with potential use in parenteral formulations of this drug: acetate, citrate, and phosphate. The apparent first-order degradation rate constants were calculated using a linear regression procedure. A pH range over which minimum degradation occurred was ascertained. General acid and specific base catalysis was demonstrated for the degradation of carmustine. From the data at 5, 22, and 37 degrees, the apparent activation energies for carmustine degradation in buffered aqueous media were computed and were strongly pH dependent.


Spectrophotometric determination of cephapirin, a cephalosporin antibacterial. A simple and specific method for the quantitative determination of cephapirin, a cephalosporin antibacterial, in finished bulk and dosage forms is reported. The method is based on reproducible degradation under controlled conditions to an unidentified species, which is measured spectrophotometrically at 375 nm. The procedure can be performed manually on a short series of samples in about 15 min or can be automated for large runs. Precursors and related substances show minimal interference. The coefficient of variation of the automated system is about 1% within days and 1.3% among days.


High-pressure liquid chromatographic analysis of cimetidine, a histamine H2-receptor antagonist, in blood and urine. A method is described for extraction of cimetidine, a histamine H2-receptor antagonist, from whole blood and urine with subsequent analysis by high-pressure liquid chromatography (HPLC). The drug is extracted from biological fluids with 1-octanol and back-extracted into dilute acid and then into a small volume of ethanol by saturation with potassium carbonate. HPLC analysis is performed on a column of 5-micrometer silica with a mixed mobile phase consisting primarily of acetonitrile. The method measures concentrations of cimetidine as low as 0.05 microgram/ml and is reproducible. Blood levels and urinary excretion data obtained with the analytical procedure are given for a group of human subjects who received 200-mg oral doses of cimetidine.


Rate and proposed mechanism of anhydrotetracycline epimerization in acid solution. The pathway through which the toxic tetracycline degradation product epianhydrotetracycline forms in solution was studied using high-performance liquid chromatography and circular dichroism, taking advantage of the large difference in ellipticity between the reactant and the product at 285 nm. The epimerization of anhydrotetracycline followed a reversible first-order process, and both analytical methods yielded the same rate constants. The rate constants indicate that anhydrotetracycline epimerization is faster than tetracycline epimerization. The equilibrium favored anhydrotetracycline, and the activation energies for the forward and reverse rates were almost the same as those for tetracycline epimerization. The epimerization was catalyzed by phosphate. Activation energies in 0.1 and 1 M phosphate were essentially the same. The equilibrium constants for both anhydrotetracycline and tetracycline favored the natural configuration rather than the epi series. Possible rationalization based on conformational and hydrogen bonding effects is presented.


Fluorometric determination of nadolol in human serum and urine. To determine nadolol, a new beta-adrenergic blocking agent, in serum and urine, the drug is extracted into n-butyl acetate or ether from alkaline potassium chloride saturated samples. After back-extraction into 0.1 N HCl, the drug is oxidized with periodic acid; the resulting aldehyde is coupled with o-phenylenediamine to produce a fluorescent compound. The method can measure as little as 0.01 microgram of nadolol/sample.


Degradation kinetics of a substituted carbinolamine in aqueous media. The apparent first-order breakdown of the medicinally active agent 3-(p-chlorophenyl)-2-ethyl-2,3,5,6-tetrahydroimidazo[2,1-b]thiazol-3-ol was studied in aqueous solutions where dehydration gave the unsaturated compound 3-(p-chlorophenyl)-5,6-dihydro-2-ethylimidazo[2,1-b]thiazole. This thiazole was the primary solvolytic product produced in approximately quantitative yields for the temperature range studied and ostensibly underwent no further reaction in acidic media even on prolonged heating. Investigations were carried out at various pH values in standard buffers at constant ionic strength. The ionization constants of the compounds are reported as well as the apparent activation energies for the degradation in acid and acetate buffers. The influence of ionic strength on the velocity constant was determined.


An in vivo assay for the analysis of the biological potency and structure-activity relationships of narcotics: serum testosterone depletion in the male rat. An in vivo method for the assessment of the agonistic-antagonistic activities and structure-activity relationships of the narcotics is described. The method, which is based on the depletion of serum testosterone levels by the narcotics in the male, satisfies all of the criteria necessary for a valid narcotic assay: 1) The ability of the narcotics to deplete serum testosterone levels is stereospecific; 2) naloxone competitively inhibits their effects; 3) and the relative potencies of the drugs in depressing serum testosterone levels parallels their activity in inhibiting electrically induced contractions of the guinea-pig ileum, the binding of [3H]opiates to "receptors" in rat brain homogenates and their relative effectiveness as analgesics. Although only rats were used in the current studies, the method can be easily adapted for use in any species, including man.


Direct and beta adrenergic blocking actions of nadolol (SQ 11725) on electrophysiologic properties of isolated canine myocardium. Effects of nadolol, a new beta adrenergic antagonist, were determined on transmembrane potentials of canine Purkinje fibers and ventricular and atrial muscle. Significant alterations in Purkinje fiber potentials occurred only with nadolol concentrations of 10(-4) M or greater. After 1 hour exposure to 10(-4) M, resting potential and action potential amplitude were reduced; at 3 hours, action potential rate of rise, phase 2 duration and action potential duration at 75% repolarization also were decreased. 10(-3) M nadolol also was depressant and, additionally, consistently reduced membrane responsiveness. Nadolol (10(-8)-10(-4) M) had no significant effects on effective refractory period. Ventricular and atrial muscle were less sensitive to all drug concentrations. Nadolol (10(-8)-10(-4) M) had little action on automaticity in normal, ouabain-treated or stretched Purkinje fibers but markedly decreased catecholamine-enhanced automaticity. In isoproterenol-treated Purkinje fibers, pA2 for nadolol was 8.2 and 7.7 for propranolol. Nadolol (10(-3) M or less) did not affect isometric force developed by isolated canine atrial trabeculae. We suggest that direct membrane depressant effects of nadolol do not contribute to its antiarrhythmic activity and that its beta adrenergic blocking ability is beneficial in catecholamine-related cardiac ectopia.


Synthesis and stereospecific antipsychotic activity of (-)-1-cyclopropylmethyl-4-(3-trifluoromethylthio-5H-dibenzo[a,d]cyclohepten-5-ylidene)piperidine. The synthesis and resolution of 3-iodocyproheptadine [(+/-)-5a] and 1-cyclopropylmethyl-4-(3-iodo-5H-dibenzo-[a,d]cyclohepten-5-ylidene)piperidine [(+/-)-5b] are described. The resulting atropisomers undergo reaction with trifluoromethylthiocopper to give optically active products without extensive racemization. In this manner, optically pure (+)- and (-)-3-trifluoromethylthiocyproheptadine [(+)-6a and (-)-6a, respectively] and (+)- and (-)-1-cyclopropylmethyl-4-(3-trifluoromethylthio-5H-dibenzo[a,d]cyclohepten-5-ylidene)piperidine [(+)-6b and (-)-6b, respectively] have been prepared. The influence of a chiral europium shift reagent on the proton and fluorine resonance signals as a diagnostic tool for the determination of the optical purities of these atropisomers is discussed. The four compounds, (+)-6a, (-)-6a, (+)-6b, and (-)-6b, were studied in squirrel monkeys for their ability to block conditioned avoidance responding. All of the antiavoidance activity was found to reside solely in the levorotatory compounds (-)-6a and (-)-6b. Further comparison of the enantiomers (-)-6b and (+)-6b showed that the ability to antagonize apomorphine-induced stereotyped behavior is confined to the levorotatory isomer (-)-6b while weak central anticholinergic activity resides solely in the dextrorotatory isomer (+)-6b. Neither (-)-6b has significant peripheral anticholinergic activity.


An approach to peripheral vasodilator-beta-adrenergic blocking agents. The syntheses of 2-phenyl- and 2-pyridyl-4-trifluoromethylimidazoles having a 3-tert-butylamino-2-hydroxypropoxy moiety attached to the aryl or heteroaryl substituent are described. Structure--activity relationships based on results from an evaluation of these compounds for antihypertensive, vasodilating, and beta-adrenergic blocking activities are discussed.


Adrenergic agents. 6. Synthesis and potential beta-adrenergic agonist activity of some meta-substituted p-hydroxyphenylethanolamines related to salbutamol. Salbutamol, an adrenergic receptor agonist with selectivity for tracheobronchial vs. cardiac muscle, differs from the catecholamine N-tert-butylnorepinephrine in that it bears a hydroxymethyl, rather than a phenolic, group in the meta position. In a search for new bronchodilating agents with minimal cardiovascular side effects, a series of derivatives, in which this m-hydroxymethyl group is modified, was prepared. These compounds were examined for potential bronchodilator activity in an in vitro test that measures relaxation of guinea pig tracheal smooth muscle. Potential cardiac stimulant activity was evaluated in vitro by monitoring changes in the rate of contraction of spontaneously beating guinea pig right atria. Although many of these compounds retained a high degree of potency, all were less effective than salbutamol in the tracheal test. Several of the derivatives, notably ones bearing 1-hydroxyethyl (1d), 1,2-dihydroxyethyl (1f), 1-hydroxy-2-methoxyethyl (1g), and 2-hydroxy-1-methoxyethyl (1h) substituents in place of the parent's m-hydroxymethyl group, however, were considerably more selective for tracheobronchial vs. cardiac muscle in the in vitro tests utilizing guinea pig tracheal and right atrial muscle.


Central nervous system activity of a novel class of annelated 1,4-benzodiazepines, aminomethylene-2,4-dihydro-1H-imidazo[1,2-a][1,4]benzodiazepin-1-ones. The synthesis and CNS activity of a noval class of annelated 1,4-benzodiazepines, the aminomethylene-2,4-dihydro-1H-imidazo[1,2-a][1,4]benzodiazepines, are described. An investigation of the structure--activity relationships in the series derived from 8-chloro-2,4-dihydro-2-dimethylaminomethylene-6-phenyl-1H-imidazo[1,2-a][1,4]-benzodiazepin-1-one (10) led to the synthesis of a group of compounds with potent minor tranquillizer activity.


Congenital total hemihypertrophy and carcinoma of undescended testicle: a case report. The first case of germinal cell tumor of the testis in a patient with congenital total hemihypertrophy is reported. The literature is discussed with emphasis placed on the frequent association of hemihypertrophy with oncogenesis and teratogenesis. We conclude that because of the high incidence of malignancy in the undescended testis prophylactic orchiectomy should be considered in a patient with hemihypertrophy and cryptorchidism.


Evaluation of cimetidine (tagamet). An antagonist of hydrochloric acid secretion. Cimetidine, a potent H2-receptor antagonist, has been introduced for the treatment of various conditions associated with acid peptic digestion in the gastrointestinal tract. Preliminary results are encouraging. Adverse reactions do not appear to be substantial; however, the effect of long-term use has yet to be ascertained.


Neural blockade by local anesthetics.

Neural blockade by local anesthetics. Local anesthetics block nerve impulse propagation by occluding transmembrane sodium channels, so preventing depolarization. First, the uncharged lipid-soluble anesthetic base pentrates the membrane; then the positively charged cation binds to anionic components of the sodium channel's internal axoplasmic mouth. Though primarily a carrier, the base contributes to blockade by causing the membrane to swell, so pinching the sodium channels. Dissolved in water, local anesthetic salt crystals dissociate into anesthetic cation and base-proportional to the drug's fixed pKa and the tissue's variable pH. The cation-base concentration ration is critical to optimal neural blockade. If there is too little base, few anesthetic molecules will penetrate to the neural target; if too little cation, few sodium channels will be plugged.


Aspiration of metallic mercury. A 22-year follow-up. Rupture of the mercury-filled bag of an intestinal tube resulted in aspiration of metallic mercury 22 years ago in the patient reported. Immediate respiratory distress was treated with vigorous suctioning and postural drainage. Follow-up chest roentgenograms persistently showed radiopaque particles in the lungs. Signs and symptoms of chronic respiratory disease have since developed in the patient. Postmortem findings from the lungs included globules of metallic mercury surrounded by extensive fibrosis and granuloma formation, which we believe to be the result of local irritative effects of mercury.


L-asparaginase--antileukemia agent.

L-asparaginase--antileukemia agent. This paper describes the most recent development in the field of biosynthesis of L-Asparaginase by means of various culture, purification by Column Chromatography, informations of the Catabolite repression of L-Asparaginase and its metabolism in mycobacteria. The separation, purification studies, idea about the effect of different PH on activity, quaternary structure and the effect of specific antibodies on catalytic activity of L-Asparaginase is discussed. This material shows the role of L-Asparaginase in acute lymphocytic leukemia.


Pathogenesis of vascular injury in rejecting rat renal allografts. Unmodified rejection of rat renal allografts was characterized by the early onset and rapid progression of endothelial damage in venules and capillaries which culminated in ischemic cortical necrosis. This pattern of endothelial injury correlated with lymphocyte accumulation in vascular lumens and could not be duplicated by renal perfusion with alloantibodies or prevented by C'3 depletion. In contrast, endothelial integrity and normal graft function were maintained over study intervals extending to 200 days when Brown Norway (BN) rat kidneys were transplanted into Lewis (Le) rat kidney recipients subjected to neonatal thymectomy or lymph drainage. Vascular lesions occurred when syngeneic thoracic duct lymphocytes were transfused into these recipients, but irreversible endothelial injury could be prevented by simultaneous injections of immune plasma. These findings indicate that the destruction of donor endothelium is mediated by thymus-dependent immune mechanisms which can be altered by thoracic duct drainage to promote indefinite survival of renal allografts across major histocompatibility loci.


[Hemodynamic changes after angiotensin II blockade by saralasin (author's transl)]. Saralasin, an angiotensin II inhibitor was infused in 10 hypertensive patients. A blood pressure reduction was achieved after stimulation of the renin-angiotensin-system by salt depletion. Heart rate and cardiac output failed to compensate for reduction of blood pressure. Thus circulatory reflex-mechanisms are inhibited by saralasin. A direct influence on baroreceptor mechanism and/or catecholamines is probable. Failure of the hypotensive effect of saralasin in salt-depleted patients after administration of beta-blockers supports this hypothesis.


Fine structural changes in dog myocardium exposed to lowered pH in vivo. A surgical preparation that allows chemical alteration of the blood supplied to specific area of the intact heart was used to investigate the effects of lowered tissue pH on the fine structure of myocardial cells. Controlled infusion of isotonic saline acidified with HCl to a pH value between 1.0 and 2.0 produced a lowering of blood pH to values of less than 7.0. This drop in the pH of the perfusing blood resulted in a corresponding decrease in pH of the relevant area of myocardium. When this decrease occurred without a concomitant increase in tissue lactate level, the myocardial cells of the acid-perfused area showed focal glycogen depletion, moderate relaxation of myofibrils and clumping of nuclear chromatin, and mild mitochondrial change, which did not include marked swelling although intramitochondrial electron-dense inclusions were often detectable. When the fall in pH was accompanied by a large increase in lactate concentration, however, the cells showed more severe glycogen depletion and clumping of nuclear chromatin, and mitochondrial change, which did include marked swelling, together with the presence of well developed inclusions.


Metal complexes of poly(alpha-amino acids). A potentiometric and circular dichroism investigation of Cu(II) complexes of poly(L-lysine), poly(L-ornithine), and poly(L-diaminobutyric acid). The conformational properties of cupric complexes of poly(L-lysine), poly(L-ornithine), and poly(L-diaminobutyric acid) have been investigated by potentiometric, visible and UV absorption, and circular dichroism (CD) techniques. The three polymers form two kinds of complexes stable at pH less than 8.5 (type I complexes) and at pH less than 8.5 (type II complexes). It has been found that in the low pH complexes of poly(L-diaminobutyric acid) at least one deprotonated amido nitrogen is coordinated to cupric ions. Type II complexes involve always amide nitrogens in the coordination sphere of Cu(II). Evidence is presented that the structure of such complexes is not compatible with the alpha-helical conformation of the peptide backbone.


The microbiology of chronic middle ear effusions in children. Serous otitis media is the most common cause of hearing loss in childhood. Traditionally, this entity has been felt to be a sterile process. This study was undertaken to determine the incidence of infected middle ear fluid in patients with asymptomatic middle ear effusions. The external auditory canal, nasopharynx and middle ear fluids of 57 patients undergoing myringotomy were cultured. Of 57 patients, 26 had positive cultures; 67% of patients less than 36 months of age had positive culture, with a bacterial flora closely resembling that of acute otitis media. Further investigation is required to determine the significance of such infection together with the optimum therapy for chronic effusions of the middle ear in young children.


[Pathogenesis of hepatic encephalopathy (author's transl)]. This contribution presents data from the literature as well as our own results concerning the mechanisms of hepatic encephalopathy (HE). 1. Blood chemistry: In patients with liver cirrhosis, the plasma levels of ammonia, phenylalanine, tyrosine, phenolic acids, and octopamine correlated with the stages of HE. Methionine and free tryptophan concentrations were increased only in stages 2-4. Further, branched chain amino acids were below the normal range. Experimental findings in animals elucidated some mechanisms of these changes. 2. Effects of administered substances: With ammonia, methionine, methanethiol, tryptophan, phenolic substances, and fatty acids central nervous disturbances were observed. 3. Interactions: Anemia, methanethiol, and fatty acids favored ammonia toxicity. Alkalosis diminished cerebral symptoms. 4. Neurotransmitters: HE was accompanied by an enhanced turnover of serotonin and by increased amounts of false neurotransmitters (like octopamine) in the brain. 5. Oxydative brain metabolism: Disorders of cerebral oxygen and glucose utilization were mainly documented in cases of long term HE with EEG alterations. 6. Structural changes of the brain: Most of them are irreversible.


Effects of cyanide on peanut lipoxygenase. Acidic and alkaline lipoxygenase isozymes were separated and assayed for activity while incubated in the presence of various concentrations of cyanide. Inhibition due to cyanide per se was not found. Instead, cyanide caused an increase in pH, which decreased the activity of the acidic isozyme to a rate corresponding to the same pH on the activity curve for the cyanide-free reaction.


Alkaloids of papaver genus IX. Alkaloids of Glaucium vitellinum Boiss and Buhse, population Seerjan and Glaucium pulchrum Stapf, population Elika. Glaucium vitellinum Boiss and Buhse. population Seerjan was shown to contain three major alkaloids, isocorydine (0.44%), protopine (0.42%), dicentrine (0.24%), and four minor alkaloids, tetrahydropalmatine (0.13%), muramine (0.12%), bulbocapnine (0.06%) and glaucine (0.01%). Glaucium pulchrum Stapf popllation Elika was shown to contain two major alkaloids, corydine (0.3%) and bulbocapnine (0.18%) and three minor alkaloids N-methylindcarpine (0.1%), isocorydine (0.03%) and protopine (0.01%). N-methyllindcarpine was found for the first time in the Papaveraceae and tetrahydropalmatine was detected for the first time in Glaucium.


Simple assay for the condensation component enzyme (beta-ketoacyl synthetase) of fatty acid synthetase. A simple assay is described for estimating the activity of the condensation component enzyme (beta-ketoacyl synthetase) of the yeast fatty acids synthetase complex. The radioactivity liberated as 14CO2 from [1,3-14C]malonyl-CoA was trapped in phenethylamine and measured by liquid scintillation spectroscopy. Three enzyme-catalysed steps are involved: acetyl-CoA transacylase, malonyl-CoA transacylase and beta-ketoacyl synthetase; however, beta-ketoacyl synthetase is rate-limiting. beta-Ketoacyl synthetase activity was made independent of subsequent enzyme activities of the complex by excluding NADPH from the assay, thus blocking beta-ketoacyl reductase and preventing fatty acid synthesis. By this assay beta-ketoacyl synthetase activity was about 0.28 of the activity of the complex for fatty acid synthesis, compared with approximately 0.001 for published assays. Several pyridine nucleotides and derivatives were tested after it was discovered that NADH stimulated beta-ketoacyl synthetase activity to a greater extent than could be accounted for by its reactivity in providing a pathway from acetoacetyl-enzyme to fatty acid synthesis. Presumably, the release of acetoacetate from the central sulphydryl of the complex is the rate-limiting step in the assay procedure.