Membranes of Rhodopseudomonas sphaeroides. V. Identification of bacteriochlorophyll alpha-depleted cytoplasmic membrane in phototrophically grown cells. The separation of membrane fragments was investigated in extracts of phototropically grown Rhodopseudomonas sphaeroides to determine if the plasma membrane contains discrete regions. A highly purified fraction of bacteriochlorophyll alpha-deficient membrane fragments was isolated by differential centrifugation, chromatography on Sepharose 2B, reaggregation, and isopycnic sedimentation on sucrose gradients. Significant levels of b- and c-type cytochromes and succinate dehydrogenase were demonstrated in the isolated membrane fragments and their appearance in electron micrographs, their polypeptide profile in dodecyl sulfate-polyacrylamide gel electrophoresis, and overall chemical composition were essentially identical to a similar fraction isolated from aerobically grown cells. Their polypeptide profiles were distinct from those of the intracytoplasmic chromatophore and outer membranes, and on the basis of bacteriochlorophyll content the phototrophic fraction was contaminated with chromatophores by less than 9%. The membrane fragments contained no diaminopimelic acid or glucosamine. It is condluded that the membrane fragments isolated from phototrophically growing Rp. sphaeroides have arisen from photosynthetic pigment-depleted regions of the plasma membrane structurally and functionally differentiated from the intracytoplasmic chromatophore membrane. These regions represent conserved chemotrophic cytoplasmic membrane whose synthesis continues under photoheterotrophic conditions.


Identification of the globin chain and the amino acid residue responsible for polymerization of bullfrog (Rana catesbeiana) hemoglobin with iodo[14C]acetamide. The bullfrog (Rana catesbeiana) major hemoglobin dissociates into its constituent globin chains (alpha and beta) which are separated by Sulfopropyl-Sephadex C-25 column chromatography after alkylation with iodo[14C]acetamide. Each globin chain has two cysteine residues and those of the beta-globin chain in the tetramer are preferentially alkylated with iodoacetamide.


Application of cell electrophoretic methods (preparative and analytical) in demonstrating the surface heterogeneity of T-lymphocytes in man. The preparative electrophoretic method allows separation of several cell populations, which can be characterized by the usual membrane markers. This qualitative study shows a heterogeneous distribution of the T-lymphocytes from the most rapid to the slowest migration zones. The electrophoretic mobility (E.M.) of each fraction has been studied by analytical electrophoresis, which confirms the heterogeneity of T-cells. The experiments used total blood lymphocytes and rosette-forming cells with sheep red blood cells. The results obtained with the two methods were well correlated. The average migration speed of the separated populations showed a regular variation from one extreme fraction to the other. The E.M. speeds that characterize the main populations are--0.90 micrometer.sec.--1v.--1cm for the B-lymphocytes; for the T-lymphocytes,--1.10 (previously identified as the "low affinity" E-rosette FC),--1.20 and a small population centered around--1.35 (corresponding to active E rosette FC). In the T-lymphocytes separated after E-rosetting, the progresssive variation in the mean E.M. of each fraction is confirmed, as well as the existence of the previous 3 T-cell populations. Further studies have to be done, to show that each separated population does correspond with a functional differentiation.


Cytochrome P-450 from mitochondria of bovine adrenal cortex: comparison of cholesterol side-chain cleavage P-450 with steroid 11beta-hydroxylation P-450 and immunochemical cross-reactivity between adrenal mitochondrial and liver microsomal cytochromes P-450. Adrenocortical mitochondrial cytochrome P-450 specific to the cholesterol side-chain cleavage (desmolase) reaction differs from that for the 11beta-hydroxylation reaction of deoxycorticosterone. The former cytochrome appears to be more loosely bound to the inner membrane than the latter. Upon ageing at 0 degrees C or by aerobic treatment with ferrous ions, the desmolase P-450 was more stable than the 11beta-hydroxylase P-450. By utilizing artificial hydroxylating agents such as cumene hydroperoxide, H2O2, and sodium periodate, the hydroxylation reaction of deoxycorticosterone to corticosterone in the absence of NADPH was observed to a comparable extent with the reaction in the presence of adrenodoxin reductase, adrenodoxin and NADPH. However, the hydroxylation reaction of cholesterol to pregnenolone was not supported by these artificial agents. Immunochemical cross-reactivity of bovine adrenal desmolase P-450 with rabbit liver microsomal P-450LM4 was also investigated. We found a weak but significant cross-reactivity between the adrenal mitochondrial P-450 and liver microsomal P-450LM4, indicating to some extent a homology between adrenal and liver cytochromes P-450.


Isolation of glycosaminoglycans from lymphocytes in chronic lymphoic leukemia. Glycosaminoglycans were isolated from type B chronic lymphoid leukemia lymphocytes. Their studies showed chondroitin sulfate A (C) and under-sulfated chondroitin sulfate, no heparan sulfate was found. These results were confirmed with 12 other chronic lymphoid leukemia patients. By the same method polyanionic glycopeptides distinct from glycosaminoglycans were obtained from lymphocyte glycoproteins. They showed similarities in composition with known cell receptors. The largest part of chondroitin sulfate was liberated from cells by mild hydrolysis with trypsin and thus appeared to be a cell-surface component.


Trasylol (aprotinin) as an anti-tumour agent. Growth of a syngeneic adenocarcinoma in C3H mice was investigated in animals receiving twice-daily subcutaneous injections of the antiprotease Trasylol. Development of primary tumours was not influenced by Trasylol treatment and the antiprotease did not affect concomitant ant--tumour immunity as assessed by resistance to secondary footpad challenge. From histological examination of tumours and a study of GVH (graft-versus-host) activity no evidence was gained that Trasylol treatment augments the host cellular immune response.


Differential effect of hydrocortisone on lymphocyte populations in the mouse spleen. The cellular composition of the spleen from CBA mice was investigated two days after administration of increasing doses of hydrocortisone acetate (HC), this using the presence of surface-immunoglobulin and the electrophoretic mobility as criteria for identification of lymphocyte populations. The B cell content was found to decrease more rapidly than did the T cell content. However, a significant diminution of the modal volume of both B and T cells was observed which suggests that HC acted similarly on each of these lymphocyte compartments by removing the larger elements. On the other hand, the in vitro reactivities to the B cell mitogens LPS and PPD were strongly depressed by HC injection while the reactivities to the T cell mitogens Con A and PHA remained relatively unchanged. Therefore, HC treatment appears to affect both B and T lymphocyte populations in the mouse spleen but the former to a greater extent than the latter.


In vitro restoration by thymosin of T lymphocytes from patients with lung tuberculosis. The percentage of E-rosette forming T-lymphocytes is considerably decreased in a number of patients with severe form of lung tuberculosis, particularly in those with tuberculous empyema of the pleura (28.5% to 38%). After in vitro incubation of lymphocytes from those patients with thymosin V fraction T-lymphocytes restored normal.


Birefringence measurements of structural inhomogeneities in Rana pipiens rod outer segments. The birefringence (deltan) of Rana pipiens rod outer segments (ROS) reveals microstructure inhomogeneities not seen with other techniques. In the basal 20-30-micron length of the ROS there is a nearly linear axial gradient in deltan of approximately equal to -2 x 10(-5)/micron. No consistent deltan gradients are found in the distal 20-30 micron of the ROS. Using glycerol imbibition to separate the intrinsic and form birefringence components, we found that the basal deltan gradient was principally due to a gradient of the intrinsic birefringence component. The disk membrane volume fraction decreases uniformly from the basal end to the distal end, while the disk membrane refractive index increases. The contributions of these changes to the form birefringence approximately cancel, so that the form component is fairly uniform along the ROS axis. Because its axial distance from the inner segment is a measure of the time since a disk membrane was formed, these gradients may reflect a disk membrane aging process. Occasionally a highly birefringent, 2-micron-wide band is seen at the basal end ot the ROS, possibly where the envelope membrane folds to form new disk membranes.


[Calcium ion uptake by the skin fibroblast culture under normal conditions and in systemic diseases of the connective tissue]. Radioactive calcium (45Ca) uptake and the effect of adrenaline on this process was studied on the suspension of cells derived from the monolayer cultures of fibroblasts of the skin of healthy donors and of patients with systemic scleroderma and rheumatic fever. A significant increase of calcium uptake was shown in case of scleroderma in comparison with the normal and rheumatism cases. Adrenaline (2 X 10(-7)--2 X 10(-5) M) enhanced the rate of calcium uptake by fibroblasts in control and had the opposite effect in scleroderma. The data obtained indicated alteration of the fibroblast membrane function in systemic scleroderma which could underlay the change in the metabolic activity of these cells.


[Tolerance-like condition developing in mice under the effect of antigen of erythrocyte lysate]. Lysate of sheep red blood cells obtained by the treatment of these cells with distilled water and purified by ultracentrifugation in cold possessed a weak immunogenicity. Its administration to mice caused the state of hyporeactivity to sheep red blood cells (a reduction of the immune response level to 10-25% of control. The capacity of the mise spleen cells to respond by immune reaction to the red blood cells following adoptive transfer was not disturbed. At the early periods after the lysate administrations the mouse spleen cells possessed a weak supressive activity in case of their transfer to the intact animals. The blood serum of mice treated with the lysate possessed a blocking activity which disappeared after the serum absorption with sheep red blood cells. A conclusion was drawn that hyporeactivity originating in mice after the lysate administration was caused by the presence in the serum of antibodies inhibiting the immune response.


[Role of thymus-dependent cells in splenic colony formation]. It was shown that treatment of the bone marrow with the serum reacting with the theta-antigen, irrespective of the presence of the complement, sharply decreases the capacity of its cells to form splenic colonies. Administration of the thymus cells together with the bone marrow to the recipient largely elminates the effect of this system, significantly increasing the splenic colony formation. It is supposed that the antiserum in the bone marrow inactivates the population of cells necessary for the formation of the colonies in the spleen, but differing from the pluripotent stem cells, possibly the population of T-cells.


Ultrastructural study of acute lymphocytic leukemia: comparison with immunologic studies. Leukemic cells from 29 cases of acute lymphocytic leukemia (ALL), studied for T and B cell markers by the use of sheep erythrocyte rosetting and surface immunoglobulin determinations, were examined by electron microscopy. The majority of patients (76%) were found to have non-T, non-B neoplasms composed predominantly of relatively small, inactive-appearing cells with frequent nuclear folds. T cell cases (21%) were associated with mediastinal masses and were predominantly composed of large, active-appearing cells with nuclear irregularity and little rough endoplasmic reticulum. One case of B cell origin was not morphologically distinct from the non-T, non-B cell cases.


The local anaesthetic activity of a benzotriazinium salt. 1 The local anaesthetic properties of 2-n-propyl-4-p-tolylamino-1,2,3-benzotriazinium iodide (TnPBI) were compared with those of lignocaine hydrochloride on intact and desheathed sciatic nerves of the frog, on the phrenic nerve-hemidiaphragm preparation of the rat, and by the intradermal wheal test in the guinea-pig. 2 Both TnPBI and lignocaine were more potent on desheathed than on intact sciatic nerves. The potency of TnPBI was affected more than that of lignocaine by the presence of the sheath in intact nerves. 3 Both drugs inhibited conduction in the rat phrenic nerve, as shown by the reduction in twitch tension of the diaphragm elicited by nerve stimulation. TnPBI also caused an initial augmentation of the twitch tension of the diaphragm when applied directly to the muscle. 4 TnPBI was shown to be approximately twice as potent as lignocaine by the guinea-pig intradermal wheal test. 5 These results are discussed in view of the known effects of TnPBI on intracellular calcium storage.


Neurotrophic control of RNA synthesis in amphibian striated muscle. When frog sartorius muscles are incubated in culture for 3 days, a 98% increase in RNA synthesis by the muscles is seen. A portion of this increase (60--100% in different experiments) is prevented by the presence of a 2 cm distal nerve stump left attached to the muscle, or by the addition to the culture medium of nerve and spinal cord extracts from frog, rat or cow. The effect of the nerve stump and the extract is seen under conditions of neuro-muscular blockade by 50 mg/l gentamicin sulfate present in the culture medium. Autoradiography of 1 micrometer sections of muscles previously labeled with [3H]uridine shows that isotope incorporation is largely confined to muscle fibers rather than connective tissue. Muscles cultured for 3 days display normal action and resting potentials either in the presence or absence of extract. The results are discussed in terms of control of muscle RNA synthesis by substances present in motor neurons.


Differential synthesis of the polypeptides of aldehyde dehydrogenase and NAD(P)H:flavin oxidoreductase in the bioluminescent bacterium Beneckea harveyi. The proteins of the bioluminescent bacterium Beneckea harveyi have been labelled with [3H]leucine prior to the induction of bioluminescence, and with [14C]leucine during the development of the bioluminescent system. An aliphatic aldehyde dehydrogenase and a NAD(P)H:flavin oxidoreductase, two enzymes that may be directly involved in the metabolism of the substrates (aldehyde, FMNH2) for the luminescent reaction catalyzed by luciferase, were purified and the isotope ratios of their respective polypeptide chains determined after sodium dodecyl sufate gel electrophoresis. A comparison of these isotope ratios to (a) the isotope ratios of the induced polypeptide chains of luciferase, purified in the same experiment, and (b) the average isotope ratio for the proteins synthesized in concert with growth has provided direct evidence that the synthesis of aldehyde dehydrogenase but not NAD(P)H:flavin oxidoreductase is induced during the development of bioluminescence.


Antagonism by phosphate buffer of the twitch loss in isolated muscle fibers produced by calcium-free solutions. In 37 of 41 isolated frog skeletal muscle fiber preparations (one, two, or three fibers) the twitch was eliminated or reduced to less than 10% of control by exposing the fibers to a O-calcium, bicarbonate-buffered solution for 10 min or less. Replacing the bicarbonate by a phosphate buffer either prevented twitch inhibition or increased the O-calcium exposure time required for its production. It is concluded that surface membrane-bound calcium ions (presumably in the t-tubules) are required to couple the action potential to the mechanical response and that phosphate ions inhibit the loss of the membrane-bound calcium ions into an external calcium-free solution.


Flocculation and adsorption of enzymes during growth of a moderate halophile, Micrococcus varians var. Halophilus. Flocculation of a moderate halophile, Micrococcus varians ATCC 2197, occurred during growth in complex medium containing 3 M NaCl and a concentration of MgSO4 and KH2PO4 greater than 40 and 14 mM, respectively. Extracellular nuclease activity was absent in the flocculated cultures. Repeated washing of flocs by Mg2+-free Tris buffer containing 3 M NaCl, lowering of pH value of floc suspension below 6.3, or addition of ethylenediaminetetraacetic acid resulted in complete dissociation of the flocs and release of Mg2+ ions as well as nuclease and amylase. Inhibition of extracellular enzyme production accompanied by flocculation appeared to be the result of adsorption of enzyme proteins to surfaces of the flocs, but not of inhibition of biosynthesis. Floc formation could also occur in media containing 18 mM CaCl2 and 3.0 mM KH2PO4, but the Ca flocs were not deflocculated by washing with Ca2+-free buffer, suggesting that the affinity of Ca2+ for cell envelopes was stronger than that of Mg2+. It was also observed that most halophilic Planococcus and Micrococcus flocculated in the presence of MgSO4 and phosphate but halophilic Pseudomonas, Acinetobacter, and Bacillus did not.


Potentiation by 2'-deoxycoformycin of the inhibitory effect by 3'-deoxyadenosine (cordycepin) on nuclear RNA synthesis in L1210 cells in vitro. The effect of the adenosine deaminase inhibitor, 2'-deoxycoformycin, on the inhibitory effect of cordycepin on nuclear RNA synthesis was examined in L1210 cells in vitro. The median inhibitory dose for the effect of deoxycoformycin on adenosine deaminase was 4 X 10(-8) M, and 100% inhibition was achieved at 5 X 10(-7) M. Pretreatment of cells for 30 min with 1 X 10(-6) M deoxycoformycin resulted in a reduction of the median inhibitory dose for cordycepin from 2.5 X 10(-4) to 1.8 X 10(-5) M, as assessed by the measurement of [3H]uridine incorporation into total RNA. Measurement of the synthesis of nuclear ribosomal RNA, nonpolyadenylic acid heterogeneous RNA, and polyadenylic acid heterogeneous RNA revealed potentiation by 2'-deoxycoformycin of the inhibitory effect of cordycepin on all species of RNA, as well as on polyadenylic acid synthesis. No differences were noted in the size of nuclear polyadenylic acid obtained from cells treated with cordycepin in either the presence or the absence of the adenosine deaminase inhibitor. These results suggest that the potentiation by 2'-deoxycoformycin of the cytotoxic and antitumor effects of cordycepin on L1210 cells in vivo is related to inhibition of nuclear RNA synthesis.


Role of intracisternal A-particles and of RNA molecules in plasmacytoma-associated immunodeficiency. A subcellular fraction from murine plasmacytoma cells was shown to suppress the primary antibody response when injected into normal mice. The active subcellular fraction copurified with intracisternal A-particles. The RNA extracted from subcellular fractions enriched in A-particles was also immunosuppressive. This activity was due to a population of RNA molecules that contained polyadenylic acid. Upon fractionation on a sucrose gradient, two populations of immunosuppressive RNA were obtained with sedimentation velocities of 12 to 18S and 40 to 50S. The 40 to 50S RNA was shown to be a thermolabile aggregate of molecules that contained the 12 to 18S RNA molecules. Plasmacytoma-derived material with similar physicochemical characteristics had previously been shown to induce in normal mouse lymphocytes surface immunoglobulins with the plasmacytoma idiotype, supporting the possibility that one of the mechanisms responsible for the development of immunological deficiency is the change of surface immunoglobulins of nonmalignant B-cells.


Illness associated with "package tours": a combined Spanish-Scottish study. The occurrence of three deaths and substantial morbidity among members of a "package tour" who stayed at the same hotel in Benidorm, Spain, during the summer of 1973 provided an opportunity to study the experiences of persons undertaking this type of holiday and some of the factors that might affect their health. Of 252 tourists, 164 (65%) reported illness, 86 of those affected (52%) having respiratory symptoms. The three persons who died had predisposing factors and all had lobar pneumonia. In the control group of 157 persons, who stayed at other hotels in Benidorm, 82 (53%) experienced ill-health during their stay but fewer (38%) had respiratory illnesses. The study demonstrated some of the hazards encountered by the inexperienced traveller and also indicated the need to improve surveillance of what appears to be an unexpectedly large amount of illness among international tourists.


A comparison of Freon- and water-based insecticidal aerosols for aircraft disinsection. In Miami, USA, passenger compartments of wide-bodied jet aircraft were disinsected at "blocks away" or while parked. The application of 2% Freon- and water-based formulations of (+)-phenothrin at about 35 g/100 m(3) was highly effective against mosquitos and flies in all instances. More basic research is needed on water-based aerosols.


A simplified technique for counting onchocercal microfilariae in skin snips. A new method for counting microfilariae in skin snips is described. The method, in which slide specimens are reconstituted after drying, is compared quantitatively with two existing methods that are used when the counting of microfilariae has to be postponed. Counts obtained by the new method were closely correlated with those taken in fresh specimens. Other advantages of the new method are discussed and its use in onchocerciasis field surveys is recommended.


Evaluation of an experimental mollusciciding programme to control Schistosoma mansoni transmission in St Lucia. The size and number of colonies of Biomphalaria glabrata were reduced after four years of a surveillance/treatment snail control programme using an emulsifiable concentrate of niclosamide (25% active ingredient). Surveys among the human population showed that the incidence of new Schistosoma mansoni infections in 0-10 year-old children fell from 22% to 4.3%, while in a comparison area the incidence remained at 20%. With reduced transmission over four years, the prevalence of infection in a cohort of children examined in 1971 and 1975 fell from 34% to 23%. The fall in prevalence and intensity of infection led to a reduction of 66% in the index of potential contamination, which was reflected in a reduced rate of infection among sentinel snails and representative samples of B. glabrata collected during surveillance searches.The overall annual cost of the programme was US $3.24 per capita.


Advantages and disadvantages of killed and live poliomyelitis vaccines. Decision-making on the use of poliomyelitis vaccines in the WHO Expanded Immunization Programme, and particularly in the developing nations, needs to be based on an understanding of the epidemiology of poliomyelitis in different parts of the globe. Even with two safe and effective kinds of poliomyelitis vaccine available, poliomyelitis has by no means been eradicated from the world. In developed countries that are considered well-vaccinated, certain sectors of the population may be inadequately protected against risk of infection by indigenous or imported wild polioviruses. In developing nations that are in transition toward an epidemic phase of poliomyelitis, wild polioviruses will continue to be a threat until thorough immunization is established and maintained. Killed-virus poliomyelitis vaccines have proved to be effective in certain countries that have used them exclusively; these are small countries with excellent public health systems, where coverage by the killed vaccine has been wide and frequent. Live vaccines, administered to hundreds of millions of persons during the past decade, have also been remarkably safe and effective. However, in certain warm-climate countries induction of antibodies in a satisfactorily high proportion of vaccinees has been difficult to accomplish. The advantages and disadvantages of each kind of poliomyelitis vaccine need to be weighed with respect to the particular setting in which a vaccine has been or will be used.


Species complexes in the Simuliidae.

Species complexes in the Simuliidae. The recent discovery that Simulium damnosum in Africa is not a single species but a complex of sibling species has important implications for the epidemiology and control of onchocerciasis. This paper sets out the current situation with regard to classification within the S. damnosum complex and the smaller S. neavei group. The nomenclature of the S. damnosum complex and current morphological and enzyme electrophoretic methods of differentiating among sibling species are discussed. Finally, recommendations are made on priorities for future research.


Ocular onchocerciasis.

Ocular onchocerciasis. Well over 20 million people in the world are infected with Onchocerca volvulus and it is probable that 200 000-500 000 people are blind as a result of this infection, which is the most important cause of blindness in certain areas of Africa and Latin America.Treatment of the disease is difficult and often produces serious adverse reactions in the patient. Combined use of diethylcarbamazine citrate and suramin is still the most suitable form of treatment. Screening for the early detection of cases at high risk of ocular manifestations must be organized, and their treatment undertaken, if blindness is to be avoided.Prevention of ocular onchocerciasis is feasible, using vector control methods to reduce transmission, but the procedures are costly and may have to be maintained for many years.Research is needed to improve treatment and to find a chemoprophylactic agent or a preventive vaccine.


Occupational health hazards of mine workers. Mining has always been among the most hazardous of occupations, and with the increasing demand for coal and minerals safety in mines assumes even greater importance. This article describes the present situation with regard to conditions in mines, the diseases and disabilities resulting from them, and measures that can be taken to prevent or treat them. The hazards covered are: accidents, dust (including poisoning by certain ores), high temperature and humidity, noise and vibration, toxic gases, and miscellaneous other hazards.


Interferon and other antiviral agents, with special reference to influenza: a memorandum. Recent progress in the production of interferons and in knowledge of their mode of action has opened new possibilities for their prophylactic or therapeutic use, for example, in the treatment of influenza. Such use still needs further evaluation as do the presently available chemical antiviral drugs. The activity of interferon against other viral infections is also promising and preliminary trials on their effect on certain tumours have begun at a number of centres. There is, however, a need for quality control of interferon preparations and recommendations for the production and use of interferon as well as for further research are included in this Memorandum.


Ebola haemorrhagic fever in Sudan, 1976. Report of a WHO/International Study Team. A large outbreak of haemorrhagic fever (subsequently named Ebola haemorrhagic fever) occurred in southern Sudan between June and November 1976. There was a total of 284 cases; 67 in the source town of Nzara, 213 in Maridi, 3 in Tembura, and 1 in Juba. The outbreak in Nzara appears to have originated in the workers of a cotton factory. The disease in Maridi was amplified by transmission in a large, active hospital. Transmission of the disease required close contact with an acute case and was usually associated with the act of nursing a patient. The incubation period was between 7 and 14 days. Although the link was not well established, it appears that Nzara could have been the source of infection for a similar outbreak in the Bumba Zone of Zaire.In this outbreak Ebola haemorrhagic fever was a unique clinical disease with a high mortality rate (53% overall) and a prolonged recovery period in those who survived. Beginning with an influenza-like syndrome, including fever, headache, and joint and muscle pains, the disease soon caused diarrhoea (81%), vomiting (59%), chest pain (83%), pain and dryness of the throat (63%), and rash (52%). Haemorrhagic manifestations were common (71%), being present in half of the recovered cases and in almost all the fatal cases.Two post mortems were carried out on patients in November 1976. The histopathological findings resembled those of an acute viral infection and although the features were characteristic they were not exclusively diagnostic. They closely resembled the features described in Marburg virus infection, with focal eosinophilic necrosis in the liver and destruction of lymphocytes and their replacement by plasma cells. One case had evidence of renal tubular necrosis.Two strains of Ebola virus were isolated from acute phase sera collected from acutely ill patients in Maridi hospital during the investigation in November 1976. Antibodies to Ebola virus were detected by immunofluorescence in 42 of 48 patients in Maridi who had been diagnosed clinically, but in only 6 of 31 patients in Nzara. The possibility of the indirect immunofluorescent test not being sufficiently sensitive is discussed.Of Maridi case contacts, in hospital and in the local community, 19% had antibodies. Very few of them gave any history of illness, indicating that Ebola virus can cause mild or even subclinical infections. Of the cloth room workers in the Nzara cotton factory, 37% appeared to have been infected, suggesting that the factory may have been the prime source of infection.


Ebola haemorrhagic fever in Zaire, 1976. Between 1 September and 24 October 1976, 318 cases of acute viral haemorrhagic fever occurred in northern Zaire. The outbreak was centred in the Bumba Zone of the Equateur Region and most of the cases were recorded within a radius of 70 km of Yambuku, although a few patients sought medical attention in Bumba, Abumombazi, and the capital city of Kinshasa, where individual secondary and tertiary cases occurred. There were 280 deaths, and only 38 serologically confirmed survivors.The index case in this outbreak had onset of symptoms on 1 September 1976, five days after receiving an injection of chloroquine for presumptive malaria at the outpatient clinic at Yambuku Mission Hospital (YMH). He had a clinical remission of his malaria symptoms. Within one week several other persons who had received injections at YMH also suffered from Ebola haemorrhagic fever, and almost all subsequent cases had either received injections at the hospital or had had close contact with another case. Most of these occurred during the first four weeks of the epidemic, after which time the hospital was closed, 11 of the 17 staff members having died of the disease. All ages and both sexes were affected, but women 15-29 years of age had the highest incidence of disease, a phenomenon strongly related to attendance at prenatal and outpatient clinics at the hospital where they received injections. The overall secondary attack rate was about 5%, although it ranged to 20% among close relatives such as spouses, parent or child, and brother or sister.Active surveillance disclosed that cases occurred in 55 of some 550 villages which were examined house-by-house. The disease was hitherto unknown to the people of the affected region. Intensive search for cases in the area of north-eastern Zaire between the Bumba Zone and the Sudan frontier near Nzara and Maridi failed to detect definite evidence of a link between an epidemic of the disease in that country and the outbreak near Bumba. Nevertheless it was established that people can and do make the trip between Nzara and Bumba in not more than four days: thus it was regarded as quite possible that an infected person had travelled from Sudan to Yambuku and transferred the virus to a needle of the hospital while receiving an injection at the outpatient clinic.Both the incubation period, and the duration of the clinical disease averaged about one week. After 3-4 days of non-specific symptoms and signs, patients typically experienced progressively severe sore throat, developed a maculopapular rash, had intractable abdominal pain, and began to bleed from multiple sites, principally the gastrointestinal tract. Although laboratory determinations were limited and not conclusive, it was concluded that pathogenesis of the disease included non-icteric hepatitis and possibly acute pancreatitis as well as disseminated intravascular coagulation.This syndrome was caused by a virus morphologically similar to Marburg virus, but immunologically distinct. It was named Ebola virus. The agent was isolated from the blood of 8 of 10 suspected cases using Vero cell cultures. Titrations of serial specimens obtained from one patient disclosed persistent viraemia of 10(6.5)-10(4.5) infectious units from the third day of illness until death on the eighth day. Ebola virus particles were found in formalin-


Quality control in laboratory investigations on Schistosoma mansoni on St Lucia, West Indies: a staff assessment scheme. A method is described for the supervision of technicians engaged in microscopical screening of large numbers of stools for Schistosoma mansoni ova. The scheme presents graphically a regularly updated longitudinal evaluation of both individual and group standards of technical competence.


Trials of ecological and chemical measures for the control of Schistosoma haematobium transmission in a Volta Lake village. Urinary schistosomiasis is highly prevalent along the shores of the Volta Lake in Ghana, where transmission occurs focally in man-water contact sites. The intermediate host, Bulinus truncatus rohlfsi, prefers to harbour in Ceratophyllum, a common aquatic weed in the lake. Removal of this weed reduced the density of both infected and uninfected snails, but not sufficiently to interrupt transmission.Niclosamide was applied at 1, 0.7, and 0.5 mg/l in combination with weed removal at five water contact sites in December, January, and February, respectively. Plastic sheets were used to isolate treated sites from the main body of the lake. Snail surveys were carried out at short intervals to assess the effectiveness of these intervention measures. Niclosamide at 0.5 mg/l applied after weed removal was effective in killing the snails. The overall results indicate that snail control along the entire lake shore is impossible but that focal control of cercarial transmission at water contact sites is attainable and shows promise.


Effect of doxycycline on transmission of Vibrio cholerae infection among family contacts of cholera patients in Calcutta. Doxycycline was used among the family contacts of hospitalized cholera patients in Calcutta to determine its effectiveness in controlling transmission of V. cholerae infection. A total of 137 such contacts were given a single oral dose of doxycycline in graded doses according to age. A similar group of 139 contacts received a single dose of multivitamins as placebo. All 276 contacts were examined bacteriologically daily for 10 days for the presence of V. cholerae in their stools. The results showed that doxycycline was effective in significantly reducing the load of V. cholerae infection for up to 5 days following treatment.


Effect of potassium cardioplegia on myocardial ischemia and post arrest ventricular function. To assess the effects of moderate potassium cardioplegia (37 mEq/l KCl) on the severity of myocardial ischemia during arrest and on post arrest ventricular function, 32 isolated, isovolumic feline hearts were studied before, during and 1 hour after ischemic arrest. Normothermia (37 degrees C) was maintained in the remaining 16 hearts, eight without KCl and eight with KCl. Hypothermia (27 degrees C) was maintained in the remaining 16 hearts, eight with KCl and eight without KCl. Myocardial oxygen (PmO2) and carbon dioxide tensions (PmCO2) were measured by mass spectrometry. Maximum developed intraventricular pressure (max DP) and max dP/dt were used as indices of performance. Compared with normothermic or hypothermic arrest alone, the addition of potassium cardioplegia resulted in a significant reduction in the peak PmCO2 measured during the arrest period. Hypothermia alone resulted in morphologic evidence of improved myocardial preservation and a significant reduction in peak PmCO2 compared with normothermia. Post arrest ventricular function was best with the combination of hypothermic arrest and potassium cardioplegia (max DP = 96 +/- 6% of control and max dP/dt = 99 +/- 5% of control). These data suggest that the beneficial effects of postassium cardioplegia and 27 degrees hypothermia are additive, and that reduction in myocardial ischemia as evidenced by a reduction in peak PmCO2 correlated with improvement in ventricular performance in the post arrest period and with preservation of myocardial structure.


Vestibular function in benign intracranial hypertension. Investigation of vestibular function in 34 patients with benign intracranial hypertension was performed and a follow-up examination 3-20 years after the primary examination, when the symptoms of intracranial hypertension had disappeared. A primarily affected vestibular function was found in 19 patients (56%), viz. hyporeactivity in 9, preponderance in 9, and hyperreactivity in 2. Except for 2 patients, all had normal vestibular reactions at follow-up. These patients exhibited no evidence of what is called 'congestive inner ear'. An account is given also of the role of vestibular testing in connection with brain death criteria and recent experience of continuous monitoring of intracranial pressure.


Accumulation of thyroid antigen-reactive T lymphocytes in the gland of patients with subacute thyroiditis. Blood and thyroid-infiltrating lymphocytes from patients with de Quervain's subacute thyroiditis were tested in the leucocyte migration inhibition test for cell-mediated immunity (CMI) to thyroid antigen. Blood leucocytes were positive for CMI in eleven out of thirteen (85%) patients in the acute phase of the disease. In five cases tested repeatedly this reactivity disappeared after 7-11 months. Thyroid-infiltrating lymphocytes were obtained from eight patients by fine-needle aspiration biopsy. There was a relative accumulation of T lymphocytes (90 +/- 5%) in the thyroid gland as compared to lower numbers of T cells (65 +/- 5%) in the blood. When thyroid-infiltrating lymphocytes were tested for CMI to thyroid antigen, a significantly stronger inhibition was demonstrated with the infiltrating lymphocytes as compared to that of blood leucocytes. We conclude that thyroid antigen-reactive T cells accumulate in the glands of patients with subacute thyroiditis. This observation is in accordance with the presumed viral aetiology of this disorder.


Enzyme differentiation of lymphocyte subpopulations in sections of human lymph nodes, tonsils and periodontal disease. Five enzymes have been studied to differentiate between T and B lymphocytes in sections of human lymph nodes, tonsils and chronic inflammatory periodontal disease. The presence of acid phosphatase, beta-glucuronidase N-acetyl-beta-glucosaminidase, non-specific esterase and fluoride-resistant esterase activity was determined histochemically. The results indicate that cells in the B area of both lymph nodes and tonsils are negative for enzyme activity, while those in the T area show a single intense granule of activity. These enzymes were unable to differentiate between T blasts, B blasts and plasma cells in the sections studied. The majority of the lymphoid cells in the lesions of chronic inflammatory periodontal disease are enzyme-negative and probably of B-cell origin.


Susceptibility of inbred strains of mice to Trypanosoma congolense: correlation with changes in spleen lymphocyte populations. A comparison was made of the susceptibility of eight inbred strains of mice to infection with Trypanosoma congolense. Marked differences in susceptibility as judged by survival were found between the different strains. The capacity of certain strains to survive longer than others appeared to be related to their ability to limit the numbers of trypanosomes in the circulation. There was no difference in the infectivity of T. congolense for mice of high and low susceptibility. Furthermore, the findings of similar prepatent periods suggested that the initial replication rate was similar in the different strains. These findings suggested that the level of parasitaemia in different strains of may reflect differences in the nature of quality of the immune response to the trypanosome. In all of the strains of mice a marked increase in splenic B and null lymphocytes was found. This, allied to the finding of an increase in the background plaque-forming cells to sheep erythrocytes, indicated, as suggested by other workers, that trypanosome infection results in a non-specific polyclonal activation of lymphocytes, and that this affects primarily B lymphocytes. In strains of mice which survived longest, i.e. C57B1/6J and AKR/A, the increase in splenic B and null cells was less marked. Whether this is associated with a decreased susceptibility of these strains to polyclonal activation induced by trypanosome infection, or whether it is merely the result of lower levels of parasitaemia, remains to be determined. By comparing T. congolense infection in three strains of mice congenic at the H-2 locus, representing H-2a, H-2b and H-2k haplotypes, it was found that the susceptibility was not associated with the H-2 haplotype. The finding that (A/J X C57B1/6J)F1 hybrids were of similar susceptibility as the C57B1/6J parents indicated that the relative resistance of this strain is inherited as a dominant trait, although in the early stages of infection the F1 hybrids consistently showed somewhat higher levels of parasitaemia than the C57B1/6J mice. Athymic nude mice and surgically splenectomized mice were found to be more susceptible to T. congolense infection than intact mice of the same strain. However, the effect of splenectomy was much less pronounced in C57B1/6J mice than in the relatively more susceptible BALB/c/A mice.


Suppression of rat adjuvant disease by cyclophosphamide pretreatment: evidence for an antibody mediated component in the pathogenesis of the disease. Cyclophosphamide (Cy), given intraperitoneally at a dose of 100 mg per kg body weight 3 days before adjuvant, was found to abolish the development of adjuvant disease in the PVG/c rat. This treatment, however, enhanced the delayed hypersensitivity responses to purified protein derivative of tuberculin (PPD) developed by these animals. Lower doses of Cy caused a partial inhibition of arthritis which was dose-related. When the time between giving Cy and the injection of adjuvant was increased, a gradual time-dependent recovery of the response was observed. The arthritic response was restored by the passive transfer of 7.6 x 10(7) to 1.5 x 10(8) normal syngeneic spleen cells, although the development of secondary lesions was delayed by 7-14 days. The response could also be restored by the transfer of small amounts of serum from arthritic, but not normal, rats. Large amounts of serum failed to restore the response. Additional evidence that pretreatment with Cy preferentially depleted the B lymphocytes was obtained by the histological examination of the lymphoid tissue. It was also shown that the primary antibody response to sheep erythrocytes was abolished by Cy, but that skin allograft rejection was unaffected. A partial inhibition of the acute inflammatory reaction to carrageenan was observed 3 days after giving Cy. It is suggested that the pathogenesis of adjuvant arthritis involves an immune complex-mediated phase, whihc initiates the joint lesions. Once these lesions have formed, cell-mediated immune mechanisms predominate in the development of the disease. It is not known whether the persistence of immune complexes is necessary to maintain the lesions.


Augmentation of mitogen responsiveness in human lymphocytes by a humoral factor obtained from thymic epithelial cultures. Supernatants derived from thymic epithelial cultures were studied for their effect in augmenting mitogen responsiveness in human thymocytes and lymphocytes. Incubation of these cells for 20 hr in diluted supernatants obtained from 14 to 25 day old cultures of thymic epithelium resulted in a significant increase in the response to Con A. The epithelial nature of the cells was confirmed by electron microscopy. Supernatants from fibroblast cultures or thymic epithelial cultures overgrown by fibroblasts were not effective, nor were supernatants from secondary epithelial outgrowths. The molecular weight of the active fraction appeared to be between 17,000 and 45,000 daltons. The data indicated that human thymus epithelium produced one or more humoral factors which were identical to, or shared properties with, thymic hormones.


Receptors for IgM: a feature of subpopulations of both T and B human lymphocytes. Receptors for IgM were detected on peripheral blood and tonsil human lymphocytes by a rosette technique with ox red blood cells (ORBC) coated with anti-ORBC rabbit IgM. It was found that the receptors are very sensitive to handling procedures of cells and to low temperatures. An overnight incubation period at 37 degrees C was the optimal condition for the maximum expression of receptors for IgM, but the use of IgM-free media in these cultures was neither an essential nor favourable factor for an optimal rosette formation, when ORBC heavily coated with rabbit IgM were used. The great majority of chronic lymphocytic leukaemia (CLL) patients presented a high number of EA(IgM)-RFC, either on freshly drawn or cultured lymphocytes. By fractionation procedures of normal peripheral blood and tonsil lymphocytes, it was found that a subpopulation of B cells, like T cells, also posses a lower avidity for IgM than those present on T lymphocytes.


Migration inhibition of T lymphocytes from human peripheral blood by specific antigen and lymphokines. The inhibition of migration of human peripheral blood cells in the presence of PPD was studied. It was found that migration inhibition of peripheral blood mononuclear cells (MN) from Mantoux-positive donors was far greater than the migration inhibition of peripheral blood leucocytes (PBL). Moreover, MN cells and T lymphocytes showed larger and more uniform areas of migration. In contrast, the migration of B lymphocytes and monocytes was poor. Further analysis using purified subpopulations of MN cells showed that PPD inhibited the migration of T lymphocytes but not of B lymphocytes and monocytes. Corresponding to these findings, lymphokine-containing supernatants also inhibited the migration of purified T cells from Matoux-negative donors. It was concluded that the T lymphocyte was the predominant cell in the MN cell population, which migrated, and was subject to inhibition by PPD or lymphokines. These results imply that the movement of human T lymphocytes may be influenced by soluble factors from antigen-activated sensitized cells.


Age-dependent changes of human blood lymphocyte subpopulations. T- and B-lymphocyte populations were enumerated at four stages of life: at the newly born, infant, adult and aged stages. The proportion of T cells detected by E rosettes and an anti-human T-lymphocyte antigen (HTLA) serum incresed from new-born children to adults, then decreased with ageing. The antiserum detected less mature T cells in aged people. The percentages of cell forming 'active' E rosettes increased with ageing. Lower numbers of B cells bearing surface immunoglobulins were found in adults.Complement receptor-bearing lymphocytes (percentages and absolute numbers) decreased from new-born children to aged humans. Finally, the number of monocytes were significantly greater in the young than in adult and aged people. Such results bring new data concerning the age-dependent changes of lymphocyte subpopulations and concerning the significance of various techniques used together to detect mononuclear cell populations in the human peripheral blood.


Catabolic rate of alpha1-antitrypsin of of Pi types S, and MMalton and of asialylated M-protein in man. 1. Human alpha1-antitrypsin was isolated with preserved microheterogeneity from subjects of Pi types M, S and MMalton. The M-protein was partially (20%) and completely desialylated. The proteins were labelled with either 125I or 131I. 2. The disappearance rate of these alpha1-antitrypsins was studied after simultaneous injection of the two types of labelled protein into Pi M subjects. The fractional catabolic rates of S- and MMalton-protein were 0.36 and 0.34 day(-1) respectively compared with 0.28 day(-1) for M-protein. The ratio of extravascular to plasma pools was 1.4 for S- protein and 1.6 for MMalton-protein. The 20% desialylated M-protein showed an increase of about 100% in its fractional catabolic rate. The disappearance rate of completely desialylated alpha1-antitrypsin was extremely rapid. 3. The slightly higher fractional catabolic rate of S- than of M-protein can only partly explain the 40% lower plasma concentration in subjects of Pi type S. Similarly the slight increase in catabolic rate of Pi MMalton- protein is too small to explain why the alpha1-antitrypsin content of the blood in Pi MMalton subjects is only 15% of that normally found. A low hepatic secretion seems to be the major cause of the low alpha1-antitrypsin concentration found in subjects of types Pi S and MMalton, as in Pi type Z.


Hemorrhagic gastritis: current concepts. Hemorrhagic gastritis is a common cause of upper GI hemorrhage, in which the clinical course may vary from a mild, self-limited illness when associated with gastric irritants, to lifethreatening massive hemorrhage when associated with stress in critically ill patients. Treatment is based upon early endoscopic diagnosis, withdrawal of offending gastric irritants, intensive organ system support, and reduction of gastric acidity. Angiographic techniques and surgery are restricted to patients who fail these supportive measures, often after blood replacement beyond the usual criteria for operative intervention, usually in patients with stress. Newer pharmacologic and endoscopic therapeutic modalities are under evaluation and may help improve survival among patients with massive hemorrhage from stress ulcer.


Diagnosis of pneumonia due to Pneumocystis by subsegmental pulmonary lavage via the fiberoptic bronchoscope. To assess the accuracy of pulmonary lavage in diagnosing pneumonia due to Pneumocystis, we used animals as a model and then prospectively studied 33 immunosuppressed adults with diffuse pulmonary infiltrates. In rats treated with cortisone, Pneumocystis organisms could be found in the effluent from lavage as early as in sections of pulmonary tissue, and the effluent from lavage remained diagnostic throughout the ten weeks of observation. Subsegmental lavage in adult patients was performed through the wedged fiberoptic bronchoscope. Pneumocystis organisms were demonstrated in seven patients by lavage, and no false-negative results were recorded. Pneumocystis organisms were readily identified among the sheets of alveolar macrophages seen in smears of the effluent from lavage that were stained with methenamine silver. Subsegmental lavage via the fiberoptic bronchoscope is an accurate and safe technique for establishing the diagnosis of pneumonia due to Pneumocystis in patients whose respiratory embarrassment or thrombocytopenia makes biopsy of the lung hazardous.


Trimethoprim with sulfamethoxazole for treatment of infection with Pneumocystis carinii in renal insufficiency. Pneumonia due to Pneumocystis carinii developed in an immunosuppressed patient with a renal transplant who had substantial renal failure. Successful treatment was acheived with administration  of trimethoprim plus sulfamethoxazole at a dosage adjusted to the degree of renal failure. There were no significant adverse effects from this therapy.


[Antimycotic sensitivity of yeast from clinical specimens (author's transl)]. The sensitivity against 5 antimycotics was examined with 353 yeast strains of 13 species from clinical material. The tests were carried out by means of the disc diffusion test on Bacto yeast morphology agar under standardized conditions. 33 (6.3%) of the isolates were resistant against 5-fluorocytosine, 11 (2.1%) strains showed no sensitivity against the imidazolyl antimycotics clotrimazole or miconazole. The resistant strains belonged to the species Candida albicans and Candida tropicalis. All of the isolates were sensitive to the polyene antimycotics nystatin and pimaricin (natamycin).


[The effectiveness of stimulation of skin receptors in the treatment of prolonged pain (author's transl)]. In two groups of 30 patients pain relief due to transcutaneous nerve stimulation was compared with a placebo, under similar external conditions. The two groups were approximately similar in age, sex, and the origin of pain. Only patients with continuous pain of the limbs or trunk were included in the investigation. There was no statistically significant difference in the success rate of the two groups.


[Thymomas (author's transl)].

[Thymomas (author's transl)]. Immunological investigations concerning pathological autoantibodies and defects of humoral immunity were performed in 7 patients with thymomas, 5 of which showed invasive growth. The number of B and T lymphocytes in blood was determined at the same time using membrane markers as well as blood lymphocyte stimulation with phytohaemagglutinine. Two of the 3 patients with auto-antibodies against striated muscles or nuclei showed the clinical signs of accompanying disease (myasthenia gravis, lupus erythematodes). A humoral immunodisturbance with IgM deficiency was demonstrable in one patient and was accompanied by clinical symptoms. Lymphopenia with decreased numbers and functional disturbance of T and B lymphocytes could be shown in the majority of patients. Immunological investigations simplify proof of accompanying diseases in thymomas. These represent an important prognostic criterium in the same way as does invasive growth.


The Zellweger syndrome: subcellular pathology, neuropathology, and the demonstration of pneumocystis carinii pneumonitis in two siblings. The first Scandinavian cases of Zellweger syndrome (ZS) are described. A brother and sister, children of first cousins, had the typical clinical symptoms and pathological findings. Extensive metabolic studies in the boy were negative. Pipecolic acid in the urine was not elevated. Both children died at 14 weeks of age. Two months earlier the girl had suffered severe intestinal bleeding. Both had pneumocystic carinii pneumonia at autopsy although no evidence of immune deficiency had been found in the boy. The girl had used up her visible iron depots while the boy still had abundant but probably physiologic amounts of hemosiderin in the  RES. Most of the cerebral abnormalities are unspecfic and possibly related to anoxia or other causes of delayed maturation. The white matter abnormalities in ZS patients may only be quantitatively different from the common "fatty metamorphosis" in infants. Previously reported ultrastructural abnormalities (absence of peroxisomes and very sparse smooth endoplasmic reticulum, as well as mitochondrial abnormalities) which are possibly unique for ZS, are confirmed. It is stressed that these were seen despite phenobarbital treatment which normally stimulates the formation of smooth endoplasmic reticulum.


Factors influencing surfactant composition in the newborn infant. In order to evaluate the surfactant maturation of the neonate, tracheal aspirates were analyzed in 84 newborn infants with 12h of birth. Using 2-dimensional thin-layer chromatography, 9 different phospholipids were identified. Dynamic surface tension measurements were performed with a modified Wilhelmy balance. Five different groups of infants with typical phospholipid patterns were characterized: i.e., 1. Normal term newborn. 2. RDS in the preterm infant. 3. Acceleration of lung maturity in preterm infants without RDS. 4. Retardation in term infants with RDS. 5. Therapeutic induction of pulmonary maturity in preterm infants following maternal glucocorticoid administration. Mature lung effluent contains high concentrations of phosphatidylcholine (PC) and phsophatidylglycerol (PG). In infants with RDS, PC is low and PG absent. Accelerated lung maturity was observed after chronic prenatal stress, such as prolonged rupture of the membranes, chronic vaginal bleeding, and maternal hepatitis or drug addiction. Retardation of pulmonary maturity was seen in infants with alpha-1-AT-deficiency, maternal diabetes and maternal hypothyroidism. Administration of methylprednisolone to the mother 24 h to 72h before birth induced both the synthesis of PC and PG in the preterm infants, resulting in an almost full-term phospholipid pattern as early as 31 weeks of gestation. The significance of these factors on the pathogenesis of RDS is discussed.


Surface immunoglobulin phenotype of murine spleen cells which form B cell colonies in agar. Separation of IgD-bearing B lymphocytes from the spleens of 3-week-old or adult mice has been achieved using cellular affinity columns and an anti-delta allotype serum. The results demonstrate that the majority of cells capable of forming B lymphocyte colonies in agar are delta-positive although they lose detectable IgD upon stimulation. It is proposed that these cells are the descendants of IgD-negative cells found in the fetal liver and neonatal spleen.


Growth of canine T-lymphocyte colonies in vitro. Canine lymphocytes from peripheral blood, lymph nodes, thymus and bone marrow were stimulated with phytohemagglutinin-P (PHA) or concanavalin-A (CON-A) to form colonies in methylcellulose. Lymphocytes exposed to mitogens in liquid phase formed clumps the size of colonies. Lymphocyte clumping was eliminated by plating cells directly into methylcellulose, but high concentrations of mitogens (CON-A or PHA is greater than 10 mg/10(6) lymphocytes) were required in order to get subsequent colony formation. Thus, in contrast to published reports, exposure of lymphocytes to mitogen prior to plating was not required for cloning of canine peripheral blood lymphocytes. Colonies from thymus, lymph node, or peripheral blood consisted predominantly of T lymphocytes, whereas cultures from bone marrow also produced colonies with macrophage morphology and surface-adherent colonies with mesenchymal morphology.


Effects of alloxan on orotic acid and glycogen content in various vertebrate species. Alloxan treatment induces a decrease of orotic acid content in various organs of carp, frog, pigeon and rat, parallel to a decrease of liver and muscle glycogen content. Loss of orotic acid and glycogen cannot be prevented by orotic acid and carbamyl phosphate given i.p. Mice, rats and pigeons use up and excrete exogenous orotic acid rapidly, but carps and frogs accumulate it.


Somatostatin reduces the release of colony-stimulating activity (CSA) from PHA-activated mouse spleen lymphocytes. PHA-activated lymphocytes release colony-stimulating activity (CSA) for macrophage-granulocyte precursor cells (colony forming units, CFUc) in the culture medium. Somatostatin, known to interfer with ribosomal protein synthesis, was demonstrated to reduce the release of CSA from PHA-treated mouse spleen lymphocytes.


Effect of immunosuppressive agents on the blastogenic response of mouse spleen T and B lymphocytes. I. Culture conditions and the evaluation system. A microplate culture system has been standardized for blastic transformation of lymphocytes, using various mitogens more or less specific to T lymphocytes (concanavalin A, phytohaemagglutinin, erythrogenic toxin) or B lymphocytes (lipopolysaccharides and their lipids A, pokeweed mitogen). The mitogen-stimulated lymphocyte activation was examined by the use of the nucleolar test determining the state of nucleolar RNA synthesis by morphologic criteria. In this introductory paper to a series of papers analyzing the specificity of action of the various chemical and biological immunosuppressive agents on the blastogenic responses of T and B lymphocytes, optimal concentrations of cells and blastogenic substances and other parameters were tested, and the kinetics of transformation was investigated in detail.


Immunological studies in psoriasis.

Immunological studies in psoriasis. The mean values of serum immunoglobulins A, G, M and D levels in 42 psoriasis patients showed no significant difference from those of control subjects. Analysis of the same sera for the presence of antiglobulin antibodies also yielded negative results. Antinuclear factor could be demonstrated in only 4.8% of the cases. The possibility that serum antiglobulins get drained into the lesions making their detection in the serum difficult, is supported by our preliminary findings of focal deposition in the stratum corneum of immune complexes of IgG, IgM and complement in tissue sections of psoriatic lesions from these patients.


Summertime actinic lichenoid eruption.

Summertime actinic lichenoid eruption. A distinct summertime skin disorder seen in 25 Indian patients is described under a tentative designation of summertime actinic lichenoid eruption (SALE). The eruption predominantly affects females in their 2nd to 4th decade. The initial lesions are in the form of closely aggregated pruritic lichenoid papules starting during summer months on the exposed parts of the body. The lesions respond favourably to sun protection and corticosteroid cream but the recurrences are common during the subsequent summers when the lesions tend to become more persistent, pruritic and lichenified. Histopathologic features include early basal cell degeneration, spongiotic vesiculation with or without focal parakeratosis and an intense lymphocytic infiltrate in the dermis. In spite of strong clinical evidence of relation to sunlight, the lesions are not easily reproducible by artificial light.


[The effect of hydrocortisone and 6-methylprednisolone on the cytoplasmic membrane of rat thymocytes in an vitro system]. Hydrocortisone and 6-methylprednisolone increase the binding constant of anilinenaphthalene-sulphonate  (ANS) with the cytoplasmatic thymocytes membrane following a 2--3 hour long incubation in vitro. Over the same period these compounds stimulate the liberation of potassium ions  from the thymocytes. A supravital staining of the thymocytes with Trypan blue did not change during the whole time of the hormones action. The effect of the hydrocortisone and 6-methylprednisolone action on the ANS binding constant and potassium release depende upon the duration of incubation and concentration of the steriod in the medium. Hydrocortisone produces a greater influence than does 6-methylprednisolone.


Gastrointestinal hemorrhage from ileal leiomyoma: utility of the complete reflux small bowel examination. A case of recurrent gastrointestinal hemorrhage with nondiagnostic conventional barium X-ray and endoscopic studies is described. A complete reflux small bowel examination clearly demonstrated an ilial leiomyoma. Details of the methodology of the reflux small bowel are reviewed.


Comparison of barium swallow and left gastric angiography in the diagnosis of esophageal varices, with special reference to the management of variceal bleeding. Seventy-five patients with portal hypertension and esophageal varices were studied by means of barium swallow and selective left gastric angiography. The relative merits and limitations of these techniques in demonstrating the presence, extent, and severity of the varices are discussed, with reference to the selection of patients for prophylactic transection of the esophagus to prevent bleeding from the varices.


Radiologic demonstration of Meckel's diverticulum. A case of bleeding Meckel's diverticulum with a positive barium meal examination, selective mesenteric arteriography and 99mTc-pertechnetate scanning is presented. Complementary radiologic examinations are needed in the visualization of Meckel's diverticulum due to difficulties in making the correct diagnosis.


Esophageal hematoma associated with thrombocytopenia. Esophageal hematoma secondary to thrombocytopenia has only recently been described in the literature in a single case report. This article presents the clinical manifestations and radiographic findings of 4 additional cases of esophageal hematoma secondary to thrombocytopenia. Three patients were receiving treatment for leukemia, and the other patient had aplastic anemia. Previously reported cases of esophageal hematomas from other causes are reviewed.


Postsplenectomy abscess leading to acute gastric hemorrhage. A case is presented of a 14-year-old boy with massive gastric bleeding 8 months after a splenectomy. A linear gastric ulcer was found associated with perforation leading to an abscess in the former splenic site. The vulnerability of proximal greater curvature to perforation after splenectomy, and the possibility of extrinsic erosion causing gastric or duodenal ulceration are emphasized.


Genetic basis of gluten-sentitive enteropathy. Families of patients with gluten-sensitive enteropathy (GSE) were typed for HLA-A and -B antigens as well as for HLA-DW3 antigen(s) using appropriate typing alloantisera. In addition, patients and families were typed for GSE-associated B cell antigens using alloantisera obtained from mothers and wives of patients. The data obtained suggest that disease occurs within families when two conditions were fulfilled: (1) the family member is homozygous for the GSE-associated B cell antigens and (2) the family member also bears the HLA-DW3 antigen(s) or an antigen usually associated with DW3. In addition, with the family studies it was possible to show that the gene(s) controlling the DW3 antigen(s) and those controlling the GSE-associated B cell antigens are separate nonlinked genetic loci, a fact which leads to the conclusion that GSE has a genetic basis in at least two genes. It is speculated that the genes responsible for GSE code for surface proteins which are physically associated on lymphoid cell membranes and which form receptors important to the initiation of disease.


Immune haemolytic anaemia after exchange transfusion. Two cases of immune haemolytic anaemia in newborn infants who had received exchange transfusion on the first day of life because of hyperbilirubinaemia probably due to a conjugation defect, are reported. The first baby, born in the 30th week of gestation presented erythrocyte-fixed IgM antibodies + C, neutropenia, and circulating leucocytotoxic antibodies. The second baby, born at term to a diabetic mother showed erythrocyte-fixed IgG + IgM antibodies + C and a selective IgA deficit. In both cases the anaemia improved and the Coombs test became negative spontaneously in the 4th and 5th months of life, respectively. These haematologic alterations are similar to a graft versus host (GVH) reaction due to the persistence in the neonate blood of the donor's immunocompetent cells.


Clinical evaluation of 5-hydroxy-L-tryptophan as an antidepressant drug. Effectiveness of 5-hydroxy-L-trytophan as an antidepressant drug was studied with 59 patients with depressive symptoms using Rating Scale for Depression made by Clinico-Psychopharmacology Research Group in Japan for a preparatory step of a double blind clinical study of 5-hydroxy-L-tryptophan treatment of depression. A daily dose of 150--300 mg of 5-hydroxyl-L-tryptophan was administered for three weeks. Favorable responses were observed in 40 patients (67.8%), of whom 13 patients were markedly improved. These effects were noticed in 32 patients (80% of the improved patients) within a week of the treatment. Analysis of General Improvement Rating in the various subtypes of depressive symptoms indicated that endogenous depression and involutional or senile depression were the preferable indication of 5-hydroxy-L-tryptophan loading. The main side effects of 5-hydroxy-L-tryptophan were gastrointestinal disturbances which were minimized by the simultaneous administration of metoclopromide or trihexyphenidyl.


[Methods and possibilities of surgical endoscopy]. In a number of diseases, operative or therapeutic endoscopy permits an effective therapy without the necessity to open up the abdominal wall and the digestive organs themselves. Included here are the removal of foreign bodies, haemostasis, papillotomy with concrement dislodgement, polypectomy and the obliteration of the excretory pancreas. The evaluation of the achieved results and the examination of operative endoscopy in respect of its clinical value cooperation with the pathologisz should not be neglected. The refined endoscopic methods and the highly developed technology can only be applied successfully, when it is possible to get enough skillful endoscopists and to prevent an everyman's endoscopy by which the new methods could be brought into discredit.


Studies on the influence of Trasylol on the partition of trypsin between the human plasma protease inhibitors in vitro. The distribution of trypsin between the protease inhibitors of human serum with and without Trasylol was studied in vitro. 1) Trypsin was preferentially bound by alpha2-macroglobulin on addition of small amounts of the enzyme to normal serum in both the presence and absence of Trasylol in a molar concentration equal to that of alpha2-macroglobulin. 2) On saturation of alpha2-macroglobulin, a considerable amount of trypsin was bound by Trasylol even when most of the serum alpha1-antitrypsin was in a free form. 3) In reaction mixtures containing small amounts of trypsin, Trasylol was identified in a free form as well as in complex with trypsin-alpha2-macroglobulin complex and to a limited extent with trypsin. 4) With larger amounts of trypsin, sufficient to saturate alpha2-macroglobulin, increasing amounts of Trasylol were bound to trypsin. The relative amount of Trasylol bound to trypsin-alpha2-macroglobulin complexes was now smaller. This was explained by a higher affinity (or binding rate) of Trasylol for trypsin than for trypsin-alpha2-macroglobulin complexes. 5) Trypsin-Trasylol complexes showed no signs of dissociation after 5 h incubation at 37 degrees C in serum.


Kartagener's syndrome in sibs: clinical and immunologic investigations. In a sibship of ten children descending from a first cousin's marriage, two sibs were affected by Kartagener's syndrome with the typical symptoms of situs inversus, bronchiectasis, and polyposis nasi. Clinical investigation of the entire family revealed chronic infections of the paranasal sinus in five sibs and the mother, two of whom had bronchiectasis as well. Immunologically, a persistent cellular or humoral defect could not be detected in any of the family members. In the HLA system, only the two sibs with Kartagener's syndrome had identical HLA-types; all other family members had different combinations. A linkage between the loci for the HLA system and Kartagener's syndrome is discussed.


Antibody forming cell precursors among glass-adherent peritoneal exudate cells. Among glass-adherent peritoneal exudate cells (gaPEC), induced by an inoculation of 1% glycogen solution, about 4.5% were classified as antibody-forming cell precursors (AFCP) on the first day of culture by means of anti-mouse B-cell antibody (anti-B-Ab). They proliferated and differentiated into IgG-forming plasma cells when cultured with antigen and thymic RNA in vitro. Pretreatment of PEC with anti-B-Ab and complement suppressed the formation of plasma cells. AFCP had receptors for  IgM-antigen complexes and for complement, both of which were  independent of Ca++ and MG++ and resistant to treatment by pronase or phospholipase C. Cells bearing detectable receptors for EA (IgM) an EAC diminished by the 6th day when gaPEC were cultured with thymic RNA, but persisted longer in cultures without  thymic RNA. The same percentages of cells demonstrated tartrate-resistant acid phosphatase activities but were devoid of esterase. Twenty to thirty percent of anti-B-Ab sensitive cells ingested latex particles. The proliferation kinetics of IgG-forming cells were studied through the 21st day of culture by means of peroxidase-labeled antibody staining methods.


Antibody production and protection against influenza virus in immunodeficient mice. The roles of T and B cells in the immune response to influenza virus were studied by using mice deficient in either T cells (athymic nude) or immunoglobulin production (CBA/N). The serological responses of these mice to either whole or disrupted A/Aichi/2/68 influenza virus vaccines were examined, and the protective effect of these inoculations was tested by challenge infection with mouse-adapted A/Aichi/2/68 influenza virus. In contrast to normal mice, neither strain of immunodeficient mouse produced detectable serum antibody after inoculation with either type of vaccine. CBA/N mice immunized with intact virus vaccine were protected, however, against subsequent lethal challenge. CBA/N mice inoculated with disrupted virus vaccine and nude mice inoculated with either disrupted or whole virus vaccine were not protected against viral challenge. Evidence of immunological memory was observed in CBA/N and nude mice that had survived live virus challenge after immunization with inactivated vaccine.


Propagation of Pneumocystis carinii in Vero cell culture. Pneumocystis carinii derived from infected murine lung was found to be capable of limited growth in Vero African green monkey kidney cell cultures. The observed increase in the number of cyst forms was influenced by the ratio of cysts to host cells in the inocula, duration of passage, and formulation of the cell culture media. Maximum growth was achieved by inoculating 1.3 X 10(5) cysts per 75-cm2 flask containing about 2 X 10(7) Vero cells (cyst-to-cell ratio=1:154) maintained on minimal essential medium supplemented with 2% fetal bovine serum. Under these conditions of culture, a 10.8-fold increase in cyst forms was observed during a 7-day passage in cell culture utilizing cyst-to-cell ratios ranging from 1:28 to 1:2,778.


Modulation of immune responses in newborn and adult mice by interferon. Interferon was found to have both suppressive and enhancing effects on the antibody response in newborn and adult mice. Evidence was obtained that these effects are primarily evoked during the initial steps controlling cell proliferation. Stimulation of thymus and spleen cells with a T-cell mitogen was enhanced by low doses and suppressed by high doses of interferon. Treatment of parental spleen cells with interferon before injecting them into immunized F1 hybrid mice resulted in an enhanced allogeneic effect. These results are compatible with the hypothesis that interferon affects T cells and has an immunoregulatory role, either by inhibiting the action of suppressor cells or by promoting immunological maturation.


Role for macrophages and thymus-derived lymphocytes in cholera toxin-induced immunosuppression. The exo-enterotoxin derived from Vibrio cholerae bacilli has marked immunomodulating activities, both in vivo and in vitro. In the present study, the mechanism whereby cholera toxin depresses the antibody-forming ability of murine splenocytes was investigated by in vitro reconstitution experiments. Spleen cells derived from mice treated with cholera toxin 2 days earlier were markedly deficient in their ability to respond to sheep erythrocytes upon challenge immunization in vitro. Addition of graded numbers of normal spleen cells to spleen cell cultures from toxin-treated mice partially restored the antibody response. Adherent splenocyte populations were even more effective in restoring antibody formation. Normal peritoneal exudate cells rich in macrophages were also capable of restoring the antibody-forming ability of toxin-pretreated splenocytes. Furthermore, thymus (T)-derived spleen cells from normal mice, as well as sheep erythrocyte "educated" T cells, were capable of restoring antibody formation to normal levels. The importance of T lymphocytes in restoring immune competence of spleen cell cultures from toxin-treated mice was shown by additional experiments in which T-depleted cell preparations were found to be ineffective in restoring antibody activity. These studies point to macrophages and T-derived lymphocytes as a major target for cholera toxin-induced immunosuppression.


Stimulation of subpopulations of human lymphocytes by a vaccine strain of Francisella tularensis. When purified T lymphocytes from individuals vaccinated with a viable, attenuated strain of Francisella tularensis were incubated in vitro in the presence of heat-killed bacteria or a membrane preparation of the vaccine strain, they were stimulated to form blast cells and to synthesize deoxyribonucleic acid. The blast cells had the characteristics of T cells, being devoid of surface immunoglobulin and able to form rosettes with sheep erythrocytes. The stimulation occurred only when monocytes were present. A lymphocyte preparation enriched in B lymphocytes did not respond to the heat-killed bacteria or to the membrane preparation. In a stimulated mononuclear leukocyte preparation, about 70% of the blast cells formed rosettes with sheep erythrocytes, and 10 to 20% of them had surface immunoglobulin. The results show that there is an enlarged population of specifically committed T lymphocytes after tularemia vaccination. It is suggested that the lymphocyte stimulation test measures mainly T-lymphocyte reactivity when membranes or whole bacteria of F. tularensis LVS are used as antigen, and that the stimulation of human T lymphocytes by whole bacteria or bacterial membranes is completely monocyte or macrophage dependent. The present experimental procedure may provide a model for study of antigen-induced stimulation of human lymphocytes under controlled conditions. The technique used gave a reproducible, extremely purified preparation of T lymphocytes and a preparation of monocytes especially suitable for microcultures.


Experimental Haemophilus influenzae type b meningitis: immunological investigation of the infant rat model. Infant rats inoculated intraperitoneally with Haemophilus influenzae type b develop bacteremia and meningitis. Rats were infected at 10 to 12 days of age and studied for the development of serum anticapsular antibody and bactericidal and opsonizing activity. Seven and 11 weeks after inoculation, convalescent animals showed a higher frequency of anticapsular antibody responses than uninfected controls, but 35 to 40% of the infected group had undetectable levels of anticapsular antibody (<0.10 mug/ml). In contrast, all of the convalescent animals, but none of the controls, showed moderate titers of serum bactericidal activity; and bactericidal activity persisted after absorption of the convalescent sera with type b capsule. Bactericidal activity was detected primarily in the eluted fraction corresponding to a molecular weight of 150,000 and was present in the offspring of convalescent females. Offspring of convalescent females were protected against challenge with H. influenzae type b, and control offspring could also be protected by passive immunization with convalescent serum which lacked detectable anticapsular antibody. Convalescent serum samples efficiently opsonized H. influenzae type b, and this activity persisted after absorption of the serum with capsular antigen. These data are consistent with the hypothesis that antibody to the noncapsular surface antigens of H. influenzae type b play an important role in host defenses.


Trypanosoma rhodesiense infection in congenitally athymic (nude) mice. Athymic nude mice (nu/nu), heterozygous litter mates (nu/+), and thymic reconstituted homozygous animals (nu/nu TxR) were infected with Trypanosoma rhodesiense. A reduced parasitemia and prolonged survival were observed in the nu/nu animals after infection and cure or after immunization with irradiated organisms. These studies indicate that resistance of mice to T. rhodesiense infection is relatively independent of thymic lymphocyte function.


Two small lymphocyte subpopulations in human peripheral blood. I. Purification and surface marker profiles. By means of simple rosette sedimentation methods two subsets from human peripheral blood lymphocytes have been isolated: (1) (E, Fc)- and (2) (E, Ig)-. The first subset was obtained by centrifuging suspensions of macrophage-depleted PBL in which E and EA rosettes had been allowed to form simultaneously. The dominant marker of these E- Fc- cells was surface Ig, and during 4 days of culture this population did not alter its surface markers. Subset 2 was obtained in two ways following rosette centrifugation with AET-treated SRBC and rabbit anti-human Ig-coated autologous RBC. This 'Null cell population' was shown to be highly variable as judged by the surface markers applied after 4 days of culture, and it is suggested that Null cells contain a number of immature lymphoid cells that may acquire their surface marker during culture. It is concluded that the methods described for purification of small subsets of human lymphocytes are effective and easy to perform and might be used to purify cells for functional studies.


Lymphocyte-potentiating factor from human peripheral blood adherent cells. Low-molecular-weight factor, which can restore the diminished concanavalin A (Con A) response of adherent-cell-depleted human peripheral blood lymphocytes, was partially purified from the culture supernatant of human peripheral blood adherent cells by ion-exchange column chromatography, gel filtration and preparative paper chromatography. This factor apparently has a mechanism of action distinct from that of 2-mercaptoethanol. It affects the cells just before the onset of DNA synthesis possibly as one of the second signals in the Con-A-induced activation of human peripheral blood lymphocytes. However, it was demonstrated that the effective activation of lymphoyctes also required the presentation of Con A by adherent cells to lymphocytes at the triggering stage of the activation, indicating that adherent cells have two distinct functions in the Con-A-induced activation of lymphocytes, i.e. the the presentation of Con A to lymphoyctes and the elaboration of soluble factors.


Initial observations on the isolated retinal pigment epithelium-choroid of the cat. An in vitro preparation of a mammalian retinal pigment epithelium (RPE)-choroid was developed, that of the cat, in order to study the RPE in isolation from the neural retina. The purpose of these initial experiments was to evaluate the electrical characteristics of the tissue and the ionic properties of the RPE apical membrane. They were designed so that results would be directly comparable to those previously obtained on a more extensively studied cold-blooded preparation, the bullfrog. The electrical characteristics of the best cat tissues were similar to those routinely obtained in bullfrog, whereas the average cat apical membrane potential was 20 mV more depolarized than that of frog. The apical membrane of cat resembled the frog's in having a large sensitivity to potassium, but it had a relatively smaller sensitivity to bicarbonate and a relatively larger sensitivity to sodium. The cat, like the frog, also had a ouabain-sensitive mechanism on its apical membrane that directly contributed to membrane potential. Two factors contributing to the lower apical membrane potential in cat were the higher potassium concentration of the mammalian salt solution and the sodium conductance of the apical membrane.


Selective embolization of the inferior mesenteric artery in massive colonic bleeding. A case is described in which massive rectal bleeding was controlled by selective embolization of the superior hemorrhoidal arteries. Selective therapeutic arterial embolization in colonic bleeding represents an important alternative to procedures such as surgery and infusion of vasospastic agents which carry a prohibitive risk in severely ill patients.


Effects of L-5-hydroxytryptophan in autistic children. Behavioral effects of L-5-Hydroxytrophan (L-5-HTP), administered in combination with carbidopa, were evaluated in three autistic children using direct behavioral observation and parent ratings. Children were assessed under each of four experimental conditions: Baseline, Placebo I, L-5-HTP plus carbidopa, and Placebo II. During the 20-week study two children showed behavioral change that appeared to be unrelated to drug treatment. The findings did not support the hypothesis that a functional deficit in brain 5-HT underlies the autistic syndrome.


Viral R plasmid Rphi6P: properties of the penicillinase plasmid prophage and the supercoiled, circular encapsidated genome. Properties of the viral R plasmid Rphi6P are described. As a temperate bacteriophage, it plaques on the facultative phototroph Rhodopseudomonas sphaeroides. Under aerobic conditions the phage had a latent period of 180 min, a burst time of 200 min, and a burst size of 15 to 20 particles per infective center. The encapsidated viral genome occurred as a supercoiled, circular DNA duplex with a mean contour length of 16.5 +/- 10 micron. Percent guanine plus cytosine, as calculated from thermal denaturation profiles, was 63.5. Mitomycin C-induced loss of the prophage suggested an extrachromosomal location in the host cell. Use of this curing agent enabled the isolation of a plasmid-free strain of R. sphaeroides. Biophysical analysis of the plasmid-free strain lysogenized with Rphi6P confirmed that the prophage occurred as a plasmid in the host cell.


[Ultrastructural study of odontogenesis in the frog (Rana temporaria L.)]. Differentiation of the cells of the dental bud which produce the hard tissues of teeth was studied at the ultrastructural level; the activity of these cells (odontoblasts and ameloblasts), the laying down of organic matrices elaborated by them, the mineralization of these matrices, and the maturation of dentine and enamel are described. Injections of tritiated proline enabled us by the autoradiographic method, to confirm and to indicate more precisely the activity of these cells. Dental calcified tissues of Rana exhibit the general properties of these tissues among Vertebrates: high content of apatite, low content of organic materials (collagenous in dentine, non-collagenous in enamel), precise orientation of crystals, and true but non-prismatic enamel. The enamel of Anura as well as enamel of Urodela is a true enamel of epithelial origin.


Topological studies of the steroid hydroxylase complexes in bovine adrenocortical mitochondria. The topology of the steroid hydroxylase complexes in bovine adrenocortical mitochondria was studied by using nonpenetrating artificial electron acceptors and the impermeable protein reagent diazobenzenesulfonate. Inhibition of steroid hydroxylase activity by ferricyanide and dichlorophenolindophenol sulfonate was only observed in mitochondria which had been damaged by various techniques. Intact mitochondria were not inhibited by these reagents. The reaction was monitored by oxygen uptake due to hydroxylation of deoxycorticosterone, as well as P-450 reduction and corticosterone formation. The results obtained were similar regardless of how the activity was measured. Labeling of the mitochondria with the nonpenetrating protein reagent diazobenzenesulfonate also inhibited P-450 reduction and corticosterone formation in mitochondria which had been damaged prior to addition of this reagent. Intact mitochondria which were labeled with this reagent showed very little inhibition of both activities. These results strongly suggest that all protein components of the steroid 11beta-hydroxylase system are located on the matrix side of the mitochondrial inner membrane. The inability of ferricyanide, dichlorophenolindophenol sulfonate, and diazobenzenesulfonate to inhibit the malate-dependent reduction of P-450 in intact mitochondria implies that all the P-450-dependent mitochondrial steroid hydroxylase systems are located on the matrix side of the inner mitochondrial membrane.


The active form of cytochrome P-45011beta from adrenal cortex mitochondria. Cytochrome P-45011beta has been solubilized and partially purified from bovine adrenal cortex mitochondria by means of chromatography on Octyl-Sepharose CL-4B or DEAE-Sepharose CL-6B. The partially purified P-450 preparations were about 90% pure as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but had a low specific content of P-450 (between 1 and 2 nmol of P-450 per mg of protein). In the presence of purified preparations of adrenodoxin reductase and adrenodoxin, the partially purified P-450 preparations catalyzed NADPH-supported 11beta-hydroxylation of unconjugated and sulfoconjugated deoxycorticosterone. In the reconstituted system the hydroxylation of deoxycorticosterone sulfate proceeded at a much higher rate than in intact mitochondria, indicating that in the former case interactions between the hydrophilic substrate and P-450 were facilitated. In the presence of Triton X-100 the partially purified cytochrome P-45011beta had a Stokes radius of 4.5 nm, a sedimentation coefficient of 3.1 S, and a partial specific volume of about 0.85 cm3/g. These results indicate that the cytochrome P-45011beta . Triton X-100 complex had a molecular weight of about 100,000 and that P-45011beta bound about 1.1 g of Triton X-100 per g of protein. The P-45011beta . Triton X-100 complex was catalytically active in hydroxylation reactions supported by NADPH or the hydroxylating agent ortho-nitroiodosobenzene, suggesting that the monomer of cytochrome P-45011beta is the active form of the protein.


Proteolytic inactivation of the luciferase from the luminous marine bacterium Beneckea harveyi. The enzymatic activity of bacterial luciferase from Beneckea harveyi (a heterodimer, Mr = approximately 79,000) is rapidly lost upon treatment with trypsin or chymotrypsin. Under nondenaturing conditions, the proteolytically inactivated molecule has the same apparent molecular weight as the native enzyme, and appears to be relatively stable to further proteolytic degradation. Gel electrophoresis in sodium dodecyl sulfate of the products of this digestion shows that only the alpha subunit is degraded during the time of these experiments, and its rate of loss is the same as the rate of loss of light-producing activity. The action of either protease produces a species with mobility indicative of a molecular weight of about 28,000 and smaller fragments, and an unaltered beta subunit.


Primary structure of human alpha1-protease inhibitor. The complete amino acid sequence of cyanogen bromide fragment II. The complete amino acid sequence of a CNBr-fragment from human alpha1-protease inhibitor has been determined and is shown below. The peptide consists of 109 amino acid residues with 1 oligosaccharide unit. The 2 glutamic acid residues which have previously been shown to be substituted by lysine in the Z and by valine in the S mutant proteins are both located in this CNBr fragment. (formula: see text).


The cyclic behavior of a cytoplasmic factor controlling nuclear membrane breakdown. The activity of a cytoplasmic factor (MPF), capable of inducing nuclear membrane breakdown (germinal vesicle breakdown) when injected into amphibian oocytes, has been studied during the course of early cleavage in amphibian embryos. Mature egg cytoplasm was found to contain high levels of this activity, but this was quickly lost after fertilization or artificial activation. MPF activity later reappeared in the egg cytoplasm and started to cycle with time. The peak of embryonic MPF activity during each cycle coincided with the time the embryonic nuclei were entering the G2-M transition, i.e., mitosis. However, in colchicine-arrested embryos, this activity remained at an elevated level and no longer oscillated. The timing of the appearance and disappearance of this activity appeared to be under the control of the cytoplasm because such behavior was still observed in enucleated eggs. Continued protein synthesis in the embryo was required for the reappearance, but not for the disappearance, of this activity. MPF, previously thought to be restricted to oocyte maturation, may play a more general role in controlling nuclear membrane breakdown during mitosis as well as meiosis.


Reinnervation of muscle fiber basal lamina after removal of myofibers. Differentiation of regenerating axons at original synaptic sites. Axons regenerate to reinnervate denervated skeletal muscle fibers precisely at original synaptic sites, and they differentiate into nerve terminals where they contact muscle fibers. The aim of this study was to determine the location of factors that influence the growth and differentiation of the regenerating axons. We damaged and denervated frog muscles, causing myofibers and nerve terminals to degenerate, and then irradiated the animals to prevent regeneration of myofibers. The sheath of basal lamina (BL) that surrounds each myofiber survives these treatments, and original synaptic sites on BL can be recognized by several histological criteria after nerve terminals and muscle cells have been completely removed. Axons regenerate into the region of damage within 2 wk. They contact surviving BL almost exclusively at original synaptic sites; thus, factors that guide the axon's growth are present at synaptic sites and stably maintained outside of the myofiber. Portions of axons that contact the BL acquire active zones and accumulations of synaptic vesicles; thus by morphological criteria they differentiate into nerve terminals even though their postsynaptic targets, the myofibers, are absent. Within the terminals, the synaptic organelles line up opposite periodic specializations in the myofiber's BL, demonstrating that components associated with the BL play a role in organizing the differentiation of the nerve terminal.


Observations of the marginal band system of nucleated erythrocytes. The marginal band (MB) of nucleated erythrocytes (thos of nonmammalian vertebrates) is a continuous peripheral bundle of microtubules normally obscured by hemoglobin. Treatment of these elliptical cells with modified microtubule polymerization media containing Triton X-100 yields a semilysed system in which MB, nucleus, and trans-MB material (TBM) are visible under phase contrast. The TBM apparently interconnects structural components, passing around opposite sides of the nucleus and suspending it in native position. In uranyl acetatestained whole whole mounts (goldfish) examined by transmission electron microscopy, the TBM appears as a network. MBs of semilysed cells are relatively planar initially, but twist subsequently into a range of "figure-8" shapes with one of the two possible mirror-image configurations predominant. Nuclei and MBs can be released using proteolytic enzymes, to which the TBM seems most rapidly vulnerable. MBs thus freed are birefringent, generally untwisted, and much more circular than they are in situ. As a working hypothesis, it is prosposed that the flattened, elliptical shape of nucleated erythrocytes is a result of TBM tension applied asymmetrically across an otherwise more circular MB, and that the firure-8 configuration occurs when there is extreme TBM shrinkage or contraction.


Amphibian cells in culture. I. Nutritional studies. Nutritional requirements of amphibian cells in culture were studied for the purpose of modifying a minimal medium in which frog cells could proliferate and which could be used for obtaining drug-resistant and auxotrophic variants. The serum, purine, CO2, and amino acid requirements for ICR 2A (a Rana pipiens haploid cell strain) have been investigated employing two different media: L-15, a nonbicarbonate, amino acid-buffered medium and Eagle's MEM, a bicarbonate-buffered medium. In this paper we present evidence to support the following conclusions: (1) With L-15 as the base medium, 10% fetal calf serum (FCS) supports optimal cell growth during exponential phase. Calf serum, whole, dialyzed, or heat-inactivated, cannot substitute for FCS and, in fact, is inhibitory. (2) Purines are required by ICR 2A cells only if grown in a nonbicarbonate-buffered medium, since the cells under these conditions cannot produce enough endogenous CO2 to support de novo purine synthesis. (3) In addition to the amino acids considered essential for mammalian cells in culture, ICR 2A cells depend upon exogenous asparagine. Glutamine and/or aspartic acid cannot replace the asparagine requirement. However, ICR 2A cells do utilized exogenous glutamine as an oxidative substrate.


Amphibian cells in culture. II. Isolation of drug-resistant variants and an asparagine-independent variant. With L-15 as the base medium, drug-resistant variants were isolated from two amphibian tissue culture strains: the Xenopus laevis A8 diploid cell line and the ICR 2A cell line of Rana pipiens. Four different classes of variants were obtained: (1) A8 cells resistant to chloramphenicol, an inhibitor of mitochondrial protein synthesis; (2) A8 cells resistant to ouabain, an inhibitor of the Na+/K+-activated ATPase of the plasma membrane;(3) ICR 2A cells resistant to low (20 microgram/ml) and high (300 microgram/ml) levels of bromodeoxyuridine (BUdR), a thymidine analog which interferes with the pyrimidine salvage pathway; and (4) ICR 2A cells resistant to 2,6-diaminopurine (DAP), an adenine analog which interferes with the purine salvage pathway. Unlike the other variants, isolation of BUdR resistant cells is a 2-step process. Resistance to low levels of BUdR is phenotypically expressed by a reduction in thymidine transport activities while resistance to high levels of this compound is evidenced by greatly reduced levels of thymidine kinase activity. DAP-resistant cells, which are characterized by reduced levels of adenine phosphoribosyl transferase (APRT) activity, do not die in AAT (adenine, aminopterin, thymidine) selection medium. This suggests that these cells utilize adenine efficiently. With MEM as the base medium, an asparagine independent clone was isolated from the ICR 2A cell line. When compared with the wild type, this variant exhibited a slightly reduced growth rate in the presence or absence of asparagine.