The author has conducted tissue culture of rabbit liver by roller tube method. The pH changes in culture media were estimated. The rise of hydrogen ion concentration in the course of culture was parallel to the rate of cell increase. The most appropriate medium has been a fluid medium consisting of 70 % Hank's solution, 20 % rabbit serum and 10 % chick embryo extract. Without chick embryo extract the cells found to grow do not easily grow out, but in its high concentration fibroblasts grow very well. Amoung the various sera rabbit serum is the most accelarative to the growth of liver cells. Human serum is second and horse serum worse in promoting activity. The ascites with stomach cancer almost dose not show promoting activity to liver cells. In addition of small amount of rabbit serum to the medium liver cells exhibit an excellent growth, but the more the serum becomes concentrated, the more fibroblasts grow.
By means of roller tube culture of rabbit liver the author studied the tissue growth pattern and various cells appearing in the growth area during the culture. 1) One to 2 days after the start of culture wandering cells and fibroblasts grow and wander out from the explants, and then from the 3 rd day or thereabout liver cells grow. The tissue growth continues up to the 6 th day, but therafter cells begin to degenerate unless the medium is replaced with a fresh one. 2) By the degree of the growth of liver cells and fibroblasts that appear in the growth area, the growth pattern can be classified in A and B type. A consists mainly of liver cells, dividing into 2 subtypes furthermore, and B type consists mainly of fibroblasts, dividing into 5 subtypes. 3) By tissue culture the number of binucleated liver cells decrease to 1/41/5 of number of normal rabbit on 4 th day, and increase gradually followed the duration of culture. On 10th day the number of binucleated cells recover to 1/3 of normal rabbit. The decrease of the number of binucleated liver cells indicates the accerelation of growth or regeneration of liver cells.
Three cases of WPW syndrome were reported. One was A-type and other two were B-type. In one of B-type supraventricular tachyçardia was seen ln E. K. G.. In another one of B-type, WPW was converted into normal pattcrn by administration of procaine amide.
Author did the experiments of tissue cultures infected with the agents (mitani strain) of Umayado Diseases, and got the results as follows: 1) Author did the tests of sensitivity to R. tsutsugamushi infection in the established culture-cells and knew they have the big spectrum of infection to the cells like FL cells, HELA cells and liver cells (Chang). They showed the specific cytopathic effects (CPE) with multiplication of R. tautsugamushi also. 2) It was possible to get the positive results in the infection tests to mouse with the agents which passaged in FL cells, and it is possible to do the quantitative tests of R. tsutsugamushi by inspecting of CPE as an infection indicators. These specific CPE supported to know the process of infection and growth of R. tsutsugamushi. 3) Growth process of R. tsutsugamushi is very similar with the bacterial cell division system (binary fission) and is very slow in time and specific in morphology.
Author mentioned the results of several problems on adsorption, growth and morphology of vegetative forms of R. tsutsugamushi in FL cells as the host cells of them in vitro. The process of growth of R. tsutsugamnshi was different from the process of growth of viruses. Binary fission system of R. tsutsugamushi were suggested in growth process of them from the results of growth curve and morphology of them. Also author knew that there are the release of rickettsia from the infected cells and the infection of rickettsia from cell to cell on some periods of infection process.
The general characteritics of R. tsutsugamushi which passaged on cells of tissue culture were always constant and showed the same charaçters with original physical characteristics of rickettsia. Gradually, growth of intracellular rickettsia was observed clearly and CPE was kept constantlly after passages in long period. Such CPE by R. tsutsugamushi was a infection indicator and showed a specific infection system.
Bleeding tendency following extracorporeal circulation was studied mainly in the view of vascular disturbances. 1) Pre-and post-operative changes in hemostatic mechanism were investigated in 61 patients, who underwent open heart surgery under extracorporeal circulation with Kay-Anderson heart lung machine. After perfusion the most noticeable and constant changes in hemostatic mechanism were prolongation of bleeding time, decreased capillary resistance, increased capillary permeability and reduction of platelet count which showed vascular disturbance. No correlation was seen between these changes and perfusion time. Among our clinical series there were 2 cases with spontaneous petechiae which, with certainty, were due to bleeding tendency caused by vascular disturbances. 2) Platelet count was markedly reduced during perfusion and return rapidly to normal after neutralization of heparin. This decrease of platelet count was proved to be partly caused not only by mechanical destruction of heart lung machine, but also by heparin itself. Beside vascular disturbances due to peripheral circulatory insufficiency caused by perfusion (atony of peripheral vessels, stagnation of the blood and erythrocyte sludging etc.), heparin was revealed to be a factor of postperfusional vascular damage. For prophylaxis of vascular disturbances due to peripheral circulatory insufficiency, low molecular weight dextran was found effective Prolongation of bleeding time was considered to be developed because of decreassed capillary resistance. 3) Heparin was proved to have side-effects of decreasing platelet count and capillary resistance, and of increasing capillary permeability. The decrease of platelet count was experimentally found to be reversible and the mechanism of this deficiency was attributed to platelet clump formation by heparin. Decrease of capillary resistance was elicited to be due to dysfunction of the vascular endothelium caused by heparin. Because it was found that heparin had effects to develop petechial hemorrhage, leucocytic sticking to the endothelium and increased extravasation of leucocytes. Prolonged bleeding time and decreased capillary resistance were observed in rabbits in which only platelet count deliberately decreased. Therefore, there must be correlation among them, but it is conceivable that capillary resistance decreased by heparin might be not only due to decrease of platelet count, but also to direct influence on capillary vessels by heparin.
1) Of 18 strains of Escherichia coli, which acquiréd Streptomycin resistance in vitro, 4 strains acquired Sulfanilamide resistance simultaneously in contamination with Streptomycin. 2) But no Chlorampheuicol nor tetracycline resistant bacteria is observed. 3) 2 strains of the 18 Streptomycin resistaut strains, acquired severe Penicillin sensitivity. On the reasons, authors are researching now. It may be phenomenon of “Collateral Sensitivty (SZYBALSKI)”.
Toxicity of drin compounds currently on market was investigated and the following results were obtained. 1. Endrin. The dose of LD50 was 5.2 mg/kg for mice and 13, 7 mg/kg for rabbits when given subcutaneously, 6, 6 mg/kg for mice and 5.2 mg/kg for rabbits when orálly, and 54.8 mg/kg rats when given percutaneously. 2. Aldrin. The dose of LD50 was 49.2 mg/kg for mice and 56.6 mg/kg for rabbits when given subcutaneously, 64.8 mg/kg for mice and 36.7 mg/kg for rabbits when given orally, and 141.4 mg/kg for rats when given percutaneously. 3. Dieldrin. The dose of LD50 was 68.0 mg/kg for mice and 59.6 mg/kg for rabbits when given subcutaneously, 44.4 mg/kg for mice and 37.1 mg/kg for rabbits when given orally, and 186, 6 mg/kg for rats when given percutaneously. 4. Toxic symptoms observed were episodic generalized seizure, salivation, restlessness, inactivity, loss of appetite, irritability, epiphora, fever, tachycardia, paralysis of four limbs, dyspneá, and cyanosis,
The author iuduced acute, subacute, or chronic drin intoxication in rabbits and studied their peripheral blood pictures, myelograms, and histological changes. The results obtained are as follows. 1. Erythropoietic activities as reflected by red cell counts, hemoglobin, and reticulocyte counts were not significantly altered in comparison to controls by acute or chronic intoxication of drins. 2. In acute intoxication, particularly of endrin, there were leukocytosis and pseudoeosinophilia with relative lymphopenia and eosinopenia. Chronic drin intoxication caused no significant changes in the white blood cell series. 3. Hematopoietic activities of the bone marrow were hardly damaged in either acute or chronic intoxication of drins. 4. Histological changes induced by drins were almost identical in acute and chronic intoxication. The brain and spinal cord showed degeneration and necrosis of the ganglion cells, edema, congestion, and proliferation of the glia cells. In the liver there were congestion and degeneration of the liver cells. The kidneys revealed hemorrhagic glomerulonephritis and degeneration of tubular epithelium. The lungs also showed edema and hemorrhagic pneumonitis.
1. Endrin was responsible for all the cases of drin intoxication reported in Japan except one attributed to aldrin. No dieldrin intoxication has been reported. 2. Endrin was ingested with the suicidal purpose in 86% of the cases, intoxication due to its spray was rare. 3. Many of the accidents occurred betweeu the month of May and Novenber, and many of the victims were between the age of 20 and 30. More females were killed than males. 4. Oral intake of 3cc or 5cc of endrin was fatal in one case, respectively, and ingestion of its 10cc killed two persons. In these cases the fatal doses were estimated at 14.2, 19.5 and 48.8 mg/kg, respectively. 5. Cardinal symptoms included generalized seizure, loss of consciousness, vomiting, cyanosis, salivation, and agony. 6. Treatment consists of gastrolavage injection of cardiotonics and liver-protecting agents, and infnsion. Symptomatically sedatives and spasmolytics are also beneficial.
An attempt has been made to analyse electroencephalogram according to frenguecy analysis, Fluothane concentration in blood and clinical observation of anaesthetic depth under 2% Fluothane anaesthesia. Forty-two adults in both sexes were utilized whose electroencephalograms has been revealed normal under awakness. Demerol, Scoplamine and Pentobarbital were preliminarily given and semi-closed method of anaesthesia was done with either a mase or a tube which was inserted under 2% Xylocaine block of superiorlaryngeal nerves and of larynx directly through neck. Those premedications were hardly found to affect the E. E. G., but, under Fluothane anaesthesia, gradual decrement of the freguency and increment of the amplitude were to be noted. Frequency analysis also revealed under a tendency of gradual increment from βf zone to δs zone. It is to be noted that slow frequency zone was not necessarily to reach the basic freguency to gain the sufficient anaesthetic stage for surgery. Such E. E. G. findings were found to be in parallel with the Fluothane concentration in blood, and to correspond with the anaesthetic depth in clinical category.
The excitability of anterior horn cells was examined with an analysis of Magladery H-wave obtained by the method of provocation E. M. G. under Fluothane, Ether and Thiopentobarbital anaesthesias. Twenty-six adults in both sexes were selected for this study in which no abnormal findings were foune. H-wave was obtained at gastrocnemius muscle under awake and anaesthetized stages, and recovery curves were made of H-wave amplitudes, responded to successive stimuli of conditioning and testing through posterior tibial nerve. Prolongation of the duration of complete depression at the recovery curve or decrement of the excitability was found under Fluothane and Ether, and in contrant, no change of the excitability was found in case of Thiopentobarbital anaesthesia. No variation of M-wave was found in all cases of the anaesthesias, which revealed no influence on peripheral nerves and muscle fibers.
So many studies on the sex chromatin have been made so far in various fields since it was discovered as a nucleolar satellite by Barr and Bertram in 1949, but very little has been described about the sex chromatin in exfoliated cells of oral neoplasms. Thus the author started this investgation of the sex chromatin in oral smears in 60 normal persons (30 males and 30 females), 35 benign tumors (17 males and 18 females), 35 malignant tumors (22 males and 13 females) and 14 sexual anomalies. All smear specimens were stained with Moore and Barr's cresyl echt violet technigue, and a sex chromatin was always located along the inner surface of the nuclear membrane of interphase nuclei. The results obtained were as follows; 1) NORMAL CASES Microscopic examination was made on the sex chromatin in smear specimens taken from buccal, gingival and lingual epithelia of the same individual. Incidence of the sex chromatin in females was on an average 23.5±6.50 per cent in buccal smears, 22.0±5.65 per cent in gingival and 21.6±5.76 per cent in lingual, while that in males was ranging from 0 to 2 per cent in smears taken from each area. Minute examinations of sex chromatin showed that they were noticed in a variety of forms, such as triangular, planoconvex, flat and others, of which triangular form was most commonly seen. The average size of sex chromatin in these epithelial cells was abont 1.4×1.0 μ 2) TUMOR CASES There was almost no difference between sex chromatin incidences of tumor and its host tissue, except two cases of benign neoplasam, i.e. fibroma and follicular cyst, in females, whose sex chromatin incidences were 4 and 6 per cent. A relationship between sex chromatin incidence and Broders' classification could not be noticed in cases of carcinoma cells. 3) INTERSEX AND SEXUAL ANOMALIES As the result of investigations of sex chromatin in cases of pseudohermaphrotidism, infantilism, Turner's syndrome and hypospadias, the sex chromatin pattern did not always agree with its genetic sex in the patients associated with sex chromosome aberrations, therefore, it will be desirable to make analysis of sex chromosome in determining the sex in the above mentioned patients.
In the study on the appearance of L. E. cells and other phagocytic phenomena conducted with 54 patients consisting of hepatitis and liver cirrhosis the following results were obtained. 1. Among them four cases proved to be positive to the L. E. cell phenomenon. In the group who did not show the appearance of L. E. cells, those with liver cirrhosis revealed the highest incidence of pseudo-L. E. cell appearance. 2. With the four cases proved to be positive to the L. E. cell phenomenon a series of studies were carried out on their clinical pictures, the results of clinical tests, liver tissue pictures and serological tests and it was found that the Case 1 and 2 belonged to the category of lupoid hepatitis, the Case 3 aside from the L. E. cell phenomenon, did not differ so markedly from general liver cirrhosis, and the Case 4 appeared to be the case of an incomplete type of systemic lupus erythematosus complicated with liver disorders due to blood transfusion but the histological picture of the liver revealed peculiar, active hepatitis showing granuloma formation. 3. In these four cases the manner of L. E. cell appearance and the progress of disease did not parallel with each other. While the relation to the anti-liver antibody remains obscure, three out of the four showed a marked precipitin are of β2M-globulin. 4. A noteworthy fact was that those negative to the L. E. cell phenomenon but with persistent appearance of pseudo-L. E. cells among L. E. cell negative group mostly showed an aggravation and change of serum transaminase. However, relations between the appearance of pseudo-L. E. cells and other nucleophagocytes on one hand and anti-liver antibody and serum gamma globulin on the other could not be verified. 5. Likewise the relation between the appearance of pseudo-L. E. cells and the histological picture of the liver could not be clarified.
With patients of liver diseases mainly composed of hepatitis and liver cirrhosis as the subjects of the study, the assay of serum proteins was conducted by immunoelectrophoresis and simultaneously observations were carried out to see the clinical correlation of anti-liver antibody and changes in the quantities of β2M-globulin. The results were presented as follows. 1. It was noted that β2A- and β2M-globulin were increased markedly, especially the latter. Both of them were increased markedly already at the convalescent stage of acute hepatitis, and following this its rate of the increase was in the order of chronic hepatitis, precirrhosis and liver cirrhosis as the disease advanced more chronic. 2. The albumin precipitin arc in the seven cases of liver cirrhosis was shortened, and in one case of lupoid hepatitis complicated with marked hypergammaglobulinemia and, hypoalbuminemia it was strikingly elongated, reaching as far as the β1-globulin area. In the majority of liver cirrhosis the quantities of α2-1 or α2-2 globulin, β1B, β1C-globulins were decreased or were totally lacking. 3. In the entire cases with a positive titer of anti-liver antibody β2M-globulin was increased, whereas in those with a negative titer 23.3 per cent of them proved to be β2M-globulin negative. 4. Change or aggravation of the clinical coure was greater in those cases with an increase in β2M-globulin than in those without such an increase. Such a tendency was still more marked in those with appearance of anti-liver antibody. 5. In the liver histologic pictures necrosis of parenchymal cells, stellate cell reaction and infiltration into the interstitium were more maked in the cases with the increase of β2M-globulin than in those without, but there could be recognized no appreciable difference in between those showing anti-liver antibody and those without it. 6. There could be recognized differences in the values of S-GOT and S-GPT between those with increased β2M-globulin and those without the increase, and likewise a similar difference could be observed between those with increase of β2M-globulin accompanied by the appearance of anti-liver antibody and those without anti-liver antibody. A similar tendency was also observable in the decrease of A/G ratio, in the decreasing tendency of albumin, particularly in the increase of gamma globulin. 7. In the cases having increased β2M-globulin level accompanied by anti-liver antibody, there could be observed a tendency of decrease in the erythrocyte and leukocyte counts and an increase in lymphocytes. 8. In the observations of change in anti-liver antibody and β2M-globulin conducted for about three months the quantities of β2M-globulin were increased in all the cases with persistent appearance of anti-liver antibody and even in those with intermittent appearance of antiliver antibody, and also in those cases whose anti-liver antibody titer proved to be persistently negative. Steroid hormone therapy and splenectomy did not affect these changes.
The electron microscopic observation in patients and animals with drug induced cholestasis, extrahepatic jaundice and intrahepatic cholestasis have been discussed by many authors. But these investigations were almost about the patients and animals with clearly obssrved jaundice. So, in this study, I had the purpose to elucidate the microscopic pathological findings of the bile canaliculus and bile pigment in the liver cell, in the serum bilirubin I to 2 mg/dl and searched the mechanism of the latent jaundice. In order to accomplish this purpose a small piece of the biopsied spicimens was fixed in Palade's cold solution (bufferd osmium tetraoxide) and embedded in methacrylate. The materials comprised in this study was such cases as acute viral hepatitis, chronic viral hepatitis, liver cirrhosis, hemosiderosis and fat liver. The results are as follows: 1. The bile pigments were observed only in the liver cell or intracellular space of four cases from 24 cases of latent jaundice on light microscope. These cases were in the state of visible jaundice followed by latent jaundice. 2. But on submicroscopic observations, 17 cases from 24 of latent jaundice had the bile pigments in the liver cells. 3. The pathologic morphologic findings of the bile canaliculus in latent jaundice were dilatation of the lumen, the cytoplasmic substsances with condenced zone in the canaliculus, the channel in the cytoplasm, outpouching of canaliculus into the cytoplasm of the liver cell and the prolifelated bile canaliculus represented by a lumen bounded by many liver cell menbranes. These findings of the bile canaliculus were thought to mention the rising hydrostatic pressures within canaliculi. 4. The occluded canaliculus with distorted microvilli, vacuoles or mitochondria and the continuous and convoluted canaliculus were also observed. These were thought to represent the cause of rising pressure within canaliculus. 5. Both findings of the canaliculus which is seemed to be the cause of rising pressure within caualiculus or the state of the rising pressure were often observed in a same case. These morphological findings of the canaliculus in the same case were thought to be characteristic in latent jaundce. 6. The communication of the canaliculus with intercellular space was observed at the point of edematous microvilli in the canaliculi and occurence of latent jaundice may be interpreted by these two morphological changes of the bile canalculi.
The purpose of this study was to elucidate the submicroscopic pathological findings of the Kupffer's cell observed to be swollen and increased in number on light microscope. In order to accomplish this purpose a small piece of the biopsied spicimens was fixed in Palade's cold buffered osmium tetraoxide and embedded in methacrylate. The material comprised in this study was such cases as normal, diabetes mellitus, hepatoses, and chronic viral hepatitis. Further, in order to investigate the function of the Kupffer's cell the correlation with morphological findings such as phagocytosis and snbsequent disintegration was studied. Intravenonsly administration of S. F. O. was carried out in the rats injured by carbon tetrachloride injection in advance. The animals were killed 24 hours or one month after the treatment. The results are as follows: 1) Normal liver. Both cytoplasmic and nuclear menbranes of the Kupffer's cell were relatively even. The smooth and rough surfaced endoplasmic reticulum were relatively poor in the development. Mitochondria and Golgi apparatus remained small in size. 2) Diabetes mellius. The Kupffer's cell was provided with numerous smooth surfaced endoplasmic reticulum. Increased reticular fibers often were found in the space of Disse. The fat-storing cell (Ito) having large fat vacuoles (inclusious) was observed between the liver cells and the endothelial lining. This was clearly differenciated from the Kupffer's cell. 3) Chronic viral hepatitis. The swelling of the Kupffer's cell and the endothelial lining occluded the lumen of the sinusoid. Both cytoplasmic and nuclear menbranes had irregular surfaces. Numerous electron dense vacuoles and smooth surfaced endoplasmic reticulum were observed. These findings were thought to represent the state of hyperfunction. 4) The Kupffer's cell intoxicated by carbon tetrachloride and injected of S. F. O. showed increased numher of mitochondria, both rough and smooth surfaced endoplasmic reticulum and vacuoles, in comparison with those of normal rat liver. Phagocytosed S. F. O, were found to locate as aggregates with single menbrane or as electrou dence particles in the vacuoles, in normal rat, besides in the intoxicated cases, S. F. O. were scattered in to the hepatic cellular cytoplasm. The latter was observed to be in the state of hyperfunction in phagocytosis. In the case of chronic carbon tetrachloride intoxication, iron aggregates remained in ferritin as long as one month while disappeared mostly in the case of normal rat liver.