Okayama University Medical School

For the purpo3e to determine exactly what stage of cell specialization the DNA level of erythroid cell nuclei begins to decline, the author observed the DNA level of erythroblasts in mitosis by microspectrophotometry and the DNA synthesis by flash labeling with H3-thymidine. The cell samples were obtained from the bone marrow of normal, blood-depleted and phenylhydrazine-treated animals, and the anemic animals received a mass red cell transfusion, all the animals being injected with colchicine 4 hours before obtaining the bone marrow sample. DNA level was measured on the smeared cells stained by Feulgen reaction and DNA synthesis by autoradiography on the smeared cells. Besides these, chromosome number was observed on the anemic rat erythroblasts at metaphase by air dry method. The observations indicated that the DNA level begins to decrease at polychromatic stage being accompanied by a decrease in TDH3-incorporation into DNA, reaching minimum level at orthochromatic cell both in DNA contents and synthesis. Chromosome numbers of erythroblasts of rat were irregular being distri buted between 42 to 20. The data have suggested that the DNA level of erythroblasts decreases only in the later stages of cell specialization, and at polychromatic stage the chromosome number may also decrease in rabbit at polychromatic stage by the cell division with an incomplete DNA replication. The high DNA level of the erythroblasts of rabbit, in severe anemia where most of the cells are denucleated at polychromatic and late basophilic stages, has been discussed from the view point of the insufficient DNA replication at polychromatic stages.

The body wall of the cercaria of Schistosoma japonicum is covered with a thin integument which is connected to epithel cells located under the uscle layer. On the outer and the basal surfaces of the integument are seen thin limiting membranes. In the matrix of the integument are distributed numerous dense granules, vacuoles and spines. The rootlet of the spine is attached to the basement membrane of the integument. The circular and longitudinal muscle layers, both underlying the integument, have smooth muscle fibers composed of thick and thin myofilaments. The cercaria possesses five pairs of secretion gland cells which are divided into two groups of three anterior and two posterior pairs. Both gland cells are filled with secretion balls. The tail of cercaria is likewise covered with a thin integumen t, whose structure is identically the same as the body integument. Beneath the integument are located thin circular and longitudinal muscle layers. The circular muscle cells have smooth muscle fibers, but the longitudinal muscle cells have striated muscle fibers. These muscle cells contain many large mitochondria. On observing the cross-sections of the tail at the flame cell level the arrangement of these muscle can be divided into four muscle groups and each muscle group reveals four or five muscle cells. The excretory system is well developed and has flame cells, excretory canal and bladder.

To obtain some information of the biological action of Kankohso 101dinicotinate and Kankohso 301-nicotinate, observations were made on the binding mode of these substances with protein, chondroitin sulfate and nucleic acids and the following results were obtained; 1. Kankohso 10 I-dinicotinate binds reversively with bovine serum albumin or serum r-globulin, resulting in metachromasia. By binding with proteins the absorption maximum of the dye shifts toward the long wave length side and the absorbance decreased distinctly. The data show that there are more than one kind of binding sites and the binding with bovine serum albumin is weak in acidic solution and strong in alkaline solution. 2. Kankohso 10 I-dinicotinate produces strong metachromasia with sodium chondroitin sulfate and the color of the solution changes from violet blue to reddish violet. The absorption maximum at 592 mp. decreases without shifting its wave length ,and the shoulder appears at 555 mp. be. comes distinct peak. The strongest metachromatical changes occurs at the concentration of the chondroitinsulfate whose sulfonate radicals is equal to the molecules of Kankohso 10 I-dinicotinate. 3. Kankohso IOI-dinicotinate produces metachromasia with nucleic acid, where absorption spectrum is shifted toward long wave length and absorbance is decreased at a certain concentration. 4. Kankohso 301.nicotinate binds weakly with bovine serum albumin, the binding of which is reversible and the maximum binding number is 1.1 per molecule of albumin. Metachromasia cannot be produced by binding. Kankohso 30I.nicotinate does not bind with bovine serum γ-globulin. This compund does not produce metachromasia with sodium chondroitin sulfate but produces weak metachromasia with nucleic acid, indicating some affinity to nucleic acid.

In order to ascertain whether black-crowned night herons (BCNH), white heron (Plumed Egrets (PE)) and domestic fowls are infected by JE virus and they serve as infection source ofJE, hemoagglutination inhibiting antibody and its 2·ME sensitive antibody in the sera of these birds were determined. Physico-chemical nature of fowl's antibody of JE produced by natural infection and their maternal antibody in the sera of chicks were examined. The results are briefly summarized as follows. 1) As to the herons captured in Tsudaka Town, two out of six adult night herons and three out of the four chicks showed positive HI reaction. On the other hand, HI reaction in the sera of two adult white herons and three chicks were negative. 2) As to the herons captured in Okayama City, twenty out of thirtytwo adult night herons and seven out of seventy white herons showed positive HI reaction in 1966 around the time when JE was prevalent in Okayama Prefecture. And six out of eleven night herons and one out of seven white herons showing positive HI reaction, responded positively to 2-ME sensitivity test. 3) The results indicate that white herons can be also infection source ofJE though less than in the case of night herons. 4) In the domestic fowls (white leghorn) kept at Takahashi District, eight out of twenty-seven fowls showed positive HI reaction. And six out of seven domestic fowls showing positive HI reaction responded positively to 2-ME sensitive reaction. 5) Transformation of JE antibody in the serum of hen from IgM to IgG was recognized. 6) Domestic chicken's sera having 1 : 640 of HI titer in the original serum and 1 : 320 of HI titer after 2-ME treatment were fractionated by gel filtration on Sephadex G-200 and the antibody activities present in the various fractions were determined. HI antibody activities occurred in both IgM and IgG classes of immunoglobulins. 7) Maternal HI antibodies reacting with JE virus were found in newly hatched domestic chickens from the eggs laid by hens with natural infection ofJE. And half life of HI antibodies in chicks was four days. 8) HI antibodies of JE in the serum of maternal immune-hens and chicken having maternal antibody were located in r-globulin fraction by starch block electrophoresis. 9) The results from 4) to 8) indicate the presence of natural infection ofJE in the domestic fowls. And domestic fowls can be infection source ofJE.

Exfoliative cytologic studies on normal oral cavity and on the postexodontic wound healing of infants were carried out and the following results were obtained: 1) The keratinization of cells was found marked in such regions as buccal mucosa, mucobuccal fold, gingiva and palate in that order. 2) As for changes in the distribution of cells and leucocytes, the cell distribution in the period of 15-21 postexodontic days proved to be identical with that of normal exfoliated cells. Namely, the wound healing from exfoliative cytologic aspects takes place during the period of 15-21 postexodontic days.

Die Autoren berichten Uber die Forschungen mit Hilfe von Kernreaktoren auf dem Grenzgebiet der Medizin und Strahlenphysik (Reaktorphysik). Es wurden kurz ihre eigene bisherige Ergebnisse, das Ziel dieser Forschungen und einzelne Zukunftsmoglichkeiten erwahnt. Nachdem wurden in mehreren Teilen die Kernreaktoren beschrieben. 1m erstem Teil werden die allgemeinen physikalischen Grundlagen eines Kernreaktors dargestellt. Mit Hilfe der Kernkrafte und der Bindungsenergie wird gezeigt, wieso eine Kernspaltung auftreten kann und wieso aus den nicht spaltbaren Isotopen U-238 die spaltbaren Isotope Pu-239 bzw. U-233 "erbrutet" werden konnen. -Das Zustandekommen einer Kettenreaktion, die die Grundlage einer Energiegewinnung aus der Kernspaltung ist, wird erlautert, wobei sich auf die Wechselwirkungen zwischen Atomkernen und Neutronen hingewiesen wird. 1m AnschluB daran werden die Vorgange im Kernreaktor im Einzelnen naher verfolgt. Dabei werden mehrere Moglichkeiten beschriebenk die es erlauben, die Neutronenzahl, die fur die Kettenreaktion entscheidend ist, zu erhohen. Einer allgemeinen Dbersicht uber die verschiedenen Reaktortypen und ihreu Einsatz in Forschung und Industrie schlie13t sich eine Beschreibung verschiedener deutscher Reaktoren und ihrer speziellen Eigenschaften und Vorteile an. Zum SchluB wird noch besonders auf die groBen Projekte der Entwicklung von Thorium-Brutern und von schnellen Brutern eingegangen.

For the purpose to obtain the information of the mechanism of protein uptake by the tumor cells, some cytochemical and electron microscopic observations were carried out on Ehrlich ascites tumor cells incubated with horseradish peroxidase (basic hemoprotein, molecular weight approximately 40,000) in vitro. In the earlier periods of the incubation peroxidase was found to be adsorbed on some area of surface of the tumor cells forming a thin protein layer, where an active pseudopodia formation was observed. With the lapse of time, the protein was taken in the deep cytoplasm by the infoldings of the cell membrane and accumulated in the cytoplasmic vesicles having limiting membrane. Concerning the accumulation of the protein into the vesicles, small tubular structures in the cytoplasm connecting the cell surface and the vesicles, were considered to participate in the intracellular transportation of peroxidase taken up. In cold environment (2°C), the formation of pseudopodia and deep inward infoldings of the cell membrane was inhibited and simultaneously the uptake of peroxidase stopped. Iodoacetate and sodium fluoride also effected to suppress the pseudopodia and infoldings formation moderetely, as well as uptake of peroxidase, though they did not stop completely. These facts have indicated that horseradish peroxdase is taken up by Ehrlich ascites tuimor cells through pinocytosis which involves energy-requiring process dependent upon glycolytic metabolism of the tumor cells.

For the purpose to reveal the changes in the metabolism of erythroblast in varied specialization stages the author observed the Feulgen DNA level of rabbit erythroblasts by microspectrophotometry. Observations were made on normal and anemic animals, and those receiving a mass red cell transfusion at the recovery stage of anemia where the early denucleation is stimulated. Observations have revealed that in normal erythropoiesis the DNA contents are kept at n to 2 n level from the proerythroblast to late basophilic stage, but in later stages, polychromatic and orthochromatic, DNA level per cell decreases gradually with advance of the cell specialization reaching the minimum level, nearly haploid level, at orthochromatic stage where most cells are believed to be denucleated. In blood depleted animals nearly the same pattern of DNA level was observed in connection with erythroid specialization as that in normal animal, except a relatively high DNA level in the later specialization. In the cases of the hemolytic anemia a similar tendency has been observed but the minimum level of DNA remains at a higher level, hypo-diploid level, in poly- and orthochromatic stages. Twenty-four hours after the mass red cell transfusion by which severe anemia has been recovered to the original level within one hour, the pattern of the DNA level of the erythroblast returns to the normal one showing a very low DNA level at the polyand the orthochromatic stages. The data indicate that the DNA synthesis of erythroblast kept at n to 2 n levels until the late basophilic stage begins to decline at polychromatic stage and reaches nearly haploid level at orthochromatic stage, but in active hemopoiesis the DNA synthesis is stimulated and the DNA contents are kept at a high level even in the late specialization stages, showing no relation between the denucleation and the low DNA level.

Localization of IgM and IgG sypylitic antibodies in the sera of patients and the experimental syphylitic rabbits was examined by the gel filtration on Sephadex G.200 column. I) In the case of late syphylitic patients; OGATA test-reactive antibodies were contained in IgM and IgG fractions. On the other hand, RPCF test-reactive antibody was contained only in IgG fraction. This discrepancy may be due to the difference in antigens; Cardiolipin.resicin and T. P. Reiter protein. 2) In the case of the experimental syphylitic rabbits; The results were as follows. a) Variation in the level of the titer. The peaks of the titer were seen 3-4 weeks after inoculation of T. P. Nichols by OGATA test, VDRL test and RPCF test, thereafter titers decreased. On the other hand, the titer kept on rising up to 2 months and maintained high level during the periods of 3, 4 and 5 months by TPHA test. b)Transformation of antibody from IgM to IgG. Transformation of antibody from IgM to IgG was seen 3-4 weeks after inoculation by all four tests; OGATA test, VDRL test, RPCF test and TPHA test, and such a transformation was completed 3 months after inoculation.

1) Normal karyotype of Donryu strain rat was determined according to the classification of KURITA et al. (8). Namely, the number of chromosomes was 42 in diploid cells, and chromosomes were divided into 3 groups (A, B and C) according to the position of centromere. A.group was consisted of 7 pairs of metacentric chromosomes, B-group 4 pairs of sub· meta.subtelocentrics and C.group 10 pairs of telocentrics and Y. 2) Among all chromosome pairs a pair of the longest telocentric chromosomes (C.l), 4 pairs of all the B.group, and the Y chromosome were recognizable. 3) The presence of polymorphism was demonstrated in the smallest submetacentric chromosomes (BA),: namely, (I) a homologous submeta. centric pair, (II) a homologous subtelocentric pair and (III) a heteromor. phic submeta and subtelocentric pair which seemed to be a hybrid from (I) and (II). To distinguish the polymorphism in their genotype from phenotype was impossible. 4) Animals with type III B·4 chromosomes were produced by type I and type II animals. 5) By checking the chromosomes of the inbred Donryu strain rats maintained over 40 generations by brother.sister mating at Nihon Rat Co., polymorphism in BA chromosomes was also recognized.

The role of -SH groups in mitochondrial energy transfer reaction was studied by observing the reduction of a disulphide, 5, 5'-dithiobis (2-nitrobenzoic acid), DTNB, a specific analytical agent for the estimation of -SH groups in biological materials, by addition of it to the isolated rat liver mitochondria in various respiratory states, as defined by CHANCE and WILLIAMS. 1. In the various respiratory states, states 1 to 5, the reduction of DTNB proceeds most rapidly at state 5, and most slowly at state 3. DTNB reduction at state 5 is suppressed by the partial oxidation of respiratory carriers with oxygen (state 4) and the addition of respiratory substrate does not affect the DTNB reduction. 2. The retardation in the reduction rate at state 3 is relieved partially by a respiratory inhibitor, KCN, and is intensified markedly by oligomycin, an inhibitor of oxidative phosphorylation. An uncoupler for oxidative phosphorylation, DNP, does not affect the reduction rate at state 3. At state 4 the reduction is stimulated by DNP and KCN, but is unaffected by oligomycin. The results suggest that the alteration in the functions of the energy transfer reaction in mitochondria is accompanied by changes in the occurrence and the functioning of -SH groups which can be detected by the reactivity with DTNB. The data suggest also that there are at least two kinds of -SH groups reacting with DTNB: the one is the -SH group which reacts DTNB actively when the respiratory carriers are kept reduced, and the other is the one which reacts actively when the respiratory carriers are kept oxidized, participating in the phosphorylating system and its reactivity with DTNB diminishes in the actively phosphorylating states (states 2 and 3).

1) It has been found that the peripheral blood of hybrid adult dogs contains about 29.4 % of lymphocytes in average. When such a blood is passed through the glass wool column the leucocytes containing 68-90 % (81.6% in average) lymphocytes are obtained. 2) In the single culture of such lymphocytes alone and mixed culture both live lymphocytes and sonicated lymphocytes in the presence of 1% (vIv) PHA, the peak of the blastformation of lymphocytes is observed at culture hour 72. 3) In the abscence of PHA both single culture and mixed culture of lymphocytes show hardly any blastformation. 4) In the mixed culture of live lymphocytes with homogenate of sonicated lymphocytes with addition of 1% (vIv) PHA the rate of blast. formation observable at culture hour 72 and the rejection of kidney trans· plant 7 days after its transplantation of hybrid adult dogs show a direct correlation, demonstrating that the mixed lymphocyte reaction reflects accurately the difference in donor histocompatibility antigens against the recipient.

Eine Salieyl-Chinin-Lithium-Kombination wurde im Siemens Unterriehtsreaktor SUR 100 BE des Instituts fur Kernteehnik der Teehnisehen UniversiUit Berlin mit Neutronen aktiviert. AnsehlieBend erfolgte die Messung der spektralen Verteilung der Gamma-Zahlrate zu zwei versehiedenen Zeitpunkten naeh der Aktivierung. Es wird festgestellt, daB diese Salieyl-Chinin-Lithium-Kombination nur sehwaeh im SUR-Reaktor aktivierbar ist, vergliehen mit einer Goldsonde etwa 200 mal sehwaeher. Trotzdem konnten mit Hilfe eines 400-Kanal-Analysators die spektralen Verteilungen der Gamma-Energien dieses Praparates noch sehr genau bestimmt werden. Ein ausgepragter Gamma-Peak tritt auf bei der Energie 0.53 MeV. Er klingt ab mit einer Halbwertszeit von 1.6 h und ist vermut· lich auf ein Element der Tablettierhilfsstoffe zurUckzufUhren. Vom strahlenphysikalischen Gesichtspunkt ist das Praparat fUr die Human. medizin gut geeignet.

With the purpose to clarified the mode of localization and mechanisms of activation of ATPase in the mitochondrial membrane, analyses were made on the properties of mitochondrial ATPase from the structural and functional aspects. The activation of ATPase by DNP and Mg++ and the oligomycin sensitivity were investigated in a series of inner membrane fragment samples obtained by ultrasonic irradiation and those samples obtained in the processes of isolation and purification of ATPase from rat liver mitochondria and beef heart mitochondria in parallel with electron microscope observations. As a result it has been found that the membrane fragments obtained from rat liver and beef heart mitochondria by ultrasonication exhibited high respiratory activity and unmasked ATPase activity which was charac· terized by remarkable stimulation by Mg++ and inhibition by oligomycin and azide. Therefore, mitochondrial ATPase seems to be bound fairly closely to the inner mitochondrial membrane. In the membrane fragments prepared by ultrasonication of intact mitochondria, ATPase activity was stimulated by DNP, but in the supernatant fractions was not. On the other hand, the supernatant fraction obtained from BHM and inner membrane fragments by severe sonication exhibits a marked ATPase activity and the activity incresed in each step of the purification on the treatments with acid, protamine and heat. Especially in the case of membrane fragments the protamine treatment can be omitted. Electron microscope observation of the fractions in each step of the purification proved the head pieces to be ATPase. The ATPase activity of solubilized head pieces is insensitive to oligo. mycin and coincides with the soluble ATPase of PULLMAN etat. (8) in the points of its cold labile property and optimum pH, but it shown no accele. ration of ATPase activity by DNP.

1. Mitochondria isolated from human liver, hepatoma and gastric cancer contain DNA. The DNA content per mitochondrial protein is about ten times as much in cancer as in normal liver. 2. Human liver, hepatoma and gastric cancer contain circular DNA molecules in their mitochondria. Circular DNAs from normal liver and cancer mitochondria are mostly about 5 μ long, and the frequency of circular DNAs of multiple or shorter length is higher in cancer mitochondrial DNA. The outline of the present paper was presented at the 26th Congress of Japanese Cancer Association (1967) (52, 53).

The amygdalofugal fibers were studied III the cat with the silver method of NAUTA-GYGAX. 1. The amygdalofugal fibers are distributed by way of the stria terminalis, the longitudinal association bundle, the inferior thalamic peduncle, and the medial forebrain bundle. 2. The amygdalofugal fibers running through the longitudinal association bundle arise in the lateral principal, intermediate principal nuclei and the lateral and possibly intermediate parts of the periamygdaloid cortex, and terminate in the lateral preoptic nucleus, the bed nucleus of the anterior commissure, the olfactory tubercle, the nucleus of the diagonal band of Broca, the nucleus accumbens, the medial and posterior septal nuclei and the basal part of the head of the caudate nucleus. In addition, there are scattered fibers coursing along the longitudinal association bundle proper. These fibers may have a widespread origin from the amygdaloid complex. The longitudinal association bundle contributes no fibers to the medial forebrain bundle. 3. The fibers, originating from the lateral principal, intermediate principal and medial principal nuclei, join the medial forebrain bundle to distribute widely in the lateral hypothalamic nucleus. A few fibers are seen to reach the ventromedial hypothalamic nucleus, and are considered to arise in the medial principal nucleus. 4. By way of the inferior thalamic peduncle some fibers from the amygdaloid complex course dorsally into the medial part of the dorsomedial thalamic nucleus at its caudal levels. They may arise widely from the amygdaloid complex. A few of them extend farther dorsally to reach the lateral habenular nucleus and the parataenial nucleus. They probably originate from the lateral principal nucleus. 5. The fibers forming the stria terminalis originate from the medial principal nucleus, the medial nucleus, the periamygdaloid cortex and the cortical nucleus, and are distributed in the bed nucleus of the stria terminalis and the lateral preoptic nucleus (preoptic component), as well as the medial preoptic nucleus, the anterior hypothalamic nucleus and the ventromedial hypothalamic nucleus (supracommissural component). The cortical nucleus, particularly its caudal part, and possibly the medial part of the periamygdaloid cortex are regarded as the main sources of the stria terminalis fibers ending in the hypothalamic region. The intermediate principal and lateral principal nuclei do not appear to contribute fibers to the stria terminalis. 6. The ventromedial hypothalamic nucleus receives amygdalofugal fibers both from the medial principal nucleus by way of the medial forebrain bundle, and from the cortical nucleus via the stria terminalis. 7. In addition to intrinsic internuclear fibers within the amygdaloid complex, some of the fibers from the complex are distributed to the ventralmost part of the putamen, the medial part of the claustrum, the periamygdaloid cortex, the prepiriform area and the anterior amygdaloid area, but do not reach the hippocampus.

As a step towards the elimination of Japanese encephalitis virus in natural surroundings, we inoculated pigs, rabbits and chicks with inactivated Japanese encephalitis vaccine supplemented with complete or incomplete Freund's adjuvant twice at one-week interval. Subsequently, we compared HI antibody titers of the groups inoculated with vaccine containing complete Freund's adjuvant (pigs, rabbits, chicks), of the group inoculated with vaccine containing incomplete adjuvant (rabbits), ar;d of the groups inoculated with vaccine containing no adjuvant (pigs, rabbits, chicks), and also observations on changes in the antibody titers due to natural infection. In a certain portion of these animals neutralizing antibody titers were also determined. The results of this study are briefly summarized as follows. 1. In the groups of pigs and rabbits inoculated with vaccine containing complete Freund's adjuvant, titers of HI antibody and neutralizing antibody were higher than those inoculated with vaccine containing no adjuvant and their high titers persisted. Further, in the group of chicks inoculated with inactivated Japanese encephalitis vaccine containing complete Freund's adjuvant, HI antibody titers were higher and persistent as compared with the antibody titers in the chicks inoculated with inactivated Japanese encephalitis vaccine alone. 2. In the rabbits inoculated with inactivated Japanese encephalitis vaccine contammg incomplete adjuvant, HI antibody titers were lower than in those receiving the vaccine with complete adjuvant, but it has been demonstrated clearly that vaccination of inactivated Japanese encephalitis vaccine supplemented with incomplete adjuvant brings about less sideeffects. Hence such a method of vaccination can be applied as the vaccination with least side-effects. 3. With respect to natural infection of swine, on August 27 when the pigs were thought to have been infected, there was observed a rise in antibody titers. And on being infected with Japanese encephalitis, the antibodies formed in those pigs inoculated with inactivated Japanese ence- phalitis vaccine with or without complete adjuvant proved to be all 2-ME resistant type, whereas the antibodies produced in the control groups not receiving such a vaccination were 2-ME sensitive antibody.

Five (21 per cent) out of 24 mongrel dogs were found to be refractory to compound 48/80 and also to sinomenine (cross-tolerance). These nonreactor dogs responded normally to PVP and tween 20 and showed normal sensitivity to histamine. The incidence was similar in both sexes. Mechanisms of this type refractoriness were discussed.

Die physikalischen Grundlagen eines Kernreaktors werden beschrieben. Als spezielles Beispiel wird Aufbau, Kontrolle und Betrieb des Siemens-Unterrichtsreaktors SUR 100 beschrieben. Dieser homogene polyathylenmoderierte Nullleistungsreaktor hat eine Leistung von nur 100 mW. Trotzdem-oder gerade deswegen-ist dieser Reaktor fur Ausbildungszwecke und als Ubungsmoglichkeit auf dem Gebiet der Reaktortheorie und der Kernenergie sehr gut geeignet, denn die Leistungsbeschrankung erlaubt eine einfache Installierung und Betriebsweise des Reaktors. Neben seiner Verwendung als Ausbildungsinstrument kann dieser Reaktor aber auch als Strahlenquelle benutzt werden. Hiermit wurden verschiedene medizinische Praparate, die auch im Strahlenschutz Verwendung finden, bestrahlt, und anschlieBend ihre Dosisrate bestimmt. AuBerdem werden noch weitere Anwendungsmoglichkeiten des Siemens-Unterrichtsreaktors SUR 100 deschrieben.