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From these findings, we confirmed that the tongue-like process formations of the cells which are still believed as the platelets formation by many investigators, would be nothing but the presentation of the cell degeneration and platelets are separated only from the tips of tentacles.

In an attempt to pursue the relationship of the fine structure of a cell to the biochemical function, the author at first tried to demonstrate cytochemically the actual sites of activity of enzymes in the terminal electron transport system involved in energy production with the use of the electron microscope. Namely, cytochemical reactions were performed by using potassium tellurite, a heavy metal salt, and then the author succeeded in the electron microscopic detection of the enzymes by freezing-drying method and by means of formalin fixation, strong reducing agents and osmium tetroxide fixation. As the results the author has been able to verify that the reactions of the enzymes belonging to the terminal electron transport system are found localizing in the mitochondria being arranged fairly densely and continuously on the critae and partially on the membrane, although some differences in the grade of the activity are found in each mitochondria even in one cell and a marked difference between the mitocndria belonging to the different kinds of cells. Furthermore, it has been clarified that the activity of the endogenous dehydrogenase system (mainly DPNH- or TPNH-dehydrogenase and others) is chiefly strong in cristae, and that the succinoxidase system exists both in cristae and membrane.

With an improved method of tissue culture of peripheral leucocytes of our own design, the authors carried out systematic observations on the peripheral leucocyte culture from the normal, various kinds of leukemias, leukemoid reactions, and hypoplastic anemia. As for the culture method we have devised a method of silicon oil coating on the blood containers, a method which will least affect the cell function as compared with the conventional culture method. As the results we have found that the tissue growth in the case of peripheral leucocyte culture of normal persons ceases after six-hour culture and also we have recognized a peculiar finding, a growth like a corona-shape, in which an empty space appears in the inner part of growth area along with the lapse of time. In every leukemic case, without presenting growth area like a corona, the cell density is high and the outer zone of growth area becomes sharply demarcated as in the case with bone-marrow culture and also the growth continues even after 12 hours. At this instance we have noticed many mitoses of immature cells. Moreover, the maturation of immature cells has been observed and it has also been possible to distinguish to what type these immature cells belonged. In the leukemoid reaction no growth pattern Characteristic to the leukemia can be recognized, and at a glance it can easily be differentiated from the leukemia. In the case of hypoplastic anemia although the growth area is like that of the normal presenting a corona-like shape, the cell density is lower and the function of leucccytes is lesser than the normal. From these results we believe that the peripheral leucocyte culture is clinically useful.

New histochemical and cytochemical methods for the demonstration of cytochrome c oxidase and of cytochrome c-cytochrome oxidase system are described, using neotetrazolium chloride in the presence of p phenylenediamine with or without additional cytochrome c. These enable the cytochemical visualization of the sites of enzyme activity at the intracellular level in fresh cell suspensions and in fresh tissue blocks under aerobic conditions, and permit the histochemical visualization of the distribution of the enzyme activity in various tissues in frozen tissue sections. The colorimetric estimation of the enzyme activity is also possible in the combination of the methods previously described.

1. Considering an imaginable important role of histamine in inflammations or allergic reactions, some clinical and experimental studies were made for the purpose of findings the relationship of the pathological changes of A. P. to local histamine. 2. Gingivae in the case of A. P. showed a remarkable quantitative increase of histamine, compared with those in the case of normal healthy controls. It could be noticed that this increase had an increasing tendency in proportion to the degree of inflammatory pathologic changes in the gingiva. 3. In the gingivae of A. P. an increase of mast cells and their morphologic changes, particularly disintegration, were seen and the grade of changes was almost parallel to an increase of histamine. 4. When a histamine solution was repeatedly injected into the mucobuccal folds of animals, a remarkable increase of histamine and mast cells in the injected gingivae were recognized with inflammatory changes. An increase of gingival histamine could be hardly recognized by means of a simple mechanical stimulation or local anaphylaxis alone. However, in the combination of two, the increase of histamine was seen and relatively remarkable inflammatory changes suggestive of A. P. were macroscopically and microscopically noticed. 5. Topical applications of the ointment mixed with sinomenine, histamine-liberating substance, and benadryl, antihistamine substance, repeated every other day in the gingival pockets, showed favorable effects in slight or early cases of A. P. 6. These observations suggest the existence of both possible role of histamine in the development of local pathologic condition of A. P. and allergic processes in etiologic factors of the disease.

Mast cells in osteoarthritic and rheumatoid arthritic synovial tissues of the human knee.

Helicobacter pylori (H. pylori) infection in the stomach is etiologically closely associated with chronic active gastritis, peptic ulcer, gastric cancer and gastric mucosa-associated lymphoid tissue lymphoma. In this study, we examined the antibody responses and cytokine profiles of three strains of mice (BALB/c, C3H/He, and C57BL/6) infected with H. pylori. Following this, correlations between host-immune reactions and intensity of inflammation were analyzed. H. pylori (ATCC43504) was intragastrically administered once a week to the mice from 4 weeks of age, and they were sacrificed at the ages of 4 and 7 months. In these mice, we examined the histology of the stomach, antibody titers against H. pylori, and serum levels of cytokines (IL-4, IL-10, TNF-alpha, IL-2 and Interferon-gamma). In BALB/c mice, inflammation of the stomach was minimal. Inflammation was observed in 63.6% of C57BL/6 mice and 33.3% of C3h/He mice. In C57BL/6 and C3H/He mice, all the cytokines tended to increase. In contrast, BALB/c mice were inactive in cytokine production except for IL-2. Two C3H/He mice developed severe inflammation with lymph follicles; one showed a response largely typical of Th-1, and the other showed a response largely typical of Th-2. Although a definite correlation was not shown between Th-1/Th-2 response evaluated by cytokine production and intensity of inflammation, it appears that in H. pylori-induced inflammation both cell-mediated (Th-1) and humoral (Th-2) immunity play a role in pathogenesis.

The hematological findings of 233 patients with brucellosis are presented and the possible pathologies discussed. Anemia was present in 128 patients (55%), leukopenia in 49 (21%) and thrombocytopenia in 59 (26%). Bone marrow aspirates of 18 patients (8%) with pancytopenia were examined. The bone marrow was hypercellular in 15 and normocellular in 3 patients. Granulomatous lesions were detected in 12 cases (67%), and slight to moderate cytophagocytosis of erythrocytes, granulocytes and platelets existed in all patients. Blood cell counts reverted to normal within 2-3 weeks of initiating chemotherapy with recovery from the disease.

To test transdifferentiation of retinal pigment epithelial (RPE) cells in suspension culture, chick embryonic RPE sheets and dissociated RPE cells were cultured for two months in a non-adherent dish for suspension culture. RPE cells, isolated as a sheet, aggregated immediately and remained the same size with their differentiated characteristics for two months. The presence of basic fibroblast growth factor (bFGF) at concentrations of 10 ng/ml or higher induced the formation of a spherical lentoid structure which was positive for crystallin and bFGF receptor. In contrast, dissociated RPE cells did reaggregate but did not develop the lentoid structure even in the presence of bFGF. The transdifferentiation of RPE cells to the lentoid structure in this study was in contrast to their transdifferentiation to the retina, as reported in a previous study.

To study the expression of the amiloride-sensitive sodium channel, a putative mechano-receptor in the rat eye, reverse transcriptase-polymerase chain reaction and in situ hybridization were done. The gene for the alpha subunit of the amiloride-sensitive sodium channel was shown by polymerase chain reaction to be expressed in mRNA isolated from the whole eye tissue. In situ hybridization demonstrated that the gene was expressed in basal layers of the corneal and conjunctival epithelium, ciliary epithelial cells, lens epithelial cells at the equator, retinal and iris pigment epithelial cells, ganglion cells and cells in the inner and outer nuclear layers of the retina. The results suggest that the amiloride-sensitive sodium channel plays a role in maintaining sodium balance as well as in possible mechanosensation in these ocular tissues.

Intrathymic (i.t.) injection of allogenic cells without administration of anti-lymphocyte serum (ALS) in neonatal recipients has induced donor-specific tolerance to subsequent cardiac allografts in rats. This study examines whether similar tactics can be successfully applied to a hamster-to-rat cardiac xenotransplantation model. Lewis neonates on their first day of life underwent i.t., subcutaneous (s.c.), intraperitoneal (i.p.), or intravenous (i.v.) injections of 5 x 10(7) Golden Syrian hamster splenocytes. After six weeks, the rats underwent heterotopic cardiac transplantation of hamster hearts. Cyclophosphamide (CyP) was administered on the day before surgery and postoperatively to suppress antibody-mediated graft rejection. Rats given splenocytes with 80 mg/kg of CyP had the following graft survival times: 8 to 12 days for i.t. injection (mean, 9.4 days); 5 to 7 days for s.c. injection (mean, 6.6 days); 4 to 11 days for i.p. injection (mean, 7.4 days); and 4 to 13 days for i.v. injection (mean, 7.9 days). Only the extension of graft survival produced by i.t. injection was statistically significant in comparison with the rats given only CyP treatment (mean, 7.5 days; P < 0.05). Thus, it appears that i.t. injection of xenogenic splenocytes in neonatal recipients with administration of CyP, but without ALS, can prolong xenograft survival. This biological intervention may be most useful in pediatric xenotransplantation when combined with other immunomodulation techniques.

In order to elucidate factors influencing the prognosis of small-cell lung cancer (SCLC), we reviewed the records of 253 patients with SCLC and evaluated 20 pretreatment prognostic factors by univariate analysis and Cox's multiple regression analysis. Recursive partitioning and amalgamation (RPA) was employed to identify subgroups with similar survival rates. Cox's multiple regression analysis identified five significant factors: extent of disease, number of metastatic sites, serum albumin, serum lactate dehydrogenase, and presence of weight loss. Among these, extent of disease was the most influential factor. RPA analysis revealed three subgroups predicting significantly different prognoses. The median survival time and 3-year survival rate were 18.4 months and 20.6%, respectively for the good-risk group (limited disease without weight loss), 13.5 months and 9.1%, respectively for the intermediate-risk group (limited disease with weight loss or extensive disease with less than two metastatic sites), and 9.2 months and 0%, respectively for the poor-risk group (extensive disease with two or more metastatic sites). These results will be useful for development of new staging system or subsequent stratification for randomized trials.

POSSUM, a Physiological and Operative Severity Score for the enUmeration of Mortality and morbidity, is a scoring system which assesses perioperative surgical risks (Copeland GP et al.: Br J Surg, 1991, Vol 78, 356-360). The POSSUM scoring system consists of two categories of assessment to assess the risk of surgery. A 12-factor (age, cardiac status, pulse rate, systolic blood pressure, respiratory status, Glasgow Coma Score, serum concentration of urea, potassium and sodium, hemoglobin concentration, white cell count and findings on electrocardiography) and 4-grade physiological score (PS) were developed. This was combined with a 6-factor (type of surgical procedure, number of procedures, blood loss, peritoneal soiling, presence of malignancy and mode of surgery) and 4-grade operative severity score (OSS). The present paper attempts to validate it retrospectively. Postoperative hospitalization period and duration of antibiotics administration were both significantly correlated with OSS, but not with PS. These results suggest that the POSSUM scoring system is useful for predicting the postoperative clinical course.

The ultrastructural and biochemical changes in the brain tissue of 20 mice were studied. The mice, separated into acute and chronic groups, were injected with phorbol myristate acetate (PMA) to trigger the release of free radicals. Malondialdehyde measurement and electron microscopic examination were applied for the evaluation of the effects of the free radicals. The level of lipid peroxide in the chronic PMA group was found to be significantly higher than it was in the acute PMA group (P < 0.005). An electron microscopic examination of the acute group revealed disruption of the mitochondrial cristae and dilatation of the endoplasmic reticulum in the neurons. Myelin sheaths of the nerve fibers exhibited focal structural changes. Neurons and neuroglial cells in the chronic group, however, exhibited distinct ultrastructural alterations. The ultrastructural and biochemical findings showed that free radicals lead to brain damage.

Thomsen-Friedenreich antigen (T antigen) has been supposed to be a cancer-specific carbohydrate antigen. We have previously shown that one third of the Japanese population normally expressed T antigen in gastric surface epithelia and the other two thirds expressed fucosyl-T antigen. Their sialylation and blocked-synthesis were associated with diseased conditions. In the present study, we studied gastric surface epithelial expression of monosaccharide antigen Tn, i.e., a precursor of T antigen, and sialyl-Tn. Normal fundic and pyloric epithelia, respectively, expressed Tn supranucleally and cytoplasmically, but did not express sialyl-Tn. In the intestinal metaplasias and intestinal-type adenomas, goblet cells expressed sialyl-Tn in their vacuoles, and absorptive cells expressed Tn apically. In gastric-type adenomas, Tn, but not sialyl-Tn, was detected. Intestinal-type cancers expressed Tn and sialyl-Tn more often than the diffuse-type cancers. Five cancers did not express Tn, sialyl-Tn, or the T-related antigens. In these, four were diffuse-type cancers. We concluded that: a) normal gastric epithelial cells express Tn; b) metaplastic differentiation is associated with sialylation of Tn and c) expression of Tn and sialyl-Tn is depressed in the gastric cancers.

Rat liver and simian virus 40 (SV40) chromatin were reconstituted in vitro under physiological conditions of ionic strength and temperature. The nucleosome assembly under the conditions was mediated in the presence of chromatin extracts, rich in nicking-closing activity, from rat liver or cultured CV-1 cells. The frequency of nucleosome assembly on DNA was dependent on both the incubation time and the weight ratio of histone to DNA. The regulatory effects of host cellular histones on the transcription of SV40 DNA were investigated by using reconstituted SV40 chromatin containing or lacking histone H1. The cellular histones composing the chromatin were prepared from permissive CV-1 cells. Transcription of chromatin was analyzed in vitro using Escherichia coli DNA-dependent RNA polymerase. The rate of incorporation of ribonucleoside triphosphates into RNA synthesized on SV40 chromatin containing Hl as the template was 5 to 10% of the rate for RNA synthesized on supercoiled SV40 DNA. The rate of incorporation for SV40 chromatin lacking Hl was approximately 40 to 50% of that for SV40 DNA. RNA products transcribed from both these chromatin and SV40 DNA were fairly homogeneous 16 to 28S species with several identical peaks.

The distribution of corticotropin releasing factor (CRF) and arginine vasopressin (AVP) in hypothalamic nuclei were examined in control and estrogen-treated female rats. CRF activity was measured using monolayer cultured rat anterior pituitary cells and AVP by radioimmunoassay. Hypothalamic nuclei were punched out according to the method of Palkovits. The distribution of CRF activity in 5 different hypothalamic nuclei was similar to that of AVP in intact female rats. CRF activity in hypothalamic nuclei, pituitary ACTH content and plasma ACTH levels in estrogen-treated rats were not significantly different from those in control rats. However, significant elevation of AVP content was observed in the supraoptic and paraventricular nuclei of estrogen-treated rats. These results indicate that CRF and AVP are distributed in similar hypothalamic nuclei, but that they are not identical.

Chromosome studies were conducted on two patients with adult T-cell leukemia. In both patients, a marker chromosome 14q+ and a structural change involving chromosome 1 with trisomy of the q arm were found in peripheral blood leukocytes. The 14q+ marker chromosome had resulted from translocation from #5p in one patient and #5q in the other patient. The present and previous studies suggest that the donor chromosomes involved in the 14q+ translocation are variable. This indicates that the 14q+ marker chromosome rather than the donor chromosome is intimately related with adult T-cell leukemia.

The distribution of corticotropin releasing factor (CRF) and arginine vasopressin (AVP) in hypothalamic nuclei were examined in control and estrogen-treated female rats. CRF activity was measured using monolayer cultured rat anterior pituitary cells and AVP by radioimmunoassay. Hypothalamic nuclei were punched out according to the method of Palkovits. The distribution of CRF activity in 5 different hypothalamic nuclei was similar to that of AVP in intact female rats. CRF activity in hypothalamic nuclei, pituitary ACTH content and plasma ACTH levels in estrogen-treated rats were not significantly different from those in control rats. However, significant elevation of AVP content was observed in the supraoptic and paraventricular nuclei of estrogen-treated rats. These results indicate that CRF and AVP are distributed in similar hypothalamic nuclei, but that they are not identical.