In vivo release of neuronal histamine in the hypothalamus of rats measured by microdialysis

Summary Using an in vivo intracerebral microdialysis method coupled with an HPLC-fluorometric method, we investigated the extracellular level of endogenous histamine in the anterior hypothalamic area of urethaneanaesthetized rats. The basal rate of release of endogenous histamine in the anterior hypothalamic area measured by this method was 0.09 + 0.01 pmol/20 min. When the anterior hypothalamic area was depolarized by infusion of 100 mM K⁺ through the dialysis membrane or electrical stimulation at 200 A was applied through an electrode implanted into the ipsilateral tuberomammillary nucleus, histamine release increased to 175% and 188%, respectively, of the basal level. These increases were completely suppressed by removal of extracellular Ca²⁺. The basal release of histamine was also suppressed after infusion of 10^–6 M tetrodotoxin or i.p. administration of 100 mg/kg of -fluoromethylhistidine. On the other hand, 3-fold increase in the basal release was observed after i. p. administration of 5 mg/kg thioperamide. These results clearly indicate that both the basal and evoked release of histamine measured by our method are of neuronal origin. Send offprint requests to T. Mochizuki at the above address     

Cytokine production in surgical stress

Surgical injury influences the function of mononuclear cells, leading to various systemic responses. Proinflammatory cytokines such as tumor necrosis factor alpha (TNF alpha), interleukin (IL)-1, -6, -8, and the antiinflammatory cytokine IL-10, which are mainly produced by mononuclear cells, are known to play an important role in the response to and pathogenesis of surgical stress. TNF alpha production by monocytes is extremely upregulated, but monocyte HLA-DR antigen expression is suppressed in patients with surgical stress. While production of Th1 cytokines such as IL-12 and interferon-gamma by mononuclear cells is suppressed, production of Th2 cytokines and IL-10 is upregulated during surgical stress. Immune suppression following surgical stress has been clarified recently in terms of Th1 and Th2 cytokine production balance mainly caused by mononuclear cells. It is thought to be very important to maintain immunological function after surgical stress by controlling cytokine production and balance.     

Development of a miniature undulation pump for the distributed artificial heart

Research of the distributed artificial heart is important not only to acquire the means of individual organ perfusion but also to clarify the characteristics of the organ and the mechanism of blood distribution. To investigate the distributed artificial heart, the miniature undulation pump was developed. The outer diameter and the thickness of the developed pump were 38 mm and 11 mm, respectively. The priming volume of the pump was 3.2 ml. The total size including the motor unit was 38 mm in diameter and 32 mm in length. The total weight was 67.5 g. The total volume was 27.5 ml. The pump was driven with pulse width modulation by using a 1 chip motor controller. More than 5 L/min of continuous output could be obtained. The results showed that the developed miniature undulation pump system had enough performance for individual organ perfusion.     

Selection of Human Ovarian Carcinoma Cells with High Dissemination Potential by Repeated Passage of the Cells in vivo into Nude Mice, and Involvement of Lex-determinant in the Dissemination Potential

Cells of the human tumor cell line RMG-1, derived from a clear-cell adenocarcinoma of the ovary, were injected intraperitoneally into nude mice, and the cells obtained from the tumor nodules in the mesenterium were found to form a larger number of, and larger-sized, tumor nodules than the original RMG-1 cells. The RMG-1-h cells, transferred into culture from the tumor nodules after a 4th in vivo passage, showed a dissemination potential as high as that of cells disseminating directly from the tissues, and exceedingly higher than that of RMG-1 cells. To assess the molecular bases of the different biological properties of RMG-1 and RMG-1-h cells, we compared the content and expression of various carbohydrate antigens in both cells. The chromosomal profile of RMG-1-h cells revealed their human origin and was identical to that of the original RMG-1 cells. In contrast to the broad histogram for the Le^x-bearing cells among RMG-1 cells in flow cytometry, the weakly and moderately positive cells toward anti-Le^x antibody were found to be eliminated from the histogram for the RMG-1-h cells, resulting in the enrichment of cells strongly expressing Le^x, which may account for the high dissemination potential. In addition, the adhesion of RMG-1 cells to mesothelial cells was found to be significantly inhibited by pretreatment of the cells with anti-Le^x antibody, indicating Le^x-mediated cell-to-cell interaction between ovarian cancer cells and mesothelial cells. By TLC-immunostaining, two Le^x-glycolipids, III³Fucα-nLc₄Cer and V³Fucα-nLc₆Cer were detected in both RMG-1 and RMG-1-h cells, and their total concentrations were not significantly different from each other. However, the hydrophobic moieties of Le^x-glycolipids in RMG-1-h cells were different from those in RMG-1 cells, suggesting that a difference in the structure of the hydrophobic moieties of Le^x is partly involved in the enhanced reactivity of RMG-1-h cells toward anti-Le^x antibody. Thus, the high dissemination potential of ovarian cancer cells was shown to be mediated by the Le^x-determinant and the Le^x-bearing cells are enriched by repeated in vivo passage of the cells into nude mice.     

Diffuse Lewy body disease searched out from 114 patients with parkinsonism]

From 114 patients who had been previously diagnosed as Parkinson's disease, we diagnosed six cases as clinically definite "diffuse Lewy body disease (DLBD)" according to McKeith's criteria with more strict modifications. Besides a central feature, dementia, and core features including parkinsonism, fluctuating cognition, and recurrent visual hallucinations, the patients presented some of supportive features, that is, repeated falls (4 cases), syncope (5 cases), and transient loss of consciousness (all cases). Autopsy, which was performed in 2 of the cases, revealed Lewy bodies in various nervous tissues including autonomic nervous systems in both cases. 7 cases of probable DLBD and 8 cases of possible DLBD, which lacked fluctuating cognition and/or visual hallucinations, demonstrated neither of repeated falls, syncope, nor transient loss of consciousness. Episodes of these supportive features, which seem to be associated with autonomic dysfunctions and/or fluctuating cognition, should be important in the differential diagnosis of DLBD.     

Molecular diversity of TEX101, a marker glycoprotein for germ cells monitored with monoclonal antibodies: variety of the molecular characteristics according to subcellular localization within the mouse testis

TEX101 was characterized as a unique germ cell marker molecule using the specific monoclonal antibody (mAb), TES101. Although this mAb has strong affinity/specificity for TEX101, TES101 mAb loses its reactivity under reducing conditions. In this study, we have generated new mAbs against TEX101 to compensate for the shortcomings of the TES101 mAb using different approaches. First, we immunized mice with the antigen on a baculovirus expression system and isolated new anti-TEX101 mAbs, 6002 and 6035. Second, we raised the mAb Ts4 from spleen cells of an immunologically naive old mouse. Western blot analysis revealed that the new mAbs possess immunoreactivity under reducing/non-reducing conditions. Immunopositive staining of the mAbs against Bouin-fixed sections was observed in spermatocytes, spermatids and testicular spermatozoa, but not in other cells, similar to paraformaldehyde (PFA)-fixed frozen sections stained with TES101 as previously reported. However, whereas the mAbs 6002/6035 mainly showed immunoreactivity only in spermatocytes in PFA-fixed frozen sections, the reactivity of the mAbs to spermatids and testicular spermatozoa was clearly recovered when the PFA-fixed sections were autoclaved or treated with SDS. Peptide mapping and deglycosylation analysis indicated that the epitopes for TES101, 6002 and 6035 are located within TEX101(25-94), whereas Ts4 recognized N-linked carbohydrate moieties on TEX101 in Triton X-100-soluble mouse testicular extracts but not in the extracellular or water-soluble fractions. These results suggest strongly that the molecular association or structure of N-linked carbohydrate moieties of TEX101 varies according to its subcellular localization within the seminiferous tubules. These new mAbs will be valuable tools for further analysis of TEX101, including its function(s).     

Correlation between a Bedridden Status and the Long-term Outcome in Hemodialysis Patients after Intracerebral Hemorrhaging

Objective The quality of life and activities of daily living (ADL) are generally poor among dialysis patients after intracerebral hemorrhaging, and their precise clinical course remains unclear. In addition, the association between the severity of cerebral hemorrhaging and the long-term prognosis in these patients has not been fully elucidated. This study aimed to evaluate the subsequent prognosis of hemodialysis patients who survived the acute phase of intracerebral hemorrhaging. Methods We included hemodialysis patients who were admitted to Nagasaki University Hospital between 2007 and 2015 for intracerebral hemorrhaging treatment. After excluding cases of in-hospital death, survivors were classified using the 5-point modified Rankin Scale (mRS), which specifically measures the ADL in patients with cerebrovascular diseases. The patients were followed up at the medical facilities to which they were transferred in the same medical zone until 2017. Results Out of 91 patients with cerebral hemorrhaging (65±11 years old, 66% men, hemodialysis duration 108±91 months), 62 survived until discharge. Twenty-one patients died during observation, largely due to infectious diseases, such as sepsis and pneumonia (n=16, 76%). Compared to patients with mRS 0-4 (n=31), those with mRS 5 (n=31) showed a significantly poorer prognosis. The hazard ratio adjusted for age and antiplatelets was 13.7 (95% confidence interval: 3.88-63.7, p<0.001). Conclusion Hemodialysis patients with intracerebral hemorrhaging who were bedridden showed poor outcomes. The major causes of death were infections. Therefore, these patients should be carefully monitored for infections in order to improve their prognosis.