Scanning tunneling microscopy study of cytochrome P450 2B4 incorporated in proteoliposomes

In the present paper, the application of scanning tunneling microscopy in cytochrome P450s membrane topology is discussed. The method enables visualization of heme location in the lipid-bilayer-incorporated protein. It is supposed that the membrane-bound cytochrome P450 on the tunneling microscope substrate should behave as 'molecular diode'. A model explaining the liposome and the proteoliposome images observed is proposed.     

Bone marrow failure in the Fanconi anemia group C mouse model after DNA damage

Fanconi anemia (FA) is a pleiotropic inherited disease that causes bone marrow failure in children. However, the specific involvement of FA genes in hematopoiesis and their relation to bone marrow (BM) failure is still unclear. The increased sensitivity of FA cells to DNA cross-linking agents such as mitomycin C (MMC) and diepoxybutane (DEB), including the induction of chromosomal aberrations and delay in the G2 phase of the cell cycle, have suggested a role for the FA genes in DNA repair, cell cycle regulation, and apoptosis. We previously reported the cloning of the FA group C gene (FAC) and the generation of a Fac mouse model. Surprisingly, the Fac -/- mice did not show any of the hematologic defects found in FA patients. To better understand the relationship of FA gene functions to BM failure, we have analyzed the in vivo effect of an FA-specific DNA damaging agent in Fac -/- mice. The mice were found to be highly sensitive to DNA cross-linking agents; acute exposure to MMC produced a marked BM hypoplasia and degeneration of proliferative tissues and caused death within a few days of treatment. However, sequential, nonlethal doses of MMC caused a progressive decrease in all peripheral blood parameters of Fac -/- mice. This treatment targeted specifically the BM compartment, with no effect on other proliferative tissues. The progressive pancytopenia resulted from a reduction in the number of early and committed hematopoietic progenitors. These results indicate that the FA genes are involved in the physiologic response of hematopoietic progenitor cells to DNA damage.     

Molecular-biologic aspects of interaction between nervous and immune systems

The problem of the neuro-immuno interactions on the level of the protein trans-factors, stimulating interleukin-2 (IL-2) gene expression was discussed. The physico-chemical and functional parameters of the low molecular nuclear proteins (SP and BP- 14, 18, 19 kDs) isolated from splenic and brain cells of immunized rats were studied. The binding of these proteins to the regulatory region of IL-2 gene in vitro and stimulation of the IL-2mRNA synthesis in splenic T-lymphocytes culture in normal conditions were shown. The protective effect of SP and BP on the IL-2mRNA synthesis in stressful conditions and by the T-cells treatment with the CsA was demonstrated.     

Differential expression of Fos protein after transection of the rat infraorbital nerve in the trigeminal nucleus caudalis

To determine the effects of nerve injury on Fos expression, temporal and spatial distributions of Fos-positive neurons in the trigeminal nucleus caudalis were examined after tissue injury for isolation of the infraorbital nerve as controls and transection of this nerve as well as noxious chemical stimulation by formalin injection in adult rats. Fos immunoreactivity was markedly elevated in laminae I and II of the only ipsilateral nucleus caudalis 2 h after these surgical procedures and noxious chemical stimulation. The distributions of Fos-positive neurons were restricted rostro-caudally following formalin injection and tissue injury compared to transection of the infraorbital nerve. One day after tissue injury and nerve transection, however, Fos-positive neurons were distributed bilaterally in laminae III and IV extending rostro-caudally and medio-laterally in this nucleus, and this persisted over the 2-week study period. The number of Fos-positive neurons in the side ipsilateral to nerve transection was markedly less than that in the contralateral side whereas positive neurons in the tissue injured rats were distributed symmetrically along the rostro-caudal axis. There was no difference in the contralateral sides between nerve transection and tissue injury groups. The rostro-caudal level showing reduction in Fos expression corresponded roughly to the sites of central termination of the injured nerve in this nucleus, suggesting a role for the primary afferents in the reduction of Fos expression in laminae III and IV neurons of the ipsilateral nucleus caudalis.     

Alpha-galactosidase of the marine bacterium Pseudoalteromonas sp. KMM 701

An alpha-galactosidase that inactivates the group specificity of B erythrocytes (group III) of human blood and does not affect A erythrocytes (group II) was isolated from the marine bacterium Pseudoalteromonas sp. KMM 701. The enzyme preparation did not contain lectin, hemolytic, sialidase, endoglycanase, or glycosidase activities. The enzyme is stable at 20 degreesC for 24 h, has pH optimum for catalysis within the range of 6.7-7.7, and is stable to high concentrations of NaCl. It is 4-fold more efficient than the alpha-galactosidase from green coffee beans. At pH 7.0 the Km for p-nitrophenyl-alpha-D-galactopyranoside is 0.29 mM. The molecular weight of the enzyme determined by gel-filtration is 195 +/- 5 kD. The alpha-galactosidase is denatured by urea and guanidine hydrochloride. Its activity does not depend on the presence of metal ions. It contains a sulfhydryl group essential for its catalytic activity.     

The ecological cause of the higher accumulation of 90Sr and 137Cs in the food of northern inhabitants

PURPOSE: To quantify S-antigen-specific (S-Ag) T cells in the retina after adoptive transfer, and to evaluate their role in the initiation and progress of destructive ocular inflammation in experimental autoimmune uveoretinitis (EAU). METHODS: Lewis rats were administered 10 x 10(6) S-Ag-specific T cells from the SP35 cell line or 10 x 10(6) concanavalin A-stimulated syngeneic spleen cell lymphoblasts labeled with lipophilic PKH26 fluorescent dye immediately before intravenous inoculation. Labeled cells in each retina were counted at various times from 4 to 120 hours after cell transfer by fluorescence microscopic analysis of each dissociated retina. Recipient eyes were examined within the same period by light and confocal microscope. RESULTS: SP35 T cells showed a biphasic distribution in the retina. The first peak of 160 cells/retina was noted at 24 hours. A steady decline of labeled cells at 48 and 72 hours was followed by a rapid increase at 96 and 120 hours. Concanavalin A-stimulated, control-labeled cell populations showed an identical peak at 24 hours but a persistent decline thereafter; only two or three T cells were present in each retina at 120 hours. Concurrent inoculation of SP35 cells and nonspecific T cell blasts did not produce more SP35 cells than control cells in the retina at any time. Microscopic analysis showed mononuclear cell infiltration of the iris, ciliary body, and aqueous humor at 48 hours, which intensified rapidly and persisted through 120 hours. Retinal inflammation did not begin until 80 hours. Mononuclear cell adherence to vascular endothelium and perivascular macrophage infiltration of the innermost layers progressed to edema, and profound destructive inflammation and loss of retinal stratification were observed at 120 hours. CONCLUSIONS: There is no evidence of a blood-ocular or blood-retinal barrier to activated T cell blasts. Autologous S-Ag does not provoke a more rapid entry of specific T cells at that site. The data confirm that anterior segment inflammation precedes retinal inflammation, even though S-Ag-specific T cells were present in the retina within a few hours after cell transfer. Because S-Ag is clearly present in the retina, delay in antigen presentation at that site may account for the temporal difference between retinal and anterior segment inflammation.     

In vivo platelet and T-lymphocyte activities during pulmonary tuberculosis

Platelets have been suggested to play a role in the inflammatory response, including defence against bacteria. The aims of this study were to determine in vivo platelet activity during the clinical course of pulmonary tuberculosis and to investigate whether or not there is a correlation between the magnitude of platelet activation and the extent of the pulmonary disease. T-lymphocyte activity was also analysed in the patients. Platelet factor-4 (PF4) and soluble interleukin-2 receptor-alpha (sIL-2Ralpha) concentrations were used as markers of platelet and T-lymphocyte activation, respectively. Twenty-five patients with pulmonary tuberculosis were studied. Fifteen healthy subjects served as a control group. The levels of both sIL-2Ralpha (3,000+/-1,948 pg x mL(-1)) and PF4 (103.1+/-6.7 IU x mL(-1)) were significantly higher in the patients with tuberculosis than in the control group (984+/-360 pg x mL(-1) and 78.2+/-23.9 IU x mL(-1), respectively) (Mann-Whitney U-test, p<0.001 for both comparisons). The plasma PF4 levels were found to be well correlated with the extent of pulmonary lesions on chest radiography (the Spearman's bivariate correlation analysis, r=0.65, p<0.001). However, sIL-2Ralpha concentrations did not correlate with the extent of disease. In conclusion, it has been suggested that platelet and T-lymphocyte activation occurs during pulmonary tuberculosis. The good correlation between platelet activation and the extent of pulmonary tuberculosis might be ascribed to a pathophysiological role of platelets in pulmonary tuberculosis.