Research review: DNA polymerases as molecular markers of the regenerating capacity of hepatocytes

Hepatocyte regeneration has been widely investigated, with the mitotic index and the incorporation of [³H]thymidine being used as regeneration markers. We focused on the induction of DNA replication enzymes, particularly DNA polymerases (pol) α, δ, and ε. Using rat models, we have shown that the activity of pol α in crude liver extract well represents the regenerating capacity of hepatocytes. Using pol α as an indicator, we analyzed liver regeneration in rat models under various conditions: obstructive jaundice, external or internal biliary drainage, and the obstruction of portal vein branches. It has been revealed that the ligation of the common bile duct alone induces a certain amount of hepatocyte proliferation. It was striking that external biliary drainage suppressed regeneration capacity in cholestatic rat liver after partial hepatectomy. The strong regeneration in nonligated lobes induced by portal branch ligation was similar to the liver regeneration seen after partial hepatectomy with respect to the induction of DNA polymerases. Taken together, the aspects of DNA replication, particularly the induction of DNA polymerases, may contribute to shedding new light on the regeneration of human liver. This work was supported in part by a Grant-in-Aid for General Scientific Research and for Cancer Research from the Ministry of Education, Science and Culture, Japan, and by grants from the Uehara Memorial Foundation     

人酪氨酸羟化酶基因真核表达载体的构建及在哺乳动物细胞中的表达

用限制性内切酶EcoRI从pKS(-)HTH_1切下全长为1.9 kb的人酪氨酸羟化酶基因,在T_4DNA连接酶的作用下连接在真核表达载体pCDNA_3的EcoR Ⅰ位点,构建成重组质粒pcD-NA_3HTH_1,该质粒转染COS-7细胞,免疫荧光组织化学染色证实酪氨酸羟化酶在其中的表达。     

Cisplatin-associated anaemia in patients with solid tumours

We have reviewed the incidence of cisplatin-induced anaemia in patients affected with solid tumours treated with at least three courses of first-line cisplatincontaining regimens. In our experience, a low percentage (5%) of patients required transfusions of red blood cells. We think it is of the utmost importance to adopt uniform criteria in monitoring and treatment of patients at risk of developing cisplatin anaemia and to identify subsets of patients to eventually treat with erythropoietin.     

Human Y-79 retinoblastoma cells express functional corticotropin-releasing hormone receptors

In human Y-79 retinoblastoma cells corticotropin-releasing hormone (CRH) produces a marked and rapid increase of adenylate cyclase activity. The concentration of the peptide producing half-maximal stimulation is 60 nM. The effect of CRH is significantly antagonized by the specific CRH receptor antagonist alpha-helical CHR 9-41 and is mimicked by sauvagine and urotensin I, two peptides displaying sequence homology with CRH. These results demonstrate the presence of functional CRH receptors in human Y-79 retinoblastoma cells and suggest that this cell line may be a suitable model in which to study the action of CRH on human retinal cell function.     

No evidence for an altered mRNA expression or protein level of haematopoietic cell phosphatase in CD34+ bone marrow progenitor cells or mature peripheral blood cells in polycythaemia vera

Abstract: Polycythaemia vera (PV) is a myeloproliferative disorder characterized by haematopoietic progenitor cells being hypersensitive to cytokines such as erythropoietin, interleukin-3, stem cell factor and insulin-like growth factor 1, which results in an increased production of mature blood cells. The pathogenetic cellular mechanism(s) behind this hypersensitivity to cytokines is unknown, but the number of cytokine receptors and the interaction between ligand and receptor are normal in PV. Interest has therefore focused on post-receptor mechanism(s). Haematopoietic cell phosphatase (HCP) is an intracellular tyrosine phosphatase that has been demonstrated to regulate proliferative signals negatively induced by the cytokines mentioned above. Moreover, motheaten mice that genetically lack HCP have an increased amount of erythroid progenitors that are hypersensitive to Epo, and patients with familial polycythaemia have been shown to exhibit a mutation of the Epo receptor gene that includes the docking site for HCP. We therefore studied mRNA expression of HCP in pure populations of CD34⁺ cells, granulocytes, platelets and lymphocytes from patients with PV, chronic myeloid leukaemia (CML) or essential thrombocythemia (ET), as well as healthy controls. Using a polymerase chain reaction analysis employing specific primers for HCP, we failed to detect any abnormalities of HCP expression in PV in any of the cell populations that were examined. Moreover, HCP mRNA expression was similar in ET and CML compared to controls. Finally, Western blot analysis revealed a normal HCP protein content in PV granulocytes and platelets. We therefore conclude that neither an impaired expression of the HCP gene nor a defect in HCP protein synthesis is present in PV, and does not seem to play a role in the aetiology of this disorder.     

Anti-Oxidative Therapy with Oral Dapsone Improved HCV Antibody Positive Annular Elastolytic Giant Cell Granuloma

A 72-year-old fisherman who was positive for the HCV antibody developed an annular, erythematous, infiltrated lesions on sun-exposed areas. The lesions were diagnosed as annular elastolytic giant cell granuloma both clinically and histologically. Topical corticosteroid and cryotherapy with liquid nitrogen for several months failed to improve the lesions. We then started dapsone, a known anti-oxidant, at 50 mg/day. A month later, the margins of the erythematous lesions faded, and the infiltration gradually decreased. No recurrence has been observed for one year after the start of the therapy. Anti-oxidative therapy appears to be effective for annular elastolytic giant cell granuloma and could be an alternate therapy for refractory granulomatous disease.     

Surface behavior of axolemma monolayers: Physico-chemical characterization and use as supported planar membranes for cultured Schwann cells

The axolemma membrane forms a stable and reproducible monomolecular layer at the air-aqueous interface. The major lipids and proteins are present in this monolayer in molar ratios similar to the original membrane. Acetylcholinesterase and Na-K-ATPase activities are preserved in the monolayer to levels of 64% and 25%, respectively. The total lipid fraction forms a homogeneously mixed phase. The presence of proteins in the monolayer introduces surface inhomogeneties. Among other features, this is revealed by the presence of two values of lateral pressure at which the monolayer shows partial or total collapse: a broad partial collapse at surface pressures between 13 to 30 mN/m and a sharp collapse point at 46 mN/m. The average molecular areas, the broad collapse point, and the variation of the surface potential per molecule suggest the relocation of protein components at surface pressures between 13 to 30 mN/m. The behavior is consistent with the extrusion and exposure of proteins toward the aqueous medium that depends on the lateral pressure. Schwann cells grown on coverslips coated with axolemma monolayers at 13 mN/m (beginning of the broad collapse) and 34 mN/m (above the broad collapse) recognize the difference in the surface organization of axolemma caused by the lateral pressure which affects their proliferation, morphology, and spatial pattern of organization. Our results show for the first time that response of Schwann cells depends on the intermolecular organization of the axolemma surface with which they interact. These results suggest that the local expression of putative surface molecules of axolemma that may mediate membrane recognition and the signalling of morphological and proliferative changes can be modulated by long range supramolecular properties. © 1993 Wiley-Liss, Inc.     

Gastro-duodenal digestion products of the major peanut allergen Ara h 1 retain an allergenic potential

BACKGROUND: The process of gastro-duodenal digestion may play a role in determining the allergenic properties of food proteins. The sensitizing and allergenic potential of digestion products of highly degraded allergens, such as the major peanut allergen Ara h 1, is currently under debate. We evaluated the effect of in vitro gastro-duodenal digestion of Ara h 1 on T cell reactivity and basophil histamine release. METHODS: An in vitro model of gastro-duodenal digestion was used to investigate changes in the allergenic properties of Ara h 1 using in vitro assays monitoring T cell reactivity (proliferation, cytokine production) and histamine release of basophils from peanut allergic individuals. The digestion process was monitored using an SDS-PAGE gel. RESULTS: In vitro gastric digestion led to rapid degradation of Ara h 1 into small fragments M(r) L5600. Gastric digestion did not affect the ability of Ara h 1 to stimulate cellular proliferation. Gastro-duodenal digestion significantly reduced its ability to stimulate clonal expansion (P<0,05; Wilxocon's signed rank test). The Th-2 type cytokine polarization of T cells from peanut allergic donors (IFN-gamma/IL-13 ratio and IFN-gamma/IL-4 ratio of CFSE(low) CD4(+) T cells) remained unchanged regardless of the level of digestion. Histamine release of basophils from peanut allergic individuals was induced to the same extent by native Ara h 1 and its digestion products. CONCLUSION: Gastro-duodenal digestion fragments of Ara h 1 retain T cell stimulatory and IgE-binding and cross-linking properties of the intact protein.