槲皮素、别嘌醇对高尿酸血症大鼠血清尿酸水平及肝、肾功能影响的对比研究

对比研究槲皮素、别嘌醇对高尿酸血症大鼠的治疗作用并观察对肝、肾功能的影响。雄性SD大鼠,连续灌胃给药7 d,第5天采用次黄嘌呤法制备大鼠高尿酸血症模型。采用比色法、连续监测法、化学氧化法、酶联免疫吸附法等测定大鼠血清中尿酸(UA)、丙氨酸氨基转移酶(ALT)、天门冬氨酸氨基转移酶(AST)、总胆红素(TBIL)、直接胆红素(DBIL)、β₂-微球蛋白(β₂-MG)、胱抑素C(Cys-C)、尿素及肌酐(Cr)含量。结果显示:别嘌醇能够显著降低大鼠血清尿酸水平(P<0.01),而槲皮素对血清尿酸无影响;槲皮素和别嘌醇显著降低大鼠ALT和AST水平(P<0.01),对TBIL和DBIL水平无明显影响,显著提高β₂-MG,Cys-C水平(P<0.01),别嘌醇治疗组大鼠血清尿素和Cr水平明显高于正常对照组(P<0.01);造模组和给药组可见大鼠肾脏轻度病理组织学改变。结果表明:槲皮素对大鼠血清尿酸水平无明显影响,而别嘌醇降尿酸作用显著。造模和给药对肝功能均无明显影响,但造模可能导致肾功能不同程度的损害,槲皮素对轻度肾损伤未见明显的保护作用,别嘌醇给药后加重肾功能损伤。     

Enhanced self-renewal capability in hepatic stem/progenitor cells drives cancer initiation

BACKGROUND & AIMS: Transformed hematopoietic stem/progenitor cells with an enhanced or acquired self-renewal capability function as leukemic stem cells. In a variety of solid cancers, stem/progenitor cells could be also targets of carcinogenesis. However, it remains unclear whether disruption of stem cell function directly contributes to cancer initiation. We sought to elucidate the mechanisms of self-renewal in hepatic stem/progenitor cells and the relation between stem cell function and hepatocarcinogenesis. METHODS: Functional analyses of polycomb-group protein Bmi1 and Wnt/beta-catenin, the molecules that are responsible for the self-renewal capability of many types of stem cells, were conducted in c-Kit(-)CD29(+)CD49f(+/low)CD45(-)Ter-119(-) hepatic stem/progenitor cells using retrovirus- or lentivirus-mediated gene transfer. The tumorigenicity of these cells transduced with the indicated retroviruses was also assessed by transplantation into nonobese diabetic/severe combined immunodeficient mice. RESULTS: Forced expression of Bmi1 and constitutively active beta-catenin mutant similarly promoted the self-renewal of hepatic stem/progenitor cells. The transplantation of Bmi1- or beta-catenin-transduced cells clonally expanded from single hepatic stem/progenitor cells produced tumors, which exhibited the histologic features of combined hepatocellular and cholangiocarcinoma. CONCLUSIONS: These observations imply that the dysregulated self-renewal of hepatic stem/progenitor cells serves as an early event in hepatocarcinogenesis, and they highlight the important roles of Bmi1 and the Wnt/beta-catenin pathway in regulating the self-renewal of normal or cancer stem cells in liver.     

IL-33 receptor ST2 deficiency attenuates renal ischaemia–reperfusion injury in euglycaemic,but not streptozotocin-induced hyperglycaemic mice

Aim

Kidney hypoxia can predispose to the development of acute and chronic renal failure in diabetes. Ischaemia–reperfusion injury (IRI) causes inflammation, and diabetes is known to exacerbate this inflammatory response in the kidney, whereas alarmin IL-33 could act as an innate immune mediator during kidney IRI. Thus, the present study examined the impact of genetic IL-33 receptor ST2 deficiency (ST2?/?) on renal IRI in euglycaemic and hyperglycaemic mice.

Molecular analysis of HPRT1+ somatic cell hybrids derived from a carrier of an HPRT1 mutation responsible for Lesch‐Nyhan syndrome

Heterozygous carriers of HPRT1 mutations responsible for Lesch‐Nyhan syndrome can be detected by analysis of somatic cell hybrids derived from peripheral blood lymphocytes and Hprt1‐negative cells of rodent origin followed by selection in culture medium containing hypoxanthine, aminopterine, and thymidine (HAT). The parental origin of the X chromosome containing the normal HPRT1 allele in HPRT1⁺ hybrid cell lines can be determined by molecular haplotyping attributable to highly polymorphic X‐linked markers. We used this procedure to study a presumed carrier whose paternal active X chromosome always segregated in the cell hybrids derived from her. Conversely, her maternal X chromosome was systematically absent in most cell hybrids, or when present, it was inactive and coexisted with an active, paternal X chromosome. These results clearly demonstrated that the proband was a heterozygous carrier of a mutation responsible for HPRT1 deficiency. © 2001 Wiley‐Liss, Inc.     

Production and Characterization of Monoclonal Antibodies against Bovine Luteinizing Hormone

Five mouse hybridoma cell lines producing monoclonal antibody against bovine luteinizing hormone (LH) have been established and the respective antibodies characterized by radioimmunoassay, immunofluorescence and immunoelectrophoresis. All antibodies belong to the IgG class and bind to staphylococcus protein A. Intraspecies cross-reactivity studies revealed no reaction with bovine follicle stimulating hormone (FSH). However, all antibodies showed partial cross-reaction with bovine thyroid stimulating hormone (TSH) suggesting a close conformational similarity between bovine LH and TSH. Studies on interspecies cross-reactivity (rat and human) showed that three of these five antibodies strongly react with rat LH but not at all with either rat FSH or rat TSH thus representing monospecific reagents for investigations concerning LH in this species. One of these three antibodies also strongly binds to human LH and to the same extent to human chorionic gonadotropin (CG) but not to human FSH or TSH. It was concluded that at least three different epitopes on the bovine LH molecule are recognized and that they are located on the β-chain of the hormone.     

Molecular analysis of a Japanese family with Lesch-Nyhan syndrome: identification of mutation and prenatal diagnosis

Complete deficiency of hypoxanthine guanine phosphoribosyltransferase (HPRT) causes Lesch-Nyhan syndrome. We examined the HPRT gene mutation for prenatal diagnosis in a Japanese family. A single nucleotide substitution of C to T in exon 3 was identified by direct sequencing analysis of the HPRT gene of a Lesch-Nyhan patient. This substitution resulted in a nonsense mutation, CGA (Arg) to TGA (stop), at codon 51. Utilizing an Xho I restriction site which was lost in the mutation as an indicator, a family study showed that the mother was heterozygous, but the grandmother normal. By the same method, prenatal genetic diagnosis was performed using chorionic villus samples (CVS), and showed that the fetus had the mutant allele.     

Antibody producing human-human hybridomas. I. Technical aspects

Technical aspects of generation of antibody-secreting human-human hybridomas are evaluated as based on 100 human-human fusions with a human B-lymphoma cell line (RH-L4) or the SKO-007 myeloma cell line as malignant fusion partners, and compared with similar fusion conditions in the mouse hybridoma system. The yield of hybrids was significantly lower when normal peripheral blood lymphocytes were used as fusion partners as compared with spleen lymphocytes, but could be substantially improved by increasing the amount of mitotic active B-lymphocytes by mitogen stimulation of the lymphocytes, preferably in HAT medium, prior to fusion. Furthermore, human hybrids grew slower and had a higher degree of chromosomal instability than usually observed in the mouse hybridoma system. Thus, out of 72 fusions, only 3 stable hybrids with antibody production against a predefined antigen were established. The importance of improved sources of human B-lymphocytes for human-human hybridoma production is discussed and methods of obtaining such improvement suggested.     

Is skeletal muscle damaged by the oxidative stress following anaerobic exercise?

We investigated whether the injury of skeletal muscle owing to the action of free radicals and the subsequent oxidative damage to tissues occurred during anaerobic exercise. To estimate injury to skeletal muscle, we determined certain indices of oxidative damage to skeletal muscle; i.e., leukocyte counts, concentrations of hypoxanthine, xanthine, urate, tissue- and serum-type CK-M isoforms, myoglobin, and total antioxidant capacity (TAC) of serum. Blood for these tests was collected at 3 min post-exercise. Post-anaerobic exercise concentrations of lactate were significantly increased from pre-exercise. The neutrophil and lymphocyte counts and alanine concentration were significantly increased by anaerobic exercise, even when the results were corrected for plasma volume changes; the plasma concentrations of hypoxanthine, urate, and TAC of serum were also significantly increased. The plasma concentration of xanthine was negatively correlated with TAC of serum. The activities of tissue- and serum-type CK-M were significantly increased post-exercise. When the hypoxanthine, urate, TAC of serum, myoglobin, and tissue- and serum-type CK-M were corrected for plasma volume changes, the post-exercise increases were no longer significantly different from the pre-exercise results. We suggest that these latter test results following anaerobic exercise exclude the presence of oxidative damage to skeletal muscle.     

ATP breakdown products in human skeletal muscle during prolonged exercise to exhaustion

Summary. To study changes in muscle energy state during prolonged exercise, especially in relation to fatigue, muscle biopsies were obtained from seven healthy males working until exhaustion on a cycle ergometer at 68% (63–74%) of their maximal oxygen uptake. Biopsies were taken at rest, after 15 and 45 min of exercise and at exhaustion, and analysed for ATP, ADP, AMP, inosine monophosphate (IMP) and hypoxanthine content by high performance liquid chromatography (HPLC), and for creatine phosphate (CP), lactate and glycogen by enzymatic fluorometric techniques. Glycogen content at exhaustion was approximately 30% of the pre-exercise level. The CP content decreased steeply during the first 15 min of exercise (P<0·01) and continued to decrease during the rest of the exercise period (P<0·05). Pronounced increases in contents of IMP (64%P<0·001) and hypoxanthine (69%, P<0·05) were found when exhaustion was approaching. Furthermore, energy charge [EC; (ATP+0·5 ADP)/(ATP+ADP+AMP)] was decreased at exhaustion (P<0·05). The increases in IMP and hypoxanthine which occurred when exhaustion was approaching during prolonged submaximal exercise together with the decrease in EC during this phase of exercise suggest a failure of the exercising skeletal muscle to regenerate ATP at exhaustion.