Role of Helicobacter pylori CagA+ infection in determining oxidative DNA damage in gastric mucosa

BACKGROUND: Although Helicobacter pylori is a risk factor for gastric cancer, the role of the bacterium in the development of this malignancy is not defined precisely. Reactive oxygen species (ROS) could play an important role in carcinogenesis by inducing DNA damage. The aims of the present study were: 1) to assess the production of ROS and 8-hydroxy-2'-deoxyguanosine (8-OHdG), a sensitive marker of oxidative DNA injury, in gastric mucosa, according to H. pylori status and cytotoxic associated gene product A (CagA); 2) to determine the relationship between ROS generation and amount of 8-OHdG. METHODS: Gastric biopsy specimens were obtained from 60 consecutive patients. ROS generation was measured by luminol enhanced chemiluminescence. 8-OHdG detection was performed by an immunoperoxidase method, using a specific anti 8-OHdG monoclonal antibody. RESULTS: 40/60 patients (67%) were H. pylori-positive. ROS generation was significantly higher in patients positive for H. pylori infection as compared to negative. 8-OHdG detection was performed in 30 patients in which CagA presence was also investigated. High expression of 8-OHdG was detected in 14/20 (70%) H. pylori-positive patients (13 CagA+ and 1 CagA-) and in 2/10 (20%) H. pylori-negative patients. A significant correlation was found between ROS production and 8-OHdG content. CONCLUSION: H. pylori infection by a CagA+ strain is associated with the highest production of ROS to which a severe oxidative DNA damage corresponds. This sequence of events could support the hypothesis that the oxygen-free radicals-mediated damage due to H. pylori cytotoxic strains could be a driving force that leads from chronic gastritis to gastric carcinoma.     

Treatment of hypoplastic anemia in mice with placental transplants

A genetic mutation in mice (W/Wv) causes an autosomal recessive disease characterized by hypoplastic anemia which lasts throughout life. Double- dominant W/Wv anemic mice were sublethally irradiated to facilitate repopulation of marrow with transplanted cells and were injected intravenously with suspensions of 5-10 million placental cells of 15 days gestation derived from normal, isogeneic donors. Red cell counts fell promptly after irradiation and then rose progressively over a period of weeks, reaching normal levels of the nonmutant. Mean corpuscular volume and hemoglobin electrophoresis patterns of red cells in recipient W/Wv mice resembled those of normal donor animals. The therapeutic effect lasted for the duration of the observation period, in some instances over 9 mo. W/Wv mice that were administered Hanks' solution or fetal blood, instead of placental transplants, remained anemic. Late gestation placentas (18 days) were also ineffective.     

Preferentially expressed genes in chronic myelogenous leukemia

The predominant circulating cells in chronic myelogenous leukemia (CML) morphologically resemble normal myeloid precursors; however, certain characteristics indicate the two are not identical. Approximately 88% of the patients with clinically typical CML present with a cytogenetic abnormality known as the Philadelphia chromosome (Ph1). Additionally, the leukocyte alkaline phosphatase (LAP) value is decreased in CML. To investigate if there are selected genes expressed in the CML cell population, poly(A+)RNA from a chronic-phase, Ph1-positive CML patient was used for construction of a complementary DNA (cDNA) library. Recombinant clones representing moderately to abundantly transcribed sequences were selected by annealing [32P]-cDNA transcribed from homologous RNA to the library sequences and assessing radioactivity in the hybrids. From an initial 729 colonies, 417 (57.2%) displayed a hybridization signal more intense than controls, indicating these recombinant plasmids contained sequences homologous to moderately or highly expressed RNAs from this particular patient. Screening of the 417 clones--utilizing 32P-cDNAs derived from normal human placenta, an acute myelomonocytic leukemia (AMML), and two other CML samples--was used to select clones likely to represent sequences preferentially expressed in CML. Sixteen recombinants were initially selected that repeatedly failed to display hybridization with the placenta and AMML- derived probes. Further analysis of eight of these clones indicated that six contain sequences preferentially expressed in CML. One clone, C-A3, has been studied with 63 different RNA samples. This sequence is found to be highly expressed in peripheral blood cells from the chronic phase of both Ph1-positive and Ph1-negative CML as well as in a Ph1- positive acute myelogenous leukemia (AML). Expression is reduced in lymphoblastic crisis of CML (L BC-CML) and essentially absent in myeloblastic crisis of CML (M BC-CML). While preliminary, the results suggest that this probe may be useful as an aid in diagnosing Ph1- negative CML and in distinguishing M BC-CML from L BC-CML and Ph1- positive AML.     

Glucocorticoids and lymphocytes. III. Effects of glucocorticoid administration on lymphocyte glucocorticoid receptors

To determine the effects of glucocorticoid administration on the number of measured lymphocyte glucocorticoid receptor sites and the duration of such effects, seven normal volunteers were studied. Glucocorticoid receptor levels of the lymphocytes circulating in the blood of each volunteer were determined. Glucocorticoid was then administered in a regimen of a total of four doses of dexamethasone 4 mg p.o. every 6 hr. Determinations of the number of receptors were performed at 6 hr and at various subsequent times after the end of dexamethasone administration. When compared to baseline receptor numbers, six volunteers showed a decrease in receptor number after glucocorticoid administration (median maximum decrease 2,046 sites/cell). The fall in receptor number occurred rapidly, reaching a nadir within 30 hr from the end of glucocorticoid administration. The return of receptor number to baseline was more gradual, requiring from 3 to as long as 17 days in one subject. Our results suggest that in order to accurately interpret glucocorticoid receptor numbers in human lymphoid cells, glucocorticoid should not have been administered for 3 wk prior to determinations of receptor levels.     

胰岛素在严重烧伤大鼠急性肺损伤中的保护作用

目的:观察胰岛素对严重烧伤所致的急性肺损伤的保护效应。方法:实验于2006-01/08在解放军第四军医大学西京医院全军烧伤中心实验室完成。取成年雄性SD大鼠36只,随机分成3组,每组12只:①烫伤 胰岛素组:水浴锅94℃,20s,制备30%总体表面积全层皮肤烫伤模型,伤后即刻腹腔注射生理盐水40mL/kg,并皮下注射胰岛素3U/kg。②烫伤组:造模和腹腔注射同烫伤 胰岛素组,皮下注射等体积生理盐水。③假烫组:用室温水浴模拟烫伤过程,伤后不给予补液。烫伤24h后,收集动脉血测定超氧化物歧化酶活性,收集支气管肺泡灌洗液测定蛋白含量,取肺组织进行苏木精-伊红染色观察病理变化,并测定髓过氧化物酶活性,同时电镜观察胸主动脉血管内皮细胞变化情况。结果:36只大鼠进入结果分析。①肺病理变化:烫伤 胰岛素组肺脏渗出和水肿较烫伤组明显减轻。②肺泡灌洗液中蛋白的质量浓度:烫伤组和烫伤 胰岛素组高于假烫组[(702.9±169.5),(486.5±149.2),(240.5±140.7)mg/L,P<0.05],烫伤 胰岛素组低于烫伤组(P<0.05)。③肺脏髓过氧化物酶活性:烫伤 胰岛素组低于烫伤组[(36.01±8.17),(59.51±12.50)nkat/g,P<0.05],与假烫组比较差异不显著。④血清超氧化物歧化酶活性:烫伤组和烫伤 胰岛素组低于假烫组[(2.27±0.18),(2.63±0.19),(2.81±0.21)mkat/L,P<0.01,0.05],烫伤 胰岛素组高于烫伤组(P<0.01)。⑤电镜下可见烫伤 胰岛素组内皮细胞损伤较烫伤组轻。结论:严重烧伤早期外源性胰岛素干预后可减轻急性肺损伤,具有明显的肺脏保护作用,这种作用可能与胰岛素的内皮细胞保护效应有关。     

Rapid freezing of whole blood or buffy coat samples for polymerase chain reaction and cell culture analysis: application to detection of human immunodeficiency virus in blood donor and recipient repositories. The Transfusion Safety Study Group

Storage of lymphocytes for later use in prospective epidemiologic studies of blood donors and transfusion recipients has been limited by the cost of separating peripheral blood mononuclear cells (PBMCs). When the Transfusion Safety Study began in 1985, it was decided to establish a cell repository of cryopreserved buffy coat (BC) samples, and thus far over 20,000 samples have been accumulated from enrolled subjects. To determine if these specimens could be used for polymerase chain reaction, a simple thawing and pelleting technique for recovering hemoglobin-free total white cells (WBCs) was developed. To validate the technique, parallel analysis was conducted of BCs, whole blood (WB), and PBMC samples from human immunodeficiency virus type 1 (HIV-1)- seropositive subjects. Immediate postthaw cell courts of 29 frozen- thawed (F-T) WB and BC samples averaged 90 percent of the prefreeze (input) values. Representative WBC populations were obtained by immediate pelleting. Amplification of HIV-1 gag sequences from F-T BCs and F-T WB was 94 and 75 percent, respectively, which is as sensitive as that obtained with freshly separated PBMC lysates. Quantitative HIV- 1 proviral load analysis by serial dilution of 23 F-T BCs and 8 WB lysates showed results comparable to those obtained with lysates of fresh PBMCs. Values for WBC differential and immunophenotyping could be applied to express viral load relative to total WBCs, PBMCs, or CD4+ cells. These results establish the basis for simplified virologic analysis of cryopreserved BC or WB specimens.     

A venetoclax and azacitidine bridge-to-transplant strategy for NPM1-mutated acute myeloid leukaemia in molecular failure

NPM1-mutated acute myeloid leukaemia (NPM1^mut AML) represents a mostly favourable/intermediate risk disease that benefits from allogeneic haematopoietic stem cell transplantation (HSCT) in case of measurable residual disease (MRD) relapse or persistence after induction chemotherapy. Although the negative prognostic role of pre-HSCT MRD is established, no recommendations are available for the management of peri-transplant molecular failure (MF). Based on the efficacy data of venetoclax (VEN)-based treatment in NPM1^mut AML older patients, we retrospectively analysed the off-label combination of VEN plus azacitidine (AZA) as bridge-to-transplant strategy in 11 NPM1^mut MRD-positive fit AML patients. Patients were in MRD-positive complete remission (CR_MRDpos) at the time of treatment: nine in molecular relapse and two in molecular persistence. After a median number of two cycles (range 1–4) of VEN–AZA, 9/11 (81.8%) achieved CR_MRD-negative (CR_MRDneg). All 11 patients proceeded to HSCT. With a median follow-up from treatment start of 26 months, and a median post-HSCT follow-up of 19 months, 10/11 patients are alive (1 died from non-relapse mortality), and 9/10 patients are in MRD_neg status. This patient series highlights the efficacy and safety of VEN–AZA to prevent overt relapse, achieve deep responses and preserve patient fitness before HSCT, in patients with NPM1^mut AML in MF.