A randomized prospective comparison of oral versus intraperitoneal ofloxacin as the primary treatment of CAPD peritonitis

Summary: Oral ofloxacin has been successfully used in our centres for the primary treatment of peritonitis complicating continous ambulatory peritoneal dialysis (CAPD). In view of the progressive rise in the resistance rate to ofloxacin among peritoneal bacterial isolates, a study was conducted to determine if oral ofloxacin remains a viable first line treatment for CAPD peritonitis in our centres and if the result can be improved by changing from an oral to an intraperitoneal (i.p.) route. In patients on three 2 L daily CAPD exchanges, ofloxacin given at the i.p. dosage of 200 mg loading followed by 25 mg/L of peritoneal dialysate achieved overnight trough peritoneal levels which are at least four times the minimal 90% inhibitory concentration (MIC90) of most bacterial pathogens without significant accumulation in the systemic circulation. This i.p. dosage was therefore chosen for the clinical study and the result was compared to that using ofloxacin given in the oral dosage of 400 mg loading followed by 300 mg once daily as maintenance. of all the recruited episodes, 35 were eligible for analysis. the overall primary cure rate including primary failures and relapses was 55.6% (10/18) in the oral treatment group and 70.6% (12/17) in the i.p. treatment group. the corresponding figures for gram positive bacterial (g +) infections were 36.4% and 50%, for gram negative bacterial (g -) infections were 66.7 and 80% and for culture negative infections were 75 and 80%. In culture positive cases, all treatment failures were due to resistant infections which were observed in 42.3% of all bacterial isolates, 47.1% of g + isolates and 33.3% of g - isolates. Due to the high background level of bacterial resistance among our CAPD population, ofloxacin monotherapy given either by the oral or the i.p. route can no longer be recommended for the primary treatment of CAPD peritonitis.     

阿克拉霉素A聚氰基丙烯酸异丁酯毫微粒冻干针剂体内外抗肝癌活性

阿克拉霉素A聚氰基丙烯酸异丁酯毫微粒的冻干针剂,能明显抑制体外培养人肝癌细胞株7703的生长,IC50为0.28μg·ml^-1。在0.8μg·ml^-1浓度时,克隆形成抑制率为90%,抑制作用有明显剂量依赖关系而未见明显时间依赖关系。静脉给药后,对常位移植人肝癌模型裸小鼠的抑瘤率为86.84%,肿瘤细胞增殖活性阳性率为20.83%。体内外均显示明显的抗肝癌活性,且体内抗肝癌活性比阿克拉霉素A冻干针剂强。     

原代培养鼠胚脊髓运动神经元活力状态与肌萎灵注射液的干预作用

目的:从细胞水平,观察扶元起萎,养荣生肌的中药制剂肌萎灵注射液对原代培养鼠胚脊髓运动神经元的保护作用。方法:实验于2004-03/2005-03在解放军第三军医大学完成。实验分组:清洁级SD雌性孕鼠,孕期10～14d。应用密度梯度离心法分离鼠胚脊髓运动神经元进行原代培养,运动神经元培养72h后,按培养板及孔分为4.5μg/L肌萎灵组(肌萎灵注射液,含生药0.9g/mL)、9μg/L肌萎灵组、45μg/L肌萎灵组、力如太组(力如太R利鲁唑片,应用时经0.9%NaCl-0.01NHCl溶解)和对照组。实验处理:各组分别加入用新鲜培养基稀释为0.5%(终浓度4.5μg/L)、1%(终浓度9μg/L)、5%(终浓度45μg/L)的肌萎灵注射液,力如太组加入利鲁唑(终浓度10μmo/L),对照组加入等量新鲜培养基。实验评估:①共同培养3d用四甲基偶氮唑盐法观察其对细胞活力的影响,以吸光度表示。②采用NF-200免疫组织化学染色并进行图像分析,测定神经突起主干长度。结果:①培养的运动神经元活力状态比较:4.5,9,45μg/L肌萎灵组培养运动神经元活力显著增强,与对照组相比,差异有显著性意义(0.317±0.054,0.396±0.087,0.329±0.097,0.230±0.130,P<0.05)。力如太组细胞生长与对照组相比无显著差异(0.266±0.141,0.230±0.130,P>0.05)。②脊髓运动神经元突起生长情况结果:4.5μg/L肌萎灵组和9μg/L肌萎灵组可促进其生长,与对照组相比,差异有显著性意义[(315.96±32.32),(373.46±80.24),(159.71±48.95)μm,P<0.05]。结论:肌萎灵注射液可增强运动神经元活力,促进脊髓运动神经元突起的生长。     

Erythrocyte membrane proteins reactive with IgG (warm-reacting) anti- red blood cell autoantibodies: II. Antibodies coprecipitating band 3 and glycophorin A

In our initial immunochemical study of the red blood cell (RBC) membrane proteins targeted in 20 cases of warm-antibody autoimmune hemolytic anemia (AHA), RBC eluates of 6 patients mediated immunoprecipitation (IP) of both band 3 and glycophorin A (GPA). This dual IP pattern had previously been observed with murine monoclonal antibodies (MoAbs) against the high frequency blood group antigen, Wrb (Wright), suggesting that the Wrb epitope may depend on a band 3-GPA interaction. Earlier, anti-Wrb had been identified serologically as a prominent non-Rh specificity of AHA autoantibodies. In the present study, 6 autoantibody eluates immunoprecipitating band 3 and GPA from common Wr(b+) RBCs were retested, in parallel with murine anti-Wrb MoAbs, against very rare Wr(a+b-)En(a+)RBCs. One patient's autoantibodies were unreactive with the Wr(b-) RBCs by either IP or indirect antiglobulin test (IAT) and were judged to have "pure" anti- Wrb specificity. Two other patients' autoantibodies displayed both IP and serologic evidence for anti-Wrb as a major component in combination with one or more additional specificities. However, among 3 other patients whose autoantibodies coprecipitated band 3 and GPA, there was no reduction in IP or IAT reactivity with Wr(b-) RBCs in 2 and only slight reduction in the third. We conclude (1) that human anti-Wrb autoantibodies, like their murine monoclonal counterparts, coprecipitate band 3 and GPA from human RBCs; but (2) that not all antibodies with this IP behavior have anti-Wrb serologic specificity, as defined by this donor's Wr(b-) RBCs. The possibility of an additional (non-Wrb) RBC epitope dependent on a band 3-GPA interaction is raised.     

Mutations of conserved arginines in the membrane domain of erythroid band 3 lead to a decrease in membrane-associated band 3 and to the phenotype of hereditary spherocytosis

To elucidate the molecular basis of band 3 deficiency in a recently defined subset of patients with autosomal dominant hereditary spherocytosis (HS), we screened band 3 cDNA for single-strand conformation polymorphism (SSCP). In 5 of 17 (29%) unrelated HS subjects with band 3 deficiency, we detected substitutions R760W, R760Q, R808C, and R870W that were all coinherited with the HS phenotype. The involved arginines are highly conserved throughout evolution. To examine whether or not the product of the mutant allele is inserted into the membrane, we studied one HS subject who was doubly heterozygous for the R760Q mutation and the K56E (band 3sMEMPHIS) polymorphism that results in altered electrophoretic mobility of the band 3 Memphis proteolytic fragments. We detected only the band 3MEMPHIS in the erythrocyte membrane indicating that the protein product of the mutant, R760Q, band 3 allele is absent from the red blood cell membrane. These findings suggest that the R760Q substitution, and probably the other arginine subsitutions, produce band 3 deficiency either by precluding incorporation of the mutant protein into the red blood cell membrane or by leading to loss of mutant protein from differentiating erythroid precursors.     

Acquired immunodeficiency syndrome-associated non-Hodgkin's lymphomas and other malignancies in patients with hemophilia

Non-Hodgkin's lymphoma (NHL) is the most common human immunodeficiency virus (HIV)-associated malignancy in hemophiliacs. We studied the incidence and clinicopathologic features of NHL in 3,041 hemophiliacs followed at 18 US Hemophilia Centers between 1978 and 1989. Of the 1,295 (56.6%) who were HIV(+), 253 (19.5%) developed acquired immunodeficiency syndrome (AIDS), of whom 14 (5.5%) developed NHL. Three NHL occurred in HIV(-) hemophiliacs, for a 36.5-fold greater risk in HIV(+) than HIV(-) hemophiliacs (P < .001). The NHL incidence rate was 29-fold greater than in the US population by Surveillance, Epidemiology, and End Results (SEER) estimates (P < .001). Between 0 and 4 lymphomas have been observed per year between 1978 and 1989. At presentation 13 (92.9%) of the HIV(+) NHL were extranodal. Ten were stage IV, 1 stage II, and 3 stage IE. Ten (71.4%) were high-grade, 3 (21.4%) intermediate-grade, and 1 (7.1%) was a low-grade B-cell lymphoma. Epstein-Barr virus (EBV) DNA was detected in 36% by in situ hybridization, including one central nervous system (CNS) lymphoma. The mean CD4 cell count at NHL diagnosis was 64/mm3, the mean latency from initial HIV infection was estimated to be 59 months, and the median survival was 7 months. The incidence of basal cell carcinoma in HIV(+) hemophiliacs was 18.3-fold greater than in HIV(-) hemophiliacs (P < .001) and 11.4-fold greater than in the US population (P < .001). In conclusion, incidence rates of NHL and basal cell carcinoma in HIV(+) hemophiliacs are significantly increased over rates in HIV(-) hemophiliacs and over rates in the US population. Clinicopathologic presentation of NHL in HIV(+) hemophiliacs is similar to that in HIV(+) homosexual men.     

Production and characterization of monoclonal antibodies to the subunits of human phosphofructokinase: new tools for the immunochemical and genetic analyses of isozymes

Recently we have demonstrated that human phosphofructokinase (PFK; ATP: D-fructose-6-P, 1-phosphotransferase; EC.2.7.1.11) is under the control of three structural loci that code for M (muscle-type), L (liver-type), and P (platelet-type) subunits: random tetramerization of these subunits produces various isozymes. In this study, we have produced and characterized BALB/c hybridoma antibodies to the M- and L-type subunits of human PFK. The specific antibodies were detected by an enzyme- immunoprecipitation assay using Staphylococci-bearing protein A as an immunoadsorbent. Of the wells tested using red blood cell (RBC) PFK (M + L), 61% were positive. Only one M-specific hybridoma was identified. The one anti-M and 4 anti-L antibodies were characterized for their biochemical and immunochemical specificities. To define the combining specificities of these antibodies, we compared their reactivity and that of monospecific rabbit anti-M antiserum with muscle and liver PFKs from 15 different vertebrate species. The rabbit anti-M shows strong cross-reactivity with the muscle PFKs from all the species studied. In contrast, the monoclonal anti-M reacts exclusively with muscle PFKs from primates. Two of four anti-L antibodies react only with human L- PFK, whereas the other two react with that from a few other vertebrate species as well. Taken together, these data suggest that primate- specific antibodies recognize evolutionarily, recently acquired antigenic determinants, whereas the antibodies reactive with PFKs from distantly related species recognize conserved determinants. The differential immunoreactivities of muscle and liver PFKs strongly suggest the presence of distinct isozymes in all the vertebrate species studied. These studies demonstrate that it is feasible to produce and characterize monoclonal antibodies that distinguish among isozymes with structural and functional similarities. These antibodies provide sensitive tools in the analyses of isozyme structure, genetics, and related fields.     

Defective platelet-fibrinogen interaction in hereditary canine thrombopathia

A unique, intrinsic, hereditary canine platelet disorder attributable to abnormal fibrinogen receptor availability is described. Thrombopathic platelets from 13 severely affected basset hounds failed to aggregate in response to all agonists tested except thrombin. Normal platelet interaction with the various stimuli was inferred on the basis of their ability to elicit unimpaired shape change in thrombopathic platelets. No quantitative differences in major platelet membrane glycoproteins, intraplatelet fibrinogen, adenine nucleotides, or serotonin uptake were detected. Dense granule secretion was impaired. The ultrastructural appearance of thrombopathic platelets was normal. Fibrinogen-platelet interaction was evaluated by reacting platelet-rich plasma (PRP) with fibrinogen coupled to polymeric acrylonitrile beads and scoring the extent of stimulus-induced agglutination. The aggregatory responses of normal and thrombopathic platelets were closely correlated with fibrinogen receptor availability. In contrast to human platelets, epinephrine-stimulated canine platelets did not interact with immobilized fibrinogen, and arachidonate generally induced only weak agglutination. Thrombopathic platelets agglutinated fibrinogen beads at reduced rates when stimulated with physiologic doses of thrombin and high-dose calcium ionophore, A23187. Our data suggest that thrombin-mediated induction of canine platelet fibrinogen receptors may proceed by pathway(s) alternate to those shared by other platelet agonists, and/or that secreted granule constituents may act synergistically with thrombin to overcome inhibition of signal-response- coupled reactions mediating the interaction of fibrinogen with its receptor. This congenital platelet defect provides further evidence, in a species other than human, for the pivotal role of fibrinogen receptor induction in platelet aggregation.     

Detection of maternal cells in human umbilical cord blood using fluorescence in situ hybridization

Cord blood is a potential source of hematopoietic stem cells for transplantation and is being used on a growing number of patients. However, there are concerns that cord blood might be contaminated with maternal cells that could lead to graft-versus-host disease. To ascertain the extent to which maternal cell contamination of cord blood occurs, we examined 49 cord blood samples from male babies for maternal cells by fluorescence in situ hybridization using probes to the X and Y chromosomes. A minimum of 1,000 nuclei were scored from each sample, and maternal cells were found in 7 of the 49 cord bloods, at levels ranging from 0.04% to 1.0%. In addition, in 39 and 27 of the cord blood samples, respectively, we examined the CD8+ and CD34+ cell populations for maternal cells. Maternal cells were found in 5 of the 39 CD8 fractions and in 1 of the 27 CD34 fractions, at levels similar to that found in the unfractionated cord blood. In sum, maternal cells were found in either the unseparated mononuclear fraction or the CD8 or CD34 fractions in 10 of the 49 cord blood samples (20%). These results show that maternal cells are present in a substantial number of cord bloods, and that some of these maternal cells are T cells.