Murine T cells expressing high activity of prolyl endopeptidase are susceptible to activation-induced cell death

Prolyl endopeptidase (PEP) is widely distributed and thought to play an important role in the degradation of peptide hormones and neuropeptides, but its biological role is totally unknown. In this study, we examined PEP activity in subpopulations of murine T cells and found that PEP activity was significantly higher in immature thymocytes than in mature thymocytes or in peripheral T cells. Stimulation of murine peripheral T cells time-dependently increased PEP activity. Although murine T cell hybridomas exhibited high PEP activity, the PEP activity was fully inhibited by treatment with PEP inhibitor. The pretreated T cells were found to be resistant to activation-induced cell death (AICD). Similar results were obtained in murine thymocytes as well as in activated peripheral T cells. PEP activity in T cell hybridomas remained unchanged during AICD. These results suggest that T cells expressing high PEP activity are susceptible to ACID.     

Self-Enhancement of Hepatitis C Virus Replication by Promotion of Specific Sphingolipid Biosynthesis

Lipids are key components in the viral life cycle that affect host-pathogen interactions. In this study, we investigated the effect of HCV infection on sphingolipid metabolism, especially on endogenous SM levels, and the relationship between HCV replication and endogenous SM molecular species. We demonstrated that HCV induces the expression of the genes (SGMS1 and 2) encoding human SM synthases 1 and 2. We observed associated increases of both total and individual sphingolipid molecular species, as assessed in human hepatocytes and in the detergent-resistant membrane (DRM) fraction in which HCV replicates. SGMS1 expression had a correlation with HCV replication. Inhibition of sphingolipid biosynthesis with a hepatotropic serine palmitoyltransferase (SPT) inhibitor, NA808, suppressed HCV-RNA production while also interfering with sphingolipid metabolism. Further, we identified the SM molecular species that comprise the DRM fraction and demonstrated that these endogenous SM species interacted with HCV nonstructural 5B polymerase to enhance viral replication. Our results reveal that HCV alters sphingolipid metabolism to promote viral replication, providing new insights into the formation of the HCV replication complex and the involvement of host lipids in the HCV life cycle.     

Inhibition of rosette formation by antithymocyte globulin

Summary The peripheral blood leukocytes from 29 patients with adenocarcinoma of the colon were studied sequentially for the T-cell level, the rosette-inhibition titer of antithymocyte globulin and the blastogenic response to PHA and Con A to evaluate the T-cell immunocompetence. The level of E-rosette-forming cells and the blastogenic response did not reflect the immunocompetence of the T-cell population. Fluctuations in the rosette-inhibition titer of antithymocyte globulin (ATG) were observed; an increased ATG titer indicated an unfavorable clinical course, while a decreased ATG titer was observed with patients who had a favorable clinical course. The rosette-inhibition titer with antithymocyte globulin was observed to be an important indicator of competent T cells in patients with colorectal cancer.Supported by the Physicians' Medical Education and Research Foundation and General Research Supports FundsAbbreviations used that are not spelled out in the text are: AET 2-aminoethylisothiouronium bromide; ATG Antithymocyte globulin; ALG Antilymphocyte globulin; E-RFC Erythrocyte rosette-forming cells; PHA Phytohemagglutinin; Con A Concanavalin A; E Sheep erythrocyte     

Improving effect of ethyl eicosapentanoate on statin-induced rhabdomyolysis in Eisai hyperbilirubinemic rats

The effect of ethyl eicosapentanoate (EPA-E) on statin-induced rhabdomyolysis was investigated by co-administration of EPA-E and pravastatin (PV), as a typical statin, to Eisai hyperbilirubinemic rats (EHBR). It was confirmed that the plasma PV concentration was not affected by simultaneous administration of EPA-E, and there was no cumulative increase of PV during prolonged co-administration of EPA-E and PV. Muscular degeneration was prominent (incidence 5/5; average grade 3.5 (range 2-4)) in EHBR treated with PV alone at 200 mg/kg/day for 14 days, but co-administration of EPA-E at doses of 100, 300, and 1000 mg/kg/day decreased the average grades to 1.4 (range 0.3-3.0), 0.5 (0.2-1.0), and 0.6 (0.0-1.7), respectively. Creatine phosphokinase (CPK) and myoglobin levels in plasma were well correlated with the grade of skeletal muscle degeneration. Thus, EPA-E appears to reduce the severity of statin-induced rhabdomyolysis.     

禽多杀性巴氏杆菌C48-3株ompH基因敲除突变株的构建及其生物学功能

[目的]探讨ompH基因在禽多杀性巴氏杆菌致病过程中的作用.[方法]利用同源重组原理构建中间为四环素抗性基因,两侧为ompH基因上下游同源序列同源的敲除载体pWSK29△ompH,将敲除载体电击转入C48-3株感受态细胞中,通过四环素抗性和菌落PCR筛选ompH基因的敲除突变株,并通过组合PCR、逆转录PCR和DNA测序对突变株进行验证.用生物学功能实验比较野生株、互补株和突变株在生长速率、荚膜结构、粘着能力和致病性等方面的差异.[结果]组合PCR、逆转录PCR和DNA测序结果证实ompH基因的敲除突变株C48-3△ompH构建成功,电镜观察结果证实ompH基因的缺陷影响细菌的荚膜合成能力,粘附实验结果显示与野生株C48-3和互补株C48-3C相比突变株C48-3△ompH对CEF细胞的粘附能力显著降低(P＜0.01),而小鼠毒力实验结果表明突变株C48-3△ompH的致病性相对减弱.[结论]本实验构建的突变株C48-3△ompH,为进一步研究多杀性巴氏杆菌的致病机理奠定基础.     

Structure of a novel highly branched α-glucan enzymatically produced from maltodextrin

The bacterial strain PP710, isolated from soil and identified as Paenibacillus species, produced a low-digestibility α-glucan containing a large amylase-resistant portion. This α-glucan was obtained in high yields from maltodextrin (dextrose equivalent 3) by using the condensed culture supernatant of the strain as the enzyme preparation. The water-soluble dietary fiber content of the low-digestibility α-glucan was 80.2%, and showed resistance to a rat intestinal enzyme preparation. The α-glucan was found to be a novel highly branched α-glucan by acid hydrolysis, NMR analysis, gel permeation chromatography, methylation analysis, and enzymatic digestion.     

Development and characterization of an anti-rituximab monoclonal antibody panel

During the development of monoclonal antibodies (mAbs) and other therapeutic proteins, immunogenicity, in particular the induction of anti-drug antibodies (ADAs), is an important concern, and thus immunogenicity assessment is a requirement for their approval. Establishment of appropriate methods for detecting and characterizing ADAs is necessary for immunogenicity assessment, but the lack of commonly available reference standards makes it difficult to compare and evaluate the methods. It is also difficult to compare the data with those obtained by other methods or facilities without reference standards. Here, we developed a panel of ADAs against anti-CD20 rituximab (Rituxan®, MabThera®); the panel consisted of eight clones of recombinant human-rat chimeric mAbs that target rituximab. The anti-rituximab mAbs showed different binding properties (specificity, epitope and affinity), and different neutralization potencies for CD20 binding, complement-dependent cytotoxicity and antibody-dependent cell-mediated cytotoxicity. The molecular size of the immune complex consisting of rituximab and the anti-rituximab mAb differed among the clones, and was well correlated with their level of Fcγ-receptor activation. These results suggest that the ADAs chosen for the newly developed panel are suitable surrogates for human ADAs, which exhibit different potential to affect the efficacy and safety of rituximab. Next, we used this panel to compare several ADA-detecting assays and revealed that the assays had different abilities to detect the ADAs with different binding characteristics. We conclude that our panel of ADAs against rituximab will be useful for the future development and characterization of assays for immunogenicity assessment.     

Distribution of manno-heptulose and sedoheptulose in plants

Upon investigation of the distribution of heptuloses in plants, most of the plants which were positive to the preliminary PC of the heptulose were found to contain manno-heptulose by the confirmative tests with descending PC and GLC of TMS derivatives after purification on the thick paper chromatogram. The amounts of manno-heptulose in many plants were comparable to those of sedoheptulose, and often higher than that of the latter.