Significance of low-level DSA detected by solid-phase assay in association with acute and chronic antibody-mediated rejection

We sought to clarify the controversial issue of whether detecting low‐level anti‐donor‐specific HLA antibody (HLA‐DSA) by single‐antigen flow‐bead assay (SAFB) may have a potential role in reducing acute and chronic antibody‐mediated rejection (AMR). We retrospectively studied the preoperative serum of ABO‐compatible living kidney transplantation recipients transplanted between 2001 and 2004 by SAFB using a Luminex platform. HLA‐DSA was detected only by SAFB in 24 patients, although all of them showed negative T‐cell and B‐cell complement‐dependent cytotoxicity (CDC) crossmatches. The HLA‐DSA patients went on to have surprisingly high levels of acute and chronic AMR despite being only weakly sensitized (acute AMR, 33.3%; chronic AMR, 41.7%). After 2005, we implemented SAFB routinely and any patient having a positive HLA‐DSA was considered to be a desensitization candidate. The 52 patients found to have HLA‐DSA underwent kidney transplantation after prior treatment with a single dose of rituximab (RIT) and three or four sessions of double‐filtration plasmapheresis (DFPP) in addition to regimens commonly used between 2001 and 2004. After 2005, there was a significant reduction in the occurrence of acute and chronic AMR (acute AMR, 4.7%, P < 0.001; chronic AMR, 4.7%, P < 0.001). The 5‐year graft survival rate also improved after implementing SAFB (83.3–98.1%, P = 0.032). The RIT/DFPP‐induction protocol may improve graft survival even in patients with low‐level DSA.     

Hepatitis B virus wild‐type and rtN236T populations show similar early HBV DNA decline in adefovir refractory patients on a tenofovir‐based regimen

Summary. Hepatitis B virus (HBV) pol/RT mutations that confer clinical resistance to tenofovir disoproxil fumarate (TDF) have not been detected to date. In vitro, the rtN236T adefovir dipivoxil (ADV)‐associated resistance mutation confers low‐level cross‐resistance to tenofovir: 3‐ to 13‐fold changes in EC₅₀ from wild type. This study evaluated the clinical response of rtN236T mutant viruses by comparing their early viral load decay kinetics to wild‐type viruses in chronic HBV monoinfected patients harbouring rtN236T prior to initiating TDF or emtricitabine (FTC)/TDF therapy. Baseline samples (n = 105) from adefovir refractory patients were tested for the presence of rtN236T using a highly sensitive allele‐specific PCR assay with an rtN236T detection cut‐off of 0.5%. The rtN236T mutation was detected at baseline in 14.3% (14/98) of analysable patient samples (0.5–93.2%, rtN236T percentage range). The median change in total HBV DNA at week 24 was comparable for patients with rtN236T detected at baseline (?3.7 log₁₀ copies/mL, n = 14) as compared to patients with wild‐type HBV (?3.2 log₁₀ copies/mL, n = 90). In patients with rtN236T, wild‐type and rtN236T mutant virus showed similar rates of HBV DNA decline with no statistically significant difference observed at week 4. Moreover, the proportion of rtN236T remained unchanged in patients in either arm of the study during treatment. In conclusion, the rtN236T mutant virus showed similar HBV DNA decline kinetics to wild‐type virus in adefovir refractory patients who switched to TDF or FTC/TDF. Despite low levels of cross‐resistance in vitro, TDF similarly suppresses wild‐type and rtN236T mutant viruses in vivo.     

Persistent hepatitis C virus (HCV) infection impairs HCV‐specific cytotoxic T cell reactivity through Mcl‐1/Bim imbalance due to CD127 down‐regulation

Summary. In persistent hepatitis C virus (HCV) infection, HCV‐specific cytotoxic T lymphocyte (CTL) reactivity is impaired and this affects HCV control. Interleukin‐7 receptor (CD127) expression on these cells could regulate CTL reactivity through Mcl‐1/Bim balance modulation. Bim is a pro‐apoptotic molecule blocked by the action of Mcl‐1. Mcl‐1/Bim expression and T cell reactivity on HCV‐specific CTLs were compared according to CD127 phenotype. Peripheral blood lymphocytes (PBL) from HLA‐A2⁺ HCV⁺ patients were obtained. HCV‐specific CTLs were visualized by staining PBL with anti‐CD8 and HLA‐A2/peptide pentameric complexes (pentamer). Mcl‐1/Bim/CD127 phenotype of HCV‐specific CTLs was tested by staining detectable CD8⁺/pentamer⁺ cells with anti‐Mcl‐1/Bim/CD127 antibodies. HCV‐specific CTL proliferation ability after specific in vitro challenge was tested in the presence and absence of pancaspase inhibitor z‐VAD‐fmk. All stained cells were analysed by flow cytometry. CD127^low‐expressing HCV‐specific CTLs associated with high HCV viraemia, while CD127^high correlated with undetectable viral loads (P < 0.001). Directly ex vivo, pentamer⁺ cell frequency was similar according to CD127 expression level. Nevertheless, CD127^low pentamer⁺ cell proliferation after specific in vitro challenge was impaired (P < 0.05), although this was corrected by z‐VAD‐fmk treatment (P < 0.05). Mcl‐1 expression was low directly ex vivo (P < 0.01), and Bim was up‐regulated after antigen encounter (P < 0.05) of CD127^low pentamer⁺ cells. The ex vivo difference between Mcl‐1 and Bim expression on pentamer⁺ cells correlated positively with CD127 expression level (P < 0.001) and with pentamer⁺ cell reactivity (P < 0.05). In summary, a low ex vivo Mcl‐1 expression and Bim up‐regulation after antigen encounter are involved in CD127^low HCV‐specific CTL hyporeactivity during chronic infection, but it can be overcome by apoptosis blockade.     

七氟烷对成年大鼠认知功能影响的研究

目的探讨不同浓度（1%,2%,3%）七氟烷对成年大鼠的认知功能及血清、海马组织中神经元特异性烯醇化酶（NSE）的影响。方法将40只成年雄性SD大鼠随机分为1%七氟烷组、2%七氟烷组、3%七氟烷组及对照组（吸入空气）,七氟烷吸入3h。处理后第1,2,3,4,5天进行Morris水迷宫的定位航行试验和第6天的空间探索试验。第6天测试完毕后取大鼠的血清和海马组织测定NSE的含量。结果吸入不同浓度七氟烷组与对照组相同时间点比较,逃避潜伏期和总路程差异无统计学意义（P＞0.05）。同时,不同浓度七氟烷组间比较,逃避潜伏期和总路程差异也无统计学意义（P＞0.05）。吸入七氟烷或空气后第6天撤台,各组大鼠的目标象限停留时间百分比和穿越平台次数之间差异也无统计学意义（P＞0.05）。吸入1%和3%七氟烷组大鼠血清中NSE水平低于对照组（P＜0.01）;同时,吸入2%和3%七氟烷组大鼠血清中NSE水平高于吸入1%七氟烷组（P＜0.01,P＜0.05）。海马组织中的NSE含量各组间差异无统计学意义（P＞0.05）。结论单次吸入1%,2%和3%的七氟烷,短期不影响成年大鼠的空间学习记忆能力。同时,吸入1%和3%七氟烷可以降低大鼠血清中NSE水平。     

金属硫蛋白对大鼠心肌缺血后再灌注损伤的影响——电镜形态计量学研究

用离体大鼠心脏缺血后再灌注模型,根据心肌亚微结构改变及线粒体比表面测定,揭示金属硫蛋白对缺血后再灌注损伤有明显保护作用,提示金属硫蛋白是一种细胞保护剂,可望用于临床,控制缺血后再灌注损伤。     

Monocyte-derived dendritic cells from highly atopic individuals are not impaired in their pro-inflammatory response to toll-like receptor ligands

BACKGROUND: Toll-like receptor (TLR) agonists are widely used as adjuvants in specific immune therapy protocols for patients with atopic disposition. Monocyte-derived dendritic cells (mDCs) are thought to be important target cells for these compounds. OBJECTIVES: To compare surface markers, TLR expression, TLR functionality after ligand stimulation, and genetic polymorphisms in the TLR 2-, 3-, and 4-genes in mDCs from atopic vs. non-atopic patients. METHODS: mDCs from highly atopic individuals (total serum IgE >1000 IU/mL) and healthy control persons (total serum IgE <75 IU/mL) were screened for TLR 1-10 expression by real-time PCR. Receptor function was analysed by IL-12 and TNF-alpha production after incubation with the respective ligands peptidoglycan (PGN) (TLR 2), polyriboinosinic-polyribocytidylic acid (poly IC) (TLR 3), lipopolysaccharide (LPS) (TLR 4), flagellin (TLR 5), and CpG-DNA/non-CpG-DNA (TLR 9). Haplotype-tagging single-nucleotide polymorphisms of the TLR 2-, 3-, and 4- genes were analysed for genetic associations. RESULTS: mDC from atopic patients showed a very similar pattern of TLR expression as controls with strong expression of TLR 2, 4, 5, 6, and 8, moderate expression of TLR 1 and 3, and no or very low expression of TLR 7, 9, and 10. After stimulation with TLR ligands, mDCs from atopic patients acquired a mature phenotype with a tendency towards a higher up-regulation of the co-stimulatory molecules CD80, CD83, and CD86 than control mDCs. IL-12 and TNF-alpha were produced at a similar level in both groups of DCs. Among the different TLR agonists, poly IC showed the strongest activation of DCs, followed by LPS, PGN, and flagellin. This was paralleled by a strong functional expression of protein kinase R and retinoid-inducible gene-I (RIG-I), two additional poly IC-sensing receptors in both groups. Genetic analysis of single-nucleotide polymorphisms in the TLR 2-, 3-, and 4-genes in both groups revealed no major allele or genotype differences. CONCLUSIONS: mDC from atopic patients are not restricted in their response to TLR-ligands. TLR agonists seem to be suitable to induce pro-inflammatory immune responses and maturation in mDCs from highly atopic individuals and represent reasonable adjuvants for specific immunotherapy reagents.     

应用ELISA检测尿螨病患者血清总IgE和粉螨特异性IgE水平

目的:探讨 IgE和粉螨特异性IgE在尿螨病诊断中的应用价值。方法:用 ELISA法对69例尿螨病患者进行血清总IgE、螨特异性IgE的检测。结果:尿螨病患者血清总IgE及螨特异性IgE水平分别为89±22IU/ml、71±22IU/ml,而正常对照组血清总IgE及螨特异性IgE水平分别为35±27IU/ml、29±18IU/ml,两者相比差异有显著性（P<0.01～0.05）。其中轻度、中度、重度螨感染者的血清总IgE及螨特异性IgE水平分别为82±18IU/ml、95±24IU/ml、106±26IU/ml和 65±19IU/ml、78±24IU/ml、84±23IU/ml,相互比较差异有显著性（P<0.01）。结论:尿螨病患者血清总IgE和粉螨特异性IgE水平均明显增高,其病变程度与血清总IgE和粉螨特异性IgE水平似有一定关系。