Selection of Stable Reference Genes for Quantitative Real-Time PCR on Herbaceous Peony (<i>Paeonia lactiflora</i> Pall.) in Response to Drought Stress

Herbaceous peony (Paeonia lactiflora Pall.), as a high-end cut flower in the international market, has high ornamental and medicinal values. But in Northern China, drought is a major environmental factor influencing the growth and development of P. lactiflora. Quantitative real-time polymerase chain reaction (qRT-PCR) can evaluate gene expression levels under different stress conditions, and stable internal reference is the key for qRT-PCR. At present, there is no systematic screening of internal reference for correcting gene expressions of P. lactiflora in response to drought stress. In this study, 10 candidate genes [ubiquitin (UBQ2), UBQ1, elongation factor 1-α (EF-1α), Histidine (His), eukaryotic initiation factor (eIF), tubulin (TUB), actin (ACT), UBQ3, ACT2, RNA polymerase II (RNA Pol II)] were chosen, and 4 analysis methods were used to compare the stabilities for these 10 genes coping with drought stress. Due to the difference of operation methods, the results of different analysis were distinct, and the final comprehensive analysis indicated that EF-1α was a relatively stable internal reference gene for P. lactiflora under drought stress. Also, UBQ1 and UBQ2 were the best reference gene combination according to GeNorm analysis. This study will lay a foundation for screening the key genes of P. lactiflora in response to drought stress.     

Associations between whole blood trace elements concentrations and HbA1c levels in patients with type 2 diabetes

BioMetals - Previous researches have been conducted to study the associations of trace elements on Type 2 diabetes (T2D) risk. The present study focuses on the evaluation of potential associations...     

Molecular characterization of a plant FKBP12 that does not mediate action of FK506 and rapamycin

Immuonosuppressive drugs FK506 and rapamycin block a number of signal transduction pathways in eukaryotic systems. The 12 kDa FK506 binding protein (FKBP12) mediates the action of both FK506 and rapamycin against their functional targets. In this report, we cloned, sequenced and characterized a gene encoding FKBP12 in Vicia faba ( Vf FKBP12). While Vf FKBP12 is highly homologous to animal and yeast FKBP12, it does not mediate the action of FK506 and rapamycin. There are unique features in plant FKBP12 sequences that cause the variation in their function. One lies in the domain that is critical for interaction with calcineurin (CaN), the mammalian and yeast target of FKBP12-FK506 complex. Protein–protein interaction assays revealed a low-affinity and unstable Vf FKBP12-FK506-CaN ternary complex. In the genetic assay, Vf FKBP12 did not restore the sensitivity of yeast FKBP12 mutant to rapamycin or FK506, supporting that plant FKBP12-ligand complexes are unable to block the function of the drug target. Also unique to plant FKBP12 proteins, a pair of cysteines is spatially adjacent to potentially form disulfide linkage. Treatment of Vf FKBP12 with reductant dithiothreitol (DTT) abolished the formation of Vf FKBP12-FK506-CaN ternary complex. Site-directed mutagenesis to substitute one of the cysteines, Cys²⁶, with Ser produced a similar effect as DTT treatment. These results indicate that an intramolecular disulfide bond is a novel structural feature required for the low–affinity interaction between plant FKBP12 and CaN. In conclusion, plant FKBP12 proteins have evolved structural changes that modify their protein-protein interacting domains and cause loss of function against the drug targets.     

脐带间充质干细胞治疗熊去氧胆酸应答不佳的原发性胆汁性胆管炎的临床观察

目的探讨脐带间充质干细胞（UC-MSCs）输注治疗熊去氧胆酸（UDCA）应答不佳的原发性胆汁性胆管炎（PBC）患者的安全性和有效性，分析影响UC-MSCs疗效应答的相关因素。 方法选取解放军总医院第五医学中心2010年8月至2017年10月接受UC-MSCs输注治疗UDCA应答不佳的PBC患者29例。患者均以4周为间隔给予3次外周静脉输注细胞1.0×10⁶个/kg。通过实验室指标、生命体征及不良事件发生情况评估UC-MSCs治疗的安全性。通过患者临床症状、肝功指标和Child-Pugh评分评估治疗的有效性。以"巴黎Ⅰ标准"作为疗效标准，评价患者UC-MSCs治疗后的疗效应答情况，比较有效患者及无效患者基线临床症状和肝脏功能差异，分析影响UC-MSCs疗效的相关因素。采用独立样本t检验分析年龄；采用Mann-Whitney U检验比较两组UDCA治疗时间、激素治疗时间、实验室数据等，采用Wilcoxon秩和检验比较组间数据；采用χ²检验比较性别、临床症状和Child-Pugh分级等指标。多因素Cox回归分析对影响UC-MSCs疗效的相关因素。 结果1例患者在治疗后因合并严重感染出现高热，所有患者未出现UC-MSCs相关严重不良事件。UC-MSCs输注后与基线相比，患者的血清碱性磷酸酶（ALP）[281.00 （182.50，428.50）比201.00 （149.50，402.00）]、γ-谷氨酰转移酶（GGT）[156.00 （73.00，390.00）比84.00 （43.50，312.50）]、总胆固醇（TC）[5.10 （3.14，7.69）比3.94 （3.00，6.01）]均下降（P < 0.05）。其中，9例（31﹪）患者治疗后疗效明显，达到"巴黎Ⅰ标准"，与无效组患者基线相比，有效组患者天冬氨酸转氨酶（AST）[93.50 （77.75，100.75）比53.00 （46.00，78.00）]、ALP[342.00 （237.25，516.00）比185.00 （152.50，295.50）]、总胆红素（TBIL）[58.50 （33.45，69.33）比13.10 （11.25，20.25）]均下降（P < 0.05）。多因素Cox回归分析显示，血清TBIL是影响UC-MSCs疗效的重要独立因素[HR为0.817 （95﹪CI：0.715 ~ 0.935），P < 0.05]。 结论UC-MSCs输注对UDCA治疗应答不佳的PBC患者是安全可行的且耐受性良好，部分患者肝功能得到一定的改善。血清TBIL是影响UC-MSCs治疗疗效的独立重要因素，提示在疾病进展早期TBIL较低的阶段进行UC-MSCs治疗可能有效改善和减缓患者疾病进程。