目的 应用转录组学技术分析伴与不伴糖尿病视网膜病变(DR)和糖尿病肾病(DN)的T2DM患者的差异表达基因,寻找DR伴DN相关基因.方法 入组T2DM患者14例,其中8例伴DR和DN纳入DNDR组、6例不伴DN和DR纳入DM组.采集患者外周血,分离白细胞,抽提总RNA,反转录为cDNA,构建转录组文库,Illumina?HiSeqTM2000系统测序,取得每份样本转录组数据.以两组间差异基因表达量均数和中位数差异倍数上下调均≥2.0倍为标准,筛选差异表达基因.并行基因本体(GO)显著性富集分析、生化代谢与信号转导通路(Pathway)显著性富集分析、基因关系网络分析.实时荧光定量PCR(RT-qPCR)检测外周血白细胞蛋白激酶C?delta结合蛋白(PRKCDBP)基因、CD177抗原(CD177)基因的mRNA水平,验证转录组测序结果.结果 与DM组比较,DNDR组筛选出差异表达基因98个,其中上调42个,下调56个.GO富集分析显示,差异基因在分子功能、细胞组分、生物学过程中富集最多的条目分别为结合功能、细胞和细胞成分、单一生物过程.Pathway显著性富集分析显示差异基因参与85个Pathway,抗原处理和递呈为富集最显著的Pathway之一,下调最显著的杀伤细胞免疫球蛋白样受体2DS1(KIR2DS1)参与其中.基因相互作用网络分析示差异表达基因中淋巴因子激活的杀伤T细胞来源蛋白激酶(PBK)与PRKCDBP、Ras关联域家族成员6(RASSF6)之间存在相互作用.杀伤细胞免疫球蛋白样受体2DL1(KIR2DL1)与KIR2DS1有相互作用关系.RT-qPCR示DNDR组PRKCDBPmRNA表达量低于DM组、CD177mRNA表达量高于DM组.结论 两组外周血白细胞中有多种差异表达基因,下调基因中的KIR2DL1、KIR2DS1、SPINK4、PRKCDBP,上调基因中的ABHD11-AS1、WNT3、CD177是否与DNDR的发病和进展有关,有待于进一步验证.

目的 利用转录组学技术研究有无糖尿病视网膜病变(DR)患者差异表达基因,寻找DR相关基因.设计 前瞻性比较性病例系列.研究对象 14例2型糖尿病(T2DM)患者,其中8例合并中度以上非增生性DR(NPDR)或增生性DR(PDR)的T2DM患者纳入DR组,6例不合并DR的T2DM患者纳入DM组.方法 采集每例患者外周血,分离白细胞,抽提总RNA,反转录为cDNA,构建转录组文库,Illumina HiSeqTM2000系统测序,对测序结果进行生物信息学分析.RT-qPCR检测外周血白细胞钙调蛋白1(CNN1)、溶血磷脂酸受体3(LPAR3)mRNA水平,验证转录组测序结果.以两组间差异基因表达量均数和中位数差异倍数上下调均≥2.0倍为标准筛选差异表达基因.主要指标 两组间差异表达基因,差异基因本体(GO)注释和富集、生化代谢与信号转导通路(Pathway)注释和富集、差异基因相互作用网络.两组间外周血白细胞CNN1、LPAR3的mRNA水平差异.结果 与DM组相比,DR组筛选出差异表达基因103个,其中上调41个,下调62个.下调最明显的为杀伤细胞免疫球蛋白样受体2DS1(KIR2DS1),上调最明显的为U105B小核仁RNA基因(RNU105B).GO富集分析显示在生物学过程(GO-P)、分子功能(GO-F)中显著富集的GO条目为细胞过程、单一生物过程、结合功能.谷胱甘肽S转移酶mu1(GSTM1)、早期生长应答1(EGR-1)、LPAR3、CNN1在以上条目均有富集.Pathway显著性富集分析显示,差异基因富集最多的为抗原处理和呈现、自然杀伤细胞介导的细胞毒性通路,KIR2S1均参与其中.差异基因相互作用网络分析示,差异表达基因中GAS1与多个基因之间存在相互作用.RT-qPCR结果显示,与DM组比较,DR组中的CNN1和LPAR3的mRNA含量均升高(P均＜0.05).结论 采用转录组学技术发现,DR组患者外周血白细胞存在多种差异表达基因,下调的基因中KIR2DS1、GSTM1、EGR-1、GAS1,上调的基因中RNUI05B、LPAR3、CNN1是否与DR的发生发展有关有待于进一步验证.

Objective:To explore the efficacy and safety of methimazole combined with levothyroxine in the treatment of patients with Graves disease (GD), and provide theoretical basis for clinical practice.Methods:Sixty-eight patients with confirmed GD who admitted to the 903rd Hospital of PLA from January 2013 to January 2018 were selected and divided into control group and observation group by random number table method.The control group was given propylthiouracil combined with levothyroxine.The observation group was treated with methimazole and levothyroxine.The clinical features of hyperthyroidism, changes in hormone levels, and bone metabolism were compared between the two groups before and after treatment.Results:After treatment, the incidence of exophthalmos, goiter and thyrotropin receptor antibody (TRAb) positive rate were significantly reduced in the two groups (the control group changed from 10 cases to 2 cases, and the observation group changed from 10 cases to 0 case), the differences were statistically significant (χ 2=27.1, 16.2, all P<0.05). The differences in TSH, FT 3, and FT 4 before and after treatment were statistically significant [the control group: TSH changed from (0.02±0.02)mU/L to (3.01±0.94)mU/L, FT 3 from (16.92±2.25)pmol/L to (10.29±1.68)pmol/L, FT 4 from (52.61±10.22)pmol/L to (19.82±4.11)pmol/L; the observation group: TSH from (0.02±0.01)mU/L to (1.97±1.27)mU/L, FT 3 from (17.09±2.72)pmol/L to (3.95±0.84)pmol/L, and FT 4 from (53.82±10.11)pmol/L to (12.65±3.31)pmol/L], and the improvement of TSH, FT 3 and FT 4 in the observation group were better than those in the control group ( t=3.24, 9.51, 16.31, all P<0.05). After treatment, the levels of PTH, CT, blood calcium, blood phosphorus were increased [the control group: PTH changed from (38.32±11.41)ng/L to (42.83±14.22)ng/L, CT changed from (8.66±2.22)ng/mL to (8.01±4.55)ng/mL, blood calcium level changed from (2.01±0.12)pmol/L to (2.53±0.20)pmol/L, blood phosphorus level changed from (1.12±0.08)pmol/L to (1.37±0.09)pmol/L; the observation group: PTH changed from (38.31±12.52)ng/L to (46.33±15.03)ng/L, CT changed from (8.45±2.21)ng/mL to (11.49±7.33)ng/mL, the calcium level changed from (2.02±0.98)pmol/L to (2.82±0.87)pmol/L, the blood phosphorus level changed from (1.10±0.07)pmol/L to (1.42±0.16)pmol/L]. The improvement of PTH and CT in the observation group was better than those in the control group ( t=6.51, 7.31, all P<0.05). Conclusion:Methimazole combined with levothyroxine in the treatment of GD has good therapeutic effect on the clinical characteristics of hyperthyroidism, changes in hormone levels, and bone metabolism.The clinical efficacy is reliable and there are few adverse reactions, which deserves clinical reference.

Objective To screen potential DR-related plasma markers by profiling the plasma proteomics in patients with diabetic retinopathy (DR). Methods A total of 8 patients with moderate or higher non-proliferative diabetic retinopathy (NPDR) or proliferative diabetic retinopathy (PDR) enrolled in our hospital are included in the DR group and 6 patients without DR as NDR group. Plasma samples from these 14 patients were subjected to protein-labeled-free quantification and parallel liquid-phase tandem mass spectrometry (LC-MS/MS) in order to calculate the relative protein abundance of each protein detected in plasma. The results were statistically analyzed to find significant differences between the two groups of proteins. Further, bioinformatics analysis was performed, including gene ontology (GO) and Pathway annotation, GO and Pathway enrichment analysis, and protein interaction network analysis. Results A total of 41 differential proteins were identified with mean and median ratios ≥2.5(up/down). Among them, 26 were up-regulated and 15 were down-regulated. GO analysis showed that the binding protein in the differential protein was dominant, the single-organism process in the biological process accounted for the highest proportion, and the cell and cell part in the cell component accounted for the highest proportion. Pathway enrichment analysis indicated that the most significant up-regulated protein, tropomyosin 4 (TPM4), is associated with regulation of muscle contraction; while the most prominent down-regulated protein, platelet membrane glycoproteins V (GPV), plays an important role in extracellular matrix (ECM) receptor interactions. Protein interaction network analysis showed that there were potential interactions among glyceraldehyde-3-phosphate dehydrogenase (GAPDH), sulfhydryl oxidase 1 (QSOX1), immunoglobulin kappa locus (IGK@), TPM4, apolipoproteins C2 (APOC2), and immunoglobulin heavy variable 4-31(IGHV4-31); another potential interaction between pregnancy zone protein (PZP) and tissue inhibitor of metalloproteinase 2 (TIMP2). Conclusion Profiling the proteomics of DR vs NDR with label-free quantification technology successfully identified differentially expressed proteins in DR and NDR. These differential proteins are potential DR associated plasma markers and maybe a new target for early prevention and treatment of DR. DOI: 10.11855/j.issn.0577-7402.2019.01.08

糖尿病视网膜病变(DR)和糖尿病肾病(DN)是糖尿病的两个主要微血管并发症,严重影响患者的生活质量.糖尿病患者同时伴有DN的情况普遍,本研究应用Labelfree蛋白质组定量技术,筛选DR伴DN患者血浆中差异表达蛋白,结合生物信息学分析,寻找DR、DN共病的机制和共同的干预靶点. 1 对象与方法 1.1 研究对象 本研究经解放军第903医院伦理委员会审查批准.选择2型糖尿病患者14例,其中8例伴DR和DN(DNDR组),6例不伴DN和DR(DM组),两组间患者的年龄、糖化血红蛋白、病程的差异均无统计学意义(P值均＞0.05).所有患者均符合1999年WHO糖尿病诊断标准,血糖控制稳定.