视网膜母细胞瘤的遗传学特征及临床筛查与治疗

视网膜母细胞瘤(RB)是儿童最常见的眼内恶性肿瘤,多发于婴幼儿,可损害患儿视力、眼球,甚至危及生命。其发生和发展与基因组和表观基因组有很大关系,遗传性RB占所有病例的45%。RB的治疗和预后取决于疾病的初始阶段,当疾病被早期诊断和治疗时,存活率超过95%,当RB在晚期发生眼外播散时,存活率低于50%。因此RB的治疗重点是通过早期肿瘤的监测和预防挽救患儿生命,其次要目标是保存眼球,并最大水平地保留视力。对RB遗传学的研究有助于通过基因筛查、监测查找特异性靶点,对提高患儿的生存率、研究RB靶向治疗十分必要。     

儿童弱视715眼治疗效果的临床分析

目的：探讨儿童弱视临床特点和综合治疗效果。

方法：选取2015-07/2017-07我院确诊收治的弱视儿童393例715眼,建立弱视治疗档案,给予弱视综合疗法治疗,观察其治疗效果,并对其年龄、弱视类型、弱视程度、弱视注视性质和治疗依从性等影响疗效的临床特点进行分析。

结果：患儿393例715眼中,基本治愈者520眼(72.7%),进步者117眼(16.4%),无效者 78眼(10.9%),总有效率为89.1%。年龄、弱视类型、弱视程度、注视性质、治疗依从性等临床特点影响弱视治疗效果。

结论：儿童弱视的总体治疗效果较好。与患儿年龄、弱视类型、弱视程度、弱视注视性质和治疗依从性等临床特点关系密切。年龄偏大,弱视类型为屈光参差性弱视、斜视性弱视,弱视程度重、旁中心注视及依从性差的患儿治疗效果差。     

Molecular analysis of emerging radiation leukemia virus variants of C57BL/Ka mice

Molecular cloning of several primary or passaged RadLV variants and their biological characterization has allowed us to propose a model of their emergence following X-ray irradiation of C57BL/6 mouse.     

FGF‐1/‐3/FGFR4 signaling in cancer‐associated fibroblasts promotes tumor progression in colon cancer through Erk and MMP‐7

Cancer‐associated fibroblasts (CAFs), as the activated fibroblasts in the tumor stroma, are important modifiers of tumour progression. In the present study, we observed that azoxymethane and dextran sodium sulfate treatments induced increasingly severe colorectal mucosal inflammation and the intratumoural accumulation of CAFs. Fibroblast growth factor (FGF)‐1 and FGF‐3 were detected in infiltrating cells, and FGFR4, the specific receptor for FGF‐1 and FGF‐3, was detected in colon cancer tissues. The phosphorylation of FGFR4 enhanced the production of metalloproteinase (MMP)‐7 and mitogen‐activated protein kinase kinase (Mek)/extracellular signal‐regulated kinase (Erk), which was accompanied by excessive vessel generation and cell proliferation. Moreover, we separated CAFs, pericarcinoma fibroblasts (PFs), and normal fibroblasts (NFs) from human colon tissue specimens to characterize the function of CAFs. We observed that CAFs secrete more FGF‐1/‐3 than NFs and PFs and promote cancer cell growth and angiogenesis through the activation of FGFR4, which is followed by the activation of Mek/Erk and the modulation of MMP‐7 expression. The administration of FGF‐1/‐3‐neutralizing antibodies or the treatment of cells with FGFR4 siRNA or the FGFR4 inhibitor PD173074 markedly suppressed colon cancer cell proliferation and neovascularization. These observations suggest a crucial role for CAFs and FGF signaling in the initiation and progression of colorectal cancer. The inhibition of the FGF signaling pathway may be a useful strategy for the treatment of colon cancer.     

胸段食管癌IGRT分次内误差对靶区剂量的影响

目的 使用CBCT测量食管癌IGRT分次内误差,评价其对靶区和周围OAR剂量的影响。方法 应用CBCT采集 23例胸段食管癌放疗前后摆位图像,获取分次内误差。将分次内误差模拟治疗计划2和计划3,与原始计划1比较,分析其对靶区和OAR剂量学影响,并用单因素方差分析和配对t检验。结果 胸上段、胸中段、胸下段食管癌IGRT分次内平均误差在左右方向分别为(1.2±1.5)、(1.0±1.0)、(1.0±1.0) mm (P=0.138),在上下方向分别为(1.2±1.0)、(1.1±1.0)、(1.2±1.0) mm (P=0.656),在前后方向分别为(1.3±1.1)、(1.2±1.0)、(0.8±0.7) mm (P=0.003)。全部患者3 mm内误差发生频率在左右、上下、前后方向分别占95.2%、94.5%、93.9%。计划3与计划1相比,GTV V_100%下降5.55%,有 3例患者PTV D_95%下降超过原始计划处方的5%。计划3中全肺 V₃₀为(15.24±2.24)%,低于计划1的(15.67±2.28)%(P=0.033)。计划2中 4例脊髓>4500 cGy (D_max为4517.2 cGy),计划3中 19例脊髓>4500 cGy (D_max为5045.2 cGy)。结论 IGRT分次内误差对患者靶区剂量分布有一定影响。脊髓为串行器官,对分次内误差相对敏感,可能会导致部分患者脊髓超过最大耐受量。     

泻肺利水法对冠心病心力衰竭患者心衰超声指数的影响

目的:评价泻肺利水法组方的心衰合剂对冠心病心力衰竭患者心衰超声指数的影响。方法:采用随机对照研究,选取2016年11月至2017年12月首都医科大学附属北京中医医院心血管科住院的冠心病HF、心功能NYHAⅢ-Ⅳ级患者60例,分为观察组和对照组,每组30例,观察组予西医常规治疗加用心衰合剂,对照组予西医常规治疗。疗程14 d。观察2组患者治疗前后中医证候积分和心衰超声指数积分。结果:治疗后2组患者中医证候积分均较治疗前显著改善(P0.05);组间比较,观察组较对照组改善更为显著(P0.05)。治疗后观察组的心室充盈、左室射血分数、肺动脉收缩压、房室重构、心衰超声指数积分均较治疗前改善(P0.05);治疗后对照组的心室充盈、左室射血分数、肺动脉收缩压、心衰超声指数积分较治疗前提高(P0.05)。治疗后观察组的瓣膜反流、肺动脉收缩压、心衰超声指数较对照组改善更明显(P0.05)。结论:在西医规范治疗基础上联合使用泻肺利水法组方的心衰合剂,可以改善冠心病心力衰竭患者的中医证候积分,改善患者的心衰超声指数。     

可弯曲电子胸腔镜在不明原因胸水患者诊断中的应用价值（附121例）

目的:探讨尖端可弯曲内科胸腔镜检查术对未明原因的胸腔积液患者的诊断价值。方法:回顾性分析121例不明原因胸腔积液患者的胸腔镜检查资料。结果:121例患者胸膜病检结果:结核性54例（44.6%）,肿瘤46例（38.0%）,非特异性炎症17例（14.0%）,感染性3例（2.5%）,不明原因1例（0.8%）。术后胸膜活检部位少量出血2例,轻度疼痛118例,无其他严重并发症。结论:内科胸腔镜检查术可以明确胸腔积液患者的病因,简单安全,费用不高。     

肌萎缩侧索硬化的临床诊治进展

肌萎缩侧索硬化（amyotrophic lateral sclerosis,ALS）是一种致死性神经系统变性疾病,主要累及锥体束、脑干和脊髓前角运动神经元,临床表现为进行性加重的肌肉萎缩、无力及延髓麻痹,最终因呼吸衰竭死亡。随着研究的深入,ALS除运动症状外,其非运动症状也越来越受到重视。尽管目前该病的诊断主要基于病史、临床表现和电生理,辅助检查仅作为协助排除临床表现相似的其他疾病,遗传学和生物标记物的研究进展为ALS的早期诊断、病理生理机制的揭示、疾病进展及预后的评估提供了客观的参考。虽然目前ALS的治疗药物并不能有效阻止疾病的进展,然而以基因修饰及干细胞移植为代表的新的治疗方法在未来的治疗中显示了巨大的应用前景。     

急性髓系白血病Ki-67的表达及其临床意义

目的：探讨增殖相关抗原Ki-67在急性髓系白血病（AML）中的表达及其临床意义。方法选取2012年10月至2016年1月首都医科大学附属复兴医院血液内科住院AML患者45例,其中初治36例,复发9例;以20例健康志愿者作为健康对照。采用流式细胞术（FCM）检测骨髓原始细胞群Ki-67抗原的表达,分析Ki-67水平与患者临床特征的相关性。结果初治、复发AML患者和健康对照者的骨髓原始细胞群Ki-67阳性率分别为（10.38±8.41）%、（20.99±11.49）%和（40.77±11.97）%,初治和复发AML患者的Ki-67阳性率均低于健康对照者（均P＜0.05）,初治AML患者Ki-67阳性率低于复发AML患者（P＝0.006）。初治AML患者的Ki-67水平与患者的年龄、FAB亚型、白细胞计数、有无骨髓增生异常综合征（MDS）病史、乳酸脱氢酶水平、骨髓原始细胞比例、NPM1基因突变、FLT3内部串联重复（ITD）、染色体核型、诱导化疗反应均无关（均P＞0.05）。初治AML患者Ki-67高水平组与低水平组的总生存时间分别为（780±110）d和（788±118）d,两组差异无统计学意义（P＝0.927）。结论 AML患者原始细胞群增殖活性较健康对照低,检测Ki-67水平可能为临床选择细胞周期特异性化疗药物提供依据;对AML患者动态监测Ki-67水平有助于监测疾病进展,预测复发。     

Epigenetic analysis of cell-free DNA by fragmentomic profiling

Cell-free DNA (cfDNA) fragmentation patterns contain important molecular information linked to tissues of origin. We explored the possibility of using fragmentation patterns to predict cytosine-phosphate-guanine (CpG) methylation of cfDNA, obviating the use of bisulfite treatment and associated risks of DNA degradation. This study investigated the cfDNA cleavage profile surrounding a CpG (i.e., within an 11-nucleotide [nt] window) to analyze cfDNA methylation. The cfDNA cleavage proportion across positions within the window appeared nonrandom and exhibited correlation with methylation status. The mean cleavage proportion was ∼twofold higher at the cytosine of methylated CpGs than unmethylated ones in healthy controls. In contrast, the mean cleavage proportion rapidly decreased at the 1-nt position immediately preceding methylated CpGs. Such differential cleavages resulted in a characteristic change in relative presentations of CGN and NCG motifs at 5′ ends, where N represented any nucleotide. CGN/NCG motif ratios were correlated with methylation levels at tissue-specific methylated CpGs (e.g., placenta or liver) (Pearson’s absolute r > 0.86). cfDNA cleavage profiles were thus informative for cfDNA methylation and tissue-of-origin analyses. Using CG-containing end motifs, we achieved an area under a receiver operating characteristic curve (AUC) of 0.98 in differentiating patients with and without hepatocellular carcinoma and enhanced the positive predictive value of nasopharyngeal carcinoma screening (from 19.6 to 26.8%). Furthermore, we elucidated the feasibility of using cfDNA cleavage patterns to deduce CpG methylation at single CpG resolution using a deep learning algorithm and achieved an AUC of 0.93. FRAGmentomics-based Methylation Analysis (FRAGMA) presents many possibilities for noninvasive prenatal, cancer, and organ transplantation assessment.

Fragmentation patterns of cell-free DNA (cfDNA) molecules contain a wealth of molecular information related to their tissues of origin (1). For instance, compared with the background DNA molecules that are mainly derived from the hematopoietic system (2, 3), size shortening of fetal and tumoral DNA molecules occurs in the plasma DNA of pregnant women and cancer patients, respectively (4–6). In addition, a series of 10-bp periodicities were present in fetal and tumoral DNA molecules below 146 bp, with a relative reduction in the major peak at 166 bp (1). Such characteristic size profiles suggest that the fragmentation of cfDNA may be associated with nucleosome structures (5, 7). Many important characteristics pertaining to cfDNA fragmentation have been unveiled recently, such as nucleosome footprints (8, 9), fragment end motifs (10), preferred ends (7, 11), and jagged ends (12), which are examples of fragmentomic markers (1).cfDNA fragmentomics is an emergent and actively pursued area, with wide-ranging biological and clinical implications. It has been reported that the use of fragmentation patterns of cfDNA could inform the expression status of genes (13, 14). Using mouse models, DNA nucleases (e.g., DNASE1L3) were found to play important roles in the generation of plasma DNA molecules (15, 16). Fragmentomic features, such as cfDNA end motifs and jagged ends, were further demonstrated to be useful for monitoring DNA nuclease activities, providing biomarkers for autoimmune diseases (e.g., systemic lupus erythematosus) (17, 18). In addition, the deficiencies of nuclease activities in a mouse model resulted in altered DNA methylation profiles of plasma DNA molecules (19). However, how cfDNA fragmentation patterns interplay with DNA methylation in human individuals under different pathophysiological conditions, such as pregnancy and oncogenesis, and in healthy patients without nuclease deficiency, is unknown. It is also not known whether fragmentomic features can be used to deduce cfDNA methylation status.A widely employed way to assess DNA methylation is through bisulfite sequencing (20). A key limitation of this approach is the severe degradation of DNA molecules caused by the bisulfite treatment (21), which greatly increases the sampling variation when analyzing rare target molecules (e.g., tumoral cfDNA at early stages of cancer). Many efforts have been made toward overcoming this issue. For example, Vaisvila et al. developed enzymatic methyl sequencing for which DNA molecules were treated using tet methylcytosine dioxygenase 2 and T4 phage β-glucosyltransferase, followed by the apolipoprotein B mRNA editing enzyme catalytic subunit 3A (APOBEC3A) treatment. Cytosine conversion based on enzymatic processes was reported to be much less destructive (22). Recently, researchers developed approaches making use of third-generation sequencing technologies such as single-molecule real-time sequencing (Pacific Biosciences) (23) and nanopore sequencing (24) to analyze cytosine-phosphate-guanine (CpG) methylation patterns in native DNA molecules, theoretically overcoming the above-mentioned limitation. However, compared with second-generation sequencing (also called next-generation sequencing [NGS]) technologies, the throughput of third-generation sequencing technologies is generally lower and the sequencing cost per nucleotide (nt) is much higher, thus restricting its immediate application in clinical settings. Here, we explore the feasibility of enabling the assessment of DNA methylation using fragmentomic characteristics of cfDNA molecules deduced from NGS results without the use of bisulfite or enzymatic treatment. If successful, such an approach could leverage the high throughput of NGS while obviating the use of chemical/enzymatic conversion and could potentially be readily integrated into currently used NGS-based platforms for cfDNA analysis.In this study, we utilize the fragmentation patterns proximal to a CpG site for deducing its methylation status. The fragmentation pattern is depicted by the frequency of cfDNA fragment ends at each position within a certain nt range relative to a CpG of interest, termed a cleavage profile (Fig. 1). Such a cleavage profile varies according to the methylation status of the CpG site of interest, providing the basis for methylation analysis by using fragmentomic features. We further correlated two types of end motifs (CGN and NCG; N represents any nucleotide of A, C, G, or T) resulting from differential cutting in the measurement window related to DNA methylation, attempting to construct a simplified approach for methylation analysis. Modeling CpG methylation using cfDNA fragmentation may facilitate noninvasive prenatal testing, cancer detection, and tissue-of-origin analysis (Fig. 1). Furthermore, we explore the feasibility of using deep learning to deduce the methylation status at single CpG resolution through the cleavage profile (Fig. 1). We refer to this FRAGmentomics-based Methylation Analysis as FRAGMA in this study.Open in a separate windowFig. 1.Schematic for FRAGMA of cfDNA molecules. cfDNA molecules were sequenced by massively parallel sequencing and aligned to the human reference genome. The cleavage proportion within an 11-nt window (the cleavage measurement window) was used to measure the cutting preference of cfDNA molecules. The patterns of cleavage proportion within a window (the cleavage profile) depended on the methylation status of one or more CpG sites associated with that window. For example, a methylated CpG site might confer a higher probability of cfDNA cutting at the cytosine in the CpG context, but an unmethylated site might not. Such methylation-dependent differential fragmentation within a cleavage measurement window resulted in the change in CGN/NCG motif ratio. Thus, the CGN/NCG motif ratio provided a simplified version for reflecting CpG methylation, allowing cfDNA tissue-of-origin analysis of cfDNA and cancer detection. Furthermore, the great number of cleavage profiles derived from cfDNA molecules might provide an opportunity to train a deep learning model for methylation prediction at the single CpG resolution.