罗非昔布对胃癌细胞上清液中培养的树突状细胞分化成熟和功能的影响

目的：从免疫学角度分析罗非昔布（Rofecoxib）抗癌作用的机制。方法：将人外周血分离的单核细胞加入含重组人粒细胞巨噬细胞集落刺激因子和重组人白细胞介素的培养液中,随机分为空白组（PBS培养液）,干预组（胃癌细胞上清液＋罗非昔布,150mg/L）,对照组（胃癌细胞上清液）使其分化为树突状细胞（denstritic cell,DC）。混合培养48h后,动态观察DC成熟过程中的形态学变化,流式细胞术检测DC成熟表型（CD40、CD80）,混合T淋巴细胞增殖反应检测DC功能。结果：与对照组比较,空白组、干预组DC的CD40、CD80表达明显在增加（P〈0.05）,T细胞增殖作用明显增强（P〈0.05）。结论：罗非昔布可通过促进DC成熟,并增加DC的免疫活性而发挥抗癌作用。     

Effects of different probiotic strains of Lactobacillus and Bifidobacterium on bacterial translocation and liver injury in an acute liver injury model

Septic complications represent frequent causes of morbidity in liver diseases and following hepatic operations. Most infections are caused by the individual own intestinal microflora. The intestinal microflora composition is important in physiological and pathophysiological processes in the human gastrointestinal tract, but their influence on liver in different situations is unclear. We therefore studied the effect of different Lactobacillus strains and a Bifidobacterium strain on the extent of liver injury, bacterial translocation and intestinal microflora in an acute liver injury model.Sprague–Dawley rats were divided into five groups: acute liver injury control, acute liver injury+B. animalis NM2, acute liver injury+L. acidophilus NM1, acute liver injury+L. rhamnosus ATCC 53103, and acute liver injury+L. rhamnosus DSM 6594 and L. plantarum DSM 9843. The bacteria were administered rectally daily for 8 days. Liver injury was induced on the 8th day by intraperitoneal injection of -galactosamine (1.1 g/kg BW). Samples were collected 24 h after the liver injury. Liver enzymes and bilirubin serum levels, bacterial translocation (to arterial and portal blood, liver and mesenteric lymph nodes (MLNs)), and intestinal microflora were evaluated.L. acidophilus NM1; L. rhamnosus ATCC 53103, and L. rhamnosus DSM 6594+L. plantarum DSM 9843 decreased bacterial translocation compared to the liver injury control group. B. animalis NM2 increased bacterial translocation to the mesenteric lymph nodes. The levels of alanine aminotransferase (ALAT) were significantly lower in the L. acidophilus, L. rhamnosus ATCC 53103, L. rhamnosus DSM 6594+L. plantarum DSM 9843 groups compared to the liver injury group. The L. rhamnosus and L. rhamnosus+L. plantarum groups significantly reduced ALAT levels compared to the B. animalis group. All administered bacteria decreased the Enterobacteriaceae count in the cecum and colon.Administration of different lactobacilli and a Bifidobacterium strain in an acute liver injury rat model, has shown different effects on bacterial translocation and hepatocellular damage. L. acidophilus, L. rhamnosus, and L. rhamnosus+L. plantarum reduced bacterial translocation and hepatocellular damage. B. animalis NM2 increased bacterial translocation to the mesenteric lymph nodes and did not affect hepatocellular damage.     

TP53 Targeted Deep Sequencing of Cell-Free DNA in Esophageal Squamous Cell Carcinoma Using Low-Quality Serum: Concordance with Tumor Mutation

Circulating cell-free DNA (cfDNA) is emerging as a potential tumor biomarker. CfDNA-based biomarkers may be applicable in tumors without an available non-invasive screening method among at-risk populations. Esophageal squamous cell carcinoma (ESCC) and residents of the Asian cancer belt are examples of those malignancies and populations. Previous epidemiological studies using cfDNA have pointed to the need for high volumes of good quality plasma (i.e., >1 mL plasma with 0 or 1 cycles of freeze-thaw) rather than archival serum, which is often the main available source of cfDNA in retrospective studies. Here, we have investigated the concordance of TP53 mutations in tumor tissue and cfDNA extracted from archival serum left-over from 42 cases and 39 matched controls (age, gender, residence) in a high-risk area of Northern Iran (Golestan). Deep sequencing of TP53 coding regions was complemented with a specialized variant caller (Needlestack). Overall, 23% to 31% of mutations were concordantly detected in tumor and serum cfDNA (based on two false discovery rate thresholds). Concordance was positively correlated with high cfDNA concentration, smoking history (p-value = 0.02) and mutations with a high potential of neoantigen formation (OR; 95%CI = 1.9 (1.11–3.29)), suggesting that tumor DNA release in the bloodstream might reflect the effects of immune and inflammatory context on tumor cell turnover. We identified TP53 mutations in five controls, one of whom was subsequently diagnosed with ESCC. Overall, the results showed that cfDNA mutations can be reliably identified by deep sequencing of archival serum, with a rate of success comparable to plasma. Nonetheless, 70% non-identifiable mutations among cancer patients and 12% mutation detection in controls are the main challenges in applying cfDNA to detect tumor-related variants when blindly targeting whole coding regions of the TP53 gene in ESCC.     

细支气管肺泡癌的光镜及电镜研究

目的：探讨细支气管肺泡癌的组织起源及亚型分类，方法：选择符合WHO肺肿瘤组织学分型标准的细支气管肺泡癌30例，进行光镜及电镜观察。结果：30例细支气管肺泡癌（其中14例行电镜检查）中，10例为粘液型，18例为非粘液型，2例为混合性粘液与非粘液细胞（中间细胞）型；电镜检查观察到粘液细胞，Clara细胞及Ⅱ型肺泡细胞的超微结构特征。结论：细支气管肺泡癌可起源于粘液细胞、Clara细胞及Ⅱ型肺泡细胞，组织学分型宜分为粘液型、非粘液型及混合型（中间细胞型）三种亚型。     

从噬菌体抗体库中筛选出抗原结合活性相近、表达水平不同的单链抗体

从抗人膀胱癌改形噬菌体抗体库中筛选出一组相对亲和力常数和细胞结合活性相近的单链抗体，而它们的结合活性均明显高于起始的鼠源单链抗体。在亲和力淘选的过程中，这一组抗体的表达量存在显著差异，这在一定程度上影响到筛选过程中ELISA鉴定的结果，使整个筛选过程并不仅仅是对高亲和力抗体的筛选，也同时得到了高表达量的抗体。     

The Effects of the CCR6/CCL20 Biological Axis on the Invasion and Metastasis of Hepatocellular Carcinoma

Chemokines and their receptors have recently been shown to play major roles in cancer metastasis. Chemokine receptor 6 (CCR6) and its ligand, CCL20, were highly expressed in a variety of human cancers. In our present study, we aimed to clarify whether CCR6/CCL20 was correlated with the migration of hepatocellular carcinoma (HCC). RT-PCR and Western blot results showed that CCR6 was overexpressed in different invasive potential HCC cell lines (p < 0.05), while the expression of CCL20 had no obvious difference (p > 0.05). CCR6 was suppressed by siRNA in HCCLM6, and then the biological behaviors of HCCLM6 cells were observed. The results showed that the CCR6/CCL20 biological axis increased the capacity of proliferation and adhesion, as well as the chemotactic migration and the level of cytokines related to degraded extracellular matrix. In conclusion, these findings indicate that CCR6 indeed participates in regulating the migration and invasion of HCC, and it might become a prognostic factor of HCC.     

In vivo and in vitro evaluation of the cytotoxic effects of Photosan-loaded hollow silica nanoparticles on liver cancer

This study aimed to compare the inhibitory effects of photosensitizers loaded in hollow silica nanoparticles and conventional photosensitizers on HepG2 human hepatoma cell proliferation and determine the underlying mechanisms. Photosensitizers (conventional Photosan-II or nanoscale Photosan-II) were administered to in vitro cultured HepG2 hepatoma cells and treated by photodynamic therapy (PDT) with various levels of light exposure. To assess photosensitizers'' effects, cell viability was determined by 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. In addition, apoptotic and necrotic cells were measured by flow cytometry and the expression of caspase-3 and caspase-9 evaluated by western blot. Finally, the in vivo effects of nanoscale and conventional photosensitizers on liver cancer were assessed in nude mice. Nanoscale Photosan-II significantly inhibited hepatoma cell viability in a concentration-dependent manner and this effect was more pronounced with high laser doses. Moreover, nanoscale photosensitizers performed better than the conventional ones under the same experimental conditions (p < 0.05). Flow cytometry data demonstrated that laser-induced cell death was markedly increased after treatment with nanoscale Photosan-II in comparison with free Photosan-II (p < 0.05). Activated caspase-3 and caspase-9 levels were significantly higher in cells treated with Photosan-II loaded in silica nanoparticles than free Photosan-II (p < 0.05). Accordingly, treatment with nanoscale photosensitizers resulted in improved outcomes (tumor volume) in a mouse model of liver cancer, in comparison with conventional photosensitizers. Hollow silica nanoparticles containing photosensitizer more efficiently inhibited hepatoma cells than photosensitizer alone, through induction of apoptosis, both in vivo and in vitro.     

Simultaneous induction of calcium transients in embryoid bodies using microfabricated electrode substrates

Precise control of differentiation processes of pluripotent stem cells is a key component for the further development of regenerative medicine. For this purpose, combining a cell-aggregate-size treatment for regulating intercellular signal transmissions and an electrical stimulation technique for inducing cellular responses is a promising approach. In the present study, we developed microfabricated electrode substrates that allow simultaneous stimulation of embryoid bodies (EBs) of P19 cells. Mouse embryonal carcinoma P19 cells can be induced to differentiate into three germ layers and serve as a promising stem cell model. Microcavity–array patterns were fabricated onto indium–tin–oxide (ITO) substrates using a standard photo-lithography technique, and uniform-sized EBs of P19 cells were inserted into each microcavity. Electrical stimulation was applied to the EBs through substrate electrodes and stimulus-induced intracellular calcium transients were monitored. We confirmed that the developed electrode device could simultaneously stimulate smaller (200 μm diameter) and larger (500 μm diameter) EBs inserted in the microcavities and induce specific spatio-temporal patterns of intracellular calcium transients in the EBs with fine reproducibility. We concluded that the developed microcavity array with embedded electrodes could simultaneously and effectively stimulate uniform-sized EBs inserted in it. Therefore, it is a promising experimental tool for precisely controlling cell differentiation processes.