Future directions in managing aniridia-associated keratopathy

Congenital aniridia is a panocular disorder that is typically characterized by iris hypoplasia and aniridia-associated keratopathy (AAK). AAK results in the progressive loss of corneal transparency and thereby loss of vision. Currently, there is no approved therapy to delay or prevent its progression, and clinical management is challenging because of phenotypic variability and high risk of complications after interventions; however, new insights into the molecular pathogenesis of AAK may help improve its management. Here, we review the current understanding about the pathogenesis and management of AAK. We highlight the biological mechanisms involved in AAK development with the aim to develop future treatment options, including surgical, pharmacological, cell therapies, and gene therapies.     

核因子κB受体活化因子配体和肿瘤坏死因子α经炎性牙周膜干细胞外泌体促进破骨细胞分化

目的　慢性牙周炎表现的病理性骨吸收非常常见，牙周膜干细胞（PDLSCs）的外泌体（Exo）对骨吸收的作用和影响尚不明确，本研究分析了该Exo蛋白组分对破骨细胞分化的影响。方法　分别从正畸前拔牙患者和慢性牙周炎患者的牙周膜组织中提取PDLSCs，通过流式细胞术检测表面标记物；通过差速离心分别提取2种细胞的Exo，即Exo-WT和Exo-CP，并通过Western blot、透射电子显微镜（TEM）、蛋白浓度检测、纳米粒径追踪检测Exo特征；通过蛋白质谱检测2种Exo的蛋白组分；通过酶联免疫吸附（ELISA）验证了差异表达蛋白肿瘤坏死因子α（TNF-α）、核因子κB受体活化因子配体（RANKL）、白细胞介素（IL）-1α、转化生长因子β（TGF-β）、骨形态发生蛋白2（BMP-2）表达水平；将10、100、1 000 μg·mL^-1的Exo-CP或Exo-WT加入RAW264.7培养基中并于5 d时通过实时荧光定量聚合酶链反应（RT-qPCR）、Western blot、抗酒石酸酸性磷酸酶（TRAP）染色检测破骨分化相关指标表达情况；采用SPSS 24.0软件对实验数据进行统计学分析。结果　Exo-CP富集的差异表达蛋白主要与破骨细胞分化的TNF信号通路、核转录因子κB（NF-κB）信号通路相关；ELISA实验证实了Exo-CP中高表达TNF-α、RANKL、IL-1α，低表达TGF-β1、BMP-2（P<0.05）；Exo-CP作用于RAW264.7，显著提高了细胞的破骨分化相关基因及蛋白的表达水平，TRAP染色可见分化的破骨细胞，且呈现浓度依赖性，100、1 000 μg·mL^-1浓度Exo-CP对破骨细胞分化的促进作用显著高于10 μg·mL^-1浓度组（P<0.05）。结论　慢性牙周炎的病理性骨吸收可能由炎性PDLSCs所分泌的Exo通过促进破骨细胞分化引起，Exo中主要的作用蛋白可能为RANKL和TNF-α，本研究为慢性牙周炎骨吸收的发病机制提供了新视角。     

Methodological advances in the design of peptide-based vaccines

The tremendous advances in genomics, recombinant DNA technology, bioengineering and nanotechnology, in conjunction with the development of high-end computations, have been instrumental in the process of rational design of peptide-based vaccines. The use of peptide vaccines was limited owing to their inherent instability when systemically administered; however, advanced formulation techniques have been developed for their systemic delivery, thereby overcoming their degradation, clearance, cellular uptake and off-target effects. With the rise of sophisticated immunological predictors and experimental techniques, several methodological advances have occurred in this field. This review examines contemporary methods to identify and optimize epitopes, engineer their immunogenic properties and develop their safe and efficient delivery into the host.     

新化合物Elixir-X对肝切及辐射所致衰老小鼠的抗衰老作用

目的 探讨蜂巢提取物Elixir-X对自然衰老及辐射衰老小鼠的影响，为Elixir-X的进一步研发提供药理学基础。方法 采用小鼠肝脏切除术建立自然衰老小鼠模型，将其分为空白对照组、Elixir-X组、白藜芦醇组及模型组，连续给药7 d后，检测小鼠70%肝脏切除后的再生过程中，衰老细胞、肝脏功能恢复与肝脏再生的变化，观察Elixir-X对自然衰老的影响；采用60CO辐射法建立辐射衰老小鼠模型，将其分为模型组、空白对照组、白藜芦醇组、Elixir-X高、低剂量组，通过检测小鼠肠隐窝干细胞再生、小肠绒毛结构以及衰老细胞的变化，对比不同剂量Elixir-X对辐射后衰老的影响。结果 在肝脏切除术的小鼠模型中，Elixir-X组的肝功能水平明显低于模型组的肝脏自然再生小鼠，显著促进了肝功能恢复（P＜0.05）；同时与模型组相比，Elixir-X组清除了肝脏切除后再生过程中产生的衰老细胞（P＜0.05），Ki-67的着色斑块明显增加，促进肝脏的再生，且肝血窦的扫描电镜影像结果中Elixir-X组肝血窦再生加快。60CO辐照小鼠模型中，与模型组相比，Elixir-X组有效促进辐照后肝脏功能恢复（P＜0.05），Elixir-X高剂量组体重也恢复到正常水平，与空白对照组无显著差异；与模型组相比，Elixir-X低剂量组小肠的β-Gal着色细胞减少，Elixir-X高剂量组甚至能更有效地减少衰老细胞（P＜0.05）；小肠HE染色的结果中，Elixir-X组小肠绒毛结构得到恢复；Ki-67的IHC染色中，与模型组相比，Elixir-X组着色板块更多，且着色区域都在肠隐窝干细胞位置。结论 Elixir-X能有效清除肝脏切除术引起的自然衰老与辐射引起的损伤性衰老细胞，对衰老具有显著地抑制作用。     

Newly recruited intraepithelial Ly6A+CCR9+CD4+ T cells protect against enteric viral infection

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Semi-perfusion cultures of suspension MDCK cells enable high cell concentrations and efficient influenza A virus production

Control and prevention of rapid influenza spread among humans depend on the availability of efficient and safe seasonal and pandemic vaccines, made primarily from inactivated influenza virus particles. Current influenza virus production processes rely heavily on embryonated chicken eggs or on cell culture as substrate for virus propagation. Today’s efforts towards process intensification in animal cell culture could innovate viral vaccine manufacturing using high-yield suspension cells in high cell density perfusion processes. In this work, we present a MDCK cell line adapted to grow as single cell suspension with a doubling time of less than 20 h, achieving cell concentrations over 1 × 10⁷ cells/mL in batch mode. Influenza A virus titer obtained in batch infections were 3.6 log₁₀(HAU/100 µL) for total- and 10⁹ virions/mL for infectious virus particles (TCID₅₀), respectively. In semi-perfusion mode concentrations up to 6 × 10⁷ cells/mL, accumulated virus titer of 4.5 log₁₀(HAU/100 µL) and infectious titer of almost 10¹⁰ virions/mL (TCID₅₀) were possible. This exceeds results reported previously for cell culture-based influenza virus propagation by far and suggests perfusion cultures as the preferred method in viral vaccine manufacturing.