Isolation and characterization of new human carrier peptides from two important vaccine immunogens

In the preparation of glycoconjugate vaccines in clinical practice, two highly immunogenic carrier proteins, CRM₁₉₇ and tetanus toxoid (TT), are predominantly conjugated with the capsular polysaccharides (CPSs) of bacterial pathogens. In addition, TT has long been used as an effective vaccine to prevent tetanus. While these carrier proteins play an important role in immunogenicity and vaccine design alike, their defined human major histocompatibility complex class II (MHCII) T cell epitopes are inadequately characterized. In this current work, we use mass spectrometry to identify the peptides from these carrier proteins that are naturally processed and presented by human B cells via MHCII pathway. The MHCII-presented peptides are screened for their T cell stimulation using primary CD4+ T cells from four healthy adult donors. These combined methods reveal a subset of eleven CD4+ T cell epitopes that proliferate and stimulate human T cells with diverse MHCII allelic repertoire. Six of these peptides stand out as potential immunodominant epitopes by responding in three or more donors. Additionally, we provide evidence of these natural epitopes eliciting more significant T cell responses in donors than previously published TT peptides selected from T cell epitope screening. This study serves toward understanding carrier protein immune responses and thus enables the use of these peptides in developing novel knowledge-based vaccines to combat persisting problems in glycoconjugate vaccine design.     

乙肝肝硬化肝胆湿热及肝肾阴虚证物质基础研究＊

目的 运用代谢组学技术分析乙肝肝硬化患者肝肾阴虚及肝胆湿热两种典型证候（同病异证）的血清差异代谢产物及其代谢通路，探寻虚、实两种典型证候的内在物质基础，以期从代谢水平上为中医证候分类提供客观依据。方法 对符合纳入标准的111例不同证候的乙肝肝硬化患者（肝胆湿热证40例，肝肾阴虚证41例，隐证(无证可辨）者30例）中医症状及体征进行描述性分析，发现两种不同证型的临床信息分布规律及证候特征；采用气相色谱-飞行时间质谱联用（GC-TOF/MS）技术对乙肝肝硬化患者，以及与之相匹配的60例健康人的血清样本进行检测，经非监督的主成分分析（Principal Components Analysis，PCA）、有监督的偏最小二乘判别分析（Partial Least Square Discriminant Analysis，PLS-DA）及监督的正交偏最小二乘法（Orthogonal Partial Least SquareDiscriminant Analysis，OPLS-DA）分析，找出与乙肝肝硬化疾病本身及其两种典型证候相关的差异性物质；运用MetaboAnalyst 3.0数据库，寻找并解析肝胆湿热及肝肾阴虚虚实两种证候间差异性物质的相关代谢通路。结果 （1）肝胆湿热证中出现频率较高（50%以上）的症状为小便色黄，口干，口苦，口臭或有异味等。肝肾阴虚证中出现频率较高的症状为口干、腰酸、乏力、腿软等。两证共见症/征为口干、尿黄、易怒、舌红。（2）各组间丙氨酸氨基转移酶（Alanine Aminotransferase,ALT）数值无统计学差异（P > 0.05）；与健康组比较，隐证组中白蛋白（Albumin，ALB），肝胆湿热证中总胆红素（Total Bilirubin，TBil）、直接胆红素（Direct Bilirubin，DBil）、谷草转氨酶（Aspartate Transaminase，AST）、碱性磷酸酶（Alkaline Phosphatase，ALP）、谷氨酰转肽酶（Gamma-Glutamyl Transpeptidase，GGT）、总胆汁酸（Total Biliary Acid，TBA）及ALB，肝肾阴虚证TBil、ALP、GGT、TBA、ALB值差异均有统计学意义（P < 0.05）；与隐证比较，肝胆湿热证TBil、DBil、AST、ALP、TBA、ALB，肝肾阴虚证TBA、ALB差异有统计学意义（P < 0.05）；肝胆湿热证与肝肾阴虚证相比，TBil、DBil差异有统计学意义（P < 0.05）。（3）代谢组学检测及代谢通路分析，发现各组之间代谢谱均有良好的区分，并获得各组间的差异性物质。发现肝胆湿热及肝肾阴虚两典型证的共同物质10个，去除疾病（隐证）的信息，则得到两证共同物质6个，涉及的代谢通路为甘氨酸、丝氨酸及苏氨酸代谢和苯丙氨酸代谢；同时，分别获得两证各自特异性的代谢物质各8个，分别涉及亚油酸代谢和甘氨酸、苏氨酸及丝氨酸代谢。结论 运用代谢组学技术，发现肝胆湿热及肝肾阴虚不同证之间既存在病的共同物质（同病），也存在证的差异物质（异证），从而在代谢层面上为中医证候分类的科学性提供科学依据。     

The activity prediction of indole inhibitors against HCV NS5B polymerase

Non‐structural viral protein 5B (NS5B) is a viral protein in hepatitis C virus. Although various inhibitors against NS5B have been found, the activity prediction of similar untested inhibitors is still highly desirable. In this respect, the Tchebichef moments (TMs) calculated from the images of molecular structures were regarded as the independent variables while the inhibitory activity (pIC₅₀) was the dependent variable, and the predictive model was established by means of stepwise regression. The R‐squared of leave‐one‐out cross‐validation (Q²) for the training set and the R‐squared of prediction () for external independent test set were 0.919 and 0.927, respectively. The obtained model was also evaluated strictly. Compared with the multivariate curve resolution with alternating least squares (MCR‐ALS) and the QSAR approaches derived from the literature, the proposed method is more accurate and reliable. This study not only provides an effective approach to predict the biological activity of RNA replication's inhibitors, but also extends the QSAR modeling technique.     

PELP1 signaling contributes to medulloblastoma progression by regulating the NF-κB pathway

Medulloblastoma (MB) is the most common and deadliest brain tumor in children. Proline-, glutamic acid-, and leucine-rich protein 1 (PELP1) is a scaffolding protein and its oncogenic signaling is implicated in the progression of several cancers. However, the role of PELP1 in the progression of MB remains unknown. The objective of this study is to examine the role of PELP1 in the progression of MB. Immunohistochemical analysis of MB tissue microarrays revealed that PELP1 is overexpressed in the MB specimens compared to normal brain. Knockdown of PELP1 reduced cell proliferation, cell survival, and cell invasion of MB cell lines. The RNA-sequencing analysis revealed that PELP1 knockdown significantly downregulated the pathways related to inflammation and extracellular matrix. Gene set enrichment analysis confirmed that the PELP1-regulated genes were negatively correlated with nuclear factor-κB (NF-κB), extracellular matrix, and angiogenesis gene sets. Interestingly, PELP1 knockdown reduced the expression of NF-κB target genes, NF-κB reporter activity, and inhibited the nuclear translocation of p65. Importantly, the knockdown of PELP1 significantly reduced in vivo MB progression in orthotopic models and improved the overall mice survival. Collectively, these results suggest that PELP1 could be a novel target for therapeutic intervention in MB.     

急性乙型肝炎患者pDCs上CD86表达变化及意义

目的 探讨外周血浆细胞样树突状细胞(pDCs)在急性乙型肝炎(AHB）患者临床转归中的变化及作用。方法 纳入2015年6月至2017年5月收治的急性乙型肝炎(AHB组)患者40例， 给予退黄、降酶、保肝等药物，并加用恩替卡韦0．5 mg空腹口服，1次/d。另纳入健康体检者26例为健康对照组（ HC组）。HC组于体检当天、AHB组于治疗前及治疗后6周采集外周静脉全血，采用ELISA法检测2组血浆HBV DNA、HBsAg、HbeAg及肝功能指标水平，并应用流式细胞仪检测pDCs的频数及其功能分子CD86的表达水平。结果 治疗6周后，AHB组临床症状显著缓解，血清学指标和乙型肝炎标志物水平显著下降（P<0.01）。治疗6周后，AHB组HBV DNA转阴率为82.5%，HBsAg清除率为72.5%，HBeAg血清学转换率为75.0%，与治疗前比较差异均有统计学意义（P<0.01）, HC组pDCs频数显著高于AHB组治疗前。AHB组治疗后CD86+pDCs显著高于治疗前。HC组CD86ABC显著低于AHB组治疗前。结论 在急性乙型肝炎患者外周血pDCs的数量降低。急性乙型肝炎患者在治疗后CD86+pDCs增多。急性乙型肝炎患者pDCs上CD86表达上调。     

Neurofibromatosis type 2 in Phelan-McDermid syndrome: Institutional experience and review of the literature

Phelan-McDermid syndrome (PMS) is a rare neurodevelopmental disorder caused by rearrangements on chromosome 22q13.3 or sequence variants in SHANK3. Individuals with PMS caused by a 22q terminal deletion and a ring chromosome are at increased risk for Neurofibromatosis type 2 (NF2). However, the prevalence of NF2 in individuals with PMS and a r (22) is unknown.Individuals with PMS and a r (22) chromosome evaluated at the Greenwood Genetic Center (GGC) or by international collaborators, or identified through the PMS International Registry (PMSIR) were contacted and participated in a clinical questionnaire. Forty-four families completed the questionnaire and consented for the study. Of the individuals with a r (22), 7 (16%) carried a diagnosis of NF2. The average age of diagnosis of r (22) was 18 years old in individuals with NF2 and three years old in individuals without NF2 (p-value <0.001). Clinical findings were similar among all individuals in our sample with the exception of hearing loss, present in 57% of individuals with NF2 and 8% of individuals without NF2 (p-value <0.01).This is the largest clinical report of individuals with PMS and a r (22) chromosome. We show a diagnosis of NF2 in individuals with r (22) is not uncommon and may be under ascertained. Moreover, the presentation of NF2 in this cohort is variable and lifelong routine screening for features of NF2 in this population should be considered.     

A single dose glycoprotein D-based subunit vaccine against pseudorabies virus infection

Pseudorabies Virus (PRV) is the causative agent of Pseudorabies (PR), also known as Aujeszky’s Disease, one of the most important infectious diseases in swine, resulting in huge economic losses to the swine industry globally. The emergence of mutant PRV strains after 2011 resulted in a sharp decrease in the efficacy of available commercial vaccines. To develop a more effective vaccine that can prevent the spread of PRV, glycoprotein B (gB), glycoprotein C (gC) and glycoprotein D (gD) from recent PRV isolates were expressed in a baculovirus system and their protective efficacy was tested in mice and piglets. Neutralizing antibody titers (NAs) in mice vaccinated with gB, gC and gD peaked at 28 days after immunization and then slowly declined. NAs in the mice immunized with gD were remarkably higher than other groups. After a lethal challenge of 5 LD₅₀ with mutant PRV-HNLH strain, the survival rates of gB and gD were 100% and 87.5% respectively, which was significantly higher than gC group (50%). Piglets vaccinated with the gD and gB + D vaccines developed the highest NAs 7 days post immunization. No piglets in these two groups exhibited clinical symptoms, high body temperature or virus shedding following challenge with 10^6.6 TCID₅₀ with the mutant PRV-HNLH strain. Histopathology and immunohistochemistry showed remarkably reduced pathological damage and viral loads in gD and gB + D groups. Furthermore, the duration of the NAs induced by gD vaccine could maintain as long as four months after a single dose. The current study indicates that a gD-based vaccine could be developed for the efficient control of PRV.     

Benign breast disorders

Benign breast disorders encompass a wide range of presentations and account for approximately 90% of breast outpatient referrals. Although the conditions discussed in this article are labelled ‘benign’, they can cause much anguish for the patient. A specialist breast clinician frequently has the privilege of being able to reassure patients in a single trip to the breast department and unburden them from their worry. One-stop breast clinics were designed to ensure that patients with worrying symptoms are promptly seen to aide early detection of breast cancer and optimize patient outcomes. The diagnostic multidisciplinary team consists of a specialist breast clinician, breast radiologist, pathologist and breast care nurses. Most patients with benign breast disorders can be reassured, with surgery limited to a few to aide definite diagnosis or as a last resort when conservative measures have failed. This is a comprehensive overview of benign breast disorders and covers all that is required for the MRCS examination and serves as a revision aide for FRCS candidates.