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Yu e Xue,1,* Dongmei Zhang,1,* Shuaixian Du,2,* Du Chen,3,* Shihan Liu,2 Tianfeng Peng,4 Chong Li,5 Jianchu Zhang,1 Xiaorong Wang1 1Department of Respiratory and Critical Care Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, People’s Republic of China; 2Department of Clinical Laboratory, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, People’s Republic of China; 3Department of Neurology, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong, People’s Republic of China; 4Emergency Department, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, People’s Republic of China; 5Gastrointestinal Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, People’s Republic of China*These authors contributed equally to this workCorrespondence: Xiaorong Wang; Jianchu Zhang, Department of Respiratory and Critical Care Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Number 1277, Jie Fang Rode, Wuhan, Hubei, 430022, People’s Republic of China, Tel + 86-27-85726707, Email rong-100@163.com; zsn0928@163.comPurpose: To investigate the molecular epidemiology and risk factors of carbapenem-resistant Klebsiella pneumoniae (CRKP) infection.Patients and Methods: Patient’s clinical data and CRKP strains were collected from November 2017 to December 2018 at a tertiary hospital in Wuhan, China. The antimicrobial susceptibilities, carbapenem-resistant genes, multi-locus sequence typing (MLST), homologous analysis, and risk factors for CRKP were determined.Results: A total of 203 CRKP strains were isolated, and 98.5% (200/203) of patients were nosocomially infected. The mortality rate was 17.7% (36/203). All 203 strains were confirmed as carbapenemases -producing strains. The most predominant carbapenemase gene was blaIMP-4 (81.3%, 165/203), followed by blaKPC-2 (25.1%, 51/203) and blaNDM-1 (23.2%, 47/205). Of the 203 strains, 28 (13.8%) had both blaKPC-2 and blaIMP-4 genes, 23 (11.3%) had both blaIMP-4 and blaNDM-1 genes, 20 (9.9%) had blaKPC-2, blaIMP-4 and blaNDM-1 three genes. MLST analysis showed that there were 48 ST typologies (including 7 new STs), of which ST-11 was the most prevalent (59.6%, 121/203). Phylogenetic analysis showed that 203 CRKP isolates came from 7 clusters and exhibited a strong correlation with the isolation source. eBURST analyses indicated that CRKP isolates have undergone different evolutionary processes. Patients with ST-11 CRKP underwent more mechanical ventilation (50% vs 32.9%, P=0.020) and gastric catheterization (15.7% vs 6.1%, P=0.042) within 3 months before sample collection, and also had higher drug-resistance rate than non-ST-11 CRKP. Comparing with CSKP (carbapenem-sensitive Klebsiella pneumoniae), gastrointestinal disease (odds ratio [OR]=6.168, P=0.003), nosocomial infection (OR=5.573, P=0.012), antibiotic exposure (OR=4.131, P=0.004), urinary catheterization (OR=3.960, P=0.031) and venous/arterial catheterization (OR=2.738, P=0.026) within the preceding 3 months were independent risk factors for CRKP infection.Conclusion: The IMP-4 was the most predominant carbapenemase and blaIMP-4 bearing Klebsiella pneumoniae ST-11 was spreading in the hospital. Nosocomial infections, antibiotic exposure, and urinary and venous/arterial catheterization within 3 months were the risk factors for developing CRKP infection.Keywords: Klebsiella pneumoniae, carbapenemase, multi-locus sequence typing, MLST, ST-11, IMP-4

Huasheng Huang,1– 3 Shayong Peng,1– 3 Yongguang Wei,1– 3 Chenlu Lan,1– 3 Wei Qin,1– 3 Xiwen Liao,1– 3 Cheng-Kun Yang,1– 3 Guangzhi Zhu,1– 3 Xin Zhou,1– 3 Tao Peng1– 3 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Zhuang Autonomous Region, People’s Republic of China; 2Guangxi Key Laboratory of Enhanced Recovery After Surgery for Gastrointestinal Cancer, Nanning, People’s Republic of China; 3Key Laboratory of Early Prevention & Treatment for Regional High Frequency Tumor (Guangxi Medical University), Ministry of Education, Nanning, Guangxi Zhuang Autonomous Region, People’s Republic of ChinaCorrespondence: Tao Peng; Xin Zhou, Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Shuang Yong Road 6#, Nanning, 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China, Tel +86-771-5356528, Fax +86-771-5350031, Email pengtaogmu@163.com; zhouxin_gxmu@163.comBackground: This study aimed to identify dendritic cells (DCs) related genes in hepatocellular carcinoma (HCC) patients, establish DC-related subtypes and signatures, and correlate them with prognosis and treatment response.Methods: DC-related genes were screened using Weighted Gene Co-expression Network Analysis (WGCNA) based on RNA sequencing from the TCGA (374 samples), GSE14520 (242 samples), and GSE76427 datasets (115 samples), following immune infiltration assessment by the TIME method. Two DC-related subtypes in HCC were identified through unsupervised clustering. A DC-related signature (DCRS) predictive of overall survival was constructed using LASSO and Cox regression models, and validated across the three datasets. Additionally, genetic mutation characteristics, immune infiltration levels, and treatment sensitivity were explored in DCRS risk groups. The expression levels of DCRS genes and risk scores were validated in the transcriptome of 13 HCC patients receiving combined targeted therapy and immunotherapy in the Guangxi cohort using Wilcoxon test.Results: A signature consisting of 13 genes related to DCs was constructed, and the superior prognostic consistency of the low DCRS risk group was validated across the TCGA (P=0.003), GSE76427 (P=0.005), and GSE14520 (P=0.047) datasets. Furthermore, in the 147-sample transarterial chemoembolization (TACE) treatment dataset GSE104580, the response group exhibited lower risk scores than the non-response group (P=0.01), whereas in the 140-sample Sorafenib treatment dataset GSE109211 (P=0.041) and the 17-sample anti-PD-1 treatment dataset GSE202069 (P=0.027), the risk scores were higher in the response group. We also validated the gene expression levels of DCRS and the higher risk scores in the response group of the Guangxi cohort (P=0.034).Conclusion: A DCRS consisting of 13 genes was established in HCC, facilitating the prediction of patient prognosis and responsiveness to TACE, targeted therapy, and immunotherapy. Keywords: hepatocellular carcinoma, dendritic cells, prognosis, immunotherapy, targeted therapy

Ying-Xue Sun,1,* Tian Tang,1,* Jin-Yu Zou,1,* Qian-Qian Yue,1 Li-Feng Hu,1 Tong Peng,1 Xin-Ru Meng,1 Ge-Hui Feng,1 Li-Li Huang,1 Ying Zeng1– 3 1Department of International and Humanistic Nursing, School of Nursing, Hengyang Medical School, University of South China, Hengyang, People’s Republic of China; 2Hunan Province Key Laboratory of Tumor Cellular & Molecular Pathology, Cancer Research Institute, Hengyang Medical School, University of South China, Hengyang, People’s Republic of China; 3Hunan Engineering Research Center for Early Diagnosis and Treatment of Liver Cancer, Cancer Research Institute, Hengyang Medical School, University of South China, Hengyang, People’s Republic of China*These authors contributed equally to this workCorrespondence: Ying Zeng, Department of International and Humanistic Nursing, School of Nursing, Hengyang Medical School, University of South China, Hengyang, People’s Republic of China, Tel +86 17773486339, Fax + 86 0734-8281344, Email zengying2003@126.comBackground: Colorectal, and gastric cancers have the second, and fourth mortality rates worldwide, respectively. Endoscopic screening is a crucial diagnostic tool for colorectal, and gastric cancers. Effective interventions can improve adherence to endoscopic screening in high-risk populations, which is important for cancer prevention and mortality reduction. This study aimed to identify interventions that could improve adherence to endoscopic screening for cancer in high-risk populations.Methods: Combination keywords including colorectal cancer, gastric cancer, screening adherence, and interventions were used to search for articles in PubMed, Web of Science, Cochrane Library, and MEDLINE Complete. The review methodology was guided by the Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews (PRISMA-SCR).Results: A total of 12 articles were included in this review: 9 randomized controlled trials(RCT) and 3 quasi-experimental studies(QEDs). Among the extracted studies, 11 were about colorectal cancer, and 1 was about gastric cancer. Most studies used lecture-based or Information Technology-based health education interventions. Narrative interventions have proven to be novel and effective approaches for promoting adherence to endoscopic screening. Health education interventions included cancer epidemiology, cancer risk factors, warning symptoms, and screening methods.Conclusion: All interventions involved were effective in increasing individual knowledge of cancer-related endoscopic screening, willingness to undergo screening, and screening behaviors. These findings provide a reference for designing endoscopy-related cancer screening interventions.Keywords: colorectal cancer, gastric cancer, screening adherence, high-risk population

Junjie Chen,1,* Xiwen Liao,1,2 Yining Wu,1,* Shenjian Ou,1 Wei Qin,1 Chengkun Yang,1,2 Yufeng Tan,1 Quan Lao,1 Minhao Peng,1 Tao Peng,1,2 Xinping Ye1 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, People’s Republic of China; 2Guangxi Key Laboratory of Enhanced Recovery After Surgery for Gastrointestinal Cancer, Nanning, 530021, People’s Republic of China*These authors contributed equally to this workCorrespondence: Xinping Ye; Tao Peng, Tel +86-771-5356528, Fax +86-771-5350031, Email yexinping@gxmu.edu.cn; pengtaogmu@163.comAbstract: Hepatocellular carcinoma (HCC) recurrence, which encompasses both true recurrence resulting from cancer spread and de novo tumors developing within the same cancer-prone liver, presents a complication in approximately 70% of cases within a 5-year timeframe. The efficacy of neoadjuvant therapy for recurrence after hepatectomy for hepatocellular carcinoma is still unclear. We report a case of recurrent massive advanced hepatocellular carcinoma with pathological complete remission was treated by continuous hepatic arterial infusion chemotherapy (HAIC) and sequential transcatheter arterial embolization (TAE) combined with secondary operation. One month after resection, the patient recurred (massive type 141mm× 76mm). After 4 times of HAIC sequential TAE conversion therapy, the tumor shrank significantly (70mm× 29mm), alpha-fetoprotein(AFP) and protein induced by vitamin K absence or antagonist-II (PIVKA-II) levels decreased significantly, residual liver volume[left half liver accounted for 39.85% of standard liver volume(left half liver + right anterior lobe) accounted for 80.17% of standard liver volume] and Indocyanine green 15-minute retention(ICG R15 8.0%) complies with surgical requirement.The second operation was performed, and the tumor was completely resected after hepatic blood flow occlusion Requirement. The postoperative pathological results showed complete remission (PCR) of the tumor, and no recurrence was found during the follow-up of 16 months. In this case, HAIC sequential TAE conversion therapy has good short-term effect on patients with postoperative recurrence of hepatocellular carcinoma, tumor burden is significantly reduced, the second surgery pathology achieves complete remission, safety and tolerance, it is worthy of study and promotion.Keywords: hepatocellular carcinoma, recurrence after resection, two-stage hepatectomy, complete remission, sequential therapy

Yongguang Wei,1– 3,* Chenlu Lan,1– 3,* Xiangkun Wang,4 Xin Zhou,1– 3 Xiwen Liao,1– 3 Huasheng Huang,1– 3 Zhongliu Wei,1– 3 Tianman Li,1– 3 Tao Peng,1– 3 Guangzhi Zhu1– 3 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, People’s Republic of China; 2Guangxi Key Laboratory of Enhanced Recovery After Surgery for Gastrointestinal Cancer, Nanning, 530021, People’s Republic of China; 3Key Laboratory of High-Incidence-Tumor Prevention & Treatment (Guangxi Medical University), Ministry of Education, Nanning, 530021, People’s Republic of China; 4Departments of Hepatobiliary and Pancreatic Surgery, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, 450052, People’s Republic of China*These authors contributed equally to this workCorrespondence: Guangzhi Zhu, Tel +86-771-5356528, Email zhuguangzhi0792@hotmail.comBackground: RAD51 associated protein 1 (RAD51AP1) is shown to regulate cell proliferation and cancer progression. However, the immune-infiltrating correlation and the therapeutics guidance of RAD51AP1 in hepatocellular carcinoma (HCC) still need further investigation.Methods: In this study, comprehensive bioinformatic analysis of RAD51AP1 on differential expression, clinicopathologic correlation, prognostic value, and function enrichment were performed in The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO; GSE14520 and GSE76427), and International Cancer Genome Consortium (ICGC) datasets. Besides, the Guangxi cohort containing 50 pairs HCC and adjacent non-cancerous samples from First Affiliated Hospital of Guangxi Medical University was served as validation cohort. Moreover, we explored the predictive value of RAD51AP1 to therapeutics response and its underlying correlation with HCC immunoinfiltration.Results: RAD51AP1 was significantly overexpressed in HCC tissues and had a high diagnostic value of HCC. The shorter survival time and poorer clinical features were showed when RAD51AP1 upregulated, and then a nomogram featuring RAD51AP1 expression and other clinicopathologic factors was established to predict prognosis. In CIBERSORT analysis, higher T cells follicular helper but lower T cells CD4+ memory resting infiltration levels were exhibited when RAD51AP1 upregulated. The ssGSEA analysis demonstrated that high-RAD51AP1 expression subgroup had higher macrophages, Th2 and Treg cells infiltration levels, but lower type II IFN response function. Furthermore, high-RAD51AP1 expression subgroup exhibited the upregulated expression levels of immune-related checkpoint genes, but lower IPS and TIDE scores which suggested a possibly better immunotherapy response. The drug sensitivity analysis showed the high-expression subgroup may be more susceptible to Bexarotene, Doxorubicin, Gemcitabine and Tipifarnib.Conclusion: Taken together, RAD51AP1 is a potential diagnostic and prognostic biomarker. It may be related to the immunosuppressive microenvironment and could be an underlying HCC treatment strategy. However, the conclusions still require further validation studies.Keywords: hepatocellular carcinoma, RAD51AP1, prognostic signature, bioinformatics, immune filtration, drug sensitivity

IntroductionAs with any other radial glia in the central nervous system, Müller glia derive from the same neuroepithelial precursors, perform similar functions, and exhibit neurogenic properties as radial glia in the brain. Müller glial cells retain progenitor-like characteristics in the adult human eye and can partially restore visual function upon intravitreal transplantation into animal models of glaucoma. Recently, it has been demonstrated that intracellular communication is possible via the secretion of nano-sized membrane-bound extracellular vesicles (EV), which contain bioactive molecules like microRNA (miRNA) and proteins that induce phenotypic changes when internalised by recipient cells.MethodsWe conducted high-throughput sequencing to profile the microRNA signature of EV populations secreted by Müller glia in culture and used bioinformatics tools to evaluate their potential role in the neuroprotective signalling attributed to these cells.ResultsSequencing of miRNA within Müller EV suggested enrichment with species associated with stem cells such as miR-21 and miR-16, as well as with miRNA previously found to play a role in diverse Müller cell functions in the retina: miR-9, miR-125b, and the let-7 family. A total of 51 miRNAs were found to be differentially enriched in EV compared to the whole cells from which EV originated. Bioinformatics analyses also indicated that preferential enrichment of species was demonstrated to regulate genes involved in cell proliferation and survival, including PTEN, the master inhibitor of the PI3K/AKT pathway.DiscussionThe results suggest that the release by Müller cells of miRNA-enriched EV abundant in species that regulate anti-apoptotic signalling networks is likely to represent a significant proportion of the neuroprotective effect observed after the transplantation of these cells into animal models of retinal ganglion cell (RGC) depletion. Future studies will seek to evaluate the modulation of putative genes as well as the activation of these pathways in in vitro and in vivo models following the internalisation of Müller-EV by target retinal neurons.

Tuochao Peng, Shuangquan Qu, Zhen Du, Zheng Chen, Ting Xiao, Ru Chen Department of Anesthesia and Surgery, Hunan Children’s Hospital, Changsha, 410007, People’s Republic of ChinaCorrespondence: Tuochao Peng, Department of Anesthesia and Surgery, Hunan Children’s Hospital, Changsha, 410007, People’s Republic of China, Tel +86 13755074295, Email 258014811@qq.comPurpose: We performed this systematic review to summarize the psychometric properties of Face, Legs, Activity, Cry and Consolability (FLACC) scale in pediatric patients in different settings.Methods: Two investigators independently searched PubMed, EMBASE, OVID and China National Knowledge Infrastructure (CNKI) for eligible studies through July 2021. We assessed the psychometric properties using the modified critical appraisal tool (CAT). Finally, we systematically reviewed the results of the included studies.Results: A total of 15 studies were eventually included. The overall quality of each eligible study was low to moderate. The FLACC scale has been available in different versions and in different settings. Although eligible studies have demonstrated significant clinical benefit in assessing postoperative pain in pediatric patients aged 0 to 10 years from post-anesthetic care unit (PACU), pediatric intensive care unit (PICU) and inpatient unit, and in assessing procedural pain in pediatric patients aged 0.5 to 7 years from emergency unit, immunization center and PICU, mostly without test-retest analysis.Conclusion: Although the absence of a gold standard of pain assessment, the currently available data support the usefulness of the FLACC from the perspective of criterion validity. Therefore, the FLACC scale can be considered for measuring observational pain in infants and children. However, further studies are still needed to provide more robust evidence.Keywords: children, FLACC scale, infants, pain assessment, systematic review

Chenlu Lan,1,2,* Xinlei Huang,1,2,* Xiwen Liao,1,2 Xin Zhou,1,2 Kai Peng,1,2 Yongguang Wei,1,2 Chuangye Han,1,2 Tao Peng,1,2 Jianyao Wang,3 Guangzhi Zhu1,2 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Zhuang Autonomous Region, People’s Republic of China; 2Key Laboratory of High-Incidence-Tumor Prevention & Treatment (Guangxi Medical University), Ministry of Education, Nanning, 530021, People’s Republic of China; 3Department of General Surgery, Shenzhen Children’s Hospital, Shenzhen, Guangdong Province, People’s Republic of China*These authors contributed equally to this workCorrespondence: Guangzhi Zhu, Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China, Tel +86-771-5356528, Fax +86-771-5350031, Email zhuguangzhi0792@hotmail.com Jianyao Wang, Department of General Surgery, Shenzhen Children’s Hospital, Lianhua District, Shenzhen, 518026, Guangdong Province, People’s Republic of China, Email yaozi66@126.comObjective: The mechanisms of pseudouridine synthase (PUS) are not definite in hepatocellular carcinoma (HCC), the objective of this study is to investigate the effect of PUS genes in HCC.Materials and Methods: Differentially expressed and prognostic gene of PUS enzymes was identified based on The Cancer Genome Atlas (TCGA), International Cancer Genome Consortium (ICGC) and Gene Expression Profiling Interactive Analysis (GEPIA) databases. For the identified gene, pseudouridine synthase 1 (PUS1), was used for further research. The clinicopathological feature of PUS1 was analyzed by Student’s t-test. Prognostic significance was explored by Kaplan-Meier (KM) analysis and Cox proportional hazards regression model. Receiver operating characteristic (ROC) curve was applied to appraise diagnostic and prognostic value. The Database for Annotation, Visualization, and Integrated Discovery (DAVID) and Gene Set Enrichment Analysis (GSEA) were implemented to explore mechanism of PUS1. A Guangxi cohort was applied to verify differential expression. In vitro cell experiments were implemented to investigate the influence for proliferation, reactive oxygen species (ROS) level, migration, and invasion of HCC cells after a knockdown of PUS1.Results: PUS1 was significantly overexpressed in HCC tissues, and patients with high PUS1 were related to unpromising clinicopathological features. Survival analysis revealed high PUS1 expression was associated with a poor overall survival (OS) and 1 year-recurrence free survival (RFS), was an independent risk factor. Meanwhile, ROC curve showed that PUS1 had a diagnostic and prognostic significance to HCC. Functional enrichment analysis implied that PUS1 may be involved in metabolic pathways, mitochondrial function, non-alcoholic fatty liver disease (NAFLD), and some important carcinogenic pathways. Cell assays revealed that knockdown of PUS1 significantly constrained the migration, proliferation, invasion and improved the ROS level of HCC cells.Conclusion: PUS1 may be a prognostic biomarker and a underlying treatment target for HCC.Keywords: PUS1, pseudouridine synthase, poor prognosis, gene knockout, HCC

Sheng Ye,1 Huichun Xiao,1 Jian Chen,1 Di Zhang,1 Li Qi,1 Ting Peng,2 Yanyang Gao,2 Qianbing Zhang,3 Jinqing Qu,2 Lei Wang,4 Ruiyuan Liu1 1Guangdong Provincial Key Laboratory of Medical Image Processing, School of Biomedical Engineering, Southern Medical University, Guangzhou, Guangdong, People’s Republic of China; 2School of Chemistry and Chemical Engineering, South China University of Technology, Guangzhou, Guangdong, People’s Republic of China; 3Cancer Research Institute, School of Basic Medical Sciences, Southern Medical University, Guangzhou, Guangdong, People’s Republic of China; 4College of Materials and Chemical Engineering, Key Laboratory of Inorganic Nonmetallic Crystalline and Energy Conversion Materials, China Three Gorges University, Yichang, People’s Republic of ChinaCorrespondence: Jinqing Qu; Ruiyuan Liu, Email cejqqu@scut.edu.cn; ruiyliu@smu.edu.cnIntroduction: The high concentration of glutathione (GSH) and hydrogen peroxide (H2O2) levels within the tumor microenvironment (TME) are the major obstacle to induce the unsatisfactory anticancer treatment efficiency. The synergistic cancer therapy strategies of the combination the GSH depletion enhanced chemodynamic therapy (CDT) with photothermal therapy (PTT) have been proved to be the promising method to significantly improve the therapeutic efficacy.Methods: The copperphosphotungstate was incorporated into polyanilines to design copperphosphotungstate doped polyaniline nanorods (CuPW@PANI Nanorods) via chemical oxidant polymerization of aniline. The low long-term toxicity and biocompatibility were evaluated. Both in vitro and in vivo experiments were carried out to confirm the GSH depletion enhanced CDT/NIR-II PTT synergistic therapy.Results: CuPW@PANI Nanorods feature biosafety and biocompatibility, strong NIR-II absorbance, and high photothermal-conversion efficiency (45.14%) in NIR-II bio-window, making them highly applicable for photoacoustic imaging and NIR-II PTT. Moreover, CuPW@PANI Nanorods could consume endogenous GSH to disrupt redox homeostasis and perform a Fenton-like reaction with H2O2 to produce cytotoxic •OH for the enhanced CDT. Furthermore, NIR-II photothermal-induced local hyperthermia accelerates •OH generation to enhance CDT, which realizes high therapeutic efficacy in vivo.Conclusion: This study provides a proof of concept of GSH-depletion augmented chemodynamic/NIR-II photothermal therapy.Keywords: copperphosphotungstate, NIR-II absorption nanorods, NIR-II photothermal therapy, GSH depletion, chemodynamic therapy, synergistic therapy

Abstract Müller glia play very important and diverse roles in retinal homeostasis and disease. Although much is known of the physiological and morphological properties of mammalian Müller glia, there is still the need to further understand the profile of these cells during human retinal development. Using human embryonic stem cell-derived retinal organoids, we investigated the transcriptomic profiles of CD29+/CD44+ cells isolated from early and late stages of organoid development. Data showed that these cells express classic markers of retinal progenitors and Müller glia, including NFIX, RAX, PAX6, VSX2, HES1, WNT2B, SOX, NR2F1/2, ASCL1 and VIM, as early as days 10–20 after initiation of retinal differentiation. Expression of genes upregulated in CD29+/CD44+ cells isolated at later stages of organoid development (days 50–90), including NEUROG1, VSX2 and ASCL1 were gradually increased as retinal organoid maturation progressed. Based on the current observations that CD24+/CD44+ cells share the characteristics of early and late-stage retinal progenitors as well as of mature Müller glia, we propose that these cells constitute a single cell population that upon exposure to developmental cues regulates its gene expression to adapt to functions exerted by Müller glia in the postnatal and mature retina.

Yining Wu,1,* Shenjian Ou,1,* Xiwen Liao,1,2 Chuangye Han,1,2 Chengkun Yang,1,2 Wei Qin,1 Yufeng Tan,1 Quan Lao,1 Tao Peng,1,2 Xinping Ye1 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Zhuang Autonomous Region, 530021, People’s Republic of China; 2Guangxi Key Laboratory of Enhanced Recovery After Surgery for Gastrointestinal Cancer, Nanning, Guangxi Zhuang Autonomous Region, 530021, People’s Republic of China*These authors contributed equally to this workCorrespondence: Xinping Ye; Tao Peng, Tel +86-771-5356528, Fax +86-771-5350031, Email yexinping@gxmu.edu.cn; pengtaogmu@163.comAbstract: Situs inversus totalis (SIT) is a rare congenital condition in which abdominal and thoracic organs are transposed from normal positions. Two-stage hepatectomy (TSH) combined with translational therapy for hepatocellular carcinoma (HCC) with SIT has been rarely reported. We report a 41-year-old man with giant hepatocellular carcinoma (71 mm × 55 mm × 51 mm) whose future residual liver (FLR) and standard liver volume (SLV) ratio at first diagnosis was 37.4%. Preoperative volume assessment of portal vein ligation (PVL) revealed inadequate hypertrophy of FLR. After a multidisciplinary group discussion (MDT), the patient decided to follow conversion therapy. Three months later, ratio of the FLR/SLV increased from 37.4% to 71% after operation, which met the surgical requirements. Second hepatectomy, right lobectomy was successful. There was no recurrence after six months of follow-up. In our case, conversion therapy appears to be effective in maintaining residual liver hyperplasia, reducing tumor load, and preventing tumor progression in patients with large HCC during TSH.Keywords: situs inversus totalis, two-stage hepatectomy, hepatocellular carcinoma, future residual liver, multidisciplinary group discussion

Huasheng Huang,* Xiwen Liao,* Guangzhi Zhu, Chuangye Han, Xiangkun Wang, Chengkun Yang, Xin Zhou, Tianyi Liang, Ketuan Huang, Tao Peng Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China*These authors contributed equally to this workCorrespondence: Tao Peng; Xiwen Liao, Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Shuang Yong Road 6, Nanning, 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China, Tel +86-771-5356528, Fax +86-771-5350031, Email pengtaogmu@163.com; LXWen1991@hotmail.comBackground: The aim of our study was to evaluate the potential of expression and single nucleotide polymorphism of Acyl-CoA binding domain containing 4 (ACBD4) gene as prognosis biomarkers in patients with hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) after hepatectomy.Methods: HBV-related HCC patients from the First Affiliated Hospital of Guangxi Medical University and GSE14520 were included in the current study, as well as The Cancer Genome Atlas (TCGA) HCC verification cohort. Prognostic analysis and multiple functional enrichment analysis methods were used to evaluate the prognostic value and potential biological functions of the ACBD4 gene in HBV-related HCC.Results: We found that ACBD4 gene is highly expressed in normal liver tissues and markedly down-regulated in HBV-related HCC tissues. ACBD4 gene was significantly related to overall survival (OS) of HCC in TCGA and GSE14520 cohorts, and patients with low ACBD4 expression were markedly related to poor OS. Rs4986172 was observed as an OS biomarker after hepatectomy in the Guangxi HBV-related HCC cohort. The OS of rs4986172 GG genotype was worse than that of HCC patients with A allele (AA and AG genotypes). Multifunctional enrichment analysis suggested that ACBD4 gene is closely related to the metabolic, peroxisome proliferator-activated receptor and cytochrome P450 pathway. Through connectivity map, we also identified eight compounds that may be used as targeted therapeutic agents for ACBD4 gene in HBV-related HCC; these compounds were scopoletin, alfaxalone, bephenium hydroxynaphthoate, apramycin, 4,5-dianilinophthalimide, DL-thiorphan, aminohippuric acid and quinidine. Immune microenvironment analysis revealed that there were significant differences in immune scores of HBV-related HCC tumor tissues with different ACBD4 expression levels.Conclusion: Our study reveals that ACBD4 expression and rs4986172 can be serve as biomarkers of OS in HBV-related HCC patients after hepatectomy.Keywords: rs4986172, ACBD4, HBV-related HCC, overall survival, hepatectomy

Yong-Guang Wei, Cheng-Kun Yang, Zhong-Liu Wei, Xi-Wen Liao, Yong-Fei He, Xin Zhou, Hua-Sheng Huang, Chen-Lu Lan, Chuang-Ye Han, Tao Peng Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi Zhuang Autonomous Region, People’s Republic of ChinaCorrespondence: Tao PengDepartment of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Shuang Yong Road 6, Nanning, Guangxi Zhuang Autonomous Region, People’s Republic of ChinaTel +86-771-5356528Fax +86-771-5350031Email pengtaogmu@163.comBackground: The protein high-mobility group AT-hook 1 (HMGA1) has been demonstrated that modulated cellular proliferation, invasion, and apoptosis with a poor prognosis in miscellaneous carcinomas. However, the mechanism of circumstantial carcinogenesis and association with the immune microenvironment of HMGA1 in hepatocellular carcinoma (HCC) had not been extensively explored.Methods: The gene expression, clinicopathological correlation, and prognosis analysis were performed in the data obtained from TCGA. The results were further validated by ICGC and GEO database and external validation cohort from Guangxi. The HMGA1 protein expression was further examined in the HPA database. Biological function analyses were conducted by GSEA, STRING database, and Coexpedia online tool. Using TIMER and CIBERSORT method, the relationship between immune infiltrate and HMGA1 was investigated.Results: In HCC, HMGA1 had much higher transcriptional and proteomic expression than in corresponding paraneoplastic tissue. Patients with high HMGA1 expression had a poor prognosis and unpromising clinicopathological features. High HMGA1 expression was closely related to the cell cycle, tumorigenesis, substance metabolism, and immune processes by regulating complex signaling pathways. Notably, HMGA1 may be associated with TP53 mutational carcinogenesis. Moreover, increased HMGA1 expression may lead to an increase in immune infiltration and a decrease in tumor purity in HCC. CIBERSORT analysis elucidated that the amount of B cell naive, B cell memory, T cells gamma delta, macrophages M2, and mast cell resting decreased when HMGA1 expression was high, whereas T cells follicular helper, macrophages M0, and Dendritic cells resting increased.Conclusion: In conclusions, HMGA1 is a potent prognostic biomarker and a sign of immune infiltration in HCC, which may be a potential immunotherapy target for HCC.Keywords: hepatocellular carcinoma, HMGA1, prognostic signature, bioinformatics, immune filtration

Xin Zhou,1,2,* Tian-Man Li,1– 3,* Jian-Zhu Luo,1,2,* Chen-Lu Lan,1,2 Zhong-Liu Wei,1,2 Tian-Hao Fu,1,2 Xi-Wen Liao,1,2 Guang-Zhi Zhu,1,2 Xin-Ping Ye,1,2 Tao Peng1,2 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China; 2Key Laboratory of Early Prevention & Treatment for Regional High Frequency Tumor (Guangxi Medical University), Ministry of Education, Nanning, 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China; 3Department of Hepatobiliary Surgery, The Sixth Affiliated Hospital of Guangxi Medical University, Yulin, 537000, Guangxi Zhuang Autonomous Region, People’s Republic of China*These authors contributed equally to this workCorrespondence: Tao Peng Email pengtaogmu@163.comBackground: Cytochrome P450 2C8 (CYP2C8) gene is one of the members of the cytochrome P450 enzymes (CYPs) gene family. The aim of this study was to reveal the function of CYP2C8 in hepatocellular carcinoma (HCC) and its effect on the sorafenib resistance.Methods: Differential expression analysis in multiple HCC datasets all suggested that CYP2C8 expression was significantly decreased in HCC tissues, compared with para-carcinoma liver tissues. The expression level of CYP2C8 was subsequently compared between HCC tissues and para-carcinoma liver tissues of 70 patients form Guangxi, China, with the result consistent with the above. Survival analysis and ROC analysis indicated that CYP2C8 was equipped with satisfactory diagnostic and prognostic value in HCC. To examine the effect of CYP2C8 on the malignant phenotype of HCC cells, stable transcriptional cell lines with CYP2C8 over-expression were established, and then Cell Counting Kit-8 (CCK8) assay, colony formation assay, cell cycle assay, cell invasion assay and wound healing assay were performed.Results: The results of aforementioned assays suggested that CYP2C8 over-expression restricted the proliferation, clonality, migration, invasion and cell cycle of HCC cells but had no significant effect on cell apoptosis. The enrichment analysis in terms of sequencing data of HCC cell lines with stable CYP2C8 over-expression suggested that CYP2C8 might be related to PI3K/Akt/p27Kip1 axis. The inhibition of CYP2C8 over-expression on PI3K/Akt/p27Kip1 axis was subsequently demonstrated with Western blot assay. In the rescue experiment, it was observed that both P27 inhibitor and PI3K agonist counteracted the repressed malignant phenotype caused by CYP2C8 over-expression, which further demonstrated that CYP2C8 played a role in HCC cells via PI3K/Akt/p27Kip1 axis.Discussion: The results demonstrated that CYP2C8 enhances the anticancer activity of sorafenib in vitro assays and in tumor xenograft model, with Ki-67 down-regulation and PI3K/Akt/p27Kip1 axis inhibition. In conclusion, these findings hinted that CYP2C8 restricted malignant phenotype and sorafenib resistance in HCC via PI3K/Akt/p27kip1 axis.Keywords: hepatocellular carcinoma, CYP2C8, sorafenib resistance, cell cycle

Lifang Wei,1,* Ye Li,2 Yue Yu,1 Minmin Xu,1 Huilan Chen,1 Lijie Li,1 Ting Peng,1 Kang Zhao,1 Yongze Zhuang3,* 1Department of Nephrology, The Third People’s Hospital Affiliated to Fujian University of Traditional Chinese Medicine, Fuzhou, Fujian, People’s Republic of China; 2The Third People’s Hospital Affiliated to Fujian University of Traditional Chinese Medicine, Fuzhou, Fujian, People’s Republic of China; 3Department of Nephrology, 900 Hospital of the Joint Logistics Team, PLA, Fujian University of Traditional Chinese Medicine, Fuzhou, Fujian, People’s Republic of China*These authors contributed equally to this workCorrespondence: Lifang Wei; Yongze Zhuang Tel +86 591 62091295Fax +86 591 22869300Email liwlf2008@163.com; Doczhuang@aliyun.comAbstract: Obesity-related glomerulopathy (ORG) is a secondary glomerular disease caused by obesity, with clinical manifestations such as proteinuria and glomerulomegaly. Currently, the high incidence of obesity brings a change in the spectrum of kidney diseases across the globe, including China. ORG has become another important secondary nephropathy leading to end-stage renal disease (ESRD), and its incidence has increased significantly. This trend is bound to bring about a serious socioeconomic burden. Therefore, it is urgent to study its pathogenesis and intervention measures. Currently, the occurrence and development mechanisms in ORG are complicated by many factors, which are still unclear. In the past 20 years, with the continuous intensive research on mechanisms such as hypoxia in the metabolic process, immune inflammation, and pyroptosis, there have been new advances in the mechanism of ORG, especially the important role of inflammation in podocyte injury and its impact on the progress of ORG. Here, we briefly review the possible pathogenic role of the inflammasome in the podocyte damage in ORG and summarize the possible therapeutical strategies targeting inflammasome.Keywords: obesity, NLRP3 inflammasome, ORG, podocyte, Chinese herbal medicines

Chunyang Pan, Yi Zhao, Haojie Xie, Yongyan Zhou, Ranran Duan, Yanfei Li, Yanjie Jia, Tao Peng Department of Neurology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, People’s Republic of ChinaCorrespondence: Tao PengDepartment of Neurology, The First Affiliated Hospital of Zhengzhou University, Erqi District, Zhengzhou, Henan Province, People’s Republic of ChinaTel +861 367 366 5072Email zdyfy0420@163.comPurpose: To describe and compare the clinical features of patients with first-episode neuromyelitis optica spectrum disorder (NMOSD) in a normal complement C4 group and a low complement C4 group, and explore the mechanism by which low complement C4 affects the clinical features of patients with NMOSD.Patients and Methods: We retrospectively analyzed clinical data of 169 aquaporin-4 (AQP4) antibody positive patients with NMOSD from the First Affiliated Hospital of Zhengzhou University from December 2013 to March 2021. Prior to treatment, the blood was drawn for detection, and the patients underwent a 3.0 Tesla MRI examination. A low complement C4 level was defined as a serum complement C4 level < 0.14 g/L. Depending on whether the complement C4 level was reduced, it was divided into the normal complement C4 group and low complement C4 group. The basic demographics, clinical manifestations, laboratory examinations, and imaging findings of the two groups were compared.Results: Among the 169 AQP4 antibody positive patients, 54 were low-complement C4 patients and 115 were normal. There were no significant differences in the demographics, clinical manifestations, treatment options, or admission Expanded Disability Status Scale (EDSS) score between two groups (P > 0.05). The median of discharged EDSS was the same (4 vs 4), but the difference between the two was statistically significant (P = 0.019). Compared with the normal complement C4 group, the blood uric acid level (225 vs 179; P = 0.001) and the C3 level (1.06 vs 0.87, P = 0.000) of the low complement C4 group were significantly lower. The incidence of brainstem lesions in patients with low complement C4 was higher (53.7% vs 33%, P = 0.01).Conclusion: The treatment effect of the first-episode AQP4 antibody positive NMOSD low complement C4 group was poor, the blood-brain barrier was more severely damaged, and the disease changes were likely to involve the brainstem.Keywords: blood-brain barrier, brainstem, humoral immunity, optic neuritis

Primary open-angle glaucoma (POAG) is highly heritable, yet not well understood from a genetic perspective. Here, the authors perform a meta-analysis of genome-wide association studies in 34,179 POAG cases, identifying 44 previously unreported risk loci and mapping effects across multiple ethnicities.

Tiequan Zhang, Mi Li, Haofeng Lu, Tao Peng Department of Hepatobiliary Surgery, The First People’s Hospital of Jingzhou, Jingzhou 434000, People’s Republic of ChinaCorrespondence: Tiequan ZhangDepartment of Hepatobiliary Surgery, The First People’s Hospital of Jingzhou, No. 8 Hangkong Road, Shashi District, Jingzhou, Hubei 434000, People’s Republic of ChinaTel +86-716-8115036Email ztq08250825@163.comBackground: Accruing evidences have pointed out that abnormal expression of circular RNAs (circRNAs) was closely related to the development of many malignancies. The present study intended to disclose the role of circRNA eukaryotic translation initiation factor 6 (circEIF6; hsa_circ_0060055) in pancreatic cancer progression.Methods: Quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to determine the expression of circEIF6, EIF6 messenger RNA (mRNA), microRNA-557 (miR-557) and solute carrier family 7 member 11 (SLC7A11) mRNA. Cell proliferation ability, migration and invasion abilities and apoptosis were evaluated by Cell Counting Kit 8 (CCK8) assay, transwell migration and invasion assays and flow cytometry. Western blot assay was performed for the expression determination of all proteins. The predicted interaction between miR-557 and circEIF6 or SLC7A11 was confirmed by dual-luciferase reporter assay. Xenograft tumor model was used for exploring the biological function of circEIF6 in vivo.Results: CircEIF6 abundance was aberrantly up-regulated in pancreatic tumor tissues and cell lines. Cell proliferation, migration and invasion were significantly restrained while cell apoptosis was induced with the silencing of circEIF6 in pancreatic cancer cells. CircEIF6 silencing also hampered the activation of phosphatidylinositol 3-kinase (PI3K)/AKT serine/threonine kinase (AKT) pathway. CircEIF6 bound to miR-557, and circEIF6 silencing elevated the expression of miR-557 in pancreatic cancer cells. MiR-557 knockdown partly overturned circEIF6 silencing-induced effects in pancreatic cancer cells. SLC7A11 was a target of miR-557, and miR-557 overexpression suppressed malignant potential of pancreatic cancer cells partly through reducing the expression of SLC7A11. CircEIF6 knockdown blocked xenograft tumor growth in vivo.Conclusion: CircEIF6 aggravated pancreatic cancer development through promoting cell proliferation, migration and invasion and suppressing cell apoptosis through targeting miR-557/SLC7A11/PI3K/AKT signaling.Keywords: pancreatic cancer, EIF6, miR-557, SLC7A11, PI3K/AKT signaling

Guangyou Duan,* Kai Wang,* Taotao Peng, Zhuoxi Wu, Hong Li Department of Anesthesiology, Second Affiliated Hospital, Army Medical University, Chongqing 400037, People’s Republic of China*These authors contributed equally to this workCorrespondence: Hong LiDepartment of Anesthesiology, Second Affiliated Hospital, Army Medical University, Chongqing 400037, People’s Republic of ChinaTel +8613608380123Fax +862368774997Email lh78553@163.comObjective: The study aimed to investigate the effects of intraoperative dexmedetomidine on postoperative sleep disturbance for different surgical patients and compare such effects between different dose of dexmedetomidine.Methods: A total of 7418 patients undergoing nine types of non-cardiac major surgeries were retrospectively studied. Patients were separated into DEX (dexmedetomidine) or Non-DEX (Non-dexmedetomidine) groups based on the use of dexmedetomidine during surgery. The patients who reported they could not fall asleep during the night or woke up repeatedly during the most of the night at the day of the surgery and whose NRS were > 6 were defined as cases with severe sleep disturbance. Propensity score matched analysis based on all preoperative baseline data was performed along with logistic regression analysis including different surgery types and dosage of dexmedetomidine use.Results: In both of the unmatched cohort (OR, 0.49 [95% CI: 0.43– 0.56]) and matched cohort (0.49 [95% CI: 0.42– 0.58]), the DEX group had a significantly lower incidence of severe sleep disturbance than the Non-DEX group. In the subgroup analysis, for gynecological and urological surgery population, the ORs for DEX-group reached 0.21 (95% CI, 0.13– 0.33; P< 0.0001) and 0.30 (95% CI,0.19– 0.47; P< 0.0001), respectively. In addition, low-dose dexmedetomidine (0.2– 0.4 μg·kg− 1·h− 1) showed the greatest effect with an odds ratio of 0.38 (95% CI: 0.31– 0.44; P< 0.0001), and the incidence of severe sleep disturbance in the low-dose group was significantly lower (11.5% vs. 17.7% vs. 16.5%, P< 0.0001) than that in the medium- (0.4– 0.6 μg·kg− 1·h− 1) and high-dose (0.6– 0.8 μg·kg− 1·h− 1) groups.Conclusion: Intraoperative dexmedetomidine use can significantly decrease the incidence of severe sleep disturbance on the day of surgery for patients undergoing non-cardiac major surgery, and the effects were most significant in patients receiving gynecological and urological surgery. Furthermore, low-dose dexmedetomidine (0.2– 0.4 μg·kg− 1·h− 1) is most effective for prevention of postoperative sleep disturbance.Keywords: dexmedetomidine, intraoperative use, postoperative sleep disturbance

Summary: Vision impairment (VI) can have wide ranging economic impact on individuals, households, and health systems. The aim of this systematic review was to describe and summarise the costs associated with VI and its major causes. We searched MEDLINE (16 November 2019), National Health Service Economic Evaluation Database, the Database of Abstracts of Reviews of Effects and the Health Technology Assessment database (12 December 2019) for partial or full economic evaluation studies, published between 1 January 2000 and the search dates, reporting cost data for participants with VI due to an unspecified cause or one of the seven leading causes globally: cataract, uncorrected refractive error, diabetic retinopathy, glaucoma, age-related macular degeneration, corneal opacity, trachoma. The search was repeated on 20 January 2022 to identify studies published since our initial search. Included studies were quality appraised using the British Medical Journal Checklist for economic submissions adapted for cost of illness studies. Results were synthesized in a structured narrative. Of the 138 included studies, 38 reported cost estimates for VI due to an unspecified cause and 100 reported costs for one of the leading causes. These 138 studies provided 155 regional cost estimates. Fourteen studies reported global data; 103/155 (66%) regional estimates were from high-income countries. Costs were most commonly reported using a societal (n = 48) or healthcare system perspective (n = 25). Most studies included only a limited number of cost components. Large variations in methodology and reporting across studies meant cost estimates varied considerably. The average quality assessment score was 78% (range 35–100%); the most common weaknesses were the lack of sensitivity analysis and insufficient disaggregation of costs. There was substantial variation across studies in average treatment costs per patient for most conditions, including refractive error correction (range $12–$201 ppp), cataract surgery (range $54–$3654 ppp), glaucoma (range $351–$1354 ppp) and AMD (range $2209–$7524 ppp). Future cost estimates of the economic burden of VI and its major causes will be improved by the development and adoption of a reference case for eye health. This could then be used in regular studies, particularly in countries with data gaps, including low- and middle-income countries in Asia, Eastern Europe, Oceania, Latin America and sub-Saharan Africa.

Summary: Background: We undertook a Grand Challenges in Global Eye Health prioritisation exercise to identify the key issues that must be addressed to improve eye health in the context of an ageing population, to eliminate persistent inequities in health-care access, and to mitigate widespread resource limitations. Methods: Drawing on methods used in previous Grand Challenges studies, we used a multi-step recruitment strategy to assemble a diverse panel of individuals from a range of disciplines relevant to global eye health from all regions globally to participate in a three-round, online, Delphi-like, prioritisation process to nominate and rank challenges in global eye health. Through this process, we developed both global and regional priority lists. Findings: Between Sept 1 and Dec 12, 2019, 470 individuals complete round 1 of the process, of whom 336 completed all three rounds (round 2 between Feb 26 and March 18, 2020, and round 3 between April 2 and April 25, 2020) 156 (46%) of 336 were women, 180 (54%) were men. The proportion of participants who worked in each region ranged from 104 (31%) in sub-Saharan Africa to 21 (6%) in central Europe, eastern Europe, and in central Asia. Of 85 unique challenges identified after round 1, 16 challenges were prioritised at the global level; six focused on detection and treatment of conditions (cataract, refractive error, glaucoma, diabetic retinopathy, services for children and screening for early detection), two focused on addressing shortages in human resource capacity, five on other health service and policy factors (including strengthening policies, integration, health information systems, and budget allocation), and three on improving access to care and promoting equity. Interpretation: This list of Grand Challenges serves as a starting point for immediate action by funders to guide investment in research and innovation in eye health. It challenges researchers, clinicians, and policy makers to build collaborations to address specific challenges. Funding: The Queen Elizabeth Diamond Jubilee Trust, Moorfields Eye Charity, National Institute for Health Research Moorfields Biomedical Research Centre, Wellcome Trust, Sightsavers, The Fred Hollows Foundation, The Seva Foundation, British Council for the Prevention of Blindness, and Christian Blind Mission. Translations: For the French, Spanish, Chinese, Portuguese, Arabic and Persian translations of the abstract see Supplementary Materials section.

[This corrects the article DOI: 10.1371/journal.pgen.1009497.].

Abstract Glaucoma is one of the leading causes of blindness, and there is an ongoing need for new therapies. Recent studies indicate that cell transplantation using Müller glia may be beneficial, but there is a need for novel sources of cells to provide therapeutic benefit. In this study, we have isolated Müller glia from retinal organoids formed by human induced pluripotent stem cells (hiPSCs) in vitro and have shown their ability to partially restore visual function in rats depleted of retinal ganglion cells by NMDA. Based on the present results, we suggest that Müller glia derived from retinal organoids formed by hiPSC may provide an attractive source of cells for human retinal therapies, to prevent and treat vision loss caused by retinal degenerative conditions. Stem Cells Translational Medicine 2019;8:775&784

Linya Guo,1,* Yanbing Li,2,3,* Ying Tian,4,5,* Shaoxin Gong,6 Xi Chen,1 Tianhong Peng,1 Aiping Wang,1,4,5,7 Zhisheng Jiang71Clinical Anatomy & Reproductive Medicine Application Institute, School of Medicine, University of South China, Hengyang 421001, People’s Republic of China; 2National Key Discipline of Human Anatomy, Southern Medical University, Guangzhou 510000, Guangdong, People’s Republic of China; 3Guangdong Engineering Research Center for Translation of Medical 3D Printing Application, Guangzhou, 510000, Guangdong, People’s Republic of China; 4Institute of Clinical Research, Affiliated Nanhua Hospital, University of South China, Hengyang 421002, Hunan, People’s Republic of China; 5Postdoctoral Research Institute on Basic Medicine, University of South China, Hengyang, 421001, Hunan, People’s Republic of China; 6Department of Pathology, First Affiliated Hospital, University of South China, Hengyang 421001, Hunan, People’s Republic of China; 7Key Laboratory for Arteriosclerology of Hunan Province, Institute of Cardiovascular Disease, University of South China, Hengyang 421001, Hunan, People’s Republic of China*These authors contributed equally to this workPurpose: Eukaryotic initiation factor 2α (eIF2α) plays important roles in the proliferation and survival of pulmonary artery smooth muscle cells (PASMCs) in animal hypoxia-induced pulmonary hypertension models. However, the underlying mechanism remains unknown at large. Autophagy has been reported to play a key role in the vascular remodeling in pulmonary arterial hypertension (PAH). The purposes of this study are to determine the functions of eIF2α and autophagy in the vascular remodeling of the monocrotaline-induced PAH rats and to clarify the correlation between eIF2α and autophagy.Methods: We established a rat model of monocrotaline-induced PAH, and we established a cell model of platelet derived growth factor (PDGF)-induced PASMCs proliferation. The vascular morphology and the expression of eIF2α, LC3B, and p62 were assessed in the pulmonary arterial tissue of Sprague-Dawleyrats and PDGF-induced PASMCs.Results: Autophagy was significantly active in monocrotaline model group (MCT)-induced PAH rats, which obviously promotes vascular remodeling in MCT-induced PAH rats. Furthermore, the proliferation of PASMCs was induced by PDGF in vitro. The expression of LC3B, eIF2α was increased in the PDGF-induced PASMCs proliferation, and the expression of p62 was reduced in the PDGF-induced PASMCs proliferation. Moreover, eIF2α siRNA downregulated the expression of eIF2α and LC3B, and upregulated the expression of p62 in PDGF-induced PASMCs proliferation. eIF2α siRNA inhibited the PDGF-induced PASMCs proliferation. Finally, chloroquine can upregulate the protein expression of LC3B and p62, it also can inhibit proliferation in PDGF-induced PASMCs.Conclusion: Based on these observations, we conclude that eIF2α promotes the proliferation of PASMCs and vascular remodeling in monocrotaline-induced PAH rats through accelerating autophagy pathway.Keywords: eIF2α, autophagy, PASMCs, PAH, monocrotaline

Optical Coherence Tomography (OCT) enables non-invasive imaging of the retina and is used to diagnose and manage ophthalmic diseases including glaucoma. We present the first large-scale genome-wide association study of inner retinal morphology using phenotypes derived from OCT images of 31,434 UK Biobank participants. We identify 46 loci associated with thickness of the retinal nerve fibre layer or ganglion cell inner plexiform layer. Only one of these loci has been associated with glaucoma, and despite its clear role as a biomarker for the disease, Mendelian randomisation does not support inner retinal thickness being on the same genetic causal pathway as glaucoma. We extracted overall retinal thickness at the fovea, representative of foveal hypoplasia, with which three of the 46 SNPs were associated. We additionally associate these three loci with visual acuity. In contrast to the Mendelian causes of severe foveal hypoplasia, our results suggest a spectrum of foveal hypoplasia, in part genetically determined, with consequences on visual function.

The iron-bearing ore, existing in the form of oolite, was mainly composed of hematite, limonite, daphnite, and collophane. The harmful element phosphorus content was 1.56%, belonging to high phosphorus ooliticiron ore in western Hubei. In this study, segregation roasting and low intensity magnetic separation techniques were applied for upgrading iron and removing phosphorus. The ores, the chlorinating agent, and the reducing agent were mixed into the roasting furnace for segregation roasting. After being transferred from the weak magnetic minerals to the strong ones, the iron was recovered by low intensity magnetic separation. During segregation roasting, new ore phases, metallic iron (Fe), a small amount of ferroferric oxide (Fe3O4), and ferrous oxide (FeO) could be observed. The results showed that the iron concentrate with the Fe content of 90.3%, the phosphorus content of 0.15%, and the iron recovery of 92.9% were obtained under the segregation roasting temperature of 1273 K, and the roasting time of 90 min, CaCl2 (calcium chloride) 20%, Ca (ClO)2 (calcium hypochlorite) 3%, the dosage of coke 20%, and low intensity magnetic separation field intensity 0.12 T.

Current estimates suggest 50% of glaucoma blindness worldwide is caused by primary angle-closure glaucoma (PACG) but the causative gene is not known. We used genetic linkage and whole genome sequencing to identify Spermatogenesis Associated Protein 13, SPATA13 (NM_001166271; NP_001159743, SPATA13 isoform I), also known as ASEF2 (Adenomatous polyposis coli-stimulated guanine nucleotide exchange factor 2), as the causal gene for PACG in a large seven-generation white British family showing variable expression and incomplete penetrance. The 9 bp deletion, c.1432_1440del; p.478_480del was present in all affected individuals with angle-closure disease. We show ubiquitous expression of this transcript in cell lines derived from human tissues and in iris, retina, retinal pigment and ciliary epithelia, cornea and lens. We also identified eight additional mutations in SPATA13 in a cohort of 189 unrelated PACS/PAC/PACG samples. This gene encodes a 1277 residue protein which localises to the nucleus with partial co-localisation with nuclear speckles. In cells undergoing mitosis SPATA13 isoform I becomes part of the kinetochore complex co-localising with two kinetochore markers, polo like kinase 1 (PLK-1) and centrosome-associated protein E (CENP-E). The 9 bp deletion reported in this study increases the RAC1-dependent guanine nucleotide exchange factors (GEF) activity. The increase in GEF activity was also observed in three other variants identified in this study. Taken together, our data suggest that SPATA13 is involved in the regulation of mitosis and the mutations dysregulate GEF activity affecting homeostasis in tissues where it is highly expressed, influencing PACG pathogenesis.

Xinping Ye,1,* Guanghui Li,1,* Chuangye Han,1 Quanfa Han,1 Liming Shang,1 Hao Su,1 Bowen Han,1 Yizhen Gong,1 Guodong Lu,2 Tao Peng1 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Province, China; 2Department of Toxicology, School of Public Health, Guangxi Medical University, Nanning, Guangxi Province, China *These authors contributed equally to this work Background: This study aimed to determine if the number of circulating tumor cells (CTCs) and changes in their numbers affected tumor recurrence and metastasis after surgical resection in patients with hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC). Methods: The primary endpoints were overall survival (OS) and progression-free survival (PFS). A total of 42 patients with HCC were selected from the First Affiliated Hospital of Guangxi Medical College from 2014 to 2017. CTCs were counted 1 day prior to and 30 days after surgical excision of HCC using the CanPatrol™ system. Results: Numbers of CTCs (> 2 CTCs and > 5 CTCs per 5 ml peripheral blood) were significantly associated with Edmondson stage in HBV-related HCC prior to surgery (P = 0.004 and 0.014, respectively). However there were no significant associations between other tested clinicopathological factors and CTC counts. Postoperative CTC counts (> 2 and > 5) and pre/postoperative change in CTC counts were significantly associated with PFS (P = 0.02, 0.009, and 0.001, respectively), but not with OS. Receiver operating characteristic curve analysis showed that pre/postoperative changes in the CTC count were a better predictor of performance than absolute count. The postoperative CTC count was also significantly associated with positive TP53 expression (P < 0.05). Conclusion: These results demonstrate that postoperative CTC counts (> 2 and > 5) and changes in CTC counts may be independent prognostic indicators for PFS in patients with HBV-related HCC, with the change in number of CTCs showing better predictive performance. Keywords: circulating tumor cell, hepatocellular carcinoma, progression-free survival, overall survival

Abstract Background Sustained drug delivery is a large unmet clinical need in glaucoma. Here, we incorporated a Myocardin-Related Transcription Factor/Serum Response Factor inhibitor, CCG-222740, into slow release large unilamellar vesicles derived from the liposomes DOTMA (1,2-di-O-octadecenyl-3-trimethylammonium propane) and DOPC (1,2-dioleoyl-sn-glycero-3-phosphocholine), and tested their effects in vitro and in vivo. Results The vesicles were spherical particles of around 130 nm and were strongly cationic. A large amount of inhibitor could be incorporated into the vesicles. We showed that the nanocarrier CCG-222740 formulation gradually released the inhibitor over 14 days using high performance liquid chromatography. Nanocarrier CCG-222740 significantly decreased ACTA2 gene expression and was not cytotoxic in human conjunctival fibroblasts. In vivo, nanocarrier CCG-222740 doubled the bleb survival from 11.0 ± 0.6 days to 22.0 ± 1.3 days (p = 0.001), decreased conjunctival scarring and did not have any local or systemic adverse effects in a rabbit model of glaucoma filtration surgery. Conclusions Our study demonstrates proof-of-concept that a nanocarrier-based formulation efficiently achieves a sustained release of a Myocardin-Related Transcription Factor/Serum Response Factor inhibitor and prevents conjunctival fibrosis in an established rabbit model of glaucoma filtration surgery.

Xiwen Liao,1 Chuangye Han,1 Xiangkun Wang,1 Ketuan Huang,1 Tingdong Yu,1 Chengkun Yang,1 Rui Huang,2 Zhengqian Liu,1 Quanfa Han,1 Tao Peng1 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China; 2Department of Hematology, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China Background: The aim of the current study was to investigate the potential prognostic value of minichromosome maintenance (MCM) genes in patients with early-stage pancreatic ductal adenocarcinoma (PDAC) after pancreaticoduodenectomy by using the RNA-sequencing dataset from The Cancer Genome Atlas (TCGA).Methods: An RNA-sequencing dataset of 112 early-stage PDAC patients who received a pancreaticoduodenectomy was obtained from TCGA. Survival analysis was used to identify potential prognostic values of MCM genes in PDAC overall survival (OS).Results: Through mining public databases, we observed that MCM genes (MCM2, MCM3, MCM4, MCM5, MCM6, and MCM7) were upregulated in pancreatic cancer tumor tissue and have a strong positive coexpression with each other. Multivariate survival analysis indicated that a high expression of MCM4 significantly increased the risk of death in patients with PDAC, and time-dependent receiver operating characteristic analysis showed an area under the curve of 0.655, 0.587, and 0.509 for a 1-, 2-, and 3-year PDAC OS prediction, respectively. Comprehensive survival analysis of MCM4 using stratified and joint effects survival analysis suggests that MCM4 may be an independent prognostic indicator for PDAC OS. Gene set enrichment analysis indicated that MCM4 may participate in multiple biologic processes and pathways, including DNA replication, cell cycle, tumor protein p53, and Notch signaling pathways, thereby affecting prognosis of PDAC patients.Conclusions: Our study indicates that MCM2–7 were upregulated in pancreatic cancer tumor tissues, and mRNA expression of MCM4 may serve as an independent prognostic indicator for PDAC OS prediction after pancreaticoduodenectomy. Keywords: minichromosome maintenance, mRNA, pancreatic ductal adenocarcinoma, prognosis, pancreaticoduodenectomy

Xiwen Liao,1 Xiangkun Wang,1 Ketuan Huang,1 Chengkun Yang,1 Tingdong Yu,1 Chuangye Han,1 Guangzhi Zhu,1 Hao Su,1 Rui Huang,2 Tao Peng1 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China; 2Department of Hematology, The First Affiliated Hospital of Guangxi Medical University, Nanning 530021, Guangxi Zhuang Autonomous Region, People’s Republic of China Background: The aim of the study was to investigate potential prognostic microRNA (miRNA) biomarkers for patients with early stage pancreatic ductal adenocarcinoma (PDAC) after pancreaticoduodenectomy using a miRNA-sequencing (miRNA-seq) data set from The Cancer Genome Atlas (TCGA). A miRNA expression-based prognostic signature was generated, and the potential role of target genes in overall survival (OS) in patients with PDAC was examined.Methods: A miRNA-seq data set of 112 PDAC patients who underwent pancreaticoduodenectomy was obtained from TCGA. Survival analysis was performed to identify potential prognostic biomarkers. Results: Eleven miRNAs (hsa-mir-501, hsa-mir-4521, hsa-mir-5091, hsa-mir-24-1, hsa-mir-126, hsa-mir-30e, hsa-mir-3157, hsa-let-7a-3, hsa-mir-133a-1, hsa-mir-4709, and hsa-mir-421) were used to construct a prognostic signature using the step function. The 11-miRNA prognostic signature showed good performance for prognosis prediction (adjusted P

Tingdong Yu, Xiangkun Wang, Guangzhi Zhu, Chuangye Han, Hao Su, Xiwen Liao, Chengkun Yang, Wei Qin, Ketuan Huang, Tao Peng Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, China Objective: The activities of four cytochrome P3A (CYP3A) subfamily members (CYP3A4, CYP3A5, CYP3A7, and CYP3A43) are well documented in drug metabolism. However, the association between CYP3A subfamily members and hepatocellular carcinoma (HCC) remains unclear. This study investigated the prognostic value of CYP3A subfamily mRNA expression levels with HCC prognosis. Materials and methods: Data from a total of 360 HCC patients were retrieved from The Cancer Genome Atlas database, and data from 231 HCC patients were retrieved from the Gene Expression Omnibus database. Kaplan–Meier analysis and Cox regression models were utilized to determine median survival, overall survival, and recurrence-free survival. Hazard ratios and 95% CI were calculated. Results: Low expression of CYP3A4, CYP3A5, and CYP3A43 in the tumor tissue was associated with short median survival (crude p=0.004, 0.001, and 0.001; adjusted p=0.022, 0.005, and 0.013, respectively). Joint-effects combination analysis of CYP3A4, CYP3A5/CYP3A4, CYP3A43/CYP3A5, and CYP3A43 revealed that high expression groups of two genes (group C, group c, group 3) were associated with a reduced risk of death, as compared to low expression of two genes (group A, group a, group 1), and the adjusted p values were 0.001, 0.004, and 0.001, respectively. Joint-effects analysis of CYP3A4, CYP3A5, and CYP3A43 showed that groups III and IV had a reduced risk of death, as compared to group I (adjusted p=0.024 and 0.002, respectively). Conclusion: CYP3A4, CYP3A5, and CYP3A43 mRNA expression levels are potential prognostic markers of HCC. Keywords: mRNA expression, CYP3A subfamily, hepatocellular carcinoma, prognosis, biomarker

Xinping Ye,* Xiangkun Wang,* Liming Shang, Guangzhi Zhu, Hao Su, Chuangye Han, Wei Qin, Guanghui Li, Tao Peng Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Province, China *These authors contributed equally to this work Objective: Acetaldehyde dehydrogenase 2 (ALDH2) and cytochrome P450 2E1 (CYP2E1) have been associated with hepatocellular carcinoma (HCC) susceptibility and prognosis. The polymorphisms ALDH2 rs671 and CYP2E1 rs2031920 are reportedly correlated with the prevalence of HCC in other countries. The aim of this study was to investigate associations between ALDH2 and CYP2E1, and HCC susceptibility in a population of Guangxi, southern China, an area with a high incidence of HCC.Patients and methods: The study cohort included 300 HCC cases, 292 healthy controls for HCC susceptibility analysis, and another 20 HCC cases and 10 healthy controls for ascertainment. Genotyping was performed using the polymerase chain reaction-restriction fragment length polymorphism method.Results: The study results demonstrated that mutant genotypes of ALDH2 (G/A and A/A) led to significant differences in HCC susceptibility, as compared with the wild genotype (G/G) with the same C1/C1 genotype in non-drinking individuals (adjusted P=0.010, OR=0.20, 95% CI=0.06–0.68). The mutant genotypes of CYP2E1 (C1/C2 and C2/C2) brought about significant differences in HCC susceptibility, as compared with the wild genotype (C1/C1) and the same G/G genotype (adjusted P=0.025, OR=0.42, 95% CI=0.20–0.90). Drinking plays a role in HCC susceptibility in the same G/G genotype individuals (adjusted P=0.004, OR=0.32, 95% CI=0.15–0.69), but had no impact when combined with CYP2E1 for analysis (all P>0.05).Conclusion: These results suggest that the mutant genotypes of ALDH2 and CYP2E1 may be protective factors for HCC susceptibility in Guangxi province, China. Keywords: hepatocellular carcinoma susceptibility, genetic polymorphism, ALDH2, CYP2E1, Chinese population

Xiwen Liao,1,* Long Yu,1,* Xiaoguang Liu,1,2 Chuangye Han,1 Tingdong Yu,1 Wei Qin,1 Chengkun Yang,1 Guangzhi Zhu,1 Hao Su,1 Tao Peng1 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, People’s Republic of China; 2Department of Hepatobiliary Surgery, Affiliated Hospital of Guangdong Medical University, Zhanjiang, People’s Republic of China *These authors contributed equally to this work Background: The aim of this investigation was to identify candidate single nucleotide polymorphisms (SNPs) and molecular pathways associated with tumor protein p53 (TP53) expression status in hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC), clarify their potential mechanisms, and generate SNP-to-gene to pathway hypothesis. Materials and methods: Identify candidate Causal SNPs and Pathways (ICSNPathway) was used to perform pathway analysis based on the results of our previous genome-wide association study of TP53 expression status in 387 HBV-related HCC patients. Results: Through the ICSNPathway analysis, we identified 18 candidate SNPs and 10 candidate pathways that are associated with TP53 expression status in HBV-related HCC. The strongest mechanism involved the modulation of major histocompatibility complex, class II, DP beta 1 (human leukocyte antigen [HLA]-DPB1-rs1042153), major histocompatibility complex, class II, DQ beta 1 (HLA-DQB1-rs1130399, HLA-DQB1-rs1049056, HLA-DQB1-rs1049059, and HLA-DQB1-rs1049060), and major histocompatibility complex, class II, DR beta 1 (HLA-DRB1-rs35445101). SNPs consequently affected regulatory roles in all the candidate pathways except hematopoietic cell lineage pathways. Association analysis using the GSE14520 data set, Gene Multiple Association Network Integration Algorithm, and Search Tool for the Retrieval of Interacting Genes/Proteins suggests that all genes of the candidate SNPs were associated with TP53. Survival analysis showed that the collagen type VI alpha 3 chain (COL6A3) rs111231885 and COL6A3-rs113155945 and COL6A3 block 4 CC haplotypes with TP53 negative status may have protective effects in HBV-related HCC patients after hepatectomy. Conclusion: Our pathway analysis identified 18 candidate SNPs and 10 candidate pathways that were associated with TP53 expression status in HBV-related HCC. Among these candidate SNPs, the genetic variation of COL6A3 may be a potential prognostic biomarker of HBV-related HCC. Keywords: TP53, hepatocellular carcinoma, hepatitis B virus, genome-wide association study, pathway analysis

Xiaoguang Liu,* Chuangye Han,* Xiwen Liao, Long Yu, Guangzhi Zhu, Hao Su, Wei Qin, Sicong Lu, Xinping Ye, Tao Peng Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Zhuang Autonomous Region, China *These authors contributed equally to this work Background: The upregulated expression of versican (VCAN) promotes the proliferation, invasion, and metastasis of various types of human cancer cells, including hepatocellular carcinoma (HCC) cells. Patients and methods: In this study, genetic variants in the exon region of VCAN were genotyped by DNA sequencing. Prognostic values of VCAN exon single nucleotide polymorphisms (SNPs) were assessed by Kaplan–Meier with the log-rank test, and uni- and multivariate Cox proportional hazard regression model. Results: A total of 111 patients with resected hepatitis B virus-associated early-stage HCC were collected for genotyping VCAN exon SNPs using Sanger DNA sequencing. Haplotype analysis was performed using Haploview 4.2. Survival data were analyzed using Kaplan–Meier curves and Cox proportional hazards regression analyses. The rs2652098, rs309559, rs188703, rs160278, and rs160277 SNPs were significantly associated with overall patient survival (p

Xiwen Liao,1 Guangzhi Zhu,1 Rui Huang,2 Chengkun Yang,1 Xiangkun Wang,1 Ketuan Huang,1 Tingdong Yu,1 Chuangye Han,1 Hao Su,1 Tao Peng1 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, People’s Republic of China; 2Department of Hematology, The First Affiliated Hospital of Guangxi Medical University, Nanning, People’s Republic of China Background: The aim of the present study was to identify potential prognostic microRNA (miRNA) biomarkers for hepatocellular carcinoma (HCC) prognosis prediction based on a dataset from The Cancer Genome Atlas (TCGA). Materials and methods: A miRNA sequencing dataset and corresponding clinical parameters of HCC were obtained from TCGA. Genome-wide univariate Cox regression analysis was used to screen prognostic differentially expressed miRNAs (DEMs), and multivariable Cox regression analysis was used for prognostic signature construction. Comprehensive survival analysis was performed to evaluate the prognostic value of the prognostic signature. Results: Five miRNAs were regarded as prognostic DEMs and used for prognostic signature construction. The five-DEM prognostic signature performed well in prognosis prediction (adjusted P < 0.0001, adjusted hazard ratio = 2.249, 95% confidence interval =1.491–3.394), and time-dependent receiver–operating characteristic (ROC) analysis showed an area under the curve (AUC) of 0.765, 0.745, 0.725, and 0.687 for 1-, 2-, 3-, and 5-year HCC overall survival (OS) prediction, respectively. Comprehensive survival analysis of the prognostic signature suggests that the risk score model could serve as an independent factor of HCC and perform better in prognosis prediction than other traditional clinical indicators. Functional assessment of the target genes of hsa-mir-139 and hsa-mir-5003 indicates that they were significantly enriched in multiple biological processes and pathways, including cell proliferation and cell migration regulation, pathways in cancer, and the cyclic adenosine monophosphate (cAMP) signaling pathway. Conclusion: Our study indicates that the novel miRNA expression signature may be a potential prognostic biomarker for HCC patients. Keywords: miRNA, TCGA, hepatocellular carcinoma, prognosis, biomarker

Chengkun Yang,1,* Hao Su,1,* Xiwen Liao,1 Chuangye Han,1 Tingdong Yu,1 Guangzhi Zhu,1 Xiangkun Wang,1 Cheryl Ann Winkler,2 Stephen J O’Brien,3–5 Tao Peng1 1Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Province, China; 2Basic Research Laboratory, CCR, NCI, Leidos Biomedical Research, Frederick National Laboratory, Frederick, MD, USA; 3Laboratory of Genomic Diversity, National Cancer Institute, NIH, Frederick, MD, USA; 4Theodosius Dobzhansky Center for Genome Bioinformatics, St. Petersburg State University, St Petersburg, Russia; 5Guy Harvey Oceanographic Center, Halmos College of Natural Sciences and Oceanography, Nova Southeastern University, Fort Lauderdale, FL, USA *These authors contributed equally to this work Abstract: Hepatocellular carcinoma (HCC) is the most frequent malignancy of the liver. Transforming growth factor beta 1 (TGFB1) and marker of proliferation Ki-67 (MKI67) regulate cell proliferation, differentiation, and growth. The association between MKI67 and TGFB1 expression and its clinical implications in HCC remain unknown.Methods: Public databases were used to analyze TGFB1 and MKI67 expression in different pathologic grades/stages and tissue types of HCC. The association between MKI67 and TGFB1 expression was explored using pathway analysis and in a HepG2 cell line treated with TGFB1. Survival analysis was performed to evaluate the prognostic value of TGFB1 and MKI67 expression in patients with hepatitis B virus (HBV)-related HCC. Results: We identified that MKI67 expression was upregulated in liver cancer tissues. MKI67 and TGFB1 expression levels were different in various stages and tissue types of liver cancer. Furthermore, MKI67 expression was associated with TGFB1 expression in liver cancer tissues and HepG2 cells. Patients with HBV-related HCC and a higher level of MKI67 expression had a worse prognosis. Moreover, a nomogram was conducted to predict the clinical outcomes of patients with HBV-related HCC. Conclusion: MKI67 expression level was associated with TGFB1 expression in liver cancer tissues and a HepG2 cell line. MKI67 expression level can predict the clinical outcomes of patients with HBV-related HCC. Keywords: MKI67, TGFB1, HBV-related HCC, nomogram

Abstract Fibrosis-related events play a part in most blinding diseases worldwide. However, little is known about the mechanisms driving this complex multifactorial disease. Here we have carried out the first genome-wide RNA-Sequencing study in human conjunctival fibrosis. We isolated 10 primary fibrotic and 7 non-fibrotic conjunctival fibroblast cell lines from patients with and without previous glaucoma surgery, respectively. The patients were matched for ethnicity and age. We identified 246 genes that were differentially expressed by over two-fold and p

Abstract The myocardin-related transcription factor/serum response factor (MRTF/SRF) pathway represents a promising therapeutic target to prevent fibrosis. We have tested the effects of new pharmacological inhibitors of MRTF/SRF signalling in a preclinical model of fibrosis. CCG-222740, a novel MRTF/SRF inhibitor, markedly decreased SRF reporter gene activity and showed a greater inhibitory effect on MRTF/SRF target genes than the previously described MRTF-A inhibitor CCG-203971. CCG-222740 was also five times more potent, with an IC50 of 5 μM, in a fibroblast-mediated collagen contraction assay, was less cytotoxic, and a more potent inhibitor of alpha-smooth muscle actin protein expression than CCG-203971. Local delivery of CCG-222740 and CCG-203971 in a validated and clinically relevant rabbit model of scar tissue formation after glaucoma filtration surgery increased the long-term success of the surgery by 67% (P

There is a need to prolong drug residence time using a biocompatible formulation in the subconjunctival space after surgery to treat glaucoma. Drug releasing discs were prepared with 2-(hydroxyethyl)methacrylate (HEMA) and 2-methacryloyl-oxyethyl phosphorylcholine (MPC). The ratio of bound water (Wb) to free water (Wf) ratio increased from 1:0.3 to 1:6.8 with increasing MPC (0 to 50%, w/w). The optimal balance between water content, SR and mechanical strength were obtained with 10% MPC (w/w) hydrogels. Water-alcohol mixtures were examined to facilitate loading of poorly soluble drugs, and they showed greater hydrogel swelling than either water or alcohol alone. The SR was 1.2 ± 0.02 and 3.3 ± 0.1 for water and water:ethanol (1:1) respectively. HEMA-MPC (10%) discs were loaded with dexamethasone using either water:ethanol (1:1) or methanol alone. Drug release was examined in an outflow rig model that mimics the subconjunctival space in the eye. Dexamethasone loading increased from 0.3 to 1.9 mg/disc when the solvent was changed from water:ethanol (1:1) to methanol with the dexamethasone half-life (t½) increasing from 1.9 to 9.7 days respectively. These encouraging results indicate that HEMA-MPC hydrogels have the potential to sustain the residence time of a drug in the subconjunctival space of the eye.

Hyunggun Kim,1 George L Britton,1 Tao Peng,1 Christy K Holland,2 David D McPherson,1 Shao-Ling Huang1 1Division of Cardiovascular Medicine, Department of Internal Medicine, The University of Texas Health Science Center at Houston, Houston, TX, USA; 2Division of Cardiovascular Diseases, Department of Internal Medicine, and Department of Biomedical Engineering and Radiology, University of Cincinnati, Cincinnati, OH, USA Abstract: Delayed cerebral vasospasm following subarachnoid hemorrhage causes severe ischemic neurologic deficits leading to permanent neurologic dysfunction or death. Reduced intravascular and perivascular nitric oxide (NO) availability is a primary pathophysiology of cerebral vasospasm. In this study, we evaluated NO-loaded echogenic liposomes (NO-ELIP) for ultrasound-facilitated NO delivery to produce vasodilation for treatment of vasospasm following subarachnoid hemorrhage. We investigated the vasodilative effects of NO released from NO-ELIP both ex vivo and in vivo. Liposomes containing phospholipids and cholesterol were prepared, and NO was encapsulated. The encapsulation and release of NO from NO-ELIP were determined by the syringe/vacuum method and ultrasound imaging. The ex vivo vasodilative effect of NO-ELIP was investigated using rabbit carotid arteries. Arterial vasodilation was clearly observed with NO-ELIP exposed to Doppler ultrasound whereas there was little vasodilative effect without exposure to Doppler ultrasound in the presence of red blood cells. Penetration of NO into the arterial wall was determined by fluorescent microscopy. The vasodilative effects of intravenously administered NO-ELIP in vivo were determined in a rat subarachnoid hemorrhage model. NO-ELIP with ultrasound activation over the carotid artery demonstrated effective arterial vasodilation in vivo resulting in improved neurologic function. This novel methodology for ultrasound-controlled delivery of NO has the potential for therapeutic treatment of vasospasm following subarachnoid hemorrhage. This ultrasound-controlled release strategy provides a new avenue for targeted bioactive gas and therapeutic delivery for improved stroke treatment. Keywords: vasospasm, vasodilation, subarachnoid hemorrhage, nitric oxide, liposomes, ultrasound

BackgroundTrachoma causes blindness through a conjunctival scarring process initiated by ocular Chlamydia trachomatis infection; however, the rates, drivers and pathophysiological determinants are poorly understood. We investigated progressive scarring and its relationship to conjunctival infection, inflammation and transcript levels of cytokines and fibrogenic factors.Methodology/principal findingsWe recruited two cohorts, one each in Ethiopia and Tanzania, of individuals with established trachomatous conjunctival scarring. They were followed six-monthly for two years, with clinical examinations and conjunctival swab sample collection. Progressive scarring cases were identified by comparing baseline and two-year photographs, and compared to individuals without progression. Samples were tested for C. trachomatis by PCR and transcript levels of S100A7, IL1B, IL13, IL17A, CXCL5, CTGF, SPARCL1, CEACAM5, MMP7, MMP9 and CD83 were estimated by quantitative RT-PCR. Progressive scarring was found in 135/585 (23.1%) of Ethiopian participants and 173/577 (30.0%) of Tanzanian participants. There was a strong relationship between progressive scarring and increasing inflammatory episodes (Ethiopia: OR 5.93, 95%CI 3.31-10.6, pConclusions/significanceScarring progressed in the absence of detectable C. trachomatis, which raises uncertainty about the primary drivers of late-stage trachoma. Chronic conjunctival inflammation appears to be central and is associated with enriched expression of pro-inflammatory factors and altered expression of extracellular matrix regulators. Host determinants of scarring progression appear more complex and subtle than the features of inflammation. Overall this indicates a potential role for anti-inflammatory interventions to interrupt progression and the need for trichiasis disease surveillance and surgery long after chlamydial infection has been controlled at community level.

BackgroundTrachomatous trichiasis (TT) needs to be managed to reduce the risk of vision loss. The long-term impact of epilation (a common traditional practice of repeated plucking of lashes touching the eye) in preventing visual impairment and corneal opacity from TT is unknown. We conducted a randomized controlled trial of epilation versus surgery for the management of minor TT (fewer than six lashes touching the eye) in Ethiopia. Here we report the four-year outcome and the effect on vision and corneal opacity.Methodology/ principal findings1300 individuals with minor TT were recruited and randomly assigned to quality trichiasis surgery or repeated epilation using high quality epilation forceps by a trained person with good near vision. Participants were examined six-monthly for two-years, and then at four-years after randomisation. At two-years all epilation arm participants were offered free surgery. At four-years 1151 (88.5%) were re-examined: 572 (88%) and 579 (89%) from epilation and surgery arms, respectively. At that time, 21.1% of the surgery arm participants had recurrent TT; 189/572 (33%) of the epilation arm had received surgery, while 383 (67%) declined surgery and had continued epilating ("epilation-only"). Among the epilation-only group, 207 (54.1%) fully controlled their TT, 166 (43.3%) had minor TT and 10 (2.6%) had major TT (>5 lashes). There were no differences between participants in the epilation-only, epilation-to-surgery and surgery arm participants in changes in visual acuity and corneal opacity between baseline and four-years.Conclusions/ significanceMost minor TT participants randomised to the epilation arm continued epilating and controlled their TT. Change in vision and corneal opacity was comparable between surgery and epilation-only participants. This suggests that good quality epilation with regular follow-up is a reasonable second-line alternative to surgery for minor TT for individuals who either decline surgery or do not have immediate access to surgical treatment.