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Jun Wang,1,* Yulong Li,1,* Chunyu Tan,1,* Jinlian Shao,1 Weitai Tang,2 Quan Kong,3 Wenqianjun Sheng,1 Zhiquan Ding,1 Feng Li,1 Jifeng Piao,1 Dingyi Lv,1 Libin Hu,1 Qinghua Wang,1 Xiaodan Jiang1 1Neuromedicine Center, The National Key Clinical Specialty, The Engineering Technology Research Center of Education Ministry of China on Diagnosis and Treatment of Cerebrovascular Disease, Guangdong Provincial Key Laboratory on Brain Function Repair and Regeneration, The Neurosurgery Institute of Guangdong Province, Guangdong-Hong Kong-Macao Greater Bay Area Center for Brain Science and Brain-Inspired Intelligence, Zhujiang Hospital, Southern Medical University, Guangzhou, 510282, People’s Republic of China; 2Department of Neurosurgery, The People’s Hospital of Luoding & Affiliated Luoding Hospital of Guangdong Medical University, Luoding, 527200, People’s Republic of China; 3Neurosurgery Department of Zengcheng Campus, Nanfang Hospital, Southern Medical University, Guangzhou, People’s Republic of China*These authors contributed equally to this workCorrespondence: Qinghua Wang; Xiaodan Jiang, Email wqh1123@126.com; jiangxiao_dan@163.comBackground: Neuroinflammatory reactions are crucial factors in secondary brain damage following intracerebral hemorrhage (ICH). Although previous studies have shown that IRAK3 is involved in immune responses, the potential effects of IRAK3 on ICH remain unclear.Methods: Collagenase IV–induced ICH mouse model. Western blotting was used to determine the expression of IRAK3 at different time points following ICH. Immunofluorescence was used to investigate the cellular localization of IRAK3. The ICH model was treated with recombinant human IRAK3 (rh-IRAK3) or IRAK3 siRNA via an intracerebroventricular injection. The effect of IRAK3 on ICH mice was assessed by Western blotting and short-term and long-term neurological function evaluation. RNA-seq was performed to explore the mechanism by which IRAK3 promotes inflammation after ICH. The mechanisms of IRAK3 and neuroinflammation will be further investigated by Western blotting, qRT-PCR and immunofluorescence. Recombinant IL-17A was used to investigate the connection between IRAK3 and the NF-κB/IL-17A signaling pathway in vivo and in vitro experiments.Results: The expression of IRAK3 increased, peaking at 24 h, followed by a subsequent decrease after ICH. IRAK3 is mainly expressed in the microglia. RNA-seq analysis revealed 1,797 differentially expressed genes around the perihematomal brain tissue after IRAK3 siRNA treatment, with multiple inflammatory pathways being downregulated. Rh-IRAK3 treatment resulted in upregulation of the levels of inflammatory cytokines around the perihematomal tissue and exacerbated neurological function deficits. Furthermore, IRAK3 siRNA treatment markedly decreased the expression of inflammatory cytokines and microglial activation via the NF-κB/IL-17A signaling pathway. Recombinant IL-17A exacerbated the inflammatory response in vivo and in vitro; however, IRAK3 knockdown reversed this process.Conclusion: IRAK3 aggravates neuroinflammation by activating the NF-κB/IL-17A signaling pathway, thereby exacerbating neurological deficits following ICH. Therefore, inhibition IRAK3 may be a promising approach for treating ICH.Keywords: IRAK3, neuroinflammation, intracerebral hemorrhage, microglia, NF-κB/IL-17A pathway

Weibei Sheng,1 Yaohang Yue,1 Tiantian Qi,1 Haotian Qin,1 Peng Liu,1 Deli Wang,1 Hui Zeng,2 Fei Yu1 1Department of Bone & Joint Surgery, National & Local Joint Engineering Research Center of Orthopaedic Biomaterials, Shenzhen Key Laboratory of Orthopaedic Diseases and Biomaterials Research, Peking University Shenzhen Hospital, Shenzhen, 518036, People’s Republic of China; 2Shenzhen Second People’s Hospital, The First Affiliated Hospital of Shenzhen University, Shenzhen, 518035, People’s Republic of ChinaCorrespondence: Hui Zeng; Fei Yu, Email zenghui_36@163.com; oscarfyu@163.comAbstract: Osteoarthritis (OA) is a chronic degenerative joint disease characterized by the degradation of joint cartilage, subchondral bone sclerosis, synovitis, and structural changes in the joint. Recent research has highlighted the role of various genes in the pathogenesis and progression of OA, with nuclear factor erythroid 2-related factor 2 (NRF2) emerging as a critical player. NRF2, a vital transcription factor, plays a key role in regulating the OA microenvironment and slowing the disease’s progression. It modulates the expression of several antioxidant enzymes, such as Heme oxygenase-1 (HO-1) and NAD(P)H oxidoreductase 1 (NQO1), among others, which help reduce oxidative stress. Furthermore, NRF2 inhibits the nuclear factor kappa-B (NF-κB) signaling pathway, thereby decreasing inflammation, joint pain, and the breakdown of cartilage extracellular matrix, while also mitigating cell aging and death. This review discusses NRF2’s impact on oxidative stress, inflammation, cell aging, and various cell death modes (such as apoptosis, necroptosis, and ferroptosis) in OA-affected chondrocytes. The role of NRF2 in OA macrophages, and synovial fibroblasts was also discussed. It also covers NRF2’s role in preserving the cartilage extracellular matrix and alleviating joint pain. The purpose of this review is to provide a comprehensive understanding of NRF2’s protective mechanisms in OA, highlighting its potential as a therapeutic target and underscoring its significance in the development of novel treatment strategies for OA.Keywords: nuclear factor erythroid 2-related factor 2, osteoarthritis, chondrocytes, inflammation, oxidative stress

Qingyi Liu,1,2 Qiongdan Zhang,1,2 Zhijian Yao,1,2 Gangqiang Yi,2 Yeonseok Kang,3 Yixing Qiu,1,2 Yupei Yang,1,2 Hanwen Yuan,1,2 Ronggeng Fu,2 Wenbing Sheng,1,2 Lidong Cheng,4 Wei Wang,1,2 Huizhen Wang,1,2 Caiyun Peng1,5 1TCM and Ethnomedicine Innovation & Development International Laboratory, School of Pharmacy, Hunan University of Chinese Medicine, Changsha, People’s Republic of China; 2School of Pharmacy, Hunan University of Chinese Medicine, Changsha, People’s Republic of China; 3College of Korean Medicine, Wonkwang University, Jeonbuk, Korea; 4Hunan Yirentang Chinese Herbal Pieces Co., Ltd, Changde, People’s Republic of China; 5Institute of Innovation and Applied Research in Chinese Medicine Hunan University of Chinese Medicine, Changsha, People’s Republic of ChinaCorrespondence: Huizhen Wang; Caiyun Peng, Tel +86-151-1648-1295 ; +86-138-7480-7689, Email whuiz0086@163.com; caiyunpeng@hnucm.edu.cnAbstract: Extracellular vesicles (EVs) are microparticles released from cells in both physiological and pathological conditions and could be used to monitor the progression of various pathological states, including neoplastic diseases. In various EVs, tumor-derived extracellular vesicles (TEVs) are secreted by different tumor cells and are abundant in many molecular components, such as proteins, nucleic acids, lipids, and carbohydrates. TEVs play a crucial role in forming and advancing various cancer processes. Therefore, TEVs are regarded as promising biomarkers for the early detection of cancer in liquid biopsy. However, the currently developed TEV detection methods still face several key scientific problems that need to be solved, such as low sensitivity, poor specificity, and poor accuracy. To overcome these limitations, DNA walkers have emerged as one of the most popular nanodevices that exhibit better signal amplification capability and enable highly sensitive and specific detection of the analytes. Due to their unique properties of high directionality, flexibility, and efficiency, DNA walkers hold great potential for detecting TEVs. This paper provides an introduction to EVs and DNA walker, additionally, it summarizes recent advances in DNA walker-based detection of TEVs (2018– 2024). The review highlights the close relationship between TEVs and DNA walkers, aims to offer valuable insights into TEV detection and to inspire the development of reliable, efficient, simple, and innovative methods for detecting TEVs based on DNA walker in the future. Keywords: EVs, TEVs, DNA walker, detection

Wen Sheng, Shuanghu Fang School of Educational Science, Anhui Normal University, Wuhu, People’s Republic of ChinaCorrespondence: Shuanghu Fang, Email fsh9075@163.comPurpose: The current study examined the association between moral elevation and college students’ sense of meaning of life, along with the potential mediating effects of gratitude and perceived social support on this relationship.Methods: Using the convenience sampling method, the Moral Elevation Scale, the Sense of Meaning of Life Questionnaire, the Gratitude Questionnaire, and the Multidimensional Scale of Perceived Social Support were used to conduct questionnaire surveys on 1088 college students (Mage=19.59 years, SD=1.46). SPSS25.0 data statistics software was used to analyze the data, and the PROCESS macro for SPSS was used to conduct the mediation effect test.Results: (1) Moral elevation was significantly and positively associated with college students’ sense of meaning of life (β = 0.43,p < 0.001). (2) Both gratitude and perceived social support partially mediated the relation between moral elevation and sense of meaning of life. The indirect effect of moral elevation on sense of meaning of life via gratitude was 0.11 (95% CI = [0.0709, 0.1453]), and the mediating effect accounted for 52.71%. The indirect effect of moral elevation on sense of meaning of life via perceived social support was 0.06 (95% CI = [0.0385, 0.0863]), and the mediating effect accounted for 29.41%. (3) Gratitude and perceived social support sequentially mediated the association between moral elevation and sense of meaning of life, the indirect effect through gratitude and perceived social support was 0.04 (95% CI = [0.0237, 0.0512]) and accounted for 17.88%.Conclusion: This study revealed the chain mediating roles of gratitude and perceived social support in the relation between moral elevation and sense of meaning of life for Chinese college students, which is of great theoretical and practical significance for the formation of a positive sense of meaning of life and the cultivation of moral elevation among college students.Keywords: moral elevation, sense of meaning of life, gratitude, perceived social support, college students

Parhat Yasin,1 Muradil Mardan,2 Dilxat Abliz,3 Tao Xu,1 Nuerbiyan Keyoumu,4 Abasi Aimaiti,4 Xiaoyu Cai,1 Weibin Sheng,1 Mardan Mamat1 1Department of Spine Surgery, The First Affiliated Hospital of Xinjiang Medical University, Urumqi, Xinjiang, 830054, People’s Republic of China; 2School of Medicine, Tongji University, Shanghai, 200092, People’s Republic of China; 3Department of Orthopedic, The Eighth Affiliated Hospital of Xinjiang Medical University, Urumqi, Xinjiang, 830054, People’s Republic of China; 4Department of Anesthesiology, The First Affiliated Hospital of Xinjiang Medical University, Urumqi, Xinjiang, 830054, People’s Republic of ChinaCorrespondence: Mardan Mamat, Tel +86 0991-4365316, Email mardanmmtmx@163.comBackground: Pyogenic spondylitis (PS) and Brucella spondylitis (BS) are common spinal infections with similar manifestations, making their differentiation challenging. This study aimed to explore the potential of CT-based radiomics features combined with machine learning algorithms to differentiate PS from BS.Methods: This retrospective study involved the collection of clinical and radiological information from 138 patients diagnosed with either PS or BS in our hospital between January 2017 and December 2022, based on histopathology examination and/or germ isolations. The region of interest (ROI) was defined by two radiologists using a 3D Slicer open-source platform, utilizing blind analysis of sagittal CT images against histopathological examination results. PyRadiomics, a Python package, was utilized to extract ROI features. Several methods were performed to reduce the dimensionality of the extracted features. Machine learning algorithms were trained and evaluated using techniques like the area under the receiver operating characteristic curve (AUC; confusion matrix-related metrics, calibration plot, and decision curve analysis to assess their ability to differentiate PS from BS. Additionally, permutation feature importance (PFI; local interpretable model-agnostic explanations (LIME; and Shapley additive explanation (SHAP) techniques were utilized to gain insights into the interpretabilities of the models that are otherwise considered opaque black-boxes.Results: A total of 15 radiomics features were screened during the analysis. The AUC value and Brier score of best the model were 0.88 and 0.13, respectively. The calibration plot and decision curve analysis displayed higher clinical efficiency in the differential diagnosis. According to the interpretation results, the most impactful features on the model output were wavelet LHL small dependence low gray-level emphasis (GLDN).Conclusion: The CT-based radiomics models that we developed have proven to be useful in reliably differentiating between PS and BS at an early stage and can provide a reliable explanation for the classification results.Keywords: Brucella spondylitis, Pyogenic spondylitis, machine learning, radiomics, model interpretation

Rui Zhang,1 Weixuan Sheng,2 Feiran Liu,1 Jin Zhang,1 Wenpei Bai1 1Department of Obstetrics and Gynaecology, Beijing Shijitan Hospital, Capital Medical University, Beijing, People’s Republic of China; 2Department of Anesthesiology, Beijing Shijitan Hospital, Capital Medical University, Beijing, People’s Republic of ChinaCorrespondence: Wenpei Bai, Email baiwp@bjsjth.cnObjective: This study aimed to investigate the risk factors of cesarean section and establish a prediction model for cesarean section based on the characteristics of pregnant women.Methods: The clinical characteristics of 2552 singleton pregnant women who delivered a live baby between January 2020 and December 2021 were retrospectively reviewed. They were divided into vaginal delivery group (n = 1850) and cesarean section group (n = 702). These subjects were divided into training set (2020.1– 2021.6) and validation set (2021.7– 2021.12). In the training set, univariate analysis, Lasso regression, and Boruta were used to screen independent risk factors for cesarean section. Four models, including Logistic Regression (LR), K-Nearest Neighbor (KNN), Classification and Regression Tree (CART), and Random forest (RF), were established in the training set using K-fold cross validation, hyperparameter optimization, and random oversampling techniques. The best model was screened, and Sort graph of feature variables, univariate partial dependency profile, and Break Down profile were delineated. In the validation set, the confusion matrix parameters were calculated, and receiver operating characteristic curve (ROC), precision recall curve (PRC), calibration curve, and clinical decision curve analysis (DCA) were delineated.Results: The risk factors of cesarean section included age and height of women, weight at delivery, weight gain, para, assisted reproduction, abnormal blood glucose during pregnancy, pregnancy hypertension, scarred uterus, premature rupture of membrane (PROM), placenta previa, fetal malposition, thrombocytopenia, floating fetal head, and labor analgesia. RF had the best performance among the four models, and the accuracy of confusion matrix parameters was 0.8956357. The Matthews correlation coefficient (MCC) was 0.753012. The area under ROC (AUC-ROC) was 0.9790787, and the area under PRC (AUC-PRC) was 0.957888.Conclusion: RF prediction model for caesarean section has high discrimination performance, accuracy and consistency, and outstanding generalization ability.Keywords: caesarean section, machine learning, confusion matrix, univariate partial dependence profile

Abstract Background and Aim Acute severe ulcerative colitis (ASUC) remains a significant cause of morbidity and healthcare utilization. This study aimed to characterize the total healthcare costs of ASUC, explore factors associated with significant cost over the 12 months following an index admission, and document outcomes including corticosteroid exposure. Methods Patients admitted from January 2016 until January 2021 for ASUC to a tertiary inflammatory bowel disease (IBD) center in Australia were identified via retrospective chart review. Costs were calculated over a 12‐month period following index admission. Results Seventy‐two patients (30 [42%] female, median age 39 [IQR 27–54] years) were included. The median length of stay of index admission was 6 days (IQR 5–10 days). The median cost of index admission was 7829 AUD, which was driven by the initial length of stay (P

Haopeng Luan, Qiang Deng, Weibin Sheng, Mardan Mamat, Hailong Guo, Huaqiang Li Department of Spine Surgery, XinJiang Medical University Affiliated First Hosptial, Urumqi, XinJiang, People’s Republic of ChinaCorrespondence: Qiang DengDepartment of Spine Surgery, Xinjiang Medical University Affiliated First Hospital, Urumqi, XinJiang, People’s Republic of ChinaTel +86 13325636562Email dengqiang_abc@163.comObjective: The present study aimed to investigate the surgical efficacy of staged posterior–anterior combined surgery for the treatment of cervicothoracic segmental tuberculosis (TB) with kyphosis in pediatric patients.Methods: The clinical data of 15 pediatric patients admitted to our hospital from January 2010 to December 2017 who underwent staged posterior–anterior combined surgery for cervicothoracic segmental TB with kyphosis were collected. A posterior median incision was made for patients after general anesthesia. Autologous bone particles or allogeneic bone particles were taken, trimmed, and placed in the articular eminence of the diseased vertebral body. Fifteen pediatric patients underwent second-stage lesion removal using the anterior approach. The left sternocleidomastoid muscle was selected as the medial oblique incision approach. The abscess and caseous necrotic material were removed and sent for pathological examination. The sagittal and coronal parameters (including the local Cobb angle, the sagittal vertical axis [SVA], and the coronal balance distance [CBD]) were measured at three time points: preoperatively, postoperatively, and at the final follow-up. The American Spinal Injury Association’s spinal-cord injury classification, the Japanese Orthopaedic Association’s (JOA) cervical spine function score, the neck disability index (NDI), and the visual analogue score (VAS) for cervicothoracic segment pain were adopted for the assessment of functional improvement and quality of life.Results: All 15 pediatric patients completed the surgery successfully, with an operation duration of 3.56 ± 0.68 h, an intraoperative hemorrhage of 289.7 ± 84.3 mL, an average fixation of 7.3 ± 1.8 segments, and a follow-up duration of 28.1 ± 9.7 months. The preoperative and postoperative sagittal local Cobb angle was 67.06 ± 17.54° vs 19.48 ± 2.32° (P < 0.01), the SVA was 35.19 ± 10.69 mm vs 7.67 ± 1.40 mm (P < 0.01), and CBD was 22.58 ± 7.59 mm vs 8.99 ± 1.25 mm (P < 0.01). The levels of the postoperative erythrocyte sedimentation rate and C-reactive protein were significantly lower in all patients. The preoperative and postoperative JOA scores were 8.93 ± 3.51 and 14.67 ± 1.34, respectively, the preoperative and postoperative VAS was 7.40 ± 1.35 and 2.67 ± 0.62, respectively, and the preoperative and postoperative NDI was 32.67 ± 4.83 and 13.73 ± 2.08, respectively. There were statistically significant differences in the above indicators before and after surgery (P < 0.05).Conclusion: In the surgical treatment of cervicothoracic TB with kyphosis in pediatric patients, staged posterior–anterior combined surgery significantly corrects deformity, achieves the safe and effective neurological decompression of the spinal cord, and obtains good neurological recovery and bone-graft fusion according to the extent of the involved segments of kyphosis, the characteristics of the lesion, and the degree of neurospinal injury.Keywords: spinal tuberculosis, cervicothoracic segment, posterior and anterior combined surgery, pediatric patient, therapeutic effect

Wenlong Sheng,1 Shijun Weng,2 Fei Li,1 Yun Zhang,1 Qiuxia He,1 Wenxiang Sheng,1 Ying Fu,3 Haiyue Yan,4 Kechun Liu1 1Biology Institute, Qilu University of Technology (Shandong Academy of Sciences), Jinan, People’s Republic of China; 2State Key Laboratory of Medical Neurobiology, Institutes of Brain Science, Department of Ophthalmology, Zhongshan Hospital, Fudan University, Shanghai, People’s Republic of China; 3Shandong Science and Technology Exchange Center, Jinan, People’s Republic of China; 4Shandong Institute of Scientific and Technical Information, Jinan, People’s Republic of ChinaCorrespondence: Wenlong Sheng; Kechun Liu Email shengwenlong1121@163.com; liukechun2000@163.comBackground: Melatonin (N-acetyl-5-methoxytryptamine), a significant indoleamine neuromodulator implicated in circadian rhythms and sleep patterns, regulates diverse rhythmic functions via activating its high-affinity G-protein-coupled receptors. However, the detailed cellular expression of the Mel1a receptor in the retina is still a research gap.Methods: The expression of the Mel1a receptor in pigeon retina was assessed using Western blot analysis and immunofluorescent staining. The cellular localization of the Mel1a receptor was studied using double immunofluorescent staining and laser-scanning confocal microscopy.Results: Our data suggested that the Mel1a receptor was extensively expressed in the outer segment of Rho4D2-labeled rod and L/M-opsin-labeled red/green cone and in the somata of the CB-labeled horizontal cell, TH-labeled dopaminergic amacrine cell, ChAT-labeled cholinergic amacrine cell, PV-labeled AII amacrine cell, Brn3a-labeled conventional ganglion cell, melanopsin-containing ganglion cell and CRALBP-labeled Müller glial cell. In addition, the Mel1a receptor was diffusely distributed throughout the full thickness of the inner plexiform layer. However, the outer segment of S-opsin-labeled blue cone, the somata of ChX-10-labeled bipolar cell and outer plexiform layer seemed to lack immunoreactivity of the Mel1a receptor.Conclusion: The finding that multiple types of retinal cells express the Mel1a receptor provides a new neurobiological basis for the participation of melatonin in the regulation of retinal functions through activating the Mel1a receptor.Keywords: circadian rhythm, retinal Mel1a receptor, cellular localization, diurnal animal

Bin Wang,* Siyuan Jiang,* Lizhe Zhu, Wei Sheng, Yan Qiao, Huimin Zhang, Jian Zhang, Yang Liu, Na Hao, Xiaoxia Ma, Can Zhou, Yu Ren Department of Breast Surgery, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, People’s Republic of China*These authors contributed equally to this workCorrespondence: Can Zhou; Yu RenDepartment of Breast Surgery, The First Affiliated Hospital of Xi’an Jiaotong University, 277 West Yanta Road, Xi’an, Shaanxi, People’s Republic of ChinaTel +86 13700222161Email zhoucanz2005@126.com ; renyyyyy@126.comIntroduction: Nipple discharge is a common symptom of breast disease. We aimed to perform a descriptive statistical analysis of the cases we evaluated and establish a model to predict intraductal tumors.Materials and Methods: We conducted a retrospective study of patients from 2007 to 2019. In total, 1333 patients who completed the fiberoptic ductoscopy (FDS) were evaluated. The variables were analyzed by χ2 test. Logistic regression was used to analyze the relationship between the patient’s clinical characteristics and intraductal tumors and establish a predictive model. Receiver operating characteristic (ROC) curve analysis was used to assess the sensitivity and specificity of the predictive ability of the model. Calibration curves and decision curve analysis (DCA) were used to evaluate the model.Results: Patients with spontaneous, single-duct, bloody discharge and a smooth ductal wall were more likely to be diagnosed with tumors by ductoscopy. A model was established based on five variables: age, side of discharge, spontaneous discharge status, duration of discharge, and color of discharge. The model was subsequently validated in 183 patients with complete data on the variables in the validation cohort. The area under the ROC curve (AUC) was calculated to be 0.716, indicating good predictive ability.Conclusion: Patients with the clinical characteristics of unilateral, bloody, single-duct, spontaneous discharge and a smooth ductal wall were more likely to have intraductal tumors by ductoscopy. Our nomogram can effectively predict intraductal tumors in patients with nipple discharge.Keywords: nipple discharge, ductoscopy, nomogram, breast tumor

Fei Ren,1,2 Weiwei Weng,1,2 Qiongyan Zhang,1,2 Cong Tan,1,2 Midie Xu,1,2 Meng Zhang,1,2 Lei Wang,1,2 Weiqi Sheng,1,2 Shujuan Ni,1,2 Dan Huang1,21Department of Pathology, Fudan University Shanghai Cancer Center, Shanghai, People’s Republic of China; 2Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, People’s Republic of ChinaBackground: High serum levels of alpha-fetoprotein (AFP) are observed in some gastrointestinal cancers. However, primary AFP-producing colorectal cancer (CRC) is extremely rare and causes confusion among clinicians. In this study, we analyzed the clinicopathological features and clinical outcomes of AFP-producing CRC and provide a brief view of this rare carcinoma.Patients and methods: Twenty patients with AFP-producing CRC were enrolled at the Fudan University Shanghai Cancer Center from 2012 to 2015. Clinical information, including serum AFP and CEA levels, and outcomes were collected. Tumors were divided into three histologic types: the common adenocarcinoma (COM) type, mucinous adenocarcinoma type and hepatoid type (HPT). Immunohistochemical (IHC) staining of GPC3, Hepa-1, SALL4 and Arg-1 was performed. Additionally, mutations of the KRAS, NRAS and BRAF genes were examined. Finally, another 40 stage-matched patients with traditional CRC were enrolled as controls for survival analysis.Results: AFP-producing CRC was more likely to occur in males (60%) and arose mainly from the ascending (40%) and sigmoid (35%) colon. In addition, the majority of patients with AFP-producing CRC had poor differentiation (50%), advanced local invasion (80%) and lymph node (LN) metastasis (60%). Synchronous distant metastasis was commonly observed (35%). Interestingly, serum AFP levels were closely associated with LN metastasis. Histopathologically, the COM type was the most common pattern. In IHC staining, the HPT pattern was the most distinct due to high positivity rates of GPC3, Hepa-1 and Arg-1. One patient had mismatch repair deficiency, and another had a KRAS mutation. Patients with AFP-producing CRC had worse progression-free and overall survival than patients with traditional CRC.Conclusion: AFP-producing CRC has unique clinical and histopathological characteristics, showing an aggressive biological behavior and worse prognosis than traditional CRC.Keywords: AFP-producing colorectal cancer, clinicopathological features, serum AFP, prognosis

Qiongyan Zhang,1–3,* Lei Wang,1,2,* Dan Huang,1,2 Midie Xu,1,2 Weiwei Weng,1,2 Shujuan Ni,1,2 Cong Tan,1,2 Weiqi Sheng1,2 1Department of Pathology, Fudan University Shanghai Cancer Center, Shanghai 200032, China; 2Department of Oncology, Shanghai Medical College, Fudan University, Shanghai 200032, China; 3Department of Pathology, Zhongshan Hospital, Fudan University, Shanghai 200032, China *These authors contributed equally to this work Background: Risk grade assessment determines therapy in patients with submucosal invasive colorectal carcinoma (CRC). However, treatment decisions are often difficult due to a lack of consensus on which risk factors should be considered. We aimed to identify predictive risk factors for lymph node metastasis (LNM) in a large cohort of submucosal invasive CRC patients from China.Patients and methods: Following collection of clinicopathological data and disease-free survival (DFS) rates from 290 patients who underwent radical intestinal resection with regional lymphadenectomy, we immunohistochemically assessed expression of DNA mismatch repair (MMR) proteins and p53. The correlation between clinicopathological parameters, MMR expression, p53 status, and LNM status was determined using chi-squared tests and logistic analysis. Receiver operator characteristic curve analysis was used to compare the predictive values. The DFS curves were plotted using the Kaplan–Meier method.Results: LNM was detected in 15.5% of the cases (45/290 patients). Three pathological characteristics, high tumor differentiation grade, lymphovascular invasion (LVI), and tumor budding, were all positively related to LNM in univariate and multivariate analyses (P0.05). Overexpression of p53 was associated with tumor budding status (P=0.036). With a negative predicative value of 0.92 and area under the curve of 0.76 (95% CI: 0.68–0.85), the combination of these three factors provided optimal predictive ability. Patients with all three risk factors had poorer DFS (P

Peng Qi,1–3 Wan-run Lin,4 Meng Zhang,1–3 Dan Huang,1–3 Shu-juan Ni,1–3 Xiao-li Zhu,1–3 Qian-ming Bai,1–3 Wei-qi Sheng,1–3 Xiang Du,2 Xiao-yan Zhou1–3 1Department of Pathology, Fudan University Shanghai Cancer Center, Shanghai, China; 2Institute of Pathology, Fudan University, Shanghai, China; 3Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China; 4Department of Pathology, University of Texas Southwestern Medical Center, Dallas, TX, USA Background: Long noncoding RNAs (lncRNAs) have been shown to play important regulatory roles in human cancer. We previously verified that the lncRNA long stress-induced noncoding transcript 5 (LSINCT5) is overexpressed in gastric cancer (GC) cells and closely correlated with cell proliferation and patient prognosis. However, whether aberrant LSINCT5 expression has an important effect on GC progression is unclear, and the potential mechanisms remain unknown. In GC, E2F1 expression is also aberrant, but the biological functions of E2F1 are controversial, and the correlation between E2F1 and lncRNAs remains unknown. Materials and methods: Expression of LSINCT5 was analyzed in metastatic GC tissues compared with nonmetastatic tissues using quantitative real-time PCR (qRT-PCR) assays. Gain and loss of function approaches were used to investigate the biological role of LSINCT5 in GC cell migration and invasion. A computational screen of LSINCT5 promoter was conducted to search for transcription factor-binding sites. LSINCT5 promoter activities were examined by ChIP and luciferase reporter assays. qRT-PCR and western blotting assays were performed to detect the expression of multiple EMT markers in cells in which LSINCT5 was overexpressed or knocked down. Results: An integrated quantitative analysis revealed that LSINCT5 was significantly overexpressed in metastatic GC tissues. Forced LSINCT5 expression promoted cell migration and invasion, whereas loss of LSINCT5 function decreased cell migration and invasion. Mechanistic investigations showed that LSINCT5 is a direct transcriptional target of E2F1. Moreover, LSINCT5 overexpression was found to play an important role in the epithelial-to-mesenchymal transition by regulating the expression of E-cadherin, N-cadherin, vimentin, and matrix metalloproteinase-2. Conclusion: These data suggest that E2F1-mediated activation of LSINCT5, a regulator of cell migration and invasion, constitute the mechanistic link between the E2F1-mediated pathway and lncRNA that regulates cell migration and invasion. Thus, LSINCT5 may be a target for new GC therapies. Keywords: E2F1, LSINCT5, metastasis, gastric cancer

Yanni Yu,1 Yongpei Hu,1 Xiufang Li,1 Yu Liu,1 Mingzhong Li,1 Jicheng Yang,2 Weihua Sheng2 1National Engineering Laboratory for Modern Silk, College of Textile and Clothing Engineering, Soochow University, 2Cell and Molecular Biology Institute, College of Medicine, Soochow University, Suzhou, People’s Republic of China Abstract: The development of a novel cationized polymer used as a gene delivery carrier that can conveniently and effectively transfect cells resulting in a stably expressed target gene remains a challenge. Antheraea pernyi silk fibroin (ASF) is a cytocompatible and biodegradable natural polymer, and it possesses Arg–Gly–Asp sequences but a negative charge. In order to render ASF amenable to packaging plasmid DNA (pDNA), spermine was used to modify ASF to synthesize cationized ASF (CASF), which was used as a gene delivery carrier. CASF was characterized using trinitrobenzene sulfonic acid assay, the zeta potential determination, and a Fourier transform infrared analysis, and the results of these characterizations indicated that the –NH2 in spermine effectively reacts with the –COOH in the side chains of ASF. Spermine grafted to the side chains of ASF resulted in the conversion of the negative charge of ASF to a positive charge. CASF packaged pDNA and formed CASF/pDNA complexes, which exhibited spherical morphology with average particle sizes of 215–281 nm and zeta potential of approximately +3.0 mV to +3.2 mV. The results of the MTT assay, confocal laser scanning microscopy, and flow cytometry analysis in a human endothelial cell line revealed that CASF/pDNA complexes exhibited lower cytotoxicity and higher transfection efficiency compared to the pDNA complexes of polyethyleneimine. These results indicate that our synthesized CASF, a cationized polymer, is a potential gene delivery carrier with the advantages of biodegradability and low cytotoxicity. Keywords: silk fibroin, spermine, cationized polymer, gene delivery carrier

Pinellia ternata (Thunb.) Briet. is a well-known traditional Chinese herbal medicine that has many beneficial effects such as anti-tumor, anti-fertility, blood fat-reducing, and liver-protective. The comprehensive utilization value of P. ternata is high and the development prospect is broad. However, low tuber production resulting from "sprout tumble" (ST) due to high temperature and light intensity has largely reduced its use. Therefore, it is necessary to explore the functional gene responses to high temperature and light. Real-time quantitative PCR (RT-qPCR) is commonly used for accurately gene expression detection during gene function exploration. However, because the RT-qPCR results may vary depending on the conditions and environment, an internal reference gene (IRG) is needed to normalize the RT-qPCR data. Analyzing IRGs in P. ternata can help with subsequent functional characterization and verification of the genes regulating P. ternata growth. In this study, we screened 8 suitable IRGs, and determined the most suitable IRGs using the Ct value, GeNorm, NormFinder, and BestKeeper under shade and high-temperature conditions, and in different tissues. The results showed that 18S was a suitable IRG for P. ternata studies. This discovery has laid the foundation for further research on P. ternata, and is also beneficial for the exploration of more IRGs in medicinal plants. [ABSTRACT FROM AUTHOR]

Citrus is highly sensitive to salt stress, and little efforts have been successful microbiologically to mitigate such abiotic stress. In this background, trifoliate orange [Poncirus trifoliata (L.) Raf.] seedlings were inoculated with Diversispora versiformis and exposed to 100 mM NaCl treatment for 85 days under controlled conditions. The NaCl application, though, strongly inhibited root mycorrhizal colonization in seedlings, but mycorrhizal inoculation considerably increased the root projected area and number of second- and third-order lateral roots under 100 mM NaCl treatment. Mycorrhizal-inoculated seedlings showed significantly higher soluble protein concentration, ornithine decarboxylase, arginine decarboxylase, and superoxide dismutase activity in leaves and roots, irrespective of NaCl concentration. While mycorrhizal seedlings displayed significantly lower polyamine oxidase activity and diamine oxidase activity in leaves and roots, irrespective of NaCl concentration. These results, thus, suggested that mycorrhizal plants were physiologically activated through mycorrhizal inoculation to downplay the adverse effect of salt stress.