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Hui Rao,1 Xianfang Wang,2 Yu Luo,3 Liu Liang,1 Wei Ye,1 Xuemin Guo1 1Department of Laboratory Medicine, Meizhou People’s Hospital, Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China; 2Center for Cardiovascular Diseases, Meizhou People’s Hospital, Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China; 3Department of Gynaecology, Meizhou People’s Hospital, Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of ChinaCorrespondence: Xuemin Guo, Department of Laboratory Medicine, Meizhou People’s Hospital, No. 63 Huangtang Road, Meijiang District, Meizhou, People’s Republic of China, Email Guoxm12@126.comBackground: The role of aldehyde dehydrogenase 2 (ALDH2) in cardiovascular diseases has been gradually studied. However, it is unclear whether ALDH2 polymorphism is associated with the risk of early onset (onset age ≤ 55 years old in men and ≤ 65 years old in women) coronary artery stenosis (CAS). The association between ALDH2 single nucleotide polymorphism (SNP) rs671 and risk in patients with early onset CAS was investigated in this study.Methods: The study included 213 early onset CAS patients and 352 individuals without CAS were set as controls. The ALDH2 rs671 polymorphism was genotyped by polymerase chain reaction (PCR) - microarray. Differences in ALDH2 rs671 genotypes and alleles between patients and controls were compared. Multiple logistic regression analysis was performed after adjusting for gender, body mass index (BMI), smoking history, drinking history, and diabetes mellitus to assess the relationship between ALDH2 rs671 genotypes and early onset CAS risk.Results: The frequency of the ALDH2 rs671 G/G genotype was lower in the early onset CAS patients (43.7% vs 55.3%, p=0.007) than that in the controls. The frequency of the ALDH2 rs671 A allele was higher (32.9% vs 25.0%) than that in the controls (p=0.005). After adjusting for other confounding factors, multivariate logistic regression showed that ALDH2 rs671 A/A genotype (A/A vs G/G: odds ratio (OR) 2.508, 95% confidence interval (CI): 1.130– 5.569, p=0.024), overweight (BMI≥ 24.0 vs 18.5– 23.9: OR 5.047, 95% CI: 3.275– 7.777, p< 0.001), history of smoking (yes vs no: OR 2.813, 95% CI: 1.595– 4.961, p< 0.001), and diabetes mellitus (yes vs no: OR 2.191, 95% CI: 1.397– 3.437, p=0.001) were the independent risk factors of early onset CAS.Conclusion: In men ≤ 55 years old and women ≤ 65 years old, individuals with ALDH2 rs671 A/A genotype, overweight (BMI ≥ 24.0 kg/m2), smoking history, and diabetes mellitus increased risk of developing CAS.Keywords: aldehyde dehydrogenase 2, gene polymorphism, coronary artery stenosis, early onset

Hao Rao Hubei University of Chinese Medicine, Wuhan, Hubei, People’s Republic of ChinaCorrespondence: Hao Rao, Email 3290656729@qq.com

Qiang Fu,1 Na Han,1 Na Li,2 Ling Gui,3 Chen Shi,4 Peipei Rong,5 Fan Zeng,1 He Rao,6 Yuan Chen1 On behalf of Cancer Rehabilitation and Palliative Treatment Professional Committee of Hubei Anti Cancer Association1Department of Oncology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, 430030, People’s Republic of China; 2Cancer Center, Renmin Hospital of Wuhan University, Wuhan, Hubei, 430060, People’s Republic of China; 3Department of Pharmacy, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, 430030, People’s Republic of China; 4Department of Pharmacy, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, People’s Republic of China; 5Department of Pharmacy, Renmin Hospital of Wuhan University, Wuhan, Hubei, 430060, People’s Republic of China; 6Department of Oncology, Zhongnan Hospital of Wuhan University, Wuhan, Hubei, 430071, People’s Republic of ChinaCorrespondence: Qiang Fu; Yuan Chen, Department of Oncology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, No. 1095 Jiefang Avenue, Qiaokou District, Wuhan, Hubei, 430030, People’s Republic of China, Email fqiang2016@163.com; chenyuan008@163.comAbstract: Pain is one of the most common clinical symptoms of cancer patients, seriously affecting the quality of life of patients and bringing heavy mental and economic burden to families and society. The treatment of cancer pain in China is facing numerous challenges, one of which includes the irrational usage of analgesic drugs in clinical practice. As a strong opioid analgesic, transdermal fentanyl patch has been widely used due to its convenient clinical application and obvious therapeutic effect. Several basic-level hospitals and even general hospitals in China fail to appropriate the application of drugs in clinical application due to the lack of understanding of the pharmacological characteristics and clinical application of fentanyl transdermal patch by medical staff, seriously affecting the treatment quality. Therefore, it is imperative to strengthen the rational use and management of fentanyl transdermal patches. Accordingly, the initiation by the Cancer Rehabilitation and Palliative Treatment Professional Committee of the Hubei Anti-cancer Association launched the compilation of the “Guidelines for Rational Clinical Use of Fentanyl Transdermal Patch” (from now on referred to as the “Guidelines”) in Hubei Province, China. The experts in the preparation group are experts in many disciplines, such as medicine, pharmacy, and nursing. The expert group determines the outline, prepares the required regulations, and revises it repeatedly. Moreover, these experts put forward suggestions for revision to strictly control the accuracy and scientific authenticity of the contents of the “Guide”. Finally, all experts of the preparation team certify and finalize the draft. This “Guide” prepared by experts of the Cancer Rehabilitation and Palliative Treatment Professional Committee of the Hubei Anti-cancer Association and the expert advisory group with joint efforts, aims to play a positive role in promoting the rational clinical use of fentanyl transdermal patch, reducing the mental and economic burden of patients, and ensuring medical quality and medical safety.Keywords: fentanyl transdermal patch, reasonable application, medication guidelines

Yu Luo,1,2 Ru Pan,1,2 Hui Rao,2,3 Xing Chen,4 Haikun Yang1,2 1Department of Gynaecology, Meizhou People’s Hospital, Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China; 2Meizhou Municipal Engineering and Technology Research Center for Molecular Diagnostics of Major Genetic Disorders, Meizhou People’s Hospital, Meizhou, People’s Republic of China; 3Department of Laboratory Medicine, Meizhou People’s Hospital, Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China; 4Data Center, Meizhou People’s Hospital, Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of ChinaCorrespondence: Haikun Yang, Department of Gynaecology, Meizhou People’s Hospital, Meizhou Academy of Medical Sciences, Add: No. 63 Huangtang Road, Meijiang District, Meizhou, People’s Republic of China, Email 13923023911@139.comObjective: To investigate the relationship between BRCA1/2 gene mutation and clinicopathological features in ovarian cancer patients, so as to develop precise individualized treatment plan for patients.Methods: Patients diagnosed with ovarian cancer between January 2018 and July 2023 who underwent BRCA1/2 genetic testing were retrospectively analyzed. The clinicopathological characteristics (age, body mass index (BMI), family history of ovarian cancer, pregnancy history, menopause status, tumor size, histopathology, Federation of Gynecology and Obstetrics (FIGO) staging, and ascites) of non-carriers and BRCA1/2 variant carriers were compared. Logistic regression analysis was used to explore the relationship between BRCA1/2 variants and clinicopathological characteristics of ovarian cancer.Results: A total of 284 ovarian cancer patients were collected, and the subjects were divided into two groups, 197 non-carriers and 87 BRCA1/2 variants carriers. The proportion of serous ovarian carcinoma in BRCA1/2 variant carriers is higher than that in non-BRCA variant carriers (78.2% vs 60.9%, p=0.015). There were 51 patients with BRCA pathogenic or likely pathogenic variant, 22 patients with BRCA likely benign variant, and 14 patients with BRCA variants of uncertain significance (VUS). The proportion of serous ovarian carcinoma in patients with BRCA pathogenic/likely pathogenic variant is higher than that in patients with BRCA likely benign variant and BRCA VUS (94.1% vs 50.0% and 64.3%. p< 0.001). There were no statistically significant differences in BMI, family history of ovarian cancer, pregnancy history, menopause status, maximum diameter of the tumor lesion, FIGO stage, and ascites among patients with different grades of variants. Multivariate logistic regression analysis showed that serous ovarian carcinoma was related to BRCA mutation (Serous carcinoma vs non-serous carcinoma: OR 2.145, 95% CI: 1.044– 4.407) (p=0.038).Conclusion: Patients with BRCA1 variant develop ovarian cancer at a younger age than those with the BRCA2 variant. The proportion of FIGO stage III–IV in patients with BRCA pathogenic + likely pathogenic variant was significantly higher than those in patients with other variants. Germline BRCA1/2 variants were most frequently identified in serous ovarian carcinoma patients.Keywords: ovarian cancer, BRCA1, BRCA2, clinicopathological characteristics

Tianxin Mao,1,2 Ya Chai,3 Bowen Guo,1 Peng Quan,3,4 Hengyi Rao1,3,5 1Center for Magnetic Resonance Imaging Research & Key Laboratory of Brain-Machine Intelligence for Information Behavior (Ministry of Education and Shanghai), School of Business and Management, Shanghai International Studies University, Shanghai, People’s Republic of China; 2School of Psychology, South China Normal University, Guangzhou, People’s Republic of China; 3Center for Functional Neuroimaging, Department of Neurology, University of Pennsylvania, Philadelphia, PA, USA; 4Research Center for Quality of Life and Applied Psychology, Guangdong Medical University, Dongguan, People’s Republic of China; 5Division of Sleep and Chronobiology, Department of Psychiatry, University of Pennsylvania, Philadelphia, PA, USACorrespondence: Hengyi Rao, Center for Functional Neuroimaging & Department of Neurology, University of Pennsylvania Perelman School of Medicine, Room D502, Richards Medical Research Building, 3700 Hamilton Walk, Philadelphia, PA, 19104-4202, USA, Email hengyi@gmail.comPurpose: Many studies have investigated the cognitive, emotional, and other impairments caused by sleep restriction. However, few studies have explored the relationship between cognitive performance and changes in sleep structure and electroencephalography (EEG) during sleep. The present study aimed to examine whether changes in sleep structure and EEG can account for cognitive impairment caused by sleep restriction.Patients and Methods: Sixteen young adults spent five consecutive nights (adaptation 9h, baseline 8h, 1st restriction 6h, 2nd restriction 6h, and recovery 10h) in a sleep laboratory, with polysomnography recordings taken during sleep. Throughout waking periods in each condition, participants completed the psychomotor vigilance test (PVT), which measures vigilant attention, and the Go/No-Go task, which measures inhibition control.Results: The results showed that sleep restriction significantly decreased the proportion of N1 and N2 sleep, increased the proportion of N3 sleep, and reduced the time spent awake after sleep onset (WASO) and sleep onset latency. Poorer performance on the PVT and Go/No Go task was associated with longer WASO, a larger proportion of N3 sleep, and a smaller proportion of N2 sleep. Additionally, the power spectral density of delta waves significantly increased after sleep restriction, and this increase predicted a decrease in vigilance and inhibition control the next day.Conclusion: These findings suggest that sleep architecture and EEG signatures may partially explain cognitive impairment caused by sleep restriction.Keywords: sleep restriction, sleep architecture, sleep EEG, vigilance, inhibition control

Yijin Chen,1,2,&ast; Hongtao Liu,2,3,&ast; Zhikang Yu,2,4 Yang Yang,1,2 Qingyan Huang,2,4 Changqing Deng,1,2 Hui Rao,2,4 Heming Wu2,4 1Department of Gastroenterology, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China; 2Guangdong Provincial Key Laboratory of Precision Medicine and Clinical Translational Research of Hakka Population, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China; 3Department of Gastrointestinal Surgery, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China; 4Center for Precision Medicine, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Heming Wu, Center for Precision Medicine, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Academy of Medical Sciences, No. 63 Huangtang Road, Meijiang District, Meizhou, 514031, People’s Republic of China, Tel +86 753-2131-591, Email wuheming1986@126.comBackground: Alcoholics are prone to alcoholic cirrhosis (ALC). Aldehyde dehydrogenase 2 (ALDH2) is involved in alcohol metabolism. Herein, the relationship between ALDH2 genotypes and ALC was analyzed among Hakka alcoholics in southern China.Methods: A total of 213 alcoholics and 214 non-alcoholics were included in the study. The ALDH2 gene rs671 polymorphism was analyzed, life history, disease history, and auxiliary examination results of these participants were collected.Results: The alcoholics had higher level of total serum protein, and serum globulin, lower level of serum albumin, serum albumin/globulin ratio, serum prealbumin, neutrophil-to-lymphocyte ratio (NLR), lymphocyte-to-monocyte ratio (LMR), and platelet-to-lymphocyte ratio (PLR) than non-alcoholics. In the 213 alcoholics, 180 developed ALC. There were 206 non-ALC persons in the 214 non-alcoholics. The proportion of the ALDH2 rs671 G/G homozygous (&ast;1/&ast;1) was significantly lower in ALC patients (83.3%) than that of other groups (100.0% in non-ALC in alcoholics, 95.6% in non-ALC in non-alcoholics), while the proportion of the G/A heterozygous (&ast;1/&ast;2) was significantly higher in ALC patients (16.7%) than that of other groups (0% in non-ALC in alcoholics, 4.4% in non-ALC in non-alcoholics). Logistic regression analysis indicated that participants with low level of NLR (adjusted OR 5.543, 95% CI 2.964– 10.368, P< 0.001), LMR (adjusted OR 9.256, 95% CI 4.740– 18.076, P< 0.001), and PLR (adjusted OR 6.047, 95% CI 3.372– 10.845, P< 0.001), and ALDH2 G/A genotype (adjusted OR 6.323, 95% CI 2.477– 16.140, P< 0.001) had a significantly higher risk of ALC.Conclusion: ALDH2 polymorphism rs671 &ast;1/&ast;2 genotype is a potential risk factor for the development of ALC among Hakka alcoholics.Keywords: ALDH2, alcoholics, alcoholic liver cirrhosis, gene polymorphism, Hakka

Yu Luo,1,2,&ast; Heming Wu,2,3,&ast; Qingyan Huang,2,3 Hui Rao,2,3 Zhikang Yu,2,3 Zhixiong Zhong2,3 1Department of Gynaecology, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China; 2Guangdong Provincial Key Laboratory of Precision Medicine and Clinical Translational Research of Hakka Population, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China; 3Center for Precision Medicine, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Academy of Medical Sciences, Meizhou, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Zhikang Yu; Zhixiong Zhong, Center for Precision Medicine, Guangdong Provincial Key Laboratory of Precision Medicine and Clinical Translational Research of Hakka Population, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Academy of Medical Sciences, No. 63 Huangtang Road, Meijiang District, Meizhou, 514031, People’s Republic of China, Tel +753-2131-591, Email yuzhikang01@126.com; zhongzhixiong01@126.comObjective: To investigate the frequencies of BRCA1 and BRCA2 mutations in Chinese Hakka patients with ovarian cancer.Methods: The protein coding regions and exon intron boundary regions of the BRCA gene were sequenced using genomic DNA isolated from the lymphocytes of patients with next-generation sequencing. The patients’ family history and clinical records were collected.Results: A total of 195 patients with ovarian cancer were included in the study, and 52 distinct variants of the BRCA gene were identified. It was found that 64 patients (64/195, 32.8%) had BRCA gene mutations, including 32 patients (50.0%) with BRCA1 mutation, 27 patients (42.2%) with BRCA2 mutation, and 5 patients (7.8%) with both mutations. Furthermore, 22 pathogenic mutations were detected in 26 patients, 2 likely pathogenic variants in 2 patients, 12 variants of uncertain significance in 20 patients, and 16 likely benign variants in 24 patients. The mutations were mainly found to occur in exons 8, 14, and 17 of BRCA1 and exons 10, 11, 14, and 15 of BRCA2. The results showed that the BRCA genes possess different mutation hotspots in different ethnic groups. In addition, recurrent mutations were noted in many patients. BRCA1 c.536 A>T, considered a founder mutation, was identified in 10 patients (15.63%, 10/64), followed by BRCA1 c.2635 G>T (6.25%, 4/64) and BRCA2 c.2566 T>C (6.25%, 4/64).Conclusion: The BRCA1 c.536 A>T could be considered to be a founder mutation in this ovarian cancer population. This recurrent BRCA1 mutation has rarely been observed in other ethnic groups. Our findings are expected to provide valuable data for clinical consultation and for designing individualized treatment for ovarian cancer.Keywords: BRCA gene, ovarian cancer, variants, Hakka population

Chunting Liang &ast;, Tongtong Huang &ast;, Xiaofeng Zhang, Huaqing Rao, Zhiru Jin, Xiaoxiong Pan, Jingtao Li, Yingying Mo, Yongzhi Cai, Ji Wu Department of Ultrasonic Medicine, The First Hospital Affiliated to Guangxi Medical University, Nanning, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Ji Wu, Department of Ultrasonic Medicine, The First Hospital Affiliated to Guangxi Medical University, Nanning, 530021, People’s Republic of China, Email gxnnwuji@126.comPurpose: To study the thrombolytic effect and safety of cRGD urokinase liposomes (cRGD-UK-LIP) in rats with acute pulmonary microthromboembolism (APMTE), and explore the application value of echocardiography (ECHO) in animal models.Patients and Methods: Ninety-six SD rats were randomized into 6 groups (16/group): normal control, sham operation, APMTE, normal saline (NS), free urokinase (UK), cRGD-UK-LIP. Four groups (APMTE, NS, UK, cRGD-UK-LIP) of rats were injected with autologous thrombus to induce APMTE. Samples were injected into 3 groups (NS, UK, cRGD-UK-LIP) of rats after modeling. Echocardiography was used to assess right ventricle (RV) function and morphology in rats. Six rats in each group were randomly selected and pulmonary artery pressure (PAP) of them was measured through ECHO-guided transthoracic puncture. Finally, the rats were killed and their tissues were taken for pathological examination.Results: Compared with normal control or sham operation group, rats in APMTE group had enlarged RV, decreased RV function, increased PAP, and lung tissue of them showed postthromboembolic appearance. There was no significant difference between NS group and APMTE group. RV morphology and function of rats in the UK group and cRGD-UK-LIP group were better and vessels with residual thrombus in these 2 groups were less than APMTE group, especially in the cRGD-UK-LIP group. In terms of PAP, only cRGD-UK-LIP group was significantly lower than APMTE group. No hyperemia, bleeding and swelling were observed in heart, liver and kidney of rats in each group.Conclusion: A rat model of APMTE was successfully established. cRGD-UK-LIP has better thrombolytic effect than free urokinase and it is safe. Echocardiography is not merely an important way to evaluate the morphology and function of RV, transthoracic puncture measurement under the guidance of it can be an effective way to monitor PAP in animal models.Keywords: cRGD urokinase liposomes, acute pulmonary microthromboembolism, targeted thrombolysis, right ventricle, echocardiography, pulmonary artery pressure

Abstract Multiple myeloma (MM) is a heterogeneous disease characterized by significant genomic instability. Recently, a causal role for the AID/APOBEC deaminases in inducing somatic mutations in myeloma has been reported. We have identified APOBEC/AID as a prominent mutational signature at diagnosis with further increase at relapse in MM. In this study, we identified upregulation of several members of APOBEC3 family (A3A, A3B, A3C, and A3G) with A3G, as one of the most expressed APOBECs. We investigated the role of APOBEC3G in MM and observed that A3G expression and APOBEC deaminase activity is elevated in myeloma cell lines and patient samples. Loss-of and gain-of function studies demonstrated that APOBEC3G significantly contributes to increase in DNA damage (abasic sites and DNA breaks) in MM cells. Evaluation of the impact on genome stability, using SNP arrays and whole genome sequencing, indicated that elevated APOBEC3G contributes to ongoing acquisition of both the copy number and mutational changes in MM cells over time. Elevated APOBEC3G also contributed to increased homologous recombination activity, a mechanism that can utilize increased DNA breaks to mediate genomic rearrangements in cancer cells. These data identify APOBEC3G as a novel gene impacting genomic evolution and underlying mechanisms in MM.

Tianxin Mao,1,2 David Dinges,3 Yao Deng,1,2 Ke Zhao,2 Zijing Yang,2,4 Hui Lei,2 Zhuo Fang,2 Fan Nils Yang,2 Olga Galli,3 Namni Goel,5 Mathias Basner,3 Hengyi Rao1– 3 1Key Laboratory of Applied Brain and Cognitive Sciences, School of Business and Management, Shanghai International Studies University, Shanghai, People’s Republic of China; 2Center for Functional Neuroimaging, Department of Neurology, University of Pennsylvania, Philadelphia, PA, USA; 3Division of Sleep and Chronobiology, Department of Psychiatry, University of Pennsylvania, Philadelphia, PA, USA; 4School of Medicine, Shanghai Jiaotong University, Shanghai, People’s Republic of China; 5Biological Rhythms Research Laboratory, Department of Psychiatry and Behavioral Sciences, Rush University Medical Center, Chicago, IL, USACorrespondence: Hengyi RaoCenter for Functional Neuroimaging & Department of Neurology, University of Pennsylvania Perelman School of Medicine, Room D502, Richards Medical Research Building, 3700 Hamilton Walk, Philadelphia, PA, 19104-4202, USATel +1 215-746-2470Email hengyi@pennmedicine.upenn.eduPurpose: Sleep loss impairs a range of neurobehavioral functions, particularly vigilant attention and arousal. However, the detrimental effects of sleep deprivation on inhibition control and its relationship to vigilant attention impairments remain unclear. This study examined the extent to which vigilant attention deficits contribute to inhibition control performance after one night of total sleep deprivation (TSD) and two nights of partial sleep restriction (PSR).Participants and Methods: We analyzed data from N = 49 participants in a one-night of TSD experiment, N=16 participants in a control experiment without sleep loss, and N = 16 participants in a two-nights of PSR experiment (time in bed, TIB = 6 h for each night). Throughout waking periods in each condition, participants completed the psychomotor vigilance test (PVT), which measures vigilant attention, and the Go/No-Go task, which measures inhibition control.Results: After TSD and PSR, participants displayed significantly slower reaction times (RT) and more lapses in PVT performance, as well as slower Go RT and more errors of omission during the Go/No-Go task. PVT deficits accounted for 18.0% of the change in Go RT and 12.4% of the change in errors of omission in the TSD study, and 23.7% of the change in Go RT and 20.3% of the change in errors of omission in the PSR study.Conclusion: Both TSD and PSR impaired inhibition control during the Go/No-Go task, which can be partly accounted for by vigilant attention deficits during the PVT. These findings support the key role of vigilant attention in maintaining overall neurobehavioral function after sleep loss.Keywords: total sleep deprivation, partial sleep restriction, vigilant attention, inhibition control

Hua-chun Rao,1,* Zhao-ke Wu,1,* Si-da Wei,1 Yun Jiang,1 Qing-xin Guo,1 Jia-wen Wang,1 Chang-xian Chen,1 Hui-yong Yang2 1Quanzhou Orthopedic-Traumatological Hospital, Fengze District, Quanzhou, Fujian, People’s Republic of China; 2School of Medicine, Institute of Molecular Medicine, Huaqiao University, Quanzhou, People’s Republic of China*These authors contributed equally to this work.Correspondence: Chang-xian Chen Tel +86 13636913112Email ccx6070@qq.comHui-yong Yang Tel +86 15359531722Email shyhy@hqu.edu.cnPurpose: Moesin-ezrin-radixin-like protein (Merlin) has been identified as a tumor suppressor in several types of cancers. However, the biological function of Merlin in osteosarcoma remains unclear. MicroRNAs (miRNAs) can influence cancer progression by targeting oncogenes or anti-oncogenes. In this study, we sought to evaluate the regulation of Merlin expression by miR-25-3p and the role of the miR-25-3p/Merlin axis in osteosarcoma progression, with the aim of identifying a potential therapeutic target for osteosarcoma.Materials and Methods: TCGA (The Cancer Genome Atlas) database was used to analyze the correlation between Merlin expression and prognosis. RT-qPCR and Western blotting analyses were performed to compare Merlin expression between normal and malignant cells. A dual-luciferase reporter assay was performed to evaluate the direct targeting of Merlin by miR-25-3p. We overexpressed miR-25-3p, or/and Merlin, in U-2 OS and 143B cells, and studied their cellular functions in vitro. MTT and colony formation assays were performed to determine the effects on cell growth. EdU and cell cycle assays were performed to analyze the effects in cell replication. We used annexin V-fluorescein isothiocyanate and propidium iodide to stain apoptotic cells, and analyzed the cells using flow cytometry. The effects on cell metastasis were studied in wound healing and transwell assays. Lastly, the underlying mechanism was determined in RT-qPCR and Western blotting experiments.Results: Low Merlin expression was linked to poor prognosis. miR-25-3p was observed to directly target Merlin and downregulate its expression. miR-25-3p promoted cell growth, migration, and invasion, and inhibited apoptosis induced by cisplatin. Moreover, the overexpression of Merlin reversed the abovementioned effects of miR-25-3p. Further, the miR-25-3p/Merlin axis was observed to play an important role in the Hippo pathway, and regulated the expression of genes such as BIRC5, CTGF, and CYR61.Conclusion: miR-25-3p functions as an oncogenic microRNA in osteosarcoma by targeting Merlin, and may serve as a potential therapeutic target for osteosarcoma.Keywords: Merlin, miR-25-3p, osteosarcoma, proliferation, apoptosis, metastasis

Chang Jiang,1,* Yu-Hua Huang,2,* Jia-Bin Lu,2 Yuan-Zhong Yang,2 Hui-Lan Rao,2 Bei Zhang,1 Wen-Zhuo He,1 Liang-Ping Xia1 1VIP Region, Sun Yat-Sen University Cancer Center, State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Guangzhou, Guangdong 510060, People’s Republic of China; 2Department of Pathology, Sun Yat-Sen University Cancer Center, State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Guangzhou, Guangdong 510060, People’s Republic of China *These authors contributed equally to this work Purpose: Tumor vessels supported by perivascular cells have been implicated in the failure of some anti-angiogenic agents. The relationship between perivascular cell coverage (PC) and bevacizumab efficacy in metastatic colorectal cancer (mCRC) was analyzed.Patients and methods: A total of 284 consecutive mCRC patients who received first-line chemotherapy with or without bevacizumab from 2007–2014 in Sun Yat-Sen University Cancer Center were analyzed. Immunohistochemical double-stain for the perivascular cell marker alpha-smooth muscle actin and endothelial cell (cluster of differentiation 31) was performed to characterize the intratumoral microvascular density. Multispectral image capturing and computerized image analyses were used to quantify the microvessels supported by the perivascular cells. The patients were divided into high and low PC group according to a median cutoff value of 0.55.Results: No significant differences in overall survival (OS) and progression-free survival (PFS) were noted between the high and low PC group. In the low PC group, the patients with bevacizumab treatment had favorable OS (P=0.03), but without PFS benefit. In the high PC group, neither OS nor PFS was significantly different between the B+C and C subgroup. Tumors with perineural invasion had high PC (P=0.03).Conclusion: The data showed that a low PC value could be a predictor for bevacizumab benefit. Keywords: perivascular cell coverage, bevacizumab, predictive marker, overall survival, metastatic colorectal cancer

Interactions between the inflammatory chemokine CCL20 and its receptor CCR6 have been associated with colorectal cancer growth and metastasis, however, a causal role for CCL20 signaling through CCR6 in promoting intestinal carcinogenesis has not been demonstrated in vivo. In this study, we aimed to determine the role of CCL20-CCR6 interactions in spontaneous intestinal tumorigenesis. CCR6-deficient mice were crossed with mice heterozygous for a mutation in the adenomatous polyposis coli (APC) gene (APCMIN/+ mice) to generate APCMIN/+ mice with CCR6 knocked out (CCR6KO-APCMIN/+ mice). CCR6KO-APCMIN/+ mice had diminished spontaneous intestinal tumorigenesis. CCR6KO-APCMIN/+ also had normal sized spleens as compared to the enlarged spleens found in APCMIN/+ mice. Decreased macrophage infiltration into intestinal adenomas and non-tumor epithelium was observed in CCR6KO-APCMIN/+ as compared to APCMIN/+ mice. CCL20 signaling through CCR6 caused increased production of CCL20 by colorectal cancer cell lines. Furthermore, CCL20 had a direct mitogenic effect on colorectal cancer cells. Thus, interactions between CCL20 and CCR6 promote intestinal carcinogenesis. Our results suggest that the intestinal tumorigenesis driven by CCL20-CCR6 interactions may be driven by macrophage recruitment into the intestine as well as proliferation of neoplastic epithelial cells. This interaction could be targeted for the treatment or prevention of malignancy.