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Yuan-Jie Liu,1,* Jing-Xiao Li,1,* Jie-Pin Li,1 Yi-Dou Hu,2 Zhi-Bin Ma,3 Wei Huang,1 Shen-Lin Liu,1 Xi Zou1 1Affiliated Hospital of Nanjing University of Chinese Medicine, Jiangsu Province Hospital of Chinese Medicine, Nanjing, Jiangsu, 210029, People’s Republic of China; 2Zhangjiagang TCM Hospital Affiliated to Nanjing University of Chinese Medicine, Zhangjiagang, Jiangsu, 215600, People’s Republic of China; 3Nanjing YOUMENG Biology Science and Technology Co. Ltd, Nanjing, Jiangsu, 210029, People’s Republic of China*These authors contributed equally to this workCorrespondence: Xi Zou; Shen-lin Liu, Email fsyy00670@njucm.edu.cn; lsljsszyy@126.comBackground: Hepatocellular carcinoma (HCC) remains a leading cause of cancer-related mortality, underscoring the need for novel therapeutic targets. This study aimed to elucidate the role of endoplasmic reticulum membrane protein complex subunit 1 (EMC1) in HCC progression and its therapeutic potential.Methods: Publicly available sequencing data and biopsy specimens were analyzed to assess EMC’s clinical value and functions in HCC. In vitro experiments validated EMC functions, and multiplex immunofluorescence analysis examined EMC-associated sorafenib resistance mechanisms. EMC1 expression was knocked down in HCC cell lines, followed by cell viability, wound healing, and transwell migration assays. Tumor growth and response to sorafenib treatment were evaluated in mouse models. Metabolomic analysis assessed changes in the TCA cycle.Results: EMC genes were aberrantly expressed in HCC, and high EMC1 expression correlated with poorer survival rates. EMC1 disruption enhanced HCC cells’ sensitivity to sorafenib, reducing cell viability, increasing apoptosis, and decreasing tumor size and weight. EMC1 maintained cancer cell stemness and promoted M2 macrophage infiltration. Metabolomic analysis revealed significant changes in the TCA cycle, indicating EMC1’s role in HCC metabolic reprogramming. Importantly, EMC1 is highly associated with sorafenib resistance, potentially linked to CTNNB1 mutation or activation.Conclusion: EMC1 plays a critical role in regulating the sorafenib resistance in HCC. Targeting EMC1 may improve HCC treatment efficacy.Keywords: endoplasmic reticulum membrane complex subunit 1, catenin beta 1, tumor stemness, M2 macrophage, sorafenib resistance

Pin Wu,1,* Lu Zhu,1,* Shi-Yu Zheng,1 Jun-Xiong Li,1 Meng-Die Wu,1 Wen-Jia Wang,1 Yu-Chao Hou,1 Jing Li,1 Huan-Gan Wu1,2 1Department of Acupuncture, Yueyang Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai, People’s Republic of China; 2Department of Key Laboratory of Acupuncture and Immunological Effects, Shanghai University of Traditional Chinese Medicine, Shanghai, People’s Republic of China*These authors contributed equally to this workCorrespondence: Jing Li, Department of Acupuncture, Yueyang Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai University of Traditional Chinese Medicine, No. 110 Ganhe Road, Hongkou District, Shanghai, 200437, People’s Republic of China, Tel +021-65161782, Email 1971921250@qq.com Huan-Gan Wu, Department of Key Laboratory of Acupuncture and Immunological Effects, Shanghai University of Traditional Chinese Medicine, No. 650 South Wanping Road, Xuhui District, Shanghai, 200437, People’s Republic of China, Tel +021-64383453, Email wuhuangan@126.comBackground: Cancer-related pain is one of the primary symptoms of patients with hepatocellular carcinoma (HCC). Previous studies have shown that transcutaneous electrical acupoint stimulation (TEAS) is effective in treating patients with acute or chronic pain. In China, it is indispensable to evaluate the efficacy of TEAS in combination with opioids for the treatment of moderate to severe HCC-related pain.Methods/Design: This is a single-center clinical, prospective randomized controlled clinical trial protocol. 104 patients will be randomly divided into the observation group and the control group in a ratio of 1:1.In addition to routine cancer pain medication, the two groups of patients will receive TEAS treatment twice a day for one week. Acupoints will include bilateral Hegu(LI4), Neiguan(PC6), Zusanli(ST36), Taichong(LR3), Ganshu(BL18), Geshu(BL17), Qimen(LR14), and Zhangmen(LR13). The treatment time is from 9:00 a.m. to 11:00 p.m. and from 4:00 p.m. to 6:00 p.m. The primary outcome measures are the Numerical Rating Scale (NRS) and the secondary outcome measures include the Brief Pain Inventory(BPI), dosage and administration duration of opioid drugs, frequency of nausea, vomiting and defecation, Karnofsky Performance Status Scale (KPS), Quality of life scale (QOL), Brief Fatigue Inventory (BFI). The outcome measures will be evaluated at baseline, during treatment and 1 week after treatment.Discussion: Results of this trial are expected to clarify the value of TEAS stimulation performed on specific points in the management of moderate to severe pain in HCC.Trial registration: Chinese clinical trial registry, ChiCTR2100044615 (http://www.chictr.org.cn), Registered on 24 March 2021.Keywords: protocol, hepatocellular carcinoma, TEAS, pain

Jing-Xiao Li,1 Jin-Shu Pang,2 Bin-Tong Yin,1 Gang Chen,1 Jun-Hong Chen,1 Jia-Yuan Luo,1 Xia Yang,1 Li-Ting Qin,1 Jiang-Hui Zeng,3 Peng Chen,4 Jia-Bo Chen,4 Deng Tang1 1Department of Pathology, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Zhuang Autonomous Region, People’s Republic of China; 2Department of Medical Ultrasonics, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Zhuang Autonomous Region, People’s Republic of China; 3Department of Clinical Laboratory, The Third Affiliated Hospital of Guangxi Medical University/Nanning Second People’s Hospital, Nanning, Guangxi Zhuang Autonomous Region, 530031, People’s Republic of China; 4Department of Pediatric Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Zhuang Autonomous Region, 530031, People’s Republic of ChinaCorrespondence: Deng TangDepartment of Pathology, The First Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Zhuang Autonomous Region, People’s Republic of ChinaTel +86-7715356534Email tdnever@163.comPurpose: The molecular mechanisms and signal pathways of ferroptosis in hepatoblastoma (HB) have not yet been clarified. In previous studies, activating transcription factor 3 (ATF3) was reported to be correlated with several tumors, but the clinical significance of ATF3 has never been determined. Herein, we investigated the clinicopathological value and mechanisms of ATF3 in regulating ferroptosis in HB.Methods: The mRNA microarray and RNA-sequencing data of 402 samples from our hospital and public databases were used to estimate ATF3 expression and assess its clinical role in HB. The standard mean difference (SMD) and summary receiver operating characteristic curves were utilized to judge the discrimination ability of ATF3 between HB and non-HB liver tissues. We examined the expression variation of ATF3 in HB cells after the treatment with erastin. We also predicted the target genes of ATF3 as a transcriptional factor from public Chromatin Immunoprecipitation-sequencing data and selected the ferroptosis-related genes for a signaling pathway analysis.Results: In ten series, the pooled SMD for ATF3 was − 0.91, demonstrating that ATF3 expression was predominantly lower in HB than in non-HB liver tissues. ATF3 down-regulation showed moderate potential to distinguish HB from non-HB liver tissues (area under curves = 0.83, 95% confidence interval = 0.79– 0.86). Altogether, 4855 putative targets of ATF3 as a transcriptional factor were collected, among which, 60 genes were ferroptosis-related.Conclusion: The down-regulated ATF3 expression may play a vital role in the occurrence of HB possible partially by regulating ferroptosis.Keywords: transcription factor, ATF3, hepatoblastoma, ferroptosis

Hui-Zhou Li,1 Jie Tan,1 Tian Tang,2 Tian-Zhi An,3 Jun-Xiang Li,4 Yu-Dong Xiao1 1Department of Radiology, The Second Xiangya Hospital of Central South University, Changsha, 410011, People’s Republic of China; 2Department of Interventional Radiology, The Affiliated Cancer Hospital of Xiangya School of Medicine, Central South University, Changsha, 410013, People’s Republic of China; 3Department of Interventional Radiology, The Affiliated Hospital of Guizhou Medical University, Guiyang, 550002, People’s Republic of China; 4Department of Interventional Radiology, Guizhou Medical University Affiliated Cancer Hospital, Guiyang, 550004, People’s Republic of ChinaCorrespondence: Yu-Dong XiaoDepartment of Radiology, The Second Xiangya Hospital of Central South University, No. 139 Middle Renmin Road, Changsha, 410011, People’s Republic of ChinaTel +86 13637403027Fax +86 731-85292116Email xiaoyudong222@csu.edu.cnPurpose: Transarterial chemoembolization (TACE) is recommended in patients with unresectable HCC beyond the Milan criteria (MC). However, the long-term efficacy of TACE remains unsatisfactory. Percutaneous microwave ablation (MWA) is a curative therapy for early-stage HCC that provides better local tumor control than TACE; however, MWA is limited for large or multifocal lesions. We aimed to compare treatment efficacy and downstaging rate following combined TACE-MWA and TACE alone in patients with unresectable HCC beyond the MC.Patients and Methods: Patients with unresectable HCC beyond the MC who underwent either TACE-MWA (n=91) or TACE alone (n=140) at four medical institutions were included. Potential influencing factors on overall survival (OS) and progression-free survival (PFS) were included in the Cox regression analysis. Propensity-score matching of patients treated with TACE-MWA and TACE alone was performed. Differences in OS and PFS were compared with the Log rank test. Patients who met the University of California, San Francisco criteria were eligible for assessment of the probability of downstaging within the MC. Downstaging rate was compared between the two groups.Results: In multivariate analysis, treatment with TACE alone was an independent predictor of poor PFS (P=0.011) and OS (P< 0.001). Both PFS (P=0.043) and OS (P=0.002) were significantly higher in patients treated with TACE-MWA than those treated with TACE alone. The downstaging rate was higher in patients treated with TACE-MWA than those treated with TACE alone (P=0.039).Conclusion: Compared with TACE alone, TACE-MWA may offer a survival benefit in terms of OS and PFS in HCC patients beyond the MC. Additionally, TACE-MWA may provide higher probability of downstaging within the MC than TACE alone, thereby increasing the possibility of liver transplantation.Keywords: microwave ablation, transarterial chemoembolization, survival, propensity score-matching, Milan criteria

Tian-Cheng Wang,1,2 Tian-Zhi An,3 Jun-Xiang Li,4,* Peng-Fei Pang1,* 1Department of Interventional Medicine, The Fifth Affiliated Hospital, Sun Yat-sen University, Zhuhai, Guangdong, People’s Republic of China; 2Department of Radiology, The Second Xiangya Hospital of Central South University, Changsha, Hunan, People’s Republic of China; 3Department of Interventional Radiology, The Affiliated Hospital of Guizhou Medical University, Guiyang, Guizhou, People’s Republic of China; 4Department of Interventional Radiology, The Affiliated Cancer Hospital of Guizhou Medical University, Guiyang, Guizhou, People’s Republic of China*These authors contributed equally to this workCorrespondence: Peng-Fei Pang; Jun-Xiang Li Email pangpf@mail.sysu.edu.cn; 357936984@qq.comBackground: Mounting evidence has shown that systemic inflammation response index (SIRI), a novel prognostic biomarker based on peripheral lymphocyte, neutrophil and monocyte counts, is associated with poor prognosis for several tumors. However, the prognostic value of SIRI in patients with hepatocellular carcinoma (HCC) undergoing transarterial chemoembolization (TACE) is elusive. Herein, we aimed to evaluate the correlation between SIRI and clinical outcomes in these patients.Methods: A total of 194 consecutive patients who underwent TACE were included in this study. Patients were stratified into high and low SIRI groups based on the cut-off value using receiver operating characteristic (ROC) analysis. Independent risk factors for tumor response were analyzed using forward stepwise logistic regression. A one-to-one propensity score matching (PSM) was conducted to compare progression-free survival (PFS) and overall survival (OS) between low and high SIRI patients. The discriminatory power of the combination of number of tumors and SIRI in predicting initial TACE response was evaluated by ROC analysis.Results: Patients were divided into high SIRI (> 0.88) and low SIRI (≤ 0.88) groups. High SIRI (p = 0.003) and more than three tumors (p = 0.002) were significantly related to poorer tumor response. Moreover, the low SIRI group had longer PFS and OS than the high SIRI group (both P < 0.05) before and after PSM. Combination of SIRI and number of tumors can improve the predictive ability to predict initial TACE response with an area under the curve (AUC) of 0.678.Conclusion: Pretreatment peripheral blood SIRI was found to be an independent predictor of tumor response and clinical outcomes in patients with HCC undergoing TACE. Patients with high SIRI may have a poor prognosis.Keywords: systemic inflammation response index, hepatocellular carcinoma, transarterial chemoembolization, biomarker, survival

Since the first plasma realized in 2020, a series of key systems on HL-3 (known as HL-2M before) tokamak have been equipped/upgraded, including in-vessel components (the first wall, lower divertor, and toroidal cryogenic/water-cooling/baking/glow discharge systems, etc.), auxiliary heating system of 11 MW, and 28 diagnostic systems (to measure the plasma density electron temperature, radiation, magnetic field, etc.). Magnet field systems were commissioned firstly for divertor plasma discharges. During the 2nd experimental campaign of HL-3 tokamak, several great progresses have been achieved. Firstly, the successful operation with plasma current larger than 1 MA was achieved under a divertor configuration. Secondly, the advanced divertor concept with two distinct snowflake configurations was realized. It is found that the distribution of ion saturation current and heat flux on bottom plate becomes wide due to magnetic surface expansion, demonstrating the advantage of such configuration in the heat flux mitigation. In addition, using the combination of NBI, ECRH and LHCD, the standard sawtoothing high confinement mode of megampere plasma was firstly accessed on the HL-3. The successful commissioning of HL-3 is beneficial for the initial operation of ITER.

Nan Xiao,1 Wei Gai,2 Wei-Guo Hu,1 Jian-Xing Li,1 Yan Zhang,2 Xiu-Ying Zhao11Department of Clinical Laboratory, Beijing Tsinghua Changgung Hospital, School of Clinical Medicine, Tsinghua University, Beijing, People’s Republic of China; 2National Engineering Research Center for Beijing Biochip Technology, Beijing, People’s Republic of ChinaBackground: Mycoplasma is an opportunistic pathogen causing both urogenital and extragenital infections. The lack of cell wall renders Mycoplasma difficult to culture and identify with ordinary methods. Next-generation sequencing (NGS) is a new technology helping a lot in the diagnosis of infective diseases. In this case, NGS played a key role in the diagnosis of Mycoplasma infection.Case presentation: A mid-aged man suffering from renal cyst underwent cyst incision followed by invasive treatments to eliminate hematoma caused by renal artery hemorrhage. After the cyst incision operation, the patient had a persistent high temperature. The persistent increase of blood neutrophile granulocyte count and C-reaction protein suggested an unresolved infection. The empirically chosen anti-infective agents were meropenem and linezolid since the ordinary bacterial cultures of surgical site drainage and blood yielded a negative result. At postoperation day (POD) 17, NGS result of his drainage clearly indicated the pathogen was Mycoplasma hominis. At POD 24, the drug sensitivity test showed resistance to quinolones, clarithromycin and erythromycin, but intermediate to azithromycin. Since then, the antimicrobial agents were changed into azithromycin and kept unchanged until the patient was fully recovered and discharged at POD 39.Conclusion: When the ordinary laboratory diagnostic methods failed, NGS diagnosis could reduce the hospitalization expenses and shorten the lengths of hospital stay.Keywords: NGS, Mycoplasma, renal cyst, infection

Li-Hua Yang,1,2,* Ye Wang,1,3,* Shi Qiao,1,* Min-Jun Wang,1 Fei Chen,1 Xiao-Yuan Zi,1 Jian-Xiu Li,1 Hai-Bin Zhang,4 Bing Yu,1,4 Yi-Ping Hu1 1Department of Cell Biology, Center for Stem Cell and Medicine, Second Military Medical University, Shanghai, People’s Republic of China; 2Department of Cell Biology and Neurobiology, Xuzhou Medical University, Xuzhou, People’s Republic of China; 3Department of Urology, Changhai Hospital, Second Military Medical University, Shanghai, People’s Republic of China; 4Department of Hepatic Surgery V, Eastern Hepatobiliary Surgery Hospital, Shanghai, People’s Republic of China *These authors contributed equally to this work Purpose: Liver cancer stem cells (CSCs) are known to be associated with the development, survival, proliferation, metastasis, and recurrence of liver tumors. The aim of this study was to investigate the association of liver-enriched activator protein 1 (LAP1) with hepatocellular carcinoma (HCC) and liver CSCs (LCSCs) and explore the impact of LAP1 on LCSCs. Materials and methods: Differences in LAP1 expression in liver cancer tissues versus matched para-tumoral liver tissues and LCSCs versus non-CSCs were analyzed by Western blotting, real-time polymerase chain reaction, immunohistochemistry, and flow cytometry. The effect of LAP1 on liver cancer cells was evaluated by the expression of CSC markers, oncosphere formation, proliferation, migration, and invasion in vitro. Cell cycle distribution and the number of apoptotic cells were analyzed to assess cell cycle and cell apoptosis. Furthermore, a mouse subcutaneous tumor implant model was established to explore the role of LAP1 in the development of HCC in vivo. Finally, the expression of CSC markers in paraffin-embedded sections was evaluated by immunofluorescence. Results: LAP1 was weakly expressed in HCC tumors and cell lines and even weaker in LCSCs. LAP1 inhibited the expression of stem cell–associated genes and reduced the abilities of oncosphere formation, proliferation, migration, and invasion in vitro. Cell cycle assay revealed that LAP1 induced G1/G0 arrest. Furthermore, LAP1 decreased subcutaneous tumor-formation ability and the expression of CSC markers and Ki67 in vivo. Conclusion: LAP1 suppressed the stem cell features of HCC, indicating that it possessed an antitumor effect in liver cancer, both in vitro and in vivo; therefore, LAP1 may prove to be a potential target in liver CSC-targeted therapy. Keywords: LAP1, liver-enriched activator protein 1, hepatocellular carcinoma, HCC, liver cancer stem cell

Jin-Xiu Li,1,2,* Wei-Ping Fu,3,* Jing Zhang,4 Xiao-Hua Zhang,1,2 Chang Sun,1,5 Lu-Ming Dai,3 Li Zhong,1,5,6 Li Yu,1,2 Ya-Ping Zhang1,7 1State Key Laboratory for Conservation and Utilization of Bio-Resource in Yunnan, 2Key Laboratory for Animal Genetic Diversity and Evolution of High Education in Yunnan Province, School of Life Sciences, Yunnan University, 3Department of Respiratory Critical Care Medicine, 4Department of Thoracic Surgery, The First Affiliated Hospital of Kunming Medical University, Kunming, 5College of Life Sciences, 6Provincial Demonstration Center for Experimental Biology Education, Shaanxi Normal University, Xi’an, 7State Key Laboratory of Genetic Resources and Evolution, and Yunnan Laboratory of Molecular Biology of Domestic Animals, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, China *These authors contributed equally to this work Background: Previous studies have suggested that β2-adrenergic receptor (ADRB2) is associated with COPD. However, the role of genetic polymorphisms in ADRB2 on COPD has not been evaluated yet. Methods: In this study, SNaPshot genotyping, luciferase assay, chromatin immunoprecipitation and real-time polymerase chain reaction were adopted to investigate the association between ADRB2 genetic polymorphisms and COPD, comprehensively. Results: One single nucleotide polymorphism (rs12654778), located upstream of ADRB2, showed a significant association with COPD by the logistic regression analysis after adjusting for age, sex and smoking history (p=0.04) in 200 COPD patients and 222 controls from southwest Chinese population. Furthermore, the luciferase assay indicated that rs12654778-A allele reduced the relative promoter activity by ~26% compared with rs12654778-G allele (p=0.0034). The chromatin immunoprecipitation analysis demonstrated that rs12654778 modulated the binding affinity of transcription factor neurofibromin 1. In addition, a significantly reduced expression of ADRB2 in COPD patients was observed, compared with normal controls (p=0.017). Conclusion: Our findings suggest a previously unknown mechanism linking allele-specific effects of rs12654778 on ADRB2 expression to COPD onset, for the first time. Keywords: β2-adrenergic receptor, ADRB2, FEV1, lung, polymorphism

Cheng Wang,1 Jun-Xiong Li,1 Dang Tang,2 Jian-Qing Zhang,1 Li-Zhou Fang,1 Wei-Ping Fu,1 Ling Liu,1 Lu-Ming Dai1 1Second Department of Respiratory Medicine, 2First Department of Neurosurgery, The First Affiliated Hospital of Kunming Medical University, Kunming, People’s Republic of China Background: Metabolomics is the global unbiased analysis of all the small-molecule metabolites within a biological system. Metabolic profiling of different high-resolution computed tomography (HRCT) phenotypes of COPD patients before and after treatment may identify discriminatory metabolites that can serve as biomarkers and therapeutic agents. Patients and methods: 1H nuclear magnetic resonance spectroscopy (1H-NMR)-based metabolomics was performed on a discovery set of plasma samples from 50 patients with stable COPD. Patients were assigned into two groups on the basis of HRCT findings including phenotype E (n=22) and phenotype M (n=28). After budesonide–formoterol treatment (160/4.5 µg ×2 inhalations twice daily for 3 months), clinical characteristics and metabolites were then compared between phenotype E pretreatment and posttreatment, phenotype M pretreatment and posttreatment, phenotype E pretreatment and phenotype M pretreatment, and phenotype E posttreatment and phenotype M posttreatment. Results: Inhaled budesonide–formoterol therapy for both phenotype E (emphysema without bronchial wall thickening) and phenotype M (emphysema with bronchial wall thickening) was effective. However, phenotype E and phenotype M were different in response to therapy. Patients with phenotype M in response to therapeutic effects were significantly greater compared with phenotype E. Certain metabolites were identified, which were closely related to the treatment and phenotype. Metabolic changes in phenotype E or phenotype M after treatment may be involved with adenosine diphosphate (ADP), guanosine, choline, malonate, tyrosine, glycine, proline, l-alanine, l-valine, l-threonine leucine, uridine, pyruvic acid, acetone and metabolism disturbance. Metabolic differences between phenotype E and phenotype M in pretreatment and posttreatment covered glycine, D-glucose, pyruvic acid, succinate, lactate, proline, l-valine and leucine. Conclusion: Bronchial wall thickening in COPD may be an indicator for predicting the better response to the treatment with bronchodilator and corticosteroid. The identification of metabolic alterations provides new insights into different HRCT phenotypes and therapeutic assessment of COPD. Keywords: COPD, metabolomics, budesonide–formoterol, HRCT

Ye Wang,1,2,* Xiao-Yuan Zi,1,* Juan Su,1 Hong-Xia Zhang,1 Xin-Rong Zhang,3 Hai-Ying Zhu,1 Jian-Xiu Li,1 Meng Yin,3 Feng Yang,3 Yi-Ping Hu,11Department of Cell Biology, 2School of Clinical Medicine, 3Department of Pharmaceuticals, Second Military Medical University, Shanghai, People's Republic of China*Authors contributed equally.Abstract: In the rapid development of nanoscience and nanotechnology, many researchers have discovered that metal oxide nanoparticles have very useful pharmacological effects. Cuprous oxide nanoparticles (CONPs) can selectively induce apoptosis and suppress the proliferation of tumor cells, showing great potential as a clinical cancer therapy. Treatment with CONPs caused a G1/G0 cell cycle arrest in tumor cells. Furthermore, CONPs enclosed in vesicles entered, or were taken up by mitochondria, which damaged their membranes, thereby inducing apoptosis. CONPs can also produce reactive oxygen species (ROS) and initiate lipid peroxidation of the liposomal membrane, thereby regulating many signaling pathways and influencing the vital movements of cells. Our results demonstrate that CONPs have selective cytotoxicity towards tumor cells, and indicate that CONPs might be a potential nanomedicine for cancer therapy.Keywords: nanomedicine, selective cytotoxicity, apoptosis, cell cycle arrest, mitochondrion-targeted nanomaterials

BACKGROUND AND PURPOSE Imidazoline I(2) receptors have been implicated in several CNS disorders. Although several I(2) receptor agonists have been described, no simple and sensitive in vivo bioassay is available for studying I(2) receptor ligands. This study examined I(2) receptor agonist-induced hypothermia as a functional in vivo assay of I(2) receptor agonism. EXPERIMENTAL APPROACH Different groups of rats were used to examine the effects of I(2) receptor agonists on the rectal temperature and locomotion. The pharmacological mechanisms were investigated by combining I(2) receptor ligands and different antagonists. KEY RESULTS All the selective I(2) receptor agonists examined (2-BFI, diphenyzoline, phenyzoline, CR4056, tracizoline, BU224 and S22687, 3.2-56 mg·kg(-1) , i.p.) dose-dependently and markedly decreased the rectal temperature (hypothermia) in rats, with varied duration of action. Pharmacological mechanism of the observed hypothermia was studied by combining the I(2) receptor agonists (2-BFI, BU224, tracizoline and diphenyzoline) with imidazoline I(2 ) receptor/ α(2) adrenoceptor antagonist idazoxan, selective I(1) receptor antagonist efaroxan, α(2) adrenoceptor antagonist/5-HT(1A) receptor agonist yohimbine. Idazoxan but not yohimbine or efaroxan attenuated the hypothermic effects of 2-BFI, BU224, tracizoline and diphenyzoline, supporting the I(2) receptor mechanism. In contrast, both idazoxan and yohimbine attenuated hypothermia induced by the α(2) adrenoceptor agonist clonidine. Among all the I(2) receptor agonists studied, only S22687 markedly increased the locomotor activity in rats. CONCLUSIONS AND IMPLICATIONS Imidazoline I(2) receptor agonists can produce hypothermic effects, which are primarily mediated by I(2) receptors. These data suggest that I(2) receptor agonist-induced hypothermia is a simple and sensitive in vivo assay for studying I(2) receptor ligands. [ABSTRACT FROM AUTHOR]