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IRE1α regulates the PTHrP-IHH feedback loop to orchestrate chondrocyte hypertrophy and cartilage mineralization

Genes & Diseases 2024年第1期11卷 464-478页

作者： Mengtian Fan Nana Geng Xingyue Li Danyang Yin Yuyou Yang Rong Jiang Cheng Chen Naibo Feng Li Liang Xiaoli Li Fengtao Luo Huabing Qi Qiaoyan Tan Yangli Xie fengjin guo Laboratory of Developmental Biology Department of Cell Biology and GeneticsSchool of Basic Medical ScienceChongqing Medical UniversityChongqing 400016China Laboratory of Stem Cells and Tissue Engineering School of Basic Medical ScienceChongqing Medical UniversityChongqing 400016China Department of Orthopedics The 1st Affiliated Hospital of Chongqing Medical UniversityChongqing 400016China Department of Wound Repair and Rehabilitation Medicine State Key Laboratory of TraumaBurns and Combined InjuryTrauma CenterResearch Institute of SurgeryDaping HospitalArmy Medical UniversityChongqing 400042China

Cartilage development is controlled by the highly synergistic proliferation and differentiation of growth plate chondrocytes,in which the Indian hedgehog(IHH)and parathyroid hormone-related protein-parathyroid hormone-1 receptor(PTHrP-PTH1R)feedback loop is crucial.The inositol-requiring enzyme 1a/X-box-binding protein-1 spliced(IRE1α/XBP1s)branch of the unfolded protein response(UPR)is essential for normal cartilage development.However,the precise role of ER stress effector IRE1α,encoded by endoplasmic reticulum to nucleus signaling 1(ERN1),in skeletal development remains unknown.Herein,we reported that loss of IRE1α accelerates chondrocyte hypertrophy and promotes endochondral bone growth.ERN1 acts as a negative regulator of chondrocyte proliferation and differentiation in postnatal growth plates.Its deficiency interrupted PTHrP/PTH1R and IHH homeostasis leading to impaired chondrocyte hypertrophy and differentiation.XBP1s,produced by p-IRE1α-mediated splicing,binds and up-regulates PTH1R and IHH,which coordinate cartilage development.Meanwhile,ER stress cannot be activated normally in ERN1-deficient chondrocytes.In conclusion,ERN1 deficiency accelerates chondrocyte hypertrophy and cartilage mineralization by impairing the homeostasis of the IHH and PTHrP/PTH1R feedback loop and ER stress.ERN1 may have a potential role as a new target for cartilage growth and maturation.

关键词： Cartilage development ER stress ERN1 IHH PTHrP/PTH1R

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Progranulin regulation of autophagy contributes to its chondroprotective effect in osteoarthritis

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Genes & Diseases 2023年第4期10卷 1582-1595页

作者： Yiming Pan Yuyou Yang Mengtian Fan Cheng Chen Rong Jiang Li Liang Menglin Xian Biao Kuang Nana Geng Naibo Feng Lin Deng Wei Zheng Fengmei Zhang Xiaoli Li fengjin guo Department of Cell Biology and Genetics Core Facility of Development BiologyChongqing Medical UniversityChongqing 400016China Department of Orthopaedics The 1st Affiliated Hospital of Chongqing Medical UniversityChongqing 400016China Laboratory of Stem Cells and Tissue Engineering Chongqing Medical UniversityChongqing 400016China Department of Orthopaedics The 2nd Affiliated Hospital of Chongqing Medical UniversityChongqing 400016China Chongqing Medical University Laboratory Animal Center Chongqing Medical UniversityChongqing 400016China

Progranulin(PGRN)is a multifunctional growth factor involved in many physiolog-ical processes and disease states.The apparent protective role of PGRN and the importance of chondrocyte autophagic function in the progression of osteoarthritis(OA)led us to investi-gate the role of PGRN in the regulation of chondrocyte autophagy.PGRN knockout chondro-cytes exhibited a deficient autophagic response with limited induction following rapamycin,serum starvation,and IL-1b-induced autophagy.PGRN-mediated anabolism and suppression of IL-1b-induced catabolism were largely abrogated in the presence of the BafA1 autophagy inhibitor.Mechanistically,during the process of OA,PGRN and the ATG5eATG12 conjugate form a protein complex;PGRN regulates autophagy in chondrocytes and OA through,at least partially,the interactions between PGRN and the ATG5eATG12 conjugate.Furthermore,the ATG5eATG12 conjugate is critical for cell proliferation and apoptosis.Knockdown or knockout of ATG5 reduces the expression of ATG5eATG12 conjugate and inhibits the chondroprotective effect of PGRN on anabolism and catabolism.Overexpression of PGRN partially reversed this effect.In brief,the PGRN-mediated regulation of chondrocyte autophagy plays a key role in the chondroprotective role of PGRN in OA.Such studies provide new insights into the pathogen-esis of OA and PGRN-associated autophagy in chondrocyte homeostasis.

关键词： Anabolism ATG12 ATG5 Autophagy Catabolism Osteoarthritis PGRN

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BRF1 ameliorates LPS-induced inflammation through autophagy crosstalking with MAPK/ ERK signaling

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Genes & Diseases 2018年第3期5卷 226-234页

作者： Weiwei Xie Wei Zheng Min Liu Qizhong Qin Yunpeng Zhao Zhi Cheng fengjin guo Department of Cell Biology and Genetics Core Facility of Development BiologyChongqing Medical UniversityChongqing400016China Department of Orthopaedic Surgery Qilu HospitalShandong UniversityJinan250012China

Inflammation is indispensable for host defense,whereas excessive inflammation often develop inflammatory diseases.Autophagy is thought to be engaged in many extracellular stress responses,such as starvation and innate immunity.Thus,autophagy plays an important role in maintaining homeostasis.The purpose of this study was to elucidate the function of BRF1 in the regulation of inflammation and autophagy response in macrophages.We found that BRF1 inhibited the LPS-induced inflammatory factors expression and the autophagy flux in macrophage.Furthermore,inhibition autophagy with 3-MA can attenuate the suppressive effect of BRF1 on LPS-mediated inflammation.In addition,MAPK/ERK signaling pathway was involved in the BRF1 inhibition inflammation and autophagy in macrophages.These findings indicate that BRF1 attenuates LPS-induced inflammatory factors secretion through autophagy,at least in part,through MAPK/ERK signaling pathway.

关键词： Autophagy BRF1 Inflammation Macrophage MAPK/ERK

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Differentially expressed gene in osteosarcoma cell lines with different metastatic potentials

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Journal of Nanjing Medical University 2009年第5期23卷 352-358页

作者： Xinzhi Li Lin Meng Anming Chen fengjin guo Zhenqiang Luo Heng Zeng Department of Orthopaedics Renhe Hospital Three Gorges University Yichang 443001 China Department of Orthopaedics Tongji Hospital Tongji Medical College Huazhong University of Science and Technology Wuhan 430030 China

Objective： To study the expression of osteosarcoma metastasis associated gene using a cDNA microarray, and screen new candidate genes related＇to the development, progress and osteosarcoma metastasis. Methods： Total RNA of a low metastatic osteosarcoma and a high metastatic osteosarcoma （M6 and M8 cell lines, respectively） was extracted, purified to mRNA and then reverse transcribed to cDNA. M6 was used as the experimental group and M8 as the control group, and the gene expression of cells from both of these two sublines was investigated using cDNA microarrays containig 8064 cDNA clones. The cDNA of M6 was labeled with cy3 and the cDNA of M8 was labeled with cyS. The two sublines were hybridized with the cDNA microarray. The hybridization signals were scanned with a Generation HI array scanner and analyzed by Imagequant 5.0 software. Results： There were 330 differentially expressed genes between M6 and M8. In the M6 subline,152 genes were up-regulated and 178 genes were down-regulated compared to the M8 subline. These genes could be classified according to their function. Cell growth-related genes that were down-regulated included CCNG1, CDC2, APC10,and RPA3, while expression of the tumor suppressor genes, CDKN1A and CDKN2D, was up-regulated. Other genes that were differentially expressed included those that have been implicated in the regulation of signal transduction, metabolism and apoptosis. Conclusion： This study exploits a cDNA microarray approach to identifying genes that may be associated with metastasis. The gene expression profiles of osteosarcoma cell lines is a potentially important index in the search of new candidate genes related to tumor occurrence, development and metastasis.

关键词： Osteosarcoma Gene expression cDNA microarray

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参附注射液对脓毒症大鼠血管内皮损伤的影响及机制

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现代中西医结合杂志 2024年第1期33卷 60-65页

作者：郭倩刘丰进康海郑鑫寅青岛大学医学部山东青岛266300 青岛大学附属烟台毓璜顶医院山东烟台264000

目的探讨早期应用参附注射液对脓毒症大鼠血管内皮损伤的保护作用及其机制。方法将48只成年SD雄性大鼠随机分为假手术组、脓毒症组、参附注射液组,每组16只。脓毒症组、参附注射液组大鼠采用盲肠结扎穿孔法制备脓毒症模型,假手术组仅... 详细信息

目的探讨早期应用参附注射液对脓毒症大鼠血管内皮损伤的保护作用及其机制。方法将48只成年SD雄性大鼠随机分为假手术组、脓毒症组、参附注射液组,每组16只。脓毒症组、参附注射液组大鼠采用盲肠结扎穿孔法制备脓毒症模型,假手术组仅开腹暴露盲肠后还纳腹腔。造模后参附注射液组每隔12 h腹腔注射1次参附注射液(6 mL/kg),假手术组和脓毒症组腹腔注射等体积生理盐水。观察各组大鼠术后48 h内的行为学表现,统计大鼠存活率;术后48 h处死各组存活大鼠,HE染色观察主动脉组织病理学形态,ELISA法检测血清中肿瘤坏死因子-α(TNF-α)、白细胞介素-10(IL-10)、组织因子(TF)、血栓调节蛋白(TM)、血管性血友病因子(vWF)、纤溶酶原激活物抑制剂-1(PAI-1)水平,Western blot法检测主动脉组织中核因子κB抑制蛋白α(IKBα)、核因子-κB(NF-κB)p65蛋白及各自磷酸化情况。结果脓毒症组大鼠毛发束立,对外界反应迟钝,腹泻,鼻及眼部可见渗血;肠道黏膜充血水肿、坏死,光镜下显示主动脉内皮细胞增生、肿胀、脱落,内弹力膜断裂。与脓毒症组比较,参附注射液组大鼠精神和毛发状态、活动度、腹泻、鼻及眼部渗血情况、肠道水肿和坏死情况、光镜下主动脉病理改变均明显好转。术后48 h,参附注射液组大鼠存活率明显高于脓毒症组(P<0.05)。脓毒症组和参附注射液组大鼠外周血中TNF-α、IL-10、TF、TM、vWF、PAI-1水平和主动脉组中p-IKBα、p-NF-κB p65蛋白相对表达量均明显高于假手术组(P均<0.05),参附注射液组外周血中TNF-α、IL-10、TF、TM、vWF、PAI-1水平和主动脉组中p-IKBα、p-NF-κB p65蛋白相对表达量均明显低于脓毒症组(P均<0.05)。结论早期应用参附注射液可抑制脓毒症大鼠机体炎症反应,减轻血管内皮损伤,对脓毒症导致的凝血功能障碍有一定保护作用,机制可能与抑制IKBα/NF-κB信号通路活化有关。

关键词：脓毒症炎症反应内皮损伤凝血功能核因子-κB信号通路

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Expression of OPGmRNA and ODFmRNA in the patients of hip fracture due to osteoporosis

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Journal of Nanjing Medical University 2007年第5期21卷 321-323页

作者： Dan Chen Anmin Chen fengjin guo Department of orthopedics Shengli Oil-field Central Hospital Dongying 257034 Shandong province China Department of orthopedics the Affiliated Tongji Hospital of Tongji Medical College Huazhong University of Science and Technology Wuhan 430030 Hubei province China

Objective：To investigate the gene expression of osteoprotegerin（OPG） and osteoclast differentiation factor（ODF） in the bone tissue of patients with hip fracture due to osteoporosis. Methods：OPGmRNA and ODFmRNA in the bone tissue in 50 cases of osteoporosis sufferers（over 50 years old） with hip fracture（Observer Group） and 30 cases of hip facture sufferers with no osteoporosis（Control group） were analyzed with the Semi-Quantitative RT-PCR method. Results：The mRNA expressed of ODF, OPG were both high in the patients with hip fracture. In the control group, the expression of OPG mRNA was observed, while the expression of ODF mRNA was very slight. Conclusion：Aged patients contained all signals including OPG, ODF that are essential for inducing osteoclastogenesis and promoting bone resorption.

关键词： osteoporosis fracture osteoprotegerin osteoclast differentiation factor

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骨肉瘤中PTEN蛋白表达及临床意义的研究(英文)

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The Chinese-German Journal of Clinical Oncology 2008年第5期7卷 296-299页

作者： Yubin Wang Anmin Chen fengjin guo Yujun Xia Department of Orthopaedics Tongji Hospital Tongji Medical College Huazhong University of Science and Technology Wuhan 430030 China Department of Orthopaedics Central Hospital of Shengli Oil-field Dongying 257034 China Medical College of Qingdao University Qingdao 266011 China

调查癌症的表示和意义的目的在骨肉瘤的禁止的基因 PTEN 蛋白质。在骨肉瘤的不同组织学的分类分析它的表示的水平。决定为诊断骨肉瘤作为标记基因拿 PTEN 蛋白质的可能性。为了观察 PTEN 的临床的值，表达式为骨肉瘤分类作为一个引用索... 详细信息

调查癌症的表示和意义的目的在骨肉瘤的禁止的基因 PTEN 蛋白质。在骨肉瘤的不同组织学的分类分析它的表示的水平。决定为诊断骨肉瘤作为标记基因拿 PTEN 蛋白质的可能性。为了观察 PTEN 的临床的值，表达式为骨肉瘤分类作为一个引用索引铺平。43 个标本从骨肉瘤收集了切除的方法被学习。从操作作为控制组被拿的骨头(骨软骨瘤) 的良性的损害在一样的时期期间收集的 30 个标本。Immunohistochemistry 染色(Elivison™二个步骤方法) 被用来在骨肉瘤的 43 种情况中检测 PTEN 蛋白质的表示。社会科学统计套装软体 10.0 在统计分析被使用。染色的结果 Immunohistochemistry 证明 PTEN 蛋白质的阳性反应都对细胞质面向，它是棕色或微黄色褐的小粒。通过 χ2 测试，在骨头之间的 PTEN 蛋白质的积极表情的有效差量良性的损害和骨肉瘤(χ2= 7.976， P s =−0.4922， P < 0.01 ) 。结论 PTEN 蛋白质可以被用作癌症的候选人基因禁止的基因：PTEN 蛋白质是癌症压制或在骨头肿瘤有表示的基因蛋白质。它可能不仅在肺的癌和神经胶质细胞瘤的学习，而且在骨头肿瘤的学习被使用；PTEN 的表示与骨头肿瘤的亲切和他们区别的度有关。PTEN 的表示断然与区别的度被相关。

关键词： PTEN/MMAC1/TEP1蛋白骨肿瘤免疫组织化学治疗方法

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不同转移潜能MG63骨肉瘤细胞亚株裸鼠肺转移模型的建立(英文)

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The Chinese-German Journal of Clinical Oncology 2007年第5期6卷 484-486页

作者： Wentao Zhu fengjin guo Tao Xu Anming Chen Department ofOrthopedics Tongji Hospital Tongji Medical College Huazhong University of Science and Technology Wuhan 430030 China Department of Rehabilitation Medicine Tongji Hospital Tongji Medical College Huazhong University of Science and Technology Wuhan 430030 China

Objective: To establish a nude mice model of human osteosarcoma lung metastasis. Methods: The growth of human osteosarcoma cell sublines M8 and M6 was determined by MTT assay. 2 × 107 cells were injected into the tail vein of nude mice. Mice were sacrificed started on week 4 after injection, and lung metastases were evaluated under both mac-roscopic and microscopic observation with HE staining. Results: The growth of low-metastatic subline M6 was lower than high-metastatic sublines M8. Seventeen mice after injected M8 had occurred lung metastases while only one mice had oc-curred in M6 group. Moreover, M8 cells within metastases were arrangement disorder with variable nuclear hyperchromasia. Conclusion: A mouse model for human osteosarcoma cancer lung metastasis can be established by injection different ability of metastasis MG63 cells into tail vein.

关键词：转移潜能 MG63 骨肉瘤细胞亚株裸鼠肺转移

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Study on Inhibitionof Growth of ACHN Cells via shRNA Expression Vector-Mediated Cycline1 Gene Silencing

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Chinese Journal of Clinical Oncology 2009年第1期6卷 29-35页

作者： Jiang Wang Anmin Chen fengjin guo Department of Orthopedics Tongji HospitalTongji Medical College Huazhong Universityof Science and Technology Wuhan 430030Hubei Province. China.

OBJECTIVE To explore the inhibition of ACHN cells via shRNAexpression vector mediated cyclinE1 gene silencing.METHODS The shRNA targeting at cyclinE1 gene was designedand synthesized. By ligation, the fragment was inserted intopGenesil-1-U6 to construct the recombinant plasmid pGenesil-1-U6-cyclinE1. The identified recombinant plasmid was introducedinto ACHN cells with lipofectamine 2000. The inhibition ofcyclinE1 mRNA and protein expression were analyzed by RT-PCRand western-blotting. MTT method was used for observing cellproliferation and drawing growth curve. The cell cycle and ratiosof apoptotic cell were assessed by flow cytometric detection. Theability of invasion and speed of cell migration were detected bytranswell chamber invasive models and cell scratch method.RESULTS The inhibition of expression of cyclinE1 in ACHN cellsmediated by recombinant vector (0.0933 ± 0.05) was significantlylower than that in the group of transfected with empty vector(0.8827 ± 0.04) and the control group (0.9021 ± 0.03) (P < 0.05).Flow cytometry showed that recombinant cells were blocked inthe G_1 phase and the apoptotic ratio was increased significantly(11.15 ± 4.00)% (P < 0.05). The curves of cell growth indicated thatthe proliferation of cell transfected with recombinant plasmid wasinhibited significantly compared with that in control group (P <0.05). The results of transwell and cell scratch suggested that theabilities of invasion and migration of the cells transfected withrecombinant plasmid were decreased conspicuously (P < 0.05).CONCLUSION The expression of cyclinE1 could be inhibitedsuccessfully by RNA interference induced by shRNA expressionvector. This consequently inhibits the cell growth and inducesapoptosis. Our study provided a preliminary result in searching ofRNA interference (RNAi) therapy for renal cell carcinoma.

关键词：细胞表达 ACHN 载体介导生长曲线基因沉默 Transwell 流式细胞仪检测 Western印迹

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Inhibition of Proliferation of Human Osteosarcoma Cells Transfected with PIN1 Antisense Gene

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The Chinese-German Journal of Clinical Oncology 2006年第4期5卷 294-297页

作者： XIONG Wenhua CHEN Anmin guo fengjin Department of Orthopaedics Tongji Hospital Tongji Medical College Huazhong University of Science and Technology Wuhan 430030 China

客观：与 Pin1 反感觉基因评估人的 osteosarcomacells transfected 的增长的抑制。方法：反的不同剂量察觉到 Pin1 基因(0， 20,50， 100， 200， 250 μ L ) 是进骨肉瘤 MG-63 房间的 transfected。在 transfection 前后的房间和文化... 详细信息

客观：与 Pin1 反感觉基因评估人的 osteosarcomacells transfected 的增长的抑制。方法：反的不同剂量察觉到 Pin1 基因(0， 20,50， 100， 200， 250 μ L ) 是进骨肉瘤 MG-63 房间的 transfected。在 transfection 前后的房间和文化上层清液是镇定的。房间生长的曲线被 MTTmethod 做。细胞生长骑车， apoptosis 被 FCM 检测。Pin1 的表示被检测由西方污点并且由聚合酶链反应(RT-PCR ) 的 Pin1 mRNA 的 respectively.Results：MTT 和 FCM 试金显示由反的 transfection 察觉到 Pin1 基因能 inhibitMG-63 增长和 induce apoptosis。西方污点的试金表明反察觉到房间有与反感觉 Pin1gene 的没有 transfected 的比那些的更弱的染色，并且染色紧张的 Pin1gene-transfected MG-63 否定地与剂量被联系。由基因的不同剂量的房间 transfected (0， 20， 50， 100， 200， 250 μ L ) 有的不同吸收度评价：0.854 ± 0 .136,0.866 ± 0 .138， 0.732 ± 0 .154， 0.611 ± 0 .121， 0.547 ± 0 .109， 0.398 ± 0 .113， 0.320 ± 0 .151 分别地，与由 F 和 q 是重要的差别测试(P < 0.05 ) 。类似的结果和它的吸收度评估的有的 Pin1 mRNA 的表示是 0.983 ± 0 .125， 0.988 ± 0 .127， 0.915 ± 0 .157,0.786 ± 0 .125， 0.608 ± 0 .124， 0.433 ± 0 .130， 0.410 ± 0 .158 分别地(P < 0.05 ) 。结论：在 MG-63 房间的 Pin1 mRNA 的 Theexpression 能被反感觉 Pin1 基因禁止，因此减少 Pin1 的表示并且压抑人的骨肉瘤房间 MG-63 的增长。

关键词：骨肉瘤肿瘤细胞核苷酸基因转染

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