Publications by authors named "Xishuai Tong"

10 Publications

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Beclin 1 positively regulates osteoprotegerin-induced inhibition of osteoclastogenesis by increasing autophagy in vitro.

Differentiation 2021 Aug 23;121:35-43. Epub 2021 Aug 23.

Institutes of Agricultural Science and Technology Development, Joint International Research Laboratory of Agriculture and Agri-Product Safety of the Ministry of Education of China, Yangzhou University, Yangzhou, 225009, Jiangsu, PR China; College of Veterinary Medicine, Yangzhou University, Yangzhou, 225009, Jiangsu, PR China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, 225009, Jiangsu, PR China; Jiangsu Key Laboratory of Zoonosis, Yangzhou, 225009, Jiangsu, PR China. Electronic address:

Osteoclastogenesis is induced by receptor activator of nuclear factor-κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF), and can be suppressed by osteoprotegerin (OPG). Beclin1 has a dual role in osteoclastogenesis. However, the role of Beclin1-mediated autophagy during OPG-induced inhibition of osteoclastogenesis remains unclear. Here, we found that Beclin1 and matrix metalloproteinase 9 (MMP-9) expression were increased during osteoclastogenesis. OPG (20, 40, and 80 ng/mL) decreased Src and MMP-9 expression, but augmented Beclin1 expression and fluorescence intensity. Similarly, treatment with the autophagy activator rapamycin increased Beclin1 expression during OPG-induced inhibition of osteoclastogenesis. Further, Beclin1 knockdown restored osteoclast numbers by reducing autophagy during OPG-induced inhibition of osteoclastogenesis. These results indicate that Beclin1 has a positive role during OPG-induced inhibition of osteoclastogenesis by regulating autophagy, which might provide a potential basis for osteoclastogenesis.
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http://dx.doi.org/10.1016/j.diff.2021.08.003DOI Listing
August 2021

Cadmium Toxicity on Chondrocytes and the Palliative Effects of 1α, 25-Dihydroxy Vitamin D in White Leghorns Chicken's Embryo.

Front Vet Sci 2021 10;8:637369. Epub 2021 Feb 10.

College of Veterinary Medicine, Yangzhou University, Yangzhou, China.

Cadmium (Cd) can causes osteoporosis and joint swelling. However, the mechanism of Cd toxicity in chondrocytes and how to alleviate Cd poisoning to chondrocytes are still unclear. Herein, we evaluated the toxicity of Cd to chicken chondrocytes, and whether vitamin D can relieve the toxicity of Cd to chondrocytes. Primary chondrocytes were collected from knee-joint cartilage of 15-day-old chicken embryos. They were treated with (0, 1, 2, and 4) μM Cd alone, 10 M 1α,25-(OH)D alone, or 2 μM Cd combined with 10 M 1α,25-(OH)D. We found that Cd significantly inhibited and mRNA expression, which are markers for chondrocyte differentiation, downregulated the mitochondrial membrane potential, upregulated the Bax/B-cell lymphoma 2 ratio. Furthermore, Cd significantly promoted matrix metalloproteinase (MMP)-9 expression, thus accelerating the degradation of extracellular matrix. And Cd also inhibited the expression of main macromolecular protein of extracellular matrix, Collagen type IIα1 (COL2A1) and acid mucopolysaccharide. However, 1α,25-(OH)D pretreatment significantly alleviated the toxicity effects of Cd on the differentiation, apoptosis and extracellular matrix gene expression in primary chondrocytes. Conclusively, Cd exposure could inhibited chicken embryo chondrocytes differentiation, extracellular matrix gene expression, and induced chondrocyte apoptosis. However, these toxic effects of Cd are alleviated by the pretreatment of chondrocytes with 1α,25-(OH)D.
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http://dx.doi.org/10.3389/fvets.2021.637369DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7902530PMC
February 2021

Overexpression of c-Fos reverses osteoprotegerin-mediated suppression of osteoclastogenesis by increasing the Beclin1-induced autophagy.

J Cell Mol Med 2021 01 4;25(2):937-945. Epub 2020 Dec 4.

Institutes of Agricultural Science and Technology Development, Joint International Research Laboratory of Agriculture and Agri-Product Safety of the Ministry of Education of China, Yangzhou University, Yangzhou, China.

Osteoclastogenesis requires the involvement of transcription factors and degrading enzymes, and is regulated by upstream and downstream signalling. However, c-Fos how regulates osteoclastogenesis through autophagy remain unclear. This study aimed to explore the role of c-Fos during osteoprotegerin (OPG)-mediated suppression of osteoclastogenesis. We found that the number of osteoclasts and the expression of c-Fos, MMP-9, CAⅡ, Src and p62 were decreased after treated with OPG, including attenuation the PI3K/Akt and the TAK1/S6 signalling pathways, but the expression of Beclin1 and LC3Ⅱ were increased. Knockdown of Beclin1 could reverse the expression of c-Fos and MMP-9 by activating the PI3K/Akt signalling pathway, but inhibiting the autophagy and the TAK1/S6 signalling pathway. In addition, inhibition of autophagy using the PI3K inhibitor LY294002 did not rescues OPG-mediated suppression of osteoclastogenesis, but caused reduction of the expression of c-Fos and CAⅡ by attenuating the autophagy, as well as the PI3K/Akt and the TAK1/S6 signalling pathways. Furthermore, continuous activation of c-Fos could reverse OPG-mediated suppression of osteoclastogenesis by activating the autophagy and the PI3K/Akt and the TAK1/S6 signalling pathways. Thus, overexpression of c-Fos could reverse OPG-mediated suppression of osteoclastogenesis via activation of Beclin1-induced autophagy, indicating c-Fos might serve as a new candidate for bone-related basic studies.
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http://dx.doi.org/10.1111/jcmm.16152DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7812271PMC
January 2021

p53 positively regulates osteoprotegerin-mediated inhibition of osteoclastogenesis by downregulating TSC2-induced autophagy in vitro.

Differentiation 2020 Jul - Aug;114:58-66. Epub 2020 Jun 21.

College of Veterinary Medicine, Yangzhou University, Yangzhou, 225009, Jiangsu, PR China; Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, 225009, Jiangsu, PR China; Jiangsu Key Laboratory of Zoonosis, Yangzhou, 225009, Jiangsu, PR China; Joint International Research Laboratory of Agriculture and Agri-Product Safety, The Ministry of Education of China, Yangzhou University, Yangzhou, 225009, Jiangsu, PR China. Electronic address:

Osteoclasts are terminally multinucleated cells that are regulated by nuclear factor-activated T cells c1 (NFATc1), and are responsible for bone resorption while the tartrate resistant acid phosphatase (TRAP) enzymes releases into bone resorption lacunae. Furthermore, tumor suppressor p53 is a negative regulator during osteoclastogenesis. Osteoprotegerin (OPG) inhibits osteoclastogenesis and bone resorption by activating autophagy, however, whether p53 is involved in OPG-mediated inhibition of osteoclastogenesis remains unclear. In the current study, OPG could enhance the expression of p53 and tuberin sclerosis complex 2 (TSC2). Moreover, the expression of p53 is regulated by autophagy during OPG-mediated inhibition of osteoclastogenesis. Inhibition of p53 by treated with pifithrin-α (PFTα) causing augments of osteoclastogenesis and bone resorption, also reversed OPG-mediated inhibition of osteoclastogenesis by reducing the expression of TSC2. In addition, knockdown of TSC2 using siRNA could rescue OPG-mediated inhibition of osteoclastogenesis by reducing autophagy, which is manifested by the decrease of the expression of Beclin1 and the phosphorylation of mammalian target of rapamycin (mTOR) and ribosomal protein S6 kinase beta 1 (S6K1, also known as p70S6K). Collectively, p53 plays a critical role during OPG-mediated inhibition of osteoclastogenesis via regulating the TSC2-induced autophagy in vitro.
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http://dx.doi.org/10.1016/j.diff.2020.06.002DOI Listing
July 2021

AMP-activated protein kinase (AMPK) regulates autophagy, inflammation and immunity and contributes to osteoclast differentiation and functionabs.

Biol Cell 2020 Sep 8;112(9):251-264. Epub 2020 Jun 8.

College of Veterinary Medicine, Yangzhou University, Yangzhou, Jiangsu, 225009, People's Republic of China.

Osteoclasts are multinucleated giant cells, responsible for bone resorption. Osteoclast differentiation and function requires a series of cytokines to remove the old bone, which coordinates with the induction of bone remodelling by osteoblast-mediated bone formation. Studies have demonstrated that AMP-activated protein kinase (AMPK) play a negative regulatory role in osteoclast differentiation and function. Research involving AMPK, a nutrient and energy sensor, has primarily focused on osteoclast differentiation and function; thus, its role in autophagy, inflammation and immunity remains poorly understood. Autophagy is a conservative homoeostatic mechanism of eukaryotic cells, and response to osteoclast differentiation and function; however, how it interacts with inflammation remains unclear. Additionally, based on the regulatory function of different AMPK subunits for osteoclast differentiation and function, its activation is regulated by upstream factors to perform bone metabolism. This review summarises the critical role of AMPK-mediated autophagy, inflammation and immunity by upstream and downstream signalling during receptor activator of nuclear factor kappa-B ligand-induced osteoclast differentiation and function. This pathway may provide therapeutic targets for bone-related diseases, as well as function as a biomarker for bone homoeostasis.
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http://dx.doi.org/10.1111/boc.202000008DOI Listing
September 2020

Vitamin D Inhibition of TRPV5 Expression During Osteoclast Differentiation.

Int J Endocrinol Metab 2019 Oct 14;17(4):e91583. Epub 2019 Oct 14.

College of Veterinary Medicine, Yangzhou University, Yangzhou, China.

Background: Vitamin D is an important steroid that can regulate bone metabolism including osteoclast (OC) differentiation. Transient receptor potential cation channel subfamily V member 5 (TRPV5), is a calcium channel protein involved in OC differentiation. However, the impact of vitamin D on TRPV5 expression during OC differentiation is not clear.

Objectives: To determine if 1,25-dihydroxyvitamin D3 (1,25(OH)D) regulates the expression of TRPV5 during OC differentiation.

Methods: Bone marrow mononuclear macrophage (BMMs) were induced to differentiate into OC with or without treatment with 10 nM 1,25(OH)D. The expression levels of vitamin D receptor (VDR) and TRPV5 were examined. The expression of several OC markers, including tartrate resistant acid phosphatase (TRAP), carbonic anhydrase II (Ca II), cathepsin K (CTSK), and vacuolar-type H-ATPase (V-ATPase) were also detected.

Results: We found that the VDR was expressed in murine bone marrow-derived macrophages at the early stage of OC differentiation. TRPV5 expression was increased during OC differentiation, which was down-regulated by 1,25(OH)D after a prolonged exposure. The 1,25(OH)D and TRPV5 inhibitors inhibited OC differentiation.

Conclusions: 1,25(OH)D can inhibit TRPV5 expression as well as TRPV5 inhibitors during OC differentiation. This suggests that 1,25(OH)D may suppress OC differentiation by inhibiting TRPV5 expression.
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http://dx.doi.org/10.5812/ijem.91583DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6948119PMC
October 2019

Suppression of AMP-activated protein kinase reverses osteoprotegerin-induced inhibition of osteoclast differentiation by reducing autophagy.

Cell Prolif 2020 Jan 7;53(1):e12714. Epub 2019 Nov 7.

College of Veterinary Medicine, Yangzhou University, Yangzhou, Jiangsu, China.

Objectives: Osteoclasts (OC) are unique terminally differentiated cells whose primary function is bone resorption. We previously showed that osteoprotegerin (OPG) inhibits OC differentiation in vitro by enhancing autophagy via the adenosine monophosphate-activated protein kinase (AMPK)/mTOR/p70S6K signalling pathway in vitro. Here, we aimed to elucidate the mechanism of AMPK mediated autophagy to regulate OPG-mediated inhibition of OC differentiation and identify potential therapeutic targets associated with bone loss.

Materials And Methods: We used the AMPK activator AICAR to determine the relationship between AMPK activation and OC differentiation, and studied the role of AMPK-mediated autophagy in OPG-mediated inhibition of OC differentiation by using autophagy inhibitors or AMPK knockdown.

Results: AMP-activated protein kinase activation caused LC3II accumulation and weakened OC differentiation activity. In contrast, inactivation of autophagy by 3-methyladenine or Bafilomycin A1 could attenuate OPG-mediated inhibition of OC differentiation via the AMPK/mTOR/p70S6K signalling pathway. Furthermore, the AMPK inhibitor compound C and knockdown of AMPK impaired OPG-mediated inhibition of OC differentiation by inducing autophagy.

Conclusions: These results demonstrated that the AMPK signalling pathway functions as a critical regulator in the OPG-mediated inhibition of OC differentiation, by inducing autophagy. Our results provide a basis for future bone-related studies on the AMPK signalling pathway.
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http://dx.doi.org/10.1111/cpr.12714DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6985670PMC
January 2020

3-(4-methoxyl)-1-(2-(4-coumarin)prop)-2-en-1-one inhibits the differentiation of Gaoyou duck embryonic osteoclasts in vitro.

Poult Sci 2019 Apr;98(4):1854-1860

State Key Laboratory Breeding Base of Green Pesticide and Agricultural Bioengineering, Guizhou University, Huaxi North Campus, Guiyang, CN 550025, China.

This study determined the influence of 3-(4-methoxyl)-1-(2-(4-coumarin)prop)-2-en-1-one (MCPEO) on the differentiation of Gaoyou duck embryo osteoclasts cultured in vitro. Bone marrow mononuclear cells (BM-MNCs) were harvested from 23-day-old Gaoyou duck embryos and induced by receptor activator of nuclear factor κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) in the presence of MCPEO at different concentrations (i.e., 1, 5, 10, 20, and 40 μM). Cell viability measurement, tartrate-resistant acid phosphatase (TRAP) staining, resorption activity assay, and co-staining with Tetramethylrhodamine (TRITC)-conjugated phalloidin and Hoechst 33,258 were conducted. Results indicated that MCPEO influenced the cell viability of the M-CSF + RANKL-induced BM-MNCs in a concentration-dependent manner, reduced the formation of positive multinucleated cells, and restrained the resorption capability of osteoclasts. Microfilament and nuclear staining indicated that MCPEO restricted the differentiation of BM-MNCs into large multinucleated osteoclasts. In short, MCPEO can inhibit the differentiation of BM-MNCs into mature osteoclasts in duck embryos. Therefore, MCPEO is a promising agent for the treatment of poultry osteoporosis.
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http://dx.doi.org/10.3382/ps/pey542DOI Listing
April 2019

Osteoprotegerin inhibit osteoclast differentiation and bone resorption by enhancing autophagy via AMPK/mTOR/p70S6K signaling pathway in vitro.

J Cell Biochem 2018 Sep 6. Epub 2018 Sep 6.

College of Veterinary Medicine, Yangzhou University, Yangzhou, China.

Osteoclasts are highly differentiated terminal cells formed by fusion of hematopoietic stem cells. Previously, osteoprotegerin (OPG) inhibit osteoclast differentiation and bone resorption by blocking receptor activator of nuclear factor-κB ligand (RANKL) binding to RANK indirect mechanism. Furthermore, autophagy plays an important role during osteoclast differentiation and function. However, whether autophagy is involved in OPG-inhibited osteoclast formation and bone resorption is not known. To elucidate the role of autophagy in OPG-inhibited osteoclast differentiation and bone resorption, we used primary osteoclast derived from mice bone marrow monocytes/macrophages (BMM) by induced M-CSF and RANKL. The results showed that autophagy-related proteins expression were upregulated; tartrate-resistant acid phosphatase-positive osteoclast number and bone resorption activity were decreased; LC3 puncta and autophagosomes number were increased and activated AMPK/mTOR/p70S6K signaling pathway. In addition, chloroquine (as the autophagy/lysosome inhibitor, CQ) or rapamycin (as the autophagy/lysosome inhibitor, Rap) attenuated osteoclast differentiation and bone resorption activity by OPG treatment via AMPK/mTOR/p70S6K signaling pathway. Our data demonstrated that autophagy plays a critical role in OPG inhibiting osteoclast differentiation and bone resorption via AMPK/mTOR/p70S6K signaling pathway in vitro.
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http://dx.doi.org/10.1002/jcb.27468DOI Listing
September 2018

Inhibition of osteoclast bone resorption activity through osteoprotegerin-induced damage of the sealing zone.

Int J Mol Med 2014 Sep 10;34(3):856-62. Epub 2014 Jul 10.

College of Veterinary Medicine, Yangzhou University, and Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu 225009, P.R. China.

Bone remodeling is dependent on the dynamic equilibrium between osteoclast-mediated bone resorption and osteoblast-mediated osteogenesis. The sealing zone is an osteoclast-specific cytoskeletal structure, the integrity of which is critical for osteoclast-mediated bone resorption. To date, studies have focused mainly on the osteoprotegerin (OPG)‑induced inhibition of osteoclast differentiation through the OPG/receptor activator of the nuclear factor kappa-B ligand (RANKL)/RANK system, which affects the bone resorption of osteoclasts. However, the effects of OPG on the sealing zone have not been reported to date. In this study, the formation of the sealing zone was observed by Hoffman modulation contrast (HMC) microscopy and confocal laser scanning microscopy. The effects of OPG on the existing sealing zone and osteoclast-mediated bone resorption activity, as well as the regulatory role of genes involved in the formation of the sealing zone were examined by immunofluorescence staining, HMC microscopy, quantitative reverse transcription polymerase chain reaction (RT-qPCR), western blot analysis and scanning electron microscopy. The sealing zone was formed on day 5, with belt-like protuberances at the cell edge and scattered distribution of cell nuclei, but no filopodia. The sealing zone was intact in the untreated control group. However, defects in the sealing zone were observed in the OPG-treated group (20 ng/ml) and the structure was absent in the groups treated with 40 and 80 ng/ml OPG. The podosomes showed a scattered or clustered distribution between the basal surface of the osteoclasts and the well surface. Furthermore, resorption lacunae were not detected in the 20 ng/ml OPG-treated group, indicating the loss of osteoclast-mediated bone resorption activity. Treatment with OPG resulted in a significant decrease in the expression of Arhgef8/Net1 and DOCK5 Rho guanine nucleotide exchange factors (RhoGEFs), 10 of 18 RhoGTPases (RhoA, RhoB, cdc42v1, cdc42v2, RhoU/Wrch1, RhoF/Rif, Rac2, RhoG, Rnd1 and RhoBTB1), ROCK1 and ROCK2. In conclusion, podosome distribution was affected by the OPG-induced inhibition of the expression of genes in the RhoGTPase signaling pathway. This resulted in damage to or destruction of the sealing zone, thus inhibiting osteoclast-mediated bone resorption activity.
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http://dx.doi.org/10.3892/ijmm.2014.1846DOI Listing
September 2014
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