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Crosstalk of renal cell carcinoma cells and tumor-associated macrophages aggravates tumor progression by modulating muscleblind-like protein 2/B-cell lymphoma 2/beclin 1-mediated autophagy

Published:October 14, 2022DOI:https://doi.org/10.1016/j.jcyt.2022.09.001

      Abstract

      Background aims

      M2-polarized tumor-associated macrophages contribute to the development of multiple human cancers, including renal cell carcinoma (RCC). However, the crosstalk mechanism between M2 macrophages and RCC remains unclear.

      Methods

      The authors constructed a co-culture system of M2 macrophages differentiated from THP-1 and RCC cells. Microscopic examination and quantitative real‑time polymerase chain reaction (qRT-PCR) validated the morphology and types of macrophages. The proliferation, migration and invasion of RCC cells were assessed by Cell Counting Kit 8 (Dojindo Molecular Technologies, Inc, Santa Clara, CA, USA) and Transwell assay (Corning, Corning, NY, USA). Messenger RNA (mRNA) and protein expression of target molecules was detected by qRT‑PCR and western blotting. Expression of Ki-67, E-cadherin and N-cadherin was measured by immunofluorescence staining or immunohistochemistry. Molecular interaction was evaluated by RNA pull-down, RNA immunoprecipitation and co-immunoprecipitation. A xenograft model was established to determine tumor growth in vivo.

      Results

      RCC cells triggered the activation of M2 macrophages. Functionally, M2-polarized macrophages facilitated the growth, migration, invasion and epithelial–mesenchymal transition of RCC cells by suppressing autophagy, whereas rapamycin, an activator of autophagy, significantly counteracted the tumor-promoting effects of M2 macrophages. Mechanistically, M2 macrophage-derived C-C motif chemokine 2 (CCL2) enhanced modulation of muscleblind-like protein 2 (MBNL2) expression. MBNL2 raised the stability of B-cell lymphoma 2 (Bcl-2) by directly binding to Bcl-2 mRNA, which endowed RCC cells with malignant properties via inhibition of beclin 1-dependent autophagy.

      Conclusions

      RCC-induced M2-polarized macrophages secrete CCL2 to promote the growth and metastasis of RCC cells via inhibition of MBNL2/Bcl-2/beclin 1-mediated autophagy, which provide a novel perspective for the development of a therapeutic strategy for -RCC.

      Key Words

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      References

        • Chen W.
        • Zheng R.
        • Baade P.D.
        • Zhang S.
        • Zeng H.
        • Bray F.
        • Jemal A.
        • Yu X.Q.
        • He J.
        Cancer statistics in China, 2015.
        CA Cancer J Clin. 2016; 66: 115-132
        • Siegel R.L.
        • Miller K.D.
        • Jemal A.
        Cancer statistics, 2019.
        CA Cancer J Clin. 2019; 69: 7-34
        • Qin Z.
        • Hu H.
        • Sun W.
        • Chen L.
        • Jin S.
        • Xu Q.
        • et al.
        miR-224-5p Contained in Urinary Extracellular Vesicles Regulates PD-L1 Expression by Inhibiting Cyclin D1 in Renal Cell Carcinoma Cells.
        Cancers (Basel). 2021; 13: 618-637
        • Harada K.
        • Nozawa M.
        • Uemura M.
        • Tatsugami K.
        • Osawa T.
        • Yamana K.
        • Kimura G.
        • Fujisawa M.
        • Nonomura N.
        • Eto M.
        • Shinohara N.
        • Tomita Y.
        • Kondo Y.
        • Ochi K.
        • Anazawa Y.
        • Uemura H.
        Treatment patterns and outcomes in patients with unresectable or metastatic renal cell carcinoma in Japan.
        Int J Urol. 2019; 26: 202-210
        • Choueiri T.K.
        • Motzer R.J.
        Systemic Therapy for Metastatic Renal-Cell Carcinoma.
        N Engl J Med. 2017; 376: 354-366
        • DeNardo D.G.
        • Ruffell B.
        Macrophages as regulators of tumour immunity and immunotherapy.
        Nat Rev Immunol. 2019; 19: 369-382
        • Lawrence T.
        • Natoli G.
        Transcriptional regulation of macrophage polarization: enabling diversity with identity.
        Nat Rev Immunol. 2011; 11: 750-761
        • Cannarile M.A.
        • Ries C.H.
        • Hoves S.
        • Ruttinger D.
        Targeting tumor-associated macrophages in cancer therapy and understanding their complexity.
        Oncoimmunology. 2014; 3e955356
        • Komohara Y.
        • Hasita H.
        • Ohnishi K.
        • Fujiwara Y.
        • Suzu S.
        • Eto M.
        • Takeya M.
        Macrophage infiltration and its prognostic relevance in clear cell renal cell carcinoma.
        Cancer Sci. 2011; 102: 1424-1431
        • Xie Y.
        • Chen Z.
        • Zhong Q.
        • Zheng Z.
        • Chen Y.
        • Shangguan W.
        • Zhang Y.
        • Yang J.
        • Zhu D.
        • Xie W.
        M2 macrophages secrete CXCL13 to promote renal cell carcinoma migration, invasion, and EMT.
        Cancer Cell Int. 2021; 21: 677
        • Davidsson S.
        • Fiorentino M.
        • Giunchi F.
        • Eriksson M.
        • Erlandsson A.
        • Sundqvist P.
        • Carlsson J.
        Infiltration of M2 Macrophages and Regulatory T Cells Plays a Role in Recurrence of Renal Cell Carcinoma.
        Eur Urol Open Sci. 2020; 20: 62-71
        • Jones T.M.
        • Carew J.S.
        • Nawrocki S.T.
        Therapeutic Targeting of Autophagy for Renal Cell Carcinoma Therapy.
        Cancers (Basel). 2020; 12: 1185-1200
        • Yildirim D.
        • Bender O.
        • Karagoz Z.F.
        • Helvacioglu F.
        • Bilgic M.A.
        • Akcay A.
        • Ruzgaresen N.B.
        Role of autophagy and evaluation the effects of microRNAs 214, 132, 34c and prorenin receptor in a rat model of focal segmental glomerulosclerosis.
        Life Sci. 2021; 280119671
        • Yang Z.J.
        • Chee C.E.
        • Huang S.
        • Sinicrope F.A.
        The role of autophagy in cancer: therapeutic implications.
        Mol Cancer Ther. 2011; 10: 1533-1541
        • Jin Y.
        • Huang R.
        • Xia Y.
        • Huang C.
        • Qiu F.
        • Pu J.
        • He X.
        • Zhao X.
        Long Noncoding RNA KIF9-AS1 Regulates Transforming Growth Factor-beta and Autophagy Signaling to Enhance Renal Cell Carcinoma Chemoresistance via microRNA-497-5p.
        DNA Cell Biol. 2020; 39: 1096-1103
        • He Y.H.
        • Tian G.
        Autophagy as a Vital Therapy Target for Renal Cell Carcinoma.
        Front Pharmacol. 2020; 11518225
        • Fu D.
        • Wu D.
        • Cheng W.
        • Gao J.
        • Zhang Z.
        • Ge J.
        • Zhou W.
        • Xu Z.
        Costunolide Induces Autophagy and Apoptosis by Activating ROS/MAPK Signaling Pathways in Renal Cell Carcinoma.
        Front Oncol. 2020; 10582273
        • Fu X.T.
        • Song K.
        • Zhou J.
        • Shi Y.H.
        • Liu W.R.
        • Shi G.M.
        • Gao Q.
        • Wang X.Y.
        • Ding Z.B.
        • Fan J.
        Tumor-associated macrophages modulate resistance to oxaliplatin via inducing autophagy in hepatocellular carcinoma.
        Cancer Cell Int. 2019; 19: 71
        • Fischer S.
        • Di Liddo A.
        • Taylor K.
        • Gerhardus J.S.
        • Sobczak K.
        • Zarnack K.
        • Weigand J.E.
        Muscleblind-like 2 controls the hypoxia response of cancer cells.
        RNA. 2020; 26: 648-663
        • Perron G.
        • Jandaghi P.
        • Solanki S.
        • Safisamghabadi M.
        • Storoz C.
        • Karimzadeh M.
        • Papadakis A.I.
        • Arseneault M.
        • Scelo G.
        • Banks R.E.
        • Tost J.
        • Lathrop M.
        • Tanguay S.
        • Brazma A.
        • Huang S.
        • Brimo F.
        • Najafabadi H.S.
        • Riazalhosseini Y.
        A General Framework for Interrogation of mRNA Stability Programs Identifies RNA-Binding Proteins that Govern Cancer Transcriptomes.
        Cell Rep. 2018; 23: 1639-1650
        • Charizanis K.
        • Lee K.Y.
        • Batra R.
        • Goodwin M.
        • Zhang C.
        • Yuan Y.
        • Shiue L.
        • Cline M.
        • Scotti M.M.
        • Xia G.
        • Kumar A.
        • Ashizawa T.
        • Clark H.B.
        • Kimura T.
        • Takahashi M.P.
        • Fujimura H.
        • Jinnai K.
        • Yoshikawa H.
        • Gomes-Pereira M.
        • Gourdon G.
        • Sakai N.
        • Nishino S.
        • Foster T.C.
        • Ares Jr., M.
        • Darnell R.B.
        • Swanson M.S.
        Muscleblind-like 2-mediated alternative splicing in the developing brain and dysregulation in myotonic dystrophy.
        Neuron. 2012; 75: 437-450
        • Zhang J.
        • Zhang S.
        • Shi Q.
        • Allen T.D.
        • You F.
        • Yang D.
        The anti-apoptotic proteins Bcl-2 and Bcl-xL suppress Beclin 1/Atg6-mediated lethal autophagy in polyploid cells.
        Exp Cell Res. 2020; 394112112
        • He Y.
        • Liu J.
        • Wang Y.
        • Zhu X.
        • Fan Z.
        • Li C.
        • Yin H.
        • Liu Y.
        Role of miR-486-5p in regulating renal cell carcinoma cell proliferation and apoptosis via TGF-beta-activated kinase 1.
        J Cell Biochem. 2019; 120: 2954-2963
        • Pattingre S.
        • Tassa A.
        • Qu X.
        • Garuti R.
        • Liang X.H.
        • Mizushima N.
        • Packer M.
        • Schneider M.D.
        • Levine B.
        Bcl-2 antiapoptotic proteins inhibit Beclin 1-dependent autophagy.
        Cell. 2005; 122: 927-939
        • Hsieh J.J.
        • Purdue M.P.
        • Signoretti S.
        • Swanton C.
        • Albiges L.
        • Schmidinger M.
        • Heng D.Y.
        • Larkin J.
        • Ficarra V.
        Renal cell carcinoma.
        Nat Rev Dis Primers. 2017; 3: 17009
        • Katayama H.
        • Tamai K.
        • Shibuya R.
        • Nakamura M.
        • Mochizuki M.
        • Yamaguchi K.
        • Kawamura S.
        • Tochigi T.
        • Sato I.
        • Okanishi T.
        • Sakurai K.
        • Fujibuchi W.
        • Arai Y.
        • Satoh K.
        Long non-coding RNA HOTAIR promotes cell migration by upregulating insulin growth factor-binding protein 2 in renal cell carcinoma.
        Sci Rep. 2017; 7: 12016
        • Yin Y.
        • Yao S.
        • Hu Y.
        • Feng Y.
        • Li M.
        • Bian Z.
        • Zhang J.
        • Qin Y.
        • Qi X.
        • Zhou L.
        • Fei B.
        • Zou J.
        • Hua D.
        • Huang Z.
        The Immune-microenvironment Confers Chemoresistance of Colorectal Cancer through Macrophage-Derived IL6.
        Clin Cancer Res. 2017; 23: 7375-7387
        • Wei C.
        • Yang C.
        • Wang S.
        • Shi D.
        • Zhang C.
        • Lin X.
        • Liu Q.
        • Dou R.
        • Xiong B.
        Crosstalk between cancer cells and tumor associated macrophages is required for mesenchymal circulating tumor cell-mediated colorectal cancer metastasis.
        Mol Cancer. 2019; 18: 64
        • Qian B.Z.
        • Pollard J.W.
        Macrophage diversity enhances tumor progression and metastasis.
        Cell. 2010; 141: 39-51
        • Zhao S.
        • Mi Y.
        • Guan B.
        • Zheng B.
        • Wei P.
        • Gu Y.
        • Zhang Z.
        • Cai S.
        • Xu Y.
        • Li X.
        • He X.
        • Zhong X.
        • Li G.
        • Chen Z.
        • Li D.
        Tumor-derived exosomal miR-934 induces macrophage M2 polarization to promote liver metastasis of colorectal cancer.
        J Hematol Oncol. 2020; 13: 156
        • Mantovani A.
        • Sozzani S.
        • Locati M.
        • Allavena P.
        • Sica A.
        Macrophage polarization: tumor-associated macrophages as a paradigm for polarized M2 mononuclear phagocytes.
        Trends Immunol. 2002; 23: 549-555
        • Peltanova B.
        • Raudenska M.
        • Masarik M.
        Effect of tumor microenvironment on pathogenesis of the head and neck squamous cell carcinoma: a systematic review.
        Mol Cancer. 2019; 18: 63
        • Pirila E.
        • Vayrynen O.
        • Sundquist E.
        • Pakkila K.
        • Nyberg P.
        • Nurmenniemi S.
        • Paakkonen V.
        • Pesonen P.
        • Dayan D.
        • Vered M.
        • Uhlin-Hansen L.
        • Salo T.
        Macrophages modulate migration and invasion of human tongue squamous cell carcinoma.
        PLoS One. 2015; 10e0120895
        • Gu W.
        • Gong L.
        • Wu X.
        • Yao X.
        Hypoxic TAM-derived exosomal miR-155-5p promotes RCC progression through HuR-dependent IGF1R/AKT/PI3K pathway.
        Cell Death Discov. 2021; 7: 147
        • Han W.
        • Xu X.
        • Che K.
        • Ma G.
        • Li D.
        • Zhang M.
        • Jiao W.
        • Niu H.
        Establishment and Validation of a Prognostic Risk Model for Autophagy-Related Genes in Clear Cell Renal Cell Carcinoma.
        Dis Markers. 2020; 20208841859
        • Kimmelman A.C.
        • White E.
        Autophagy and Tumor Metabolism.
        Cell Metab. 2017; 25: 1037-1043
        • Ogata M.
        • Hino S.
        • Saito A.
        • Morikawa K.
        • Kondo S.
        • Kanemoto S.
        • Murakami T.
        • Taniguchi M.
        • Tanii I.
        • Yoshinaga K.
        • Shiosaka S.
        • Hammarback J.A.
        • Urano F.
        • Imaizumi K.
        Autophagy is activated for cell survival after endoplasmic reticulum stress.
        Mol Cell Biol. 2006; 26: 9220-9231
        • Zou M.
        • Lu N.
        • Hu C.
        • Liu W.
        • Sun Y.
        • Wang X.
        • You Q.
        • Gu C.
        • Xi T.
        • Guo Q.
        Beclin 1-mediated autophagy in hepatocellular carcinoma cells: implication in anticancer efficiency of oroxylin A via inhibition of mTOR signaling.
        Cell Signal. 2012; 24: 1722-1732
        • Hang P.
        • Zhao J.
        • Su Z.
        • Sun H.
        • Chen T.
        • Zhao L.
        • Du Z.
        Choline Inhibits Ischemia-Reperfusion-Induced Cardiomyocyte Autophagy in Rat Myocardium by Activating Akt/mTOR Signaling.
        Cell Physiol Biochem. 2018; 45: 2136-2144
        • Chen W.
        • Li X.
        • Guo S.
        • Song N.
        • Wang J.
        • Jia L.
        • Zhu A.
        Tanshinone IIA harmonizes the crosstalk of autophagy and polarization in macrophages via miR-375/KLF4 pathway to attenuate atherosclerosis.
        Int Immunopharmacol. 2019; 70: 486-497
        • Guo Y.
        • Sun W.
        • Gao W.
        • Li L.
        • Liang Y.
        • Mei Z.
        • et al.
        Long Noncoding RNA H19 Derived from M2 Tumor-Associated Macrophages Promotes Bladder Cell Autophagy via Stabilizing ULK1.
        J Oncol. 2022; : 3465459
        • Daurkin I.
        • Eruslanov E.
        • Stoffs T.
        • Perrin G.Q.
        • Algood C.
        • Gilbert S.M.
        • Rosser C.J.
        • Su L.M.
        • Vieweg J.
        • Kusmartsev S.
        Tumor-associated macrophages mediate immunosuppression in the renal cancer microenvironment by activating the 15-lipoxygenase-2 pathway.
        Cancer Res. 2011; 71: 6400-6409
        • Lim S.Y.
        • Yuzhalin A.E.
        • Gordon-Weeks A.N.
        • Muschel R.J.
        Targeting the CCL2-CCR2 signaling axis in cancer metastasis.
        Oncotarget. 2016; 7: 28697-28710
        • Li X.
        • Yao W.
        • Yuan Y.
        • Chen P.
        • Li B.
        • Li J.
        • Chu R.
        • Song H.
        • Xie D.
        • Jiang X.
        • Wang H.
        Targeting of tumour-infiltrating macrophages via CCL2/CCR2 signalling as a therapeutic strategy against hepatocellular carcinoma.
        Gut. 2017; 66: 157-167
        • Chen W.
        • Gao Q.
        • Han S.
        • Pan F.
        • Fan W.
        The CCL2/CCR2 axis enhances IL-6-induced epithelial-mesenchymal transition by cooperatively activating STAT3-Twist signaling.
        Tumour Biol. 2015; 36: 973-981
        • Liu H.
        • Sun Y.
        • O'Brien J.A.
        • Franco-Barraza J.
        • Qi X.
        • Yuan H.
        • Jin W.
        • Zhang J.
        • Gu C.
        • Zhao Z.
        • Yu C.
        • Feng S.
        • Yu X.
        Necroptotic astrocytes contribute to maintaining stemness of disseminated medulloblastoma through CCL2 secretion.
        Neuro Oncol. 2020; 22: 625-638
        • Izumi K.
        • Fang L.Y.
        • Mizokami A.
        • Namiki M.
        • Li L.
        • Lin W.J.
        • Chang C.
        Targeting the androgen receptor with siRNA promotes prostate cancer metastasis through enhanced macrophage recruitment via CCL2/CCR2-induced STAT3 activation.
        EMBO Mol Med. 2013; 5: 1383-1401
        • Robinson R.L.
        • Sharma A.
        • Bai S.
        • Heneidi S.
        • Lee T.J.
        • Kodeboyina S.K.
        • Patel N.
        • Sharma S.
        Comparative STAT3-Regulated Gene Expression Profile in Renal Cell Carcinoma Subtypes.
        Front Oncol. 2019; 9: 72
        • Shapiro I.M.
        • Cheng A.W.
        • Flytzanis N.C.
        • Balsamo M.
        • Condeelis J.S.
        • Oktay M.H.
        • Burge C.B.
        • Gertler F.B.
        An EMT-driven alternative splicing program occurs in human breast cancer and modulates cellular phenotype.
        PLoS Genet. 2011; 7e1002218
        • Cai J.
        • Wang N.
        • Lin G.
        • Zhang H.
        • Xie W.
        • Zhang Y.
        • et al.
        MBNL2 Regulates DNA Damage Response via Stabilizing p21.
        Int J Mol Sci. 2021; 22: 783-796
        • Qin X.
        • Lu A.
        • Ke M.
        • Zhu W.
        • Ye X.
        • Wang G.
        • Weng G.
        DJ-1 inhibits autophagy activity of prostate cancer cells by repressing JNK-Bcl2-Beclin1 signaling.
        Cell Biol Int. 2020; 44: 937-946
        • Zhou B.
        • Yang C.
        • Yan X.
        • Shi Z.
        • Xiao H.
        • Wei X.
        • Jiang N.
        • Wu Z.
        LETM1 Knockdown Promotes Autophagy and Apoptosis Through AMP-Activated Protein Kinase Phosphorylation-Mediated Beclin-1/Bcl-2 Complex Dissociation in Hepatocellular Carcinoma.
        Front Oncol. 2020; 10606790
        • Kang R.
        • Zeh H.J.
        • Lotze M.T.
        • Tang D.
        The Beclin 1 network regulates autophagy and apoptosis.
        Cell Death Differ. 2011; 18: 571-580