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SCIENCE CHINA Materials, Volume 62, Issue 6: 873-884(2019) https://doi.org/10.1007/s40843-018-9383-3

Stiffness heterogeneity-induced double-edged sword behaviors of carcinoma-associated fibroblasts in antitumor therapy

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  • ReceivedNov 7, 2018
  • AcceptedDec 13, 2018
  • PublishedJan 4, 2019

Abstract

Carcinoma-associated fibroblasts (CAFs) function as a double-edged sword in tumor progression. However, factors affecting the transition between tumor promotion and inhibition remain to be investigated. Here, we found that the transition was determined by stiffness heterogeneity of the tumor stroma in which tumor cells and CAFs were grown. When tumor cells were grown on a rigid plastic substrate, supernatants from CAFs inhibited the cytotoxic effects of 5-fluorouracil. In contrast, when tumor cells were grown on a soft substrate (5.3 kPa), supernatants from CAFs grown on a soft substrate increased the cytotoxicity of 5-fluorouracil. The diverse effects of CAFs were mediated by mechanotransduction factors, including stroma stiffness-induced cytokine expression in CAFs and signal transduction associated with stress fiber formation of CAFs. Moreover, we found that the cytokine expression in CAFs was regulated by nuclear Yes-associated protein, which changed according to cell stiffness, as characterized by atomic force microscopy. Overall, these findings suggested that modulating the mechanotransduction of the stroma together with CAFs might be important for increasing the efficacy of chemotherapy.


Funded by

the Postdoctoral Science Foundation Program of Chinese Academy of Medical Sciences & Peking Union Medical College

the National Natural Science Foundation of China(NSFC)

and National Institutes of Health / National Cancer Institute(NIH/NCI)

CA208196.


Acknowledgment

This work was financially supported by the Postdoctoral Science Foundation Program of Chinese Academy of Medical Sciences & Peking Union Medical College, the National Natural Science Foundation of China (NSFC) (31470905), and National Institutes of Health/National Cancer Institute (NIH/NCI) Grant R21, CA208196.


Interest statement

The authors declare no competing financial interest.


Contributions statement

Rao J, Han D, He J, and Liao F designed the experiments. Feng J, Li X, Rao E, and Zhang Q conducted experiments and analyzed the data. Feng J, Sharma S, Rao E, Han D and Rao J wrote the manuscript. All authors contributed to the general discussion.


Author information

Jiantao Feng was born in 1987. He received his PhD degree in chemistry from the Department of Chemistry, Tsinghua University in 2014. He is currently doing postdoctoral research with Prof. Jie He and Prof. Jianyu Rao at Cancer Hospital, Chinese Academy of Medical Sciences. His research focuses on the biomechanopharmacology in tumor microenvironment.


Jie He is currently a professor in Cancer Hospital, Chinese Academy of Medical Sciences. He got his PhD degree in Peking Union Medical College in 1993. His research interest focuses on the surgical diagnosis and treatment of lung cancer and esophageal cancer.


Dong Han is currently a professor in the National Center for Nanoscience and Technology (NCNST). He received his clinical MSc degree (1998) and MD/PhD (2001) from China Academy of Chinese Medical Sciences. His research interest focuses on nanobiomedical imaging, functional biointerfaces and translational medicine.


Jianyu Rao is a tenured full professor and vice chair of the Department of Pathology and Laboratory Medicine at the University of California at Los Angeles (UCLA). He received his MD degree from Shanghai Medical University in 1984. His research focuses on cancer biomarker studies. He is also interested in developing new technologies for pathology especially cytopathology, including imaging and digital analysis, telepathology, nanotechnology, liquid biopsy, and AI-technology.


Supplement

Supplementary information

Supplementary data associated with this article is available in the online version of the paper.


References

[1] Mitchell MJ, Jain RK, Langer R. Engineering and physical sciences in oncology: Challenges and opportunities. Nat Rev Cancer, 2017, 17: 659-675 CrossRef PubMed Google Scholar

[2] Liu L, Zhang SX, Liao W, et al. Mechanoresponsive stem cells to target cancer metastases through biophysical cues. Sci Transl Med, 2017, 9: eaan2966 CrossRef PubMed Google Scholar

[3] Plodinec M, Loparic M, Monnier CA, et al. The nanomechanical signature of breast cancer. Nat Nanotech, 2012, 7: 757-765 CrossRef PubMed ADS Google Scholar

[4] Acerbi I, Cassereau L, Dean I, et al. Human breast cancer invasion and aggression correlates with ECM stiffening and immune cell infiltration. Integr Biol, 2015, 7: 1120-1134 CrossRef PubMed Google Scholar

[5] Hu N, Cao Y, Ao Z, et al. Flow behavior of liquid metal in the connected fascial space: Intervaginal space injection in the rat wrist and mice with tumor. Nano Res, 2017, 11: 2265-2276 CrossRef Google Scholar

[6] Kalluri R. The biology and function of fibroblasts in cancer. Nat Rev Cancer, 2016, 16: 582-598 CrossRef PubMed Google Scholar

[7] Kharaishvili G, Simkova D, Bouchalova K, et al. The role of cancer-associated fibroblasts, solid stress and other microenvironmental factors in tumor progression and therapy resistance. Cancer Cell Int, 2014, 14: 41 CrossRef PubMed Google Scholar

[8] Kalli M, Papageorgis P, Gkretsi V, et al. Solid stress facilitates fibroblasts activation to promote pancreatic cancer cell migration. Ann Biomed Eng, 2018, 46: 657-669 CrossRef PubMed Google Scholar

[9] Levental KR, Yu H, Kass L, et al. Matrix crosslinking forces tumor progression by enhancing integrin signaling. Cell, 2009, 139: 891-906 CrossRef PubMed Google Scholar

[10] Yaqoob U, Cao S, Shergill U, et al. Neuropilin-1 stimulates tumor growth by increasing fibronectin fibril assembly in the tumor microenvironment. Cancer Res, 2012, 72: 4047-4059 CrossRef PubMed Google Scholar

[11] Schrader J, Gordon-Walker TT, Aucott RL, et al. Matrix stiffness modulates proliferation, chemotherapeutic response, and dormancy in hepatocellular carcinoma cells. Hepatology, 2011, 53: 1192-1205 CrossRef PubMed Google Scholar

[12] Ulrich TA, de Juan Pardo EM, Kumar S. The mechanical rigidity of the extracellular matrix regulates the structure, motility, and proliferation of glioma cells. Cancer Res, 2009, 69: 4167-4174 CrossRef PubMed Google Scholar

[13] Nune KC, Li S, Misra RDK. Advancements in three-dimensional titanium alloy mesh scaffolds fabricated by electron beam melting for biomedical devices: Mechanical and biological aspects. Sci China Mater, 2018, 61: 455-474 CrossRef Google Scholar

[14] Feng J, Tang Y, Xu Y, et al. Substrate stiffness influences the outcome of antitumor drug screening in vitro. Clin Hemorheol Microcirc, 2013, 55: 121–131. Google Scholar

[15] Liu C, Li X, Hua W, et al. Porous matrix stiffness modulates response to targeted therapy in breast carcinoma. Small, 2016, 12: 4675-4681 CrossRef PubMed Google Scholar

[16] Sun Y. Tumor microenvironment and cancer therapy resistance. Cancer Lett, 2016, 380: 205-215 CrossRef PubMed Google Scholar

[17] Krishnan V, Schaar B, Tallapragada S, et al. Tumor associated macrophages in gynecologic cancers. Gynecol Oncol, 2018, 149: 205-213 CrossRef PubMed Google Scholar

[18] Chen J, Lin L, Yan N, et al. Macrophages loaded CpG and GNR-PEI for combination of tumor photothermal therapy and immunotherapy. Sci China Mater, 2018, 61: 1484-1494 CrossRef Google Scholar

[19] Yamamura Y, Asai N, Enomoto A, et al. Akt-Girdin signaling in cancer-associated fibroblasts contributes to tumor progression. Cancer Res, 2015, 75: 813-823 CrossRef PubMed Google Scholar

[20] Öhlund D, Elyada E, Tuveson D. Fibroblast heterogeneity in the cancer wound. J Exp Med, 2014, 211: 1503-1523 CrossRef PubMed Google Scholar

[21] Özdemir BC, Pentcheva-Hoang T, Carstens JL, et al. Depletion of carcinoma-associated fibroblasts and fibrosis induces immunosuppression and accelerates pancreas cancer with reduced survival. Cancer Cell, 2014, 25: 719-734 CrossRef PubMed Google Scholar

[22] Geng L, Feng J, Sun Q, et al. Nanomechanical clues from morphologically normal cervical squamous cells could improve cervical cancer screening. Nanoscale, 2015, 7: 15589-15593 CrossRef PubMed ADS Google Scholar

[23] Butt HJ, Cappella B, Kappl M. Force measurements with the atomic force microscope: Technique, interpretation and applications. Surf Sci Rep, 2005, 59: 1-152 CrossRef ADS Google Scholar

[24] Paszek MJ, Zahir N, Johnson KR, et al. Tensional homeostasis and the malignant phenotype. Cancer Cell, 2005, 8: 241-254 CrossRef PubMed Google Scholar

[25] Farmer P, Bonnefoi H, Anderle P, et al. A stroma-related gene signature predicts resistance to neoadjuvant chemotherapy in breast cancer. Nat Med, 2009, 15: 68-74 CrossRef PubMed Google Scholar

[26] Dupont S, Morsut L, Aragona M, et al. Role of YAP/TAZ in mechanotransduction. Nature, 2011, 474: 179-183 CrossRef PubMed Google Scholar

[27] Sharif GM, Schmidt MO, Yi C, et al. Cell growth density modulates cancer cell vascular invasion via hippo pathway activity and CXCR2 signaling. Oncogene, 2015, 34: 5879-5889 CrossRef PubMed Google Scholar

[28] Wang G, Lu X, Dey P, et al. Targeting YAP-dependent MDSC infiltration impairs tumor progression. Cancer Discov, 2016, 6: 80-95 CrossRef PubMed Google Scholar

[29] Calvo F, Ege N, Grande-Garcia A, et al. Mechanotransduction and YAP-dependent matrix remodelling is required for the generation and maintenance of cancer-associated fibroblasts. Nat Cell Biol, 2013, 15: 637-646 CrossRef PubMed Google Scholar

[30] Cole SW, Nagaraja AS, Lutgendorf SK, et al. Sympathetic nervous system regulation of the tumour microenvironment. Nat Rev Cancer, 2015, 15: 563-572 CrossRef PubMed Google Scholar

[31] Quail DF, Joyce JA. Microenvironmental regulation of tumor progression and metastasis. Nat Med, 2013, 19: 1423-1437 CrossRef PubMed Google Scholar

[32] De Palma M, Biziato D, Petrova TV. Microenvironmental regulation of tumour angiogenesis. Nat Rev Cancer, 2017, 17: 457-474 CrossRef PubMed Google Scholar

[33] Straussman R, Morikawa T, Shee K, et al. Tumour micro-environment elicits innate resistance to RAF inhibitors through HGF secretion. Nature, 2012, 487: 500-504 CrossRef PubMed ADS Google Scholar

[34] Wilson TR, Fridlyand J, Yan Y, et al. Widespread potential for growth-factor-driven resistance to anticancer kinase inhibitors. Nature, 2012, 487: 505-509 CrossRef PubMed ADS Google Scholar

[35] Holohan C, Van Schaeybroeck S, Longley DB, et al. Cancer drug resistance: An evolving paradigm. Nat Rev Cancer, 2013, 13: 714-726 CrossRef PubMed Google Scholar

[36] Wu Y, Wang D, Li Y. Understanding of the major reactions in solution synthesis of functional nanomaterials. Sci China Mater, 2016, 59: 938-996 CrossRef Google Scholar

[37] Xu X, Lu Y, Yang Y, et al. Tuning the growth of metal-organic framework nanocrystals by using polyoxometalates as coordination modulators. Sci China Mater, 2015, 58: 370-377 CrossRef Google Scholar

[38] Paraiso KHT, Smalley KSM. Fibroblast-mediated drug resistance in cancer. Biochem Pharmacol, 2013, 85: 1033-1041 CrossRef PubMed Google Scholar

[39] Brunen D, Willems SM, Kellner U, et al. TGF-β: An emerging player in drug resistance. Cell Cycle, 2013, 12: 2960-2968 CrossRef PubMed Google Scholar

[40] Leight JL, Wozniak MA, Chen S, et al. Matrix rigidity regulates a switch between TGF-β1–induced apoptosis and epithelial–mesenchymal transition. Mol Bio Cell, 2012, 23: 781-791 CrossRef PubMed Google Scholar

[41] Liu Y, He K, Hu Y, et al. YAP modulates TGF-β1-induced simultaneous apoptosis and EMT through upregulation of the EGF receptor. Sci Rep, 2017, 7: 45523 CrossRef PubMed ADS Google Scholar

[42] Kawamura M, Toiyama Y, Tanaka K, et al. CXCL5, a promoter of cell proliferation, migration and invasion, is a novel serum prognostic marker in patients with colorectal cancer. Eur J Cancer, 2012, 48: 2244-2251 CrossRef PubMed Google Scholar

[43] Wang B, Hendricks DT, Wamunyokoli F, et al. A growth-related oncogene/CXC chemokine receptor 2 autocrine loop contributes to cellular proliferation in esophageal cancer. Cancer Res, 2006, 66: 3071-3077 CrossRef PubMed Google Scholar

[44] Low BC, Pan CQ, Shivashankar GV, et al. YAP/TAZ as mechanosensors and mechanotransducers in regulating organ size and tumor growth. FEBS Lett, 2014, 588: 2663-2670 CrossRef PubMed Google Scholar

[45] Liu F, Lagares D, Choi KM, et al. Mechanosignaling through YAP and TAZ drives fibroblast activation and fibrosis. Am J Physiol-Lung Cellular Mol Physiol, 2015, 308: L344-L357 CrossRef PubMed Google Scholar

[46] Oria R, Wiegand T, Escribano J, et al. Force loading explains spatial sensing of ligands by cells. Nature, 2017, 196: 219-224 CrossRef PubMed ADS Google Scholar

[47] Zhao B, Ye X, Yu J, et al. Tead mediates YAP-dependent gene induction and growth control. Genes Dev, 2008, 22: 1962-1971 CrossRef PubMed Google Scholar

[48] Hasebe T. Tumor–stromal interactions in breast tumor progression–significance of histological heterogeneity of tumor–stromal fibroblasts. Expert Opin Therap Targets, 2013, 17: 449-460 CrossRef PubMed Google Scholar

[49] Sharma S, Santiskulvong C, Bentolila LA, et al. Correlative nanomechanical profiling with super-resolution F-actin imaging reveals novel insights into mechanisms of cisplatin resistance in ovarian cancer cells. Nanomed-Nanotechnol Biol Med, 2012, 8: 757-766 CrossRef PubMed Google Scholar

[50] Cross SE, Jin YS, Rao J, et al. Nanomechanical analysis of cells from cancer patients. Nat Nanotech, 2007, 2: 780-783 CrossRef PubMed ADS Google Scholar

[51] Adams JL, Smothers J, Srinivasan R, et al. Big opportunities for small molecules in immuno-oncology. Nat Rev Drug Discov, 2015, 14: 603-622 CrossRef PubMed Google Scholar

[52] Schmid D, Park CG, Hartl CA, et al. T cell-targeting nanoparticles focus delivery of immunotherapy to improve antitumor immunity. Nat Commun, 2017, 8: 1747 CrossRef PubMed ADS Google Scholar

[53] Tu Y. The discovery of artemisinin (qinghaosu) and gifts from Chinese medicine. Nat Med, 2011, 17: 1217-1220 CrossRef PubMed Google Scholar

[54] Liao F, Li M, Han D, et al. Biomechanopharmacology: A new borderline discipline. Trends Pharmacol Sci, 2006, 27: 287-289 CrossRef PubMed Google Scholar

  • Figure 1

    Proliferation and survival of SK-BR-3 cells in response to PCM from CCD-1095Sk cells (CAFs). (a) Fluorescent imaging of fibroblast-activating protein (FAP) expressed in CCD-1095Sk cells. (b) Fluorescent imaging of α-SMA expressed in CCD-1095Sk cells. (c) Proliferation of SK-BR-3 cells on rigid substrate (plastic) in response to PCM prepared on different substrates. (d) Proliferation of SK-BR-3 cells on soft substrate (5.3 kPa) in response to PCM prepared on different substrates. (e) SK-BR-3 cell survival rates in response to PCM prepared on different substrates at different 5-fluorouracil (5-FU) concentrations when grown on rigid substrate (plastic). (f) SK-BR-3 survival rates in response to PCM prepared on different substrates at different 5-FU concentrations when grown on soft substrate (5.3 kPa). Statistical analysis was performed using one-way ANOVA with Student-Newman-Keuls Test. *p<0.05, **p<0.01.

  • Figure 2

    Expression analysis of cytokines, Yes-associated protein (YAP), and stress fibers in CCD-1095Sk cells (CAFs) grown on substrates with stiffnesses of 5.3 and 46.7 kPa. (a) Heat map of cytokine secretion from CCD-1095Sk cells cultured on 5.3 and 46.7 kPa. (b) Relative expression of GRO in PCM/5.3 kPa and PCM/46.7 kPa groups. (c) Relative expression of epithelial-derived neutrophil-activating peptide 78 (ENA-78) in PCM/5.3 kPa and PCM/46.7 kPa groups. (d) Fluorescent characterization of YAP and stress fibers in CCD-1095Sk cells grown on substrates with stiffnesses of 5.3 and 46.7 kPa. (e) Quantification of YAP nuclear distribution in CCD-1095Sk cells with nuclear/cytoplasmic values grown on substrates with stiffnesses of 5.3 and 46.7 kPa. (f) Stiffness characterization of CCD-1095Sk cells grown on substrates with stiffnesses of 5.3 and 46.7 kPa employing atomic force microscopy. For (b) and (c), statistical analysis was performed using Student’s t-tests. For (e) and (f), statistical analysis was performed using Kruskal-Wallis tests with the NPAR1WAY procedure in SAS. *p<0.05.

  • Figure 3

    Influence of stress fiber inhibitors on cytokine secretion from CCD-1095Sk cells (CAFs) and survival of SK-BR-3 cells. (a) Heat map of cytokine secretion from CCD-1095Sk cells before and after blebbistatin (blebbist.) inhibition. (b) List of cytokines inhibited by more than 2-fold in (a). (c) Cancer cell survival rates in cells grown in PCM from CCD-1095Sk cells prepared on substrates with stiffnesses of 5.3 and 46.7 kPa after treatment with blebbist. (d) Cancer cell survival rates in cells grown in PCM from CCD-1095Sk cells prepared on substrates with stiffnesses of 5.3 and 46.7 kPa after treatment with Y27632. Statistical analysis was performed using pooled two sample t-tests. n.s., not significant.

  • Figure 4

    Influence of stress fiber inhibitors on YAP nuclear localization and stress fiber formation in CCD-1095Sk cells (CAFs) grown on substrates with stiffnesses of 5.3 and 46.7 kPa. (a) Fluorescent imaging of YAP distribution in CCD-1095Sk cells after inhibition with blebbistatin or Y27632 for cells grown on substrates with stiffnesses of 5.3 and 46.7 kPa. (b) Fluorescent imaging of stress fibers in CCD-1095Sk cells after inhibition with blebbistatin or Y27632 for cells grown on substrates with stiffnesses of 5.3 and 46.7 kPa. (c) Quantification of YAP distribution in CCD-1095Sk cells grown on substrates with stiffnesses of 5.3 and 46.7 kPa after inhibition with blebbistatin (i) or Y27632 (ii); stiffness characterization of CCD-1095Sk cells grown on substrates with stiffnesses of 5.3 and 46.7 kPa after inhibition with blebbistatin (iii) or Y27632 (iv). Statistical analysis was performed using Kruskal-Wallis tests with the NPAR1WAY procedure in SAS. n.s., not significant.

  • Figure 5

    Illustration of the mechanism of the double-edged sword effect of CAFs on tumor drug responses. CAFs sense the stiffness of the extracellular matrix by mechanotransduction of the integrin complex, which delivers the mechanical signal to stress fibers. Stress fibers transmit extracellular mechanical signals and mediate YAP nuclear translocation, which then activates the secretion of cytokines from CAFs. When tumor cells were grown on the rigid substrate, TGF-β promoted tumor survival. When tumor cells and CAFs were both grown on the soft substrate (5.3 kPa), nuclear YAP inhibited cancer cell growth by upregulation of GRO and ENA-78 expression in CAFs. The process was suppressed by blebbistatin or Y27632, which disrupted the structures of stress fibers.

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