Skip to main content

Advertisement

SpringerLink
Log in

Single-cell RNA sequencing reveals the cellular and molecular characteristics of high-grade and metastatic bladder cancer

  • Research
  • Published:
Cellular Oncology Aims and scope Submit manuscript

Abstract

Purpose

Metastatic bladder cancer (BC) has the highest somatic mutation frequency and recurrence rate of all tumors. However, the cellular and molecular characteristics of BC remain unclear.

Methods

We performed single-cell RNA sequencing (scRNA-seq) on the samples of paracancerous normal tissue (PNT), primary tumor (PT) and lymph node metastasis (LNM). The proportions and gene expression profiles of different cell types in the tumor microenvironment (TME) were investigated.

Results

In total, 50,158 cells were classified into six populations. Malignant cells of PT and LNM exhibited large mutant DNA fragments, while the cell phenotypes and gene expression profiles differed during differentiation. Metastasis was associated with a poorer prognosis than PT. Tumor-associated stromal cells and inhibitory immune cells were the main cell populations in PT and LNM. Cell-cell communication analysis revealed the roles of signaling pathways of inflammatory cancer-associated fibroblast (iCAF) and tumor-associated macrophage (TAM) in exhaustion of T cells. In addition, iCAF may recruit TAM to promote formation of the TME earlier than the differentiation of tumor cells.

Conclusion

This study through scRNA-seq enhanced our understanding of new features about the cellular and molecular similarities and differences of high-grade and metastatic bladder cancer, which might provide potential therapeutic targets in future treatment.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Data availability

All data generated or analyzed during this study are included in this published article.

References

  1. H. Sung, J. Ferlay, R.L. Siegel, M. Laversanne, I. Soerjomataram, A. Jemal, F. Bray, Global Cancer Statistics 2020: GLOBOCAN estimates of incidence and Mortality Worldwide for 36 cancers in 185 countries. CA Cancer J Clin 71(3), 209–249 (2021)

    Article  PubMed  Google Scholar 

  2. Comprehensive molecular characterization, Of urothelial bladder carcinoma. Nature 507(7492), 315–322 (2014)

    Article  Google Scholar 

  3. E.J. Pietzak, A. Bagrodia, E.K. Cha, E.N. Drill, G. Iyer, S. Isharwal, I. Ostrovnaya, P. Baez, Q. Li, M.F. Berger, A. Zehir, N. Schultz, J.E. Rosenberg, D.F. Bajorin, G. Dalbagni, H. Al-Ahmadie, D.B. Solit, B.H. Bochner, Next-generation sequencing of nonmuscle invasive bladder Cancer reveals potential biomarkers and rational therapeutic targets. Eur. Urol. 72(6), 952–959 (2017)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. L.B. Alexandrov, S. Nik-Zainal, D.C. Wedge, S.A. Aparicio, S. Behjati, A.V. Biankin, G.R. Bignell, N. Bolli, A. Borg, A.L. Børresen-Dale, S. Boyault, B. Burkhardt, A.P. Butler, C. Caldas, H.R. Davies, C. Desmedt, R. Eils, J.E. Eyfjörd, J.A. Foekens, M. Greaves, F. Hosoda, B. Hutter, T. Ilicic, S. Imbeaud, M. Imielinski, N. Jäger, D.T. Jones, D. Jones, S. Knappskog, M. Kool, S.R. Lakhani, C. López-Otín, S. Martin, N.C. Munshi, H. Nakamura, P.A. Northcott, M. Pajic, E. Papaemmanuil, A. Paradiso, J.V. Pearson, X.S. Puente, K. Raine, M. Ramakrishna, A.L. Richardson, J. Richter, P. Rosenstiel, M. Schlesner, T.N. Schumacher, P.N. Span, J.W. Teague, Y. Totoki, A.N. Tutt, R. Valdés-Mas, M.M. van Buuren, L. van ‘t Veer, A. Vincent-Salomon, N. Waddell, L.R. Yates, J. Zucman-Rossi, P.A. Futreal, U. McDermott, P. Lichter, M. Meyerson, S.M. Grimmond, R. Siebert, E. Campo, T. Shibata, S.M. Pfister, P.J. Campbell, M.R. Stratton, Signatures of mutational processes in human cancer, Nature 500(7463), 415–421 (2013)

  5. L.M. Merlo, J.W. Pepper, B.J. Reid, C.C. Maley, Cancer as an evolutionary and ecological process. Nat. Rev. Cancer 6(12), 924–935 (2006)

    Article  CAS  PubMed  Google Scholar 

  6. T. Okamoto, D. duVerle, K. Yaginuma, Y. Natsume, H. Yamanaka, D. Kusama, M. Fukuda, M. Yamamoto, F. Perraudeau, U. Srivastava, Y. Kashima, A. Suzuki, Y. Kuze, Y. Takahashi, M. Ueno, Y. Sakai, T. Noda, K. Tsuda, Y. Suzuki, S. Nagayama, R. Yao, Comparative analysis of patient-matched PDOs revealed a reduction in OLFM4-Associated clusters in metastatic lesions in Colorectal Cancer. Stem Cell. Reports 16(4), 954–967 (2021)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. J.A. Witjes, H.M. Bruins, R. Cathomas, E.M. Compérat, N.C. Cowan, G. Gakis, V. Hernández, E. Linares Espinós, A. Lorch, Y. Neuzillet, M. Rouanne, G.N. Thalmann, E. Veskimäe, M.J. Ribal, A.G. van der Heijden, European Association of Urology Guidelines on muscle-invasive and metastatic bladder Cancer: Summary of the 2020 guidelines. Eur. Urol. 79(1), 82–104 (2021)

    Article  CAS  PubMed  Google Scholar 

  8. V.G. Patel, W.K. Oh, M.D. Galsky, Treatment of muscle-invasive and advanced bladder cancer in 2020. CA Cancer J Clin 70(5), 404–423 (2020)

    Article  PubMed  Google Scholar 

  9. Y. Zheng, X. Yang, Spatial RNA sequencing methods show high resolution of single cell in cancer metastasis and the formation of tumor microenvironment, Biosci Rep 43(2) (2023)

  10. J.J. Quinn, M.G. Jones, R.A. Okimoto, S. Nanjo, M.M. Chan, N. Yosef, T.G. Bivona, J.S. Weissman, Single-cell lineages reveal the rates, routes, and drivers of metastasis in cancer xenografts, Science 371(6532) (2021)

  11. Y. Chi, J. Remsik, V. Kiseliovas, C. Derderian, U. Sener, M. Alghader, F. Saadeh, K. Nikishina, T. Bale, C. Iacobuzio-Donahue, T. Thomas, D. Pe’er, L. Mazutis, A. Boire, Cancer cells deploy lipocalin-2 to collect limiting iron in leptomeningeal metastasis. Science 369(6501), 276–282 (2020)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. A. Dobin, C.A. Davis, F. Schlesinger, J. Drenkow, C. Zaleski, S. Jha, P. Batut, M. Chaisson, T.R. Gingeras, STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29(1), 15–21 (2013)

    Article  CAS  PubMed  Google Scholar 

  13. A. Butler, P. Hoffman, P. Smibert, E. Papalexi, R. Satija, Integrating single-cell transcriptomic data across different conditions, technologies, and species. Nat. Biotechnol. 36(5), 411–420 (2018)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. C.S. McGinnis, L.M. Murrow, Z.J. Gartner, DoubletFinder: Doublet Detection in single-cell RNA sequencing data using Artificial Nearest neighbors. Cell. Syst. 8(4), 329–337.e4 (2019)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. E.Z. Macosko, A. Basu, R. Satija, J. Nemesh, K. Shekhar, M. Goldman, I. Tirosh, A.R. Bialas, N. Kamitaki, E.M. Martersteck, J.J. Trombetta, D.A. Weitz, J.R. Sanes, A.K. Shalek, A. Regev, S.A. McCarroll, Highly parallel genome-wide expression profiling of individual cells using Nanoliter Droplets. Cell 161(5), 1202–1214 (2015)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. D. Aran, A.P. Looney, L. Liu, E. Wu, V. Fong, A. Hsu, S. Chak, R.P. Naikawadi, P.J. Wolters, A.R. Abate, A.J. Butte, M. Bhattacharya, Reference-based analysis of lung single-cell sequencing reveals a transitional profibrotic macrophage. Nat. Immunol. 20(2), 163–172 (2019)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. S.V. Puram, I. Tirosh, A.S. Parikh, A.P. Patel, K. Yizhak, S. Gillespie, C. Rodman, C.L. Luo, E.A. Mroz, K.S. Emerick, D.G. Deschler, M.A. Varvares, R. Mylvaganam, O. Rozenblatt-Rosen, J.W. Rocco, W.C. Faquin, D.T. Lin, A. Regev, B.E. Bernstein, single-cell transcriptomic analysis of primary and metastatic Tumor Ecosystems in Head and Neck Cancer. Cell 171(7), 1611–1624.e24 (2017)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. C. Trapnell, D. Cacchiarelli, J. Grimsby, P. Pokharel, S. Li, M. Morse, N.J. Lennon, K.J. Livak, T.S. Mikkelsen, J.L. Rinn, The dynamics and regulators of cell fate decisions are revealed by pseudotemporal ordering of single cells. Nat. Biotechnol. 32(4), 381–386 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. S. Hänzelmann, R. Castelo, J. Guinney, GSVA: gene set variation analysis for microarray and RNA-seq data. BMC Bioinform 14, 7 (2013)

    Article  Google Scholar 

  20. S. Jin, C.F. Guerrero-Juarez, L. Zhang, I. Chang, R. Ramos, C.H. Kuan, P. Myung, M.V. Plikus, Q. Nie, Inference and analysis of cell-cell communication using CellChat. Nat. Commun. 12(1), 1088 (2021)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. P.M. Moonen, G.F. Merkx, P. Peelen, H.F. Karthaus, D.F. Smeets, J.A. Witjes, UroVysion compared with cytology and quantitative cytology in the surveillance of non-muscle-invasive bladder cancer. Eur. Urol. 51(5), 1275–1280 (2007); discussion 1280

    Article  PubMed  Google Scholar 

  22. X. Diao, J. Cai, J. Zheng, J. Kong, S. Wu, H. Yu, H. Huang, W. Xie, X. Chen, C. Huang, L. Huang, H. Qin, J. Huang, T. Lin, Association of chromosome 7 aneuploidy measured by fluorescence in situ hybridization assay with muscular invasion in bladder cancer. Cancer Commun. (Lond) 40(4), 167–180 (2020)

    Article  PubMed  PubMed Central  Google Scholar 

  23. J. Bellmunt, Stem-Like Signature Predicting Disease Progression in Early Stage Bladder Cancer. The Role of E2F3 and SOX4, Biomedicines 6(3) (2018)

  24. M. Riester, L. Werner, J. Bellmunt, S. Selvarajah, E.A. Guancial, B.A. Weir, E.C. Stack, R.S. Park, R. O’Brien, F.A. Schutz, T.K. Choueiri, S. Signoretti, J. Lloreta, L. Marchionni, E. Gallardo, F. Rojo, D.I. Garcia, Y. Chekaluk, D.J. Kwiatkowski, B.H. Bochner, W.C. Hahn, A.H. Ligon, J.A. Barletta, M. Loda, D.M. Berman, P.W. Kantoff, F. Michor, J.E. Rosenberg, Integrative analysis of 1q23.3 copy-number gain in metastatic urothelial carcinoma. Clin. Cancer Res. 20(7), 1873–1883 (2014)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. A. Veerakumarasivam, H.E. Scott, S.F. Chin, A. Warren, M.J. Wallard, D. Grimmer, K. Ichimura, C. Caldas, V.P. Collins, D.E. Neal, J.D. Kelly, High-resolution array-based comparative genomic hybridization of bladder cancers identifies mouse double minute 4 (MDM4) as an amplification target exclusive of MDM2 and TP53. Clin. Cancer Res. 14(9), 2527–2534 (2008)

    Article  CAS  PubMed  Google Scholar 

  26. X. Li, M. Xu, L. Ding, J. Tang, MiR-27a: a novel biomarker and potential therapeutic target in tumors. J. Cancer 10(12), 2836–2848 (2019)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. L. Haghverdi, M. Büttner, F.A. Wolf, F. Buettner, F.J. Theis, Diffusion pseudotime robustly reconstructs lineage branching. Nat. Methods 13(10), 845–848 (2016)

    Article  CAS  PubMed  Google Scholar 

  28. E. Sahai, I. Astsaturov, E. Cukierman, D.G. DeNardo, M. Egeblad, R.M. Evans, D. Fearon, F.R. Greten, S.R. Hingorani, T. Hunter, R.O. Hynes, R.K. Jain, T. Janowitz, C. Jorgensen, A.C. Kimmelman, M.G. Kolonin, R.G. Maki, R.S. Powers, E. Puré, D.C. Ramirez, R. Scherz-Shouval, M.H. Sherman, S. Stewart, T.D. Tlsty, D.A. Tuveson, F.M. Watt, V. Weaver, A.T. Weeraratna, Z. Werb, A framework for advancing our understanding of cancer-associated fibroblasts. Nat. Rev. Cancer 20(3), 174–186 (2020)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. T. Zhan, N. Rindtorff, M. Boutros, Wnt signaling in cancer. Oncogene 36(11), 1461–1473 (2017)

    Article  CAS  PubMed  Google Scholar 

  30. K.D. Howarth, T. Mirza, S.L. Cooke, S.F. Chin, J.C. Pole, E. Turro, M.D. Eldridge, R.M. Garcia, O.M. Rueda, C. Boursnell, J.E. Abraham, C. Caldas, P.A.W. Edwards, NRG1 fusions in breast cancer. Breast Cancer Res 23(1), 3 (2021)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. D.H. Shin, J.Y. Jo, J.Y. Han, Dual targeting of ERBB2/ERBB3 for the treatment of SLC3A2-NRG1-Mediated Lung Cancer. Mol. Cancer Ther. 17(9), 2024–2033 (2018)

    Article  CAS  PubMed  Google Scholar 

  32. C.D. Mills, K. Kincaid, J.M. Alt, M.J. Heilman, A.M. Hill, M-1/M-2 macrophages and the Th1/Th2 paradigm. J. Immunol. 164(12), 6166–6173 (2000)

    Article  CAS  PubMed  Google Scholar 

  33. Y. Pan, Y. Yu, X. Wang, T. Zhang, Tumor-Associated Macrophages in Tumor Immunity. Front. Immunol. 11, 583084 (2020)

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. C. Yunna, H. Mengru, W. Lei, C. Weidong, Macrophage M1/M2 polarization. Eur. J. Pharmacol. 877, 173090 (2020)

    Article  PubMed  Google Scholar 

  35. N. Zhang, S.H. Kim, A. Gainullina, E.C. Erlich, E.J. Onufer, J. Kim, R.S. Czepielewski, B.A. Helmink, J.R. Dominguez, B.T. Saunders, J. Ding, J.W. Williams, J.X. Jiang, B.H. Segal, B.H. Zinselmeyer, G.J. Randolph, K.W. Kim, LYVE1 + macrophages of murine peritoneal mesothelium promote omentum-independent ovarian tumor growth, J Exp Med 218(12) (2021)

  36. M.B. Buechler, W. Fu, S.J. Turley, Fibroblast-macrophage reciprocal interactions in health, fibrosis, and cancer. Immunity 54(5), 903–915 (2021)

    Article  CAS  PubMed  Google Scholar 

  37. J.E. Cheong, L. Sun, Targeting the IDO1/TDO2-KYN-AhR pathway for Cancer Immunotherapy - Challenges and Opportunities. Trends Pharmacol. Sci. 39(3), 307–325 (2018)

    Article  CAS  PubMed  Google Scholar 

  38. L.M. McLane, M.S. Abdel-Hakeem, E.J. Wherry, CD8 T cell exhaustion during chronic viral infection and Cancer. Annu. Rev. Immunol. 37, 457–495 (2019)

    Article  CAS  PubMed  Google Scholar 

  39. D. Masopust, V. Vezys, A.L. Marzo, L. Lefrançois, Preferential localization of effector memory cells in nonlymphoid tissue. Science 291(5512), 2413–2417 (2001)

    Article  CAS  PubMed  Google Scholar 

  40. S. Olalekan, B. Xie, R. Back, H. Eckart, A. Basu, Characterizing the tumor microenvironment of metastatic ovarian cancer by single-cell transcriptomics. Cell. Rep. 35(8), 109165 (2021)

    Article  CAS  PubMed  Google Scholar 

  41. S.K. Daniel, Y.D. Seo, V.G. Pillarisetty, The CXCL12-CXCR4/CXCR7 axis as a mechanism of immune resistance in gastrointestinal malignancies. Semin Cancer Biol 65, 176–188 (2020)

    Article  CAS  PubMed  Google Scholar 

  42. X. Zhang, Y. Dong, M. Zhao, L. Ding, X. Yang, Y. Jing, Y. Song, S. Chen, Q. Hu, Y. Ni, ITGB2-mediated metabolic switch in CAFs promotes OSCC proliferation by oxidation of NADH in mitochondrial oxidative phosphorylation system. Theranostics 10(26), 12044–12059 (2020)

    Article  CAS  PubMed  Google Scholar 

  43. M. Marchesi, E. Andersson, L. Villabona, B. Seliger, A. Lundqvist, R. Kiessling, G.V. Masucci, HLA-dependent tumour development: a role for tumour associate macrophages? J. Transl Med. 11, 247 (2013)

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We acknowledge the OE Biotech Co. Ltd (Shanghai, China) for providing single-cell RNA sequencing technology. And the support Funding: including the Central Guidance on Local Science and Technology Development Fund of Shanxi Province (No. YDZJSX2021C010); Nature Science Foundation of Shanxi Province (No.202103021224412); Fund Program for the Scientific Activities of Selected Returned Overseas Professionals in Shanxi Province (No.20210005); Shanxi Provincial Basic Applied Research Project (No.20210302124611).

Funding

Nature Science Foundation of Shanxi Province (No.202,103,021,224,412). The Central Guidance on Local Science and Technology Development Fund of Shanxi Province (No.YDZJSX2021C010). Fund Program for the Scientific Activities of Selected Returned Overseas Professionals in Shanxi Province (No.20,210,005). Shanxi Provincial Basic Applied Research Project (No.20,210,302,124,611).

Author information

Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Basic Medical College, Shanxi Medical University, No. 56, Xinjiang South Road, Yingze street, Yingze District, Taiyuan City, 030000, Shanxi Province, China

    Yue Zheng

  2. Department of Urology, First Hospital of Shanxi Medical University, No. 85, Jiefang South Road, Yingze street, Yingze District, Taiyuan City, 030000, Shanxi Province, China

    Xin Wang & Xiaofeng Yang

  3. Department of Urology and State Key Laboratory of Molecular Oncology, National Cancer Center, National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing City, 100021, China

    Nianzeng Xing

  4. Department of Urology, Shanxi Hospital Affiliated to Cancer Hospital, Cancer Hospital, Shanxi Province Cancer Hospital, Chinese Academy of Medical Sciences, Shanxi Medical University, Taiyuan, 030013, Shanxi Province, China

    Nianzeng Xing

Authors
  1. Yue Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Xin Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Xiaofeng Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Nianzeng Xing
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Material preparation, data collection and analysis were performed by Yue Zheng. The first draft of the manuscript was written by Yue Zheng and Xin Wang. Yue Zheng was the major contributor in designing and writing this manuscript. Xin Wang was responsible for writing the experimental part. Funding acquisition, reviewing and editing were performed by Xiaofeng Yang. All authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Xiaofeng Yang.

Ethics declarations

Competing interests

The authors declare that they have no competing interests.

Ethics approval and consent to participate

The research involving human samples have been performed in accordance with the Declaration of Helsinki. This study was approved by the Ethics Committee of the First Hospital of Shanxi Medical University (K093). Informed consent was obtained from the patients.

Consent for publication

Not applicable.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zheng, Y., Wang, X., Yang, X. et al. Single-cell RNA sequencing reveals the cellular and molecular characteristics of high-grade and metastatic bladder cancer. Cell Oncol. 46, 1415–1427 (2023). https://doi.org/10.1007/s13402-023-00820-x

Download citation

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s13402-023-00820-x

Keywords

Search

Navigation