Abstract
The pressure towards innovation and creation of new model systems in regenerative medicine and cancer research has fostered the development of novel potential therapeutic applications. Kidney injuries provoke a high request of organ transplants making it the most demanding system in the field of regenerative medicine. Furthermore, renal cancer frequently threaten patients’ life and aggressive forms still remain difficult to treat. Ethical issues related to the use of embryonic stem cells, has fueled research on adult, patient-specific pluripotent stem cells as a model for discovery and therapeutic development, but to date, normal and cancerous renal experimental models are lacking. Several research groups are focusing on the development of organoid cultures. Since organoids mimic the original tissue architecture in vitro, they represent an excellent model for tissue engineering studies and cancer therapy testing. We established normal and tumor renal cell carcinoma organoids previously maintained in a heterogeneous multi-clone stem cell-like enriching medium. Starting from adult normal kidney specimens, we were able to isolate and propagate organoid 3D-structures composed of both differentiated and undifferentiated cells while expressing nephron specific markers. Furthermore, we were capable to establish organoids derived from cancer tissues although with a success rate inferior to that of their normal counterpart. Cancer cultures displayed epithelial and mesenchymal phenotype while retaining tumor specific markers. Of note, tumor organoids recapitulated neoplastic masses when orthotopically injected into immunocompromised mice. Our data suggest an innovative approach of long-term establishment of normal- and cancer-derived renal organoids obtained from cultures of fleshly dissociated adult tissues. Our results pave the way to organ replacement pioneering strategies as well as to new models for studying drug-induced nephrotoxicity and renal diseases. Along similar lines, deriving organoids from renal cancer patients opens unprecedented opportunities for generation of preclinical models aimed at improving therapeutic treatments.
Highlights
Regenerative medicine research is based on the rapid advances in stem cell investigation, tissue-engineering and patient-derived models[1,2]
Establishment of healthy kidney organoid cultures Thirteen surgery specimens obtained from normal tissues of neoplastic patients who underwent total nephrectomy were used for culture condition optimization
After 72 hours, suspensions were plated in a specific medium composed by low-density Matrigel supplemented with EGF, basic Fibroblast Growth Factor (bFGF), Rho kinase (ROCK) inhibitor and A8301, a selective inhibitor of the Transforming Growth Factor β type I Receptor (TGF-β), the Anaplatistic lymphoma kinase 4 and 7 (ALK4 and ALK7)[16,17,28], (Fig. 1a and Supplementary Table 1)
Summary
Regenerative medicine research is based on the rapid advances in stem cell investigation, tissue-engineering and patient-derived models[1,2]. Many efforts are devoted to improve stem-cell-based and tissue regenerative technologies. Ethical limitations foster the use of adult-patient-derived-pluripotent stem models including organoids which represent recent technological breakthrough[12]. Organoid models raise expectations in tissue repair[19,20,21,22,23] and cancer therapy testing[20,24,25,26] research. Cancer-derived-organoids have been established from diverse organs[17,18,22,27,28,29,30,31]
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