Nanoparticle enhanced combination therapy for stem-like progenitors defined by single-cell transcriptomics in chemotherapy-resistant osteosarcoma

The adaptation of osteosarcoma cells to therapeutic pressure impedes the efficacy of chemotherapy for osteosarcoma.However,the characteristics and cellular organization of therapy-resistant cells in osteosarcoma tumors remain elusive.Here,we utilized single-cell transcriptomics to systematically map the cell-type-specific gene expression in a chemotherapy-resistant osteosarcoma tumor.Our data demonstrated the VEGFR2-JMJD3-abundant subsets as quiescent stem-like cells,thereby establishing the hierarchy of therapy-resistant actively cycling progenitor pools(JMJD3-abundant)in osteosarcoma.VEGFR2 inhibitor and JMJD3 inhibitor synergistically impeded osteosarcoma cell propagation and tumor growth.Although osteosarcoma cells are predisposed to apoptosis induced by the synergistic therapy through activation of the CHOP pro-apoptotic factor via the endoplasmic reticulum(ER)stress,the stem-like/progenitor cells exhibit an adaptive response,leading to their survival.Reduction in cellular glutathione levels in stem-like/progenitor cells caused by the treatment with a glutathione synthesis inhibitor increases ER stress-induced apoptosis.Importantly,the marked therapeutic improvement of synergistic therapy against stem-like/progenitor cells was achieved by using glutathione-scavenging nanoparticles,which can load and release the drug pair effectively.Overall,our study provides a framework for understanding glutathione signaling as one of the therapeutic vulnerabilities of stem-like/progenitor cells.Broadly,these findings revealed a promising arsenal by encapsulating glutathione-scavenging nanoparticles with co-targeting VEGFR2 and JMJD3 to eradicate chemotherapy-resistant osteosarcoma.

Bioadaptive Nanorod Topography of Titanium Surface to Control Cell Behaviors and Osteogenic Differentiation of Preosteoblast Cells

Titanium （Ti） nanorods fabricated using selective corrosion of Ti substrate by anodic technology show better biocompatibility with pre-osteoblast cells. The current study investigated the response of the murine pre-osteoblast cell MCST3-E1 on Ti nanorod topography and untreated Ti surfaces by means of examination of the morphology and osteogenic differentiation responsible for the pre-osteoblast reaction. The morphology of MCST3-E1 cells was observed using scanning electron microscopy, and alkaline phosphatase （ALP） activity was measured using a colorimetric assay after incubation for 7, 14, and 21 days. The expression of three osteogenic differentiation markers including ALP, osteocalcin （OCN）, and collagen type 1A1 （COL1A1） and two transcription factors including runt related transcription factor 2 （Runx2） and osterix （Osx） at different time points was detected using real-time polymerase chain reaction analysis in both groups. Osx was used to confirm the protein level. The results showed that Ti nanorod surfaces provided prolonged higher levels of ALP activity compared with unmodified Ti surface on the 14th and 21st days. Gene expression analysis of ALP, OCN, and COL1A1 showed significant upregulation with modified nanorod topography after incubation for 14 and 21 days. Osteogenic transcription factors of Runx2 and Osx exhibited changes consistent with the osteogenic differentiation markers, and this may contribute to the persistently active differentiation of MC3T3-E1 cells in the Ti nanorod group. These results demonstrated that the current nanostructured surface may be considered bioadaptive topography to control cellular behaviors and osteoblast differentiation. The in vivo performance and applicability are further required to investigate osseointegration between implant and host bone in the early stages for prevention of aseptic implant loosening.