In Vivo Multi-modal Stem Cell Tracing Of Cartilage Reconstruction Based On Up-Conversion Nanoparticles

Background and Objective:At present in the cartilage reconstruction,the combination of seed cells,scaffolds and biological reaction conditions is always the most classic model.Among them,stem cells are the first choice for seed cells,and the outcome of their survival and position distribution in vivo is the basis for their investigation.However,currently used in vivo stem cell tracing techniques in laboratories are relatively limited by such as the high operating costs of fluorescein and cytotoxicity.In the emerging stem cell tracing technology,upconversion nanoparticles(UCN)have drawn broad attention of researchers.With their unique optical properties for upconversion imaging,UCN can make multi-modal cell tracing possible when combined with CT and MRI.It has been revealed that studies based upon UCN bio-application are mainly focused on tumor cell tracking and targeted therapies,while stem cell tracing during tissue repair has been rarely reported.This study aims to explore the applicational value of up-conversion nanomaterials in stem cell labelling and tracking throughout the process of tissue reconstruction.Method:The joint-derived chondrocytes and cartilage progeintor stem cells of rats were labeled with UCN to examine the effects of UCN on cell viability,proliferation,and pluripotency,and to verify the optimal concentration,time,and conditions of co-cultivation.Cells labeled with different concentrations was imaged with 980nm upconversion fluorescence.UCN-labeled cell pellets in hydrogel were subjected to CT and MR imaging,and the intensity of the contrast signal at different UCN concentrations was measured to determine the proper concentration which can be well distinguished from osteochondral and soft tissue.The pellets were also placed under the skin of nude mice,and CT and up-conversion fluorescence were taken at D0,D14 and D28 to record the signal intensity.Results:The lethal dose of UCN towards rat chondrocytes and cartilage progenitor/stem cells(CPSC)were 1.25mg/mL and 1.5mg/mL respectively.When the labeling concentration is greater than 100?g/mL,the differentiation ability of CPSC was inhibited.Signal intensities of up-conversion fluorescence,CT are proportional to the concentrations of UCN.As for MRI,when the concentration of UCN relative to the cell pellet was greater than 2.5 mg/mL,no signal can be received in the T1 and T2 patterns.In up-conversion fluorescence imaging in vivo,UCN could be used to trace hydrogel-conjugated CPSC subcutaneously in nude mice for at least 28 days.At the labeling concentration of 100?g/mL,evident up-conversion luminescence could be obtained at the cell density of 3×106/mL.In the CT imaging,when the cell density was higher than 15×106/mL,the location could be clearly defined and relatively quantified,and when lower than the density,the position of up-conversion luminescence signal was required to be located.