CD146-positive adipose-derived stem cells subpopulation enriched by albumin magnetic sphere ameliorates knee osteoarthritis pain and promotes cartilage repair

Abstract

Background: The use of adipose stem cell (ADSCs) subpopulations in cartilage repair remains poorly characterized. In this study, we constructed an albumin magnetic sphere with specific targeting of CD146 (CD146-AMs) for sorting a subpopulation of CD146-positive ADSCs (CD146 + ADSCs) and explored the role of CD146 + ADSCs on joint pain and cartilage repair in rats with knee osteoarthritis (KOA).

Methods: CD146-AMs were prepared and analyzed in materialistic characterization tests. Subpopulations of CD146 + ADSCs were sorted using CD146-AMs. Surface labeling, viability, and proliferation of a subpopulation of CD146 + ADSCs were evaluated in vitro. Molecular characterization of mRNA and protein expression profiles was analyzed by microarray. A rat KOA pain model was established by the iodoacetic acid method, and KOA pain and the promotion of cartilage repair were assessed after treatment with bilateral joint cavity injections of CD146 + ADSCs.

Results: The CD146-AMs prepared in this study had an average particle size of 242.63 ± 6.74 nm, an average potential of 33.82 ± 3.53 mv, and high CD146 targeting and low cytotoxicity. The positive rate of enriched CD146 + ADSCs was 98.21% and showed a high level of stem cell marker expression and good cell viability. Gene and protein expression profiles showed that CD146 + ADSCs have different cellular functions, especially in regulating inflammation. In the KOA model, low, medium and high concentrations of CD146 + ADSCs were able to improve KOA pain and promote cartilage repair in a concentration-dependent trend.

Conclusions: The CD146-AMs prepared in this study were able to safely and efficiently sort out the CD146 + ADSCs subpopulation. The subpopulation of CD146 + ADSCs has a unique molecular profile that ameliorates KOA pain and repairs cartilage damage in rats, providing a new idea for KOA treatment.

Keywords: Adipose-derived stem cells; CD146; Cartilage; Knee osteoarthritis; Magnetic sphere.

Fig. 1

Preparation of CD146-AMs and experimental flowchart

Fig. 2

Characterization analysis. A Particle size test of CD146-AMs; B potential test of CD146-AMs; C magnetization curve test; D AFM observation graph of CD146-AMs; E TEM observation graph of CD146-AMs; F infrared test

Fig. 3

Target recognition properties and in vitro cytotoxicity assay of CD146-AMs. A Prussian blue staining of CD146-AMs after binding to cells; B SEM observation of CD146-AMs after binding to ADSCs cells; C exploration of the binding time of CD146-AMs to ADSCs; D effects of adding different concentrations of CD146-AMs on the growth of HUVEC cells; E effects of adding different concentrations of CD146-AMs on the growth of ADSCs cells; F BrdU staining

Fig. 4

Growth of CD146 + ADSCs cells with surface antigen detection. A Growth morphology of CD146 + ADSCs cells; B morphology of CD146 + ADSCs cells after HE staining; C growth curve of CD146 + ADSCs cells; D surface antigen expression rate of CD146 + ADSCs cells; E surface antigen expression rate of P3 generation CD146 + ADSCs cells

Fig. 5

Gene and protein expression in ADSCs and CD146 + ADSCs. A Heatmap of differential gene expression in ADSCs and CD146 + ADSCs; B inflammation-related genes that underwent significant changes in CD146 + ADSCs cells; C. volcano map showing the distribution of gene expression; D GO analysis of genes by biological process; E heatmap of cytokine expression in ADSCs and CD146 + ADSCs cell subpopulations; F GO analysis of proteins by molecular function; G KEGG pathway enrichment analysis of proteins

Fig. 6

Pressure pain threshold, thermal pain threshold and histopathological detection of knee cartilage in rats. A Pressure pain threshold measurement in rats at week 2 after modeling; B pressure pain threshold determination in rats at week 3 after modeling; C determination of pressure pain threshold in rats at week 4 after modeling; D determination of thermal pain threshold in rats at week 2 after modeling; E determination of thermal pain threshold in rats at week 3 after modeling; F determination of thermal pain threshold in rats at week 4 after modeling; G Safranine O staining of rat knee joint cartilage tissue (50×); H immunohistochemical staining of IL-6 (40×); I Mankin’s score for histopathology of rat knee joint cartilage; J analysis of IL-6 expression in cartilage tissues of rat knee joints