Cholesterol

Protocatechualdehyde (Lot: HS14510S1) and hydroxysafflor yellow A (Lot: HR21714B1) were purchased from Baoji Herbest Bio-Tech (Xi'an, China). Cholesterol (>95 % purity) and propylene glycol were obtained from Macklin Biochemical Co., Ltd (Shanghai, China). Zebrafish feed was obtained from Shanghai FishBio Co., Ltd (Shanghai, China). Oil Red O and Methyl cellulose were obtained from Sigma-Aldrich, Inc. (St. Louis, MO, USA). Simvastatin and Egg yolk powder were purchased from Shanghai source leaf Biological Technology Co., Ltd. (Shanghai, China). Dimethyl Sulfoxide (DMSO) was obtained from Solarbio Life Science (Beijing, China).

HSPC (EK20010) was purchased from A.V.T. Pharmaceutical Co. Ltd. (Shanghai, China), and the liposomes were prepared via a film dispersion method. Briefly, HSPC, cholesterol (Macklin, China), octadecylamine (Aladdin, China), and RAPA (Macklin, China) (40:10:4:6, w/w/w/w) were dissolved in chloroform and evaporated at 60°C for 30 min. The dried lipid film was then hydrated with deionized water and sonicated for 20 min to obtain dispersed multilamellar liposomes. The liposomes were then downsized by stepwise extrusion through 0.45- and 0.22-μm membrane filters (Millex, Ireland). The morphology of liposomes was observed under TEM (FEI Talos L120C, USA). In addition, the liposome size, PDI, and zeta potential were measured by dynamic light scattering using a Zetasizer (Malvern Nano-ZS, UK). To verify the cartilage-targeting ability of the cationic liposomes, a cartilage section was obtained from a freshly slaughtered pig knee joint and incubated with Dil (Beyotime, China)–labeled liposomes for 1 hour at 37°C. The section was then washed with PBS before observation using an LSCM (ZEISS, Germany). To confirm that the cationic liposomes can be internalized by chondrocytes, human chondrocyte C-28/I2 cells were incubated with Dil-labeled liposomes for 12 hours and then observed under an LSCM.

Uric acid and l-α-phosphatidylcholine (l-α-PC) were obtained from Sigma–Aldrich. Chitosan hydrochloride (standard for medical products in tissue engineering) was provided by Golden–Shell Pharmaceutical Co., Ltd (Yuhuan, China). HA-Oligo (MW < 10 kDa) was purchased from Bloomage Freda Biopharm Co., Ltd (Jinan, China). Kollidon@ 90F (polyvinyl pyrrolidone, PVP K90) was kindly donated by BASF (Ludwigshafen, Germany). Polydimethylsiloxane (PDMS) was procured from Dow Corning Co., Ltd (Midland, MI, USA). Colchicine, cholesterol, uricase, sucrose, rhodamine-6G, fluorescein5 (6)-isothiocyanate, and dextran were purchased from Shanghai Macklin Biochemical Co., Ltd (Shanghai, China). All chemicals applied to HPLC were of chromatographic grade. And all the other chemicals were of analytical grade.
Abelson murine leukemia virus-induced tumor cell line, raw264.7, was donated by the Institute of Pharmaceutical Analysis, School of Pharmaceutical Sciences (Shenzhen), Sun Yat-sen University.
Adult male Sprague–Dawley rats were obtained from Laboratory Animal Center, Sun Yat-sen University. All animal studies were performed under the protocol approved by Institutional Animal Care and Use Committee, Sun Yat-sen University (Guangzhou, China, Approval No.: SYSU-IACUC-2021-000521, 01 Sep. 2021).