A PLP-loaded ApoE-based Reconstituted High Density Lipoprotein Nanocarrier: Formation And Its Therapeutic Effect On The Mouse Model Of Systemic Lupus Erythematosus

Objective: Glucocorticoid is the most effective and commonly used drug in the treatment of systemic lupus erythematosus.However,due to the extensive effect of glucocorticoid,there are a large number of off-target effects and many side effects will occur after long-term and high-dose treatment of glucocorticoid,which severely restricts the use of glucocorticoid.Thus,in this study,we constructed a recombinant high-density lipoprotein loading prednisolone sodium phosphate--a new glucocorticoid-containing nanodrug,and applied it to the mouse model of systemic lupus erythematosus,assessing the targeting effect and the therapeutic effect.Methods: 1.Apo E3?calcium phosphate core and prednisolone disodium phosphate was used to construct the nanodrug PLP-Ca P-r HDL.The particle size and zeta potential were determined by Zetasizer.The morphology was observed by transmission electron microscopy and the encapsulation rate was detected by HPLC system.2.The cellular uptake of PLP-Ca P-r HDL by macrophage cell line RAW264.7 was detected by a highcontent screening reader.The tissue distribution of PLP-Ca P-r HDL in MRL/ lpr mice was observed by the Marven tissue imaging system and a laser confocal microscope,so as to explore the targeting effect of PLP-Ca P-r HDL.3.In vitro,real-time PCR and ELISA were used to detect the effect of PLP-Ca P-r HDL on the expression of inflammatory cytokines in macrophage cell line RAW264.7.In vivo,MRL/ lpr mice was selected as the study subjects.Short-term treatment and long-term treatment were adopted to evaluate the possible therapeutic effects of PLP-Ca P-r HDL from the aspects of clinical manifestations,lab test and pathology.Results: 1.The recombinant high-density lipoprotein loading glucocorticoids was successfully constructed.The particle size was 19.46±2.83 nm,zeta potential was-22.87±0.07 m V,and PDI was close to 0.3.The shape was almost spherical,with smooth edge and uniform distribution.The encapsulation rate of PLP-Ca P-r HDL was 82.87%.2.In vitro,PLP-Ca P-r HDL was largely uptaken by macrophage cell line RAW264.7;Inflammatory state promoted the uptake of PLP-Ca P-r HDL by macrophage cell line RAW264.7.In vivo,PLP-Ca P-r HDL was targeted to the inflammatory sites(kidney and joint)of MRL/lpr mice.It is suggested that PLP-Ca P-r HDL is able to target inflammation and macrophages.3.Compared with the common glucocorticoid,PLPCa P-r HDL reduced the expression of inflammatory cytokines in macrophage cell line RAW264.7 more effectively;In vivo,PLP-Ca P-r HDL significantly reduced the disease activity of lupus mice,which was manifested in the improvement of hyponatremia and the reduction of anti-double-stranded DNA level.MRL/lpr mice treated with PLP-Ca Pr HDL showed milder signs of nephritis,especially in the level of urinary protein,renal function and pathological damage.Joint swelling in MRL/ lpr mice was relieved as well.No obvious side effects were observed after long-term treatment.PLP-Ca P-r HDL was safe and could slow down the increase of blood glucose.Conclusion: PLP-Ca P-r HDL was successfully constructed,with desired characterization and encapsulation rate.PLP-Ca P-r HDL was targeted to the inflammatory sites of MRL/lpr mice and macrophage cell line RAW264.7.Compared with common glucocorticoid,PLP-Ca P-r HDL significantly reduced the expression of inflammatory cytokines in macrophage cell line RAW264.7 and effectively alleviated the lupus-like disease in MRL/ lpr mice,without observable cytotoxicity and side effects,even slowing down the increase of blood glucose.PLP-Ca P-r HDL has clinical value,suggesting a potential role in the treatment of systemic lupus erythematosus.