Fig 1: Evaluation of Biosafety of SM102 LNPs- Il1rn mRNA treatment. (A) H&E-stained sections of rat heart, spleen, liver, kidney, lung, and pancreas from Control, Untreated, and Treated groups after a 4-week intervention. Scale bar = 100 µm (B) Biochemical analysis of serum markers indicating organ function in different groups. The panels display levels of AST, ALT, CREA, UREA, DBIL, and TBIL. *p < 0.05; **p < 0.01; ***p < 0.001; n = 5; All data are shown as the mean ± Standard Error of the Mean (SEM). Statistical significance was determined by one-way ANOVA with Fisher’s LSD test.
Fig 2: Schematic illustrating SM102 LNPs- Il1rn mRNA-mediated therapeutic mechanisms against tendinopathy through coordinated modulation of inflammation, MMPs expression, collagen synthesis, and tenocyte migration (By Figdraw).
Fig 3: Assessment of the in vitro transfection efficiency of mRNA mediated by SM102 LNPs. (A) Schematic diagram of SM102 LNPs. (B) Dynamic light scattering distribution plot of SM102 LNPs-Il1rn. (C) Representative transmission electron microscopy image of SM102 LNPs-Il1rn. (D) Denaturing agarose gel electrophoresis of Il1rn mRNA before (lane 1) and after (lane 2) SM102 LNPs encapsulation. (E) Detection of IL-1RA protein in HEK293T cells treated with (lane 2) or without (lane 1) SM102 LNPs- Il1rn mRNA. Lane 3 was the positive control (Commercially available human IL-1RA protein) (F) Representative images showing the SM102 LNPs-mediated transfection of GFP mRNA in HEK293T cells. Scale bar: 50 μm. (G) Flow cytometry results of GFP positive cells for the control group (upper panel) and transfection group (Lower panel) at 48 h post-treatment. (H) Representative images showing the SM102 LNPs-mediated transfection of GFP mRNA in primary tendon stem cells. Scale bar: 50 μm. (I) Quantification of the fluorescence intensity of GFP from 24 h to 72 h post-transfection. (J) ELISA assay for detection of IL-1RA levels in the culture medium of tendon cells treated with 2 ng/μL SM102 LNPs- Il1rn mRNA from Day 1 to Day 7 post-transfection. (K) ELISA assay for detection of IL-1RA levels in the culture medium of tendon cells treated with different concentration of SM102 LNPs- Il1rn mRNA at 24 h post-transfection. *p < 0.05, **p < 0.01, ***p < 0.001; n = 3; All data are shown as the mean ± Standard Error of the Mean (SEM); Statistical significance was determined by one-way ANOVA with Fisher’s LSD test.
Fig 4: In vivo therapeutic effects of SM102 LNPs- Il1rn mRNA on tendon tissues at week 1. (A) Scheme of animal experiment design (By Figdraw). (B) Representative images of H&E staining (upper panel), Masson’s trichrome staining (middle panel), and Sirius red staining (lower panel) of tendons from different groups in the first week postoperatively. Scale bar = 100 µm (C,D) Quantification analysis of Masson’s trichrome staining (C) and Sirius red staining (D). (E) Representative images of immunofluorescent staining of MMP1, MMP13, IL-6, and IL-10 of sections of each group in the first week postoperatively. Scale bar = 100 µm (F–I) Quantification of MMP1, MMP13, IL-6, and IL-10 expression in different groups. (J) Elisa assay for detection of IL-6 (Left) and IL-10 (Right) expression level in tendon tissue from different groups. (K) Representative images of immunofluorescent staining of iNOS of sections of each group in the first week postoperatively. Scale bar = 100 µm (L) Quantification of iNOS expression in different groups. *p < 0.05; **p < 0.01; ***p < 0.001; n = 5; All data are shown as the mean ± Standard Error of the Mean (SEM). Statistical significance was determined by one-way ANOVA with Fisher’s LSD test.
Fig 5: In vitro anti-inflammation property of IL-1RA on tendon stem cells. (A) qPCR results of Il6 (upper), Nos2 (middle) and Il10 (Lower) in control, IL-1β and SM102 LNPs-Il1rn group. (B) Representative images showing the Col I expression in control, IL-1β, and IL-1β+SM102 LNPs-Il1rn group. Col I (Red); phalloidin (Green); Nucleus (Blue). Scale bar = 100 μm (C) Quantification analysis of Col I expression in control, IL-1β and IL-1β+ SM102 LNPs-Il1rn group. (D) Representative images showing the Col III expression in control, IL-1β and IL-1β+ SM102 LNPs-Il1rn group. Col III (Red); phalloidin (Green); Nucleus (Blue). Scale bar = 100 μm (E) Quantification analysis of Col III expression in control, IL-1β, and IL-1β+ SM102 LNPs-Il1rn group. (F) Quantification analysis of Col III/Col I expression in control, IL-1β, and IL-1β+ SM102 LNPs-Il1rn group. (G) Representative microscopic images showing the scratch closure in control, IL-1β, and IL-1β+ SM102 LNPs-Il1rn group at 0 h, 12 h, and 24 h. Scale bar = 200 μm (H) Quantification of scratch closure in different groups at 0 h, 12 h, and 24 h *p < 0.05, **p < 0.01, ***p < 0.001; n = 3; All data are shown as the mean ± Standard Error of the Mean (SEM); Statistical significance was determined by one-way ANOVA with Fisher’s LSD test (A,C,E); Statistical significance was determined by two-way ANOVA with Fisher’s LSD test (G).
Supplier Page from Abcam for Recombinant Human IL-1ra Protein Standard (His tag)