Fig 2: Anti-LDLR treatment protects mice from CCHFV pathogenesis in mice.a A flowchart of the experiment. C57BL/6 mice were pretreated with anti-IFNAR1 monoclonal antibody MAR1-5A3 (300 µg/mouse) plus control rIgG (n = 10, 100 µg/mouse) or an LDLR blocking antibody (R004) (n = 7, 100 µg/mouse) intraperitoneally 24 h before infection. Another dose of the rIgG or R004 mAb (100 µg) was administrated 1 h prior to infection. Mice were challenged with CCHFV (100 TCID50 per mouse, subcutaneously). Twenty-four hours post infection, MAR1-5A3 (200 µg/mouse) was administrated. The rIgG or R004 mAb was administrated at 100 µg quaque die for 5 days post infection. b, c, Protective effects of LDLR blocking antibody on fatality caused by CCHFV infection. As described in (a), C57BL/6 mice were treated with rIgG (n = 10) or the LDLR blocking antibody (R004) (n = 7) and subjected to CCHFV challenge. Body weight (b) and survival (c) of the mice were monitored daily. d, e Protective effects of LDLR blocking antibody on CCHFV caused pathogenesis. As described in (a), C57BL/6 mice were treated with rIgG (n = 6) or the LDLR blocking antibody (R004) (n = 6) and subjected to CCHFV challenge. Mice were necropsied and the livers and spleens were collected at day 5 post infection. Viral loads were quantified by qRT-PCR and shown as the number of viral RNA copies per microgram of organs or per mL of sera (d). H&E staining and immunostaining with anti-Gn mAb (7A11) were performed and the pathological changes were indicated (e). The extensive necrosis (white arrowheads) and necrotic cellular debris (white arrows) in liver were indicated; white pulps in the spleen (white asterisks) were marked. The bars represent 100 µm. Data are represented as mean ± SD. *P < 0.05; **P < 0.01.

Fig 3: APOO deficiency accelerated atherosclerosis in hyperlipidemic mouse models.Eight-week-old Ldlr single knockout (Apoo+/+ Ldlr−/−) and Apoo/Ldlr dKO (Apoo−/− Ldlr−/−) mice were randomly grouped (n = 16–20 for male, n = 13–15 for female) and fed an HCD diet for 12 weeks. A Plasma TC levels in Apoo+/+Ldlr−/− and Apoo−/− Ldlr−/− male and female mice. B Quantification of the aorta, thoracic aorta, and abdominal aorta lesion areas, presented as a percentage area of the entire aorta. C Representative oil-red O-stained aortas. (D, E) H&E staining was used to confirm the atherosclerotic plaque at the aortic root from both male D (n = 9 for the male Apoo+/+ Ldlr−/− group, n = 7 for the male Apoo−/− Ldlr−/− group) and female (E) (n = 10 for the female Apoo+/+ Ldlr−/− group, n = 12 for the female Apoo−/− Ldlr−/− group) mice. Scale bar = 200 µm. (F, G) F4/80 staining was used to detect macrophage infiltration at the aortic root plaque in male F and female G mice. Scale bar = 100 µm. Masson staining was used to determine the collagen composition of the aortic root plaque in male (H) and female (I) mice. Scale bar = 100 µm. Values are represented as mean ± SEM. Two-way ANOVA (B) or unpaired two-tailed Student’s t-test for other panels. *p < 0.05, **p < 0.01, ***p < 0.001. ANOVA analysis of variance, SEM standard error of the mean, APOO apolipoprotein O, HCD high cholesterol diet, H&E hematoxylin and eosin.

Fig 4: LDLR is required for CCHFV pathogenesis in mice.a, b WT (n = 11) and LDLR-knockout (n = 11) mice were pretreated with anti-IFNAR1 monoclonal antibody MAR1-5A3 (200 µg/mouse) 24 h before infection and then infected with CCHFV (10 TCID50) via the intraperitoneal route. Forty-eight hours post infection, 200 µg of MAR1-5A3 was administrated. The body weight (a) and survival (b) of the mice were monitored daily. c, d WT (n = 7) and LDLR-knockout (n = 7) mice were pretreated with anti-IFNAR1 monoclonal antibody MAR1-5A3 (200 µg/mouse) 24 h before infection and then infected with CCHFV (10 TCID50) via the intraperitoneal route. Forty-eight hours post infection, 200 µg of MAR1-5A3 was administrated. Mice were necropsied and the livers and spleens were collected at day 3 and 5 post infection. Viral loads were quantified by RT-qPCR and shown as the number of viral RNA copies per microgram of organs or per mL of sera (c). H&E staining and immunostaining with anti-Gn mAb (7A11) were performed and the pathological changes were indicated (d). The extensive necrosis (white arrowheads) and necrotic cellular debris (white arrows) in the liver were indicated; white pulps in the spleen (white asterisks) were marked. The bars represent 100 µm. Data are represented as mean ± SD. **P < 0.01; ***P < 0.001; ****P < 0.0001.

Fig 5: APOO regulated the saturation of fatty acyl chains of phosphatidylcholine (PC), which was correlated with cholesterol levels.A–D Eight-week-old Apoo−/− and Apoo+/+ mice were randomly grouped and fed an HFD diet for 12 weeks, liver lipidomics were performed for lipid composition analysis (n = 8). A Global abundance of LPS, MGDG, CerG1, PG, Co, PS, SM, PE, DG, TG, and PC revealed by lipidomics, B)analysis of fatty acyl composition of PC by the total degree of unsaturation, C, D correlation levels between the selected lipid species that were analyzed and blood TC or LDL-C levels. E The proteome of livers from male HFD-fed Apoo+/+ and Apoo−/− mice (n = 3) were performed. Heatmap of the abundance profile of proteins related to mitochondrial function was shown. F Representative TEM images of the mitochondrion in male NCD-fed Apoo+/+ and Apoo−/− mice. Quantification represents numbers of CJs in one hundred arbitrarily chosen mitochondria per group. G Oxygen consumption rate (OCR) determined by XFe96 Seahorse in hepatocytes from Apoo−/− and Apoo+/+ mice. Measures of mitochondrial respiration were calculated from the OCR trace. H Volcano plot of cholesterol metabolism-related proteins of E. I Summary of the pathways regulating bile acid metabolism of (E); green arrows indicate downregulated proteins, while red arrows indicate upregulated proteins. LDLR low-density lipoprotein receptor, SR-BI scavenger receptor class B type I, CYP7A1 cytochrome P450 family 7 subfamily A member 1, UGT1A2 UDP-glucuronosyltransferase family 1, CA2 carbonic anhydrase 2, SLCO1A1 solute carrier organic anion transporter family member 1A1; SLCO1A4: Solute carrier organic anion transporter family member 1A4; HDL high-density lipoprotein; BA bile acid. J Significantly upregulated pathway enriched in differentially expressed proteins of (E). Values are represented as mean ± SEM. Unpaired two-tailed Student’s t-test (A, B), or Mann–Whitney test (F) or two-way ANOVA (G). *p < 0.05, **p < 0.01, ***p < 0.001. LPS lysophosphatidylserine, MGDG monogalactosyldiacylglycerol, CerG1 glucocerebroside, PG phosphatidylglycerol, Co coenzyme, PS phosphatidylserine; PE phosphatidylethanolamine. DG diglycerides, TG triglyceride, PC phosphatidylcholine, PL phospholipid; TM6SF2 transmembrane 6 superfamily member 2; FDPS farnesyl pyrophosphate synthase; NSDHL sterol-4-alpha-carboxylate 3-dehydrogenase, decarboxylating, LSS lanosterol synthase; DHCR7 7-dehydrocholesterol reductase, cAV1 Caveolin-1, ACLY ATP-citrate synthase.