Targeting delivery of simvastatin using ICAM-1 antibody-conjugated nanostructured lipid carriers for acute lung injury therapy

Figures & data

Table 1. Physicochemical characterization of the NLCs.

Carriers	Conc. (mg/mL)	dn (nm)	PI	Zeta potential (mV)	EE (%)	DL (%)
NLCs-1	1	43.3 ± 11.3	0.206 ± 0.019	−19.5 ± 0.9	–	–
NLCs-2	3	85.3 ± 15.3	0.196 ± 0.028	−22.1 ± 0.5	–	–
NLCs-3	9	198.3 ± 13.3	0.193 ± 0.033	−25.4 ± 0.6	–	–
SV/NLCs-1	1	143.7 ± 2.4	0.171 ± 0.018	−23.0 ± 1.1	66.70 ± 0.88	3.29 ± 0.04
SV/NLCs-2	3	259.6 ± 9.9	0.170 ± 0.036	−22.3 ± 1.9	84.97 ± 0.43	4.15 ± 0.02
SV/NLCs-3	9	337.8 ± 25.8	0.390 ± 0.095	−30.1 ± 1.1	91.04 ± 0.87	4.43 ± 0.04
Anti-ICAM/SV/NLCs	9	354.7 ± 18.2	0.229 ± 0.051	−32.1 ± 3.2	96.78 ± 0.12	4.85 ± 0.01

The d_n, PI, EE and DL represent the hydrodynamic diameters, polydispersity index, drug encapsulation efficiency and drug loading content, respectively. The data represent the mean ± SD (n = 3).

Figure 1. The characterization of formulated SV/NLCs. (A) Representative TEM images of formulated SV/NLCs (bar = 100 nm). (B) In vitro release profile of simvastatin from NLCs. The data represent the mean ± SD (n = 3).

Figure 2. The cytotoxicity and cellular uptake of NLCs against EAs. The cells viability influence of blank NLCs (A) and SV/NLCs (B) against EAs after incubation for 48 h. (C) Cellular uptake of FITC-labeled SV/NLCs (bar = 100 μm). (D) The semi-quantitative analysis of cellular uptake by the software “Image J”. The data represent the mean ± SD (n = 3).

Figure 3. In vivo bio-distribution of non-targeted NLCs. (A) The fluorescence images of excised representative organs from mice. Where A1, A2 were the images of lung tissues merely observed in the same field of vision; A3, A4 were the images of the whole representative organs observed in the same field of vision. (B) The semi-quantitative fluorescence intensity in the organs, expressed as the average fluorescence intensity (avg signal/counts).

Figure 4. In vivo bio-distribution of anti-ICAM/SV/NLCs. (A) The fluorescence images of excised representative organs from mice. The column in each image sequentially showed the anti-ICAM/SV/NLCs bio-distribution in the representative organs of the healthy mice, saline-challenged mice and LPS-induced mice, from left to right. The non-targeted SV/NLCs-3 bio-distribution in the LPS-induced mice was reflected in the leftmost column. (B) The semi-quantified fluorescence intensity in the representative organs, expressed as the average fluorescence intensity (avg signal/counts).

Figure 5. The cellular uptake of anti-ICAM/SV/NLCs. (A) The representative immunofluorescence images revealed the expression of ICAM-1 on EAs (bar = 20 μm). (B) The quantitative analysis of ICAM-1 targeting characteristic of anti-ICAM/SV/NLCs in vitro by a flow cytometer.

Figure 6. In vivo anti-inflammatory assessments. (A) Treatment schedule. (B) TNF-α level. (C) IL-6 level. (D) Total cells counts. (E) Neutrophils counts. #p < 0.05, ##p < 0.01, ###p < 0.001 indicated the indices of corresponding groups compared with anti-ICAM/SV/NLCs treatment group after 24-h administration. *p < 0.05, **p < 0.01, ***p < 0.001 indicated the indices of corresponding groups compared with anti-ICAM/SV/NLCs treatment group after 48-h administration. The data represent the mean ± SD (n = 6).

Figure 7. Histopathological examination. Lung section of (A) control mice, (B) LPS-induced model mice, (C) free simvastatin-treated mice, (D) non-targeted SV/NLCs-3-treated mice, (E) anti-ICAM/SV/NLCs-treated mice. The blue arrows indicate alveolar wall. The red arrows depict the alveolar wall hyperemia. The black arrows indicate the neutrophils infiltration (bar = 50 μm).