Fig 1.
Expression analysis of αvβ3, αvβ5 and NRP-1 in A549 cells and xenografts.
(A) Expression analysis of αvβ3, αvβ5 and NRP-1 in A549 cells by flow cytometry. (B) Expression analysis of αvβ3, αvβ5 and NRP-1 in A549 xenografts by immunofluorescence staining. EG, experimental group; CG, control group. Both αvβ3 and αvβ5 are stained green. NRP-1 is stained red. Nuclei are stained blue. Magnification, × 400; Scale bars = 50 μm.
Fig 2.
Penetration analysis of Evans Blue in extravascular tumor tissues.
(A) The representative images of Evans Blue accumulation in normal organs and in tumors of the mice that received different treatments. The blue intensity denotes the quantity of accumulated Evans Blue. (B) Images of the tumor in (A). (C) Quantification of Evans Blue extracted from tumors and organs in (A) by OD600 measurement. n = 3; Error bars, mean ± SEM; ***p< 0.001.
Fig 3.
Therapeutic efficacy of Gemcitabine combined with iRGD for human NSCLC xenografts.
(A) The tumor volume curve during treatment. Arrows indicate the time of injection. The day when treatment started was recorded as day 0. Tumor volume was measured once every three days until day 30. (B) Average tumor weight of each group at the end of treatment. (C) The body weight shift curve of the mice during the experiment. n = 6. Error bars, mean ± SD; ns, not significant; *** p < 0.001.
Fig 4.
The expression of PCNA in tumor tissue.
(A) Immunohistochemical staining analysis. PCNA-positive cells in the sections are stained brown. The nuclei are stained blue. Representative figures from each group are shown; n = 6; Magnification, × 400; Scale bars = 50 μm. (B) The corresponding quantitative analysis results of PCNA are shown in (A). Five fields from each tumor tissue section were randomly selected for the calculation of the IOD value of the positive region through IPP software, which indicated the amount of antigen expression. (C) Western blot analysis. Total protein from the tumor tissues was extracted. PCNA was detected with β-actin as an internal control. n = 3. (D) The quantitative analysis results of PCNA are shown in (D). The intensity of each strip was analyzed by ImageJ software. The average intensities of PCNA were standardized to β-actin. Error bars, mean ± SD; ns, not significant; *** p <0.001.
Fig 5.
Induction of apoptosis by Gemcitabine combined with iRGD.
(A) Apoptotic cells in the tumor tissues were detected by a TUNEL assay. TUNEL-positive nuclei are stained brown, and TUNEL-negative nuclei are stained blue. The figures shown here are representative of the six tumors in each group. Arrows indicate the apoptotic bodies. Magnification, × 400; Scale bars = 50 μm. (B) Quantitative analysis of the apoptosis index in each group. The percentage of TUNEL-positive cells was counted from 100 randomly selected tumor cells per section. Five sections were counted per tumor. n = 6; Error bars, mean ± SD; *** p < 0.001.