Fig. 10: Silencing of MRPL17 attenuates subcutaneous and in situ NSCLC xenograft tumor growth in nude mice. | Cell Death & Disease

Fig. 10: Silencing of MRPL17 attenuates subcutaneous and in situ NSCLC xenograft tumor growth in nude mice.

From: MRPL17 is a critical regulator of mitochondrial function and a novel therapeutic target in non-small cell lung cancer

Fig. 10

Analysis of subcutaneous xenografts derived from primary pNSCLC-1 cells stably expressing control shRNA (shC) or shRNA targeting MRPL17 (kdMRPL17-sh2) engrafted into nude mice. Longitudinal monitoring of tumor volume over 48 days post-initiation (Day-0 defined as 3 weeks post-engraftment) (A), calculation of the derived tumor growth rate (B), assessment of explanted tumor mass at Day-48 (C), and longitudinal monitoring of host body weights (D). Validation of target modulation within tumor tissues explanted at Day-18 and Day-30 included qRT-PCR analysis of MRPL17, MRPL12 and COX8A mRNA levels (E, L), immunoblotting analysis with quantification of MRPL17, MRPL12 and COX8A protein levels (F, M), and immunohistochemical (IHC) assessment of MRPL17 protein expression at Day-30 (G). Analyses of mitochondrial function and redox state performed on lysates from tumors explanted at Day-18 and Day-30 encompassed measurement of mitochondrial Complex I enzymatic activity (H), quantification of total cellular ATP contents (I), determination of the cellular GSH/GSSG ratio (J), and measurement of lipid peroxidation via TBAR assay (K). Further assessments included immunohistochemical evaluation of Ki-67-positive proliferating cells in tumor sections (N), quantification of cytosolic Cytochrome c levels in tumor lysates (O), immunoblotting analysis of Cleaved-Caspase-3 and Cleaved PARP1 protein levels in tumor lysates (P), and TUNEL assay for detection of apoptotic cells in tumor sections (Q). The bar graph illustrated the incidence of tumor formation, defined as the proportion of mice that successfully formed clear in situ pNSCLC-1 tumors in both the shC control group and the kdMRPL17-sh2 group. A significant decrease in in situ pNSCLC-1 tumor formation was observed in the kdMRPL17-sh2 group (2 out of 9 mice) compared to the shC control group (5 out of 9 mice) (R). Statistical significance was assessed relative to the shC control group (P < 0.05 denoted by asterisks; N.S., non-significant, P > 0.05). Data for A–D reflect n = 10 mice per group. For E–Q analyses utilized five distinct tissues randomly selected per xenograft (n = 5). For R, n = 9 mice per group. Scale bars represent 100 µm.

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