Journal: Life Science Alliance

Article Title: Rnd3 regulates lung cancer cell invasion and migration independently of ROCK1 signaling via alpha 5 integrin modulation

doi: 10.26508/lsa.202503494

Figure Lengend Snippet: (A) Representative epifluorescence images of anti-alpha 5 integrin (green) and nuclear DAPI stain (blue) in A549 NSC and siRnd3 (oligo A), siAlpha5 (oligo A), and siRnd3:Alpha5–knockdown A549 cells, imaged with equal exposure times using a 60x oil objective; scale bar = 50 μm. (B) Analysis of alpha 5 fluorescence intensity (per biological repeat, five fields of view were captured with equal exposure time using a 40x objective per condition, 10 cells plus a section of background were defined as region of interest in Nikon NIS-Elements software, and the average intensity for each cell minus background was determined). Data are presented as individual cells with bar representing overall mean for each biological repeat ± SEM, n = 4. (C) Invasion assay across a transwell membrane coated with Matrigel, and percentage of invaded cells compared with NSC cells at 24 h. Data are presented as an individual mean for each experiment with bar representing overall mean ± SEM, n = 3. (D) Wound healing assay. Percentage wound closure at 24 h compared with wound at 0 h. Data are presented as an individual mean for each experiment with bar representing overall mean ± SEM, n = 4. (E) Representative Western blots in which decreased protein expression levels of Rnd3 (oligo A) and alpha 5 (Alpha 5) integrin (oligo A) are observed, corresponding to siRNA treatment compared with the non-silencing control. Tubulin was used as a loading control. (F, G) Quantification of representative Western blots. (F, G) Densitometric analysis was performed for each sample set over multiple biological replicates, n = 4, for Rnd3 (F) and alpha 5 (G) expression after transient transfection with siRNA oligo compared with expression levels in NSC A549 cells. Expression levels of each protein of interest were normalized to own tubulin loading control, then normalized to the expression level of that protein of interest in the NSC cells. Data are presented as an individual mean for each experiment with bar representing overall mean ± SEM. Statistical comparisons were performed using one-way ANOVA with Dunnett’s multiple test correction. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, ns, not significant; AU, arbitrary units.

Article Snippet: For epifluorescence, cells were visualized with a Nikon Ts2-FL inverted microscope equipped with a Panda sCMOS camera and images were captured with Nikon NIS-Elements.

Techniques: Staining, Knockdown, Fluorescence, Software, Invasion Assay, Membrane, Wound Healing Assay, Western Blot, Expressing, Control, Transfection

Journal: Nature Communications

Article Title: Serum-tolerant polymeric complex for stem-cell transfection and neural differentiation

doi: 10.1038/s41467-025-57278-8

Figure Lengend Snippet: a Representative fluorescent images of MSCs after FAM-miRNA (green) transfection with 25K PEI FAM-miRNA , 10K PEI FAM-miRNA , BP FAM-miRNA , and APOs@BP FAM-miRNA in medium with or without 10% FBS. b Cell viability after transfection with 25k PEI miRNA , 10k PEI miRNA , BP miRNA , and APOs@BP miRNA in medium with or without 10% FBS. c Cell proliferation in 3 days after transfection with 25K PEI miRNA , 10K PEI miRNA , BP miRNA , and APOs@BP miRNA in medium with 10% FBS. d Representative TEM images of 25K PEI miRNA , 10K PEI miRNA , BP miRNA , and APOs@BP miRNA after incubation in 10% FBS for 2 h. e Evaluation of the stability of 25K PEI miRNA , 10K PEI miRNA , BP miRNA, and APOs@BP miRNA in the presence or absence of serum by measuring particle size, respectively. f Determination of protein adsorption of polymeric complexes by SDS-PAGE. g Schematic illustration of serum-tolerant polymeric complexes to avoid serum-protein adsorption . h SPR analysis of MSC membrane binding affinity of BSA@BP miRNA and APOs@BP miRNA i CLSM images of lipid vesicles (labeled with DiI, red) after incubation with FAM labeled BSA@BP miRNA and APOs@BP miRNA (green), respectively. The solid white line denotes the straight-line distance employed for colocation analysis between DiI and FAM signals on the right. j The conformational content of APOs in APOs@BP miRNA before and after incubation with lipid vesicles. k Schematic illustration of intracellular trafficking of APOs@BP miRNA . l The dynamic colocalization of APOs@BP Cy5-miRNA (green) and lysosomes (red) in MSCs after incubation for 0.5 and 2 h, respectively. m Intracellular accumulation and ROS-triggered dissociation of Cy5-miRNA (red) from polymerized carrier (FBP, green). Data were presented as mean ± SD ( n = 5 independent samples in b ; n = 6 independent samples in c ; n = 3 independent samples in e , j ;). Two-tailed unpaired t -test was used for statistical analysis of b , j . One-way ANOVA with Dunnett’s multiple comparisons test was used for statistical analysis of c . Three independent experiments were performed with similar results and representative results are shown. Source data are provided as a Source Data file.

Article Snippet: Different polymers, including 10K PEI, 25K PEI, BP, and APOs@BP, compressed FAM-labeled miRNA (FAM-miRNA, RIBOBIO, miR01104-1-1) at their optimized N/P ratios and dissolved in DMEM containing 0 or 10% FBS to maintain a final miRNA concentration of 120 nM, which were then incubated with MSCs for 6 h. The transfected MSCs were washed three times with PBS and observed using an inverted fluorescence microscope (Eclipse Ts2-FL, Nikon Precision Inc., CA, USA).

Techniques: Transfection, Incubation, Adsorption, SDS Page, Membrane, Binding Assay, Labeling, Two Tailed Test