6 research outputs found
Mechanical Stress Downregulates MHC Class I Expression on Human Cancer Cell Membrane
In our body, cells are continuously exposed to physical forces that can regulate different cell functions such as cell proliferation, differentiation and death. In this work, we employed two different strategies to mechanically stress cancer cells. The cancer and healthy cell populations were treated either with mechanical stress delivered by a micropump (fabricated by deep X-ray nanolithography) or by ultrasound wave stimuli. A specific down-regulation of Major Histocompatibility Complex (MHC) class I molecules expression on cancer cell membrane compared to different kinds of healthy cells (fibroblasts, macrophages, dendritic and lymphocyte cells) was observed, stimulating the cells with forces in the range of nano-newton, and pressures between 1 and 10 bar (1 bar = 100.000 Pascal), depending on the devices used. Moreover, Raman spectroscopy analysis, after mechanical treatment, in the range between 700-1800 cm(-1), indicated a relative concentration variation of MHC class I. PCA analysis was also performed to distinguish control and stressed cells within different cell lines. These mechanical induced phenotypic changes increase the tumor immunogenicity, as revealed by the related increased susceptibility to Natural Killer (NK) cells cytotoxic recognition
Raman spectroscopy of different cell lines.
<p>Raman spectra for control and stressed cancer cell lines (Mel 59c (A), Mel 42a (B), Mel 103b(C) and 293T(D)) with standard deviation error bar. The spectra were performed before and after mechanical stress with micropump.</p
Protein concentration in different cell lines.
<p>Normalized area of band centred at around 1440 (top) and 1670 cm<sup>−1</sup> (down) for all the cell lines, showing the variation of protein concentration and relative α-helix content over the cell surface.</p
Western Blotting of MHC class I expression on the supernatants of treated samples: Tumor (A) and healthy cells (B) were analysed before and after mechanical stress by shock waves.
<p>MHC-I has molecular weight of 45 kDa. (C) Membrane incubated with Ponceau S red staining solution, as loading controls.</p
Increased NK susceptibility on mechanical stressed tumor cells.
<p>NK cell recognition of different tumor cell targets at different E/T (effector/target) ratio: 59c, 42a, 66b (melanoma cell lines), 293T (kidney carcinoma) and IM9 (lymphoblastoidcell lines) before (grey) and after (black) mechanical stress. The Mel 42a, Mel 66b, fibroblasts cells (panels B, E, and F) were treated with the micropump, the Mel 59c, IM9, 293 T cells (panels A, C and D) were stressed with the shock waves. As healthy target cells, in this case fibroblasts are shown. Representative experiments are reported for each cell type. Panels G and H show the statistics derived from three different functional assays, using NK lymphocytes as effectors cells (E) and IM9 and Melanoma cells as targets (T). The IM9 target cells were treated with the shock waves (panel G: n = 3, p = 0.0325), while the Melanoma target cells were stressed with the micropump (panel H, n = 3, p = 0.0186). E/T ratio 12/1, p<0.05.</p
Principal component analysis.
<p>PCA analysis on control and stress cells for various cell lines; Mel 42a, Mel 59c, Mel 103b and 293T. a) PC1 vs. PC2, b) PC2 vs. PC3 and c) PC1 vs. PC3.</p