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. Author manuscript; available in PMC: 2021 May 11.
Published in final edited form as: Kidney Int. 2020 Feb 17;97(5):1017–1031. doi: 10.1016/j.kint.2020.01.026

Figure 8. TNC activates integrin αvβ6/FAK/MAPK signaling in vitro.

Figure 8.

(a-c) TNC induced integrin αvβ6 expression in vitro. HKC-8 cells were treated with TNC (50 ng/ml) for various periods of time as indicated. Representative Western blots (a) and quantitative data (b and c) are presented. *P<0.05, **P<0.01 versus controls (n=3). (d-f) TNC induced a rapid FAK and ERK1/2 phosphorylation in vitro. HKC-8 cells were treated with TNC (50 ng/ml) for various periods of time as indicated. Representative Western blots (d) and quantitative data (e and f) are presented. *P < 0.05, **P < 0.01, ***P < 0.001 versus controls (n=3). (g-i) TNC-mediated FAK and ERK1/2 activation was dependent on αvβ6 integrin signaling. HKC-8 cells were pre-incubated with neutralizing antibody against integrin αvβ6 (10D5; 100 µg/ml) for 1 hour, followed by treatment with TNC (50 ng/ml) for 10 min. Representative Western blots (g) and quantitative data (h and i) are presented. *P < 0.05, **P < 0.01 versus controls; † P < 0.05 versus TNC alone (n=3). (j-m) Blocking of αvβ6 integrin signaling abolished the induction of partial EMT by TNC. HKC-8 cells were pre-incubated with 10D5 neutralizing antibody for 1 hour, followed by treatment with TNC for 2 days. Cell lysates were subjected to Western blot analyses with specific antibodies against fibronectin, vimentin, α-SMA and α-tubulin, respectively. Representative Western blots (j) and quantitative data (k-m) are presented. *P < 0.05 versus controls; † P < 0.05 versus TNC alone (n=3). (n-p) TNC-mediated ERK1/2 activation was dependent on FAK signaling. HKC-8 cells were pre-incubated with FAK inhibitor PF573228 (10 µM) for 1 hour, followed by treatment with TNC for 10 minutes. Representative Western blots (n) and quantitative data (o and p) are presented. *P < 0.05 versus controls; † P < 0.05, †† P < 0.01 versus TNC alone (n=3). (q-s) Blocking of FAK signalling abolished the induction of partial EMT by TNC. HKC-8 cells were pre-incubated with PF573228 for 1 hour, followed by treatment with TNC for 2 days. Representative Western blots (q) and quantitative data (r and s) are presented. *P < 0.05, **P < 0.01 versus controls; † P < 0.05, †† P < 0.01 versus TNC alone (n=3).