pan pkc Search Results


pser660 pkcβ  (Cell Signaling Technology Inc)
96
Cell Signaling Technology Inc pser660 pkcβ
Pser660 Pkcβ, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/Phospho-PKC+(pan)+(betaII+Ser660)+Antibody/pmc11310474-305-72-74
Average 96 stars, based on 1 article reviews
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phospho pkc pan  (Cell Signaling Technology Inc)
93
Cell Signaling Technology Inc phospho pkc pan
( A ) IP3R blocking inhibits PLCE1-induced signal transduction in the axis of <t>PKC/GSK-3β/β-Catenin.</t> ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with 2-APB (10 µM) for additional 18 hr. ( B ) [Ca 2+ ] blocking represses signal transduction in the axis of PKC/GSK-3β/β-Catenin induced by PLCE1. ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with BAPTA-AM (10 µM) for additional 18 hr. ( C ) PKC inhibition blocks PLCE1 stimulated signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmid expressing Plce1 . 6 hr post-transfection, cells were treated with 100 nM of Midostaurin for additional 18 hr. ( D ) PKC inhibition represses PLCE1-induced nuclear translocation of β-Catenin in ECA-109 cells. Cells were transfected with the plasmid expressing PLCE1. 6 hr post-transfection, 2-APB (10 µM), BAPTA-AM (10 µM), or Midostaurin (100 nM) was added in culture medium, and cells were cultured for additional 18 hr. Scale bar = 10 µm. Immunofluorescence was used to examine subcellular distribution of β-Catenin. ( E <t>)</t> <t>TAK1</t> counteracts PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated for 24 hr. ( F ) TAK1 reduces PLCE1-induced nuclear distribution of β-Catenin in ECA-109 cells. Cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated. Scale bar = 10 µm. ( G ) Dominant negative TAK1 (K63W) fails to block signal transduction in the axis of PKC/GSK-3β/β-Catenin/MMP2 induced by PLCE1. ECA-109 cells were transfected with the plasmids expressing Plce1 or mutated Map3k7 (TAK1 K63W) for 24 hr. ( H ) TAK1 has no effect on PLCE1 S1060A-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing PLCE1 S1060A or TAK1 for 24 hr. Protein levels were analyzed by western blot, and Actin was used as a loading control. Representative blots were shown. Figure 5—source data 1. PDF file containing original western blots for , indicating the relevant bands. Figure 5—source data 2. Original files for western blot analysis displayed in .
Phospho Pkc Pan, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/Phospho-PKC+(pan)+(gamma+Thr514)+Rabbit+mAb/pmc12017773-278-11-71
Average 93 stars, based on 1 article reviews
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polyclonal anti pkc antibody  (Elabscience Biotechnology)
96
Elabscience Biotechnology polyclonal anti pkc antibody
( A ) IP3R blocking inhibits PLCE1-induced signal transduction in the axis of <t>PKC/GSK-3β/β-Catenin.</t> ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with 2-APB (10 µM) for additional 18 hr. ( B ) [Ca 2+ ] blocking represses signal transduction in the axis of PKC/GSK-3β/β-Catenin induced by PLCE1. ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with BAPTA-AM (10 µM) for additional 18 hr. ( C ) PKC inhibition blocks PLCE1 stimulated signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmid expressing Plce1 . 6 hr post-transfection, cells were treated with 100 nM of Midostaurin for additional 18 hr. ( D ) PKC inhibition represses PLCE1-induced nuclear translocation of β-Catenin in ECA-109 cells. Cells were transfected with the plasmid expressing PLCE1. 6 hr post-transfection, 2-APB (10 µM), BAPTA-AM (10 µM), or Midostaurin (100 nM) was added in culture medium, and cells were cultured for additional 18 hr. Scale bar = 10 µm. Immunofluorescence was used to examine subcellular distribution of β-Catenin. ( E <t>)</t> <t>TAK1</t> counteracts PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated for 24 hr. ( F ) TAK1 reduces PLCE1-induced nuclear distribution of β-Catenin in ECA-109 cells. Cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated. Scale bar = 10 µm. ( G ) Dominant negative TAK1 (K63W) fails to block signal transduction in the axis of PKC/GSK-3β/β-Catenin/MMP2 induced by PLCE1. ECA-109 cells were transfected with the plasmids expressing Plce1 or mutated Map3k7 (TAK1 K63W) for 24 hr. ( H ) TAK1 has no effect on PLCE1 S1060A-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing PLCE1 S1060A or TAK1 for 24 hr. Protein levels were analyzed by western blot, and Actin was used as a loading control. Representative blots were shown. Figure 5—source data 1. PDF file containing original western blots for , indicating the relevant bands. Figure 5—source data 2. Original files for western blot analysis displayed in .
Polyclonal Anti Pkc Antibody, supplied by Elabscience Biotechnology, used in various techniques. Bioz Stars score: 96/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/Pan+PKC+Polyclonal+Antibody/pmc07227959-76-32-37
Average 96 stars, based on 1 article reviews
polyclonal anti pkc antibody - by Bioz Stars, 2026-10
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rabbit phospho pkc pan thr410 antibody  (Cell Signaling Technology Inc)
93
Cell Signaling Technology Inc rabbit phospho pkc pan thr410 antibody
( A ) IP3R blocking inhibits PLCE1-induced signal transduction in the axis of <t>PKC/GSK-3β/β-Catenin.</t> ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with 2-APB (10 µM) for additional 18 hr. ( B ) [Ca 2+ ] blocking represses signal transduction in the axis of PKC/GSK-3β/β-Catenin induced by PLCE1. ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with BAPTA-AM (10 µM) for additional 18 hr. ( C ) PKC inhibition blocks PLCE1 stimulated signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmid expressing Plce1 . 6 hr post-transfection, cells were treated with 100 nM of Midostaurin for additional 18 hr. ( D ) PKC inhibition represses PLCE1-induced nuclear translocation of β-Catenin in ECA-109 cells. Cells were transfected with the plasmid expressing PLCE1. 6 hr post-transfection, 2-APB (10 µM), BAPTA-AM (10 µM), or Midostaurin (100 nM) was added in culture medium, and cells were cultured for additional 18 hr. Scale bar = 10 µm. Immunofluorescence was used to examine subcellular distribution of β-Catenin. ( E <t>)</t> <t>TAK1</t> counteracts PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated for 24 hr. ( F ) TAK1 reduces PLCE1-induced nuclear distribution of β-Catenin in ECA-109 cells. Cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated. Scale bar = 10 µm. ( G ) Dominant negative TAK1 (K63W) fails to block signal transduction in the axis of PKC/GSK-3β/β-Catenin/MMP2 induced by PLCE1. ECA-109 cells were transfected with the plasmids expressing Plce1 or mutated Map3k7 (TAK1 K63W) for 24 hr. ( H ) TAK1 has no effect on PLCE1 S1060A-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing PLCE1 S1060A or TAK1 for 24 hr. Protein levels were analyzed by western blot, and Actin was used as a loading control. Representative blots were shown. Figure 5—source data 1. PDF file containing original western blots for , indicating the relevant bands. Figure 5—source data 2. Original files for western blot analysis displayed in .
Rabbit Phospho Pkc Pan Thr410 Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/Phospho-PKC+(pan)+(zeta+Thr410)+Rabbit+mAb/pmc09866800-116-29-43
Average 93 stars, based on 1 article reviews
rabbit phospho pkc pan thr410 antibody - by Bioz Stars, 2026-10
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ap0520  (ABclonal Biotechnology)
94
ABclonal Biotechnology ap0520
( A ) IP3R blocking inhibits PLCE1-induced signal transduction in the axis of <t>PKC/GSK-3β/β-Catenin.</t> ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with 2-APB (10 µM) for additional 18 hr. ( B ) [Ca 2+ ] blocking represses signal transduction in the axis of PKC/GSK-3β/β-Catenin induced by PLCE1. ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with BAPTA-AM (10 µM) for additional 18 hr. ( C ) PKC inhibition blocks PLCE1 stimulated signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmid expressing Plce1 . 6 hr post-transfection, cells were treated with 100 nM of Midostaurin for additional 18 hr. ( D ) PKC inhibition represses PLCE1-induced nuclear translocation of β-Catenin in ECA-109 cells. Cells were transfected with the plasmid expressing PLCE1. 6 hr post-transfection, 2-APB (10 µM), BAPTA-AM (10 µM), or Midostaurin (100 nM) was added in culture medium, and cells were cultured for additional 18 hr. Scale bar = 10 µm. Immunofluorescence was used to examine subcellular distribution of β-Catenin. ( E <t>)</t> <t>TAK1</t> counteracts PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated for 24 hr. ( F ) TAK1 reduces PLCE1-induced nuclear distribution of β-Catenin in ECA-109 cells. Cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated. Scale bar = 10 µm. ( G ) Dominant negative TAK1 (K63W) fails to block signal transduction in the axis of PKC/GSK-3β/β-Catenin/MMP2 induced by PLCE1. ECA-109 cells were transfected with the plasmids expressing Plce1 or mutated Map3k7 (TAK1 K63W) for 24 hr. ( H ) TAK1 has no effect on PLCE1 S1060A-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing PLCE1 S1060A or TAK1 for 24 hr. Protein levels were analyzed by western blot, and Actin was used as a loading control. Representative blots were shown. Figure 5—source data 1. PDF file containing original western blots for , indicating the relevant bands. Figure 5—source data 2. Original files for western blot analysis displayed in .
Ap0520, supplied by ABclonal Biotechnology, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/Phospho-PKC+(pan)+(zeta)-T410+Rabbit+pAb/pmc13124628-26-7-5
Average 94 stars, based on 1 article reviews
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pan-pkc inhibitor gö6983  (STEMCELL Technologies Inc)
90
STEMCELL Technologies Inc pan-pkc inhibitor gö6983
Effects of p38 mitogen-activated protein kinase (p38 MAPK), mitogen-activated protein kinase kinase 1/2 (MEK1/2) and protein kinase C (PKC) inhibitors on stretch-induced hypertrophic gene expression in human pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The following inhibitors were utilized: SB203580 at 10 μM to inhibit p38 MAPK, U0126 at 10 μM to inhibit MEK1/2, <t>Gö6983</t> at 1 μM to inhibit all PKC isoforms and Gö6976 at 1 μM to inhibit classical PKC isoforms. Natriuretic peptide B ( NPPB ; A), skeletal muscle alpha actin 1 ( ACTA1 ; B), galanin ( GAL ; C) and long intergenic non-protein coding RNA 648 ( LINC00648 ; D) RNA expression was measured with qRT–PCR after a 24-h cyclic mechanical stretch. The results are presented as fold change relative to the unstretched control. The data are shown as mean ± standard error of the mean, and values from individual experiments are presented as dots ( n = 5, except for SB203580 and Gö6976 n = 4; where n represents biological replicates of cells from individual differentiations). * p < 0.05, ** p < 0.01 vs. unstretched DMSO, † p < 0.05 vs. stretched DMSO, Mann–Whitney U test.
Pan Pkc Inhibitor Gö6983, supplied by STEMCELL Technologies Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/pan+pkc+inhibitor+g%C3%B66983/pmc11708431-34-23-29
Average 90 stars, based on 1 article reviews
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sotrastaurin (pan-pkc inhibitor)  (Axon Medchem LLC)
90
Axon Medchem LLC sotrastaurin (pan-pkc inhibitor)
The pan-PKC inhibitor <t>sotrastaurin</t> reduces ACE2 shedding in mouse proximal tubular cells. (A) Graph depicts effect of sotrastaurin (10-5-10-8 M) on ACE2 activity recovered from cell culture media. ∗ P < 0.005 vs. all other groups (C, vehicle-treated control); ∗∗ P < 0.05 vs. C and 10 -8 M; n = 6. (B) Graph depicts effect of varying concentrations of sotrastaurin on ACE2 shedding into culture media, determined by immunoblot analysis. Representative immunoblot is depicted above graph, showing ACE2 fragments at 90 and 70 kDa, with mouse kidney cortex ACE2 at 100 kDa. ∗ P < 0.01 vs. all other groups; ∗∗ P < 0.05 vs. 10 -7 M and vehicle-treated control (C); n = 5. (C) Graph depicts effect of sotrastaurin (10 -5 M), with or without the ADAM17 inhibitor TAPI-1 (10 -5 M), on ACE2 activity in the culture media, in basal conditions ( D -glucose 7.8 mM) or high D -glucose (28 mM). ∗ P < 0.01 vs. Control and TAPI-1 (7.8 mM D -glucose); In D -glucose 28 mM, † P < 0.001 vs. Control and TAPI-1, †† P < 0.05 vs. Control; ‡ P < 0.001 vs. Control ( D -glucose 7.8 mM); n = 4–5.
Sotrastaurin (Pan Pkc Inhibitor), supplied by Axon Medchem LLC, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/sotrastaurin++pan+pkc+inhibitor+/pmc04887483-61-0-5
Average 90 stars, based on 1 article reviews
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pan-pkc activity assay  (Enzo Biochem)
90
Enzo Biochem pan-pkc activity assay
The pan-PKC inhibitor <t>sotrastaurin</t> reduces ACE2 shedding in mouse proximal tubular cells. (A) Graph depicts effect of sotrastaurin (10-5-10-8 M) on ACE2 activity recovered from cell culture media. ∗ P < 0.005 vs. all other groups (C, vehicle-treated control); ∗∗ P < 0.05 vs. C and 10 -8 M; n = 6. (B) Graph depicts effect of varying concentrations of sotrastaurin on ACE2 shedding into culture media, determined by immunoblot analysis. Representative immunoblot is depicted above graph, showing ACE2 fragments at 90 and 70 kDa, with mouse kidney cortex ACE2 at 100 kDa. ∗ P < 0.01 vs. all other groups; ∗∗ P < 0.05 vs. 10 -7 M and vehicle-treated control (C); n = 5. (C) Graph depicts effect of sotrastaurin (10 -5 M), with or without the ADAM17 inhibitor TAPI-1 (10 -5 M), on ACE2 activity in the culture media, in basal conditions ( D -glucose 7.8 mM) or high D -glucose (28 mM). ∗ P < 0.01 vs. Control and TAPI-1 (7.8 mM D -glucose); In D -glucose 28 mM, † P < 0.001 vs. Control and TAPI-1, †† P < 0.05 vs. Control; ‡ P < 0.001 vs. Control ( D -glucose 7.8 mM); n = 4–5.
Pan Pkc Activity Assay, supplied by Enzo Biochem, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/the+pan+pkc+activity+assay/pmc03574184-269-10-14
Average 90 stars, based on 1 article reviews
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monoclonal mouse anti-human pkc (pan) antibody  (Biomol GmbH)
90
Biomol GmbH monoclonal mouse anti-human pkc (pan) antibody
The pan-PKC inhibitor <t>sotrastaurin</t> reduces ACE2 shedding in mouse proximal tubular cells. (A) Graph depicts effect of sotrastaurin (10-5-10-8 M) on ACE2 activity recovered from cell culture media. ∗ P < 0.005 vs. all other groups (C, vehicle-treated control); ∗∗ P < 0.05 vs. C and 10 -8 M; n = 6. (B) Graph depicts effect of varying concentrations of sotrastaurin on ACE2 shedding into culture media, determined by immunoblot analysis. Representative immunoblot is depicted above graph, showing ACE2 fragments at 90 and 70 kDa, with mouse kidney cortex ACE2 at 100 kDa. ∗ P < 0.01 vs. all other groups; ∗∗ P < 0.05 vs. 10 -7 M and vehicle-treated control (C); n = 5. (C) Graph depicts effect of sotrastaurin (10 -5 M), with or without the ADAM17 inhibitor TAPI-1 (10 -5 M), on ACE2 activity in the culture media, in basal conditions ( D -glucose 7.8 mM) or high D -glucose (28 mM). ∗ P < 0.01 vs. Control and TAPI-1 (7.8 mM D -glucose); In D -glucose 28 mM, † P < 0.001 vs. Control and TAPI-1, †† P < 0.05 vs. Control; ‡ P < 0.001 vs. Control ( D -glucose 7.8 mM); n = 4–5.
Monoclonal Mouse Anti Human Pkc (Pan) Antibody, supplied by Biomol GmbH, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/monoclonal+mouse+anti+human+pkc++pan++antibody/10__1074_slash_jbc__m704915200-99-40-49
Average 90 stars, based on 1 article reviews
monoclonal mouse anti-human pkc (pan) antibody - by Bioz Stars, 2026-10
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pan classical pkc inhibitor  (KAI Pharmaceuticals)
90
KAI Pharmaceuticals pan classical pkc inhibitor
Confluent IEC monolayers were wounded by introduction of a single linear scratch wound, preincubated with the specified PKC inhibitors for 1 hour (Cheleretrine, pan PKC inhibitor (5 µM, pan); Calphostin C, Classical/non-conventional PKC inhibitor (10 µM, Clas/Non-C ); KIC1-1, classical PKC inhibitor (5 µM, Clas); KIBI31-1, PKCβI inhibitor (1 µM, PKCβI); KIBII31-1 (5 µM, PKCβII); KIG31-1, PKCγ inhibitor (5 µM, PKCβγ) and stimulated with PMA (200nM). The area of the wound was measured at 12 and 24 hours. Inhibition of PKCβII, but not inhibition of other members of the classical PKCs significantly diminished PMA enhanced wound closure. N = 3 independent experiments. **significantly different from control (PMA alone, p<0.01).
Pan Classical Pkc Inhibitor, supplied by KAI Pharmaceuticals, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/pan+classical+pkc+inhibitor/pmc03569445-36-13-22
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pan-pkc inhibitor sotrastaurin  (ChemScene llc)
90
ChemScene llc pan-pkc inhibitor sotrastaurin
Confluent IEC monolayers were wounded by introduction of a single linear scratch wound, preincubated with the specified PKC inhibitors for 1 hour (Cheleretrine, pan PKC inhibitor (5 µM, pan); Calphostin C, Classical/non-conventional PKC inhibitor (10 µM, Clas/Non-C ); KIC1-1, classical PKC inhibitor (5 µM, Clas); KIBI31-1, PKCβI inhibitor (1 µM, PKCβI); KIBII31-1 (5 µM, PKCβII); KIG31-1, PKCγ inhibitor (5 µM, PKCβγ) and stimulated with PMA (200nM). The area of the wound was measured at 12 and 24 hours. Inhibition of PKCβII, but not inhibition of other members of the classical PKCs significantly diminished PMA enhanced wound closure. N = 3 independent experiments. **significantly different from control (PMA alone, p<0.01).
Pan Pkc Inhibitor Sotrastaurin, supplied by ChemScene llc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/pan+pkc/pan+pkc+inhibitor+sotrastaurin/pm34987138-88-9-12
Average 90 stars, based on 1 article reviews
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Image Search Results


( A ) IP3R blocking inhibits PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with 2-APB (10 µM) for additional 18 hr. ( B ) [Ca 2+ ] blocking represses signal transduction in the axis of PKC/GSK-3β/β-Catenin induced by PLCE1. ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with BAPTA-AM (10 µM) for additional 18 hr. ( C ) PKC inhibition blocks PLCE1 stimulated signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmid expressing Plce1 . 6 hr post-transfection, cells were treated with 100 nM of Midostaurin for additional 18 hr. ( D ) PKC inhibition represses PLCE1-induced nuclear translocation of β-Catenin in ECA-109 cells. Cells were transfected with the plasmid expressing PLCE1. 6 hr post-transfection, 2-APB (10 µM), BAPTA-AM (10 µM), or Midostaurin (100 nM) was added in culture medium, and cells were cultured for additional 18 hr. Scale bar = 10 µm. Immunofluorescence was used to examine subcellular distribution of β-Catenin. ( E ) TAK1 counteracts PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated for 24 hr. ( F ) TAK1 reduces PLCE1-induced nuclear distribution of β-Catenin in ECA-109 cells. Cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated. Scale bar = 10 µm. ( G ) Dominant negative TAK1 (K63W) fails to block signal transduction in the axis of PKC/GSK-3β/β-Catenin/MMP2 induced by PLCE1. ECA-109 cells were transfected with the plasmids expressing Plce1 or mutated Map3k7 (TAK1 K63W) for 24 hr. ( H ) TAK1 has no effect on PLCE1 S1060A-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing PLCE1 S1060A or TAK1 for 24 hr. Protein levels were analyzed by western blot, and Actin was used as a loading control. Representative blots were shown. Figure 5—source data 1. PDF file containing original western blots for , indicating the relevant bands. Figure 5—source data 2. Original files for western blot analysis displayed in .

Journal: eLife

Article Title: TAK1-mediated phosphorylation of PLCE1 represses PIP2 hydrolysis to impede esophageal squamous cancer metastasis

doi: 10.7554/eLife.97373

Figure Lengend Snippet: ( A ) IP3R blocking inhibits PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with 2-APB (10 µM) for additional 18 hr. ( B ) [Ca 2+ ] blocking represses signal transduction in the axis of PKC/GSK-3β/β-Catenin induced by PLCE1. ECA-109 cells were transfected with the plasmid expressing Plce1 for 6 hr and then treated with BAPTA-AM (10 µM) for additional 18 hr. ( C ) PKC inhibition blocks PLCE1 stimulated signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmid expressing Plce1 . 6 hr post-transfection, cells were treated with 100 nM of Midostaurin for additional 18 hr. ( D ) PKC inhibition represses PLCE1-induced nuclear translocation of β-Catenin in ECA-109 cells. Cells were transfected with the plasmid expressing PLCE1. 6 hr post-transfection, 2-APB (10 µM), BAPTA-AM (10 µM), or Midostaurin (100 nM) was added in culture medium, and cells were cultured for additional 18 hr. Scale bar = 10 µm. Immunofluorescence was used to examine subcellular distribution of β-Catenin. ( E ) TAK1 counteracts PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated for 24 hr. ( F ) TAK1 reduces PLCE1-induced nuclear distribution of β-Catenin in ECA-109 cells. Cells were transfected with the plasmids expressing Plce1 or Map3k7 as indicated. Scale bar = 10 µm. ( G ) Dominant negative TAK1 (K63W) fails to block signal transduction in the axis of PKC/GSK-3β/β-Catenin/MMP2 induced by PLCE1. ECA-109 cells were transfected with the plasmids expressing Plce1 or mutated Map3k7 (TAK1 K63W) for 24 hr. ( H ) TAK1 has no effect on PLCE1 S1060A-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin. ECA-109 cells were transfected with the plasmids expressing PLCE1 S1060A or TAK1 for 24 hr. Protein levels were analyzed by western blot, and Actin was used as a loading control. Representative blots were shown. Figure 5—source data 1. PDF file containing original western blots for , indicating the relevant bands. Figure 5—source data 2. Original files for western blot analysis displayed in .

Article Snippet: Rabbit monoclonal antibodies against TAK1 (#5206), phospho-TAK1 (Ser412, #9339), PKCα (#2056), phospho-PKC (pan) (gamma Thr514) (#38938), GSK-3β (#9315), phospho-GSK-3β (Ser9) (#5558), β-Catenin (#8480), phospho-β-Catenin (Ser33/37/Thr41) (#9561), MMP-2 (#87809), mouse monoclonal antibodies against Actin (#3700), Myc-Tag (Sepharose Bead Conjugate) (#55464), Epithelial-Mesenchymal Transition (EMT) antibody sample kit (#9782), phosphor-IKK (#2078), IKK (#61294), phosphor-JNK (#4668), JNK (#9252), phospho-ERK (#4370), ERK (#9102), phospho-P38 MAPK (#9211), P38 MPAK (#9212), and normal rabbit IgG (#2729) were from Cell Signaling Technology (Beverley, MA, USA).

Techniques: Blocking Assay, Transduction, Transfection, Plasmid Preparation, Expressing, Inhibition, Translocation Assay, Cell Culture, Immunofluorescence, Dominant Negative Mutation, Western Blot, Control

( A–F ) Quantified data for the western blots as shown in . n=3 biologically independent replicates. Data are presented as mean ± SD. Statistical significance was tested by two-tailed one-way ANOVA test. *p<0.05, **p<0.01, and ***p<0.001. Figure 5—figure supplement 7—source data 1. TAK1 mitigates PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin.

Journal: eLife

Article Title: TAK1-mediated phosphorylation of PLCE1 represses PIP2 hydrolysis to impede esophageal squamous cancer metastasis

doi: 10.7554/eLife.97373

Figure Lengend Snippet: ( A–F ) Quantified data for the western blots as shown in . n=3 biologically independent replicates. Data are presented as mean ± SD. Statistical significance was tested by two-tailed one-way ANOVA test. *p<0.05, **p<0.01, and ***p<0.001. Figure 5—figure supplement 7—source data 1. TAK1 mitigates PLCE1-induced signal transduction in the axis of PKC/GSK-3β/β-Catenin.

Article Snippet: Rabbit monoclonal antibodies against TAK1 (#5206), phospho-TAK1 (Ser412, #9339), PKCα (#2056), phospho-PKC (pan) (gamma Thr514) (#38938), GSK-3β (#9315), phospho-GSK-3β (Ser9) (#5558), β-Catenin (#8480), phospho-β-Catenin (Ser33/37/Thr41) (#9561), MMP-2 (#87809), mouse monoclonal antibodies against Actin (#3700), Myc-Tag (Sepharose Bead Conjugate) (#55464), Epithelial-Mesenchymal Transition (EMT) antibody sample kit (#9782), phosphor-IKK (#2078), IKK (#61294), phosphor-JNK (#4668), JNK (#9252), phospho-ERK (#4370), ERK (#9102), phospho-P38 MAPK (#9211), P38 MPAK (#9212), and normal rabbit IgG (#2729) were from Cell Signaling Technology (Beverley, MA, USA).

Techniques: Western Blot, Two Tailed Test, Transduction

Each mouse was intravenously injected with 1×10 6 ECA-109 cells diluted in 100 µl PBS. Mice were treated with Takinib at the dosage of 50 mg/kg/day for 15 days, mice in control group were received vehicle (corn oil). Eight weeks later, mice were sacrificed, and the lungs and livers from each group were collected and photographed. ( A ) Typical images of specimens. ( B ) Hematoxylin and eosin staining of metastatic nodules in lungs. ( C ) The number of nodules in lungs. ( D ) Takinib treatment induces signal transduction in the axis of PKC/GSK-3β/β-Catenin. Protein levels were analyzed by western blot, and Actin was used as a loading control. n=3 biologically independent replicates. ( E ) Quantitative analysis of the western blot data shown in ( D ). Data are presented as mean ± SD. Statistical significance was tested by unpaired Student’s t-test. * p < 0.05 , ** p<0.01, and *** p<0.001 . Figure 6—source data 1. Inhibition of TAK1 by Takinib promotes esophageal squamous cell carcinoma (ESCC) metastasis in nude mice. Figure 6—source data 2. PDF file containing original western blots for , indicating the relevant bands. Figure 6—source data 3. Original files for western blot analysis displayed in .

Journal: eLife

Article Title: TAK1-mediated phosphorylation of PLCE1 represses PIP2 hydrolysis to impede esophageal squamous cancer metastasis

doi: 10.7554/eLife.97373

Figure Lengend Snippet: Each mouse was intravenously injected with 1×10 6 ECA-109 cells diluted in 100 µl PBS. Mice were treated with Takinib at the dosage of 50 mg/kg/day for 15 days, mice in control group were received vehicle (corn oil). Eight weeks later, mice were sacrificed, and the lungs and livers from each group were collected and photographed. ( A ) Typical images of specimens. ( B ) Hematoxylin and eosin staining of metastatic nodules in lungs. ( C ) The number of nodules in lungs. ( D ) Takinib treatment induces signal transduction in the axis of PKC/GSK-3β/β-Catenin. Protein levels were analyzed by western blot, and Actin was used as a loading control. n=3 biologically independent replicates. ( E ) Quantitative analysis of the western blot data shown in ( D ). Data are presented as mean ± SD. Statistical significance was tested by unpaired Student’s t-test. * p < 0.05 , ** p<0.01, and *** p<0.001 . Figure 6—source data 1. Inhibition of TAK1 by Takinib promotes esophageal squamous cell carcinoma (ESCC) metastasis in nude mice. Figure 6—source data 2. PDF file containing original western blots for , indicating the relevant bands. Figure 6—source data 3. Original files for western blot analysis displayed in .

Article Snippet: Rabbit monoclonal antibodies against TAK1 (#5206), phospho-TAK1 (Ser412, #9339), PKCα (#2056), phospho-PKC (pan) (gamma Thr514) (#38938), GSK-3β (#9315), phospho-GSK-3β (Ser9) (#5558), β-Catenin (#8480), phospho-β-Catenin (Ser33/37/Thr41) (#9561), MMP-2 (#87809), mouse monoclonal antibodies against Actin (#3700), Myc-Tag (Sepharose Bead Conjugate) (#55464), Epithelial-Mesenchymal Transition (EMT) antibody sample kit (#9782), phosphor-IKK (#2078), IKK (#61294), phosphor-JNK (#4668), JNK (#9252), phospho-ERK (#4370), ERK (#9102), phospho-P38 MAPK (#9211), P38 MPAK (#9212), and normal rabbit IgG (#2729) were from Cell Signaling Technology (Beverley, MA, USA).

Techniques: Injection, Control, Staining, Transduction, Western Blot, Inhibition

ECA-109 cells were transduced with lentivirus bearing PLCE1 shRNA (LV-shPLCE1) or NC shRNA (LV-shPLCE1 NC). Each mouse was intravenously injected with the LV transduced cells (1×10 6 cells/mouse). Eight weeks later, mice were sacrificed, and the lungs and livers from each group were collected and photographed. ( A ) Typical images of lung specimens. ( B ) Hematoxylin and eosin staining of metastatic nodules in lungs. ( C ) The number of nodules in lungs. ( D ) PLCE1 knockdown represses signal transduction in the axis of PKC/GSK-3β/β-Catenin. Protein levels were analyzed by western blot, and Actin was used as a loading control. n=3 biologically independent replicates. ( E ) Quantitative analysis of the western blot data shown in ( D ). Data are presented as mean ± SD. Statistical significance was tested by unpaired Student’s t-test. ** p<0.01 and *** p<0.001 . Figure 7—source data 1. PLCE1 knockdown inhibits esophageal squamous cell carcinoma (ESCC) metastasis in nude mice. Figure 7—source data 2. PDF file containing original western blots for , indicating the relevant bands. Figure 7—source data 3. Original files for western blot analysis displayed in .

Journal: eLife

Article Title: TAK1-mediated phosphorylation of PLCE1 represses PIP2 hydrolysis to impede esophageal squamous cancer metastasis

doi: 10.7554/eLife.97373

Figure Lengend Snippet: ECA-109 cells were transduced with lentivirus bearing PLCE1 shRNA (LV-shPLCE1) or NC shRNA (LV-shPLCE1 NC). Each mouse was intravenously injected with the LV transduced cells (1×10 6 cells/mouse). Eight weeks later, mice were sacrificed, and the lungs and livers from each group were collected and photographed. ( A ) Typical images of lung specimens. ( B ) Hematoxylin and eosin staining of metastatic nodules in lungs. ( C ) The number of nodules in lungs. ( D ) PLCE1 knockdown represses signal transduction in the axis of PKC/GSK-3β/β-Catenin. Protein levels were analyzed by western blot, and Actin was used as a loading control. n=3 biologically independent replicates. ( E ) Quantitative analysis of the western blot data shown in ( D ). Data are presented as mean ± SD. Statistical significance was tested by unpaired Student’s t-test. ** p<0.01 and *** p<0.001 . Figure 7—source data 1. PLCE1 knockdown inhibits esophageal squamous cell carcinoma (ESCC) metastasis in nude mice. Figure 7—source data 2. PDF file containing original western blots for , indicating the relevant bands. Figure 7—source data 3. Original files for western blot analysis displayed in .

Article Snippet: Rabbit monoclonal antibodies against TAK1 (#5206), phospho-TAK1 (Ser412, #9339), PKCα (#2056), phospho-PKC (pan) (gamma Thr514) (#38938), GSK-3β (#9315), phospho-GSK-3β (Ser9) (#5558), β-Catenin (#8480), phospho-β-Catenin (Ser33/37/Thr41) (#9561), MMP-2 (#87809), mouse monoclonal antibodies against Actin (#3700), Myc-Tag (Sepharose Bead Conjugate) (#55464), Epithelial-Mesenchymal Transition (EMT) antibody sample kit (#9782), phosphor-IKK (#2078), IKK (#61294), phosphor-JNK (#4668), JNK (#9252), phospho-ERK (#4370), ERK (#9102), phospho-P38 MAPK (#9211), P38 MPAK (#9212), and normal rabbit IgG (#2729) were from Cell Signaling Technology (Beverley, MA, USA).

Techniques: Transduction, shRNA, Injection, Staining, Knockdown, Western Blot, Control

Journal: eLife

Article Title: TAK1-mediated phosphorylation of PLCE1 represses PIP2 hydrolysis to impede esophageal squamous cancer metastasis

doi: 10.7554/eLife.97373

Figure Lengend Snippet:

Article Snippet: Rabbit monoclonal antibodies against TAK1 (#5206), phospho-TAK1 (Ser412, #9339), PKCα (#2056), phospho-PKC (pan) (gamma Thr514) (#38938), GSK-3β (#9315), phospho-GSK-3β (Ser9) (#5558), β-Catenin (#8480), phospho-β-Catenin (Ser33/37/Thr41) (#9561), MMP-2 (#87809), mouse monoclonal antibodies against Actin (#3700), Myc-Tag (Sepharose Bead Conjugate) (#55464), Epithelial-Mesenchymal Transition (EMT) antibody sample kit (#9782), phosphor-IKK (#2078), IKK (#61294), phosphor-JNK (#4668), JNK (#9252), phospho-ERK (#4370), ERK (#9102), phospho-P38 MAPK (#9211), P38 MPAK (#9212), and normal rabbit IgG (#2729) were from Cell Signaling Technology (Beverley, MA, USA).

Techniques: Transfection, Construct, shRNA, Recombinant, Plasmid Preparation, Sequencing, Activity Assay, Enzyme-linked Immunosorbent Assay, Software

Effects of p38 mitogen-activated protein kinase (p38 MAPK), mitogen-activated protein kinase kinase 1/2 (MEK1/2) and protein kinase C (PKC) inhibitors on stretch-induced hypertrophic gene expression in human pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The following inhibitors were utilized: SB203580 at 10 μM to inhibit p38 MAPK, U0126 at 10 μM to inhibit MEK1/2, Gö6983 at 1 μM to inhibit all PKC isoforms and Gö6976 at 1 μM to inhibit classical PKC isoforms. Natriuretic peptide B ( NPPB ; A), skeletal muscle alpha actin 1 ( ACTA1 ; B), galanin ( GAL ; C) and long intergenic non-protein coding RNA 648 ( LINC00648 ; D) RNA expression was measured with qRT–PCR after a 24-h cyclic mechanical stretch. The results are presented as fold change relative to the unstretched control. The data are shown as mean ± standard error of the mean, and values from individual experiments are presented as dots ( n = 5, except for SB203580 and Gö6976 n = 4; where n represents biological replicates of cells from individual differentiations). * p < 0.05, ** p < 0.01 vs. unstretched DMSO, † p < 0.05 vs. stretched DMSO, Mann–Whitney U test.

Journal: Journal of Molecular and Cellular Cardiology Plus

Article Title: Transcriptomics reveal stretched human pluripotent stem cell-derived cardiomyocytes as an advantageous hypertrophy model

doi: 10.1016/j.jmccpl.2022.100020

Figure Lengend Snippet: Effects of p38 mitogen-activated protein kinase (p38 MAPK), mitogen-activated protein kinase kinase 1/2 (MEK1/2) and protein kinase C (PKC) inhibitors on stretch-induced hypertrophic gene expression in human pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The following inhibitors were utilized: SB203580 at 10 μM to inhibit p38 MAPK, U0126 at 10 μM to inhibit MEK1/2, Gö6983 at 1 μM to inhibit all PKC isoforms and Gö6976 at 1 μM to inhibit classical PKC isoforms. Natriuretic peptide B ( NPPB ; A), skeletal muscle alpha actin 1 ( ACTA1 ; B), galanin ( GAL ; C) and long intergenic non-protein coding RNA 648 ( LINC00648 ; D) RNA expression was measured with qRT–PCR after a 24-h cyclic mechanical stretch. The results are presented as fold change relative to the unstretched control. The data are shown as mean ± standard error of the mean, and values from individual experiments are presented as dots ( n = 5, except for SB203580 and Gö6976 n = 4; where n represents biological replicates of cells from individual differentiations). * p < 0.05, ** p < 0.01 vs. unstretched DMSO, † p < 0.05 vs. stretched DMSO, Mann–Whitney U test.

Article Snippet: The mitogen-activated protein kinase kinase 1/2 (MEK1/2) inhibitor U0126 and the p38 MAPK inhibitor SB203580 were purchased from Tocris Bioscience (Bristol, UK), the pan-PKC inhibitor Gö6983 was purchased from STEMCELL Technologies (Vancouver, Canada) and the classical PKC inhibitor Gö6976 was purchased from Merck (Darmstadt, Germany).

Techniques: Gene Expression, Derivative Assay, RNA Expression, Quantitative RT-PCR, Control, MANN-WHITNEY

The pan-PKC inhibitor sotrastaurin reduces ACE2 shedding in mouse proximal tubular cells. (A) Graph depicts effect of sotrastaurin (10-5-10-8 M) on ACE2 activity recovered from cell culture media. ∗ P < 0.005 vs. all other groups (C, vehicle-treated control); ∗∗ P < 0.05 vs. C and 10 -8 M; n = 6. (B) Graph depicts effect of varying concentrations of sotrastaurin on ACE2 shedding into culture media, determined by immunoblot analysis. Representative immunoblot is depicted above graph, showing ACE2 fragments at 90 and 70 kDa, with mouse kidney cortex ACE2 at 100 kDa. ∗ P < 0.01 vs. all other groups; ∗∗ P < 0.05 vs. 10 -7 M and vehicle-treated control (C); n = 5. (C) Graph depicts effect of sotrastaurin (10 -5 M), with or without the ADAM17 inhibitor TAPI-1 (10 -5 M), on ACE2 activity in the culture media, in basal conditions ( D -glucose 7.8 mM) or high D -glucose (28 mM). ∗ P < 0.01 vs. Control and TAPI-1 (7.8 mM D -glucose); In D -glucose 28 mM, † P < 0.001 vs. Control and TAPI-1, †† P < 0.05 vs. Control; ‡ P < 0.001 vs. Control ( D -glucose 7.8 mM); n = 4–5.

Journal: Frontiers in Pharmacology

Article Title: Protein Kinase C-δ Mediates Shedding of Angiotensin-Converting Enzyme 2 from Proximal Tubular Cells

doi: 10.3389/fphar.2016.00146

Figure Lengend Snippet: The pan-PKC inhibitor sotrastaurin reduces ACE2 shedding in mouse proximal tubular cells. (A) Graph depicts effect of sotrastaurin (10-5-10-8 M) on ACE2 activity recovered from cell culture media. ∗ P < 0.005 vs. all other groups (C, vehicle-treated control); ∗∗ P < 0.05 vs. C and 10 -8 M; n = 6. (B) Graph depicts effect of varying concentrations of sotrastaurin on ACE2 shedding into culture media, determined by immunoblot analysis. Representative immunoblot is depicted above graph, showing ACE2 fragments at 90 and 70 kDa, with mouse kidney cortex ACE2 at 100 kDa. ∗ P < 0.01 vs. all other groups; ∗∗ P < 0.05 vs. 10 -7 M and vehicle-treated control (C); n = 5. (C) Graph depicts effect of sotrastaurin (10 -5 M), with or without the ADAM17 inhibitor TAPI-1 (10 -5 M), on ACE2 activity in the culture media, in basal conditions ( D -glucose 7.8 mM) or high D -glucose (28 mM). ∗ P < 0.01 vs. Control and TAPI-1 (7.8 mM D -glucose); In D -glucose 28 mM, † P < 0.001 vs. Control and TAPI-1, †† P < 0.05 vs. Control; ‡ P < 0.001 vs. Control ( D -glucose 7.8 mM); n = 4–5.

Article Snippet: Sotrastaurin (pan-PKC inhibitor) was from Axon Medchem BV (Gronigen, Netherlands).

Techniques: Activity Assay, Cell Culture, Control, Western Blot

Confluent IEC monolayers were wounded by introduction of a single linear scratch wound, preincubated with the specified PKC inhibitors for 1 hour (Cheleretrine, pan PKC inhibitor (5 µM, pan); Calphostin C, Classical/non-conventional PKC inhibitor (10 µM, Clas/Non-C ); KIC1-1, classical PKC inhibitor (5 µM, Clas); KIBI31-1, PKCβI inhibitor (1 µM, PKCβI); KIBII31-1 (5 µM, PKCβII); KIG31-1, PKCγ inhibitor (5 µM, PKCβγ) and stimulated with PMA (200nM). The area of the wound was measured at 12 and 24 hours. Inhibition of PKCβII, but not inhibition of other members of the classical PKCs significantly diminished PMA enhanced wound closure. N = 3 independent experiments. **significantly different from control (PMA alone, p<0.01).

Journal: PLoS ONE

Article Title: Activation of PKCβII by PMA Facilitates Enhanced Epithelial Wound Repair through Increased Cell Spreading and Migration

doi: 10.1371/journal.pone.0055775

Figure Lengend Snippet: Confluent IEC monolayers were wounded by introduction of a single linear scratch wound, preincubated with the specified PKC inhibitors for 1 hour (Cheleretrine, pan PKC inhibitor (5 µM, pan); Calphostin C, Classical/non-conventional PKC inhibitor (10 µM, Clas/Non-C ); KIC1-1, classical PKC inhibitor (5 µM, Clas); KIBI31-1, PKCβI inhibitor (1 µM, PKCβI); KIBII31-1 (5 µM, PKCβII); KIG31-1, PKCγ inhibitor (5 µM, PKCβγ) and stimulated with PMA (200nM). The area of the wound was measured at 12 and 24 hours. Inhibition of PKCβII, but not inhibition of other members of the classical PKCs significantly diminished PMA enhanced wound closure. N = 3 independent experiments. **significantly different from control (PMA alone, p<0.01).

Article Snippet: PKC isoform-specific peptide inhibitors for PKCβI (KIBI31-1), PKCβII (KIBII31-1), PKCγ (KIG31-1), and a pan classical PKC inhibitor (KIC1-1) were kindly provided by KAI Pharmaceuticals (San Francisco, CA).

Techniques: Inhibition, Control

Scratch wounded IEC monolayers were allowed to migrate for 4 hours in the absence (control) or presence of PMA (PMA) and PKC isoforms inhibitors. The levels of total and phospo-paxillin were determined by immunoblotting ( A ) and quantified using densitometric analysis ( B ). The data are expressed as fold change relative to control after normalization to loading control. PMA treatment increased paxillin phosphorylation, which was attenuated by the inhibition of PKCβII (PMA+βII), but not PKCβI (PMA+βI). ( C ) Cell adhesion assay was used to examine IEC-matrix adhesion after PMA treatment in the presence or absence of specific inhibitors of PKC isoforms. PMA treatment significantly increased cell-matrix adhesion.This effect was reversed with inhibition of PKCβII, but not PKCβI. *significantly different (p<0.05). ( D ) To examine formation of focal adhesions by the cells at the leading edge (upper panels), and cells away from the wound (bottom panels) IECs were fixed and stained for F-actin (red) and phospho-paxillin (green). Representative images show increased phosphorylation of paxillin both at the lamellipodium and areas remote from the wound after PKC activation with PMA. The bar is 20 µm. ( E–F ) Quantification of the changes in the relative fluorescence intensity, as an indication of the changes in paxillin phosphorylation. ( E ) Average relative fluorescence intensity of 5 randomly placed lines (as shown in (D), bottom panels) per field of view, at least 7 fields per condition. N = 3 independent experiments. **significantly different (p<0.01). ( F ) Fluorescence intensity profiles of phospho-paxillin along representative white lines projected across fields of view (as shown on the images in (D), bottom panels) that were used for quantification in (E).

Journal: PLoS ONE

Article Title: Activation of PKCβII by PMA Facilitates Enhanced Epithelial Wound Repair through Increased Cell Spreading and Migration

doi: 10.1371/journal.pone.0055775

Figure Lengend Snippet: Scratch wounded IEC monolayers were allowed to migrate for 4 hours in the absence (control) or presence of PMA (PMA) and PKC isoforms inhibitors. The levels of total and phospo-paxillin were determined by immunoblotting ( A ) and quantified using densitometric analysis ( B ). The data are expressed as fold change relative to control after normalization to loading control. PMA treatment increased paxillin phosphorylation, which was attenuated by the inhibition of PKCβII (PMA+βII), but not PKCβI (PMA+βI). ( C ) Cell adhesion assay was used to examine IEC-matrix adhesion after PMA treatment in the presence or absence of specific inhibitors of PKC isoforms. PMA treatment significantly increased cell-matrix adhesion.This effect was reversed with inhibition of PKCβII, but not PKCβI. *significantly different (p<0.05). ( D ) To examine formation of focal adhesions by the cells at the leading edge (upper panels), and cells away from the wound (bottom panels) IECs were fixed and stained for F-actin (red) and phospho-paxillin (green). Representative images show increased phosphorylation of paxillin both at the lamellipodium and areas remote from the wound after PKC activation with PMA. The bar is 20 µm. ( E–F ) Quantification of the changes in the relative fluorescence intensity, as an indication of the changes in paxillin phosphorylation. ( E ) Average relative fluorescence intensity of 5 randomly placed lines (as shown in (D), bottom panels) per field of view, at least 7 fields per condition. N = 3 independent experiments. **significantly different (p<0.01). ( F ) Fluorescence intensity profiles of phospho-paxillin along representative white lines projected across fields of view (as shown on the images in (D), bottom panels) that were used for quantification in (E).

Article Snippet: PKC isoform-specific peptide inhibitors for PKCβI (KIBI31-1), PKCβII (KIBII31-1), PKCγ (KIG31-1), and a pan classical PKC inhibitor (KIC1-1) were kindly provided by KAI Pharmaceuticals (San Francisco, CA).

Techniques: Control, Western Blot, Phospho-proteomics, Inhibition, Cell Adhesion Assay, Staining, Activation Assay, Fluorescence

( A,B ) Subconfluent IECs monolayers (∼40% confluency) were allowed to spread on 13-mm collagen-coated coverslips in the absence (left panels) or presence (right panels) of PMA activation (200nM, 4 h), with the addition of PKCβII or PKCβI inhibitors as indicated. Cell area per field of view was measured before (t = 0 h, upper panels) and after treatment (t = 4 h, bottom panels). (A) Representative images demonstrating enhanced IEC spreading after PMA treatment. The bar is 50 µm. ( B ) Quantification of IEC spreading in the presence or absence of PKCβII (PMA+βII) or PKCβI (PMA+βI) inhibitors. Data presented as percent cell area/total area of the field of view. PMA induced cell spreading was significantly attenuated after inhibition of PKCβII. **significantly different (p<0.01). ( C ) Spreading IECs stimulated with PMA were fixed and stained for F-actin at t = 0 (left panels) and t = 4 h (right panels). Representative confocal microscopy images show most apical (upper panels) and most basal (bottom panels) distribution of F-actin. A dramatic reorganization of F-actin including loss of perijunctional actomyosin belt at the apical side, and redistribution of actin towards cell borders, as well as loss of stress fibers at the cell basal side in response to PMA treatment was observed. The bar is 20 µm.

Journal: PLoS ONE

Article Title: Activation of PKCβII by PMA Facilitates Enhanced Epithelial Wound Repair through Increased Cell Spreading and Migration

doi: 10.1371/journal.pone.0055775

Figure Lengend Snippet: ( A,B ) Subconfluent IECs monolayers (∼40% confluency) were allowed to spread on 13-mm collagen-coated coverslips in the absence (left panels) or presence (right panels) of PMA activation (200nM, 4 h), with the addition of PKCβII or PKCβI inhibitors as indicated. Cell area per field of view was measured before (t = 0 h, upper panels) and after treatment (t = 4 h, bottom panels). (A) Representative images demonstrating enhanced IEC spreading after PMA treatment. The bar is 50 µm. ( B ) Quantification of IEC spreading in the presence or absence of PKCβII (PMA+βII) or PKCβI (PMA+βI) inhibitors. Data presented as percent cell area/total area of the field of view. PMA induced cell spreading was significantly attenuated after inhibition of PKCβII. **significantly different (p<0.01). ( C ) Spreading IECs stimulated with PMA were fixed and stained for F-actin at t = 0 (left panels) and t = 4 h (right panels). Representative confocal microscopy images show most apical (upper panels) and most basal (bottom panels) distribution of F-actin. A dramatic reorganization of F-actin including loss of perijunctional actomyosin belt at the apical side, and redistribution of actin towards cell borders, as well as loss of stress fibers at the cell basal side in response to PMA treatment was observed. The bar is 20 µm.

Article Snippet: PKC isoform-specific peptide inhibitors for PKCβI (KIBI31-1), PKCβII (KIBII31-1), PKCγ (KIG31-1), and a pan classical PKC inhibitor (KIC1-1) were kindly provided by KAI Pharmaceuticals (San Francisco, CA).

Techniques: Activation Assay, Inhibition, Staining, Confocal Microscopy