Additional file 1 of Activation of the EGFR-PI3K-CaM pathway by PRL-1-overexpressing placenta-derived mesenchymal stem cells ameliorates liver cirrhosis via ER stress-dependent calcium
Creators: name: Kim, Se Ho
name: Kim, Jae Yeon
name: Park, Soo Young
name: Jeong, Won Tae
name: Kim, Jin Man
name: Bae, Si Hyun
name: Kim, Gi Jin
Publisher: name: figshare
Year: 2021
Subjects: subject: Biophysics
subject: Biochemistry
subject: Medicine
subject: Cell Biology
subject: Molecular Biology
subject: Physiology
subject: FOS: Biological sciences
subject: 39999 Chemical Sciences not elsewhere classified
subject: FOS: Chemical sciences
subject: 69999 Biological Sciences not elsewhere classified
subject: Science Policy

description: Additional file 1. Supplementary Fig. 1 Optimal concentration of thapsigargin (TG) in WB-F344. a TG was treated with each concentration (100, 500, and 1000 nM) for 6, 12, 24, and 48 h in WB-F344. XBP1 splice form was transformed upon TG 24 h treatment. GAPDH was used as loading control. GAPDH, glycraldehyde-3-phosphate dehydrogenase; TG, tapsigargin. Supplementary Fig. 2 PD-MSCsPRL-1 regulated calcium channels in a rat BDL and hepatocyte treated with TG. a–d mRNA levels of calcium channels (e.g., IP3R, GRP75, VDAC1, and MCU) in a rat model with BDL. e-i mRNA levels of calcium channels (e.g., IP3R, GRP75, VDAC1, MCU, and CaM) exposed to TG (500 nM) for 24 h in WB-F344 by qRT-PCR. Data from each group are shown as the means ± SD and were assessed using Student’s t-test. *p &lt; 0.05 vs. NTx, #p &lt; 0.05 vs. PD-MSCs. BDL, bile duct ligation; CaM, calmodulin; GRP75, glucose regulated protein 75; IP3R, inositol trisphosphate receptor; MCU, mitochondria calcium uniporter; NTx, nontransplantation; PRL-1, phosphatase of regenerating liver-1; TG, thapsigargin; VDAC1, voltage dependent anion channel 1. Supplementary Fig. 3 Increased expression of SERCA2b and STIM1 in PD-MSCsPRL-1 groups in a rat model with BDL. a, b mRNA levels of ER-specific Ca2+ channel factor SERCA2b and Ca2+ sensor factor STIM1 in a rat model with BDL. Data from each group are shown as the means ± SD and were assessed using Student’s t-test. *p &lt; 0.05 vs. NTx, #p &lt; 0.05 vs. PD-MSCs. BDL, bile duct ligation; NTx, nontransplantation; SERCA2b, sarco/endoplasmic reticulum Ca2+ ATPase; PRL-1, phosphatase of regenerating liver-1; STIM1, stromal interaction molecule 1. Supplementary Fig. 4 PRL-1 regulates EGFR-PI3K-CaM calcium signaling in a BDL-injured rat liver and rat hepatocyte treated LCA. a mRNA levels of CaM and b PI3K-p85 in a rat model with BDL. c-e mRNA levels of intracellular calcium signaling (e.g., EGFR, PI3K, and CaM) induced LCA (100 μM) and treated recombinant PRL-1 (rePRL-1; 500 pg) in WB-F344. Data from each group are shown as the means ± SD and were assessed using Student’s t-test. *p &lt; 0.05 vs. NTx, #p &lt; 0.05 vs. PD-MSCs. BDL, bile duct ligation; CaM, calmodulin; EGFR, epidermal growth factor receptor; NTx, nontransplantation; PI3K, phosphatidylinositol-3-kinase. Supplementary Fig. 5 PD-MSCsPRL-1 modulated calcium channels in a rat hepatocyte treated with LCA by qRT-PCR. a-g mRNA levels related to intracellular calcium transport channels factors (e.g., SERCA2b, IP3R, VDAC1, MCU, GRP75, CaM, and STIM1) induced LCA (100 μM) and co-cultivation with naïve PD-MSCs or PD-MSCsPRL-1 in WB-F344. Data from each group are shown as the means ± SD and were assessed using Student’s t-test. *p &lt; 0.05 vs. LCA (100 μM), #p &lt; 0.05 vs. PD-MSCs. CaM, calmodulin; GRP75, glucose regulated protein 75; IP3R, inositol trisphosphate receptor; LCA, lithocholic acid; MCU, mitochondria calcium uniporter; PRL-1, phosphatase of regenerating liver-1; SERCA2b, sarco/endoplasmic reticulum Ca2+ ATPase; STIM1, stromal interaction molecule 1; VDAC1, voltage dependent anion channel 1. Supplementary Fig. 6 PD-MSCsPRL-1 regulated calcium channels in a rat hepatocyte treated with LCA and TG by western blotting. a Protein expression and b-f their intensities of intracellular calcium transport channels factors (e.g., SERCA2b, IP3R, VDAC1, MCU, and GRP75) induced LCA (100 μM) and co-cultivation with naïve PD-MSCs or PD-MSCsPRL-1 in WB-F344. g Protein expression and h-j their intensities of intracellular calcium transport channels factors (e.g., SERCA2b, and MCU) and liver regeneration factor induced LCA (100 μM) and co-cultivation with naïve PD-MSCs or PD-MSCsPRL-1 and Pentamidine (10 μg) treated in rat primary hepatocytes. GAPDH and α-tubulin was used as loading control. Data from each group are shown as the means ± SD and assessed using Student’s t-test. *p &lt; 0.05 vs. LCA (100 μM), #p &lt; 0.05 vs. PD-MSCs, $p &lt; 0.05 vs. PD-MSCs or PD-MSCsPRL-1. ALB, Albumin; GAPDH, glycraldehyde-3-phosphate dehydrogenase; GRP75, glucose regulated protein 75; IP3R, inositol trisphosphate receptor; LCA, lithocholic acid; MCU, mitochondria calcium uniporter; PRL-1, phosphatase of regenerating liver-1; SERCA2b, sarco/endoplasmic reticulum Ca2+ ATPase; TG, Thapsigargin; VDAC1, voltage dependent anion channel 1. Supplementary Fig. 7 Recombinant PRL-1 increased calcium channels in a rat hepatocyte treated with LCA. a-g mRNA levels of intracellular calcium transport channels factors (e.g., SERCA2b, IP3R, VDAC1, MCU, GRP75, CaM, and STIM1) induced LCA (100 μM) and treated recombinant PRL-1 (rePRL-1; 500 pg) in WB-F344. Data from each group are shown as the means ± SD and assessed using Student’s t-test. *p &lt; 0.05 vs. LCA (100 μM). CaM, calmodulin; GRP75, glucose regulated protein 75; IP3R, inositol trisphosphate receptor; LCA, lithocholic acid; MCU, mitochondria calcium uniporter; PRL-1, phosphatase of regenerating liver-1; SERCA2b, sarco/endoplasmic reticulum Ca2+ ATPase; VDAC1, voltage dependent anion channel 1.