abs against prp  (Boster Bio)

Journal: International Journal of Molecular Sciences

Article Title: TUDCA-Treated Mesenchymal Stem Cells Protect against ER Stress in the Hippocampus of a Murine Chronic Kidney Disease Model

doi: 10.3390/ijms20030613

Figure Lengend Snippet: Uremic toxin-induced apoptosis in SH-SY5Y cells through induction of reactive oxygen species (ROS)-mediated endoplasmic reticulum (ER) stress. ( A ) Flow cytometry analysis for dihydroethidium (DHE) in SH-SY5Y cells after treatment with P -cresol (500 μM) for 48 h and H 2 O 2 (200 μM) for 4 h (n = 5). The filled and clear histograms represent the cells in the absence and presence of DHE, respectively. ( B ) Quantification of the percentage of DHE positive cells. ( C ) Western blot analysis for GRP78, phosphorylation of protein kinase R (PKR)-like endoplasmic reticulum kinase (p-PERK), PERK, phosphorylation of inositol-requiring enzyme 1 α (p-IRE1α), IRE1α, and activating transcription factor 4 (ATF4) in SH-SY5Y cells after treatment with P -cresol and H 2 O 2 ( n = 3). ( D ) The protein levels of ( C ) were determined by densitometry relative to β-actin. ( E ) Flow cytometry analysis for PI/Annexin staining in SH-SY5Y cells after treatment with P -cresol and H 2 O 2 ( n = 5). ( F ) Quantification of the percentage of Annexin V positive cells. ( G ) The concentration of PrP C in SH-SY5Y cells after treatment with P -cresol and H 2 O 2 , as assessed by ELISA (n = 5). Statistical analysis: Values represent the mean ± standard error of the mean (SEM). ( B ) ** p < 0.01 vs. control. ( D ) ** p < 0.01 vs. control. ( F ) ** p < 0.01 vs. control.

Article Snippet: After blocking membranes with 5% skim milk for 1 h, glucose-regulated protein 78 (GRP78) primary antibodies against PrP C , protein kinase-like endoplasmic reticulum kinase (PERK), phospho-PERK (p-PERK), inositol-requiring enzyme 1α (IRE1α), p-IRE1α, activating transcription factor 4 (ATF4), and β-actin (Santa Cruz Biotechnology, Dallas, TX, USA) were incubated overnight.

Techniques: Flow Cytometry, Western Blot, Phospho-proteomics, Staining, Concentration Assay, Enzyme-linked Immunosorbent Assay, Control

Journal: International Journal of Molecular Sciences

Article Title: TUDCA-Treated Mesenchymal Stem Cells Protect against ER Stress in the Hippocampus of a Murine Chronic Kidney Disease Model

doi: 10.3390/ijms20030613

Figure Lengend Snippet: Co-culture with TUDCA-treated CKD-hMSCs prevents apoptosis of SH-SY5Y cells in the presence of uremic toxin through protection of ER stress. In the presence of P -cresol, SH-SY5Y cells were co-cultured with hMSCs and subsequently analyzed by flow cytometry analysis and western blot. ( A ) Flow cytometry analysis for DHE staining in SH-SY5Y cells after co-culture with hMSCs ( n = 5). The filled and clear histograms represent cells in the absence and presence of DHE, respectively. ( B ) Quantification of the percentage of DHE positive cells from ( A ). ( C ) Western blot analysis for GRP78, p-PERK, PERK, p-IRE1α, IRE1α, and ATF4 in SH-SY5Y cells after co-culture with hMSCs ( n = 3). ( D ) The protein levels of ( C ) were determined by densitometry relative to β-actin. ( E ) Flow cytometry analysis following PI/Annexin V staining of SH-SY5Y cells co-cultured with hMSCs ( n = 5). ( F ) Quantification of the percentage of Annexin V positive cells from ( E ). Statistical analysis: Values represent the mean ± SEM. ( B ) ** p < 0.01 vs. co-culture with normal hMSCs, ## p < 0.01 vs. co-culture with CKD-hMSCs, $$ p < 0.01 vs. co-culture with CKD-hMSCs + si- PRNP + TUDCA. ( D ) * p < 0.05, ** p < 0.01 vs. co-culture with normal hMSCs, ## p < 0.01 vs. co-culture with CKD-hMSCs, $$ p < 0.01 vs. co-culture with CKD-hMSCs + si- PRNP + TUDCA. ( F ) ** p < 0.01 vs. co-culture with normal hMSCs, ## p < 0.01 vs. co-culture with CKD-hMSCs, $$ p < 0.01 vs. co-culture with CKD-hMSCs + si- PRNP + TUDCA.

Article Snippet: After blocking membranes with 5% skim milk for 1 h, glucose-regulated protein 78 (GRP78) primary antibodies against PrP C , protein kinase-like endoplasmic reticulum kinase (PERK), phospho-PERK (p-PERK), inositol-requiring enzyme 1α (IRE1α), p-IRE1α, activating transcription factor 4 (ATF4), and β-actin (Santa Cruz Biotechnology, Dallas, TX, USA) were incubated overnight.

Techniques: Co-Culture Assay, Cell Culture, Flow Cytometry, Western Blot, Staining

Journal: International Journal of Molecular Sciences

Article Title: TUDCA-Treated Mesenchymal Stem Cells Protect against ER Stress in the Hippocampus of a Murine Chronic Kidney Disease Model

doi: 10.3390/ijms20030613

Figure Lengend Snippet: Co-culture of SH-SY5Y cells with TUDCA-stimulated CKD-hMSCs increases the activity of anti-oxidant enzymes via upregulation of PrP C . ( A ) In healthy mice ( n = 3) or murine CKD model ( n = 3), hematoxylin and eosin (H and E; upper images) staining and dihydroethidium (DHE, red; cropped images) staining were performed on hippocampus sections. ( B ) Immunofluorescence staining for glucose-regulated protein 78 (GRP78) in the hippocampus of a CKD mouse ( n = 3). Scale bars = 100 μm. ( C ) Western blot analysis for GRP78 in the hippocampus of a healthy mouse and a CKD mouse ( n = 3). The protein levels were determined by densitometry relative to α-tubulin. ( D ) DHE staining (red) the hippocampus of a murine CKD model was injected with either phosphate buffer saline (PBS) or each type of hMSCs. For each mouse ( n = 3), the hippocampus was isolated and analyzed 25 days after the first injection. ( E ) Immunofluorescence staining for GRP78 in the hippocampus of a murine CKD model was injected with either PBS or each type of hMSCs ( n = 3). Scale bar =100 μm. ( F ) Western blot analysis for GRP78 in the hippocampus of a murine CKD model ( n = 3). The protein levels were determined by densitometry relative to α-tubulin. Statistical analysis: Values represent the mean ± SEM. (C) ** p < 0.01 vs. healthy control. ( F ) * p < 0.05, ** p < 0.01 vs. PBS, # p < 0.05, ## p < 0.01 vs. normal hMSCs, $$ p < 0.01 vs. CKD-hMSCs, && p < 0.01 vs. CKD-hMSCs + TUDCA + si- PRNP .

Article Snippet: After blocking membranes with 5% skim milk for 1 h, glucose-regulated protein 78 (GRP78) primary antibodies against PrP C , protein kinase-like endoplasmic reticulum kinase (PERK), phospho-PERK (p-PERK), inositol-requiring enzyme 1α (IRE1α), p-IRE1α, activating transcription factor 4 (ATF4), and β-actin (Santa Cruz Biotechnology, Dallas, TX, USA) were incubated overnight.

Techniques: Co-Culture Assay, Activity Assay, Staining, Immunofluorescence, Western Blot, Injection, Saline, Isolation, Control