Journal: Experimental Animals

Article Title: Phosphoglycerate kinase (PGK) 1 succinylation modulates epileptic seizures and the blood-brain barrier

doi: 10.1538/expanim.23-0019

Figure Lengend Snippet: Decreased succinylation of phosphoglycerate kinase (PGK) 1 protein expression levels but not PGK1 is decreased in a rat model of lithium-pilocarpine-induced acute epilepsy. (a) Representative Immunoprecipitation showed significantly decreased succinylation of PGK1 protein expression levels in the hippocampus of the rat model compared with normals (n=3 per group). (b) PGK1 expression levels were not significantly changed in the hippocampus of the rat model compared with normal rats (n=4 per group). All the succinylation of PGK1 protein expression levels were normalized by calculating the OD ratio of succinylation-PGK1 to PGK1 (Ksucc-PGK1/PGK1). All expression levels of PGK1 were normalized by calculating the OD ratio of PGK1 to β-tubulin (PGK1/β-tubulin) (* P <0.05).

Article Snippet: Then membranes were incubated with rabbit anti-PGK1 (1:1,000, ProteinTech Group, Chicago, IL, USA, GTX107614), mouse anti-PGK1 (2 µ g per 100–500 µ g of total protein, Santa Cruz Biotechnology, Santa Cruz, CA, USA, sc-130335), rabbit anti-angiostatin (1:1,000, Beijing Biosynthesis Biotech Co., Ltd., Beijing, China; bs-1828R), rabbit anti-succinyllysine (1:1,000, Protein Post-translational Modification Biolab, Hangzhou, China, PTM-401), mouse anti-Flag (1:1,000, ZEN BIO, Chengdu, China, 390002), mouse anti-β-Tubulin (1:5,000, ZEN BIO, 200608), rabbit anti-VEGF (1:1,000, Wanleibio, Shenyang, China, WL000096) antibodies overnight at 4°C.

Techniques: Expressing, Immunoprecipitation

Journal: Experimental Animals

Article Title: Phosphoglycerate kinase (PGK) 1 succinylation modulates epileptic seizures and the blood-brain barrier

doi: 10.1538/expanim.23-0019

Figure Lengend Snippet: The succinylation of phosphoglycerate kinase (PGK) 1 protein expression levels is regulated at K15 in vivo and in vitro . (A) Representative Immunoprecipitation shows that the succinylation of PGK1 protein expression levels was decreased significantly in the K15R variants and increased significantly in the K15E variants compared with the wild-type group by transfecting plasmid in HEK293 cells (n=3 per group). (B) Representative Western blots show that the expression of PGK1 total protein was insignificant in the groups of the K15R variants and the K15E variants compared with the wild-type group by transfecting plasmid in HEK293 cells (n=4 per group). (C) Representative Immunoprecipitation shows that the succinylation of PGK1 protein expression levels decreased significantly in the K15R variants and increased significantly in the K15E variants compared with the scramble group by transfecting with the lentivirus and a rat model of lithium-pilocarpine-induced acute epilepsy (n=3 per group). (D) Representative Western blots show that the expression of PGK1 total protein was insignificant in the groups of the K15R variants and the K15E variants compared with the scramble group by transfecting with the lentivirus and a rat model of lithium-pilocarpine-induced acute epilepsy (n=4 per group). All expression levels of the succinylation of PGK1 protein expression levels were normalized by calculating the OD ratio of the succinylation of PGK1 to PGK1 (Ksucc-PGK1/PGK1). All expression levels of PGK1 were normalized by calculating the OD ratio of PGK1 to β-tubulin (PGK1/β-tubulin)(* P <0.05).

Article Snippet: Then membranes were incubated with rabbit anti-PGK1 (1:1,000, ProteinTech Group, Chicago, IL, USA, GTX107614), mouse anti-PGK1 (2 µ g per 100–500 µ g of total protein, Santa Cruz Biotechnology, Santa Cruz, CA, USA, sc-130335), rabbit anti-angiostatin (1:1,000, Beijing Biosynthesis Biotech Co., Ltd., Beijing, China; bs-1828R), rabbit anti-succinyllysine (1:1,000, Protein Post-translational Modification Biolab, Hangzhou, China, PTM-401), mouse anti-Flag (1:1,000, ZEN BIO, Chengdu, China, 390002), mouse anti-β-Tubulin (1:5,000, ZEN BIO, 200608), rabbit anti-VEGF (1:1,000, Wanleibio, Shenyang, China, WL000096) antibodies overnight at 4°C.

Techniques: Expressing, In Vivo, In Vitro, Immunoprecipitation, Plasmid Preparation, Western Blot

Journal: Experimental Animals

Article Title: Phosphoglycerate kinase (PGK) 1 succinylation modulates epileptic seizures and the blood-brain barrier

doi: 10.1538/expanim.23-0019

Figure Lengend Snippet: The succinylation of phosphoglycerate kinase (PGK) 1 at K15 is involved in the expression of angiostatin. (a) Representative Western blots show that the expression of angiostatin total protein was decreased significantly in a rat model of lithium-pilocarpine-induced acute epilepsy compared with the normal group (n=4 per group; * P <0.05). (b) Representative Western blots show that the expression of angiostatin total protein was decreased significantly in the K15R variants and increased significantly in the K15E variants compared with the scramble group by transfecting with the lentivirus and a rat model of lithium-pilocarpine-induced acute epilepsy (n=3 per group; * P <0.05). All expression levels of angiostatin were normalized by calculating the OD ratio of angiostatin to β-tubulin (PGK1/β-tubulin).

Article Snippet: Then membranes were incubated with rabbit anti-PGK1 (1:1,000, ProteinTech Group, Chicago, IL, USA, GTX107614), mouse anti-PGK1 (2 µ g per 100–500 µ g of total protein, Santa Cruz Biotechnology, Santa Cruz, CA, USA, sc-130335), rabbit anti-angiostatin (1:1,000, Beijing Biosynthesis Biotech Co., Ltd., Beijing, China; bs-1828R), rabbit anti-succinyllysine (1:1,000, Protein Post-translational Modification Biolab, Hangzhou, China, PTM-401), mouse anti-Flag (1:1,000, ZEN BIO, Chengdu, China, 390002), mouse anti-β-Tubulin (1:5,000, ZEN BIO, 200608), rabbit anti-VEGF (1:1,000, Wanleibio, Shenyang, China, WL000096) antibodies overnight at 4°C.

Techniques: Expressing, Western Blot

Journal: Experimental Animals

Article Title: Phosphoglycerate kinase (PGK) 1 succinylation modulates epileptic seizures and the blood-brain barrier

doi: 10.1538/expanim.23-0019

Figure Lengend Snippet: The succinylation of phosphoglycerate kinase (PGK) 1 at K15 mediates vascular endothelial growth factor (VEGF) and the blood-brain barrier integrity in epilepsy. (a) Representative Western blots show that the expression of VEGF total protein was increased significantly in a rat model of lithium-pilocarpine-induced acute epilepsy compared with the normal group (n=4 per group; * P <0.05). (b) Representative Western blots show that the expression of VEGF total protein was increased significantly in the K15R variants and decreased significantly in the K15E variants compared with the scramble group by transfecting with the lentivirus and a rat model of lithium-pilocarpine-induced acute epilepsy (n=4 per group; * P <0.05). (c) ELISA data showed that the level of S100β in serum was increased significantly in a rat model of lithium-pilocarpine-induced acute epilepsy compared with the normal group. ELISA showed that the level of S100β in serum was increased significantly in the K15R variants and decreased significantly in the K15E variants compared with the scramble group by transfecting with the lentivirus and a rat model of lithium-pilocarpine-induced acute epilepsy (n=4 per group; * P <0.05, *** P <0.01). All expression levels of VEGF were normalized by calculating the OD ratio of VEGF to β-tubulin (VEGF/β-tubulin).

Article Snippet: Then membranes were incubated with rabbit anti-PGK1 (1:1,000, ProteinTech Group, Chicago, IL, USA, GTX107614), mouse anti-PGK1 (2 µ g per 100–500 µ g of total protein, Santa Cruz Biotechnology, Santa Cruz, CA, USA, sc-130335), rabbit anti-angiostatin (1:1,000, Beijing Biosynthesis Biotech Co., Ltd., Beijing, China; bs-1828R), rabbit anti-succinyllysine (1:1,000, Protein Post-translational Modification Biolab, Hangzhou, China, PTM-401), mouse anti-Flag (1:1,000, ZEN BIO, Chengdu, China, 390002), mouse anti-β-Tubulin (1:5,000, ZEN BIO, 200608), rabbit anti-VEGF (1:1,000, Wanleibio, Shenyang, China, WL000096) antibodies overnight at 4°C.

Techniques: Western Blot, Expressing, Enzyme-linked Immunosorbent Assay