Journal: Brain research bulletin

Article Title: MiR-26b-5p-modified hUB-MSCs derived exosomes attenuate early brain injury during subarachnoid hemorrhage via MAT2A-mediated the p38 MAPK/STAT3 signaling pathway.

doi: 10.1016/j.brainresbull.2021.07.014

Figure Lengend Snippet: Fig. 4. MAT2A was a direct target of miR- 26b-5p. A. StarBase3.0 (http://starbase.sysu.edu.cn/) was used to predicted the targeting sites of miR- 26b-5p and MAT2A. The wild-type and mutant sequences of MAT2A are shown in the figure. B. The relative luciferase activity was tested with wild-type and mutant-type of MAT2A, respec tively. C. The miR-26b-5p mimic or miR-26b-5p inhibitor was transfected into cells and the protein expression of MAT2A was analyzed by Western blotting. GAPDH was used as an in ternal reference. N = 6, *P < 0.01.

Article Snippet: Then, equal amounts of proteins (15 μg/lane) were separated on a 12 % sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to polyvinylidenedifluoride (PVDF) membranes (Bio-Rad, Hercules, CA, USA).The membranes were blocked in 5% (w/v) nonfat dry milk in TBST (Tris-buffered saline-0.1 % Tween) at 25 ◦C for 3 h and then incubated with the following primary antibodies: rabbit polyclonal anti-GAPDH antibody (1:1000, Abcam, ab8245), rabbit monoclonal anti-CD63 antibody (1:1500, Abcam, ab217345), mouse monoclonal anti-CD81 antibody (1:1200, Abcam, ab79559), rabbit monoclonal anti-TSG101 antibody (1:2500, Abcam, ab125011), rabbit monoclonal anti-Alix antibody (1:2000, Abcam, ab186492), Rabbit polyclonal anti-MAT2A antibody (1:2500, Abcam, ab154343), Rabbit monoclonal anti-COX-2 antibody (1:2000, Abcam, ab15191), Rabbit monoclonal anti-MCP-1 antibody (1:1000, Abcam, ab214819), Rabbit monoclonal anti-iNOS antibody (1:800, Abcam, ab178945), rabbit monoclonal anti-STAT3 antibody (1:1500, Abcam, ab68153), rabbit monoclonal anti-AKT (phospho Y705) antibody (1:5000, Abcam, ab76315).

Techniques: Mutagenesis, Luciferase, Activity Assay, Transfection, Expressing, Western Blot

Journal: Brain research bulletin

Article Title: MiR-26b-5p-modified hUB-MSCs derived exosomes attenuate early brain injury during subarachnoid hemorrhage via MAT2A-mediated the p38 MAPK/STAT3 signaling pathway.

doi: 10.1016/j.brainresbull.2021.07.014

Figure Lengend Snippet: Fig. 5. Effect of regulating MAT2A expression on oxyHb-induced PC12 cells. PC12 cells were stimulated with 50 ng/mL oxyHb (cell density was 1 × 106) for 6 h. The cells were then incubated with exosomes at a concentration of 100 μg/mL, and the overexpression vector of MAT2A was transfected into the cells. A. The protein expression of MAT2A was analyzed by Western blotting. B. MTT assay was used to detected cell proliferation. C. Apoptosis of PC12 cells were detected by flow cytometry. D. Western botting was used to detected the protein expression of COX-2, MCP-1 and iNOS. GAPDH was used as an invariant internal control for calculating protein-fold changes. N = 6, *P < 0.01.

Article Snippet: Then, equal amounts of proteins (15 μg/lane) were separated on a 12 % sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to polyvinylidenedifluoride (PVDF) membranes (Bio-Rad, Hercules, CA, USA).The membranes were blocked in 5% (w/v) nonfat dry milk in TBST (Tris-buffered saline-0.1 % Tween) at 25 ◦C for 3 h and then incubated with the following primary antibodies: rabbit polyclonal anti-GAPDH antibody (1:1000, Abcam, ab8245), rabbit monoclonal anti-CD63 antibody (1:1500, Abcam, ab217345), mouse monoclonal anti-CD81 antibody (1:1200, Abcam, ab79559), rabbit monoclonal anti-TSG101 antibody (1:2500, Abcam, ab125011), rabbit monoclonal anti-Alix antibody (1:2000, Abcam, ab186492), Rabbit polyclonal anti-MAT2A antibody (1:2500, Abcam, ab154343), Rabbit monoclonal anti-COX-2 antibody (1:2000, Abcam, ab15191), Rabbit monoclonal anti-MCP-1 antibody (1:1000, Abcam, ab214819), Rabbit monoclonal anti-iNOS antibody (1:800, Abcam, ab178945), rabbit monoclonal anti-STAT3 antibody (1:1500, Abcam, ab68153), rabbit monoclonal anti-AKT (phospho Y705) antibody (1:5000, Abcam, ab76315).

Techniques: Expressing, Incubation, Concentration Assay, Over Expression, Plasmid Preparation, Transfection, Western Blot, MTT Assay, Flow Cytometry, Control

Journal: Brain research bulletin

Article Title: MiR-26b-5p-modified hUB-MSCs derived exosomes attenuate early brain injury during subarachnoid hemorrhage via MAT2A-mediated the p38 MAPK/STAT3 signaling pathway.

doi: 10.1016/j.brainresbull.2021.07.014

Figure Lengend Snippet: Fig. 6. MiR-26b-5p-modified exosomes inhibited p38 MAPK/STAT3 signaling pathway to alleviate oxyHb-induced cell injury. PC12 cells were stimulated with 50 ng/mL oxyHb (cell density was 1 × 106) for 6 h and incubated with exosomes at a concentration of 100 μg/mL. Then, cells were transfected with miR-26b-5p inhibitor alone or together with si-MAT2A, respectively. A. Western botting was used to detect the protein expression of MAT2A as well as the phosphorylation levels of p38 MAPK and STAT3 proteins. B. MTT assay was used to detected cell proliferation. C. The protein expression of COX-2, MCP-1 and iNOS was analyzed by Western botting. GAPDH was used as an internal reference. N = 6, *P < 0.01.

Article Snippet: Then, equal amounts of proteins (15 μg/lane) were separated on a 12 % sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to polyvinylidenedifluoride (PVDF) membranes (Bio-Rad, Hercules, CA, USA).The membranes were blocked in 5% (w/v) nonfat dry milk in TBST (Tris-buffered saline-0.1 % Tween) at 25 ◦C for 3 h and then incubated with the following primary antibodies: rabbit polyclonal anti-GAPDH antibody (1:1000, Abcam, ab8245), rabbit monoclonal anti-CD63 antibody (1:1500, Abcam, ab217345), mouse monoclonal anti-CD81 antibody (1:1200, Abcam, ab79559), rabbit monoclonal anti-TSG101 antibody (1:2500, Abcam, ab125011), rabbit monoclonal anti-Alix antibody (1:2000, Abcam, ab186492), Rabbit polyclonal anti-MAT2A antibody (1:2500, Abcam, ab154343), Rabbit monoclonal anti-COX-2 antibody (1:2000, Abcam, ab15191), Rabbit monoclonal anti-MCP-1 antibody (1:1000, Abcam, ab214819), Rabbit monoclonal anti-iNOS antibody (1:800, Abcam, ab178945), rabbit monoclonal anti-STAT3 antibody (1:1500, Abcam, ab68153), rabbit monoclonal anti-AKT (phospho Y705) antibody (1:5000, Abcam, ab76315).

Techniques: Modification, Incubation, Concentration Assay, Transfection, Western Blot, Expressing, Phospho-proteomics, MTT Assay

Journal: Brain research bulletin

Article Title: MiR-26b-5p-modified hUB-MSCs derived exosomes attenuate early brain injury during subarachnoid hemorrhage via MAT2A-mediated the p38 MAPK/STAT3 signaling pathway.

doi: 10.1016/j.brainresbull.2021.07.014

Figure Lengend Snippet: Fig. 7. MiR-26b-5p-modified exosomes attenuate early brain injury in SAH rats. The animal model of SAH was established by the classical occipital cistern secondary injection method. Thirty SD rats were divided into five groups on average. The cerebellar medullary cistern of rats in sham operation group was injected with normal saline. A. Relative expression of miR-26b-5p was detected by RT-qPCR. B. Western botting was used to detect the protein expression of MAT2A as well as the phosphorylation levels of p38 MAPK and STAT3 proteins. C. Behavioral and neurological functions of SAH rats were evaluated 24 h after modeling. D. Brain edema content in SAH rats was detected 24 h after modeling. E. The protein expression of COX-2, MCP-1 and iNOS was detected by Western botting. GAPDH was used as an invariant internal control for calculating protein-fold changes. N = 6, *P < 0.01.

Article Snippet: Then, equal amounts of proteins (15 μg/lane) were separated on a 12 % sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to polyvinylidenedifluoride (PVDF) membranes (Bio-Rad, Hercules, CA, USA).The membranes were blocked in 5% (w/v) nonfat dry milk in TBST (Tris-buffered saline-0.1 % Tween) at 25 ◦C for 3 h and then incubated with the following primary antibodies: rabbit polyclonal anti-GAPDH antibody (1:1000, Abcam, ab8245), rabbit monoclonal anti-CD63 antibody (1:1500, Abcam, ab217345), mouse monoclonal anti-CD81 antibody (1:1200, Abcam, ab79559), rabbit monoclonal anti-TSG101 antibody (1:2500, Abcam, ab125011), rabbit monoclonal anti-Alix antibody (1:2000, Abcam, ab186492), Rabbit polyclonal anti-MAT2A antibody (1:2500, Abcam, ab154343), Rabbit monoclonal anti-COX-2 antibody (1:2000, Abcam, ab15191), Rabbit monoclonal anti-MCP-1 antibody (1:1000, Abcam, ab214819), Rabbit monoclonal anti-iNOS antibody (1:800, Abcam, ab178945), rabbit monoclonal anti-STAT3 antibody (1:1500, Abcam, ab68153), rabbit monoclonal anti-AKT (phospho Y705) antibody (1:5000, Abcam, ab76315).

Techniques: Modification, Animal Model, Injection, Saline, Expressing, Quantitative RT-PCR, Western Blot, Phospho-proteomics, Control

Journal: Cell metabolism

Article Title: Transsulfuration activity can support cell growth upon extracellular cysteine limitation

doi: 10.1016/j.cmet.2019.09.009

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit polyclonal anti-MAT2A , Bethyl Laboratories , Cat# A304-279A-M; RRID: AB_2620475.

Techniques: Virus, Recombinant, SYBR Green Assay, Plasmid Preparation, Software, CRISPR

Journal: BMC Cancer

Article Title: Expression of methionine adenosyltransferase 2A in renal cell carcinomas and potential mechanism for kidney carcinogenesis

doi: 10.1186/1471-2407-14-196

Figure Lengend Snippet: The mRNA level analysis of the MAT2A in RCC patients and cell lines. Total RNA of 24 RCC patients and 4 RCC cell lines was extracted and reverse transcripted to cDNA. Then, real-time qRT-PCR was carried out to determine the mRNA expression levels of MAT2A. A . Relative mRNA expression level of MAT2A in RCC cancer tissues and paired normal tissues of 24 RCC patients. B . Relative mRNA expression level of MAT2A was lower in RCC cancer tissues (C) than in paired normal tissues (N) (n = 24; P < 0.05). C . Relative mRNA expression level of MAT2A was lower in 4 RCC cell lines than in HEK293.

Article Snippet: The primary antibodies used included rabbit polyclonal antibodies to MAT2A (1:1000, Abcam, Hong Kong, China), HO-1(1:200, Santa Cruz, Shanghai, China) and β-actin (1:5,000, Abcam, Hong Kong, China).

Techniques: Quantitative RT-PCR, Expressing

Journal: BMC Cancer

Article Title: Expression of methionine adenosyltransferase 2A in renal cell carcinomas and potential mechanism for kidney carcinogenesis

doi: 10.1186/1471-2407-14-196

Figure Lengend Snippet: The protein expression level of MTA2A in RCC patients. A-D Immunohistochemical analysis of MAT2A expression. MAT2A protein content was obviously lower in cancer tissues ( C and D ) than in normal tissues ( A and B ). Magnifications × 200 ( A and C ) and × 400 ( B and D ). E Level of MAT2A protein was lower in RCC cancer samples (C) than in paired normal tissuesamples (N) (n = 55, P < 0.001). The MAT2A protein were semiquantitatively scored according to staining intensity and percentage in immunohistochemical analysis of cancer or adjacent tissues. F Western blotting analysis of MAT2A. The protein expression level of MAT2A was lower in RCC cancer tissues (C) than in paired normal tissues (N).

Article Snippet: The primary antibodies used included rabbit polyclonal antibodies to MAT2A (1:1000, Abcam, Hong Kong, China), HO-1(1:200, Santa Cruz, Shanghai, China) and β-actin (1:5,000, Abcam, Hong Kong, China).

Techniques: Expressing, Immunohistochemical staining, Staining, Western Blot

Journal: BMC Cancer

Article Title: Expression of methionine adenosyltransferase 2A in renal cell carcinomas and potential mechanism for kidney carcinogenesis

doi: 10.1186/1471-2407-14-196

Figure Lengend Snippet: The negative correlation between MAT2A and HO-1 expression. mRNA levels of HO-1 and COX-2 were analyzed with real-time qRT-PCR. The correlation analysis was performed between MAT2A and HO-1 as well as COX-2 in RCC patients. A and B Relative mRNA expression levels of HO-1 (A) and COX-2 (B) were higher in RCC cancer tissues (C) than in paired normal tissues (N) (n = 24; P < 0.05). C Relative mRNA expression level of HO-1 and COX-2 were higher in 4 RCC cell lines than in HEK293. D The western blotting analysis of MAT2A and HO-1 in cell lines.The correlation of protein content between MAT2A and HO-1 is negative. E The statistical analysis reveals a negative correlation between MAT2A and HO-1 expression in RCC patients (P < 0.01). F The statistical analysis reveals no significant correlation between MAT2A and COX-2 expression in RCC patients.

Article Snippet: The primary antibodies used included rabbit polyclonal antibodies to MAT2A (1:1000, Abcam, Hong Kong, China), HO-1(1:200, Santa Cruz, Shanghai, China) and β-actin (1:5,000, Abcam, Hong Kong, China).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: BMC Cancer

Article Title: Expression of methionine adenosyltransferase 2A in renal cell carcinomas and potential mechanism for kidney carcinogenesis

doi: 10.1186/1471-2407-14-196

Figure Lengend Snippet: The proposed model of MAT2A role on RCC development. The lower content of MAT2A level reduces the product of S-adenosylmethionine (SAMe) and then decreases the level of methylation, which leads to the reactivation of HO-1 expression to increase the cell proliferation and inhibit cell apoptosis.

Article Snippet: The primary antibodies used included rabbit polyclonal antibodies to MAT2A (1:1000, Abcam, Hong Kong, China), HO-1(1:200, Santa Cruz, Shanghai, China) and β-actin (1:5,000, Abcam, Hong Kong, China).

Techniques: Methylation, Expressing