Journal: Antioxidants

Article Title: The Antioxidant Enzyme Methionine Sulfoxide Reductase A (MsrA) Interacts with Jab1/CSN5 and Regulates Its Function

doi: 10.3390/antiox9050452

Figure Lengend Snippet: Post-translational modification of proteins by neddylation. The ubiquitin-like protein Nedd8 is covalently bound to substrate proteins by a cascade of E1 activating, E2 conjugating and E3 ligase enzymes through a process termed neddylation. Deneddylation regulates this process and is catalyzed by the c-Jun activation domain-binding protein-1 (Jab1)/Csn5 subunit active site of the COP9-signalosome, which cleaves the isopeptide linkage to release neural precursor cell expressed developmentally down-regulated 8 (Nedd8) from the substrate protein. ~, thioester intermediate. —, isopeptide linkage.

Article Snippet: Antibodies used included: anti-MsrA antibody (Proteintech Group, Rosemont, IL, USA), anti-Jab1 antibody (Thermo-Fisher Scientific, Waltham, MA, USA), anti-Cul-1 antibody (Novus, Littleton, CO, USA), anti-Nedd8 antibody (Boster Bio, Pleasanton, CA, USA), anti-P27 antibody (Proteintech Group, Rosemont, IL, USA), anti-β actin antibody (Abcam, Cambridge, UK), and HRP-conjugated secondary antibodies for Western blot analyses (Rabbit anti-mouse and Goat anti-rabbit) (Bio-Rad, Hercules, CA, USA).

Techniques: Modification, Ubiquitin Proteomics, Activation Assay, Binding Assay

Journal: Antioxidants

Article Title: The Antioxidant Enzyme Methionine Sulfoxide Reductase A (MsrA) Interacts with Jab1/CSN5 and Regulates Its Function

doi: 10.3390/antiox9050452

Figure Lengend Snippet: In vivo regulation of Jab1 activity by MsrA through monitoring Cul-1 neddylation levels in mouse brain and liver extracts. ( A ), a–d. Mouse extracts from 6 months old mice ( n = 3) were made in PBS and in the presence of protease inhibitors cocktail (Sigma-Aldrich). Immunoprecipitation (IP) experiments were performed on brain and liver extracts (500 µg of protein per extract) using either anti-Cul-1 antibody or anti-Nedd8 antibody followed by Western blot (WB) analyses using anti-Nedd8 antibody or anti-Cul-1 antibody as the primary antibody, respectively. Ae. In a unique experiment, Jab1 was used as a “fishing” probe in an IP experiment using brain extracts, followed by Western blot analysis using anti-Cul-1antibody. Only the 100-kDa neddylated Cul-1 protein was identified due to the specific pull-down of the neddylated form of Cul-1 by Jab1. ( B ) Quantification of each band identified by Western blot analyses described in Panel ( A ) a–e, as determined by using the NIH Image-J program. All the observed differences in the observed band levels between the WT and MT pairs were statistically significant as judged by student t -test analysis (* p < 0.01, n = 3 per strain). ( C ) Loading controls for the liver and brain protein levels, following Coomassie blue staining (to confirm the use of equal amount of proteins from each mouse strain per tissue in the IP experiments. WT, wild type; M, MsrA KO. ( D ) Western blot analysis of the mouse brain extracts using anti Jab1 antibody as the primary antibody and quantified by the NIH Image-J program. WT, wild type; MT, MsrA KO. Arrows shown in ( A ) a–e indicate the position of the neddylated 100-kDa Cul-1 (deneddylated form runs as ~90 kDa protein, not detected). The shown WB and Coomassie blue staining experiments are representatives of three independent experiments.

Article Snippet: Antibodies used included: anti-MsrA antibody (Proteintech Group, Rosemont, IL, USA), anti-Jab1 antibody (Thermo-Fisher Scientific, Waltham, MA, USA), anti-Cul-1 antibody (Novus, Littleton, CO, USA), anti-Nedd8 antibody (Boster Bio, Pleasanton, CA, USA), anti-P27 antibody (Proteintech Group, Rosemont, IL, USA), anti-β actin antibody (Abcam, Cambridge, UK), and HRP-conjugated secondary antibodies for Western blot analyses (Rabbit anti-mouse and Goat anti-rabbit) (Bio-Rad, Hercules, CA, USA).

Techniques: In Vivo, Activity Assay, Immunoprecipitation, Western Blot, Staining

Journal: Antioxidants

Article Title: The Antioxidant Enzyme Methionine Sulfoxide Reductase A (MsrA) Interacts with Jab1/CSN5 and Regulates Its Function

doi: 10.3390/antiox9050452

Figure Lengend Snippet: MsrA alters the kinetics of Jab1 deneddylase activity in vitro. (A) Western blot pattern of control group (time 0′). Total protein neddylation and Nedd8 levels are shown using the primary anti-His antibody. We focused on lowest molecular weight Nedd8 positive bands (Low M.W.Nedd8) as they represent the best end-product of the Nedd8 conjugates through the deneddylation process. ( B ) Quantification of lowest molecular weight Nedd8 positive bands at different time points following incubation with WT and MsrA KO mouse brain extracts ( n = 3, females, 6 months old). ( C ) Time curve of lowest molecular weight Nedd8 levels. All data were normalized to 0 min time. The shown WB are representatives of three repeated experiments. WT, Wild-type; KO, MsrA knockout; The difference between the two animal groups was statistically significant from the 5–30 min time points ( t -test, p < 0.001).

Article Snippet: Antibodies used included: anti-MsrA antibody (Proteintech Group, Rosemont, IL, USA), anti-Jab1 antibody (Thermo-Fisher Scientific, Waltham, MA, USA), anti-Cul-1 antibody (Novus, Littleton, CO, USA), anti-Nedd8 antibody (Boster Bio, Pleasanton, CA, USA), anti-P27 antibody (Proteintech Group, Rosemont, IL, USA), anti-β actin antibody (Abcam, Cambridge, UK), and HRP-conjugated secondary antibodies for Western blot analyses (Rabbit anti-mouse and Goat anti-rabbit) (Bio-Rad, Hercules, CA, USA).

Techniques: Activity Assay, In Vitro, Western Blot, Control, Molecular Weight, Incubation, Knock-Out

Journal: Molecules

Article Title: The Large Molecular Weight Polysaccharide from Wild Cordyceps and Its Antitumor Activity on H22 Tumor-Bearing Mice

doi: 10.3390/molecules28083351

Figure Lengend Snippet: WCP promotes Cyto-c/Caspase8/3 and inhibits IL-10/STAT3/Bcl2 pathway. ( A – F ) Relative mRNA expression of IL-6, IL-Iβ, NF-κB, TNF-α, Bax, and Bcl2. * p < 0.05 compared to model group.

Article Snippet: Rabbit monoclonal antibodies against Caspase8 (PTM-6085) antibody were purchased from PTM Biolab Co., Ltd. (Hangzhou, China), and signal transducer and activator of transcription3 (Stat3, BM4052), phosphorylated signal transducer and activator of transcription3 (p-STAT3 Y705, BM4835) were purchased from Boster Biological Technology Co., Ltd. (Wuhan, China).

Techniques: Expressing

Journal: Molecules

Article Title: The Large Molecular Weight Polysaccharide from Wild Cordyceps and Its Antitumor Activity on H22 Tumor-Bearing Mice

doi: 10.3390/molecules28083351

Figure Lengend Snippet: WCP promotes Cyto-c/Caspase8/3 and inhibits IL-10/STAT3/Bcl2pathway. ( A , F ) Photographs of the proteins in each group. ( B – E , G – I ) Relative protein expression of Cyto-c, Caspase8, Caspase3, p-STAT3, Bcl2, Bax, and Bax/Bcl2. * p < 0.05, ** p < 0.01, *** p < 0.001 compared to model group.

Article Snippet: Rabbit monoclonal antibodies against Caspase8 (PTM-6085) antibody were purchased from PTM Biolab Co., Ltd. (Hangzhou, China), and signal transducer and activator of transcription3 (Stat3, BM4052), phosphorylated signal transducer and activator of transcription3 (p-STAT3 Y705, BM4835) were purchased from Boster Biological Technology Co., Ltd. (Wuhan, China).

Techniques: Expressing

Journal: International journal of oncology

Article Title: Knockdown of SNHG15 suppresses renal cell carcinoma proliferation and EMT by regulating the NF-κB signaling pathway.

doi: 10.3892/ijo.2018.4395

Figure Lengend Snippet: Figure 5. Knockdown of SNHG15 affects NF‑κB entry into the nucleus. (A) Knockdown of SNHG15 reduced the protein expression levels of EMT‑associated transcription factors (Snail1, Slug and ZEB1). However, there was no difference in the total protein expression levels of NF‑κB between the SNHG15 siRNA and NC groups. *P<0.01; nsP>0.05. (B) Nuclear/cytoplasmic NF‑κB expression in ACHN and 786‑O cells transfected with siRNAs. *P<0.01. (C) Following stimulation with TNF‑α for 6 h, EMT markers were examined among the various groups. *P<0.01.

Article Snippet: Monoclonal primary antibodies were specific to Snail (1:1,000, 3879), Slug (1:1,000, 9585), Vimentin (1:1,000, 5741), E-cadherin (1:1,000, 14472), N-cadherin (1:1,000, 13116) and Histone H3 (1:2,000, 4499) (all from Cell Signaling Technology, Inc., Danvers, MA, USA), ZEB1 (1:500, BA2871‐2; Wuhan Boster Biological Technology, Ltd., Wuhan, China), β-actin (1:5,000, A1978), GAPDH (1:5,000, G9545) (both from Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) and NF-κB1 (p50) (1:1,000, 14220-1-AP; Wuhan Sanying Biotechnology, Wuhan, China).

Techniques: Knockdown, Expressing, Transfection

Journal: International journal of oncology

Article Title: Knockdown of SNHG15 suppresses renal cell carcinoma proliferation and EMT by regulating the NF-κB signaling pathway.

doi: 10.3892/ijo.2018.4395

Figure Lengend Snippet: Figure 5. Continued. (D) Nuclear immunofluorescence intensity of NF‑κB was reduced in the SNHG15 siRNA groups compared with in the NC groups. (E) Cell migration and invasion were altered among the various groups following TNF‑α stimulation and siRNA transfection (magnification, x40). *P<0.01. EMT, epithelial‑mesenchymal transition; NC, negative control; NF‑κB, nuclear factor‑κB; siRNA, small interfering RNA; SNHG15, small nucleolar RNA host gene 15; TNF‑α, tumor necrosis factor‑α; ZEB1, zinc finger E‑box‑binding homeobox 1.

Article Snippet: Monoclonal primary antibodies were specific to Snail (1:1,000, 3879), Slug (1:1,000, 9585), Vimentin (1:1,000, 5741), E-cadherin (1:1,000, 14472), N-cadherin (1:1,000, 13116) and Histone H3 (1:2,000, 4499) (all from Cell Signaling Technology, Inc., Danvers, MA, USA), ZEB1 (1:500, BA2871‐2; Wuhan Boster Biological Technology, Ltd., Wuhan, China), β-actin (1:5,000, A1978), GAPDH (1:5,000, G9545) (both from Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) and NF-κB1 (p50) (1:1,000, 14220-1-AP; Wuhan Sanying Biotechnology, Wuhan, China).

Techniques: Immunofluorescence, Migration, Transfection, Negative Control, Small Interfering RNA

Journal: International review of neurobiology

Article Title: Cell transplantation to repair the injured spinal cord

doi: 10.1016/bs.irn.2022.09.008

Figure Lengend Snippet: Transplantation of tissues and cells to provide novel neuronal connections & an anatomical relay for spinal cord repair.

Article Snippet: S.Biomedics Co., Ltd. (Korea) , Neural precursor cells derived from human embryonic stem cell line , 18–65 y.o. SCI at C4-C7 level; AIS A , 7–60 days post-injury , Phase I and II , Safety; sensory and motor function evaluated with AIS, ISNCSCI, and SCIM scores , 2021–2028 , 5 , NCT04812431.

Techniques: Transplantation Assay, Suspension, Cell Culture

Journal: International review of neurobiology

Article Title: Cell transplantation to repair the injured spinal cord

doi: 10.1016/bs.irn.2022.09.008

Figure Lengend Snippet: Transplantation of tissues and cells to provide neuroprotection of spinal cord tissue.

Article Snippet: S.Biomedics Co., Ltd. (Korea) , Neural precursor cells derived from human embryonic stem cell line , 18–65 y.o. SCI at C4-C7 level; AIS A , 7–60 days post-injury , Phase I and II , Safety; sensory and motor function evaluated with AIS, ISNCSCI, and SCIM scores , 2021–2028 , 5 , NCT04812431.

Techniques: Transplantation Assay, Derivative Assay, Cell Culture

Journal: International review of neurobiology

Article Title: Cell transplantation to repair the injured spinal cord

doi: 10.1016/bs.irn.2022.09.008

Figure Lengend Snippet: Clinical trials transplanting tissue and cells for spinal cord repair.

Article Snippet: S.Biomedics Co., Ltd. (Korea) , Neural precursor cells derived from human embryonic stem cell line , 18–65 y.o. SCI at C4-C7 level; AIS A , 7–60 days post-injury , Phase I and II , Safety; sensory and motor function evaluated with AIS, ISNCSCI, and SCIM scores , 2021–2028 , 5 , NCT04812431.

Techniques: Clinical Proteomics, Modification, Derivative Assay, Transplantation Assay, Functional Assay, Control, Cell Culture

Journal: International review of neurobiology

Article Title: Cell transplantation to repair the injured spinal cord

doi: 10.1016/bs.irn.2022.09.008

Figure Lengend Snippet: Schematic diagram highlighting potential sources of neural stem and precursor cells: the developing neural tissues (A), pluripotent embryonic stem cells (B), and induced pluripotent stem cells (C; exemplified by skin fibroblast de-differentiation/reprogramming). These cells can be engineered to produce neural stem and progenitor cells that can then be expanded (D), specific subsets selected (e.g., NRPs and GRPs), and cryopreserved for “cell banking.” These cell stores can be thawed and prepared for transplantation into the injured spinal cord (E) alone, or in combination with additional treatments (E), including rehabilitation and activity-based therapy, neural interfacing and neuromodulation, application of scaffolds/biomaterials, or additional pharmaceuticals to enhance efficacy. Figure modified from Zholudeva, L. V., & Lane, M. A. (2022). Spinal interneurons: Plasticity after spinal cord injury, 1st ed. Academic Press.

Article Snippet: S.Biomedics Co., Ltd. (Korea) , Neural precursor cells derived from human embryonic stem cell line , 18–65 y.o. SCI at C4-C7 level; AIS A , 7–60 days post-injury , Phase I and II , Safety; sensory and motor function evaluated with AIS, ISNCSCI, and SCIM scores , 2021–2028 , 5 , NCT04812431.

Techniques: Transplantation Assay, Activity Assay, Modification