atg4b (Proteintech)
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atg4b
Atg4b, supplied by Proteintech, used in various techniques. Bioz Stars score: 96/100, based on 1514 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/atg4b/Beclin+1+Antibody/pmc13003811-32-129-133
Average 96 stars, based on 1514 article reviews
Atg4b, supplied by Proteintech, used in various techniques. Bioz Stars score: 96/100, based on 1514 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/atg4b/Beclin+1+Antibody/pmc13003811-32-129-133
Average 96 stars, based on 1514 article reviews
atg4b - by Bioz Stars,
2026-09
96/100 stars
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1) Product Images from "Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response"
Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response
Journal: Journal of Inflammation Research
doi: 10.2147/JIR.S555053
Figure Legend Snippet: LPS activates autophagy-related protein expression in HBE cells in a concentration-dependent manner. ( A ) Representative Western blot bands showing the expression of ATG7, PIK3C3, ATG4B, p62, LC3B, and β-actin (loading control) in HBE cells treated with 0, 1, 10, 50, or 100 ng/mL LPS. ( B–F ) Quantitative analysis of relative protein expression (normalized to β-actin) for ATG4B ( B ), ATG7 ( C ), PIK3C3 ( D ), p62 ( E ), and LC3B-II ( F ). Data are presented as mean ± SD (n=3 independent experiments). *P<0.05 vs 0 ng/mL group.
Techniques Used: Expressing, Concentration Assay, Western Blot, Control
Figure Legend Snippet: G-Rg3 Antagonizes LPS-Induced Autophagy in HBE Cells in Concentration- and Time-Dependent Manners. ( A ) Representative Western blot bands of ATG7, PIK3C3, ATG4B, p62, LC3B, and β-actin (loading control) in HBE cells treated with LPS (100 ng/mL) alone or co-treated with 2–16 μM G-Rg3. ( B ) Representative Western blot bands of p62, LC3B, and β-actin in HBE cells treated with LPS (100 ng/mL) + 16 μM G-Rg3 for 0–30 h. ( C ) Quantitative analysis of relative p62 and LC3B-II/I expression (normalized to β-actin) in LPS-treated cells with different G-Rg3 concentrations. ( D ) Quantitative analysis of relative p62 and LC3B-II/I expression (normalized to β-actin) in LPS + 16 μM G-Rg3-treated cells at different time points. Data are presented as mean ± SD (n = 3 independent experiments). *P < 0.05 vs the corresponding control group.
Techniques Used: Concentration Assay, Western Blot, Control, Expressing
Related Articles
Concentration Assay:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Human bronchial epithelial (HBE) cells (ATCC, Cat# CCL-202) were used as the in vitro model, with strict control of cell handling and culture conditions to ensure experimental consistency: remove cryopreserved cells from −80°C storage, immediately immerse the cryovial in a 37°C water bath (Thermo Fisher, Model WB2000) and gently agitate for 1–2 minutes until only a small ice crystal remains, disinfect the vial surface with 75% ethanol (Solarbio, Cat# 10009218), transfer the cell suspension to a 15 mL centrifuge tube (Corning, Cat# 430791) containing 5 mL pre-warmed (37°C) complete Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Cat# c11965500BT) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Cat# 10099141) and 1% penicillin-streptomycin (100 U/mL penicillin, 100 μg/mL streptomycin; Solarbio, Cat# P1400), centrifuge at 1000×g for 5 minutes at room temperature (RT), discard the supernatant, resuspend the cell pellet in 5 mL complete DMEM, count cells using a hemocytometer (Thermo Fisher, Cat# 02–671-5), seed at a density of 5×10 5 cells per T75 flask (Corning, Cat# 430641), and culture in a humidified incubator (Thermo Fisher, Model 3111) set to 37°C, 5% CO 2 , and 95% relative humidity; when cells reach 80–90% confluence (typically 48–72 hours post-seeding), perform passaging by aspirating the culture medium, rinsing twice with 5 mL pre-warmed PBS (Solarbio, Cat# P1020), adding 2 mL 0.25% trypsin-EDTA (Gibco, Cat# 25200056) and incubating at 37°C for 2–3 minutes (monitoring cell detachment under an inverted microscope), neutralizing trypsin with 4 mL complete DMEM, centrifuging at 1000×g for 5 minutes, discarding the supernatant, resuspending the cell pellet in 5 mL complete DMEM, counting again, and seeding at the required density, with only cells at passages 3–8 used for experiments (to avoid cellular senescence) and cell viability confirmed to be >95% via trypan blue staining (Solarbio, Cat# T8154) before each experiment. .. Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Cell Culture:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Human bronchial epithelial (HBE) cells (ATCC, Cat# CCL-202) were used as the in vitro model, with strict control of cell handling and culture conditions to ensure experimental consistency: remove cryopreserved cells from −80°C storage, immediately immerse the cryovial in a 37°C water bath (Thermo Fisher, Model WB2000) and gently agitate for 1–2 minutes until only a small ice crystal remains, disinfect the vial surface with 75% ethanol (Solarbio, Cat# 10009218), transfer the cell suspension to a 15 mL centrifuge tube (Corning, Cat# 430791) containing 5 mL pre-warmed (37°C) complete Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Cat# c11965500BT) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Cat# 10099141) and 1% penicillin-streptomycin (100 U/mL penicillin, 100 μg/mL streptomycin; Solarbio, Cat# P1400), centrifuge at 1000×g for 5 minutes at room temperature (RT), discard the supernatant, resuspend the cell pellet in 5 mL complete DMEM, count cells using a hemocytometer (Thermo Fisher, Cat# 02–671-5), seed at a density of 5×10 5 cells per T75 flask (Corning, Cat# 430641), and culture in a humidified incubator (Thermo Fisher, Model 3111) set to 37°C, 5% CO 2 , and 95% relative humidity; when cells reach 80–90% confluence (typically 48–72 hours post-seeding), perform passaging by aspirating the culture medium, rinsing twice with 5 mL pre-warmed PBS (Solarbio, Cat# P1020), adding 2 mL 0.25% trypsin-EDTA (Gibco, Cat# 25200056) and incubating at 37°C for 2–3 minutes (monitoring cell detachment under an inverted microscope), neutralizing trypsin with 4 mL complete DMEM, centrifuging at 1000×g for 5 minutes, discarding the supernatant, resuspending the cell pellet in 5 mL complete DMEM, counting again, and seeding at the required density, with only cells at passages 3–8 used for experiments (to avoid cellular senescence) and cell viability confirmed to be >95% via trypan blue staining (Solarbio, Cat# T8154) before each experiment. .. Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Sterility:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Human bronchial epithelial (HBE) cells (ATCC, Cat# CCL-202) were used as the in vitro model, with strict control of cell handling and culture conditions to ensure experimental consistency: remove cryopreserved cells from −80°C storage, immediately immerse the cryovial in a 37°C water bath (Thermo Fisher, Model WB2000) and gently agitate for 1–2 minutes until only a small ice crystal remains, disinfect the vial surface with 75% ethanol (Solarbio, Cat# 10009218), transfer the cell suspension to a 15 mL centrifuge tube (Corning, Cat# 430791) containing 5 mL pre-warmed (37°C) complete Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Cat# c11965500BT) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Cat# 10099141) and 1% penicillin-streptomycin (100 U/mL penicillin, 100 μg/mL streptomycin; Solarbio, Cat# P1400), centrifuge at 1000×g for 5 minutes at room temperature (RT), discard the supernatant, resuspend the cell pellet in 5 mL complete DMEM, count cells using a hemocytometer (Thermo Fisher, Cat# 02–671-5), seed at a density of 5×10 5 cells per T75 flask (Corning, Cat# 430641), and culture in a humidified incubator (Thermo Fisher, Model 3111) set to 37°C, 5% CO 2 , and 95% relative humidity; when cells reach 80–90% confluence (typically 48–72 hours post-seeding), perform passaging by aspirating the culture medium, rinsing twice with 5 mL pre-warmed PBS (Solarbio, Cat# P1020), adding 2 mL 0.25% trypsin-EDTA (Gibco, Cat# 25200056) and incubating at 37°C for 2–3 minutes (monitoring cell detachment under an inverted microscope), neutralizing trypsin with 4 mL complete DMEM, centrifuging at 1000×g for 5 minutes, discarding the supernatant, resuspending the cell pellet in 5 mL complete DMEM, counting again, and seeding at the required density, with only cells at passages 3–8 used for experiments (to avoid cellular senescence) and cell viability confirmed to be >95% via trypan blue staining (Solarbio, Cat# T8154) before each experiment. .. Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Saline:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Human bronchial epithelial (HBE) cells (ATCC, Cat# CCL-202) were used as the in vitro model, with strict control of cell handling and culture conditions to ensure experimental consistency: remove cryopreserved cells from −80°C storage, immediately immerse the cryovial in a 37°C water bath (Thermo Fisher, Model WB2000) and gently agitate for 1–2 minutes until only a small ice crystal remains, disinfect the vial surface with 75% ethanol (Solarbio, Cat# 10009218), transfer the cell suspension to a 15 mL centrifuge tube (Corning, Cat# 430791) containing 5 mL pre-warmed (37°C) complete Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Cat# c11965500BT) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Cat# 10099141) and 1% penicillin-streptomycin (100 U/mL penicillin, 100 μg/mL streptomycin; Solarbio, Cat# P1400), centrifuge at 1000×g for 5 minutes at room temperature (RT), discard the supernatant, resuspend the cell pellet in 5 mL complete DMEM, count cells using a hemocytometer (Thermo Fisher, Cat# 02–671-5), seed at a density of 5×10 5 cells per T75 flask (Corning, Cat# 430641), and culture in a humidified incubator (Thermo Fisher, Model 3111) set to 37°C, 5% CO 2 , and 95% relative humidity; when cells reach 80–90% confluence (typically 48–72 hours post-seeding), perform passaging by aspirating the culture medium, rinsing twice with 5 mL pre-warmed PBS (Solarbio, Cat# P1020), adding 2 mL 0.25% trypsin-EDTA (Gibco, Cat# 25200056) and incubating at 37°C for 2–3 minutes (monitoring cell detachment under an inverted microscope), neutralizing trypsin with 4 mL complete DMEM, centrifuging at 1000×g for 5 minutes, discarding the supernatant, resuspending the cell pellet in 5 mL complete DMEM, counting again, and seeding at the required density, with only cells at passages 3–8 used for experiments (to avoid cellular senescence) and cell viability confirmed to be >95% via trypan blue staining (Solarbio, Cat# T8154) before each experiment. .. Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Inhibition:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Human bronchial epithelial (HBE) cells (ATCC, Cat# CCL-202) were used as the in vitro model, with strict control of cell handling and culture conditions to ensure experimental consistency: remove cryopreserved cells from −80°C storage, immediately immerse the cryovial in a 37°C water bath (Thermo Fisher, Model WB2000) and gently agitate for 1–2 minutes until only a small ice crystal remains, disinfect the vial surface with 75% ethanol (Solarbio, Cat# 10009218), transfer the cell suspension to a 15 mL centrifuge tube (Corning, Cat# 430791) containing 5 mL pre-warmed (37°C) complete Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Cat# c11965500BT) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Cat# 10099141) and 1% penicillin-streptomycin (100 U/mL penicillin, 100 μg/mL streptomycin; Solarbio, Cat# P1400), centrifuge at 1000×g for 5 minutes at room temperature (RT), discard the supernatant, resuspend the cell pellet in 5 mL complete DMEM, count cells using a hemocytometer (Thermo Fisher, Cat# 02–671-5), seed at a density of 5×10 5 cells per T75 flask (Corning, Cat# 430641), and culture in a humidified incubator (Thermo Fisher, Model 3111) set to 37°C, 5% CO 2 , and 95% relative humidity; when cells reach 80–90% confluence (typically 48–72 hours post-seeding), perform passaging by aspirating the culture medium, rinsing twice with 5 mL pre-warmed PBS (Solarbio, Cat# P1020), adding 2 mL 0.25% trypsin-EDTA (Gibco, Cat# 25200056) and incubating at 37°C for 2–3 minutes (monitoring cell detachment under an inverted microscope), neutralizing trypsin with 4 mL complete DMEM, centrifuging at 1000×g for 5 minutes, discarding the supernatant, resuspending the cell pellet in 5 mL complete DMEM, counting again, and seeding at the required density, with only cells at passages 3–8 used for experiments (to avoid cellular senescence) and cell viability confirmed to be >95% via trypan blue staining (Solarbio, Cat# T8154) before each experiment. .. Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Positive Control:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Human bronchial epithelial (HBE) cells (ATCC, Cat# CCL-202) were used as the in vitro model, with strict control of cell handling and culture conditions to ensure experimental consistency: remove cryopreserved cells from −80°C storage, immediately immerse the cryovial in a 37°C water bath (Thermo Fisher, Model WB2000) and gently agitate for 1–2 minutes until only a small ice crystal remains, disinfect the vial surface with 75% ethanol (Solarbio, Cat# 10009218), transfer the cell suspension to a 15 mL centrifuge tube (Corning, Cat# 430791) containing 5 mL pre-warmed (37°C) complete Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Cat# c11965500BT) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Cat# 10099141) and 1% penicillin-streptomycin (100 U/mL penicillin, 100 μg/mL streptomycin; Solarbio, Cat# P1400), centrifuge at 1000×g for 5 minutes at room temperature (RT), discard the supernatant, resuspend the cell pellet in 5 mL complete DMEM, count cells using a hemocytometer (Thermo Fisher, Cat# 02–671-5), seed at a density of 5×10 5 cells per T75 flask (Corning, Cat# 430641), and culture in a humidified incubator (Thermo Fisher, Model 3111) set to 37°C, 5% CO 2 , and 95% relative humidity; when cells reach 80–90% confluence (typically 48–72 hours post-seeding), perform passaging by aspirating the culture medium, rinsing twice with 5 mL pre-warmed PBS (Solarbio, Cat# P1020), adding 2 mL 0.25% trypsin-EDTA (Gibco, Cat# 25200056) and incubating at 37°C for 2–3 minutes (monitoring cell detachment under an inverted microscope), neutralizing trypsin with 4 mL complete DMEM, centrifuging at 1000×g for 5 minutes, discarding the supernatant, resuspending the cell pellet in 5 mL complete DMEM, counting again, and seeding at the required density, with only cells at passages 3–8 used for experiments (to avoid cellular senescence) and cell viability confirmed to be >95% via trypan blue staining (Solarbio, Cat# T8154) before each experiment. .. Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Lysis:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Human bronchial epithelial (HBE) cells (ATCC, Cat# CCL-202) were used as the in vitro model, with strict control of cell handling and culture conditions to ensure experimental consistency: remove cryopreserved cells from −80°C storage, immediately immerse the cryovial in a 37°C water bath (Thermo Fisher, Model WB2000) and gently agitate for 1–2 minutes until only a small ice crystal remains, disinfect the vial surface with 75% ethanol (Solarbio, Cat# 10009218), transfer the cell suspension to a 15 mL centrifuge tube (Corning, Cat# 430791) containing 5 mL pre-warmed (37°C) complete Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Cat# c11965500BT) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Cat# 10099141) and 1% penicillin-streptomycin (100 U/mL penicillin, 100 μg/mL streptomycin; Solarbio, Cat# P1400), centrifuge at 1000×g for 5 minutes at room temperature (RT), discard the supernatant, resuspend the cell pellet in 5 mL complete DMEM, count cells using a hemocytometer (Thermo Fisher, Cat# 02–671-5), seed at a density of 5×10 5 cells per T75 flask (Corning, Cat# 430641), and culture in a humidified incubator (Thermo Fisher, Model 3111) set to 37°C, 5% CO 2 , and 95% relative humidity; when cells reach 80–90% confluence (typically 48–72 hours post-seeding), perform passaging by aspirating the culture medium, rinsing twice with 5 mL pre-warmed PBS (Solarbio, Cat# P1020), adding 2 mL 0.25% trypsin-EDTA (Gibco, Cat# 25200056) and incubating at 37°C for 2–3 minutes (monitoring cell detachment under an inverted microscope), neutralizing trypsin with 4 mL complete DMEM, centrifuging at 1000×g for 5 minutes, discarding the supernatant, resuspending the cell pellet in 5 mL complete DMEM, counting again, and seeding at the required density, with only cells at passages 3–8 used for experiments (to avoid cellular senescence) and cell viability confirmed to be >95% via trypan blue staining (Solarbio, Cat# T8154) before each experiment. .. Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Protease Inhibitor:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Human bronchial epithelial (HBE) cells (ATCC, Cat# CCL-202) were used as the in vitro model, with strict control of cell handling and culture conditions to ensure experimental consistency: remove cryopreserved cells from −80°C storage, immediately immerse the cryovial in a 37°C water bath (Thermo Fisher, Model WB2000) and gently agitate for 1–2 minutes until only a small ice crystal remains, disinfect the vial surface with 75% ethanol (Solarbio, Cat# 10009218), transfer the cell suspension to a 15 mL centrifuge tube (Corning, Cat# 430791) containing 5 mL pre-warmed (37°C) complete Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Cat# c11965500BT) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Cat# 10099141) and 1% penicillin-streptomycin (100 U/mL penicillin, 100 μg/mL streptomycin; Solarbio, Cat# P1400), centrifuge at 1000×g for 5 minutes at room temperature (RT), discard the supernatant, resuspend the cell pellet in 5 mL complete DMEM, count cells using a hemocytometer (Thermo Fisher, Cat# 02–671-5), seed at a density of 5×10 5 cells per T75 flask (Corning, Cat# 430641), and culture in a humidified incubator (Thermo Fisher, Model 3111) set to 37°C, 5% CO 2 , and 95% relative humidity; when cells reach 80–90% confluence (typically 48–72 hours post-seeding), perform passaging by aspirating the culture medium, rinsing twice with 5 mL pre-warmed PBS (Solarbio, Cat# P1020), adding 2 mL 0.25% trypsin-EDTA (Gibco, Cat# 25200056) and incubating at 37°C for 2–3 minutes (monitoring cell detachment under an inverted microscope), neutralizing trypsin with 4 mL complete DMEM, centrifuging at 1000×g for 5 minutes, discarding the supernatant, resuspending the cell pellet in 5 mL complete DMEM, counting again, and seeding at the required density, with only cells at passages 3–8 used for experiments (to avoid cellular senescence) and cell viability confirmed to be >95% via trypan blue staining (Solarbio, Cat# T8154) before each experiment. .. Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Bicinchoninic Acid Protein Assay:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Human bronchial epithelial (HBE) cells (ATCC, Cat# CCL-202) were used as the in vitro model, with strict control of cell handling and culture conditions to ensure experimental consistency: remove cryopreserved cells from −80°C storage, immediately immerse the cryovial in a 37°C water bath (Thermo Fisher, Model WB2000) and gently agitate for 1–2 minutes until only a small ice crystal remains, disinfect the vial surface with 75% ethanol (Solarbio, Cat# 10009218), transfer the cell suspension to a 15 mL centrifuge tube (Corning, Cat# 430791) containing 5 mL pre-warmed (37°C) complete Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Cat# c11965500BT) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Cat# 10099141) and 1% penicillin-streptomycin (100 U/mL penicillin, 100 μg/mL streptomycin; Solarbio, Cat# P1400), centrifuge at 1000×g for 5 minutes at room temperature (RT), discard the supernatant, resuspend the cell pellet in 5 mL complete DMEM, count cells using a hemocytometer (Thermo Fisher, Cat# 02–671-5), seed at a density of 5×10 5 cells per T75 flask (Corning, Cat# 430641), and culture in a humidified incubator (Thermo Fisher, Model 3111) set to 37°C, 5% CO 2 , and 95% relative humidity; when cells reach 80–90% confluence (typically 48–72 hours post-seeding), perform passaging by aspirating the culture medium, rinsing twice with 5 mL pre-warmed PBS (Solarbio, Cat# P1020), adding 2 mL 0.25% trypsin-EDTA (Gibco, Cat# 25200056) and incubating at 37°C for 2–3 minutes (monitoring cell detachment under an inverted microscope), neutralizing trypsin with 4 mL complete DMEM, centrifuging at 1000×g for 5 minutes, discarding the supernatant, resuspending the cell pellet in 5 mL complete DMEM, counting again, and seeding at the required density, with only cells at passages 3–8 used for experiments (to avoid cellular senescence) and cell viability confirmed to be >95% via trypan blue staining (Solarbio, Cat# T8154) before each experiment. .. Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Western Blot:Article Title: A biomimetic multimodal nanoplatform combining neutrophil-coated two-dimensional metalloporphyrinic framework nanosheet and exendin-4 to treat obesity-related osteoporosis Article Snippet: .. WB using antibodies including: TET2 (Abcam, USA), COL1A1 (Collagen I, Abcam, UK), RUNX2 (Runt-related transcription factor 2, CST, USA), PINK1 (PTEN-induced putative kinase protein 1, Bioss, China), Parkin (E3 ubiquitin-protein ligase parkin, Servicebio, China), LC3B (Light Chain 3, Abmart, China), ATG4B (Autophagy Related 4B Cysteine Peptidase, Proteintech, China), BCL2 (B-cell lymphoma-2, Abmart, China), BECN1 (Beclin-1, Abmart, China), ACTB (Beyotime, China), Integrin β1 (MCE, USA), Integrin β2 (Bio-Techne, USA), CXCR2 (Proteintech, China), HRP-labeled Goat Anti-Rabbit IgG (H + L) (Beyotime, China), HRP-labeled Goat Anti-Mouse IgG (H + L) (Beyotime, China), KEAP1 (Kelch Like ECH Associated Protein 1, Proteintech, China),NRF2 (Nuclear factor erythroid 2-related factor 2, Proteintech, China). .. Article Title: Hexavalent chromium induced autophagy-dependent mTOR expression mediated by upregulation of HMGA2. Article Snippet: High mobility group A2 (HMGA2) overexpression is often observed in cancers.. Previously, we found that HMGA2 contributed to hexavalent chromium [Cr (VI)]-mediated autophagy.. In this study, interestingly, Cr (VI) treatment triggered both autophagy and the mammalian target of rapamycin (mTOR) in vivo ( BALB/c mice, 0.5 and 1.5 mg/kg, p.o.) and in vitro (0.1, 0.2 and 0.4 μM in A549 and HELF cells), and both autophagy and mTOR were implicated in Cr (VI)-initiated cell growth. Ubiquitin Proteomics:Article Title: A biomimetic multimodal nanoplatform combining neutrophil-coated two-dimensional metalloporphyrinic framework nanosheet and exendin-4 to treat obesity-related osteoporosis Article Snippet: .. WB using antibodies including: TET2 (Abcam, USA), COL1A1 (Collagen I, Abcam, UK), RUNX2 (Runt-related transcription factor 2, CST, USA), PINK1 (PTEN-induced putative kinase protein 1, Bioss, China), Parkin (E3 ubiquitin-protein ligase parkin, Servicebio, China), LC3B (Light Chain 3, Abmart, China), other:Article Title: Ginsenoside Rg3 Mitigates LPS-Induced Injury in Human Bronchial Epithelial Cells by Restoring Autophagic Flux and Inhibiting the TLR4/NF-κB-Mediated Inflammatory Response Article Snippet: Reagents and Antibodies Ginsenoside Rg3 (G-Rg3) with ≥98% purity was purchased from TargetMol (Cat# T3402), dissolved in dimethyl sulfoxide (DMSO; Sigma, Cat# C6628) to prepare a 10 mM stock solution, and stored at −80°C (avoiding repeated freeze-thaw cycles, with final DMSO concentration in cell culture ≤0.1% to eliminate cytotoxicity); lipopolysaccharide (LPS) from Escherichia coli 055:B5 was obtained from Solarbio (Beijing, China; Cat# L8880), reconstituted in sterile phosphate-buffered saline (PBS; Solarbio, Cat# P1020) to a 1 mg/mL stock solution, filtered through a 0.22 μm sterile filter (Millipore, Cat# SLGP033RB), and stored at −20°C; chloroquine (autophagy inhibition positive control) was purchased from Sigma (Cat# RNBH9960), prepared as a 50 mM stock solution in PBS, and stored at 4°C; primary antibodies against autophagy-related proteins and internal reference were sourced as follows: β-actin (Cat# 66009-1-Ig) and Article Title: Targeting STK26 and ATG4B: miR-22-3p as a modulator of autophagy and tumor progression in HCC Article Snippet: Antibody(IHC) , |
