The human podocytes (AB8/13) was obtained from American Type Culture Collection (ATCC, Manassas, VA, USA) and cultured in RPMI 1640 Medium supplemented with 10% fetal bovine serum (Thermo Fisher Scientific, Waltham, MA, USA), 100 μg/mL streptomycin and 100 U/mL penicillin at 37°C with 5% CO2. To mimic DN in vitro, AB8/13 cells were treated with 30 mM high glucose (HG, Sigma- Aldrich, Saint Louis, MO, USA) for 48 h.
High glucose hg
Manufactured by Merck Group
Sourced in United States
High glucose (HG) is a laboratory equipment product used for the measurement and analysis of glucose levels. It serves as a core function in various applications, such as clinical diagnostics, research, and quality control processes. The detailed specifications and intended use of this product require further information to be provided in an unbiased and factual manner.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Ab8 13, by American Type Culture Collection (1 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions)
Dmem f12, by Merck Group (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Mannitol, by Merck Group (1 mentions)
Sv40 mes13, by Procell (1 mentions)
6 protocols using high glucose hg
1
In Vitro Diabetic Nephropathy Model
2
HK-2 Cells Glucose Stimulation Model
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A human proximal tubular epithelial cell line (HK-2) was kindly provided by Dr. John Cijiang He (Icahn School of Medicine at Mount Sinai, New York, USA). The cells were cultured in DMEM/F12 (Sigma-Aldrich) medium supplemented with 10% fetal bovine serum (Gibco, Grand Island, NY), 1% penicillin, and streptomycin in an atmosphere of 5% CO2 and 95% air at 37 °C.
In all experiments, the cells were starved in a serum-free medium for 12–16 h, and then stimulated with 30 mmol/L of high glucose (HG, Sigma-Aldrich) for 24 h. Box5 (100 μM/L) or recombinant Wnt5a protein (200 ng/mL) was given for the indicated time periods. The cells were pretreated with Box5 2 h before the glucose treatment.
For gene disruption, HK-2 cells were plated in six-well plates and transiently transfected with 50 nM Wnt5a-specific small-interfering RNA (siRNA) (Sigma-Aldrich, St. Louis, MO, USA) or 1 μg CD146-specific siRNA (Thermo Fisher Scientific, Waltham, MA, USA) using Lipofectamine 3000 (Invitrogen, Grand Island, NY) according to the manufacturer’s protocol. The efficacy of knockdown was determined by real-time PCR and western blotting (Supplementary Fig. S2 ). After transfection for 48 h, these cells were treated with HG for another 24 h. Cell supernatants, lysates, and RNA were collected for subsequent experiments.
In all experiments, the cells were starved in a serum-free medium for 12–16 h, and then stimulated with 30 mmol/L of high glucose (HG, Sigma-Aldrich) for 24 h. Box5 (100 μM/L) or recombinant Wnt5a protein (200 ng/mL) was given for the indicated time periods. The cells were pretreated with Box5 2 h before the glucose treatment.
For gene disruption, HK-2 cells were plated in six-well plates and transiently transfected with 50 nM Wnt5a-specific small-interfering RNA (siRNA) (Sigma-Aldrich, St. Louis, MO, USA) or 1 μg CD146-specific siRNA (Thermo Fisher Scientific, Waltham, MA, USA) using Lipofectamine 3000 (Invitrogen, Grand Island, NY) according to the manufacturer’s protocol. The efficacy of knockdown was determined by real-time PCR and western blotting (Supplementary Fig. S
3
Glucose Stimulation Across Cell Types
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were stimulated with 30 mmol/L d ‐glucose [High Glucose, ‘HG’ (Sigma)] for 6 hours (RAW 264.7 and BMDM), 3 hours (3T3‐L1) or 30 minutes (βTC6).
4
Aortic and Carotid Artery Functional Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After mice were sacrificed by CO2 inhalation, mouse thoracic aortas and carotid arteries were dissected and cleaned in sterile phosphate-buffered saline (PBS) and then cut into segments (~2 mm) and incubated in Dulbecco’s modified Eagle’s medium (5.55 mM glucose present in DMEM, Gibco, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS, Gibco, Paisley, UK) and 1% penicillin/streptomycin (P/S, Gibco, Grand Island, NY, USA). Mannitol as osmotic control, high glucose (HG; 30 mM, 48 h, Sigma-Aldrich, ST. Louis, MO, USA), ER stress inducer tunicamycin (Tuni; 2 µg/mL, 24 h) and JAT (0.1 or 1 µM; Shanghai Aladdin Bio-Chem 96 Technology Co. Ltd., Shanghai, China) were added individually into the culture medium that bathed the aortic rings in an incubator of 5% CO2 at 37 °C. After the incubation period, segments were transferred to fresh Krebs solution for functional studies in a wire myograph and were frozen for Western blotting and fluorescence imaging.
5
Coptisine Modulates Vascular Function
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After mice were sacrificed, mouse thoracic aortas and carotid arteries were dissected in sterile PBS and then incubated in DMEM supplemented with 10% FBS and 1% penicillin/streptomycin (Gibco, Gaithersbury, MD, USA). High glucose (HG; 30 mM, 48 h; Sigma-Aldrich, St. Louis, MO, USA) and tunicamycin (tuni; 2 µg/ml, 24 h; Sigma-Aldrich) were added individually into the culture medium that bathed aortic rings in a humidified atmosphere of 5% CO2 at 37 °C. Some arterial rings were co-treated with coptisine (COP, 0.1 or 1 µM) which was purchased from Shanghai Aladdin Bio-Chem Technology Co. Ltd. (Shanghai, China) (purity > 98%). After the incubation period, segments were transferred to fresh Krebs solution for functional studies in a wire myograph and were frozen for Western blotting and fluorescence imaging.
6
Mesangial Cell Glucose Response
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Mouse glomerulus mesangial cells (SV40-MES13; Procell, Wuhan, China) were cultured in 71.25% DMEM + 23.75% Ham’s F-12 medium + 5% FBS. Cells were administered with normal glucose (NG; Sigma-Aldrich; 5.5 mmol/mL), high glucose (HG; 25 mmol/mL) or mannitol (20 mmol/L; Sigma-Aldrich) and then cultured for 24 h. Cells were cultured at 37 °C conditions containing 5% CO2.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!