cdh17 sirna silencer® select pre-designed sirna product  (Thermo Fisher)

Journal: Scientific Reports

Article Title: Intestinal alkaline phosphatase is a receptor for cholesterol-lowering pentapeptide IIAEK and regulates cholesterol homeostasis in mice

doi: 10.1038/s41598-025-04722-w

Figure Lengend Snippet: IIAEK-human IAP complex interacts with cadherin-17. ( a ) SDS-PAGE analysis to identify the transmembrane target protein interacting with the human IAP- IIAEK (Ile-Ile-Ala-Glu-Lys) complex. ( b ) Identification of the transmembrane target protein interacting with the human IAP-IIAEK complex using nano LC–MS/MS and a Mascot search engine. ( c ) SDS-PAGE analysis to observe the interaction between the IIAEK-human IAP complex and cynomolgus cadherin-17. ( d ) SPR sensorgram of the interaction between cynomolgus cadherin-17 and immobilised human intestinal alkaline phosphatase (hIAP). ( e ) Effect of the addition of IIAEK on the SPR sensorgram of the interaction between cynomolgus cadherin-17 and immobilised hIAP. The original gels are presented in Supplementary Fig. 8.

Article Snippet: Subsequently, equal amounts of serum-free DMEM containing 240 nM CDH17 siRNA (Silencer ® Select Pre-designed siRNA Product, Ambion, 4392420, Thermo Fisher Scientific) or 240 nM Control siRNA (Silencncer ® Select Negative Control #1 siRNA, Ambion, 439084, Thermo Fisher Scientific) and serum-free DMEM containing siLentFectTM Lipid Reagent (BIO-RAD, 170–3361) were mixed and incubated at room temperature for 30 min to generate siRNA-siLentFect complexes.

Techniques: SDS Page, Liquid Chromatography with Mass Spectroscopy

Journal: Scientific Reports

Article Title: Intestinal alkaline phosphatase is a receptor for cholesterol-lowering pentapeptide IIAEK and regulates cholesterol homeostasis in mice

doi: 10.1038/s41598-025-04722-w

Figure Lengend Snippet: Effect of IIAEK on fluorescently labelled cholesterol uptake in Caco-2 cells with knockdown of human cadherin 17 ( CDH17 ). Caco-2 cells were cultured in 6-well plates for 14 days post-confluence. CDH 17 (20 nM) or negative control siRNA (20 nM) was introduced into the cells for 72 h. After 72 h of introduction, the cells were reintroduced with these siRNAs for 72 h. Subsequently, the cells were treated with serum-free medium containing 2 mM IIAEK (Ile-Ile-Ala-Glu-Lys) or the vehicle for 24 h. After removing the medium, the cells were incubated with a serum-free medium containing fluorescently labeled cholesterol (NBD-cholesterol) micelle for 1 h. After incubation, the cells were washed and cultivated to measure the fluorescent intensity of NBD-cholesterol. ( a ) CDH 17 mRNA levels. ( b ) Effect of IIAEK on NBD-cholesterol uptake in Caco-2 cells introduced with negative control siRNA. ( c ) Effect of IIAEK on NBD-cholesterol uptake in Caco-2 cells introduced with CDH17 siRNA. ( a – c ) n = 3/group. Data are presented as mean ± standard error of the mean (SEM). Statistical significance was calculated using an unpaired Student’s t -test. Asterisks indicate significant differences between the experimental groups (* p < 0.05).

Article Snippet: Subsequently, equal amounts of serum-free DMEM containing 240 nM CDH17 siRNA (Silencer ® Select Pre-designed siRNA Product, Ambion, 4392420, Thermo Fisher Scientific) or 240 nM Control siRNA (Silencncer ® Select Negative Control #1 siRNA, Ambion, 439084, Thermo Fisher Scientific) and serum-free DMEM containing siLentFectTM Lipid Reagent (BIO-RAD, 170–3361) were mixed and incubated at room temperature for 30 min to generate siRNA-siLentFect complexes.

Techniques: Knockdown, Cell Culture, Negative Control, Incubation, Labeling

Journal: Scientific Reports

Article Title: Intestinal alkaline phosphatase is a receptor for cholesterol-lowering pentapeptide IIAEK and regulates cholesterol homeostasis in mice

doi: 10.1038/s41598-025-04722-w

Figure Lengend Snippet: Hypothesis of how the specific interaction of IIAEK and IAP (IIAEK receptor) ameliorates cholesterol metabolism. The cholesterol-lowering pentapeptide IIAEK (Ile-Ile-Ala-Glu-Lys) specifically interacts with the GPI-anchored receptor intestinal alkaline phosphatase (IAP). Subsequently, the IIAEK-IAP dimer complex interacts with cadherin-17 to ameliorate cholesterol metabolism by suppressing cholesterol absorption, liver cholesterol, and serum cholesterol levels and promoting fecal cholesterol excretion.

Article Snippet: Subsequently, equal amounts of serum-free DMEM containing 240 nM CDH17 siRNA (Silencer ® Select Pre-designed siRNA Product, Ambion, 4392420, Thermo Fisher Scientific) or 240 nM Control siRNA (Silencncer ® Select Negative Control #1 siRNA, Ambion, 439084, Thermo Fisher Scientific) and serum-free DMEM containing siLentFectTM Lipid Reagent (BIO-RAD, 170–3361) were mixed and incubated at room temperature for 30 min to generate siRNA-siLentFect complexes.

Techniques: