Encapsulation Alleviates the Auto-browning of Epigallocatechin-3-gallate in Aqueous Solutions through Regulating Molecular Self-Aggregation Behavior

Catechins are widely recognized for superb antioxidant capability, but their application as food antioxidants is hindered by susceptibility to auto-browning under high-moisture conditions. Here, we proposed a strategy of ordered encapsulation with cyclodextrin-based metal–organic frameworks (CD-MOFs) to alleviate the auto-browning phenomenon of catechins while preserving their antioxidant capability and demonstrated the feasibility of this strategy via selecting epigallocatechin-3-gallate (EGCG) as a model. Even in aqueous solutions, EGCG@CD-MOFs still possessed delayed browning, in contrast with pristine EGCG, characterized by suppressed efficiencies on the generation of oxidative dimers (theasinensin A) and semiquinone radicals. Mechanism insights revealed that ordered encapsulation brought dual regulations on the self-aggregation behavior of EGCG: EGCG@CD-MOFs exhibited a gradual structural collapse from the framework toward irregular aggregates as O–K bonds broke progressively, which restricted molecular mobility of EGCG, and EGCG molecular conformations became constrained by the structure of EGCG@CD-MOFs as well as rich intermolecular forces, even after structural collapse

Additional file 6: of TNFα induces Ca2+ influx to accelerate extrinsic apoptosis in hepatocellular carcinoma cells

Figure S5. The role of TNFα-mediated Ca2+ influx in normal hepatic cells. (a) and (c) Confocal microscope analysis of [Ca2+]c level using fluorescent probe Fura-2/AM in QSG-7701 cells with treatment as indicated. TNFα: 100 ng/mL; siTRPM7: siRNA target TRPM7. (b) qRT-PCR and western blot analysis of TRPM7 mRNA and protein expression levels in QSG-7701 cells transfected with siRNA as indicated. (d) Analysis of Calpain activity after TNFα stimulation for 4 h in QSG-7701 cells with treatment as indicated. Data were shown as mean ± SD. All experiments were performed at least three times. * P < 0.05; ** P < 0.01. (JPEG 664 kb

Additional file 1: of TNFÎą induces Ca2+ influx to accelerate extrinsic apoptosis in hepatocellular carcinoma cells

Supplemental materials and Methods. Table S1. Primary antibodies used for western blot. Table S2. Sequence of primers and siRNA. Table S3. Public datasets used for bioinformatic analysis. (DOC 97 kb

Additional file 2: of TNFα induces Ca2+ influx to accelerate extrinsic apoptosis in hepatocellular carcinoma cells

Figure S1. (a) and (b) The relative mRNA and protein expression of TNFR1 and TNFR2 measured by qRT-PCR or Western Blot in SNU739 and HLF cells. (c) and (d) qRT-PCR and Western Blot analysis of TNFR1 and TNFR2 mRNA and protein expression levels in SNU739 and HLF cells transfected with siRNA as indicated. (e) TNFR1 protein expression were determined by Co-immunoprecipitation (Co-IP) and western blot in HCC cells as described. (f) and (g) qRT-PCR and western blot analysis of TRPM7 mRNA and protein expression levels in SNU739 and HLF cells transfected with siRNA as indicated. (h) The relative mRNA expression levels of TRPC1, TRPC6, TRPM2, TRPM3, TRPM7, TRPV4, TRPV5, TRPV6, TRPP2, and TRPP5 in HCC tumor tissues were analyzed in public microarray data TCGA downloaded from the Gene Expression Omnibus (GEO) database. (i) The interaction effects between TNFα and TRPM7 were determined by Co-immunoprecipitation (Co-IP) and western blot in HCC cells as described. Data were shown as mean ± SD. All experiments were performed at least three times. * P < 0.05; ** P < 0.01. (JPEG 945 kb

Additional file 5: of TNFα induces Ca2+ influx to accelerate extrinsic apoptosis in hepatocellular carcinoma cells

Figure S4. The level of extracellular calcium influx is positively correlated with TNFα-mediated apoptosis. (a) Confocal microscope analysis of [Ca2+]c level using fluorescent probe Fura-2/AM in 10 kinds of HCC cells with treatment as indicated. (b) Apoptosis analysis by flow cytometry 24 h after treatment as indicated. All experiments were performed at least three times. (ZIP 2184 kb

Additional file 4: of TNFα induces Ca2+ influx to accelerate extrinsic apoptosis in hepatocellular carcinoma cells

Figure S3. Cytosolic Ca2+ sensitized HCC cells to TNFα-induced apoptosis. (a) Apoptosis analysis by flow cytometry 24 h after treatment as indicated. (b) and (c) Confocal microscope analysis of [Ca2+]c level using fluorescent probe Fura-2/AM in HLF and QSG-7701 cells with treatment as indicated. TNFα (50 ng/mL) + Iono: 50 ng/mL TNFα combined with 1 μM ionomycin, TNFα (100 ng/mL) + Iono: 100 ng/mL TNFα combined with 1 μM ionomycin, TNFα (200 ng/mL) + Iono: 200 ng/mL TNFα combined with 1 μM ionomycin. TNFα (50 ng/mL) + IP3: 50 ng/mL TNFα combined with 10 μM IP3; TNFα (100 ng/mL) + IP3: 100 ng/mL TNFα combined with 10 μM IP3; TNFα (200 ng/mL) + IP3: 200 ng/mL TNFα combined with 10 μM IP3; (d)-(f) Apoptosis analysis by flow cytometry 24 h after treatment as indicated. Data were shown as mean ± SD. All experiments were performed at least three times. * P < 0.05; ** P < 0.01. (ZIP 2448 kb