Table 3. Main characteristics and effects of protocols on oxidative stress and the expression of other proteins.
| Author | Experimental model | Groups | Bleaching gel protocol | Additional protocol | Period of analysis | Methods for outcome assessment | Main results | |
|---|---|---|---|---|---|---|---|---|
| In vitro studies using enamel/dentin discs and MSCs | ||||||||
| Dias et al. 2023 [62] | Enamel/dentin discs, and MDPC-23 cells | Negative control, PCP, 10% HP, 10% HP + PCP, 20% HP, 20% HP + PCP, 35% HP, 35% HP + PCP | 10%, 20% and 35% HP for 45 min | 10 μL of PCP (topically): before HP | 1 h | Oxidative stress: carboxy-H2DCFDA fluorescence | The groups where PCP was used before applying the bleaching gels showed lower oxidative stress in MDPC-23 cells. | |
| de Oliveira Ribeiro et al. 2022 [60] | Enamel/dentin discs, and MDPC-23 cells | Negative control, 35% HP, 10% HP, 10% HP + 2 mg/mL MnO2, 10% HP + 6 mg/mL MnO2, 10% HP + 10 mg/mL MnO2 | 10% HP for 45 min | 20 μL of 10% HP mixed with 2 mg/mL, 6 mg/mL, or 10 mg/mL MnO2 (topically): 45 min (3 applications of 15 min) | 30 min | Oxidative stress: carboxy-H2DCFDA fluorescence | MnO2 increased the degradation kinetics of the HP molecule, consequently reducing the cellular oxidative stress index. | |
| Ribeiro et al. 2022 [59] | Bovine enamel and dentin discs, and MDPC-23 cells | CG: untreated; G1: 35% HP; G2: 35% HP + 2 mg/mL MnO2; G3: 35% HP + 6 mg/mL MnO2; G4: 35% HP + 10 mg/mL MnO2 | 45 min (3 applications of 15 min) | 2, 6, and 10 mg/mL of MnO2 was incorporated into the bleaching gel: for 45 min (3 × 15 min) | Immediately | Oxidative stress: carboxy-H2DCFDA fluorescence; | The addition of MnO2 to 35% HP reduced oxidative stress. | |
| Ortecho-Zuta et al. 2019 [57] | Enamel/dentin discs, and MDPC-23 cells | Untreated, HP: 35% HP, 35% HP + HRP | 35% HP for 45 min (3 applications of 15 min) | 1 mL of 35% HP mixed with 10 mg HRP (topically): for 45 min (3 applications of 15 min) | 1 h | Oxidative stress: carboxy-H2DCFDA fluorescence | HRP combined with HP reduced oxidative stress. | |
| Soares et al. 2019 [23] | Enamel/dentin discs, and MDPC-23 cells | Untreated, HP: 35% HP, 35% HP + FS, 35% HP + MC, 35% HP + PR, 35% HP + CT | 35% HP for 45 min (3 applications of 15 min) | 40 mL of 35% HP mixed with 1 mg FS, MC, PR, or CT (topically): for 45 min (3 applications of 15 min) | 1 and 24 h | Oxidative stress: carboxy-H2DCFDA fluorescence | All chemically activated groups showed reduced oxidative stress. | |
| In vitro studies using MSCs | ||||||||
| Huang et al. 2019 [56] | HDPCs | NC: negative control, HP (50, 150, 250, 350 μM), HP+NAC: HP (250 μM) + NAC (2.5 mM), HP+CsA: HP (250 μM) + CsA (2 μM), siRNA-CypD: CypD siRNA targeting human PPIF, siRNA-CypD+HP: CypD siRNA targeting human PPIF + HP (250 μM) | 250 μM HP for 24 h | NAC or CsA reagents for 24 h; CypD siRNA-PPIF for 24 h | 24 h | Bockade of CypD: western lot analyses | NAC, CsA, and CypD siRNA-PPIF decreased the CypD expression, increasing mitochondrial membrane potential. | |
| Kim et al. 2017 [54] | HDPCs | Control: untreated, 180 μM HP, HP + 50 μM IAA, HP + 100 μM IAA, HP + 150 μM IAA, HP + 200 μM IAA, HP + 250 μM IAA, HP + 300 μM IAA | 180 μM HP for 24 h | IAA: ranging from 1 to 300 μM | 6 h | Expression of apoptotic (BAX and p53) and antiapoptotic (BCL-2 and ATF5) genes: PCR; ROS detection: fluorescence; Nrf2 and HO-1 expression: densitometric analysis; cell cycle: flow cytometry | IAA treatment protected HDPCs against HP-induced oxidative stress via increased expression of Nrf2 and HO-1. Moreover, IAA treatment rescued cell cycle and prevented apoptosis. | |
| Kim et al. 2017 [55] | HDPCs | Control: untreated, HP: 300 µM HP, CA: 20 µM CA, CA + HP: 20 µM CA and then 300 µM HP, CoPP: 20 µM CoPP, CoPP+HP: 20 µM CoPP and then 300 µM HP | 300 µM HP for 24 h | 20 µM CA: for 24 h, before HP | 6 h | ROS measurement: fluorescence analysis; expression of proteins (HO-1, Nrf2): western blot | Pre-treatment with CA protected HDPCs against HP-induced oxidative stress by enhancing the expression of HO-1 through the Nrf2 signaling pathway. | |
| Jeong et al. 2010 [46] | HDPCs | 1 mM HP, HP + 5 μM sappanchalcone, HP + 10 μM sappanchalcone, HP + 20 μM sappanchalcone, HP + 40 μM sappanchalcone, HP + 40 μM sappanchalcone + 100 μM SnPP, 100 μM SnPP, HP + 20 μM CoPP, positive control: 20 μM | 1 mM HP for 12 h | 5–40 μM sappanchalcone: 12 h | 12 h | ROS measurement: fluorescence | CoPP and 20 and 40 μM sappanchalcone inhibited ROS production. | |
| Lee et al. 2013 [24] | HDPCs | Control: untreated, 1mM HP, 1mM HP + 2.5 μM butein, 1mM HP + 5 μM butein, 1mM HP + 10 μM butein, 1mM HP + 20 μM butein, 1mM HP + 20 μM CoPP, 1mM HP+ 100 μM SnPP | 1 mM HP for 12 h | Butein, CoPP and SnPP: for 8 h or until 24 h | 8 h for ROS detection; 0, 3, 6, 12, 18 and 24 h for HO-1 expression | ROS measurement: fluorescence; HO-1 expression: western blot; Nrf2 expression: immunofluorescence, western blot | Butein inhibited HP-induced ROS production, presumably through JNK Nrf2/ARE-dependent HO-1 expression. | |
| Lee et al. 2013 [49] | HDPCs | HP+Ad/PPARγ: HP+Ad PPARγ, HP+Ad/LacZ: control, HP | 150 μmol HP for 12 d | Ad/PPARγ virus: a dose of 100 multiplicity of infection (MOI) for 24 h | 12 d | ROS measurement: flow cytometry; expression of antioxidant molecules: western blot | PPARγ in pulp cells removed cellular ROS under oxidative stress. | |
| Lee et al. 2013 [50] | HDPCs | Control: untreated, HP: 500 µM HP, 500 µM HP + 5 µM sulfuretin, 500 µM HP + 10 µM sulfuretin, 500 µM HP + 20 µM sulfuretin, 500 µM HP + 40 µM sulfuretin, sulfuretin: 5-40 µM sulfuretin, HP+CoPP: 500 µM HP + 20 µM CoPP, positive control: 20 µM CoPP | 500 µM HP for 12 h | 5-40 µM sulfuretin: for 12 h, before HP | 12 h | ROS measurement: fluorescence; expression of HO-1: western blot | Pre-treatment with sulfuretin suppressed cellular damage from oxidation caused by HP in HDPCs, presumably through HO-1 expression. | |
| Lee et al. 2013 [51] | HDPCs | HP: 150 µM HP, 150 µM HP + 15 µM pachymic acid, 150 µM HP + 5 µM pachymic acid, untreated cells | 150 µM HP for 1, 3, 5, 7 and 12 d | 15 µM pachymic acid: before 1 h prior to incubation with HP | 1, 3, 5, 7 and 12 d | Expression of inflammatory molecules and odontoblast differentiation level: western blot | The pachymic acid showed anti-inflammatory function and odontoblast differentiation via HO-1 pathway. | |
| Choi et al. 2012 [48] | hDPSCs | HP: 200 µM HP, 200 µM HP + 2 µM SOD1, 200 µM HP + 2 µM LMWP-SOD1, 2 µM SOD1, 2 µM LMWP-SOD1, untreated | 200 µM HP for 2 h | 2 µM LMWP-SOD1: for 3 h, before HP | 3 and 28 d | Transduction of LMWP-SOD1 and quantification of p53 and p21Cip1: western blot | LMWP-SOD1 conjugates were effective for attenuating cellular senescence and reversing osteoblastic differentiation of hDPSCs caused by oxidative stress inhibition. | |
Ad, adenovirus; ARE, antioxidant response element; ATF-5, activating transcription factor 5; BAX, BCL2 associated X; BCL-2, B-cell lymphoma protein 2; CA, cinnamaldehyde; CG, control group; CoPP, cobalt protoporphyrin; CsA, cyclosporine A; CT, catalase; FS, ferrous sulfate; G, group; HDPCs, human dental pulp cells; hDPSCs, human dental pulp stem cells; HO-1, heme oxygenase 1; HP, hydrogen peroxide; HRP, horseradish peroxidase; IAA, indole-3-acetic acid; JNK, c-Jun NH2-terminal kinase; LMWP, low-molecular weight protamine; MC, manganese chloride; MDPC-23, mouse dental papilla Cell-23; MOI, multiplicity of infection; MnO2, manganese oxide; MSCs, mesenchymal stem cells; NAC, N-acetylcysteine; NC, negative control; Nrf2, nuclear factor erythroid 2-related factor 2; p 53, PCP, polymeric catalyst primer; PCR, polymerase chain reaction; PPARγ, proliferator-activated receptor gamma; PPIF, peptidylprolyl isomerase F; PR, peroxidase; ROS, reactive oxygen species; siRNA-CypD, cyclophilin D small interfering ribonucleic acid; SnPP, tin protoporphyrin; SOD1, superoxide dismutase 1.