Table 2.
Included in vitro studies. Symbols of greater than (>), less than (<) or equal to (=) are used to compare the results of the tested groups.
| Studies | Materials | Type of Exposure | Parameters | Methods | Results |
| Alliot-Licht et al. (1994) [56] |
Calcium hydroxide (CH) Hydroxyapatite (HAp) |
CH particles sterilized by heating (180 °C-1 h); direct contact (materials powder in culture medium) |
Cell morphology | Light microscopy (at 3 & 5 days) | CH inhibited pulp fibroblasts growth (<cell density than control; subjective observation) HAp did not affect the cell density (≈cell density as the control; subjective observation) |
| Phagocytotic activity | SEM (at day 5) | Close contact of CH particles with fibroblasts’ membrane. HAp particles were closely bound to cell membrane or internalized by the cells. | |||
| TEM (at day 5) | Cells cultured in the presence of CH exhibited ghost cells and electron-dense spherical vesicles in the cytoplasm of living cells. TEM revealed HAp particles within the cells. | ||||
| Cell proliferation | DNA synthesis (at 1, 2, 3 & 4 days) | CH and HAp delayed the proliferation of cells at all time points. | |||
| Protein synthetic activity (at 6 days) | CH < incorporation of [3H]-leucine and [3H]-proline by pulp fibroblasts at day 6. HAp > incorporation of [3H]-leucine and [3H]-proline by the pulp fibroblasts at day 6. |
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| Cell differentiation | ALP activity (at 8 days for CH; at 5 & 8 days for HAp) | CH inhibited ALP activity of pulp fibroblasts at day 8. HAp inhibited ALP activity of pulp fibroblasts at 5 and 8 days. |
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| Min et al. (2007) [57] | Portland cement (PC) Portland cement with bismuth oxide (BPC) |
Indirect contact (SET materials) |
Cell viability | MTT assay (at 12, 24, 48 & 72 h) | PC > BPC at 12 and 24 h PC ≈ BPC at 48 and 72 h |
| Nitric oxide production | Griess reaction(at 12, 24, 48 & 72 h) | BPC > nitrite production than PC at 12 and 24 h. PC ≈ BPC nitrite production at 48 and 72 h. |
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| Ho-1 and iNOS | RT-PCR (at 12, 24, 48 & 72 h) | Ho-1: PC < BPC at all study periods iNOS: PC < BPC at all study points |
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| Min et al. (2007) [58] | Portland cement (PC) Fuji-II LC (Fuji-II, GC) Zinc-oxide Eugenol (IRM; Dentsply-Sirona) CH cement Dycal (Dentsply-Sirona) |
Direct and indirect contact (SET materials) |
Cell morphology | SEM (at 24 h) | PC: showed flattened cells close to one another and spreading across the substrate. Fuji-II, IRM, and Dycal: no living cells were seen. |
| Cell viability | MTT assay (at 12, 24, 48 & 72 h) | PC ≈ control at all study periods. PC > Fuji-II, Dycal and IRM at all study periods. Control > Fuji II LC, IRM, and Dycal at all study points. |
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| Cell differentiation | RT-PCR (ON, DSPP) (at 7 days) | ON: PC ≈ positive control group. DSPP: PC stimulated mineralization but less than the positive control. |
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| Laurent et al. (2008) [59] |
Ca3SiO5 cement (CS) Dycal (Dentsply-Sirona) Pro-Root MTA (MTA; Dentsply-Sirona) |
Indirect contact * (SET materials) [ISO-Standard–(Nr. Not mentioned)] |
Cell viability | MTT assay (at 24 h) | No contact (disk diffusion): CS ≈ MTA ≈ Dyc Indirect contact (eluates from materials): Cs ≈ MTA > Dycal |
| Cell differentiation | Immunohistochemistry (at 4 weeks) | MTA and CS expressed Nestin and Collagen I at a similar level as the control group. Both materials generated mineral deposits at a similar level as the control group. | |||
| Genotoxicity | Ames test | CS does not induce reverse mutations with/without the S9 metabolic activation system. | |||
|
Micronuclei test
Comet assay |
CS generated lymphocytes with micronuclei ≈ as the negative control. | ||||
| CS generated DNA in the tail ≈ as the negative control and < than the cytotoxic control. | |||||
| Min et al. (2009) [60] | Radiopaque Portland cement (RPC) Portland cement (PC) IRM (Dentsply-Sirona) |
Direct and indirect contact (SET materials) |
Cell morphology | SEM (at 48 h) | PC and RPC: Spread and flattened HDPCs. The density and characteristics of the HDPCs in both groups were similar to that on control samples. IRM: no living cells were seen in contact with the |
| Cell differentiation | ALP activity (at 1, 3, 7 & 14 days) |
1d: PC and RPC > control; 3d, 7d, 14d: control > PC and RPC 2wk and 3wk: PC and PCR > control DSPP: PC and RPC > control at day 14; OCN: control ≈ Pc and RPC at all study periods. |
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| ARS staining (1, 2 & 3 wk) | |||||
| RT-PCR (DSPP, ON) (at 1, 3, 7 & 14 days) | |||||
| Lee et al. (2014) [61] | ProRoot MTA (MTA; Dentsply-Sirona) α-tricalcium phosphate-based cement (α-TCP) |
Direct and indirect contact (SET materials) |
Cell morphology | SEM (at 72 h) | hDPCs in contact with MTA and α-TCP were well-spread and flattened. |
| Cell viability | MTT assay (at 1, 2, 3, 7 & 14 days) | MTA and α-TCP ≈ control until day 7 α-TCP > MTA at 14d; α-TCP ≈ control |
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| Cell differentiation | Western blot (DSPP, DMP-1 and ON) (at 3 days) | α-TCP ≈ MTA for DSPP, DMP-1 and ON. | |||
| ARS staining (at 14 days) | α-TCP ≈ MTA for DSPP, DMP-1 and ON. | ||||
| Immunofluorescence (DSPP, DMP-1 and ON) (at 7 days) | α-TCP and MTA induced higher protein signals than the control group. | ||||
| Bortoluzzi et al. (2015) [62] | Biodentine (Bd; Septodont) Theracal LC (Th; Bisco) MTA Angelus (MTA-A; Angelus) |
Indirect contact * (SET materials) |
Cell viability | XTT assay (direct and indirect eluate evaluations) Flow cytometry– Annexin V-PI (4 weekly cycles) |
Direct evaluation: 1st cycle: control > Bd > MTA-A and Th 2nd cycle: control > Bd ≈ MTA-A > Th 3rd cycle: control ≥ MTA-A ≥ Bd > Th 4th cycle: control ≈ Bd ≈ MTA-A > Th Indirect eluate evaluation: 1:1&1:10 dilutions: control > MTA-A ≈ Bd > Th; 1:100 dilution: control ≈ MTA-A ≈ Bd > Th Percentage of healthy, non-apoptotic and non-necrotic cells: control > MTA-A ≈ Bd > Th Th was the most cytotoxic material causing apoptosis and necrosis. |
| Cell differentiation | qRT-PCR (DSPP, OCN, BSP, RUNX 2, DMP-1 and ALP) (at 7 days) | ALP; OCN; BSP; DSPP; DMP-1: Bd and MTA-A > control ≈ Th RUNX 2: Bd ≈ MTA-A ≈ control ≈ Th |
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| ALP activity (at 14 days) | Bd ≈ control > MTA-A > Th | ||||
| ARS and TEM (at 21 days) | Bd > control > MTA-A > Th | ||||
| Niu et al. (2015) [63] |
ProRoot MTA (MTA; Dentsply-Sirona) Quick-Set2 (Qs; Avalon Biomed Inc) |
Direct and indirect contact (SET materials) |
Cell viability | Flow cytometry– Annexin V-PI (3 weekly cycles) Leakage of cytosolic enzyme (3 weekly cycles) Caspase-3 acitivity (3 weekly cycles) Oxidative stress (3 weekly cycles) |
Number of healthy cells: 1st cycle: control > Qs > MTA (p < 0.001) > IRM 2nd cycle: control > Qs ≈ MTA > IRM; 3rd cycle: control ⩾ MTA ⩾ Qs > IRM Percentage of cytotoxicity: 1st cycle: IRM > MTA > Qs > control; 2nd and 3rd cycles: IRM > MTA ≈ Qs > control Relative caspase-3 activity: 1st cycle: IRM > MTA > Qs > control; 2nd cycle: IRM > MTA > Qs > control 3rd cycle: IRM > MTA ≈ Qs > control Oxidative stress: 1st cycle: IRM > MTA > Qs > control; 2nd cycle: IRM > MTA ≈ Qs > control 3rd cycle: IRM > MTA ≈ Qs ≈ control |
| Cell proliferation | MTT assay (3 weekly cycles)Cellular DNA content (3 weekly cycles) | 1st cycle: control > Qs > MTA > IRM 2nd cycle: control > Qs ≈ MTA > IRM 3rd cycle: control > Qs > MTA > IRM DNA content: 1st cycle: control > Qs > MTA > IRM 2nd cycle: control > Qs ≈ MTA > IRM 3rd cycle: control > Qs ≈ MTA > IRM |
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| Öncel Torun et al. (2015) [64] |
iRoot BP Plus (iBP; Innovative Bioceramix) White MTA Angelus (MTA-A; Angelus) |
Indirect contact (SET materials) | Cell viability | XTT assay (24, 48 & 72 h) |
24 h; 1:1 and 1:2 dilutions: iBP > W-MTA-A; 1:5 and 1:10 dilutions: iBP ≈ MTA-A 48 h; 1:1 dilution: iBP > W-MTA-A; 1:2, 1:5 and 1:10 dilutions: iBP ≈ MTA-A 72 h; all concentrations: iBP ≈ MTA-A |
| Cell differentiation | qRT-PCR (BMP-2, ON, BSP, OPN, DSPP, Col I A1, HO-1 at 24 & 72 h) |
BMP-2: 24 h 1:1 and 1:5 dilutions MTA-A > iBP; 1:2 dilution: iBP ≈ MTA-A 72 h 1:1, 1:2 and 1:5 dilutions MTA-A > iBPON: 24 h 1:1 and 1:5 dilutions iBP > MTA-A; 1:2 diution: iBP ≈ MTA-A 72 h: 1:1 dilution iBP > MTA-A; 1:2 and 1:5 dilutions MTA-A > iBP BSP: 24 h: 1:1 dilution MTA-A > iBP; 1:2 and 1:5 diutions: iBP ≈ MTA-A 72 h: 1:1 and 1:2 dilutions MTA-A > iBP; 1:5 diution: iBP ≈ MTA-AOPN: 24 h: 1:2 dilution iBP > MTA-A; 1:1 and 1:5 dilutions: iBP ≈ MTA-A 72 h: 1:1 and 1:5 dilutions MTA-A > iBP; 1:2 dilution: iBP ≈ MTA-A DSPP: 24 h: 1:1 dilution iBP > MTA-A; 1:2 and 1:5 dilutions: iBP ≈ MTA-A 72 h: 1:2 dilution iBP> MTA-A; 1:1 dilution MTA-A > iBP; 1:5 dilution: iBP ≈ MTA-ACol I A1: 24 h: 1:1 dilution iBP > MTA-A; 1:2 and 1:5 dilutions: iBP ≈ MTA-A 72 h: 1:1 and 1:2 dilutions iBP > MTA-A; 1:5 dilution: iBP ≈ MTA-AHO-1: 24 h 1:1 and 1:2 dilutions MTA-A > iBP; 1:5 dilution: iBP ≈ MTA-A 72 h: 1:1, 1:2 and 1:5 dilutions MTA-A > iBP |
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| Zhang et al.(2015) [65] | iRoot BP Plus (iBP; Innovative Bioceramix) ProRoot MTA (MTA; Dentsply-Sirona) |
Indirect contact (SET materials) (ISO 10993-5) |
Cell Viability | Flow cytometry– Annexin V-PI |
iBP ≈ MTA ≈control |
| Cell Migration | Wound-healing at 24 h | iBP ≈ MTA > control | |||
| Transwell assay at 24 h | iBP = MTA > control | ||||
| Cellular adhesion and motility | Western-Blot (at 5, 10, 30 & 60 min) Cell Immunofluorescence assay (at 1 h) |
iBP led to phosphorylation of p38 MAPK, ERK 1/2, JNK, Akt, and FGFR | |||
| iBP significantly increased p–focal adhesion kinase (p-FAK), p-paxillin, and vinculin Cells treated with iBP showed highly organized and stretched stress fiber assembly | |||||
| Chung CJ et al. (2016) [66] | Dycal (Dy; Dentsply-Sirona) Endocem Zr (E-Zr, Maruchi) White ProRoot MTA (MTA; Dentsply-Sirona) Retro-MTA (R-MTA; Bio MTA) |
Indirect and direct contact; SET (s) and FRESH (f) materials |
Cell morphology/attachment | Phase microscopy (at 3 & 7 days) SEM (at 3 & 7 days) |
3d: MTA > cell morphology and attachement than R-MTA and E-Zr 7d: MTA, R-MTA and E-Zr sowed good cell morphology and attachement Dycal treated cells were dead after 3 and 7 days. Dycal was not further used |
| Cell viability | XTT assay (at 3 & 7 days) | 3 d: control ≈ MTA (s) ≈ MTA (f) > R-MTA (s) ≈ R-MTA (f) > E-Zr (s) ≈ E-Zr (f) 7 d: MTA(f) > control ≈ MTA (s) ≈ R-MTA (s) ≈ R-MTA (f) ≈ E-Zr (f) > E-Zr (s) |
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| Angiogenic properties | ELISA (VEGF, angiogenin, FGF-2) (at 3 & 7 days) |
VEGF24 h: control ≈ MTA (s) ≈ R-MTA (s) ≈ R-MTA (f) ≥ MTA (f) ≈ E-Zr (s) > E-Zr (f)
VEGF72 h: MTA (s) ≈ MTA (f) ≈ R-MTA (s) ≈ E-Zr (s) ≥ control ≥ R-MTA (f) ≈ E-Zr (f) Angiogenin 24 h: control≈ MTA(s) ≥ R-MTA (s) > MTA(f) > R-MTA (f) ≈ E-Zr(s)>E-Zr (f) Angiogenin 72 h: R-MTA (s) ≈ R-MTA (f) ≈ control > MTA(s) ≈ MTA (f) > E-Zr (s) ≈ E-Zr (f) FGF-2 24 h and 72 h: no difference among materials and control |
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| Daltoé M et al. (2016) [67] | Biodentine (Bd; Septodont) White ProRoot MTA (MTA; Dentsply-Sirona) |
Indirect contact (SET materials) (ISO 10993-5) |
Cell Viability | MTT assay (at 24 & 48 h) | 24 h: control ≈ MTA_1:100 ≈ Bd_1:100 > MTA_1:10, Bd_1:10, Bd_1:1 and MTA_1:1 48 h: control ≈ MTA_1:100 ≈ Bd_1:100 > MTA_1:10, Bd_1:10, MTA_1:1 and Bd_1:1 |
| Cell differentiation | qRT-PCR (SPP1, IBSP, DSPP, ALP 1, DMP-1 and RUNX 2 (at 24 & 48 h) | SPP1 & ALP1 & RUNX2 at 24 h: Bd and MTA ≈ control SPP1 & ALP1 & RUNX2 48 h: Bd and MTA > control IBSP & DSPP & DMP1: 24 h and 48 h: no expression |
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| Widbiller M et al. (2016) [68] | Biodentine (Bd; Septodont) GI Ketac-Molar (KM; 3M) ProRoot MTA (MTA; Dentsply-Sirona) |
Indirect and direct contact (SET materials) |
Cell morphology/attachement (only Bd) | SEM (at 24 h) | Biodentine: cells showed adhesion to and spreading onto the cement surface * Not done for the other materials. |
| Cell viability | MTT assay (at 1, 3, 5, 7, 10 & 14 days) | Bd > other materials and control at 1, 3, 5 and 7 d; Bd ≈ MTA > control > KM at 10 and 14 d MTA < viability than the control at 1d; MTA ≈ cell viability as the control at 3-5-7d; KM < cell viability than all the materials tested and the control at all time points |
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| Cell differentiation Not performed on KM |
RT-qPCR (ALP, Col-I A1, DSPP, RUNX 2) (at 7, 14 & 21 days) | Col-I A1 & ALP: upregulated at 7d, especially for MTA, and decreased steadily until 21d DSPP: upregulated for MTA and BD at 14 and 21d RUNX2: downregulated for MTA and BF throughout the whole study period |
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| ALP activity (at 3, 7 & 14 days) | ALP activity was downregulated for Bd at all times: MTA ≈ control > Bd | ||||
| Jeanneau C et al. (2017) [28] | Biodentine (Bd; Septodont) Theracal LC (Th; Bisco) Xeno III (Dentsply-Sirona) |
Indirect contact (SET materials) |
Cell proliferation | MTT assay (3, 5, & 7 days) | Bd_0.05 cm2/mL > Th_ 0.05 cm2/mL (p < 0.05) ≈ control at 3, 5 and 7 days Bd_0.5 cm2/mL > Th_ 0.5 cm2/mL (p < 0.05) ≈ control at 3, 5 and 7 days |
| Cell differentiation | Immunofluorescence (DSP and Nestin at day 7) |
Bd increased the expression of both markers, while Th had no effect | |||
| Inflammatory effect | ELISA (IL-8; 24 and 48 h) |
IL-8 expression at 24 h: Th_0.05 cm2/mL > Bd_0.05 cm2/mL ≈ control
IL-8 expression at 48 h: Th_0.05 cm2/mL > Bd_0.05 cm2/mL > control |
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| Jun S-K et al. (2017) [69] | Activa Bioactive (Activa; Pulpdent) Dycal (Dy; Dentsply-Sirona) Theracal LC (Th; Bisco) |
Indirect contact (SET materials) (ISO 10993-12) |
Cell viability | MTS assay (at 24 h) Live/dead assay (at 24 h) |
3.125% eluates: Dy > Th > Activa ≈ control; 6.25% eluates: Dy > Th ≈ Activa ≈ control 12.5% eluates: Dy ≈ Th ≈ Activa ≈ control; 25% eluates: Dy < Activa < Th < control 50% eluates: Dy ≈ Activa < Th < control 50% eluates: Dy < Activa < Th < control |
| Cell differentiation | ALP (at days 14 and 21) | 14 d: Th > Dy > Activa ≈ Osteogenic medium 21 d: Th ≈ Dy > Activa > Osteogenic medium |
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| ARS (at 21 days) | Th ≈ Dy > Activa ≈ Osteogenic medium | ||||
| Lee B-N et al. (2017) [70] | ProRoot MTA (MTA; Dentsply-Sirona) Theracal LC (Th; Bisco) |
Indirect contact (SET materials) |
Cell viability | WST-1 assay (at 24 h) | 100% concentration: Th > MTA; At 50%, 25% and 10% dilutions: Th ≈ MTA At 100% MTA: cell viability < 70% and significantly lower than Th. |
| Cell differentiation | RT-PCR (DSPP, DMP-1 at 1 & 3d) Q-PCR(DSPP, DMP-1 at 2, 5 & 7d) ALP staining (at day 7) ARS (at day 14) |
DSPP 1 d: MTA > Th ≈ control; 3 d: MTA ≈ Th > control DMP-1 at 1 and 3d: MTA ≈ Th ≈ control DSPP & DMP-1: upregulated for both materials, especially at day 7. MTA > Th ≈ control MTA > Th > control |
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| Mestieri LB et al. (2017) [71] | White MTA Angelus (MTA-A; Angelus) White Portland Cement (PC; Votoran) |
Indirect contact (SET materials) |
Cell viability | MTT assay | 1:2, 1:3, 1:4 and 1:8 dilutions: control > W-MTA-A >W-PC 1:6 dilution: MTA-A ≈ control > PC |
| Trypan blue assay | 1:2 dilution: control > MTA-A > PC 1:3 dilution: control > PC > MTA-A 1:4 and 1:6 dilutions: control > PC > MTA-A 1:8 dilution: control > PC ≈ MTA-A |
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| Cell Differentiation | ALP activity (at 1, 3 & 7d) | 1, 3 and 7 d: MTA-A ≈ PC ≈ control | |||
| Rodrigues EM et al. (2017) [72] | MTA-Plus (MTA-P; Prevest Denpro) White MTA Angelus (MTA-A; Angelus) |
Indirect contact (SET materials)(ISO-10993) |
Cell viability | MTT Assay Flow cytometry– Annexin V-PI |
1:2 concentration: MTA-P > MTA-A ≈ control 1:4 and 1:8 concentrations: MTA-P ≈ MTA-A > control MTA-A > live cells than MTA-P ≈ control MTA-A > necrotic cells than MTA-P > control |
| Cell differentiation | ALP activity (at 1, 3 & 7 days) | MTA-A < control < MTA-P after 7 days. | |||
| ARS (14d) | MTA-A > MTA-P > control | ||||
| qRT-PCR (BMP2, OC, ALP) | Day 1_BMP2 & OC: MTA-A > MTA-P > control; ALP: MTA-A ≈ MTA-P < control Day 3_ BMP2: MTA-A > MTA-P > control; OC & ALP: MTA-A ≈ MTA-P < control |
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| Sun Y et al. (2017) [73] | Biodentine (Bd; Septodont) iRoot FS (iFS; Innovative Bioceramix) |
Indirect contact (SET materials) |
Cell proliferation | CCK-8 assay (1, 3 & 7 days) | 1 d: Bd_0.2 mg/mL ≈ Bd_2 mg/mL ≈ iFS_0.2 mg/mL ≈ iFS_2 mg/mL ≈ control (p ≥ 0.05) 3 d: Bd_0.2 mg/mL ≈ Bd_2 mg/mL ≈ iFS_0.2 mg/mL ≈ iFS_2 mg/mL > control 7 d: Bd_0.2 mg/mL ≈ iFS_0.2 mg/mL > Bd_2 mg/mL ≈ iFS_2 mg/mL > control |
| Cell migration (24 h) |
Wound healing assay
Transwell migration assay |
iFS_0.2 mg/mL > iFS_2 mg/mL > control > Bd_0.2 mg/mL > Bd_2 mg/mL | |||
| iFS_0.2 mg/mL > iFS_2 mg/mL > control > Bd_0.2 mg/mL > Bd_2 mg/mL | |||||
| Cell differentiation | ALP activity (at 7, 14 d) | 7 d: iFS_0.2 mg/mL ≈ iFS_2 mg/mL ≈ Bd_0.2 mg/mL > Bd_2 mg/mL > control | |||
| 14 d: iFS_0.2 mg/mL > Bd_0.2 mg/mL > Bd_2 mg/mL ≈ iFS_2 mg/mL > control | |||||
| ARS (at 21 d) | 21 d: iFS_0.2 mg/mL > Bd_0.2 mg/mL ≈ Bd_2 mg/mL ≈ iFS_2 mg/mL ≈ control | ||||
| qRT-PCR (Col I and OCN) (at 1, 7 & 14 d) | 1 d: Col I control ≥ all materials OCN iFS_2 mg/mL ≥ iFS_0.2 mg/mL ≈ Bd_0.2 mg/mL ≈ Bd_2 mg/mL ≈ control 7 d: Col I control > iFS_0.2 mg/mL > Bd_0.2 mg/mL > Bd_2 mg/mL > iFS_2 mg/mL OCN iFS_0.2 mg/mL > control ≈ iFS_2 mg/mL > Bd_2 mg/mL ≥ Bd_0.2 mg/mL |
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| 14 d: Col I iFS_0.2 mg/mL > Bd_0.2 mg/mL ≈ Bd_2 mg/mL ≥ control ≥ iFS_2 mg/mL OCN iFS_0.2 mg/mL ≈ iFS_2 mg/m ≥ control ≥ Bd_0.2 mg/mL ≥ Bd_2 mg/mL | |||||
| Tomás -Catalá et al. (2017) [74] | MTA-repair HP Angelus (MTA-HP; Angelus) NeoMTA-Plus (N-MTA-P; Avalon Biomed Inc) White MTA Angelus (W-MTA; Angelus) |
Indirect and direct contact (SET materials) (ISO 10993-5) | Cell morphology | SEM-EDX (direct contact, 72 h) |
Cells attached and merged in all three materials, more cell monolayer structures were evident on the surface of W-MTA. EDX revealed MTA-HP ≈ N-MTA-P ≈ W-MTA in %weight of Ca, C and O. |
| Cell Viability (24, 48 & 72 h) | MTT assay | 24 h all dilutions: MTA-HP ≈ N-MTA-P ≈ W-MTA ≈ control 48 h undiluted extract: MTA-HP ≈ W-MTA > control 48 h 1:2 dilution: MTA-HP ≈ N-MTA-P ≈ W-MTA ≈ control 48 h 1:4 dilution: W-MTA > control ≈ MTA-HP > N-MTA-P 72 h undiluted extract: W-MTA > N-MTA-P > MTA-HP > control 72 h 1:2 dilution: MTA-HP ≈ N-MTA-P ≈ W-MTA ≈ control 72 h 1:4 dilution: MTA-HP < control ≈ N-MTA-P ≈ W-MTA |
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| Cell migration (24 & 48 h) | Wound healing–scratch assay |
N-MTA-P < control for all dilutions and time points
MTA-HP-A > control at 24 h_1:1/1:2 dilutions but < control at 48 h W-MTA-A > control at 24 h_all dilutions but < control at 48 h |
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| Collado-González M et al. (2018) [75] | GI Equia Forte (EF; GC) GI Ionostar Molar (IoM;Voco) |
Indirect and direct contact (SET materials) (ISO 10993-5) |
Cell morphology (indirect contact, 24 h) |
Confocal microscopy (cytoskeletal F-actin) |
1:1 extracts EF ≈ control (an organized and stretched stress fiber) 1:1 extracts IoM < control (cell numbers and stretched stress fiber) |
| Cell morphology (direct contact, 72 h) |
SEM | EF > IoM (cell attachment, morphology and growth) | |||
| Cell Viability (at 24, 48 & 72 h) | MTT assay | 24 h all concentrations: Control > EF ≈ IoM 48 h 1:1 dilution: Control ≈ IoM > EF; 48 h 1:2 dilution: IoM ≈ EF ≈ control 48 h 1:4 dilution: IoM ≈ EF ≈ control 72 h 1:1 dilution: EF ≈ control > IoM 72 h 1:2 dilution: control > IoM ≈ EF 72 h 1:4 dilution: control > EF > IoM |
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| Cell migration (24 and 48 h) | Scratch assay | Control > EF > IoM for all concentrations and study periods | |||
| Cell differentiation | Flow cytometry– Annexin V/7-AAD staining |
IoM and EF ≈ control (the percentage of positive expression of mesenchymal markers) | |||
| Pedano MS et al. (2018) [76] | Exp-PPL (PPL) Biodentine (Bd; Septodont) Nex-Cem MTA (Nex-MTA; GC) Zinc-oxide eugenol Alganol (ZnO; Kemdent) |
Indirect contact (FRESH materials) |
Cell viability (24 h) | XTT assay | 10% eluates: Bd > PPL ≈ Nex-MTA > ZnO 25% eluates: PPL > Nex-MTA > Bd > ZnO 50% eluates: PPL ≈ Nex-MTA > Bd > ZnO 100% eluates: Nex-MTA > PPL > Bd > ZnO |
| Cell proliferation (1, 4 & 7 d) | XTT assay | 10% eluates 7d: PPL ≈ Bd ≈ control > Nex-MTA > ZnO 25% eluates 7d: control > Bd > PPL > Nex-MTA > ZnO 50% eluates 7d: control > Bd > PPL > Nex-MTA > ZnO 100% eluates 7d: control > PPL ≈ Bd ≈ Nex-MTA > ZnO |
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| Cell migration (24 h) | Scratch-wound healing assay |
10% and 25% eluates: control ≈ PPL ≈ Nex-MTA > Bd
50% eluates: control ≈Nex-MTA ≈ PPL > Bd 100% eluates: control > PPL > Nex-MTA > Bd |
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| Cell differentiation (4, 10 & 14 d) | RT-PCR (ALP, OCN, DSPP) | ALP 4 d: differentiation medium > PPL ≈ Bd ≈ Nex-MTA 10 d: differentiation medium ≈ PPL ≈ Bd ≈ Nex-MTA 14 d: differentiation medium > PPL > Bd ≈ Nex-MTA OCN 14d: PPL ≈ Bd > Nex-MTA ≈ differentiation medium DSPP 10 d: PPL ≈ Bd ≈ Nex-MTA ≈ differentiation medium 14 d: Bd > PPL > differentiation medium > Nex-MTA |
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| Tomás-Catalá CJ et al. (2018) [77] |
Biodentine (Bd; Septodont)MTA Repair HP Angelus (MTA-HP-A; Angelus) NeoMTA Plus (N-MTA-P; Avalon Biomed Inc) |
Indirect and direct contact (SET materials) (ISO 10993-5) | Cell attachment | SEM-EDX (direct contact, 72 h) | SEM showed Bd revealed more cells and with better morphology than MTA-HP-A and N-MTA-P. The EDX revealed that Bd, MTA-HP-A and N-MTA-P had similar percentages of Ca, C and O. |
| Cell viability | MTT assay (24, 48 & 72 h) | Undiluted extract: Bd > MTA-HP-A > N-MTA-P > control at 48 h and 72 h 1:2 dilution: Bd > MTA-HP-A ≈ N-MTA-P ≈ control (p < 0.01) at 48 h and 72 h 1:4 dilution: Bd > N-MTA-P ≈ control > MTA-HP-A at 72 h |
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| Cell migration | Scratch assay (at 24 & 48 h) |
24 h: Bd > MTA-HP-A ≈ N-MTA-P ≈ control (p < 0.01)
48 h: Bd > control for all dilutions; control > N-MTA-P > MTA-HP-A |
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| Lee S-M et al. (2019) [78] | Smart Dentin Replacement (SDR; Dentsply-Sirona) Venus Bulk-fill (VBF; Hereaus Kulzer) Beautifil Bulk flowable (BBF; Shofu) Filtek Z350 XT Flowable (ZFF; 3M) |
Indirect contact (Set materials) (ISO 10993-5) |
Cell viability | WST assay (24 h) Live/dead Assay (direct visualization with confocal microscopy) |
2-mm-cured composite: ≈ 100% cell-viability except for BFF (49%) 4-mm-cured composite: SDR not cytotoxic at all dilutions. VBF & BBF statistically different values (71.05% and 64.43%, respectively) of cell viability at 100% concentration compared to control (p < 0.05) but no statistically different cell viability compared to control at 25% and 12.5% concentrations, respectively (~100%, p > 0.05) 6-mm-cured composite: SDR and BBF were ~69% and ~6% at 100% concentration (p < 0.05), and these resins did not show statistically different cell viability compared to control at 25% and 12.5% (~100%, p > 0.05), respectively. In contrast, VBF and ZFF did not reach non-cytotoxic levels (~100%) even at 12.5% dilution. |
| At 100% concentrations of SDR, VBF, and ZFF, 6-mm cured composite showed 5~60% live cell numbers compared to the 2-mm cured group. Another bulk-fill resin, BBF, had 5~35% live cells with some dead cells in all groups. At 12.5%, there were full of live cells at all groups while the 4-mm cured ZFF and the 6-mm cured VBF and ZFF revealed fewer live cells (~75%) than the control. | |||||
| Cell differentiation (7 days) |
ALP staining | 6-mm-cured bulk-fill resins showed significantly lower ALP staining than the differentiation media control (p < 0.05), while all 2-mm and 4-mm cured bulk-fill resins showed similar ALP staining, except for 4-mm-cured BBF. ALP staining from the bulk-fill resins was ranked as follows: 2-mm > 4-mm > 6-mm cured. The flowable resin, ZFF, exhibited the least amount of ALP staining between the experimental groups. | |||
| López-García et al. (2019) [79] | Activa Kids (Activa; Pulpdent) GI Ionolux (Voco) Riva Light Cure (Riva; SDI) |
Indirect and direct contact (Set materials) (ISO 10993-5) |
Cell morphology (indirect contact) | Immunofluorescence | Activa > cell density and spreading than Riva > Inolux |
| Cell attachment/adhesion (direct contact) | SEM | Activa showed well-adhered fibroblastic cells with multiple cytoplasmic extensions. Riva showed less density and fewer cells than Activa. Ionolux induced drastic reduction in cell density and attachement. |
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| Cell viability | MTT assay (1, 2 & 4 days) | 24 h - Undiluted extracts: Activa ≈ control > Riva > Ionolux (p < 0.01) 24 h–1:2 dilution: Activa ≈ control ≈ Riva > Ionolux 24 h–1:4 dilution: Activa ≈ control > Riva > Ionolux 48 h-Undiluted extracts: Ionolux < Activa & Riva (p < 0.01) < control (p < 0.01) 48 h–1:2 dilution: Activa & Riva & Ionolux ≈ control 48 h–1:4 dilution: Activa & Riva & Ionolux ≈ control |
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| 72 h-Undiluted extracts: Control > Activa > Riva > Ionolux 72 h–1:2 dilution: Control > Activa > Riva > Ionolux 72 h–1:4 dilution: Activa ≈ control; Riva & Ionolux < control | |||||
| Cell migration | Wound healing assay |
Activa ≈ control at all dilutions except 1:2 at 72 h
Riva < migration than control except 1:4 dilution Ionolux < migration than control except 1:4 dilution at 24 h and 48 h |
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| Dou L et al. (2020) [80] | Dycal (Dentsply-Sirona) Pro-Root MTA (MTA; Dentsply-Sirona) iRoot BP (iRoot; Innovative Bioceramix) Platelet-rich Fibrin (PRF) Concentrated Growth Factors (CGF) |
Indirect contact (Set materials) |
Cell viability | Trypan Blue Staining (1, 3 & 7 days) Flow cytometry– Annexin V-PI (1, 3 & 7 days) Cell Cycle(1, 3 & 7 days) |
Dycal < cell viability than MTA ≈ iRoot ≈ PRF ≈ CGF ≈ control at 1, 3 & 7 days |
| Dycal > apoptotic cells than MTA ≈ iRoot ≈ CGF ≈ control at 1, 3 & 7 days Days 1 & 3: no significant differences among the groups Day 7: CGF showed less cells in G0/G1-phase compared to MTA & Dycal | |||||
| Cell proliferation | CCK-8 | Day 1: Dycal < cell proliferation than all groups; MTA ≈ iRoot ≈ PRF ≈ CGF ≈ control. Day 3: PRF & CGF > cell proliferation than control & MTA, but ≈ iRoot; Dycal < all groups Day 7: CGF > cell proliferation than iRoot & MTA, but ≈ control &PRF; Dycal < all groups |
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| Cell differentiation (1,3 & 7 days) |
ALP staining | Days 1 & 3: MTA > ALP-activity than control; Control ≈ iRoot ≈ PRF ≈ CGF ≈ Dycal Day 7: Dycal < ALP-activity than CGF; CGF ≈ control ≈ MTA ≈ iRoot ≈ PRF |
* Direct contact was considered when the cells were seeded on top of the materials. When the material was placed on a transwell insert or materials’ eluates were used, it was considered INDIRECT contact.