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HOME > Restor Dent Endod > Volume 36(5); 2011 > Article
Basic Research Comparison of gene expression profiles of human dental pulp cells treated with mineral trioxide aggregate and calcium hydroxide
Yong-Beom Kim, DDS, MSD, PhD1, Won-Jun Shon, DDS, MSD, PhD2, Woocheol Lee, DDS, MSD, PhD2, Kee-Yeon Kum, DDS, MSD, PhD2, Seung-Ho Baek, DDS, MSD, PhD2, Kwang-Shik Bae, DDS, MSD, PhD2,*
J Korean Acad Conserv Dent 2011;36(5):-408.
DOI: https://doi.org/10.5395/JKACD.2011.36.5.397
Published online: September 14, 2011

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*Correspondence to: Kwang-Shik Bae, DDS, MSD, PhD., Professor, Department of Conservative Dentistry, Seoul National University School of Dentistry, Dental Research Institute, 28-2 Yongon-dong, Jongno-gu, Seoul, Korea 110-749, Tel, +82-2-2072-2651; Fax, +82-2-2072-3859; E-mail, baeks@snu.ac.kr
• Received: July 20, 2011   • Revised: August 17, 2011   • Accepted: August 21, 2011

Copyright © 2011 The Korean Academy of Conservative Dentistry

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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  • Objectives:
    This study investigated changes in gene expressions concerning of differentiation, proliferation, mineralization and inflammation using Human-8 expression bead arrays when white Mineral Trioxide Aggregate and calcium hydroxide-containing cement were applied in vitro to human dental pulp cells (HDPCs).
  • Materials and Methods:
    wMTA (white ProRoot MTA, Dentsply) and Dycal (Dentsply Caulk) in a Teflon tube (inner diameter 10 mm, height 1 mm) were applied to HDPCs. Empty tube-applied HDPCs were used as negative control. Total RNA was extracted at 3, 6, 9 and 24 hr after wMTA and Dycal application. The results of microarray were confirmed by reverse transcriptase polymerase chain reaction.
  • Results:
    Out of the 24,546 genes, 43 genes (e.g., BMP2, FOSB, THBS1, EDN1, IL11, COL10A1, TUFT1, HMOX1) were up-regulated greater than two-fold and 25 genes (e.g., SMAD6, TIMP2, DCN, SOCS2, CEBPD, KIAA1199) were down-regulated below 50% by wMTA. Two hundred thirty nine genes (e.g., BMP2, BMP6, SMAD6, IL11, FOS, VEGFA, PlGF, HMOX1, SOCS2, CEBPD, KIAA1199) were up-regulated greater than two-fold and 358 genes (e.g., EDN1, FGF) were down-regulated below 50% by Dycal.
  • Conclusions:
    Both wMTA and Dycal induced changes in gene expressions related with differentiation and proliferation of pulp cells. wMTA induced changes in gene expressions related with mineralization, and Dycal induced those related with angiogenesis. The genes related with inflammation were more expressed by Dycal than by wMTA. It was confirmed that both wMTA and Dycal were able to induce gene expression changes concerned with the pulp repair in different ways.
  • Keywords: Calcium hydroxide; Microarray; Mineral trioxide aggregate; Pulp capping; Pulp cell; Reverse transcriptase polymerase chain reaction
Figure 1.
Classification of expressed genes by wMTA treatment. The genes which were up-or down-regulated by at least twofold following a cellular treatment with wMTA at any time point were classified using the panther classification system (http://www.pantherdb.org).
jkacd-36-397f1.jpg
Figure 2.
Classification of expressed genes by Dycal treatment. The genes which were up-or down-regulated by at least twofold following a cellular treatment with Dycal at any time point were classified using the panther classification system.
jkacd-36-397f2.jpg
Table 1.
Primer sequence list used in RT-PCR
Target gene Forward Primer Reverse Product Size (bp)
GAPDH 5′-GAA GTG GAA GGT CGG AGT C-3′   5′-GAA GAT GGT GAT GGG ATT TC-3′ 313
BMP2 5′-ACT TTT GGA CAC CAG GTT GG-3′   5′-GCT GTT TGT GTT TGG CTT GA-3′ 296
IL11 5′-GCA ATC TGA GGT CAC TGC AA-3′   5′-TCA CAG GGT GAC TTG TGG AA-3′ 300
EDN1 5′-TTC CGT ATG GAC TTG GAA GC-3′   5′-TCT CCG ACC TGG TTT GTC TT-3′ 300
CEBPD 5′-AGA AGT TGG TGG AGC TGT CG-3′   5′-CCA GGC TGT AGC TTC TTT GC-3′ 300
FOSB 5′-GGC TCA ACA AGG AAA AAT GC-3′   5′-GAG GCC AGA AAT TCC AAT CA-3′ 297
TUFT1 5′-GCA GAA AGA GGC AGA AGT CG-3′   5′-CCA AGC AGA ACG AGA AAA GG-3′ 152
FOS 5′-CTT CCT GTT CCC AGC ATC AT-3′   5′-GTC AGA GGA AGG CTC ATT GC-3′ 298
THBS1 5′-CCT GTG CTG CAG ACA TTG AT-3′   5′-GAA CAC AAT CCG GAT CAG CT-3′ 148
DCN 5′-GGA CCG TTT CAA CAG AGA GG-3′   5′-AAT GCC ATC TTC GAG TGG TC-3′ 147
BMP6 5′-TTC GTG CTG GAG TTT TGT TG-3′   5′-CTA ACC CTG TGG CGT GGT AT-3′ 298
FGF5 5′-GCA AGT GCC AAG TTC ACA GA-3′   5′-AGG TGC AGA AAG GGG AAT CT-3′ 303
Table 2.
Summary of expression profiles of genes differentially regulated by wMTA and Dycal
Fold change
 Number Gene symbol
wMTA Dycal
fold > 2 fold > 2 17 NUAK1, TRIB1, SGK, FZD8, NOX4, ABCA1, SLC35F2, BPGM, IL11,
      PMEPA1, ASNS, RCAN1, BMP2, PTGS2, PTHLH, STC1, HMOX1
  fold < -2 1 EDN1
 
 2 > fold > -2
 25
 LOH3CR2A, ANGPTL4, SH3BP5L, MMP10, DENR, DACT1, SCD, SLC46A3, UAP1L1, PDPN, FOSB, TUFT1, TNS1, ZNF365, ESM1, HBEGF, NKX3-1, MAZ, AK3L1, ETV4, MT1X, FOLR3, LRRC17, THBS1, COL10A1
 
 Subtotal
 43
  
fold < -2 fold > 2 7 KIAA1199, DHRS3, CEBPD, KYNU, SMAD6, ATOH8, SOCS2
fold < -2 5 PDE5A, HERC4, DDIT4L, VCAM1, SLC25A22
 
 2 > fold > -2
 13
 TIMP2, FBN2, COL3A1, MFGE8, CNTNAP1, MFAD4, UBA7, PRRX2, VPS24, SIPA1, ASAM, PHF15, DCN
 
 Subtotal
 25
  
Table 3.
The relative mRNA expression levels of genes analyzed by RT-PCR
Symbol 6hr
 9hr
 24hr
wMTA Dycal wMTA Dycal wMTA Dycal
BMP2 1.88 2.31        
IL11     1.99 2.38    
EDN1         1.41 0.26
CEBPD         1.16 2.07
FOSB     1.28      
TUFT1     1.57      
FOS       1.77    
THBS1         1.89  
DCN         0.58  
BMP6           11.93
FGF5           0.25

The relative level of gene expression was normalized against GAPDH messenger RNA, and the control was set as 1.0. Optical density values represent the mean (n=3).

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    • Analysis of gene expression during odontogenic differentiation of cultured human dental pulp cells
      Min-Seock Seo, Kyung-Gyun Hwang, Hyongbum Kim, Seung-Ho Baek
      Restorative Dentistry & Endodontics.2012; 37(3): 142.     CrossRef

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      Comparison of gene expression profiles of human dental pulp cells treated with mineral trioxide aggregate and calcium hydroxide
      J Korean Acad Conserv Dent. 2011;36(5):397-408.   Published online September 14, 2011
      DOI: https://doi.org/10.5395/JKACD.2011.36.5.397
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    Comparison of gene expression profiles of human dental pulp cells treated with mineral trioxide aggregate and calcium hydroxide
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    Figure 1. Classification of expressed genes by wMTA treatment. The genes which were up-or down-regulated by at least twofold following a cellular treatment with wMTA at any time point were classified using the panther classification system (http://www.pantherdb.org).
    Figure 2. Classification of expressed genes by Dycal treatment. The genes which were up-or down-regulated by at least twofold following a cellular treatment with Dycal at any time point were classified using the panther classification system.

    Figure 1.

    Figure 2.

    Comparison of gene expression profiles of human dental pulp cells treated with mineral trioxide aggregate and calcium hydroxide
    Target gene Forward Primer Reverse Product Size (bp)
    GAPDH 5′-GAA GTG GAA GGT CGG AGT C-3′   5′-GAA GAT GGT GAT GGG ATT TC-3′ 313
    BMP2 5′-ACT TTT GGA CAC CAG GTT GG-3′   5′-GCT GTT TGT GTT TGG CTT GA-3′ 296
    IL11 5′-GCA ATC TGA GGT CAC TGC AA-3′   5′-TCA CAG GGT GAC TTG TGG AA-3′ 300
    EDN1 5′-TTC CGT ATG GAC TTG GAA GC-3′   5′-TCT CCG ACC TGG TTT GTC TT-3′ 300
    CEBPD 5′-AGA AGT TGG TGG AGC TGT CG-3′   5′-CCA GGC TGT AGC TTC TTT GC-3′ 300
    FOSB 5′-GGC TCA ACA AGG AAA AAT GC-3′   5′-GAG GCC AGA AAT TCC AAT CA-3′ 297
    TUFT1 5′-GCA GAA AGA GGC AGA AGT CG-3′   5′-CCA AGC AGA ACG AGA AAA GG-3′ 152
    FOS 5′-CTT CCT GTT CCC AGC ATC AT-3′   5′-GTC AGA GGA AGG CTC ATT GC-3′ 298
    THBS1 5′-CCT GTG CTG CAG ACA TTG AT-3′   5′-GAA CAC AAT CCG GAT CAG CT-3′ 148
    DCN 5′-GGA CCG TTT CAA CAG AGA GG-3′   5′-AAT GCC ATC TTC GAG TGG TC-3′ 147
    BMP6 5′-TTC GTG CTG GAG TTT TGT TG-3′   5′-CTA ACC CTG TGG CGT GGT AT-3′ 298
    FGF5 5′-GCA AGT GCC AAG TTC ACA GA-3′   5′-AGG TGC AGA AAG GGG AAT CT-3′ 303
    Fold change
    		 Number Gene symbol
    wMTA Dycal
    fold > 2 fold > 2 17 NUAK1, TRIB1, SGK, FZD8, NOX4, ABCA1, SLC35F2, BPGM, IL11,
          PMEPA1, ASNS, RCAN1, BMP2, PTGS2, PTHLH, STC1, HMOX1
      fold < -2 1 EDN1
     
    		 2 > fold > -2
    		 25
    		 LOH3CR2A, ANGPTL4, SH3BP5L, MMP10, DENR, DACT1, SCD, SLC46A3, UAP1L1, PDPN, FOSB, TUFT1, TNS1, ZNF365, ESM1, HBEGF, NKX3-1, MAZ, AK3L1, ETV4, MT1X, FOLR3, LRRC17, THBS1, COL10A1
     
    		 Subtotal
    		 43
    		  
    fold < -2 fold > 2 7 KIAA1199, DHRS3, CEBPD, KYNU, SMAD6, ATOH8, SOCS2
    fold < -2 5 PDE5A, HERC4, DDIT4L, VCAM1, SLC25A22
     
    		 2 > fold > -2
    		 13
    		 TIMP2, FBN2, COL3A1, MFGE8, CNTNAP1, MFAD4, UBA7, PRRX2, VPS24, SIPA1, ASAM, PHF15, DCN
     
    		 Subtotal
    		 25
    		  
    Symbol 6hr
    		 9hr
    		 24hr
    wMTA Dycal wMTA Dycal wMTA Dycal
    BMP2 1.88 2.31        
    IL11     1.99 2.38    
    EDN1         1.41 0.26
    CEBPD         1.16 2.07
    FOSB     1.28      
    TUFT1     1.57      
    FOS       1.77    
    THBS1         1.89  
    DCN         0.58  
    BMP6           11.93
    FGF5           0.25
    Table 1. Primer sequence list used in RT-PCR

    Table 2. Summary of expression profiles of genes differentially regulated by wMTA and Dycal

    Table 3. The relative mRNA expression levels of genes analyzed by RT-PCR

    The relative level of gene expression was normalized against GAPDH messenger RNA, and the control was set as 1.0. Optical density values represent the mean (n=3).

    Table 1.

    Table 2.

    Table 3.

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