Vitamin D3 inhibits expression and activities of matrix metalloproteinase-2 and -9 in human uterine fibroid cells

Sunil K. Halder, Kevin G. Osteen, Ayman Al-Hendy

Research output: Contribution to journalArticle

61 Citations (Scopus)

Abstract

STUDY QUESTIONCan biologically active vitamin D3 [1,25(OH)2D3] regulate the expression and activity of matrix metalloproteinases (MMPs) in human uterine fibroid cells?SUMMARY ANSWER1,25(OH)2D3 effectively reduced the expression and activities of MMP-2 and MMP-9 in cultured human uterine fibroid cells.WHAT IS KNOWN ALREADYUterine fibroids (leiomyoma) express higher levels of MMP activity than adjacent normal myometrium, and this is associated with uterine fibroid pathogenesis. However, it is unknown whether 1,25(OH)2D3 can regulate the expression and activities of MMPs in human uterine fibroid cells.STUDY DESIGN, SIZE, DURATIONSurgically removed fresh fibroid tissue was used to generate primary uterine fibroid cells.PARTICIPANTS/MATERIALS, SETTING, METHODSAn immortalized human uterine fibroid cell line (HuLM) and/or primary human uterine fibroid cells isolated from fresh fibroid tissue were used to examine the expression of several MMPs, tissue inhibitors of metalloproteinases (TIMP) 1 and 2 and the activities of MMP-2 and MMP-9 after 1,25(OH)2D3 treatment. Real-time PCR and western blots analyses were used to measure mRNA and protein expression of MMPs, respectively. Supernatant cell culture media were analyzed for MMP-2 and MMP-9 activities using a gelatin zymography assay.MAIN RESULTS AND THE ROLE OF CHANCE1-1000 nM 1,25(OH)2D3 significantly reduced mRNA levels of MMP-2 and MMP-9 in HuLM cells in a concentration-dependent manner (P < 0.5 to P < 0.001). The mRNA levels of MMP-1, MMP-3, MMP-13 and MMP-14 in HuLM cells were also reduced by 1,25(OH)2D3. 1,25(OH)2D3 significantly reduced MMP-2 and MMP-9 protein levels in a concentration- dependent manner in both HuLM and primary uterine fibroid cells (P < 0.05 to P < 0.001). Moreover, 1,25(OH)2D3 increased the mRNA levels of vitamin D receptor (VDR) and TIMP-2 in a concentration-dependent manner in HuLM cells (P < 0.05 to P < 0.01). 1,25(OH)2D3 also significantly increased protein levels of VDR and TIMP-2 in all cell types tested (P < 0.05 to P < 0.001). Gelatin zymography revealed that pro-MMP-2, active MMP-2 and pro-MMP-9 were down-regulated by 1,25(OH)2D3 in a concentration-dependent manner; however, the active MMP-9 was undetectable.LIMITATIONS, REASONS FOR CAUTIONThis study was performed using in vitro uterine fibroid cell cultures and the results were extrapolated to in vivo situation of uterine fibroids. Moreover, in this study the interaction of vitamin D3 with other regulators such as steroid hormone receptors was not explored.WIDER IMPLICATIONS OF THE FINDINGSThis study reveals an important biological function of 1,25(OH)2D3 in the regulation of expression and activities of MMP-2 and MMP-9. Thus, 1,25(OH)2D3 might be a potential effective, safe non-surgical treatment option for human uterine fibroids.

Original languageEnglish (US)
Pages (from-to)2407-2416
Number of pages10
JournalHuman Reproduction
Volume28
Issue number9
DOIs
StatePublished - Sep 2013

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Cholecalciferol
Matrix Metalloproteinase 2
Matrix Metalloproteinase 9
Leiomyoma
Matrix Metalloproteinases
Tissue Inhibitor of Metalloproteinase-2
Messenger RNA
Calcitriol Receptors
Gelatin
Cell Culture Techniques
Matrix Metalloproteinase 13
Matrix Metalloproteinase 14
Matrix Metalloproteinase 3
Matrix Metalloproteinase 1
Proteins
Tissue Inhibitor of Metalloproteinase-1
Matrix Metalloproteinase Inhibitors
Myometrium
Steroid Receptors
Culture Media

Keywords

  • MMPs
  • TIMP-2
  • VDR
  • fibroids
  • vitamin D3

ASJC Scopus subject areas

  • Reproductive Medicine
  • Obstetrics and Gynecology

Cite this

Vitamin D3 inhibits expression and activities of matrix metalloproteinase-2 and -9 in human uterine fibroid cells. / Halder, Sunil K.; Osteen, Kevin G.; Al-Hendy, Ayman.

In: Human Reproduction, Vol. 28, No. 9, 09.2013, p. 2407-2416.

Research output: Contribution to journalArticle

Halder, Sunil K. ; Osteen, Kevin G. ; Al-Hendy, Ayman. / Vitamin D3 inhibits expression and activities of matrix metalloproteinase-2 and -9 in human uterine fibroid cells. In: Human Reproduction. 2013 ; Vol. 28, No. 9. pp. 2407-2416.
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T1 - Vitamin D3 inhibits expression and activities of matrix metalloproteinase-2 and -9 in human uterine fibroid cells

AU - Halder, Sunil K.

AU - Osteen, Kevin G.

AU - Al-Hendy, Ayman

PY - 2013/9

Y1 - 2013/9

N2 - STUDY QUESTIONCan biologically active vitamin D3 [1,25(OH)2D3] regulate the expression and activity of matrix metalloproteinases (MMPs) in human uterine fibroid cells?SUMMARY ANSWER1,25(OH)2D3 effectively reduced the expression and activities of MMP-2 and MMP-9 in cultured human uterine fibroid cells.WHAT IS KNOWN ALREADYUterine fibroids (leiomyoma) express higher levels of MMP activity than adjacent normal myometrium, and this is associated with uterine fibroid pathogenesis. However, it is unknown whether 1,25(OH)2D3 can regulate the expression and activities of MMPs in human uterine fibroid cells.STUDY DESIGN, SIZE, DURATIONSurgically removed fresh fibroid tissue was used to generate primary uterine fibroid cells.PARTICIPANTS/MATERIALS, SETTING, METHODSAn immortalized human uterine fibroid cell line (HuLM) and/or primary human uterine fibroid cells isolated from fresh fibroid tissue were used to examine the expression of several MMPs, tissue inhibitors of metalloproteinases (TIMP) 1 and 2 and the activities of MMP-2 and MMP-9 after 1,25(OH)2D3 treatment. Real-time PCR and western blots analyses were used to measure mRNA and protein expression of MMPs, respectively. Supernatant cell culture media were analyzed for MMP-2 and MMP-9 activities using a gelatin zymography assay.MAIN RESULTS AND THE ROLE OF CHANCE1-1000 nM 1,25(OH)2D3 significantly reduced mRNA levels of MMP-2 and MMP-9 in HuLM cells in a concentration-dependent manner (P < 0.5 to P < 0.001). The mRNA levels of MMP-1, MMP-3, MMP-13 and MMP-14 in HuLM cells were also reduced by 1,25(OH)2D3. 1,25(OH)2D3 significantly reduced MMP-2 and MMP-9 protein levels in a concentration- dependent manner in both HuLM and primary uterine fibroid cells (P < 0.05 to P < 0.001). Moreover, 1,25(OH)2D3 increased the mRNA levels of vitamin D receptor (VDR) and TIMP-2 in a concentration-dependent manner in HuLM cells (P < 0.05 to P < 0.01). 1,25(OH)2D3 also significantly increased protein levels of VDR and TIMP-2 in all cell types tested (P < 0.05 to P < 0.001). Gelatin zymography revealed that pro-MMP-2, active MMP-2 and pro-MMP-9 were down-regulated by 1,25(OH)2D3 in a concentration-dependent manner; however, the active MMP-9 was undetectable.LIMITATIONS, REASONS FOR CAUTIONThis study was performed using in vitro uterine fibroid cell cultures and the results were extrapolated to in vivo situation of uterine fibroids. Moreover, in this study the interaction of vitamin D3 with other regulators such as steroid hormone receptors was not explored.WIDER IMPLICATIONS OF THE FINDINGSThis study reveals an important biological function of 1,25(OH)2D3 in the regulation of expression and activities of MMP-2 and MMP-9. Thus, 1,25(OH)2D3 might be a potential effective, safe non-surgical treatment option for human uterine fibroids.

AB - STUDY QUESTIONCan biologically active vitamin D3 [1,25(OH)2D3] regulate the expression and activity of matrix metalloproteinases (MMPs) in human uterine fibroid cells?SUMMARY ANSWER1,25(OH)2D3 effectively reduced the expression and activities of MMP-2 and MMP-9 in cultured human uterine fibroid cells.WHAT IS KNOWN ALREADYUterine fibroids (leiomyoma) express higher levels of MMP activity than adjacent normal myometrium, and this is associated with uterine fibroid pathogenesis. However, it is unknown whether 1,25(OH)2D3 can regulate the expression and activities of MMPs in human uterine fibroid cells.STUDY DESIGN, SIZE, DURATIONSurgically removed fresh fibroid tissue was used to generate primary uterine fibroid cells.PARTICIPANTS/MATERIALS, SETTING, METHODSAn immortalized human uterine fibroid cell line (HuLM) and/or primary human uterine fibroid cells isolated from fresh fibroid tissue were used to examine the expression of several MMPs, tissue inhibitors of metalloproteinases (TIMP) 1 and 2 and the activities of MMP-2 and MMP-9 after 1,25(OH)2D3 treatment. Real-time PCR and western blots analyses were used to measure mRNA and protein expression of MMPs, respectively. Supernatant cell culture media were analyzed for MMP-2 and MMP-9 activities using a gelatin zymography assay.MAIN RESULTS AND THE ROLE OF CHANCE1-1000 nM 1,25(OH)2D3 significantly reduced mRNA levels of MMP-2 and MMP-9 in HuLM cells in a concentration-dependent manner (P < 0.5 to P < 0.001). The mRNA levels of MMP-1, MMP-3, MMP-13 and MMP-14 in HuLM cells were also reduced by 1,25(OH)2D3. 1,25(OH)2D3 significantly reduced MMP-2 and MMP-9 protein levels in a concentration- dependent manner in both HuLM and primary uterine fibroid cells (P < 0.05 to P < 0.001). Moreover, 1,25(OH)2D3 increased the mRNA levels of vitamin D receptor (VDR) and TIMP-2 in a concentration-dependent manner in HuLM cells (P < 0.05 to P < 0.01). 1,25(OH)2D3 also significantly increased protein levels of VDR and TIMP-2 in all cell types tested (P < 0.05 to P < 0.001). Gelatin zymography revealed that pro-MMP-2, active MMP-2 and pro-MMP-9 were down-regulated by 1,25(OH)2D3 in a concentration-dependent manner; however, the active MMP-9 was undetectable.LIMITATIONS, REASONS FOR CAUTIONThis study was performed using in vitro uterine fibroid cell cultures and the results were extrapolated to in vivo situation of uterine fibroids. Moreover, in this study the interaction of vitamin D3 with other regulators such as steroid hormone receptors was not explored.WIDER IMPLICATIONS OF THE FINDINGSThis study reveals an important biological function of 1,25(OH)2D3 in the regulation of expression and activities of MMP-2 and MMP-9. Thus, 1,25(OH)2D3 might be a potential effective, safe non-surgical treatment option for human uterine fibroids.

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KW - TIMP-2

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KW - fibroids

KW - vitamin D3

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