Sodium-coupled vitamin C transporter (SVCT2): Expression, function, and regulation in intervertebral disc cells

Paresh P. Chothe, Norman Chutkan, Rajnikumar Sangani, Karl H. Wenger, Puttur D. Prasad, Muthusamy Thangaraju, Mark W. Hamrick, Carlos M. Isales, Vadivel Ganapathy, Sadanand Fulzele

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Background context: Vitamin C (ascorbic acid [AA]) is essential for the synthesis of collagen and also acts as an antioxidant in the intervertebral disc (IVD). However, there is very little information currently available on the identity of the transporter that facilitates AA entry into IVD cells and the factors that mediate the transport process. Purpose: To investigate the expression of the two known isoforms of Na+-coupled vitamin C transporter, SVCT1 and SVCT2, in IVD cells and its regulation by insulin-like growth factor 1 (IGF-1) and the steroid hormone dexamethasone. Study design: To identify the expression and functional activity of the sodium-dependent vitamin C transporter (SVCT) in the IVD. Methods: Uptake studies were carried out with rabbit annulus fibrosis and nucleus pulposus cells in 24-well plates using [14C]-AA. To characterize SVCT transporter, uptake was done in the presence and absence of Na+ in the uptake buffer. Time dependency, Na+ activation kinetics, saturation kinetics, and substrate selectivity studies were performed. Regulatory studies were performed in the presence of IGF-1 and the steroid hormone dexamethasone. Gene expression was analyzed by quantitative polymerase chain reaction. Results: Our real-time polymerase chain reaction results showed the presence of SVCT2 but not SVCT1 in IVD cells. Uptake of vitamin C in IVD cells is Na+ dependent and saturable. The Michaelis constant for the process is 96±11 μM. The activation of vitamin C uptake by Na+ exhibits a sigmoidal relationship, indicating involvement of more than one Na+ in the activation process. The uptake system does not recognize any other water-soluble vitamin as a substrate. Immunocytochemical analysis shows robust expression of SVCT2 protein in IVD cells. The growth factors IGF-1 and the steroid hormone dexamethasone upregulate the expression of SVCT2 in IVD cells. Conclusions: Our studies demonstrate that the active SVCT2 is expressed in IVD cells and that the expression of this transporter is regulated by growth factors IGF-1 and dexamethasone.

Original languageEnglish (US)
Pages (from-to)549-557
Number of pages9
JournalSpine Journal
Volume13
Issue number5
DOIs
StatePublished - May 1 2013

Fingerprint

Sodium-Coupled Vitamin C Transporters
Intervertebral Disc
Ascorbic Acid
Somatomedins
Dexamethasone
Steroids
Hormones
Intercellular Signaling Peptides and Proteins
Vitamins
Real-Time Polymerase Chain Reaction
Buffers
Protein Isoforms

Keywords

  • Annulus fibrosus cells
  • Intervertebral disc
  • Nucleus pulposus
  • SVCT2
  • Vitamin C transport

ASJC Scopus subject areas

  • Surgery
  • Orthopedics and Sports Medicine
  • Clinical Neurology

Cite this

Sodium-coupled vitamin C transporter (SVCT2) : Expression, function, and regulation in intervertebral disc cells. / Chothe, Paresh P.; Chutkan, Norman; Sangani, Rajnikumar; Wenger, Karl H.; Prasad, Puttur D.; Thangaraju, Muthusamy; Hamrick, Mark W.; Isales, Carlos M.; Ganapathy, Vadivel; Fulzele, Sadanand.

In: Spine Journal, Vol. 13, No. 5, 01.05.2013, p. 549-557.

Research output: Contribution to journalArticle

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abstract = "Background context: Vitamin C (ascorbic acid [AA]) is essential for the synthesis of collagen and also acts as an antioxidant in the intervertebral disc (IVD). However, there is very little information currently available on the identity of the transporter that facilitates AA entry into IVD cells and the factors that mediate the transport process. Purpose: To investigate the expression of the two known isoforms of Na+-coupled vitamin C transporter, SVCT1 and SVCT2, in IVD cells and its regulation by insulin-like growth factor 1 (IGF-1) and the steroid hormone dexamethasone. Study design: To identify the expression and functional activity of the sodium-dependent vitamin C transporter (SVCT) in the IVD. Methods: Uptake studies were carried out with rabbit annulus fibrosis and nucleus pulposus cells in 24-well plates using [14C]-AA. To characterize SVCT transporter, uptake was done in the presence and absence of Na+ in the uptake buffer. Time dependency, Na+ activation kinetics, saturation kinetics, and substrate selectivity studies were performed. Regulatory studies were performed in the presence of IGF-1 and the steroid hormone dexamethasone. Gene expression was analyzed by quantitative polymerase chain reaction. Results: Our real-time polymerase chain reaction results showed the presence of SVCT2 but not SVCT1 in IVD cells. Uptake of vitamin C in IVD cells is Na+ dependent and saturable. The Michaelis constant for the process is 96±11 μM. The activation of vitamin C uptake by Na+ exhibits a sigmoidal relationship, indicating involvement of more than one Na+ in the activation process. The uptake system does not recognize any other water-soluble vitamin as a substrate. Immunocytochemical analysis shows robust expression of SVCT2 protein in IVD cells. The growth factors IGF-1 and the steroid hormone dexamethasone upregulate the expression of SVCT2 in IVD cells. Conclusions: Our studies demonstrate that the active SVCT2 is expressed in IVD cells and that the expression of this transporter is regulated by growth factors IGF-1 and dexamethasone.",
keywords = "Annulus fibrosus cells, Intervertebral disc, Nucleus pulposus, SVCT2, Vitamin C transport",
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AU - Chothe, Paresh P.

AU - Chutkan, Norman

AU - Sangani, Rajnikumar

AU - Wenger, Karl H.

AU - Prasad, Puttur D.

AU - Thangaraju, Muthusamy

AU - Hamrick, Mark W.

AU - Isales, Carlos M.

AU - Ganapathy, Vadivel

AU - Fulzele, Sadanand

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N2 - Background context: Vitamin C (ascorbic acid [AA]) is essential for the synthesis of collagen and also acts as an antioxidant in the intervertebral disc (IVD). However, there is very little information currently available on the identity of the transporter that facilitates AA entry into IVD cells and the factors that mediate the transport process. Purpose: To investigate the expression of the two known isoforms of Na+-coupled vitamin C transporter, SVCT1 and SVCT2, in IVD cells and its regulation by insulin-like growth factor 1 (IGF-1) and the steroid hormone dexamethasone. Study design: To identify the expression and functional activity of the sodium-dependent vitamin C transporter (SVCT) in the IVD. Methods: Uptake studies were carried out with rabbit annulus fibrosis and nucleus pulposus cells in 24-well plates using [14C]-AA. To characterize SVCT transporter, uptake was done in the presence and absence of Na+ in the uptake buffer. Time dependency, Na+ activation kinetics, saturation kinetics, and substrate selectivity studies were performed. Regulatory studies were performed in the presence of IGF-1 and the steroid hormone dexamethasone. Gene expression was analyzed by quantitative polymerase chain reaction. Results: Our real-time polymerase chain reaction results showed the presence of SVCT2 but not SVCT1 in IVD cells. Uptake of vitamin C in IVD cells is Na+ dependent and saturable. The Michaelis constant for the process is 96±11 μM. The activation of vitamin C uptake by Na+ exhibits a sigmoidal relationship, indicating involvement of more than one Na+ in the activation process. The uptake system does not recognize any other water-soluble vitamin as a substrate. Immunocytochemical analysis shows robust expression of SVCT2 protein in IVD cells. The growth factors IGF-1 and the steroid hormone dexamethasone upregulate the expression of SVCT2 in IVD cells. Conclusions: Our studies demonstrate that the active SVCT2 is expressed in IVD cells and that the expression of this transporter is regulated by growth factors IGF-1 and dexamethasone.

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KW - Annulus fibrosus cells

KW - Intervertebral disc

KW - Nucleus pulposus

KW - SVCT2

KW - Vitamin C transport

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