Vitamin D3 metabolites regulate LTBP1 and latent TGF-?1 expression and latent TGF-?1 incorporation in the extracellular matrix of chondrocytes

Abstract

Growth plate chondrocytes make TGF-beta1 in latent form (LTGF-beta1) and store it in the extracellular matrix via LTGF-beta1 binding protein (LTBP1). 1,25-(OH)2D3 (1,25) regulates matrix protein production in growth zone (GC) chondrocyte cultures, whereas 24,25-(OH)2D3 (24,25) does so in resting zone (RC) cell cultures. The aim of this study was to determine if 24,25 and 1,25 regulate LTBP1 expression as well as the LTBP1 -mediated storage of TGF-beta1 in the extracellular matrix of RC and GC cells. Expression of LTBP1 and TGF-beta1 in the growth plate and in cultured RC and GC cells was determined by in situ hybridization using sense and antisense oligonucleotide probes based on the published rat LTBP1 and TGF-beta1 cDNA sequences. Fourth passage male rat costochondral RC and GC chondrocytes were treated for 24 h with 10(-7)-10(-9) M 24,25 and 10(-8)-10(-10) M 1,25, respectively. LTBP1 and TGF-beta1 mRNA levels were measured by in situ hybridization; production of LTGF-beta1, LTGF-beta2, and LTBP1 protein in the conditioned media was verified by immunoassays of FPLC-purified fractions. In addition, ELISA assays were used to measure the effect of 1,25 and 24,25 on the level of TGF-beta1 in the media and matrix of the cultures. Matrix-bound LTGF-beta1 was released by digesting isolated matrices with 1 U/ml plasmin for 3 h at 37 degrees C. LTBP1 and TGF-beta1 mRNAs are co-expressed throughout the growth plate, except in the lower hypertrophic area. Cultured GC cells express more LTBP1 and TGF-beta1 mRNAs than RC cells. FPLC purification of the conditioned media confirmed that RC cells produce LTGF-beta1, LTGF-beta2, and LTBP1. GC cells also produce LTGF-beta2, but at lower concentrations. 1,25 dose-dependently increased the number of GC cells with high LTBP1 expression, as seen by in situ hybridization. 24,25 had a similar, but less pronounced, effect on RC cells. 1,25 also caused a dose-dependent increase in the amount of TGF-beta1 protein found in the matrix, significant at 10(-8) and 10(-9) M, and a corresponding decrease in TGF-beta1 in the media. 24,25 had no effect on the level of TGF-beta1 in the matrix or media produced by RC cells. This indicates that 1,25 induces the production of LTBP1 by GC cells and suggests that the TGF-beta1 content of the media is reduced through the formation of latent TGF-beta1 -LTBP1 complexes which mediates storage in the matrix. Although 24,25 induced the expression of LTBP1 by RCs, TGF-beta1 incorporation into the matrix is not regulated by this vitamin D3 metabolite. Thus, vitamin D3 metabolites may play a role in regulating the availability of TGF-beta1 by modulating LTBP1 production.

Molecular and functional aspects of latent transforming growth factor-β binding protein: just a masking protein?

“…In 1995 it was found that vitamin D 3 enhances the expression of TGFβ1 and its latent binding protein in cultured breast carcinoma cells (Koli and Keski-Oja 1995). Recently, the observation that vitamin D 3 metabolites are involved in the regulation of LTBP and latent TGFβ1 expression has been confirmed in chondrocytes (Pedrozo et al 1999). Interestingly, TGFβ1 itself has been demonstrated to increase the transcription of LTBP-1 and LTBP-2 in osteoblastlike cell lines and human fetal lung fibroblasts, respectively (Dallas et al 1994;Ahmed et al 1998).…”

A review of 1α,25(OH)2D3 dependent Pdia3 receptor complex components in Wnt5a non-canonical pathway signaling

“…Growth factors and hormones regulate the postnatal growth of the long bones via a network of interacting signaling pathways [1][2][3][4], including the actions of 1a,25-dihydroxyvitamin D3 [1a,25 (OH) 2 D 3 ] and Wnt5a. 1a,25(OH) 2 D 3 stimulates differentiation of chondrocytes in the growth zone of the costochondral growth plate cartilage (GC cells) by increasing alkaline phosphatase specific activity [5].…”

1,25(OH)2-Vitamin D3 Inhibits Proliferation and Decreases Production of Monocyte Chemoattractant Protein-1, Thrombopoietin, VEGF, and Angiogenin by Human Annulus Cells In Vitro

“…4 The vitamin D metabolites also regulate latent TGF-␤ incorporation into the ECM. 8 It is also now recognized that 1,25(OH) 2 D 3 has functions in tissues, which are not viewed as traditional targets of vitamin D action. 9 Examples include the parathyroid gland, 10,11 skin, 12 thymus, 13 and ovarian cells.…”