Death receptor‐3 mediates apoptosis in human osteoblasts under narrowly regulated conditions

Abstract

We previously reported that a soluble form of the TNF-family receptor death receptor-3 (DR3) is expressed in osteoblasts. DR3 regulates death or differentiation in other tissues, and DR3 ligands occur in bone, but the function of DR3 in the osteoblast was unknown. We studied the expression of DR3 and the effects crosslinking antibodies to DR3 or of natural DR3 ligands in human osteoblasts. Western analysis showed that nontransformed osteoblasts and the MG63 osteosarcoma cell line produce both soluble decoy receptor and transmembrane isoforms of DR3. Cell surface labeling showed that low and high DR3-expressing osteoblast populations occur. Verification of by cloning showed a point mutation in DR3 from MG63 cells. Activation of DR3 by antibody crosslinking or with DR3 ligands caused apoptosis in osteoblasts and in MG63 cells, but only in low-density cell cultures. In dense cultures apoptosis did not occur, but nuclear factor-kappaB nuclear translocation was observed under some conditions. Crosslinking of DR3 in high-density MG63 cultures blocked expression of bone matrix elements. DR3 activation in high-density nontransformed osteoblasts had only minor effects on cell maturation. We conclude that DR3 activation can mediate apoptosis in osteoblasts. Its activity is, however, highly restricted by its soluble ligand-binding isoform and possibly also by alternate survival signals. In the presence of survival signals, DR3 may affect cell maturation although effects on differentiation were clearly seen only in the MG63 transformed cell line.

Modulation of osteoblast gap junction connectivity by serum, TNFα, and TRAIL

“…Activation of transmembrane receptors for TNFs should affect cell survival or differentiation. In MG63 cells, Fas crosslinking caused apoptosis and crosslinking of DR3 affects the balance of fibroblast-like growth with bone matrix production, and causes apoptosis under some conditions [13]. However, in several studies, including 12 hour treatment of serum starved cells with 100 ng/ml of TRAIL or TNFα, apoptosis was at background despite strong apoptosis in Fas-crosslinked controls ( Figure 1A).…”

“…The cysteine proteinase inhibitor leupeptin and the protein kinase C activator phorbol myristate acetate (PMA) were from Sigma (St Louis, MO). The cAMP-dependent protein kinase Inhibitor Peptide PKI (6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22), TYADFIASGRTGRRNAI was from Upstate (Lake Placid, NY). Protein was measured by alkaline reduction of copper with bicinchoninic acid relative to albumin standards measured as absorbance at 662 nm (BCA, Pierce, Rockford, IL).…”

“…Osteoblasts express the TNF-receptor family proteins TNFR-1 (p55), VEGI/TL1a (Apo3, DR3), TRAIL-R1 (DR4) and FAS (Apo1) [9,13]. Activation of transmembrane receptors for TNFs should affect cell survival or differentiation.…”

In-Silico Screening of Flavonoids targeted for Death Receptors in Cancer by Using Hex Molecular Docking

“…Among the death receptors DR3 doesn't have enough detail mechanism in activation of apoptosis, 4 also evidences that DR3 shows activation of apoptosis in low, dense population of osteoblasts, while in high density population it doesn't show apoptosis. 5 The receptor structures are obtained from Protein Data Bank (PDB) and the receptors are selected based on the previous studies. The Death Receptor 1 (1F3V), 6 Death Receptor 2 (3MX7), 7 Death Receptor 4 (1DG6), 8 Death Receptor 5 (1D4V).…”

DETERMINATION OF LEVEL EXPRESSION OF mRNA SPLICING VARIANTS FOR DR3 IN BLOOD CELLS IN INFECTIOUS MONONUCLEOSIS

“…Из них 4 мРНК кодируют мембранные формы рецептора, 8 мРНК соответствуют растворимым аналогам DR3, а 2 мРНК являются некодирующими [1,11]. Функции мембранных и растворимых вари-антов DR3 различны: мембранные формы ре- [4]. Такой функциональ-ный антагонизм различных сплайсированных вариантов DR3 играет важную роль в созревании и дальнейшей работе иммунокомпетентных кле-ток.…”