Differentiation- and maturation-dependent content, localization, and secretion of cystatin C in human dendritic cells

Abstract

Antigen-presenting cells (APC) play a pivotal role in the initiation of the T cell-mediated and antigen-specific immune response. The suggested role of endogenous inhibitor cystatin C (CyC) is to modulate cysteine proteases (cathepsins) present in human APC. To test this hypothesis, dendritic cells (DC) were generated in vitro from isolated monocytes, and changes in content, localization, and secretion of CyC and cathepsins S, L, and H (CatS, -L, and -H, repsectively) were followed in response to interleukin-4, enabling monocyte differentiation, and to tumor necrosis factor alpha (TNF-alpha), enabling DC maturation. A large increase in intracellular CyC accompanied the differentiation of monocytes to immature DC, also shown by strong immunolabeling of Golgi in immature DC. On DC maturation, intracellular CyC levels decreased, and CyC was mostly absent from the Golgi. On prolonged incubation of mature DC with TNF-alpha, CyC was found located in the proximity of the plasma membrane, indicating that the transport of CyC from Golgi was not blocked as the result of the arrested exocytosis in mature DC. The secretion of CyC ceased, consistent with the peak of the surface expression of phenotypic markers (CD40, CD54, CD80, CD83, CD86, and major histocompatibility complex class II), characteristic for the mature DC stage, whereas the secretion of cathepsins did not correlate with the maturation stage. The difference in localization of CyC and of CatS, -L, and -H in immature and mature DC shows that the regulatory potential of CyC toward CatS, -L, and -H inside DC is limited. However, these interactions may occur extracellularly in lymph, as suggested by the large excess of CyC over secreted CatS, -L, and -H, and they may facilitate DC migration to lymph nodes.

IL-6-STAT3 Controls Intracellular MHC Class II αβ Dimer Level through Cathepsin S Activity in Dendritic Cells

“…It should be pointed out that there must be a certain amount of cystatin C molecules still, even after treatment of IL-6 in DCs. Cystation C is reported to be secreted from DCs (Zavasnik-Bergant et al, 2005). Therefore, it is reasonable that the preexisting cystatin C molecules are secreted outside of DCs gradually or digested by intracellular proteases including cathepsins.…”

Dendritic Cell Maturation Results in Pronounced Changes in Glycan Expression Affecting Recognition by Siglecs and Galectins

“…Interestingly, the increase in the binding of CD22, or the ␣2,6-specific plant lectin SNA, did not correlate with the expression of the enzyme involved in the synthesis of its glycan, ST6Gal-1, which decreased dramatically. This could be explained by the maturation-triggered mobilization of molecules stored in granules, a phenomenon that has been previously described on DC (56,57), and needs to be further investigated. The interaction between DC and B lymphocytes has been shown to result in multiple effects, such as the inhibition of B cell apoptosis (58), blockage of the production of IgE (59), or Ag presentation (60).…”

Salivary Tick Cystatin OmC2 Targets Lysosomal Cathepsins S and C in Human Dendritic Cells

“…Selected bands were excised and analyzed using mass spectrometry or the proteins were transferred to PVDF membranes. The proteins from the SDS-PAGE or IEF gels were electrotransferred to PVDF membranes and immunolabeled with rabbit anti-cathepsin S pAb (Zavašnik-Bergant et al, 2005), goat anti-cathepsin S pAb (AF1183, R&D Systems), mouse anti-cathepsin C mAb (sc-74590, Santa Cruz Biotechnology), or mouse anti-β-actin antibody (A1978, Sigma-Aldrich). ECL Western blotting detection reagents (Amersham, GE Healthcare) were applied, and the imaging films (BIOMAX Light Film, Carestream) were developed (Konica Minolta SRX-101A).…”