Expression cloning of the human C3a anaphylatoxin receptor (C3aR) from differentiated U‐937 cells

164

136

Mesenchymal stem cells (MSCs) have a great potential for tissue repair, especially if they can be delivered efficiently to sites of tissue injury. Since complement activation occurs whenever there is tissue damage, the effects of the complement activation products C3a and C5a on MSCs were examined. Both C3a and C5a were chemoattractants for human bone marrow-derived MSCs, which expressed both the C3a receptor (C3aR) and the C5a receptor (C5aR; CD88) on the cell surface. Specific C3aR and C5aR inhibitors blocked the chemotactic response, as did pertussis toxin, indicating that the response was mediated by the known anaphylatoxin receptors in a Gi activation-dependent fashion. While C5a causes strong and prolonged activation of various signaling pathways in many different cell types, the response observed with C3a is generally transient and weak. However, we show herein that in MSCs both C3a and C5a caused prolonged and robust ERK1/2 and Akt phosphorylation. Phospho-ERK1/2 was translocated to the nucleus in both C3a and C5a-stimulated MSCs, which was associated with subsequent phosphorylation of the transcription factor Elk, which could not be detected in other cell types stimulated with C3a. More surprisingly, the C3aR itself was translocated to the nucleus in C3a-stimulated MSCs, especially at low cell densities. Since nuclear activation/translocation of G protein-coupled receptors has been shown to induce long-term effects, this novel observation implies that C3a exerts far-reaching consequences on MSC biology. These results suggest that the anaphylatoxins C3a and C5a present in injured tissues contribute to the recruitment of MSCs and regulation of their behavior.

Section: C3a and C5a Are Chemotactic Factors For Human Mesenchymal Stmentioning

confidence: 99%

C3a and C5a Are Chemotactic Factors for Human Mesenchymal Stem Cells, Which Cause Prolonged ERK1/2 Phosphorylation

Schraufstätter

DiScipio

Zhao

et al. 2009

164

136

“…Mouse C3aR riboprobes were transcribed from a 327-bp DNA fragment corresponding to nucleotides 553-879 of mouse C3aR cDNA (46), and human C3aR riboprobes were transcribed from a 333-bp DNA fragment corresponding to nucleotides 553-885 of human C3aR cDNA (30). Mouse C5aR riboprobes were transcribed from a 214-bp DNA fragment corresponding to nucleotides 793-1014 of mouse C5aR cDNA (61), and human C5aR riboprobes were transcribed from a 205-bp DNA fragment corresponding to nucleotides 20 -225 of human C5aR cDNA (28).…”

Section: Riboprobesmentioning

confidence: 99%

Expression of the Complement Anaphylatoxin C3a and C5a Receptors on Bronchial Epithelial and Smooth Muscle Cells in Models of Sepsis and Asthma

Drouin¹,

Kildsgaard²,

Haviland³

et al. 2001

193

149

The presence of the complement-derived anaphylatoxin peptides, C3a and C5a, in the lung can induce respiratory distress characterized by contraction of the smooth muscle walls in bronchioles and pulmonary arteries and aggregation of platelets and leukocytes in pulmonary vessels. C3a and C5a mediate these effects by binding to their specific receptors, C3aR and C5aR, respectively. The cells that express these receptors in the lung have not been thoroughly investigated, nor has their expression been examined during inflammation. Accordingly, C3aR and C5aR expression in normal human and murine lung was determined in this study by immunohistochemistry and in situ hybridization. In addition, the expression of these receptors was delineated in mice subjected to LPS- and OVA-induced models of inflammation. Under noninflamed conditions, C3aR and C5aR protein and mRNA were expressed by bronchial epithelial and smooth muscle cells of both human and mouse lung. C3aR expression increased significantly on both bronchial epithelial and smooth muscle cells in mice treated with LPS; however, in the OVA-challenged animals only the bronchial smooth muscle cells showed increased C3aR expression. C5aR expression also increased significantly on bronchial epithelial cells in mice treated with LPS, but was not elevated in either cell type in the OVA-challenged mice. These results demonstrate the expression of C3aR and C5aR by cells endogenous to the lung, and, given the participation of bronchial epithelial and smooth muscle cells in the pathology of diseases such as sepsis and asthma, the data suggest a role for these receptors during lung inflammation.

“…PCR was performed as described previously (24). The following primers were used: CD3␦ sense 5Ј-CTG GAC CTG GGA AAA CGC ATC and antisense 5Ј-GTA CTG AGC ATC ATC TCG ATC, resulting in a 309-bp product; C3aR: sense 5Ј-TGA AGC CTT CAG CTA CTG TCT CAG and antisense 5Ј-GGA CAA TGA TGGA GGG GAT GAG, based on the published sequence of the human C3aR (18). An aliquot of each PCR product was subjected to electrophoresis on a 2% agarose gel (Qualex Gold; AGS, Heidelberg, Germany), stained with ethidium bromide, visualized, and photographed under ultraviolet illumination.…”

Section: Pcrmentioning

confidence: 99%

“…The recently cloned human C3aR (3) belongs to the large family of G protein-coupled receptors with seven transmembrane segments (17)(18)(19). It represents the only as yet characterized receptor for C3a and does not bind C3a(desArg) (20).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Activated Human T Lymphocytes Express a Functional C3a Receptor

Werfel¹,

Kirchhoff²,

Wittmann³

et al. 2000

113

The C3a molecule is an anaphylatoxin of the C system with a wide spectrum of proinflammatory effects predominantly on cells of myeloid origin. In this study we investigated the expression of the high affinity receptor for C3a (C3aR) in human T lymphocytes using receptor-specific mAb. C3aR expression was detected in CD4+ and CD8+ blood- or skin-derived T cell clones (TCC) from birch pollen-sensitized patients with atopic dermatitis. No significant difference in C3aR expression in CD4+ or CD8+ TCCs could be observed. In contrast to C3a(desArg), C3a led to a transient calcium flux in TCCs expressing the C3aR, whereas C3aR-negative TCCs were unreactive. Circulating T cells from patients suffering from severe inflammatory skin diseases expressed the C3aR, whereas no expression of C3aR could be found in unstimulated T lymphocytes from patients with mild inflammatory skin diseases or from healthy individuals. Type I IFNs, which are potent stimulators of cellular immunity, were identified as up-regulators of C3aR expression in vitro in freshly isolated or cloned T lymphocytes. Moreover, C3aR+ T cells were found at the sites of injection in IFN-β-treated patients with multiple sclerosis. These data provide direct evidence for the expression of C3aR on activated human T lymphocytes; this may point to a biological function of C3a in T cell-dependent diseases.