RESUMO
A variety of G protein-coupled receptors (GPCRs) is expressed in hematopoietic stem and progenitor cells (HPCs), including the chemokine receptor CXCR4, the leukotriene receptor CysLT1, the sphingosine 1-phosphate receptor S1P1, the cannabinoid receptor CB2, and the complement receptor C3aR. While the role of CXCR4 in stem cell homing is largely established, the function of the other GPCRs expressed in HPCs is only partially understood. CXCR4 and CysLT1 inhibit their own activation after ligand binding (homologous desensitization). Stimulation of S1P1 or C3aR has been shown to activate CXCR4 in HPCs that may sensitize CXCR4-dependent stem cell homing. In contrast, activation of CXCR4 results in a loss of CysLT1 function, which is most likely mediated by protein kinase C (PKC) signaling (heterologous desensitization) and could explain the ineffectiveness of CysLT1 antagonists to mobilize HPCs in vivo. Further characterization of GPCR crosstalk will allow a better understanding of HPC trafficking.
Assuntos
Células-Tronco Hematopoéticas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animais , Movimento Celular , Humanos , Camundongos , Modelos Biológicos , Receptor Cross-Talk , Receptor CB2 de Canabinoide/metabolismo , Receptores CXCR4/metabolismo , Receptores de Complemento/metabolismo , Receptores de Leucotrienos/metabolismo , Receptores de Lisoesfingolipídeo/metabolismo , Transdução de SinaisRESUMO
The G protein-coupled receptor (GPCR) CXCR4 is involved in bone marrow tropism and survival of chronic lymphocytic leukemia (CLL) cells. The function of the GPCRs cysteinyl leukotriene receptor 1 (CysLT1) and CysLT2 remains elusive. Here we demonstrate that in CLL and normal B lymphocytes, CysLT1 mRNA is consistently expressed, in contrast to low CysLT2 levels. Similar to the CXCR4 ligand CXCL12, the cysteinyl leukotriene (cysLT) LTD(4) induces calcium fluxes, actin polymerization, and chemotaxis. These effects are blocked by specific CysLT1 antagonists. Their inhibition by pertussis toxin suggests Giα/o protein involvement. Furthermore, CysLT1 mediates MAP-kinase phosphorylation, which implicates contribution of cysLT to survival. Indeed, CysLT1 antagonists induce apoptosis and reduce viability independent of Gαi/o protein signaling. Considering the production of cysLTs in the bone marrow, our data suggest that CysLT1 induces chemokine-like effects, supports accumulation and survival of CLL cells in the bone marrow and thus represents a potential treatment target.
Assuntos
Antagonistas de Leucotrienos/farmacologia , Leucotrieno D4/farmacologia , Receptores de Leucotrienos/genética , Acetofenonas/farmacologia , Actinas/metabolismo , Idoso , Apoptose/efeitos dos fármacos , Western Blotting , Cálcio/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Quimiocinas/farmacologia , Quimiotaxia/efeitos dos fármacos , Cisteína/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Células Jurkat , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Linfocítica Crônica de Células B/patologia , Leucotrienos/farmacologia , Masculino , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosforilação/efeitos dos fármacos , Propionatos/farmacologia , Quinolinas/farmacologia , Receptores de Leucotrienos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Tetrazóis/farmacologia , Fatores de Tempo , Células Tumorais CultivadasRESUMO
To explore a new treatment strategy for urinary incontinence, human bone marrow mesenchymal stem cells (MSC) of the first in vitro passage were exposed to 5-azacytidine (AZA) to induce myogenic differentiation, and cultured for a total of six passages. Expression of stem cell surface antigens and intracellular alpha-actin was examined by flow cytometry at the end of each passage and compared to that of native MSC (not exposed to AZA) cultured in parallel. To analyze differentiation into striated muscle, expression of the transcription factor MyoD1 and myosin heavy chain (MyHC) was examined by RT-PCR. Both native and AZA-exposed MSC of all passages were negative for the progenitor/endothelial antigen CD34, leukocytic CD45, and endothelial/monocytic CD31. In contrast, the MSC markers CD73, CD90, CD105, and intracellular actin were detected in both groups of MSC throughout the culture period. After an initial increase, the expression level of MSC antigens decreased over time particularly in AZA-exposed MSC. Expression of smooth muscle alpha-actin also declined, but was greater in AZA-exposed MSC throughout the culture period. Varying percentages of MSC cultures expressed MyoD1 and MyHC mRNA. In late passages, AZA-exposed MSC tended to be more frequently positive than native MSC. In pilot experiments, transplantation of MSC into the bladder neck tissue of athymic rats was feasible; long-term analyses are pending. We conclude that independent of AZA exposure, MSC express smooth and striated muscle antigens. Treatment with AZA slightly increases myogenic differentiation, but may not be necessary in future studies of MSC as a treatment modality for urinary incontinence.