RESUMEN
BACKGROUND: Promising outcomes have been observed in multiple myeloma (MM) with the use of immunotherapies, specifically chimeric antigen receptor T (CAR-T) cell therapy. However, a portion of MM patients do not respond to CAR-T therapy, and the reasons for this lack of response remain unclear. The objective of this study was to investigate the impact of miR-34a on the immunosuppressive polarization of macrophages obtained from MM patients. METHODS: The levels of miR-34a and TLR9 (Toll-like receptor 9) were examined in macrophages obtained from both healthy individuals and patients with MM. ELISA was employed to investigate the cytokine profiles of the macrophage samples. Co-culture experiments were conducted to evaluate the immunomodulatory impact of MM-associated macrophages on CAR-T cells. RESULTS: There was an observed suppressed activation of macrophages and CD4+ T lymphocytes in the blood samples of MM patients. Overexpression of miR-34a in MM-associated macrophages dampened the TLR9 expression and impaired the inflammatory polarization. In both the co-culture system and an animal model, MM-associated macrophages suppressed the activity and tumoricidal effect of CAR-T cells in a miR-34a-dependent manner. CONCLUSION: The findings imply that targeting the macrophage miR-34a/TLR9 axis could potentially alleviate the immunosuppression associated with CAR-T therapy in MM patients.
Asunto(s)
MicroARNs , Mieloma Múltiple , Transducción de Señal , Receptor Toll-Like 9 , Mieloma Múltiple/inmunología , Mieloma Múltiple/genética , Mieloma Múltiple/terapia , Mieloma Múltiple/metabolismo , MicroARNs/genética , Receptor Toll-Like 9/metabolismo , Receptor Toll-Like 9/genética , Humanos , Animales , Ratones , Técnicas de Cocultivo , Macrófagos Asociados a Tumores/inmunología , Macrófagos Asociados a Tumores/metabolismo , Macrófagos/inmunología , Macrófagos/metabolismo , Inmunoterapia Adoptiva/métodos , Masculino , Femenino , Activación de Macrófagos/inmunología , Activación de Macrófagos/genética , Línea Celular TumoralRESUMEN
Multiple myeloma (MM) progression involves diminished tumor antigen presentation and an immunosuppressive microenvironment, characterized by diminished expression of major histocompatibility complexes (MHC) class I molecule and elevated programmed death ligand 1 (PDL1) in MM cells, along with an enriched population of regulatory T cells (Tregs). To investigate Treg's influence on MM cells, we established a co-culture system using Tregs from MM patients and the MM cell lines (MM.1S and SK-MM-1) in vitro and assessed the effects of intervening in the relevant pathways connecting Tregs and MM cells in vivo. In vitro, Tregs induced transforming growth factor beta-1 (TGF-ß1) production, downregulated MHC I members, and increased PDL1 expression in MM cells. Treg-derived TGF-ß1 suppressed the cGAS-STING pathway, contributing to the loss of MHC I molecule expression and PDL1 upregulation. Correspondingly, neutralizing TGF-ß1 or activating the cGAS-STING pathway restored MHC I and PDL1 expression, effectively countering the pro-tumorigenic effect of Tregs on MM cells in vivo. These data elucidated how Tregs influence tumor antigen presentation and immunosuppressive signal in MM cells, potentially providing therapeutic strategies, such as neutralizing TGF-ß1 or activating the cGAS-STING pathway, to address the immune escape and immunosuppressive dynamics in MM.
Asunto(s)
Antígeno B7-H1 , Antígenos de Histocompatibilidad Clase I , Proteínas de la Membrana , Mieloma Múltiple , Nucleotidiltransferasas , Transducción de Señal , Linfocitos T Reguladores , Factor de Crecimiento Transformador beta1 , Humanos , Mieloma Múltiple/metabolismo , Mieloma Múltiple/inmunología , Mieloma Múltiple/patología , Mieloma Múltiple/genética , Factor de Crecimiento Transformador beta1/metabolismo , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Antígenos de Histocompatibilidad Clase I/metabolismo , Antígenos de Histocompatibilidad Clase I/genética , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Nucleotidiltransferasas/metabolismo , Nucleotidiltransferasas/genética , Antígeno B7-H1/metabolismo , Antígeno B7-H1/genética , Línea Celular Tumoral , Animales , Regulación hacia Abajo , Ratones , Femenino , Técnicas de Cocultivo , Masculino , Regulación Neoplásica de la Expresión GénicaRESUMEN
Cytoplasmic lncRNAs have been found to directly interact with target mRNAs and regulate their stability. In this study, we aimed to study the molecular mechanism underlying the function of m6 A as a central regulator in chemoresistance and CML proliferation. In this study, we established three mice groups (control group, ADR-R group and ADR-R + shLINC00470 group). We detected PTEN mRNA expression in the presence of LINC00470 in the mice models, as well as in the KCL22 and K562 cells. LINC00470 was significantly enriched for PTEN mRNA to exhibit a negative regulatory relationship between LINC00470 and PTEN mRNA. However, the alteration of LINC00470 had no effect on the luciferase activity of PTEN promoter, while the half-life of PTEN mRNA was affected. It was further validated that LINC00470 down-regulated PTEN expression by positively regulating the m6A modification of PTEN mRNA via RNA methyltransferase METTL3. Moreover, the relative expression of LC3II, Beclin-1, ATG7 and ATG5 was all decreased in cells treated with LINC00470, and down-regulated PTEN expression was observed in chemo-resistant cells, while the expression of PTEN was rescued by the transfection of shMETTL3 into chemo-resistant cells. Moreover, the knockdown of METTL3 also restored the normal level of PTEN m6 A modification and LINC00470 expression in chemo-resistant cells. In conclusion, our results demonstrated the molecular mechanism underlying the effect of LINC00470 on CML by reducing the PTEN stability via RNA methyltransferase METTL3, thus leading to the inhibition of cell autophagy while promoting chemoresistance in CML.
Asunto(s)
Autofagia , Biomarcadores de Tumor/metabolismo , Resistencia a Antineoplásicos , Regulación Neoplásica de la Expresión Génica , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Metiltransferasas/metabolismo , ARN Largo no Codificante/genética , Animales , Antineoplásicos/farmacología , Apoptosis , Biomarcadores de Tumor/genética , Proliferación Celular , Humanos , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Masculino , Metiltransferasas/genética , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Fosfohidrolasa PTEN , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The aim of the present study was to investigate the effect of coculture with amniotic epithelial cells (AECs) on the biological characteristics of amniotic mesenchymal stem cells (AMSCs), to compare the expression of CXC motif chemokine receptor 4 (CXCR4) in cocultured AMSCs and to investigate the roles of the stromal cellderived factor1 (SDF1)/CXCR4 axis in the homing and migration of AMSCs. AMSCs were isolated from human amniotic membranes, purified and then differentiated into osteoblasts and adipocytes in vitro, which was verified by von Kossa Staining and Oil Red O staining. Cell viability was measured by Cell Counting kit8 and trypan blue assays at 24, 48 and 72 h, the expression of CXCR4 was analyzed by immunofluorescencebased flow cytometry and reverse transcriptionquantitative polymerase chain reaction, and the migration ability of AMSCs in vitro was observed by a migration assay. The results demonstrated that cell viability (at 48 and 72 h) and survival (at 24, 48 and 72 h) in the coculture and serum groups were higher compared with the serumfree group. Furthermore, CXCR4 mRNA and protein expression, and migration along the SDF1 gradient, in the coculture and serumfree groups were higher compared with the serum group. Overall, the results indicated that AMSCs cocultured with AECs exhibited enhanced proliferation activity and survival rate. In conclusion, the present study demonstrated that coculture of AMSCs with AECs upregulated CXCR4 on the surface of AMSCs and enhanced the migration ability of AMSCs in vitro. This result may improve the directional migration and homing ability of AMSCs, as well as provide a theoretical basis for the application of AMSCs in clinical practice as a novel strategy to increase the success of hematopoietic stem cell transplantation.
