RESUMEN
Myostatin (Mstn) is postulated to be a key determinant of muscle loss and cachexia in cancer. However, no experimental evidence supports a role for Mstn in cancer, particularly in regulating the survival and growth of cancer cells. In this study, we showed that the expression of Mstn was significantly increased in different tumor tissues and human cancer cells. Mstn knockdown inhibited the proliferation of cancer cells. A knockout (KO) of Mstn created by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) 9 (CRISPR/Cas9) induced mitochondria-dependent apoptosis in HeLa cells. Furthermore, KO of Mstn reduced the lipid content. Molecular analyses demonstrated that the expression levels of fatty acid oxidation-related genes were upregulated and then increased rate of fatty acid oxidation. Mstn deficiency-induced apoptosis took place along with generation of reactive oxygen species (ROS) and elevated fatty acid oxidation, which may play a role in triggering mitochondrial membrane depolarization, the release of cytochrome c (Cyt-c), and caspase activation. Importantly, apoptosis induced by Mstn KO was partially rescued by antioxidants and etomoxir, thereby suggesting that the increased level of ROS was functionally involved in mediating apoptosis. Overall, our findings demonstrate a novel function of Mstn in regulating mitochondrial metabolism and apoptosis within cancer cells. Hence, inhibiting the production and function of Mstn may be an effective therapeutic intervention during cancer progression and muscle loss in cachexia.
Asunto(s)
Apoptosis/genética , Caquexia/patología , Miostatina/genética , Especies Reactivas de Oxígeno/metabolismo , Neoplasias del Cuello Uterino/patología , Células A549 , Animales , Antioxidantes/farmacología , Sistemas CRISPR-Cas/genética , Caspasas/metabolismo , Línea Celular Tumoral , Proliferación Celular/genética , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas/genética , Citocromos c/metabolismo , Compuestos Epoxi/farmacología , Ácidos Grasos/metabolismo , Femenino , Técnicas de Inactivación de Genes , Células HEK293 , Células HeLa , Humanos , Metabolismo de los Lípidos/fisiología , Potencial de la Membrana Mitocondrial/genética , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Mitocondrias/genética , Mitocondrias/metabolismo , Oxidación-Reducción , Neoplasias del Cuello Uterino/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
OBJECTIVE: The purpose of the article is to evaluate the changes in lipid metabolism in bovine mammary-gland epithelial MAC-T cells after PKM2 knockdown. RESULTS: MAC-T cells stably expressing low levels of PKM2 were established with lentivirus-mediated small hairpin RNA. Although the knockdown of PKM2 had no effect on MAC-T cell growth, the reduced expression of PKM2 attenuated the mRNA and protein expression of key enzymes involved in sterol synthesis through the SREBP pathway. CONCLUSIONS: The downregulation of PKM2 significantly influenced lipid synthesis in bovine mammary-gland epithelial MAC-T cells. These findings extend our understanding of the crosstalk between glycolysis and lipid metabolism in bovine mammary-gland epithelial cells.
Asunto(s)
Proteínas Portadoras/genética , Metabolismo de los Lípidos/genética , Glándulas Mamarias Animales/metabolismo , Proteínas de la Membrana/genética , Proteínas de Unión a los Elementos Reguladores de Esteroles/genética , Hormonas Tiroideas/genética , Animales , Proteínas Portadoras/metabolismo , Bovinos , Células Epiteliales/metabolismo , Femenino , Técnicas de Silenciamiento del Gen , Glucólisis/genética , Lípidos/biosíntesis , Proteínas de la Membrana/metabolismo , ARN Mensajero/genética , Transducción de Señal , Proteínas de Unión a los Elementos Reguladores de Esteroles/metabolismo , Linfocitos T/metabolismo , Hormonas Tiroideas/metabolismo , Proteínas de Unión a Hormona TiroideRESUMEN
IFITM3 is involved in cell adhesion, apoptosis, immune, and antivirus activity. Furthermore, IFITM3 gene has been considered as a preferential marker for inflammatory diseases, and positive correlation to pathological grades. Therefore, we assumed that IFITM3 was regulated by different signal pathways. To better understand IFITM3 function in inflammatory response, we cloned swine IFITM3 gene, and detected IFITM3 distribution in tissues, as well as characterized this gene. Results indicated that the length of swine IFITM3 gene was 438 bp, encoding 145 amino acids. IFITM3 gene expression abundance was higher in spleen and lungs. Moreover, we next constructed the eukaryotic expression vector PBIFM3 and transfected into PK15 cells, finally obtained swine IFITM3 gene stable expression cell line. Meanwhile, we explored the effects of LPS on swine IFITM3 expression. Results showed that LPS increased IFITM3 mRNA abundance and exhibited time-dependent effect for LPS treatment. To further demonstrate the mechanism that IFITM3 regulated type I IFNs production, we also detected the important molecules expression of TLR4 signaling pathway. In transfected and non-transfected IFITM3 PK15 cells, LPS exacerbated the relative expression of TLR4-NFκB signaling molecules. However, the IFITM3 overexpression suppressed the inflammatory development of PK15 cells. In conclusion, these data indicated that the overexpression of swine IFITM3 could decrease the inflammatory response through TLR4 signaling pathway, and participate in type I interferon production. These findings may lead to an improved understanding of the biological function of IFITM3 in inflammation.