RESUMO
The tumor microenvironment affects malignancy in hepatocellular carcinoma (HCC) cells, and cancer-associated fibroblasts (CAFs) play an important role in the microenvironment. As recent studies indicated a difference between CAFs isolated from chemoresistant and non-resistant cancer tissues, therefore we investigated the intracellular mechanism in resistant HCC co-cultured CAFs and interactions between these CAFs with cancer cells. We established a sorafenib-resistant (SR) Huh7 (human HCC) cell line, and characterized it with cytokine assays, then developed CAFs by co-culturing human hepatic stellate cells with resistant or parental Huh7 cells. The 2 types of CAFs were co-cultured with parental Huh7 cells, thereafter the cell viability of these Huh7 cells was checked under sorafenib treatment. The SR Huh7 (Huh7SR ) cells expressed increased B-cell activating factor (BAFF), which promoted high expression of CAF-specific markers in Huh7SR -co-cultured CAFs, showed activated BAFF, BAFF-R, and downstream of the NFκB-Nrf2 pathway, and aggravated invasion, migration, and drug resistance in co-cultured Huh7 cells. When we knocked down BAFF expression in Huh7SR cells, the previously increased malignancy and BAFF/NFκB axis in Huh7SR -co-cultured CAFs reversed, and enhanced chemoresistance in co-cultured Huh7 cells returned as well. In conclusion, the BAFF/NFκB pathway was activated in CAFs co-cultured with cell-culture medium from resistant Huh7, which promoted chemoresistance, and increased the malignancy in co-cultured non-resistant Huh7 cells. This suggests that the BAFF/NFκB axis in CAFs might be a potential therapeutic target in chemoresistance of HCC.
Assuntos
Antineoplásicos/farmacologia , Fator Ativador de Células B/metabolismo , Fibroblastos Associados a Câncer/metabolismo , Carcinoma Hepatocelular/metabolismo , Comunicação Celular/genética , Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias Hepáticas/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais/genética , Sorafenibe/farmacologia , Fator Ativador de Células B/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Técnicas de Cocultura , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , TransfecçãoRESUMO
BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer-associated mortality worldwide with an overall five-year survival rate less than 7%. Accumulating evidence has revealed the cancer preventive and therapeutic effects of metformin, one of the most widely prescribed medications for type 2 diabetes mellitus. However, its role in pancreatic cancer is not fully elucidated. Herein, we aimed to further study the preventive and therapeutic effects of metformin in genetically engineered mouse models of pancreatic cancer. METHODS: LSL-KrasG12D/+; Pdx1-Cre (KC) mouse model was established to investigate the effect of metformin in pancreatic tumorigenesis suppression; LSL-KrasG12D/+; Trp53fl/+; Pdx1-Cre (KPC) mouse model was used to evaluate the therapeutic efficiency of metformin in PDAC. Chronic pancreatitis was induced in KC mice by peritoneal injection of cerulein. RESULTS: Following metformin treatment, pancreatic acinar-to-ductal metaplasia (ADM) and mouse pancreatic intraepithelial neoplasia (mPanIN) were decreased in KC mice. Chronic pancreatitis induced a stroma-rich and duct-like structure and increased the formation of ADM and mPanIN lesions, in line with an increased cytokeratin 19 (CK19)-stained area. Metformin treatment diminished chronic pancreatitis-mediated ADM and mPanIN formation. In addition, it alleviated the percent area of Masson's trichrome staining, and decreased the number of Ki67-positive cells. In KPC mice, metformin inhibited tumor growth and the incidence of abdominal invasion. More importantly, it prolonged the overall survival. CONCLUSIONS: Metformin inhibited pancreatic cancer initiation, suppressed chronic pancreatitis-induced tumorigenesis, and showed promising therapeutic effect in PDAC.
Assuntos
Carcinogênese/efeitos dos fármacos , Metformina/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Animais , Carcinogênese/metabolismo , Carcinoma in Situ/tratamento farmacológico , Carcinoma in Situ/metabolismo , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Queratina-19/metabolismo , Antígeno Ki-67/metabolismo , Camundongos , Camundongos Transgênicos , Pâncreas/efeitos dos fármacos , Ductos Pancreáticos/efeitos dos fármacos , Ductos Pancreáticos/metabolismo , Neoplasias Pancreáticas/metabolismo , Pancreatite Crônica/tratamento farmacológico , Pancreatite Crônica/metabolismoRESUMO
Exploring non-genetic evolution of cell states during cancer treatments has become attainable by recent advances in lineage-tracing methods. However, transcriptional changes that drive cells into resistant fates may be subtle, necessitating high resolution analysis. Here, we present ReSisTrace that uses shared transcriptomic features of sister cells to predict the states priming treatment resistance. Applying ReSisTrace in ovarian cancer cells perturbed with olaparib, carboplatin or natural killer (NK) cells reveals pre-resistant phenotypes defined by proteostatic and mRNA surveillance features, reflecting traits enriched in the upcoming subclonal selection. Furthermore, we show that DNA repair deficiency renders cells susceptible to both DNA damaging agents and NK killing in a context-dependent manner. Finally, we leverage the obtained pre-resistance profiles to predict and validate small molecules driving cells to sensitive states prior to treatment. In summary, ReSisTrace resolves pre-existing transcriptional features of treatment vulnerability, facilitating both molecular patient stratification and discovery of synergistic pre-sensitizing therapies.
Assuntos
Células Matadoras Naturais , Neoplasias Ovarianas , Humanos , Feminino , Neoplasias Ovarianas/genética , Carboplatina , Fenótipo , Linhagem Celular TumoralRESUMO
Generation of hepatocytes from human adipose-derived mesenchymal stem cells (hADSCs) could be a promising alternative source of human hepatocytes. However, mechanisms to differentiate hepatocytes from hADSCs are not fully elucidated. We have previously demonstrated that our three-step differentiation protocol with glycogen synthase kinase (GSK) 3 inhibitor was effective to improve hepatocyte functions. In this study, we investigated the activation of the nuclear factor erythroid-2 related factor 2 (Nrf2) on hADSCs undergoing differentiation to HLC (hepatocyte-like cells). Our three-step differentiation protocol was applied for 21 days (Step 1:day 1-6, Step2:day 6-11, Step3:day 11-21). Our results show that significant nuclear translocation of Nrf2 occurred from day 11 until the end of HLC differentiation. Nuclear translocation of Nrf2 and CYP3A4 activity in the GSK3 inhibitor-treated group was obviously higher than that in Activin A-treated groups at day 11. The maturation of HLCs was delayed in Nrf2-siRNA group compared to control group. Furthermore, CYP3A4 activity in Nrf2-siRNA group was decreased at the almost same level in Activin A-treated group. Nrf2 translocation might enhance the function of HLC and be a target for developing highly functional HLC. J. Med. Invest. 70 : 343-349, August, 2023.
Assuntos
Quinase 3 da Glicogênio Sintase , Fator 2 Relacionado a NF-E2 , Humanos , Fator 2 Relacionado a NF-E2/farmacologia , Quinase 3 da Glicogênio Sintase/farmacologia , Citocromo P-450 CYP3A/farmacologia , Hepatócitos , Diferenciação Celular/genética , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologiaRESUMO
BACKGROUND AND AIM: As a multiple tyrosine kinase inhibitor, sorafenib is widely used to treat hepatocellular carcinoma (HCC), but patients frequently face resistance problems. Because the mechanism controlling sorafenib-resistance is not well understood, this study focused on the connection between tumor characteristics and the Nrf2 signaling pathway in a sorafenib-resistant HCC cell line. METHODS: A sorafenib-resistant HCC cell line (Huh7) was developed by increasing the dose of sorafenib in the culture medium until the target concentration was reached. Cell morphology, migration/invasion rates, and expression of stemness-related and ATP-binding cassette (ABC) transporter genes were compared between sorafenib-resistant Huh7 cells and parental Huh7 cells. Next, a small interfering RNA was used to knock down Nrf2 expression in sorafenib-resistant Huh7 cells, after which cell viability, stemness, migration, and ABC transporter gene expression were examined again. RESULTS: Proliferation, migration, and invasion rates of sorafenib-resistant Huh7 cells were significantly increased relative to the parental cells with or without sorafenib added to the medium. The expression levels of stemness markers and ABC transporter genes were up-regulated in sorafenib-resistant cells. After Nrf2 was knocked down in sorafenib-resistant cells, cell migration and invasion rates were reduced, and expression levels of stemness markers and ABC transporter genes were reduced. CONCLUSION: Nrf2 signaling promotes cancer stemness, migration, and expression of ABC transporter genes in sorafenib-resistant HCC cells.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Antineoplásicos/farmacologia , Carcinoma Hepatocelular/metabolismo , Movimento Celular/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Expressão Gênica , Neoplasias Hepáticas/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais/genética , Sorafenibe/farmacologia , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Resistencia a Medicamentos Antineoplásicos/genética , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Fator 2 Relacionado a NF-E2/genética , Transdução de Sinais/efeitos dos fármacos , TransfecçãoRESUMO
OBJECTIVES: We aimed to determine whether responsive insulin-producing cells (IPCs) could be generated from adipose-derived stem cells (ADSCs) isolated from patients with type 1 diabetes mellitus (T1DM). METHODS: We isolated ADSCs from adipose tissue of 4 patients (one patient with T1DM and 3 nondiabetic patients), who underwent surgery and differentiated them into IPCs with using a 2-step xeno-antigen free, 3-dimensional culture method. Characteristics of isolated ADSCs, in vitro cell quality, programmed cell death ligand-1 (PDL-1) expression, and transplantation into streptozotocin induced diabetic nude mice were investigated. RESULTS: Adipose-derived stem cells from T1DM patients and commercially obtained ADSCs showed the same surface markers; CD31CD34CD45CD90CD105CD146. Moreover, the generated IPCs at day 21 demonstrated appropriate autonomous insulin secretion (stimulation index, 3.5; standard deviation, 0.8). Nonfasting blood glucose concentrations of IPC-transplanted mice were normal at 30 days. The normalized rate of IPC-transplanted mice was significantly higher than that of the sham-operated group (P < 0.05). Insulin-producing cells generated from T1DM adipose tissue expressed high levels of PDL-1. CONCLUSIONS: Insulin-producing cells obtained from adipose tissue of T1DM patients are capable of secreting insulin long-term and achieve normoglycemia after transplantation. Expression of PDL-1 suggests the potential for immune circumvention.
Assuntos
Tecido Adiposo/citologia , Diabetes Mellitus Experimental/terapia , Diabetes Mellitus Tipo 1/patologia , Células Secretoras de Insulina/citologia , Transplante de Células-Tronco/métodos , Células-Tronco/citologia , Animais , Células Cultivadas , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Glucose/farmacologia , Humanos , Insulina/metabolismo , Secreção de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/ultraestrutura , Camundongos Nus , Microscopia Eletrônica de Transmissão , Células-Tronco/metabolismoRESUMO
Resveratrol, a natural polyphenol present in most plants, inhibits the growth of numerous cancers both in vitro and in vivo. Aberrant expression of YAP has been reported to activate multiple growth-regulatory pathways and confer anti-apoptotic abilities to many cancer cells. However, the role of resveratrol in YES-activated protein (YAP) expression and that of YAP in pancreatic cancer cells' response to gemcitabine resistance remain elusive. In this study, we found that resveratrol suppressed the proliferation and cloning ability and induced the apoptosis of pancreatic cancer cells. These multiple biological effects might result from the activation of AMP-activation protein kinase (AMPK) (Thr172) and, thus, the induction of YAP cytoplasmic retention, Ser127 phosphorylation, and the inhibition of YAP transcriptional activity by resveratrol. YAP silencing by siRNA or resveratrol enhanced the sensitivity of gemcitabine in pancreatic cancer cells. Taken together, these findings demonstrate that resveratrol could increase the sensitivity of pancreatic cancer cells to gemcitabine by inhibiting YAP expression. More importantly, our work reveals that resveratrol is a potential anticancer agent for the treatment of pancreatic cancer, and YAP may serve as a promising target for sensitizing pancreatic cancer cells to chemotherapy.