RESUMO
Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations in circulating tumor deoxyribonucleic acid (ctDNA) have been reported as representative noninvasive prognostic markers for pancreatic ductal adenocarcinoma (PDAC). Here, we aimed to evaluate single KRAS mutations as prognostic and predictive biomarkers, with an emphasis on potential therapeutic approaches to PDAC. A total of 128 patients were analyzed for multiple or single KRAS mutations (G12A, G12C, G12D, G12R, G12S, G12V, and G13D) in their tumors and plasma using droplet digital polymerase chain reaction (ddPCR). Overall, KRAS mutations were detected by multiplex ddPCR in 119 (93%) of tumor DNA and 68 (53.1%) of ctDNA, with a concordance rate of 80% between plasma ctDNA and tumor DNA in the metastatic stage, which was higher than the 44% in the resectable stage. Moreover, the prognostic prediction of both overall survival (OS) and progression-free survival (PFS) was more relevant using plasma ctDNA than tumor DNA. Further, we evaluated the selective tumor-suppressive efficacy of the KRAS G12C inhibitor sotorasib in a patient-derived organoid (PDO) from a KRAS G12C-mutated patient using a patient-derived xenograft (PDX) model. Sotorasib showed selective inhibition in vitro and in vivo with altered tumor microenvironment, including fibroblasts and macrophages. Collectively, screening for KRAS single mutations in plasma ctDNA and the use of preclinical models of PDO and PDX with genetic mutations would impact precision medicine in the context of PDAC.
Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Proteínas Proto-Oncogênicas p21(ras)/genética , Biomarcadores Tumorais/genética , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/diagnóstico , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , DNA de Neoplasias/genética , Mutação , Microambiente TumoralRESUMO
Pancreatic ductal adenocarcinoma (PDAC) has a dismal prognosis due to the absence of diagnostic markers and molecular targets. Here, we took an unconventional approach to identify new molecular targets for pancreatic cancer. We chose uncharacterized protein evidence level 1 without function annotation from extensive proteomic research on pancreatic cancer and focused on proline and serine-rich 2 (PROSER2), which ranked high in the cell membrane and cytoplasm. In our study using cell lines and patient-derived orthotopic xenograft cells, PROSER2 exhibited a higher expression in cells derived from primary tumors than in those from metastatic tissues. PROSER2 was localized in the cell membrane and cytosol by immunocytochemistry. PROSER2 overexpression significantly reduced the metastatic ability of cancer cells, whereas its suppression had the opposite effect. Proteomic analysis revealed that PROSER2 interacts with STK25 and PDCD10, and their binding was confirmed by immunoprecipitation and immunocytochemistry. STK25 knockdown enhanced metastasis by decreasing p-AMPK levels, whereas PROSER2-overexpressing cells increased the level of p-AMPK, indicating that PROSER2 suppresses invasion via the AMPK pathway by interacting with STK25. This is the first demonstration of the novel role of PROSER2 in antagonizing tumor progression via the STK25-AMPK pathway in PDAC. LC-MS/MS data are available at MassIVE (MSV000092953) and ProteomeXchange (PXD045646).
Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Animais , Humanos , Proteínas Quinases Ativadas por AMP , Cromatografia Líquida , Proteômica , Proliferação de Células , Movimento Celular , Espectrometria de Massas em Tandem , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/genética , Modelos Animais de Doenças , Proteínas Serina-Treonina Quinases , Peptídeos e Proteínas de Sinalização IntracelularRESUMO
PURPOSE: Patient-derived tumor cells can be a powerful resource for studying pathophysiological mechanisms and developing robust strategies for precision medicine. However, establishing organoids from patient-derived cells is challenging because of limited access to tissue specimens. Therefore, we aimed to establish organoids from malignant ascites and pleural effusions. MATERIALS AND METHODS: Ascitic or pleural fluid from pancreatic, gastric, and breast cancer patients was collected and concentrated to culture tumor cells ex vivo. Organoids were considered to be successfully cultured when maintained for five or more passages. Immunohistochemical staining was performed to compare the molecular features, and drug sensitivity was assayed to analyze the clinical responses of original patients. RESULTS: We collected 70 fluid samples from 58 patients (pancreatic cancer, n=39; gastric cancer, n=21; and breast cancer, n=10). The overall success rate was 40%; however, it differed with types of malignancy, with pancreatic, gastric, and breast cancers showing 48.7%, 33.3%, and 20%, respectively. Cytopathological results significantly differed between successful and failed cases (p=0.014). Immunohistochemical staining of breast cancer organoids showed molecular features identical to those of tumor tissues. In drug sensitivity assays, pancreatic cancer organoids recapitulated the clinical responses of the original patients. CONCLUSION: Tumor organoids established from malignant ascites or pleural effusion of pancreatic, gastric, and breast cancers reflect the molecular characteristics and drug sensitivity profiles. Our organoid platform could be used as a testbed for patients with pleural and peritoneal metastases to guide precision oncology and drug discovery.
Assuntos
Neoplasias da Mama , Neoplasias Pancreáticas , Neoplasias Peritoneais , Humanos , Feminino , Ascite , Medicina de Precisão , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Neoplasias Peritoneais/patologia , Organoides/patologia , Neoplasias PancreáticasRESUMO
BACKGROUND & AIMS: Intrahepatic cholangiocarcinomas (iCCs) are characterized by their rarity, difficult diagnosis, and overall poor prognosis. The iCC molecular classification for developing precision medicine strategies was investigated. METHODS: Comprehensive genomic, transcriptomic, proteomic, and phosphoproteomic analyses were performed on treatment-naïve tumor samples from 102 patients with iCC who underwent surgical resection with curative intent. An organoid model was constructed for testing therapeutic potential. RESULTS: Three clinically supported subtypes (stem-like, poorly immunogenic, and metabolism) were identified. NCT-501 (aldehyde dehydrogenase 1 family member A1 [ALDH1A1] inhibitor) exhibited synergism with nanoparticle albumin-bound-paclitaxel in the organoid model for the stem-like subtype. The oncometabolite dysregulations were associated with different clinical outcomes in the stem-like and metabolism subtypes. The poorly immunogenic subtype harbors the non-T-cell tumor infiltration. Integrated multiomics analysis not only reproduced the 3 subtypes but also showed heterogeneity in iCC. CONCLUSIONS: This large-scale proteogenomic analysis provides information beyond that obtained with genomic analysis, allowing the functional impact of genomic alterations to be discerned. These findings may assist in the stratification of patients with iCC and in developing rational therapeutic strategies.
Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Proteogenômica , Humanos , Proteômica , Prognóstico , Colangiocarcinoma/genética , Colangiocarcinoma/cirurgia , Colangiocarcinoma/metabolismo , Ductos Biliares Intra-Hepáticos/patologia , Neoplasias dos Ductos Biliares/patologiaRESUMO
Owing to a lack of reliable markers and therapeutic targets, pancreatic ductal adenocarcinoma (PDAC) remains the most lethal malignant tumor despite numerous therapeutic advances. In this study, we utilized cell-SELEX to isolate a DNA aptamer recognizing the natural conformation of the target on the cell surface. PAp7T8, an aptamer optimized by size and chemical modification, exhibited specific targeting to pancreatic cancer cells and orthotopic xenograft pancreatic tumors. To confer therapeutic functions to the aptamer, we adopted a drug-conjugated oligobody (DOligobody) strategy. Monomethyl auristatin E was used as a cytotoxic drug, digoxigenin acted as a hapten, and the humanized anti-digoxigenin antibody served as a universal carrier of the aptamer. The resulting PAp7T8-DOligobody showed extended in vivo half-life and markedly inhibited tumor growth in an orthotopic pancreatic cancer xenograft model without causing significant toxicity. Therefore, PAp7T8-DOligobody represents a promising novel therapeutic delivery platform for PDAC.
Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Preparações Farmacêuticas , Linhagem Celular Tumoral , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/patologia , Anticorpos , Oligonucleotídeos/uso terapêutico , Ensaios Antitumorais Modelo de Xenoenxerto , Neoplasias PancreáticasRESUMO
To investigate the effects of agar oligosaccharides (AO) on lipid metabolism, changes in obesity phenotypes and related molecular factors were evaluated in C57BL/6N mice fed a high-fat diet (HFD). When HFD-induced obese mice were fed AO, they lost weight. Also, fat accumulation in abdominal and liver tissues was lower in the AO groups than in the Vehicle group. Lipid droplet sizes in tissue sections were reduced by AO, and these observations were mirrored by serum lipid contents. To evaluate the effects of AO on lipid metabolism, lipogenesis and lipolysis-related factors were analyzed. The mRNA expressions of genes involved in lipogenesis, such as adipocyte-protein 2 (aP2) and fatty acid synthase (FAS), were reduced by AO administration, and the expressions of lipolysis-associated proteins, including perilipin, hormone-sensitive lipase (HSL), and fat triglyceride lipase (ATGL), were increased. Taken together, our results suggest that AO should be considered a valuable natural agent that inhibits obesity.
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Dieta Hiperlipídica , Lipólise , Camundongos , Animais , Dieta Hiperlipídica/efeitos adversos , Lipogênese , Ágar/farmacologia , Camundongos Endogâmicos C57BL , Obesidade/etiologia , Obesidade/genética , Camundongos Obesos , Fígado/metabolismo , Oligossacarídeos/farmacologia , Oligossacarídeos/metabolismoRESUMO
BACKGROUND/AIM: Invasive papillary cholangio-carcinoma (IPC) is a minor subtype of extrahepatic cholangiocarcinoma. However, its etiology and characteristics remain unknown because of the unavailability of in vitro and in vivo models. We aimed to establish a novel preclinical model for translational research of IPC. MATERIALS AND METHODS: A patient-derived xenograft (PDX) was engrafted in NOG mice and the cell line National Cancer Center human IPC (NCChIPC) was subsequently established from the PDX tumors. Immunohistochemistry and RNA-sequencing were used to determine the retention of original characteristics of patient tissues. RESULTS: PDX tumors showed successful amplification, and the NCChIPC-derived xenograft largely retained the histopathological features of the original tumor with CK19, MUC1 and MUC5AC expression. Transcriptome analysis showed a high correlation between patient and preclinical models. Additionally, anticancer drugs response was analyzed in the NCChIPC PDX. CONCLUSION: These novel preclinical models here will help elucidate IPC etiology and facilitate translational research.
Assuntos
Carcinoma Papilar/genética , Colangiocarcinoma/genética , Queratina-19/genética , Mucina-5AC/genética , Mucina-1/genética , Idoso , Animais , Antineoplásicos/farmacologia , Carcinoma Papilar/tratamento farmacológico , Carcinoma Papilar/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Colangiocarcinoma/tratamento farmacológico , Colangiocarcinoma/patologia , Modelos Animais de Doenças , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Camundongos , Transcriptoma/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
RATIONALE: In recent years, the use of extracorporeal membrane oxygenation (ECMO) treatment for pediatric patients with cardiorespiratory failure has increased, with emphasis being given to the prevention of complications in ECMO-treated patients. Several studies have reported ECMO-related central nervous system complications, such as intracranial hemorrhage, cerebral infarction, and seizure. However, few cases of peripheral nerve injury have been reported in ECMO-treated adults; there have also been no reported cases of peripheral nerve injury in the pediatric population. PATIENT CONCERNS: Two pediatric patients aged 16 and 6 experienced motor weakness in the extremities after the insertion of ECMO equipment. DIAGNOSES: They were diagnosed with peripheral nerve injuries through an electrodiagnostic study that showed femoral/sciatic neuropathies and brachial plexopathy. Arteriography and doppler sonography was performed to find the cause of peripheral nerve injury, and this may be the results of vascular compromise and compressive injuries, respectively. INTERVENTIONS: Surgical embolectomy was performed to remove thrombus in one patient. Two patients received orthosis, and physical therapy and occupational therapy were performed to prevent contracture and improve strength and functional use. OUTCOMES: Two pediatric patients showed a gradual improvement in motor power and function. LESSONS: Through this case report, we present rare ECMO-related complications and emphasize the importance of early diagnosis and monitoring of peripheral nerve injury in ECMO-treated children.
Assuntos
Oxigenação por Membrana Extracorpórea , Insuficiência Cardíaca , Traumatismos dos Nervos Periféricos , Insuficiência Respiratória , Trombose , Adulto , Criança , Oxigenação por Membrana Extracorpórea/efeitos adversos , Humanos , Estudos RetrospectivosRESUMO
Mulberry leaves have antioxidant activity and antiinflammatory effects in several types of cells. However, the efficacy of mulberry leaves fermented with Cordyceps militaris remains unknown. Therefore, the present study aimed to investigate whether the ethanol extracts of mulberry leaves fermented with C. militaris (EMfC) can prevent lipopolysaccharide (LPS)induced inflammation and autophagy in macrophages. To achieve this, RAW264.7 cells pretreated with three different dose of EMfCs were subsequently stimulated with LPS, and examined for alterations in the regulatory factors of inflammatory responses and key parameters of the autophagy signaling pathway. EMfC treatment inhibited the generation of reactive oxidative species; however, significant activity was observed for 2,2diphenyl1picrylhydrazyl (DPPH) radical scavenging (IC50=579.6703 mg/ml). Most regulatory factors in inflammatory responses were significantly inhibited following treatment with EMfC, without any significant cellular toxicity. EMfCtreated groups exhibited marked suppression of nitrogen oxide (NO) levels, mRNA expression levels of iNOS/COX2, levels of all inflammatory cytokines (TNFα, IL1ß and IL6) and phosphorylation of MAPK members, as well as recovery of cell cycle progression. Furthermore, similar effects were observed in the LPSinduced autophagy signaling pathway of RAW264.7 cells. The expression levels of microtubuleassociated protein 1A/1Blight chain 3 (LC3) and Beclin exhibited a dosedependent decrease in the EMfC+LPStreated groups compared with in the Vehicle+LPStreated group, whereas the phosphorylation of PI3K and mTOR were enhanced in a dosedependent manner in the same groups. Overall, the results of the present study provide evidence that exposure to EMfC protects against LPSinduced inflammation and autophagy in RAW264.7 cells. These results indicated that EMfC is a potential candidate for treatment of inflammatory diseases.
Assuntos
Inflamação/tratamento farmacológico , Morus/metabolismo , Extratos Vegetais/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Autofagia/efeitos dos fármacos , Cordyceps/metabolismo , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Fermentação/fisiologia , Lipopolissacarídeos/efeitos adversos , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Óxido Nítrico/metabolismo , Folhas de Planta/metabolismo , Células RAW 264.7 , Espécies Reativas de OxigênioRESUMO
Several techniques have been employed for deletion of the NKX3.1 gene, resulting in developmental defects of the prostate, including alterations in ductal branching morphogenesis and prostatic secretions as well as epithelial hyperplasia and dysplasia. To investigate whether the CRISPR/Cas9-mediated technique can be applied to study prostate carcinogenesis through exon I deletion of NKX3.1 gene, alterations in the prostatic intraepithelial neoplasia (PIN) and their regulatory mechanism were observed in the prostate of NKX3.1 knockout (KO) mice produced by the CRISPR/Cas9-mediated NKX3.1 mutant gene, at the ages of 16 and 24 weeks. The weight of dorsal-lateral prostate (DLP) and anterior prostate (AP) were observed to be increased in only the 24 weeks KO mice, although morphogenesis was constant in all groups. Obvious PIN 1 and 2 lesions were frequently detected in prostate of the 24 weeks KO mice, as compared with the same age wild type (WT) mice. Ki67, a key indicator for PIN, was densely stained in the epithelium of prostate in the 24 weeks KO mice, while the expression of p53 protein was suppressed in the same group. Also, both the 16 and 24 weeks KO mice reveal inhibition of the PI3K/AKT/mTOR pathway in the prostate. However, prostate specific antigen (PSA) levels and Bax/Bcl-2 expressions were decreased in the prostate of 16 weeks KO mice, and were increased in only the 24 weeks KO mice. Taken together, the results of the present study provide additional evidence that CRISPR/Cas9-mediated exon 1 deletion of the NKX3.1 gene successfully induces PIN lesions, along with significant alterations of Ki67 expression, EGFR signaling pathway, and cancer-regulated proteins.
Assuntos
Proteínas de Homeodomínio/genética , Morfogênese/genética , Neoplasia Prostática Intraepitelial/genética , Fatores de Transcrição/genética , Proteína Supressora de Tumor p53/genética , Animais , Sistemas CRISPR-Cas/genética , Receptores ErbB/genética , Humanos , Antígeno Ki-67/genética , Masculino , Camundongos , Camundongos Knockout , Próstata/crescimento & desenvolvimento , Próstata/patologia , Neoplasia Prostática Intraepitelial/patologiaRESUMO
BACKGROUND: A novel extract of mulberry leaves fermented with Cordyceps militaris (EMfC) is reported to exert anti-obesity activity, although their molecular mechanism during hepatic steatosis has not verified. METHODS: To investigate the role of inflammation and autophagy during the anti-hepatic steatosis effects of EMfC, we measured alterations in the key parameters for inflammatory response and autophagy pathway in liver tissues of the high fat diet (HFD) treated C57BL/6N mice after exposure to EMfC for 12 weeks. RESULTS: Significant anti-hepatic steatosis effects, including decreased number of lipid droplets and expression of Klf2 mRNA, were detected in the liver of the HFD + EMfC treated group. The levels of mast cell infiltration, expression of two inflammatory mediators (iNOS and COX-2), and the MAPK signaling pathway were remarkably decreased in the liver of HFD + EMfC treated group as compared to the HFD + Vehicle treated group. Furthermore, a similar inhibitory effect was measured for the expression levels of pro-inflammatory cytokines, including IL-1ß, IL-6, TNF-α and NF-κB. The expression level of members in the AKT/mTOR signaling pathway (a central regulator in autophagy) was recovered after treatment with EMfC, and autophagy-related proteins (Beclin and LC3-II) were remarkably decreased in the HFD + EMfC treated group compared to the HFD + Vehicle treated group. Moreover, the HFD + EMfC treated group showed decreased transcript levels of autophagy-regulated genes including Atg4b, Atg5, Atg7 and Atg12. CONCLUSIONS: Taken together, findings of the present study provide novel evidences that the anti-hepatic steatosis of EMfC is tightly linked to the regulation of the inflammatory response and autophagy pathway in the liver tissue of HFD-induced obesity mice.
Assuntos
Autofagia/efeitos dos fármacos , Inflamação/tratamento farmacológico , Morus , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Extratos Vegetais/farmacologia , Animais , Cordyceps , Dieta Hiperlipídica , Modelos Animais de Doenças , Fermentação , Camundongos , Camundongos Endogâmicos C57BL , Folhas de Planta , República da CoreiaRESUMO
The mechanisms through which cancerupregulated gene 2 (CUG2), a novel oncogene, affects Wnt/ßcatenin signaling, essential for tumorigenesis, are unclear. In this study, we aimed to elucidate some of these mechanisms in A549 lung cancer cells. Under the overexpression of CUG2, the protein levels and activity of ßcatenin were evaluated by western blot analysis and luciferase assay. To examine a biological consequence of ßcatenin under CUG2 overexpression, cell migration, invasion and sphere formation assay were performed. The upregulation of ßcatenin induced by CUG2 overexpression was also accessed by xenotransplantation in mice. We first found that CUG2 overexpression increased ßcatenin expression and activity. The suppression of ßcatenin decreased cancer stem cell (CSC)like phenotypes, indicating that ßcatenin is involved in CUG2mediated CSClike phenotypes. Notably, CUG2 overexpression increased the phosphorylation of ßcatenin at Ser33/Ser37, which is known to recruit E3 ligase for ßcatenin degradation. Moreover, CUG2 interacted with and enhanced the expression and kinase activity of never in mitosis gene Arelated kinase 2 (NEK2). Recombinant NEK2 phosphorylated ßcatenin at Ser33/Ser37, while NEK2 knockdown decreased the phosphorylation of ßcatenin, suggesting that NEK2 is involved in the phosphorylation of ßcatenin at Ser33/Ser37. Treatment with CGK062, a small chemical molecule, which promotes the phosphorylation of ßcatenin at Ser33/Ser37 through protein kinase C (PKC)α to induce its degradation, reduced ßcatenin levels and inhibited the CUG2induced features of malignant tumors, including increased cell migration, invasion and sphere formation. Furthermore, CGK062 treatment suppressed CUG2mediated tumor formation in nude mice. Taken together, the findings of this study suggest that CUG2 enhances the phosphorylation of ßcatenin at Ser33/Ser37 by activating NEK2, thus stabilizing ßcatenin. CGK062 may thus have potential for use as a therapeutic drug against CUG2overexpressing lung cancer cells.
Assuntos
Carcinogênese/efeitos dos fármacos , Proteínas Cromossômicas não Histona/metabolismo , Quinases Relacionadas a NIMA/metabolismo , Neoplasias/tratamento farmacológico , beta Catenina/metabolismo , Células A549 , Acrilatos/farmacologia , Acrilatos/uso terapêutico , Animais , Carcinogênese/patologia , Cromanos/farmacologia , Cromanos/uso terapêutico , Feminino , Humanos , Masculino , Camundongos , Camundongos Nus , Quinases Relacionadas a NIMA/genética , Neoplasias/patologia , Fosforilação/efeitos dos fármacos , Estabilidade Proteica/efeitos dos fármacos , RNA Interferente Pequeno/metabolismo , Via de Sinalização Wnt/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto , beta Catenina/genéticaRESUMO
BACKGROUND: Although constipation has been researched in various neurological disorders, including Parkinson's disease (PD) and spinal cord injury (SCI), the pathological mechanism of this symptom has not been investigated in Alzheimer's disease (AD) associated with loss of nerve cells in the brain. This study was undertaken to gain scientific evidences for a molecular correlation between constipation and AD. METHODS: To understand the etiology, we measured alterations in various constipation parameters, muscarinic acetylcholine receptors (mAChRs) and endoplasmic reticulum (ER) stress response, in 11-month-old Tg2576 transgenic (Tg) mice showing AD-like phenotypes. RESULTS: A high accumulation of amyloid beta (Aß) peptides, a key marker of AD pathology, were detected in the cortex and hippocampus of Tg mice. Furthermore, significant alterations were observed in various constipation parameters including stool weight, histological structure, cytological structure and mucin secretion in Tg2576 mice. Moreover, M2 and M3 expression and the downstream signaling pathways of mAChRs were decreased in the Tg group, as compared with non-Tg (NT) group. Furthermore, activation of ER stress proteins and alteration of ER structure were also detected in the same group. CONCLUSIONS: The results of the present study provide strong novel evidence that the neuropathological constipation detected in Tg2576 mice is linked to dysregulation of the mAChR signaling pathways and ER stress response.
Assuntos
Doença de Alzheimer/complicações , Doença de Alzheimer/metabolismo , Constipação Intestinal/complicações , Constipação Intestinal/metabolismo , Estresse do Retículo Endoplasmático , Receptores Muscarínicos/metabolismo , Doença de Alzheimer/genética , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Colo/patologia , Constipação Intestinal/genética , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Mucinas/metabolismo , Fragmentos de Peptídeos/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Muscarínicos/genética , Transdução de SinaisRESUMO
The therapeutic effects of mulberry (Morus alba) leaves on lipid metabolism, including lipogenesis, lipolysis and hyperlipidemia are widely known, although their fermented products are yet to be applied. To investigate the therapeutic effects of a novel extract of mulberry leaves fermented with Cordyceps militaris (EMfC) on lipid metabolism, the lipid profile of serum, lipid accumulation, lipolytic activity and lipogenesis regulation were measured in high fat diet (HFD)-induced obese C57BL/6 mice treated with EMfC for 12 weeks. Briefly, the concentrations of low-density lipoprotein, triglyceride, total cholesterol and glucose significantly decreased in the serum of the HFD+EMfC treated group when compared with the HFD+Vehicle treated group, while the levels of high-density lipoprotein increased in the HFD+EMfC group. The amount of abdominal fat and the size of adipocytes were significantly lower in the HFD+EMfC treated group when compared with the HFD+Vehicle treated group. The weight and number of lipid droplets of liver tissue exhibited a similar decrease. Furthermore, the mRNA levels of peroxisome proliferator-activated receptor-γ for adipogenesis as well as adipocyte protein 2 and Fas cell surface death receptor for lipogenesis reduced following EMfC treatment for 12 weeks. Phosphorylation of perilipin and hormone-sensitive lipase, and in the adipose triglyceride lipase expression showed a significant increase in the HFD+EMfC treated group. These results indicated that EMfC may prevent fat accumulation in the HFD-induced obese C57BL/6 mice through the inhibition of lipogenesis and by stimulating lipolysis. Thus, the results provide evidence for the potential use of EMfC as an anti-obesity complex in the treatment of obesity.
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Cellulose in different forms has extensively been applied in biomedical treatments, including scaffolding, tissue engineering and tissue formation. To evaluate the therapeutic effects of a liquid bandage (LB) prepared with cellulose powders from Styela clava tunics (SCT) and Broussonetia kazinoki bark (BSLB) for healing cutaneous wounds, the remedial effects of a low concentration (LoBSLB) and a high concentration (HiBSLB) of BSLB on skin regeneration and toxicity in Sprague Dawley rats. Results indicated that the total area of skin involved in the surgical wound was lower in the BSLBtreated group compared with the Vehicletreated group at days 412, although some variations were observed in the HiBSLBtreated group. In addition, the BSLBtreated group showed significantly enhanced width of the reepithelialization region and epidermal thickness when compared with the Vehicletreated group. Furthermore, significant stimulation in the expression level of collagen1 and the signaling pathway of VEGF after topical application of BSLB was indicated. No liver or kidney toxicities were detected for either doses of BSLB. Overall, the results of the present study suggest that BSLB accelerates the process of wound healing in surgical skin wounds of Sprague Dawley rats through stimulation of reepithelialization and connective tissue formation, without any accompanying significant toxicity.
Assuntos
Broussonetia/química , Celulose/farmacologia , Pós/farmacologia , Pele/efeitos dos fármacos , Ferida Cirúrgica/tratamento farmacológico , Urocordados/química , Cicatrização/efeitos dos fármacos , Animais , Bandagens , Colágeno Tipo I/metabolismo , Epiderme/efeitos dos fármacos , Epiderme/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley , Pele/metabolismo , Ferida Cirúrgica/metabolismo , Engenharia Tecidual/métodosRESUMO
Dietary garlic has been suggested to possess anticancer properties, and several attempts have been made to isolate the anticancer compounds. In this study, we efficiently synthesized N-benzyl-N-methyl-dodecan-1-amine (BMDA) by the reductive amination method. We evaluated the potential anticancer activities of BMDA against A549 lung cancer cells with cancer stem cell-like phenotypes due to the overexpression of cancer upregulated gene (CUG)2. N-Benzyl-N-methyl-dodecan-1-amine treatment sensitized A549 cells overexpressing CUG2 (A549-CUG2) to apoptosis and autophagy compared with those of the control cells. The treatment with BMDA also reduced tumor development in xenografted nude mice. Furthermore, BMDA inhibited cell migration, invasion, and sphere formation in A549-CUG2 cells, in which TGF-ß signaling is involved. Further analysis showed that BMDA hindered TGF-ß promoter activity, protein synthesis, and phosphorylation of Smad2, thus decreasing the expression of TGF-ß-targeted proteins, including Snail and Twist. N-Benzyl-N-methyl-dodecan-1-amine also decreased Twist expression in vivo. In addition, BMDA inhibited Akt-ERK activities, ß-catenin expression, and its transcriptional activity. These results suggest that BMDA can be a promising anticancer agent against cancer cells overexpressing CUG2.
Assuntos
Aminas/farmacologia , Antineoplásicos/farmacologia , Proteínas Cromossômicas não Histona/genética , Alho/química , Regulação para Cima/efeitos dos fármacos , Células A549 , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Camundongos Endogâmicos BALB C , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , beta Catenina/metabolismoRESUMO
Introduction: Roots of Asparagus cochinchinensis, which have pharmacologically active ingredients, have received great attention because they show good therapeutic effects for various inflammatory diseases without specific toxicity. This study investigated the anti-asthmatic effects of a butanol extract of Asparagus cochinchinensis roots that had been fermented with Weissella cibaria (BAW) and its possible underlying cholinergic regulation. Methods: Alterations of the anti-asthmatic markers and the molecular response factors were measured in an ovalbumin (OVA)-induced asthma model after treatment with BAW. Results: Treatment with BAW decreased the intracellular reactive oxygen species (ROS) production in lipopolysaccharides (LPS) activated RAW264.7 cells. The results of the animal experiments revealed lower infiltration of inflammatory cells and bronchial thickness, and a significant reduction in the number of macrophages and eosinophils, concentration of OVA-specific IgE, and expression of Th2 cytokines in the OVA + BAW treated group. In addition, a significant recovery of goblet cell hyperplasia, MMP-9 expression, and the VEGF signaling pathway was observed upon airway remodeling in the OVA + BAW treated group. Furthermore, these responses of BAW were linked to recovery of acetylcholine esterase (AChE) activity and muscarinic acetylcholine receptor (mAChR) M3 downstream signaling pathway in epithelial cells, smooth muscle cells, and afferent sensory nerves of OVA + BAW-treated mice. Conclusion: Overall, these findings are the first to provide evidence that the therapeutic effects of BAW can prevent airway inflammation and remodeling through the recovery of cholinergic regulation in structural cells and inflammatory cells of the chronic asthma model.
RESUMO
The butanol extract of Asparagus cochinchinensis roots fermented with Weissella cibaria (BAfW) significantly suppressed the inflammatory response induced by lipopolysaccharide (LPS) treatment in RAW264.7 cells. To investigate the dose dependence and durability of BAfW on the anti-asthma effects, alterations in key parameters were measured in ovalbumin (OVA)-challenged Balb/c mice treated with the different doses of BAfW at three different time points. The number of immune cells, OVA-specific IgE level, thickness of respiratory epithelium and mucus score decreased significantly in a dose-dependent manner in response to treatment with 125 to 500 mg/kg BAfW (P<0.05), although the highest level was detected in the 500 mg/kg treated group. Moreover, the decrease in these parameters was maintained from 24 to 48 h in the 500 mg/kg of BAfW treated group. At 72 h, the effects of BAfW on the number of immune cells, OVA-specific IgE level and thickness of respiratory epithelium partially disappeared. Overall, this study provides the first evidence that the anti-asthma effect of BAfW may reach the maximum level in OVA-challenged Balb/c mice treated with 500 mg/kg and that these effects can last for 48 h.
RESUMO
To evaluate the carcinogenicity of p27 knockout (KO) mice with RNA-guided endonuclease (RGENs)-mediated p27 mutant exon I gene (IΔ), alterations in the carcinogenic phenotypes including tumor spectrum, tumor suppressor proteins, apoptotic proteins and cell cycle regulators were observed in p27 (IΔ) KO mice after treatment with 7,12-Dimethylbenz[a]anthracene (DMBA) and 12-O-tetradecanoylphorbol-13-acetate (TPA)(DT) for 5 months. The target region (544~571 nt) in exon I of the p27 gene was successfully disrupted in p27 (IΔ) KO mice using the RGEN-induced non-homologous end joining (NHEJ) technique. After DT exposure for 5 months, a few solid tumors (identified as squamous cell carcinoma) developed on the surface of back skin of DT-treated p27 (IΔ) KO mice. Also, squamous cell hyperplasia with chronic inflammation was detected in the skin dermis of DT-treated p27 (IΔ) KO mice, while the Vehicle+p27 (IΔ) KO mice and WT mice maintained their normal histological skin structure. A significant increase was observed in the expression levels of tumor suppressor protein (p53), apoptotic proteins (Bax, Bcl-2 and Caspase-3) and cell-cycle regulator proteins (Cyclin D1, CDK2 and CDK4) in the skin of DT-treated p27 (IΔ) KO mice, although their enhancement ratio was varied. Taken together, the results of the present study suggest that squamous cell carcinoma and hyperplasia of skin tissue can be successfully developed in new p27 (IΔ) KO mice produced by RGEN-induced NHEJ technique following DT exposure for 5 months.
RESUMO
Conflicting results for morusin activity during adipogenic differentiation are reported in 3T3-L1 adipocytes and cancer cells. To elucidate the influence of morusin on fat metabolism, their anti-obesity effects and molecular mechanism were investigated in 3T3-L1 cells and primary adipocytes. Morusin at a dose of less than 20 µM does not induce any significant change in the viability of 3T3-L1 adipocytes. The accumulation of intracellular lipid droplets in 3T3-L1 adipocytes stimulated with 0.5 mM 3-isobutyl-1-methylxanthine, 1 µM dexamethasone, 10 µg/mL insulin in DMEM containing 10% FBS (MDI)-significantly reduces in a dose-dependent manner after morusin treatment. The phosphorylation level of members in the MAP kinase signaling pathway under the insulin receptor downstream also decrease significantly in the MDI + morusin-treated group compared to MDI + vehicle-treated group. Also, the expression of adipogenic transcription factors (PPARγ and C/EBPα) and lipogenic proteins (aP2 and FAS) are significantly attenuated by exposure to the compound in MDI-stimulated 3T3-L1 adipocytes. Furthermore, the decrease in the G0/G1 arrest of cell cycle after culturing in MDI medium was dramatically recovered after co-culturing in MDI + 20 µM morusin. Moreover, morusin treatment induces glycerol release in the primary adipocytes of SD rats and enhances lipolytic protein expression (HSL, ATGL, and perilipin) in differentiated 3T3-L1 adipocytes. Overall, the results of the present study provide strong evidence that morusin inhibits adipogenesis by regulating the insulin receptor signaling, cell cycle and adipogenic protein expression as well as stimulating lipolysis by enhancing glycerol release and lipolytic proteins expression.