The lncRNA-MALAT1/miR-330-3p Axis Promotes Growth and Metastasis of Gastric Cancer Through Regulation of Vascular Endothelial Growth Factor A: A Genetic and Cell Evaluation Study

SUMMARY: Overexpression of metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in various tumor tissues and cell lines was found to promote tumor cell proliferation, migration, and invasion. However, the role of MALAT1 in gastric cancer (GC) is still unclear. We aimed to investigate the correlation between long-chain non-coding RNAs (lncRNAs), MALAT1, MicroRNAs (miRNA) and vascular endothelial growth factor A (VEGFA) in gastric cancer and to disclose underlying mechanism. The correlation between MALAT1 levels and clinical features was analyzed by bioinformatics data and human samples. The expression of MALAT1 was down regulated in AGS cells to detect the cell proliferation, migration, and invasion characteristics, as well as the effects on signal pathways. Furthermore, we validated the role of MALAT1/miR-330-3p axis in GC by dual luciferase reporter gene assays. Expression of MALAT1 was higher in cancer tissues than in para-cancerous tissues. The high MALAT1 level predicted malignancy and worse prognosis. Down-regulation of MALAT1 expression in AGS cells inhibited cell proliferation, migration, and invasion by targeting VEGFA. By dual luciferase reporter gene assay and miR-330-3p inhibitor treatment, we demonstrate that MALAT1 sponged miR-330-3p in GC, leading to VEGFA upregulation and activation of the mTOR signaling pathway. The MALAT1/miR-330-3p axis regulates VEGFA through the mTOR signaling pathway and promotes the growth and metastasis of gastric cancer.

Se descubrió que la sobreexpresión del transcrito 1 de adenocarcinoma de pulmón asociado a metástasis (MALAT1) en varios tejidos tumorales y líneas celulares promueve la proliferación, migración e invasión de células tumorales. Sin embargo, el papel de MALAT1 en el cáncer gástrico (CG) aún no está claro. Nuestro objetivo fue investigar la correlación entre los ARN no codificantes de cadena larga (lncRNA), MALAT1, los microARN (miARN) y el factor de crecimiento endotelial vascular A (VEGFA) en el cáncer gástrico y revelar el mecanismo subyacente. La correlación entre los niveles de MALAT1 y las características clínicas se analizó mediante datos bioinformáticos y muestras humanas. La expresión de MALAT1 se reguló negativamente en las células AGS para detectar las características de proliferación, migración e invasión celular, así como los efectos sobre las vías de señales. Además, validamos el papel del eje MALAT1/miR- 330-3p en GC mediante ensayos de genes indicadores de luciferasa dual. La expresión de MALAT1 fue mayor en tejidos cancerosos que en tejidos paracancerosos. El alto nivel de MALAT1 predijo malignidad y peor pronóstico. La regulación negativa de la expresión de MALAT1 en células AGS inhibió la proliferación, migración e invasión celular al apuntar a VEGFA. Mediante un ensayo de gen indicador de luciferasa dual y un tratamiento con inhibidor de miR-330-3p, demostramos que MALAT1 esponjaba miR-330-3p en GC, lo que lleva a la regulación positiva de VEGFA y la activación de la vía de señalización mTOR. El eje MALAT1/miR-330-3p regula VEGFA a través de la vía de señalización mTOR y promueve el crecimiento y la metástasis del cáncer gástrico.

Acteoside Attenuates Recognition Memory Impairment in CUMS Rats via Regulating Synaptic Plasticity and mTOR Signaling Pathway / El Acteoside Atenúa el Deterioro de la Memoria de Reconocimiento en Ratas ELCI Mediante la Regulación de la Plasticidad Sináptica y la Vía de Señalización mTOR

SUMMARY: We evaluated the role and mechanism of acteoside in the regulation of memory impairment induced by chronic unpredictable mild stress (CUMS). CUMS was used to induce depression in rats and the successful establishment of CUMS model were verified by forced swimming test and sucrose preference test. The Y-maze test and novel object recognition test assessed memory functions. The structural changes in the cortex and hippocampus were observed by hematoxylin and eosin (HE) staining. Immunofluorescence staining and western blotting determined the protein levels. Y-maze test and novel object recognition test showed that there was memory performance impairment in rats of CUMS group, which was improved by the acteoside treatment. HE staining showed that CUMS exposure damaged the structure in the cortex and hippocampus, while the acteoside treatment alleviated the structural changes. Compared with the control group, the levels of BNDF and CREB in the cortex and hippocampus of the CUMS group were significantly decreased. Acteoside significantly reversed the expressions of these proteins in CUMS rats. Meanwhile, compared with the control group, the levels of p-mTOR and p- P70S6K in the cortex and hippocampus of the CUMS group were significantly increased, and these changes were significantly reversed by acteoside. Nevertheless, the effect of acteoside on mTOR signaling was markedly blocked by rapamycin, a specific inhibitor of mTOR signaling. Acteoside can attenuate memory impairment and ameliorate neuronal damage and synaptic plasticity in depression rats probably via inhibiting the mTOR signaling pathway. Acteoside may serve as a novel reagent for the prevention of depression.

Evaluamos el papel y el mecanismo del acteoside en la regulación del deterioro de la memoria inducido por estrés leve crónico impredecible (ELCI). Se utilizó ELCI para inducir depresión en ratas y el establecimiento exitoso del modelo ELCI se verificó mediante una prueba de natación forzada y una prueba de preferencia de sacarosa. La prueba del laberinto en Y y la prueba de reconocimiento de objetos novedosos evaluaron las funciones de la memoria. Los cambios estructurales en la corteza y el hipocampo se observaron mediante tinción con hematoxilina y eosina (HE). La tinción por inmunofluorescencia y la transferencia Western determinaron los niveles de proteína. La prueba del laberinto en Y y la prueba de reconocimiento de objetos novedosos mostraron que había un deterioro del rendimiento de la memoria en ratas del grupo ELCI, que mejoró con el tratamiento con acteósidos. La tinción con HE mostró que la exposición a ELCI dañó la estructura de la corteza y el hipocampo, mientras que el tratamiento con actósidos alivió los cambios estructurales. En comparación con el grupo de control, los niveles de BNDF y CREB en la corteza y el hipocampo del grupo ELCI disminuyeron significativamente. Acteoside revirtió significativamente las expresiones de estas proteínas en ratas ELCI. Mientras tanto, en comparación con el grupo control, los niveles de p-mTOR y p-P70S6K en la corteza y el hipocampo del grupo ELCI aumentaron significativamente, y estos cambios fueron revertidos significativamente ELCI por el acteoside. Sin embargo, el efecto del acteoside sobre la señalización de mTOR fue notablemente bloqueado por la rapamicina, un inhibidor específico de la señalización de mTOR. El acteoside puede atenuar el deterioro de la memoria y mejorar el daño neuronal y la plasticidad sináptica en ratas con depresión, probablemente mediante la inhibición de la vía de señalización mTOR. Acteoside puede servir como un reactivo novedoso para la prevención de la depresión.

Inhibition of Multifunctional Protein p32/C1QBP Promotes Cytostatic Effects in Colon Cancer Cells by Altering Mitogenic Signaling Pathways and Promoting Mitochondrial Damage.

The protein p32 (C1QBP) is a multifunctional and multicompartmental homotrimer that is overexpressed in many cancer types, including colon cancer. High expression levels of C1QBP are negatively correlated with the survival of patients. Previously, we demonstrated that C1QBP is an essential promoter of migration, chemoresistance, clonogenic, and tumorigenic capacity in colon cancer cells. However, the mechanisms underlying these functions and the effects of specific C1QBP protein inhibitors remain unexplored. Here, we show that the specific pharmacological inhibition of C1QBP with the small molecule M36 significantly decreased the viability rate, clonogenic capacity, and proliferation rate of different colon cancer cell lines in a dose-dependent manner. The effects of the inhibitor of C1QBP were cytostatic and non-cytotoxic, inducing a decreased activation rate of critical pro-malignant and mitogenic cellular pathways such as Akt-mTOR and MAPK in RKO colon cancer cells. Additionally, treatment with M36 significantly affected the mitochondrial integrity and dynamics of malignant cells, indicating that p32/C1QBP plays an essential role in maintaining mitochondrial homeostasis. Altogether, our results reinforce that C1QBP is an important oncogene target and that M36 may be a promising therapeutic drug for the treatment of colon cancer.

Silencing ANGPT2 alleviates ulcerative colitis by regulating autophagy-mediated NLRP3 inflammasome inactivation via the mTOR signaling pathway

Ulcerative colitis (UC) is a difficult intestinal disease characterized by inflammation, and its mechanism is complex and diverse. Angiopoietin-like protein 2 (ANGPT2) plays an important regulatory role in inflammatory diseases. However, the role of ANGPT2 in UC has not been reported so far. After exploring the expression level of ANGPT2 in serum of UC patients, the reaction mechanism of ANGPT2 was investigated in dextran sodium sulfate (DSS)-induced UC mice. After ANGPT2 expression was suppressed, the clinical symptoms and pathological changes of UC mice were detected. Colonic infiltration, oxidative stress, and colonic mucosal barrier in UC mice were evaluated utilizing immunohistochemistry, immunofluorescence, and related kits. Finally, western blot was applied for the estimation of mTOR signaling pathway and NLRP3 inflammasome-related proteins. ANGPT2 silencing improved clinical symptoms and pathological changes, alleviated colonic inflammatory infiltration and oxidative stress, and maintained the colonic mucosal barrier in DSS-induced UC mice. The regulatory effect of ANGPT2 on UC disease might occur by regulating the mTOR signaling pathway and thus affecting autophagy-mediated NLRP3 inflammasome inactivation. ANGPT2 silencing alleviated UC by regulating autophagy-mediated NLRP3 inflammasome inactivation via the mTOR signaling pathway.

Long Non-Coding RNAs (lncRNAs) as Regulators of the PI3K/AKT/mTOR Pathway in Gastric Carcinoma.

Gastric cancer (GC) represents ~10% of the global cancer-related deaths, increasingly affecting the younger population in active stages of life. The high mortality of GC is due to late diagnosis, the presence of metastasis and drug resistance development. Additionally, current clinical markers do not guide the patient management adequately, thereby new and more reliable biomarkers and therapeutic targets are still needed for this disease. RNA-seq technology has allowed the discovery of new types of RNA transcripts including long non-coding RNAs (lncRNAs), which are able to regulate the gene/protein expression of many signaling pathways (e.g., the PI3K/AKT/mTOR pathway) in cancer cells by diverse molecular mechanisms. In addition, these lncRNAs might also be proposed as promising diagnostic or prognostic biomarkers or as potential therapeutic targets in GC. This review describes important topics about some lncRNAs that have been described as regulators of the PI3K/AKT/mTOR signaling pathway, and hence, their potential oncogenic role in the development of this malignancy.

The prognostic and predictive values of differential expression of exosomal receptor tyrosine kinases and associated with the PI3K/AKT/mTOR signaling in breast cancer patients undergoing neoadjuvant chemotherapy.

PURPOSE: Cancer cell-derived exosomes are the mediator of the tumor microenvironment and the molecular content of exosomes presents a promising prognostic or predictive marker in tumor progression and the treatment response of cancer patients. The aim of this study was to identify the expression levels of receptor tyrosine kinases (RTKs) and AKT1 and mTOR before and after neoadjuvant chemotherapy (NACT) in the exosomes of BC patients compared with healthy females. METHODS: After isolating exosomes in the serum of 25 BC patients and characterization by flow cytometry, the mRNA levels of FGFR2, FGFR3, PDGFRB, AKT1 and mTOR in the exosomes were analyzed by RT-PCR. RESULTS: Our preliminary findings showed that FGFR2, PDGFRB, AKT1 and mTOR levels were significantly upregulated in BC patients before NACT compared with the healthy group (p < 0.05). Furthermore, the mRNA levels PDGFRB and AKT1 were significantly down-regulated after NACT compared with control. PDGFRB expression level could predict pathological non-response and significantly correlated with tumor size after NACT. CONCLUSION: Therefore, especially FGFR2, PDGFRB and AKT1 could be a therapeutic target as a prognostic marker, whereas PDGFRB may be a promising predictive indicator of therapy response in BC patients. However, the prognostic or predictive role of RTKs and PI3K/AKT/mTOR signaling in the exosomes should be further investigated in a large patient population.

Circulating miRNA-1-3p as Biomarker of Accelerated Sarcopenia in Patients Diagnosed with Chronic Heart Failure.

Background: While sarcopenia is an important clinical finding in individuals diagnosed with chronic heart failure (CHF), efforts to identify a reliable biomarker capable of predicting the overall muscular and functional decline in CHF patients have been unsuccessful to date. Objectives: The objectives of this study were to study the diagnostic utility of MicroRNA (miRNA)-1-3p as a predictor of sarcopenia status in individuals diagnosed with CHF. Methods: In total, 80 individuals with heart failure exhibiting a left ventricular ejection fraction < 50% were enrolled in this study. All patients were analyzed to assess miR-1-3p expression levels, with body composition being evaluated through dual-energy X-ray absorptiometry and sarcopenia being defined based on the sum of appendicular lean muscle mass (ALM) divided by height in meters squared and handgrip strength (HGS). In addition, the activation of the Akt/mTOR signaling pathway was evaluated in these individuals. Results: In total, 40 of the enrolled patients (50%) exhibited sarcopenia. Sarcopenic patients presented with increased miR-1-3p expression levels as compared to non-sarcopenic individuals (1.69 ± 0.132 vs. 1.22 ± 0.106; p < 0.05). With respect to sarcopenic indices, appendicular skeletal mass index was most strongly correlated with miR-1-3p expression, which was also strongly correlated with HGS. High levels of Akt/mTOR signaling pathway components were expressed in sarcopenic individuals, highlighting a significant relationship between miR-1-3p activity and signaling through this pathway. Moreover, miR-1-3p was identified as a specific marker for sarcopenia in individuals with CHF. Conclusions: These results suggest that circulating miR-1-3p levels are related to Akt/mTOR pathway activation and can offer valuable insight into the overall physical capacity and muscular integrity of CHF patients as a predictor of sarcopenia. (Rev Invest Clin. 2022;74(5):276-83).

Circulating miRNA-1-3p as Biomarker of Accelerated Sarcopenia in Patients Diagnosed with Chronic Heart Failure

ABSTRACT Background: While sarcopenia is an important clinical finding in individuals diagnosed with chronic heart failure (CHF), efforts to identify a reliable biomarker capable of predicting the overall muscular and functional decline in CHF patients have been unsuccessful to date. Objectives: The objectives of this study were to study the diagnostic utility of MicroRNA (miRNA)-1-3p as a predictor of sarcopenia status in individuals diagnosed with CHF. Methods: In total, 80 individuals with heart failure exhibiting a left ventricular ejection fraction < 50% were enrolled in this study. All patients were analyzed to assess miR-1-3p expression levels, with body composition being evaluated through dual-energy X-ray absorptiometry and sarcopenia being defined based on the sum of appendicular lean muscle mass (ALM) divided by height in meters squared and handgrip strength (HGS). In addition, the activation of the Akt/mTOR signaling pathway was evaluated in these individuals. Results: In total, 40 of the enrolled patients (50%) exhibited sarcopenia. Sarcopenic patients presented with increased miR-1-3p expression levels as compared to non-sarcopenic individuals (1.69 ± 0.132 vs. 1.22 ± 0.106; p < 0.05). With respect to sarcopenic indices, appendicular skeletal mass index was most strongly correlated with miR-1-3p expression, which was also strongly correlated with HGS. High levels of Akt/mTOR signaling pathway components were expressed in sarcopenic individuals, highlighting a significant relationship between miR-1-3p activity and signaling through this pathway. Moreover, miR-1-3p was identified as a specific marker for sarcopenia in individuals with CHF. Conclusion: These results suggest that circulating miR-1-3p levels are related to Akt/mTOR pathway activation and can offer valuable insight into the overall physical capacity and muscular integrity of CHF patients as a predictor of sarcopenia.

Cuban Policosanol Prevents the Apoptosis and the Mitochondrial Dysfunction Induced by Lipopolysaccharide in C2C12 Myoblast via Activation of Akt and Erk Pathways.

Skeletal muscle plays crucial roles in locomotion, protein reservoir, and maintenance of metabolic homeostasis. Loss of muscle, known as muscle atrophy, causes the metabolic diseases such as type 2 diabetes mellitus, hypertension, and so on. Therefore, great efforts have been devoted to prevent the muscle atrophy. Policosanols are a mixture of long chain fatty alcohols extracted from various natural sources. They have long been used as functional foods to lower the level of serum lipids, including triacylglycerol and cholesterol, and to protect against inflammatory stress. In this study, we examine the protective effect and molecular mechanism of Cuban policosanol on skeletal muscle cell death and mitochondrial dysfunction using lipopolysaccharide-treated C2C12 cells. Our results demonstrated that policosanol significantly rescued cell survival (40% vs. 88%; LPS vs. LPS+policosanol) via activation of the Akt pathway, resulting in inhibition of apoptosis (p<0.05). Moreover, policosanol restored the LPS-induced repression of collagen by two fold (0.33±0.04 vs. 0.67±0.03 compared to that of control; LPS vs. LPS+policosanol) via activation of ERK-mTOR-p70S6K pathways. In addition, policosanol increased the mitochondrial fusion by regulating the activities of DRP1 and Mfn2, leading to ameliorate the mitochondrial dysfunction induced by LPS. Improved mitochondria function increased the oxygen consumption rate with glucose as fuel source, indicating that policosanol could shift the glucose metabolism from lactate fermentation, induced by lipopolysaccharide, to oxidative phosphorylation. Thus, policosanol is a promising agent for preventing the inflammation-induced muscle cell death and mitochondrial dysfunction.

Emerging agents that target signaling pathways to eradicate colorectal cancer stem cells.

Colorectal cancer (CRC) represents the third most commonly diagnosed cancer and the second leading cause of cancer death worldwide. The modern concept of cancer biology indicates that cancer is formed of a small population of cells called cancer stem cells (CSCs), which present both pluripotency and self-renewal properties. These cells are considered responsible for the progression of the disease, recurrence and tumor resistance. Interestingly, some cell signaling pathways participate in CRC survival, proliferation, and self-renewal properties, and most of them are dysregulated in CSCs, including the Wingless (Wnt)/ß-catenin, Notch, Hedgehog, nuclear factor kappa B (NF-κB), Janus kinase/signal transducer and activator of transcription (JAK/STAT), peroxisome proliferator-activated receptor (PPAR), phosphatidyl-inositol-3-kinase/Akt/mechanistic target of rapamycin (PI3K/Akt/mTOR), and transforming growth factor-ß (TGF-ß)/Smad pathways. In this review, we summarize the strategies for eradicating CRC stem cells by modulating these dysregulated pathways, which will contribute to the study of potential therapeutic schemes, combining conventional drugs with CSC-targeting drugs, and allowing better cure rates in anti-CRC therapy.

Signaling Pathways Involved in Nutrient Sensing Control in Cancer Stem Cells: An Overview.

Cancer cells characteristically have a high proliferation rate. Because tumor growth depends on energy-consuming anabolic processes, including biosynthesis of protein, lipid, and nucleotides, many tumor-associated conditions, including intermittent oxygen deficiency due to insufficient vascularization, oxidative stress, and nutrient deprivation, results from fast growth. To cope with these environmental stressors, cancer cells, including cancer stem cells, must adapt their metabolism to maintain cellular homeostasis. It is well- known that cancer stem cells (CSC) reprogram their metabolism to adapt to live in hypoxic niches. They usually change from oxidative phosphorylation to increased aerobic glycolysis even in the presence of oxygen. However, as opposed to most differentiated cancer cells relying on glycolysis, CSCs can be highly glycolytic or oxidative phosphorylation-dependent, displaying high metabolic plasticity. Although the influence of the metabolic and nutrient-sensing pathways on the maintenance of stemness has been recognized, the molecular mechanisms that link these pathways to stemness are not well known. Here in this review, we describe the most relevant signaling pathways involved in nutrient sensing and cancer cell survival. Among them, Adenosine monophosphate (AMP)-activated protein kinase (AMPK) pathway, mTOR pathway, and Hexosamine Biosynthetic Pathway (HBP) are critical sensors of cellular energy and nutrient status in cancer cells and interact in complex and dynamic ways.

Poricoic acid A induces apoptosis and autophagy in ovarian cancer via modulating the mTOR/p70s6k signaling axis

Due to the high mortality and rapid disease progression, ovarian cancer remains one of the most common malignancies threatening the health of women. The present study was conducted to explore the anticancer effects and the underlying mechanisms of poricoic acid A (PAA), the main components of Poria cocos, on ovarian cancer. We investigated the anticancer effects of different concentrations of PAA in the SKOV3 cell line. Cell viability and proliferation were examined by CCK-8 assay. Cellular migration and invasion were assessed by the scratch and Transwell migration assays, respectively. The effect of PPA on cell apoptosis was measured by flow cytometry and caspase-3/8/9 colorimetric assay. Western blot was performed to detect protein level changes related to apoptosis and mTOR signaling pathways. The in vivo anticancer effect of PAA was evaluated using xenograft tumorigenesis model in nude mice. Our results showed that PAA suppressed SKOV3 cellular viability, migration, and invasion in a dosage-dependent manner. Flow cytometry results demonstrated PAA treatment could induce SKOV3 cell apoptosis. In addition, increased ratio of LC3-II/LC3-I (a marker for autophagosome formation) was observed after PAA treatment, as well as inhibition of m-TOR and p70s6k phosphorylation. In nude mice, PAA treatment reduced the xenograft tumor weight by 70% (P<0.05). In conclusion, our data suggested that PAA induced apoptosis and autophagy in ovarian cancer via modulating the mTOR/p70s6k signaling axis.

Protein blend and casein supplementations before inactive phase similarly activate mechanistic target of rapamycin signaling in rat skeletal muscle.

During overnight sleep, the longest postabsorptive and inactive phase of the day causes protein catabolism and loss. However, the daytime ingestion of dairy proteins has been shown to stimulate muscle protein synthesis and growth. This study compared the effects of pre-sleep supplementation of a protein blend (PB) composed of micellar casein (MCa) and whey protein (1:1) versus isolate MCa on the plasma levels of branched-chain amino acids (BCAAs) and the activation of the mechanistic target of rapamycin (mTOR) signaling, a critical intracellular pathway involved in the regulation of muscle protein synthesis. After 10 h of fasting during the active phase, rats were fed with a single dose of PB or MCa (5.6 g protein/kg of body mass) by gavage, and samples of blood and gastrocnemius muscle were collected at 30, 90, and 450 min. PB and MCa supplementations induced an increase (~3-fold, P < 0.001) of plasma BCAAs at 30 and 90 min. Most importantly, the stimulatory phosphorylation levels of mTOR and its downstream target p70 ribosomal protein S6 kinase (p70S6K) were similarly higher (~2.5-fold, P < 0.001) 30 and 90 min after MCa and PB. Plasma levels of leucine, isoleucine, valine, and overall BCAAs were correlated with the activation of mTOR (P < 0.001) and p70S6K (P < 0.001). MCa and PB supplementations before the inactive phase of rats resulted in an anabolic milieu in the skeletal muscle by inducing a transient increase in plasma BCAAs and a similar activation of the mTOR/p70S6K axis.

miR-660-5p promotes breast cancer progression through down-regulating TET2 and activating PI3K/AKT/mTOR signaling

Breast cancer (BC) is a commonly diagnosed cancer in females. MicroRNA-660-5p (miR-660-5p) has been reported to be involved in the occurrence and development of BC. However, the regulatory network of miR-660-5p in BC has not been fully addressed. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to detect the enrichment of miR-660-5p and tet-eleven translocation 2 (TET2) in BC tissues and cells. Cell counting kit-8 (CCK8), flow cytometry, and transwell migration and invasion assays were used to measure cell proliferation, apoptosis, migration, and invasion. The target relationship between miR-660-5p and TET2 was confirmed by dual luciferase reporter assay. Protein expression was measured by western blot. The expression of miR-660-5p was elevated in BC, and high expression of miR-660-5p was closely related to lymph node metastasis, advanced TNM stage, and vascular invasion of BC tumors. miR-660-5p silencing inhibited cell proliferation and metastasis, but induced apoptosis of BC cells. TET2 was identified as a direct target of miR-660-5p, and the interference of TET2 partly reversed the suppressive effects of miR-660-5p silencing on the malignant potential of BC cells. miR-660-5p promoted BC progression partly through modulating TET2 and PI3K/AKT/mTOR signaling. miR-660-5p/TET2 axis might be a promising target for BC treatment.

Vitexin induces apoptosis through mitochondrial pathway and PI3K/Akt/mTOR signaling in human non-small cell lung cancer A549 cells.

BACKGROUND: Currently, the prognosis of patients with non-small cell lung cancer (NSCLC) remains dismal; hence, it is critical to identify effective anti-NSCLC agents with limited side effects. This study aimed to evaluate the therapeutic potential of flavonoid compound vitexin in human NSCLC cells and the underlying mechanisms. RESULTS: The experimental results indicated that vitexin reduced the viability of A549 cells in a dose-dependent manner with nearly no toxicity against normal human bronchial epithelial 16HBE cells. Vitexin also dose-dependently increased A549 cell apoptosis, accompanied by the decreased Bcl-2/Bax ratio and the increased expression of cleaved caspase-3. Moreover, the in vivo anticancer activity of vitexin was further determined in nude mice bearing A549 cells. In addition, vitexin induced the release of cytochrome c from the mitochondria to the cytosol and the loss of mitochondrial membrane potential. Vitexin also significantly reduced the levels of p-PI3K, p-Akt and p-mTOR, and the pro-apoptotic effect of vitexin on A549 cells was partly blocked by SC79, an Akt activator. CONCLUSIONS: Accordingly, we believed that vitexin could be used as a potential therapeutic agent for the treatment of NSCLC in the future.

Vitexin induces apoptosis through mitochondrial pathway and PI3K/Akt/mTOR signaling in human non-small cell lung cancer A549 cells

BACKGROUND: Currently, the prognosis of patients with non-small cell lung cancer (NSCLC) remains dismal; hence, it is critical to identify effective anti-NSCLC agents with limited side effects. This study aimed to evaluate the therapeutic potential of flavonoid compound vitexin in human NSCLC cells and the underlying mechanisms. RESULTS: The experimental results indicated that vitexin reduced the viability of A549 cells in a dose-dependent manner with nearly no toxicity against normal human bronchial epithelial 16HBE cells. Vitexin also dose-dependently increased A549 cell apoptosis, accompanied by the decreased Bcl-2/Bax ratio and the increased expression of cleaved caspase-3. Moreover, the in vivo anticancer activity of vitexin was further determined in nude mice bearing A549 cells. In addition, vitexin induced the release of cytochrome c from the mitochondria to the cytosol and the loss of mitochondrial membrane potential. Vitexin also significantly reduced the levels of p-PI3K, p-Akt and p-mTOR, and the pro-apoptotic effect of vitexin on A549 cells was partly blocked by SC79, an Akt activator. CONCLUSIONS: Accordingly, we believed that vitexin could be used as a potential therapeutic agent for the treatment of NSCLC in the future.

Effect of lncRNA HULC knockdown on rat secreting pituitary adenoma GH3 cells

Pituitary adenoma is one of the most common tumors in the neuroendocrine system. This study investigated the effects of long non-coding RNAs (lncRNAs) highly up-regulated in liver cancer (HULC) on rat secreting pituitary adenoma GH3 cell viability, migration, invasion, apoptosis, and hormone secretion, as well as the underlying potential mechanisms. Cell transfection and qRT-PCR were used to change and measure the expression levels of HULC, miR-130b, and FOXM1. Cell viability, migration, invasion, and apoptosis were assessed using trypan blue staining assay, MTT assay, two-chamber transwell assay, Guava Nexin assay, and western blotting. The concentrations of prolactin (PRL) and growth hormone (GH) in culture supernatant of GH3 cells were assessed using ELISA. The targeting relationship between miR-130b and FOXM1 was verified using dual luciferase activity. Finally, the expression levels of key factors involved in PI3K/AKT/mTOR and JAK1/STAT3 pathways were evaluated using western blotting. We found that HULC was highly expressed in GH3 cells. Overexpression of HULC promoted GH3 cell viability, migration, invasion, PRL and GH secretion, as well as activated PI3K/AKT/mTOR and JAK1/STAT3 pathways. Knockdown of HULC had opposite effects and induced cell apoptosis. HULC negatively regulated the expression of miR-130b, and miR-130b participated in the effects of HULC on GH3 cells. FOXM1 was a target gene of miR-130b, which was involved in the regulation of GH3 cell viability, migration, invasion, and apoptosis, as well as PI3K/AKT/mTOR and JAK1/STAT3 pathways. In conclusion, HULC tumor-promoting roles in secreting pituitary adenoma might be via down-regulating miR-130b, up-regulating FOXM1, and activating PI3K/AKT/mTOR and JAK1/STAT3 pathways.

Metformin Plus Caloric Restriction Show Anti-epileptic Effects Mediated by mTOR Pathway Inhibition.

Caloric restriction (CR) has anti-epileptic effects in different animal models, at least partially due to inhibition of the mechanistic or mammalian target of rapamycin (mTOR) signaling pathway. Adenosine monophosphate-activated protein kinase (AMPK) inhibits mTOR cascade function if energy levels are low. Since hyper-activation of mTOR participates in epilepsy, its inhibition results in beneficial anti-convulsive effects. A way to attain this is to activate AMPK with metformin. The effects of metformin, alone or combined with CR, on the electrical kindling epilepsy model and the mTOR cascade in the hippocampus and the neocortex were studied. Combined metformin plus CR beneficially affected many kindling aspects, especially those relating to generalized convulsive seizures. Therefore, metformin plus CR could decrease measures of epileptic activity in patients with generalized convulsive seizures. Patients that are obese, overweight or that have metabolic syndrome in addition to having an epileptic disease are an ideal population for clinical trials to test the effectiveness of metformin plus CR.

Agmatine produces antidepressant-like effects by activating AMPA receptors and mTOR signaling.

The activation of AMPA receptors and mTOR signaling has been reported as mechanisms underlying the antidepressant effects of fast-acting agents, specially the NMDA receptor antagonist ketamine. In the present study, oral administration of agmatine (0.1mg/kg), a neuromodulator that has been reported to modulate NMDA receptors, caused a significant reduction in the immobility time of mice submitted to the tail suspension test (TST), an effect prevented by the administration of DNQX (AMPA receptor antagonist, 2.5µg/site, i.c.v.), BDNF antibody (1µg/site, i.c.v.), K-252a (TrkB receptor antagonist, 1µg/site, i.c.v.), LY294002 (PI3K inhibitor, 10nmol/site, i.c.v.) or rapamycin (selective mTOR inhibitor, 0.2nmol/site, i.c.v.). Moreover, the administration of lithium chloride (non-selective GSK-3ß inhibitor, 10mg/kg, p.o.) or AR-A014418 (selective GSK-3ß inhibitor, 0.01µg/site, i.c.v.) in combination with a sub-effective dose of agmatine (0.0001mg/kg, p.o.) reduced the immobility time in the TST when compared with either drug alone. Furthermore, increased immunocontents of BDNF, PSD-95 and GluA1 were found in the prefrontal cortex of mice just 1h after agmatine administration. These results indicate that the antidepressant-like effect of agmatine in the TST may be dependent on the activation of AMPA and TrkB receptors, PI3K and mTOR signaling as well as inhibition of GSK-3ß, and increase in synaptic proteins. The results contribute to elucidate the complex signaling pathways involved in the antidepressant effect of agmatine and reinforce the pivotal role of these molecular targets for antidepressant responses.

Resistência de células de carcinoma epidermóide bucal à terapia fotodinâmica mediada pelo ácido 5-aminolevulínico / Resistance of oral squamous cell carcinomas to 5-aminolevulinic acidmediated photodynamic therapy

O carcinoma epidermóide bucal (CEC) é uma neoplasia maligna com alta morbidade e mortalidade e de difícil tratamento. O tratamento convencional para o CEC inclui cirurgia e radioterapia, seguida ou não de quimioterapia. Apesar de serem amplamente difundidos, esses tratamentos podem ser ineficazes para alguns CECs resistentes. A terapia fotodinâmica (PDT) oncológica tem sido utilizada para o tratamento adjuvante do CEC bucal, principalmente nos casos menos invasivos e que necessitam de redução do tumor para a ressecção cirúrgica. Contudo, semelhantemente aos tratamentos convencionais, a PDT pode também induzir o aparecimento de populações celulares resistentes, fato já descrito para carcinoma cutâneo, adenocarcinoma de cólon e adenocarcinoma mamário. A hipótese de que células de CEC bucal possam desenvolver resistência à PDT ainda não foi testada. Portanto, o objetivo deste trabalho foi verificar se células de CEC bucal (SCC9) desenvolvem resistência a ciclos repetidos de PDT mediada pelo ácido 5- aminolevulínico (5-ALA-PDT) e avaliar se nesse processo ocorre modificação da expressão de marcadores relacionados a sobrevivência celular (NF?B, Bcl-2, iNOS, mTOR e Akt). Foi utilizada linhagem de células de CEC bucal (SCC9), submetida às seguintes condições: 1) Controle - células cultivadas sem nenhum tratamento; 2) ALA - células incubadas com 5-ALA (1mM durante 4 horas); 3) LED - tratadas com iluminação LED (630nm, 5,86J/cm2, 22,5J, 150mW, 150s); 4) PDT - tratadas com 5- ALA-PDT, com os protocolos do grupo ALA e LED combinados, gerando dose letal de 90%. Inicialmente foi realizado somente um ciclo de PDT, sendo avaliada a viabilidade celular em todos os grupos após 24, 48, 72 e 120h da irradiação. Também foi realizado ensaio de detecção da fragmentação de DNA (TUNEL) e análise por imunofluorescência da expressão das proteínas NF?B, Bcl-2, iNOS, pmTOR e pAkt nas células viáveis...

Oral squamous cell carcinoma (SCC) is a malignant tumor with high morbidity and mortality rates, and it is difficult to treat. Conventional treatment for oral SCCs includes surgery and radiotherapy that may be followed by chemotherapy. Although these treatments are widely used, they are ineffective against some resistant tumors. Oncologic photodynamic therapy (PDT) has been used as an adjuvant treatment for oral SCCs, especially in less invasive cases that require tumor reduction before surgical resection. However, like conventional treatments, PDT can induce the occurrence of resistant cell populations such as cutaneous carcinomas and colon and breast adenocarcinomas. The hypothesis that oral SCCs develop resistance to PDT has not yet been tested. Therefore, the aims of this study were to investigate whether oral SCCs (SCC9) develop resistance to several cycles of 5-aminolevulinic acidmediated PDT (5-ALA-PDT) and to determine whether the expression of markers associated with cell survival (NF?B, Bcl-2, iNOS, mTOR, and Akt) is altered during this process. An oral SCC (SCC9) cell line was used, which was subjected to the following conditions: 1) Control: cultured without any treatment; 2) ALA: incubated with 5-ALA (1 mM for 4 h); 3) LED: treated with LED light (630 nm, 5.86 J/cm2, 22.5 J, 150 mW, 150 s); and 4) PDT: treated with 5-ALA-PDT (with the protocols of the ALA and LED groups combined) generating a lethal dose of 90%. Initially, only one cycle of PDT was administered, and cell viability was determined in all groups 24, 48, 72, and 120 h after irradiation. Subsequently, the DNA fragmentation detection assay (TUNEL) and immunofluorescence analysis of the expression of proteins NF?B, Bcl-2, iNOS, pmTOR, and pAkt were performed on viable cells. The fraction of cells that survived the first treatment with 5-ALA-PDT exhibited intense staining for pmTOR and growth potential during the testing period...