Revealing the mechanism of quinoa on type 2 diabetes based on intestinal flora and taste pathways.

To investigate the antidiabetic effects and mechanisms of quinoa on type 2 diabetes mellitus (T2DM) mice model. In this context, we induced the T2DM mice model with a high-fat diet (HFD) combined with streptozotocin (STZ), followed by treatment with a quinoa diet. To explore the impact of quinoa on the intestinal flora, we predicted and validated its potential mechanism of hypoglycemic effect through network pharmacology, molecular docking, western blot, and immunohistochemistry (IHC). We found that quinoa could significantly improve abnormal glucolipid metabolism in T2DM mice. Further analysis showed that quinoa contributed to the improvement of gut microbiota composition positively. Moreover, it could downregulate the expression of TAS1R3 and TRPM5 in the colon. A total of 72 active components were identified by network pharmacology. Among them, TAS1R3 and TRPM5 were successfully docked with the core components of quinoa. These findings confirm that quinoa may exert hypoglycemic effects through gut microbiota and the TAS1R3/TRPM5 taste signaling pathway.

Moringa oleifera leaf attenuate osteoporosis in ovariectomized rats by modulating gut microbiota composition and MAPK signaling pathway.

Moringa oleifera leaf (MLP) contains abundant complex nutrients with anti-osteoporosis potential. However, its efficacy and mechanisms against osteoporosis remain unknown. The purpose of this research is to investigate MLP's anti-osteoporotic effects and mechanisms. Animal experiments were used in this work to validate MLP's anti-osteoporotic efficacy. We investigated the mode of action of MLP, analyzed its impact on the gut microbiota, and predicted and validated its anti-osteoporosis-related molecular targets and pathways through network pharmacology, molecular docking, and western blotting. In an ovariectomized osteoporosis rat model, MLP significantly increased bone mineral density and improved bone metabolism-related indicators, bone microstructure, and lipid profile. Moreover, it improved gut microbiota composition and increased the expression of Occludin and Claudin-1 protein in the duodenum. Network pharmacology identified a total of 97 active ingredients and 478 core anti-osteoporosis targets. Of these, MAPK1 (also known as ERK2), MAPK3 (also known as ERK1), and MAPK8 (also known as JNK) were successfully docked with the active constituents of MLP. Interestingly, MLP increased ERK and VAV3 protein expression and decreased p-ERK and JNK protein expression in the femur. These findings confirm MLP's anti-osteoporotic efficacy, which could be mediated via regulation of gut microbiota and MAPK signaling.

Gut Microbiome Composition of the Fire Ant Solenopsis invicta: an Integrated Analysis of Host Genotype and Geographical Distribution.

Gut symbiotic bacteria are known to be closely related to insect development, nutrient metabolism, and disease resistance traits, but the most important factors leading to changes in these communities have not been well clarified. To address this, we examined the associations between the gut symbiotic bacteria and the host genotype and geographical distribution of Solenopsis invicta in China, where it is invasive and has spread primarily by human-mediated dispersal. Thirty-two phyla were detected in the gut symbiotic bacteria of S. invicta. Proteobacteria were the most dominant group among the gut symbiotic bacteria. Furthermore, the Bray-Curtis dissimilarity matrices of the gut symbiotic bacteria were significantly positively correlated with the geographical distance between the host ant colonies, but this relationship was affected by the social form. The distance between monogyne colonies had a significant effect on the Bray-Curtis dissimilarity matrices of gut symbiotic bacteria, but the distance between polygyne colonies did not. Moreover, the Bray-Curtis dissimilarity matrices were positively correlated with Nei's genetic distance of the host but were not correlated with the COI-based genetic distance. This study provides a scientific basis for further understanding the ecological adaptability of red imported fire ants during invasion and dispersal. IMPORTANCE We demonstrated that gut microbiota composition and diversity varied among populations. These among-population differences were associated with host genotype and geographical distribution. Our results suggested that population-level differences in S. invicta gut microbiota may depend more on environmental factors than on host genotype.

TAT&RGD Peptide-Modified Naringin-Loaded Lipid Nanoparticles Promote the Osteogenic Differentiation of Human Dental Pulp Stem Cells.

Background: Naringin is a naturally occurring flavanone that promotes osteogenesis. Owing to the high lipophilicity, poor in vivo bioavailability, and extensive metabolic alteration upon administration, the clinical efficacy of naringin is understudied. Additionally, information on the molecular mechanism by which it promotes osteogenesis is limited. Methods: In this study, we prepared TAT & RGD peptide-modified naringin-loaded nanoparticles (TAT-RGD-NAR-NPs), evaluated their potency on the osteogenic differentiation of human dental pulp stem cells (hDPSCs), and studied its mechanism of action through metabolomic analysis. Results: The particle size and zeta potential of TAT-RGD-NAR-NPs were 160.70±2.05 mm and -20.77±0.47mV, respectively. The result of cell uptake assay showed that TAT-RGD-NAR-NPs could effectively enter hDPSCs. TAT-RGD-NAR-NPs had a more significant effect on cell proliferation and osteogenic differentiation promotion. Furthermore, in metabolomic analysis, naringin particles showed a strong influence on the glycerophospholipid metabolism pathway of hDPSCs. Specifically, it upregulated the expression of PLA2G3 and PLA2G1B (two isozymes of phospholipase A2, PLA2), increased the biosynthesis of lysophosphatidic acid (LPA). Conclusion: These results suggested that TAT-RGD-NPs might be used for transporting naringin to hDPSCs for modulating stem cell osteogenic differentiation. The metabolomic analysis was used for the first time to elucidate the mechanism by which naringin promotes hDPSCs osteogenesis by upregulating PLA2G3 and PLA2G1B.

VEGF-Loaded Heparinised Gelatine-Hydroxyapatite-Tricalcium Phosphate Scaffold Accelerates Bone Regeneration via Enhancing Osteogenesis-Angiogenesis Coupling.

Background: Bone tissue defect, one of the common orthopaedicdiseases, is traumatizing and affects patient's lifestyle. Although autologous and xenograft bone transplantations are performed in bone tissue engineering, clinical development of bone transplantation is limited because ofvarious factors, such as varying degrees of immune rejection, lack of bone sources, and secondary damage to bone harvesting. Methods: We synthesised a heparinised gelatine-hydroxyapatite-tricalcium phosphate (HG-HA-TCP) scaffold loaded with sustained-release vascular endothelial growth factor (VEGF) analysed their structure, mechanical properties, and biocompatibility. Additionally, the effects of HG-HA-TCP (VEGF) scaffolds on osteogenic differentiation and vascularisation of stem cells from human exfoliated deciduous teeth (SHED) in vitro and bone regeneration in vivo were investigated. Results: HG-HA-TCP scaffold possessed good pore structure, mechanical properties, and biocompatibility. HG-HA-TCP scaffold loaded with VEGF could effectively promote SHED proliferation, migration, and adhesion. Moreover, HG-HA-TCP (VEGF) scaffold increased the expression of osteogenesis- and angiogenesis-related genes and promoted osteogenic differentiation and vascularisation in cells. In vivo results demonstrated that VEGF-loaded HG-HA-TCP scaffold improved new bone regeneration and enhanced bone mineral density, revealed byhistological, micro-CT and histochemical straining analyses. Osteogenic and angiogenic abilities of the three biological scaffolds wereranked as follows: HG-HA-TCP (VEGF) > G-HA-TCP (VEGF) > G-HA-TCP. Conclusion: HG-HA-TCP (VEGF) scaffold with good biocompatibility could create an encouraging osteogenic microenvironment that could accelerate vessel formation and osteogenesis, providing an effective scaffold for bone tissue engineering and developing new clinical treatment strategies for bone tissue defects.

ROR2 Downregulation Activates the MSX2/NSUN2/p21 Regulatory Axis and Promotes Dental Pulp Stem Cell Senescence.

Cellular senescence severely limits the research and the application of dental pulp stem cells (DPSCs). A previous study conducted by our research group revealed a close implication of ROR2 in DPSC senescence, although the mechanism underlying the regulation of ROR2 in DPSCs remains poorly understood so far. In the present study, it was revealed that the expression of the ROR2-interacting transcription factor MSX2 was increased in aging DPSCs. It was demonstrated that the depletion of MSX2 inhibits the senescence of DPSCs and restores their self-renewal capacity, and the simultaneous overexpression of ROR2 enhanced this effect. Moreover, MSX2 knockdown suppressed the transcription of NOP2/Sun domain family member 2 (NSUN2), which regulates the expression of p21 by binding to and causing the 5-methylcytidine methylation of the 3'- untranslated region of p21 mRNA. Interestingly, ROR2 downregulation elevated the levels of MSX2 protein, and not the MSX2 mRNA expression, by reducing the phosphorylation level of MSX2 and inhibiting the RNF34-mediated MSX2 ubiquitination degradation. The results of the present study demonstrated the vital role of the ROR2/MSX2/NSUN2 axis in the regulation of DPSC senescence, thereby revealing a potential target for antagonizing DPSC aging.

Investigation of a Novel LRP6 Variant Causing Autosomal-Dominant Tooth Agenesis.

BACKGROUND: The low-density lipoprotein receptor-related protein 6 (LRP6) gene is a recently defined gene that is associated with the autosomal-dominant inherited tooth agenesis (TA). In the present study, a family of four generations having TA was recruited and subjected to a series of clinical, genetic, in silico, and in vitro investigations. METHODS: After routine clinical evaluation, the proband was subjected to whole-exome sequencing (WES) to detect the diagnostic variant. Next, in silico structural and molecular dynamics (MD) analysis was conducted on the identified novel missense variant for predicting its intramolecular impact. Subsequently, an in vitro study was performed to further explore the effect of this variant on protein maturation and phosphorylation. RESULTS: WES identified a novel variant, designated as LRP6: c.2570G > A (p.R857H), harbored by six members of the concerned family, four of whom exhibited varied TA symptoms. The in silico analysis suggested that this novel variant could probably damage the Wnt bonding function of the LRP6 protein. The experimental study demonstrated that although this novel variant did not affect the LRP6 gene transcription, it caused a impairment in the maturation and phosphorylation of LRP6 protein, suggesting the possibility of the disruption of the Wnt signaling. CONCLUSION: The present study expanded the mutation spectrum of human TA in the LRP6 gene. The findings of the present study are insightful and conducive to understanding the functional significance of specific LRP6 variants.

Quercetin Efficiently Alleviates TNF-α-Stimulated Injury by Signal Transducer and Activator of Transcription 1 and Mitogen-Activated Protein Kinase Pathway in H9c2 Cells: A Protective Role of Quercetin in Myocarditis.

ABSTRACT: This study aimed to evaluate the protective effect of quercetin and its in-depth mechanism in TNF-α-stimulated cardiomyocytes. The differential expression of TNF-alpha (TNF-α) and signal transducer and activator of transcription 1 (STAT1) was analyzed based on the GEO database. H9c2 cells were stimulated with TNF-α to simulate myocarditis. Cell counting kit-8 assay and flow cytometry assay were performed to detect the cell viability and apoptosis. ELISA was used to measure the levels of proinflammatory cytokines (IL-6 and IL-17A) and anti-inflammatory cytokine (IL-10). STAT1 expression was downregulated by transfection with si-STAT1, and its expression was detected using quantitative real-time polymerase chain reaction and Western blot. Western blot was also performed to assess the expression of the mitogen-activated protein kinase (MAPK) pathway-related factors. In this article, TNF-α was highly expressed in patients with myocarditis, and TNF-α (20 µg/mL) declined the viability of H9c2 cells. Quercetin pretreatment partially alleviated the decrease of cell viability, the increase of apoptosis, and the release of inflammatory cytokines (IL-10, IL-6, and IL-17A) induced by TNF-α. In addition, TNF-α increased STAT1 expression, but quercetin prevented the TNF-α-increased STAT1 level. Remarkably, knockdown of STAT1 enhanced the protective effect of quercetin on TNF-α-injured H9c2 cells. Moreover, quercetin restrained the TNF-α-induced activation of the MAPK pathway. Also, the inhibitory effect of quercetin on the pathway was aggravated by STAT1 lacking. In summing, quercetin plays a protective role in TNF-α-stimulated H9c2 cell injury, which may be related to the regulation of STAT1 and MAPK pathway.

Efficacy of paclitaxel, carboplatin, and bevacizumab for cervical cancer: A protocol for systematic review and meta-analysis.

BACKGROUND: Cervical cancer (CC) is a very common and malignant tumor in female population. Although a variety of single medications are reported to treat this condition, they all have limited efficacy. Previous studies have reported the combination of paclitaxel, carboplatin, and bevacizumab (PCB) can be used for the treatment of patients with CC effectively. However, no systematic review has explored its efficacy and safety. This study will address its efficacy and safety systematically and comprehensively. METHODS: The following electronic databases will be retrieved from their inceptions to the January 1, 2020 to identify all potential associated studies: MEDLINE, EMBASE, Cochrane Library, Scopus, Web of Science, CINAHL, Google scholar, and Chinese Biomedical Literature Database. We will include randomized controlled trials (RCTs) of adult women (≥18 years) with CC globally. Eligible interventions will target any forms of PCB. The study methodological quality of all included studies will be appraised using Cochrane risk of bias tool. Statistical analysis will be undertaken using RevMan 5.3 software. In addition, we will perform a narrative synthesis to describe quality and content of the evidence. RESULTS: This study will summarize recent evidence and provide quality evidence for the efficacy and safety of PCB on CC. CONCLUSION: The findings of this study will seek to identify the efficacy and safety of PCB and suggest future directions for research efforts targeting CC among this population. SYSTEMATIC REVIEW REGISTRATION: INPLASY202040195.

Serum complement C1q level is associated with acute coronary syndrome.

BACKGROUND AND OBJECTIVES: The complement system plays an important role in the development of acute coronary syndrome (ACS). Complement C1q is an important initial component of the classical complement pathway and closely related to many chronic inflammatory diseases, including atherosclerosis (AS). We aimed to determine whether there was association between serum complement C1q and the severity of coronary stenosis. SUBJECTS AND METHODS: 320 patients who underwent coronary arteriography (CAG) were stratified into non-ACS group (control group, n = 74), unstable angina group (UA group, n = 197) and acute myocardial infarction group (AMI group, n = 49) according to the severity of coronary stenosis and clinical manifestations. The severity of coronary stenosis was represented in Gensini score, and serum complement C1q level was compared using immunity transmission turbidity among three groups. RESULTS: The level of complement C1q in AMI group was lower significantly than control group and UA group (P < 0.05), but there was no correlation between serum complement C1q and Gensini score (ß=-0.086, P = 0.125). In nitrate-taking patients, serum complement C1q had a negative association with Gensini score (r=-0.275, P = 0.001), and in non-smokers, there was also a negative correlation (ß=-0.159, P = 0.036). After calibrating smoking, drinking or statins, the serum complement C1q levels of control group, UA group and AMI group decreased in sequence (P <  0.05). Logistic regression analysis showed that the decreasing of serum complement C1q was an unfavorable factor for acute myocardial infarction (OR=0.984, 95 %CI=0.972∼0.997, P = 0.015) and for ACS (OR=0.984, 95 %CI=0.971∼0.984, P = 0.025) in drinking patients. Regrettably, ROC curve suggested that the accuracy in diagnosing coronary atherosclerotic heart disease by serum complement C1q was low (AUC=0.568, 95 %CI= 0.492-0.644, P = 0.076, sensitivity 73.6 %, specificity 58.1 %). CONCLUSION: Serum complement C1q in ACS patients, in particular AMI patients, showed lower level. This finding suggests further decrease of complement C1q level in ACS patients may be a contributory factor to instability or rupture of atherosclerotic plaques. Combined with other clinical indicators, it can be helpful to predict the risk and severity of coronary stenosis.

Spirometric Reference Equations for Elderly Chinese in Jinan Aged 60-84 Years.

BACKGROUND: The interpretation of spirometry varies on different reference values. Older people are usually underrepresented in published predictive values. This study aimed at developing spirometric reference equations for elderly Chinese in Jinan aged 60-84 years and to compare them to previous equations. METHODS: The project covered all of Jinan city, and the recruitment period lasted 9 months from January 1, 2017 to September 30, 2017, 434 healthy people aged 60-84 years who had never smoked (226 females and 208 males) were recruited to undergo spirometry. Vital capacity (VC), forced VC (FVC), forced expiratory volume in 1 s (FEV1), FEV1/FVC, FEV1/VC, FEV6, peak expiratory flow, and forced expiratory flow at 25%, 50%, 75%, and 25-75% of FVC exhaled (FEF25%, FEF50%, FEF75%, and FEF25-75%) were analyzed. Reference equations for mean and the lower limit of normal (LLN) were derived using the lambda-mu-sigma method. Comparisons between new and previous equations were performed by paired t-test. RESULTS: New reference equations were developed from the sample. The LLN of FEV1/FVC, FEF25-75%computed using the 2012-Global Lung Function Initiative (GLI) and 2006-Hong Kong equations were both lower than the new equations. The biggest degree of difference for FEV1/FVC was 19% (70.46% vs. 59.29%, t = 33.954, P < 0.01) and for maximal midexpiratory flow (MMEF, equals to FEF25-75%) was 22% (0.82 vs. 0.67, t = 21.303, P < 0.01). The 1990-North China and 2009-North China equations predicted higher mean values of FEV1/FVC and FEF25-75%than the present model. The biggest degrees of difference were -4% (78.31% vs. 81.27%, t = -85.359, P < 0.01) and -60% (2.11 vs. 4.68, t = -170.287, P < 0.01), respectively. CONCLUSIONS: The newly developed spirometric reference equations are applicable to elderly Chinese in Jinan. The 2012-GLI and 2006-Hong Kong equations may lead to missed diagnoses of obstructive ventilatory defects and the small airway dysfunction, while traditional linear equations for all ages may lead to overdiagnosis.

Corrigendum: BDNF val66met Polymorphism Impairs Hippocampal Long-Term Depression by Down-Regulation of 5-HT3 Receptors.

[This corrects the article on p. 306 in vol. 11, PMID: 29075179.].

Electrode Placing Sites affect Pulp Vitality Test of Human Incisors and Premolars.

OBJECTIVE: To investigate electric pulp test thresholds at different sites on healthy incisors and premolars and determine appropriate test sites. METHODS: Overall, 47 volunteers aged 20 to 30 years were recruited, and 163 incisors and 140 premolars were tested at several sites with an Electric Pulp Tester. One-way analysis of variance and a Tukey test were used to analyse the threshold values among different tooth types and sites. RESULTS: The lowest threshold value for incisors was identified on the incisal edge. The difference of threshold on the incisal edge and other sites was statistically significant in mandibular incisors. For maxillary premolars and the mandibular second premolar, the lowest response was obtained with the tester tip on the lingual slope of the buccal cusp. For the mandibular first premolar, the response at the lingual slope of the buccal cusp, as the second lowest, was slightly higher than that at the buccal cusp. CONCLUSION: The incisal edge for incisors and the lingual slope of the buccal cusp for premolars was favoured as the optimal sites for electric pulp test.

BDNF val66met Polymorphism Impairs Hippocampal Long-Term Depression by Down-Regulation of 5-HT3 Receptors.

Brain-derived neurotrophic factor (BDNF) is a key regulator of neuronal plasticity and cognitive functions. BDNF val66met polymorphism, a human single-nucleotide polymorphism (SNP) in the pro-domain of BDNF gene, is associated with deficits in activity-dependent BDNF secretion and hippocampus-dependent memory. However, the underlying mechanism remains unclear. Here we show that in the BDNFMet/Met mouse line mimicking the human SNP, BDNF expression in the hippocampus was decreased. There was a reduction in the total number of cells in hippocampal CA1 region, while hippocampal expression of mRNAs for NR2a, 2b, GluR1, 2 and GABAARß3 subunits were up-regulated. Although basal glutamatergic neurotransmission was unaltered, hippocampal long-term depression (LTD) induced by low-frequency stimulation was impaired, which was partially rescued by exogenous application of BDNF. Interestingly, 5-HT3a receptors were down-regulated in the hippocampus of BDNFMet/Met mice, whereas 5-HT2c receptors were up-regulated. Moreover, impaired LTD in BDNFMet/Met mice was reversed by 5-HT3aR agonist. Thus, these observations indicate that BDNF val66met polymorphism changes hippocampal synaptic plasticity via down-regulation of 5-HT3a receptors, which may underlie cognition dysfunction of Met allele carriers.

Combination effects of arsenic trioxide and fibroblast growth factor receptor inhibitor in squamous cell lung carcinoma.

OBJECTIVES: Lung cancer remains the top cancer killer worldwide, with squamous cell carcinoma (SCC) as the second commonest histologic subtype. Arsenic trioxide (ATO) was previously shown to suppress growth of lung cancer. Fibroblast growth factor receptor (FGFR) amplification was recently demonstrated in lung SCC, with specific FGFR inhibitor (e.g. PD173074) developed as a potential targeted therapy. Therefore the combination effects of ATO and PD173074 in SCC was studied. MATERIALS AND METHODS: The combination of ATO/PD173074 was studied in a proof-of-principle model using a lung SCC cell line with FGFR1 overexpression: SK-MES-1. The effects of ATO and/or PD173074 on cell viability and protein expression were studied by MTT assay and Western blot respectively. Cell cycle analysis, phosphatidylserine externalization and mitochondrial membrane depolarization were monitored by flow cytometry. FGFR1 knockdown was performed with siRNAs. Proteasome inhibitor (MG-132) was used to study the degradation mechanism. In vivo effect of ATO and/or PD173074 was investigated using a nude mice xenograft model. RESULTS: Combined ATO/PD173074 reduced cell viability along with increased sub-G1 population, phosphatidylserine externalization and mitochondrial membrane depolarization more significantly than single treatments. Downregulation of FGFR1, p-Akt, Akt, p-Src, Src, p-c-Raf, c-Raf, Erk and survivin as well as upregulation of p-Erk and cleaved PARP were observed upon ATO and/or PD treatment. MG-132 partially reversed the degradation of Akt, Src, c-Raf and Erk induced by ATO/PD, suggestive of ubiquitin-independent proteasome-dependent degradation. However, the mechanism of FGFR1 downregulation remained unknown. Downregulation of FGFR1, Akt, Src, c-Raf and Erk as well as cleaved PARP elevation induced by ATO and/or PD were confirmed in vivo. CONCLUSION: Massive protein degradation (FGFR1, Akt, Src, c-Raf and Erk) was induced by ATO and/or PD173074 treatment mainly mediated by activation of proteasomal degradation in SCC cell line SK-MES-1 in vitro and in vivo.

The Effect of Tumor Microenvironment on Autophagy and Sensitivity to Targeted Therapy in EGFR-Mutated Lung Adenocarcinoma.

Lung cancer is the top cancer killer worldwide. Tyrosine kinase inhibitors (TKIs), for example erlotinib, are commonly used to target epidermal growth factor receptor (EGFR)-mutated lung adenocarcinoma (ADC). Autophagy is a cellular response to stress, serving as a protective mechanism during anticancer therapy. The tumor microenvironment (TME) is composed of non-tumor cells that include fibroblasts. Our study aimed to investigate the effect of TME on autophagy and TKI sensitivity. Following cell sorting after direct co-culturing, autophagy and cytokine production were observed in both HCC827 and MRC-5 cells. The synergistic combination of erlotinib and chloroquine (autophagy inhibitor) was observed under TME. Tumor growth was significantly suppressed with combined erlotinib/chloroquine compared with erlotinib in HCC827 xenografts.

Arsenic trioxide-induced cytotoxicity in small cell lung cancer via altered redox homeostasis and mitochondrial integrity.

Arsenic trioxide (ATO) has demonstrated anticancer activity in different malignancies, especially acute promyelocytic leukemia, with a wide array of putative mechanisms. In this study, we aimed to elucidate the activity and mechanisms of ATO in small cell lung cancer (SCLC). A panel of SCLC cell lines (H841, DMS79, H526, H69 and H187) was employed to demonstrate the activity of ATO. Cell viability, apoptosis and mitochondrial membrane depolarization were assessed. Western blotting was performed to determine the alteration of pro-apoptotic and anti-apoptotic mediators. Reactive oxygen species (ROS) (hydrogen peroxide and superoxide) and intracellular glutathione (GSH) were measured. Antioxidants, N-acetyl-L-cysteine (NAC) and butylated hydroxyanisole (BHA), were applied to restore GSH content and reduce production of ROS. All SCLC cell lines were relatively sensitive to ATO with IC50 values below 10 µM. ATO induced cell death mainly through apoptosis in H841 cells in a dose-dependent manner. Hydrogen peroxide was the major ROS in SCLC cells induced by ATO. Along with GSH depletion and Bcl-2 downregulation, mitochondrial membrane permeabilization was enhanced, followed by release of AIF and SMAC from mitochondria to initiate different cell death pathways. NAC reversed cell death and molecular changes induced by ATO via restoring GSH and reducing ROS content. BHA inhibited hydrogen peroxide production completely and partially restored GSH content accounting for partial reversal of cell inhibition and mitochondrial dysfunction. Nonetheless, ATO reduced both reduced and oxidized form of thioredoxin 1 (Trx1) with no effect on Trx1 redox potential. ATO led to cell death in SCLC mainly through mitochondrial dysfunction, resulting from altered cellular redox homeostasis, namely, hydrogen peroxide generation, GSH depletion and Trx1 downregulation.

Downregulation of thymidylate synthase and E2F1 by arsenic trioxide in mesothelioma.

Malignant pleural mesothelioma is a global health issue. Arsenic trioxide (ATO) has been shown to suppress thymidylate synthase (TYMS) in lung adenocarcinoma and colorectal cancer, and induce apoptosis in acute promyelocytic leukemia. With TYMS as a putative therapeutic target, the effect of ATO in mesothelioma was therefore studied. A panel of 5 mesothelioma cell lines was used to study the effect of ATO on cell viability, protein expression, mRNA expression and TYMS activity by MTT assay, western blot, qPCR and tritium-release assay, respectively. The knockdown of TYMS and E2F1 was performed with a specific siRNA. Phosphatidylserine externalization and mitochondrial membrane depolarization were measured by Annexin V and JC-1 staining respectively. The in vivo effect of ATO was studied using a nude mouse xenograft model. Application of ATO demonstrated anticancer effects in the cell line model with clinically achievable concentrations. Downregulation of TYMS protein (except H226 cells and 1.25 µM ATO in H2052 cells) and mRNA expression (H28 cells), pRB1 (H28 cells) and E2F1 and TYMS activity (except H226 cells) were also evident. E2F1 knockdown decreased cell viability more significantly than TYMS knockdown. In general, thymidine kinase 1, ribonucleotide reductase M1, c-myc and skp2 were downregulated by ATO. p-c-Jun was downregulated in H28 cells while upregulated in 211H cells. Phosphatidylserine externalization, mitochondrial membrane depolarization, downregulation of Bcl-2 and Bcl-xL, and upregulation of Bak and cleaved caspase-3 were observed. In the H226 xenograft model, the relative tumor growth was aborted, and E2F1 was downregulated while cleaved caspase-3 was elevated and localized to the nucleus in the ATO treatment group. ATO has potent antiproliferative and cytotoxic effects in mesothelioma in vitro and in vivo, partially mediated through E2F1 targeting (less effect through TYMS targeting). There is sound scientific evidence to support the clinical application of ATO in treatment of mesothelioma.

E2F1 downregulation by arsenic trioxide in lung adenocarcinoma.

Lung cancer is one of the most common cancers worldwide. Arsenic trioxide (ATO) has been approved by the US Food and Drug Administration for the treatment of acute promyelocytic leukemia. Nonetheless preliminary data have suggested potential activity of ATO in solid tumors including lung cancer. This study aimed to examine the underlying mechanisms of ATO in the treatment of lung adenocarcinoma. Using a panel of 7 lung adenocarcinoma cell lines, the effects of ATO treatment on cell viability, expression of E2F1 and its downstream targets, phosphatidylserine externalization, mitochondrial membrane depolarization and alteration of apoptotic/anti-apoptotic factors were studied. Tumor growth inhibition in vivo was investigated using a nude mouse xenograft model. ATO decreased cell viability with clinically achievable concentrations (8 µM) in all cell lines investigated. This was accompanied by reduced expression of E2F1, cyclin A2, skp2, c-myc, thymidine kinase and ribonucleotide reductase M1, while p-c-Jun was upregulated. Cell viability was significantly decreased with E2F1 knockdown. Treatment with ATO resulted in phosphatidylserine externalization in H23 cells and mitochondrial membrane depolarization in all cell lines, associated with truncation of Bid, downregulation of Bcl-2, upregulation of Bax and Bak, caspase-9 and -3 activation and PARP cleavage. Using the H358 xenograft model, the tumor growth was suppressed in the ATO treatment group during 8 days of treatment, associated with downregulation of E2F1 and upregulation of truncated Bid and cleaved caspase-3. In conclusion, ATO has potent in vitro and in vivo activity in lung adenocarcinoma, partially mediated through E2F1 downregulation and apoptosis.

Downregulation of thymidylate synthase with arsenic trioxide in lung adenocarcinoma.

Thymidylate synthase (TYMS) is an important chemotherapeutic target in non-small cell lung cancer (NSCLC). Arsenic trioxide (ATO) has been shown to suppress TYMS in a colonic cancer model. We examined the effects of TYMS suppression by ATO in lung adenocarcinoma. A panel of 4 lung adenocarcinoma cell lines was used to determine the effects of ATO treatment on cell viability, TYMS expression (protein and mRNA), E2F1 protein expression and TYMS activity. TYMS knockdown and overexpression were performed. Tumor growth inhibition in vivo was studied using a nude mouse xenograft model. ATO showed antiproliferative effects with clinically achievable concentrations (around 1.1-6.9 µM) in 4 lung adenocarcinoma cell lines. Downregulation of TYMS protein and mRNA expression, reduced TYMS activity, and suppressed E2F1 expression were demonstrated in lung adenocarcinoma with ATO. Cell viability was reduced by 15-50% with TYMS knockdown. Overexpression of TYMS led to a 2.7-fold increase in IC50 value with ATO treatment in H358 cells, but not in H23 cells. Using a xenograft model with H358 cell line, relative tumor volume was reduced to 44% that of the control following 8 days of treatment with 7.5 mg/kg ATO, and associated with significant downregulation of TYMS protein expression. In conclusion, ATO has potent in vitro and in vivo activity in lung adenocarcinoma, and is partially mediated by transcriptional downregulation of TYMS.