Anti-ENO1 antibody combined with metformin against tumor resistance: a novel antibody-based platform.

Background: Antibody-based platforms (i.e., ADC) have emerged as one of the most encouraging tools for the cancer resistance caused by cancer stem cells (CSCs) enrichment. Our study might provide a promising therapeutic direction against drug resistance and serve as a potential precursor platform for screening ADC. Methods: The cell migration, invasion, drug resistance, and self-renewal were assessed by the cell invasion and migration assay, wound healing assay, CCK-8 assay, colony formation assay, and sphere formation assay, respectively. The expression profiles of CSCs (ALDH+ and CD44+) subpopulations were screened by flow cytometry. The western blot and cell immunofluorescence assay were used to evaluate pathway-related protein expression in both anti-ENO1 antibody, MET combined with DPP/CTX-treated CSCs. Results: In the present study, western blot and flow cytometry verified that anti-ENO1 antibody target the CD44+ subpopulation by inhibiting the PI3K/AKT pathway, while metformin might target the ALDH+ subpopulation through activation of the AMPK pathway and thus reverse drug resistance to varying degrees. Subsequently, in vitro investigation indicated that anti-ENO1 antibody, metformin combined with cisplatin/cetuximab could simultaneously target ALDH+ and CD44+ subpopulations. The combination also inhibited the CSCs proliferation, migration, invasion, and sphere formation; which may result in overcoming the drug resistance. Then, molecular mechanism exploration verified that the anti-ENO1 antibody, metformin combined with cisplatin/cetuximab inhibited the Wnt/ß-catenin signaling. Conclusions: The study preliminarily revealed anti-ENO1 antibody combined with metformin could overcome drug resistance against CSCs by inhibiting the Wnt//ß-catenin pathway and might serve as a potential precursor platform for screening ADC. More importantly, it is reasonably believed that antibody-based drug combination therapy might function as an encouraging tool for oncotherapy.

HSP90, as a functional target antigen of a mAb 11C9, promotes stemness and tumor progression in hepatocellular carcinoma.

BACKGROUND: Identification of promising targeted antigens that exhibited cancer-specific expression is a crucial step in the development of novel antibody-targeted therapies. We here aimed to investigate the anti-tumor activity of a novel monoclonal antibody (mAb) 11C9 and identify the antibody tractable target in the hepatocellular cancer stem cells (HCSCs). METHODS: The identification of the targeted antigen was conducted using SDS-PAGE, western blot, mass spectrometry, and co-immunoprecipitation. Silence of HSP90 was induced by siRNA interference. Positive cells were sorted by fluorescence-activated cell sorting. Double-immunofluorescent (IF) staining and two-color flow cytometry detected the co-expression. Self-renewal, invasion, and drug resistance were assessed by sphere formation, matrigel-coated Transwell assay, and CCK-8 assay, respectively. Tumorigenicity was evaluated in mouse xenograft models. RNA-seq and bioinformatics analysis were performed to explore the mechanism of mAb 11C9 and potential targets. RESULTS: MAb 11C9 inhibited invasion and self-renewal abilities of HCC cell lines and reversed the cisplatin resistance. HSP90 (~ 95 kDa) was identified as a targeted antigen of mAb 11C9. Tissue microarrays and online databases revealed that HSP90 was overexpressed in HCC and associated with a poor prognosis. FACS and double-IF staining showed the co-expression of HSP90 and CSCs markers (CD90 and ESA). In vitro and in vivo demonstrated the tumorigenic potentials of HSP90. The inhibition of HSP90 by siRNA interference or 17-AAG inhibitor both decreased the number of invasion, sphere cells, and CD90+ or ESA+ cells, as well as reversed the resistance. Bioinformatics analysis and western blot verified that HSP90 activated Wnt/ß-catenin signaling. CONCLUSIONS: The study preliminarily revealed the anti-tumor activity of mAb 11C9. More importantly, we identified HSP90 as a targeted antigen of mAb 11C9, which functions as an oncogene in phenotype shaping, stemness maintenance, and therapeutic resistance by activating Wnt/ß-catenin signaling.

Ghrelin promotes cardiomyocyte differentiation of adipose tissue­derived mesenchymal stem cells by DDX17­mediated regulation of the SFRP4/Wnt/ß­catenin axis.

Adipose tissue­derived mesenchymal stem cells (ADMSCs) differentiate into cardiomyocytes and may be an ideal cell source for myocardial regenerative medicine. Ghrelin is a gastric­secreted peptide hormone involved in the multilineage differentiation of MSCs. To the best of our knowledge, however, the role and potential downstream regulatory mechanism of ghrelin in cardiomyocyte differentiation of ADMSCs is still unknown. The mRNA and protein levels were measured by reverse transcription­quantitative PCR and western blotting. Immunofluorescence staining was used to show the expression and cellular localization of cardiomyocyte markers and ß­catenin. RNA sequencing was used to explore the differentially expressed genes (DEGs) that regulated by ghrelin. The present study found that ghrelin promoted cardiomyocyte differentiation of ADMSCs in a concentration­dependent manner, as shown by increased levels of cardiomyocyte markers GATA binding protein 4, α­myosin heavy chain (α­MHC), ISL LIM homeobox 1, NK2 homeobox 5 and troponin T2, cardiac type. Ghrelin increased ß­catenin accumulation in nucleus and decreased the protein expression of secreted frizzled­related protein 4 (SFRP4), an inhibitor of Wnt signaling. RNA sequencing was used to determine the DEGs regulated by ghrelin. Functional enrichment showed that DEGs were more enriched in cardiomyocyte differentiation­associated terms and Wnt pathways. Dead­box helicase 17 (DDX17), an upregulated DEG, showed enhanced mRNA and protein expression levels following ghrelin addition. Overexpression of DDX17 promoted protein expression of cardiac­specific markers and ß­catenin and enhanced the fluorescence intensity of α­MHC and ß­catenin. DDX17 upregulation inhibited protein expression of SFRP4. Rescue assay confirmed that the addition of SFRP4 partially reversed ghrelin­enhanced protein levels of cardiac­specific markers and the fluorescence intensity of α­MHC. In conclusion, ghrelin promoted cardiomyocyte differentiation of ADMSCs by DDX17­mediated regulation of the SFRP4/Wnt/ß­catenin axis.

Synthesis and antitumor activity of erysolin and its metabolites.

Erysolin and its two metabolites which were found in blood, ERY-GSH and ERY-NAC, were synthesized by alkylation, amination, isothiocyanation and oxidation reactions from 1-bromo-4-chlorobutane and sodium methyl mercaptide. The reaction temperature, time, feed ratios and purification method were also optimized. The synthesis method was simple, green, safe and low-cost. Erysolin, ERY-GSH and ERY-NAC showed good antitumor activities against MCF-7, HeLa, HepG2, A549 and SW480 cells, which suggested that the antitumor mechanism of erysolin can also be clarified from its metabolites in addition to itself.

Palladin promotes cancer stem cell-like properties in lung cancer by activating Wnt/Β-Catenin signaling.

BACKGROUND: Cancer stem cells (CSCs) are responsible for drug resistance, cancer relapse, and metastasis. Here, we report the first analysis of Palladin expression and its impacts on stem cell-like properties in lung cancer. METHODS: Tissue microarrays were used to investigate Palladin expression and its association with prognosis. Immunofluorescence (IF), flow fluorescence assay, and Western blot were performed to detect Palladin expression in 6 NSCLC cell lines. Cell phenotypes and drug resistance were evaluated. Xenograft models were constructed to confirm the role of Palladin in vivo. RESULTS: By using the tissue microarrays, Palladin was identified to be highly expressed in the cytoplasm, specifically in the cytomembrane of NSCLC, and its high expression is associated with poor prognosis. Palladin is widely expressed and enriched in the sphere cells. The in vitro and in vivo studies showed that Palladin promoted stem cell-like properties, including cell viability, invasion, migration, self-renewal abilities, taxol resistance, and tumorigenicity. Western blot revealed that Palladin promoted the accumulation of ß-catenin and activated Wnt/ß-catenin signaling. Tissue microarrays analysis further confirmed the positive correlation between Palladin and ß-catenin. Wnt/ß-catenin pathway inhibitor blocked the Palladin-induced enhancement of sphere-forming. CONCLUSIONS: Palladin might act as an oncogene by promoting CSCs-like properties and tumorigenicity of NSCLC cells via the Wnt/ß-catenin signaling pathway. Besides, Palladin was identified to have the potential as a cell surface marker for LCSCs identification. These findings provide a possible target for developing putative agents targeted to LCSCs.

Triptolide Induces Liver Injury by Regulating Macrophage Recruitment and Polarization via the Nrf2 Signaling Pathway.

Triptolide (TP) has limited usage in clinical practice due to its side effects and toxicity, especially liver injury. Hepatic macrophages, key player of liver innate immunity, were found to be recruited and activated by TP in our previous study. The nuclear factor-erythroid-2-related factor 2 (Nrf2) pathway exerts a protective role in TP-induced liver damage, but its effect on the functions of hepatic macrophage has not been elucidated. Here, we determined whether TP can regulate the recruitment and polarization of hepatic macrophages by inhibiting Nrf2 signaling cascade. Our results demonstrated that TP inhibited the Nrf2 signaling pathway in hepatic macrophages. The changes in hepatic macrophages were responsible for the increased susceptibility toward inflammatory stimuli, and hence, TP pretreatment could induce severe liver damage upon the stimulation of a nontoxic dose of lipopolysaccharides. In addition, the Nrf2 agonist protected macrophages from TP-induced toxicity and Nrf2 deficiency significantly aggravated liver injury by enhancing the recruitment and M1 polarization of hepatic macrophages. This study suggests that Nrf2 pathway-mediated hepatic macrophage polarization plays an essential role in TP-induced liver damage, which can serve as a potential therapeutic target for preventing hepatotoxicity induced by TP.

Prophylactic Effect of Simultaneous Placement of Mesh on Incidence of Parastomal Hernia After Miles' Surgical Resection of Colorectal Cancer: A Prospective Study.

INTRODUCTION: To assess the prophylactic effect of simultaneous placement of mesh and the incidence of parastomal hernia (PSH) after abdominoperineal resection of rectal cancer. METHODS: This study included real-world data of 56 surgically resected patients with colorectal cancer who were consecutively assigned to two groups: control (no mesh, n = 32) and experimental (received mesh, n = 24). An artificial patch was placed under the tunica vaginalis of rectus abdominis for patients in the experimental group, whereas those in the control group received routine sigmoidostomy. The median follow-up time was >20 mo. The difference in hazards function was analyzed by cox regression analysis. The Kaplan-Meir analysis was used to determine the survival curves. A P value of <0.05 was considered as significant. RESULTS: The postoperative incidence rate of PSH was lower in the experimental (41.7%) group than in the control group (71.9%; P = 0.045). The PSH postoperative time in the experimental group was significantly delayed compared to the control group (48 mo versus 10 mo; P < 0.001). The risk of progression from H1 to H2 was less in the experimental group compared to the control group (49.28% versus 60.86%; P = 0.14). CONCLUSIONS: Prophylactic mesh placement significantly prolonged postoperative time for the recurrence of PSH. The incidence of recurrence of H2 (severe PSH) requiring secondary surgical repair was also reduced.

Eustachian tube teratoma: A case report.

BACKGROUND: Mature teratoma composed of all three basic germ cell layers of the head and neck is a rare disease. Teratomas involving the temporal bone are particularly scarce. CASE SUMMARY: A 48-year-old male patient with a history of chronic otitis of the left ear from infancy, for which he had been operated on twice, was referred to our hospital for chronic otitis, cholesteatoma and a middle ear mass. Computed tomography (CT) scan and magnetic resonance imaging (MRI) revealed a eustachian tube teratoma, in which the anterior lower part and posterior upper part were connected by a thin membranaceous tissue. The mass was removed completely under general anesthesia by mastoidectomy. As of last follow-up (2 years post-surgery), the disease had not relapsed. CONCLUSION: Pre-operative CT and MRI are necessary for eustachian tube teratoma. Complete surgical resection provided excellent prognosis.

Mechanism of GLP-1 Receptor Agonists-Mediated Attenuation of Palmitic Acid-Induced Lipotoxicity in L6 Myoblasts.

Methods: Cells were divided into 5 groups-control, high-fat, 10 nmol/L LR + 0.6 mmol/L palmitic acid (PA) (10LR), 100 nmol/L LR + 0.6 mmol/L PA (100LR), and 1000 nmol/L LR + 0.6 mmol/L PA (1000LR). CCK-8 method to detect cell viability, GPO-PAP enzymatic method to detect intracellular triglyceride content, and reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) and western blotting methods to detect fatty acid translocase CD36 (FAT/CD36) and fatty acid binding protein 4 (FABP4) in L6 cells, glucose-regulated protein 78 (GRP78), glucose transporter 4 (GLUT4) expression at the mRNA and protein levels, respectively, were performed. Results: We found that after PA intervention for 24 h, the cell viability decreased significantly; the cell viability of the LR group was higher than that of the high-fat group (P < 0.01). After PA intervention, compared with those in the high-fat group, GRP-78, FAT/CD36, FABP4 mRNA ((4.36 ± 0.32 vs. 8.15 ± 0.35); (1.00 ± 0.04 vs. 2.46 ± 0.08); (2.88 ± 0.55 vs. 8.29 ± 0.52), P < 0.01) and protein ((3338.13 ± 333.15 vs. 4963.98 ± 277.29); (1978.85 ± 124.24 vs. 2676.07 ± 100.64); (3372.00 ± 219.84 vs. 6083.20 ± 284.70), both P < 0.01) expression decreased in the LR group. The expression levels of GLUT4 mRNA ((0.75 ± 0.04 vs. 0.34 ± 0.03), P < 0.01) and protein ((3443.71 ± 191.89 vs. 2137.79 ± 118.75), P < 0.01) increased. Conclusion: Therefore, we conclude that LR can reverse PA-induced cell inactivation and lipid deposition, which may be related to the change in GRP-78, FAT/CD36, FABP4, GLUT4, and other factors.

MYH9 is crucial for stem cell-like properties in non-small cell lung cancer by activating mTOR signaling.

The fatality rate of non-small cell lung cancer (NSCLC) has been high due to the existence of cancer stem cells (CSCs). Non-muscle myosin heavy chain 9 (MYH9) can promote the progression of various tumors, but its effect on the stem cell-like characteristics of lung cancer cells (LCCs) has not been clarified. Our research found that the stemness characteristics of LCCs were significantly enhanced by the overexpression of MYH9, and the knockout of MYH9 had the opposite effects. The in vivo with inhibitor blebbistatin further confirmed the effect of MYH9 on the stem cell-like behavior of LCCs. Furthermore, western blotting showed that the expression level of CSCs markers (CD44, SOX2, Nanog, CD133, and OCT4) was also regulated by MYH9. Mechanistic studies have shown that MYH9 regulates stem cell-like features of LCCs by regulating the mTOR signaling pathway, which was supported by sphere formation experiments after LCCs were treated with inhibitors Rapamycin and CHIR-99021. Importantly, high expression of MYH9 in lung cancer is positively correlated with poor clinical prognosis and is an independent risk factor for patients with NSCLC.

Cohort Profile: Chinese Cervical Cancer Clinical Study.

Cervical cancer is the fourth most common cancer worldwide, but its incidence varies greatly in different countries. Regardless of incidence or mortality, the burden of cervical cancer in China accounts for approximately 18% of the global burden. The Chinese Cervical Cancer Clinical Study is a hospital-based multicenter open cohort. The major aims of this study include (i) to explore the associations of therapeutic strategies with complications as well as mid- and long-term clinical outcomes; (ii) to widely assess the factors which may have an influence on the prognosis of cervical cancer and then guide the treatment options, and to estimate prognosis using a prediction model for precise post-treatment care and follow-up; (iii) to develop a knowledge base of cervical clinical auxiliary diagnosis and prognosis prediction using artificial intelligence and machine learning approaches; and (iv) to roughly map the burden of cervical cancer in different districts and monitoring the trend in incidence of cervical cancer to potentially inform prevention and control strategies. Patients eligible for inclusion were those diagnosed with cervical cancer, whether during an outpatient visit or hospital admission, at 47 different types of medical institutions in 19 cities of 11 provinces across mainland China between 2004 and 2018. In a total, 63 926 patients with cervical cancer were enrolled in the cohort. Since the project inception, a large number of standardized variables have been collected, including epidemiological characteristics, cervical cancer-related symptoms, physical examination results, laboratory testing results, imaging reports, tumor biomarkers, tumor staging, tumor characteristics, comorbidities, co-infections, treatment and short-term complications. Follow-up was performed at least once every 6 months within the first 5 years after receiving treatment and then annually thereafter. At present, we are developing a cervical cancer imaging database containing Dicom files with data of computed tomography/magnetic resonance imaging examination. Additionally, we are also collecting original pathological specimens of patients with cervical cancer. Potential collaborators are welcomed to contact the corresponding authors, and anyone can submit at least one specific study proposal describing the background, objectives and methods of the study.

HA of H1N1 enhanced the expression of ICAM-1 and IL-6 in HUVECs and pathological injury in the lungs in mice.

OBJECTIVE: Both COVID-19 and influenza are viral respiratory tract infections and the epidemics of viral respiratory tract infections remain highly prevalent with lethal consequences in susceptible individuals. Expression of ICAM-1 on vascular endothelium recruits leukocytes which initiates inflammation. IL-6 induces ICAM-1. Both ICAM-1 and IL-6 can be enhanced in influenza virus infection and COVID-19 patients. Besides initiation of virus entry host cells, whether HA alone, instead of whole virus, of influenza has the effects on expression of ICAM-1 and IL-6 in vascular endothelium with injury in the lungs, remains to be demonstrated. METHODS: RT-qPCR and Western blot as well as histopathologic examination were used to examine mRNA and protein of ICAM-1 and IL-6 as well as pathological injury in the lung tissues, respectively. RESULTS: After incubation of the Human Umbilical Vein Endothelial Cells (HUVECs) with HA of H1N1 for 24 h, the mRNA and protein of ICAM-1 and IL-6 in HUVECs were increased in group of 5 µg/ml concentration with statistical significance (p < 0.05). Pathological injury in lung tissues of the mice was shown 12 h after tail intravenous injection with 100 µl of HA (50 µg/ml and 100 µg/ml in normal saline), including widened alveolar spaces with angiotelectasis in alveolar wall, alveolar luminal and interstitial inflammatory infiltrates, alveolar luminal erythrocyte effusion. CONCLUSIONS: HA alone, instead of whole H1N1 virus, induced more expression of ICAM-1 and IL-6, two molecules involving in pathological and inflammatory responses, in HUVECs and pathological injury in lung tissues of the mice. This knowledge provides a new HA-targeted potential direction for prevention and treatment of disease related to H1N1 infection.

Intraoperative Oxygen Concentration and Postoperative Delirium After Laparoscopic Gastric and Colorectal Malignancies Surgery: A Randomized, Double-Blind, Controlled Trial.

PURPOSE: Postoperative delirium (POD) is common in elderly patients undergoing laparoscopic surgery for gastric and colorectal malignancies. POD may be affected by different fraction of inspired oxygen (FiO2). The purpose of this study was to compare the effects of different FiO2 on POD. PATIENTS AND METHODS: A randomized, double-blind controlled trial was performed in Qingdao Municipal Hospital Affiliated to Qingdao University. A total of 662 patients aged 65 to 85 years old underwent isolated laparoscopic radical gastrectomy, radical resection of colon cancer, or radical resection of rectal cancer only. A random number table method was used to divide the patients into two groups: 40% FiO2 (group A) and 80% FiO2 (group B). The primary endpoint was the incidence of POD, which was assessed by the Confusion Assessment Method (CAM) twice daily during the first 7 postoperative days, and POD severity was measured by the Memorial Delirium Assessment Scale (MDAS). The secondary endpoints were the intraoperative regional cerebral oxygen saturation (rSO2), Bispectral (BIS) index, invasive arterial blood pressure (IABP), oxygen saturation (SpO2), end-tidal carbon dioxide partial pressure (PETCO2), the number of atelectasis cases and visual analogue scale (VAS) scores on days 1-7 after surgery. RESULTS: The incidence of POD was 19.37% (122/630), including 20.38% (64/314) in group A and 18.35% (58/316) in group B. No statistical significance was found in the incidence of POD between the two groups (P > 0.05); compared with group B, SpO2, rSO2 and PaO2 decreased at T2 to T4 time point (P < 0.01), and the incidence of postoperative atelectasis decreased (P < 0.05) in group A. CONCLUSION: The incidence of POD was not significantly affected by different FiO2 and the incidence of postoperative atelectasis was decreased at low FiO2.

BCAT1 Activates PI3K/AKT/mTOR Pathway and Contributes to the Angiogenesis and Tumorigenicity of Gastric Cancer.

BACKGROUND: Focusing on antiangiogenesis may provide promising choices for treatment of gastric cancer (GC). This study aimed to investigate the mechanistic role of BCAT1 in the pathogenesis of GC, particularly in angiogenesis. METHODS: Bioinformatics and clinical samples analysis were used to investigate the expression and potential mechanism of BCAT1 in GC. BGC823 cells with BCAT1 overexpression or silencing were induced by lentiviral transduction. Cell phenotypes and angiogenesis were evaluated. The relevant proteins were quantized by Western blotting, immunohistochemistry, or immunofluorescence. Xenograft models were constructed to confirm the role of BCAT1 in vivo. RESULTS: BCAT1 was overexpressed in GC patients and associated with lower survival. BCAT1 expression was correlated with proliferation-, invasion-, or angiogenesis-related markers expression and pathways. Silencing BCAT1 expression suppressed cell viability, colony formation, cycle progression, invasion, and angiogenesis of BGC823 cells, as well as the tumor growth of xenograft models, whereas overexpressing BCAT1 had the opposite results both in vitro and in vivo. Bioinformatics analysis and Western blotting demonstrated that BCAT1 activated the PI3K/AKT/mTOR pathway. The addition of LY294002 reversed the tumor growth induced by BCAT1 overexpression, further verifying this mechanism. CONCLUSION: BCAT1 might act as an oncogene by facilitating proliferation, invasion, and angiogenesis through activation of the PI3K/AKT/mTOR pathway. This finding could aid the optimization of antiangiogenesis strategies.

Altered integrity of hepatocyte tight junctions in rats with triptolide-induced cholestasis.

Triptolide (TP), an active component of Tripterygium wilfordiiHook. f. (TWHF), has been widely used for centuries as a traditional Chinese medicine. However, the clinical application of TP has been restricted due to multitarget toxicity, such as hepatotoxicity. In this study, 28 days of oral TP administration (100, 200, or 400 µg·kg-1·d-1) induced the occurrence of cholestasis in female Wistar rats, as evidenced by increased serum levels of γ-glutamyl transpeptidase (γ-GGT), alkaline phosphatase (ALP) and hepatic total bile acids (TBAs). In addition, the heptocyte polarity associated with the strcture of tight junctions (TJs) was disrupted in both rats and sandwich-cultured primary hepatocytes. Immunoblotting revealed decreased expression of the TJ-associated proteins occludin, claudin-1, and zonula occludens protein (ZO-1), and downregulated mRNA levels of these TJs was also detected by real-time PCR. An immunofluorescence analysis showed abnormal subcellular localization of occludin, claudin-1 and ZO-1, which was also confirmed by transmission electron microscopy. Moreover, the concentration of FITC-dextran, a marker of paracellular penetration, was found to increase rapidly in bile increased rapidly (within 6 minutes) after treatment with TP, which indicated the functional impairment of TJs. Taken together, these results suggest that the administration of TP for 28 consecutive days to rats could induce cholestatic injury in the liver, and the increased paracellular permeability might play an important role in these pathological changes.

Alpha-enolase (ENO1), identified as an antigen to monoclonal antibody 12C7, promotes the self-renewal and malignant phenotype of lung cancer stem cells by AMPK/mTOR pathway.

BACKGROUND: Tumor-associated antigens (TAAs) can be targeted in cancer therapy. We previously identified a monoclonal antibody (mAb) 12C7, which presented anti-tumor activity in lung cancer stem cells (LCSCs). Here, we aimed to identify the target antigen for 12C7 and confirm its role in LCSCs. METHODS: Immunofluorescence was used for antigen localization. After targeted antigen purification by electrophoresis and immunoblot, the antigen was identified by LC-MALDI-TOF/TOF mass spectrometry, immunofluorescence, and immunoprecipitation. The overexpression or silence of ENO1 was induced by lentiviral transduction. Self-renewal, growth, and invasion of LCSCs were evaluated by sphere formation, colony formation, and invasion assay, respectively. High-throughput transcriptome sequencing (RNA-seq) and bioinformatics analysis were performed to analyze downstream targets and pathways of targeted antigen. RESULTS: Targeted antigen showed a surface antigen expression pattern, and the 43-55 kDa protein band was identified as α-enolase (ENO1). Self-renewal, growth, and invasion abilities of LCSCs were remarkably inhibited by ENO1 downregulation, while enhanced by ENO1 upregulation. RNA-seq and bioinformatics analysis eventually screened 4 self-renewal-related and 6 invasion-related differentially expressed genes. GSEA analysis and qRT-PCR verified that ENO1 regulated self-renewal, invasion-related genes, and pathways. KEGG pathway analysis and immunoblot demonstrated that ENO1 inactivated AMPK pathway and activated mTOR pathway in LCSCs. CONCLUSIONS: ENO1 is identified as a targeted antigen of mAb 12C7 and plays a pivotal role in facilitating self-renewal, growth, and invasion of LCSCs. These findings provide a potent therapeutic target for the stem cell therapy for lung cancer and have potential to improve the anti-tumor activity of 12C7.

Catalpol counteracts the pathology in a mouse model of Duchenne muscular dystrophy by inhibiting the TGF-ß1/TAK1 signaling pathway.

Duchenne muscular dystrophy (DMD) is a progressive neuromuscular disease caused by a mutation in the gene encoding the dystrophin protein. Catalpol is an iridoid glycoside found in Chinese herbs with anti-inflammatory, anti-oxidant, anti-apoptotic, and hypoglycemic activities that can protect against muscle wasting. In the present study we investigated the effects of catalpol on DMD. Aged Dystrophin-deficient (mdx) mice (12 months old) were treated with catalpol (100, 200 mg·kg-1·d-1, ig) for 6 weeks. At the end of the experiment, the mice were sacrificed, and gastrocnemius (GAS), tibialis anterior (TA), extensor digitorum longus (EDL), soleus (SOL) muscles were collected. We found that catalpol administration dose-dependently increased stride length and decreased stride width in Gait test. Wire grip test showed that the time of wire grip and grip strength were increased. We found that catalpol administration dose-dependently alleviated skeletal muscle damage, evidenced by reduced plasma CK and LDH activity as well as increased the weight of skeletal muscles. Catalpol administration had no effect on dystrophin expression, but exerted anti-inflammatory effects. Furthermore, catalpol administration dose-dependently decreased tibialis anterior (TA) muscle fibrosis, and inhibited the expression of TGF-ß1, TAK1 and α-SMA. In primary myoblasts from mdx mice, knockdown of TAK1 abolished the inhibitory effects of catalpol on the expression levels of TGF-ß1 and α-SMA. In conclusion, catalpol can restore skeletal muscle strength and alleviate skeletal muscle damage in aged mdx mice, thus may provide a novel therapy for DMD. Catalpol attenuates muscle fibrosis by inhibiting the TGF-ß1/TAK1 signaling pathway.

Enolase 1 regulates stem cell-like properties in gastric cancer cells by stimulating glycolysis.

Recent studies have demonstrated that gastric cancer stem cells (CSCs) are a rare sub-group of gastric cancer (GC) cells and have an important role in promoting the tumor growth and progression of GC. In the present study, we demonstrated that the glycolytic enzyme Enolase 1 (ENO1) was involved in the regulation of the stem cell-like characteristics of GC cells, as compared to the parental cell lines PAMC-82 and SNU16, the expression of ENO1 in spheroids markedly increased. We then observed that ENO1 could enhance stem cell-like characteristics, including self-renewal capacity, cell invasion and migration, chemoresistance, and even the tumorigenicity of GC cells. ENO1 is known as an enzyme that is involved in glycolysis, but our results showed that ENO1 could markedly promote the glycolytic activity of cells. Furthermore, inhibiting glycolysis activity using 2-deoxy-D-glucose treatment significantly reduced the stemness of GC cells. Therefore, ENO1 could improve the stemness of CSCs by enhancing the cells' glycolysis. Subsequently, to further confirm our results, we found that the inhibition of ENO1 using AP-III-a4 (ENOblock) could reduce the stemness of GC cells to a similar extent as the knockdown of ENO1 by shRNA. Finally, increased expression of ENO1 was related to poor prognosis in GC patients. Taken together, our results demonstrated that ENO1 is a significant biomarker associated with the stemness of GC cells.

Organic anion transporter 1 and 3 contribute to traditional Chinese medicine-induced nephrotoxicity.

With the internationally growing popularity of traditional Chinese medicine (TCM), TCM-induced nephropathy has attracted public attention. Minimizing this toxicity is an important issue for future research. Typical nephrotoxic TCM drugs such as Aristolochic acid, Tripterygium wilfordii Hook. f, Rheum officinale Baill, and cinnabar mainly damage renal proximal tubules or cause interstitial nephritis. Transporters in renal proximal tubule are believed to be critical in the disposition of xenobiotics. In this review, we provide information on the alteration of renal transporters by nephrotoxic TCMs, which may be helpful for understanding the nephrotoxic mechanism of TCMs and reducing adverse effects. Studies have proven that when administering nephrotoxic TCMs, the expression or function of renal transporters is altered, especially organic anion transporter 1 and 3. The alteration of these transporters may enhance the accumulation of toxic drugs or the dysfunction of endogenous toxins and subsequently sensitize the kidney to injury. Transporters-related drug combination and clinical biomarkers supervision to avoid the risk of future toxicity are proposed.

Emergence and spread patterns of antibiotic resistance genes during two different aerobic granular sludge cultivation processes.

The prevalence and accumulation of antibiotic resistance genes (ARGs) were frequently detected in biological wastewater treatment processes, which might cause potential health crisis to human. In present study, the fates of ARGs during two different aerobic granular sludge (AGS) cultivation processes were investigated. The results showed that traditional AGS (T-AGS) cultivation process and enhanced AGS (E-AGS) cultivation process had significant differences (P < 0.005) in ARGs shift patterns. E-AGS process had higher average relative abundance (0.280 ± 0.079) of ARGs than T-AGS process (0.130 ± 0.041), while the intensity of ARGs enrichment during E-AGS (1.52-5.29 fold) was lower than T-AGS (3.79-75.31 fold) process. TnpA and intI1 as two different types of mobile genetic elements (MGEs) carrying ARGs, were observed to contribute significantly to the horizontal gene transfer (HGT) during T-AGS (r = 0.902, P < 0.050) and E-AGS (r = 0.823, P < 0.001) processes, respectively. Higher HGT level took place and more possible potential hosts (25 hosts) harboring ARGs were detected during E-AGS process comparing with T-AGS process (17 hosts). Meanwhile, over large AGS might increase the propagation of several antibiotic deactivation ARGs, so it was not advised. Overall, whether during T-AGS or during E-AGS process which was applied in a pilot-scale sequencing batch reactor treating municipal wastewater, the accumulation and spread of ARGs were inevitable. It should be valued that some suitable pre-treatments of seed sludge should be executed, meanwhile, advanced treatment for removing of ARGs in AGS should be conducted to maintain the relative abundances of ARGs at relatively low level.