A new evaluation system for drug-microbiota interactions.

The drug response phenotype is determined by a combination of genetic and environmental factors. The high clinical conversion failure rate of gene-targeted drugs might be attributed to the lack of emphasis on environmental factors and the inherent individual variability in drug response (IVDR). Current evidence suggests that environmental variables, rather than the disease itself, are the primary determinants of both gut microbiota composition and drug metabolism. Additionally, individual differences in gut microbiota create a unique metabolic environment that influences the in vivo processes underlying drug absorption, distribution, metabolism, and excretion (ADME). Here, we discuss how gut microbiota, shaped by both genetic and environmental factors, affects the host's ADME microenvironment within a new evaluation system for drug-microbiota interactions. Furthermore, we propose a new top-down research approach to investigate the intricate nature of drug-microbiota interactions in vivo. This approach utilizes germ-free animal models, providing foundation for the development of a new evaluation system for drug-microbiota interactions.

Leaching characteristics and pollution risk assessment of potentially harmful elements from coal gangue exposed to weathering for different periods of time.

To explore the leaching behavior and potential degree of pollution that can result from the backfilling of goafs with different types of coal gangue (CG), fresh CG from the Hongqi Coal Mine goaf and surface CG (weathered for 1 year) were selected as the research objects in this study. A series of leaching experiments were carried out using the Ordovician limestone karst waters of the mining areas as the soaking solution. A comparative study on the dissolution characteristics of Fe3+, Mn2+, and SO42- and on the traditional water quality parameters of the two types of CG was conducted. The results showed that the soaked, weathered CG displayed a higher ion dissolution value than fresh CG. The ratio of each ion was as follows: Fe3+ was 1, Mn2+ was 2.86 ~ 68.18, and SO42- was 1.34 ~ 2.09. Over time, the ion concentration of water samples that initially contained high ion concentration values showed a decreasing trend after CG was soaked in these waters, but the values were still in the range of high ion release concentrations. The pH and oxidation‒reduction potential (ORP) values of the leachate of both CG types indicated that the leachates were weakly alkaline and weakly oxidizing, and the overall change in total dissolved solids (TDS) was small and consistent with the SO42- trend. SO42- in the leachate of the weathered CG showed a more significant correlation with the pH and TDS of the soaking solution, and it was the major pollutant. According to the geoaccumulation index evaluation, weathered CG had higher pollution potential than fresh CG. Fe3+ presented a slight and moderate risk for contamination.

Gut microbial characteristical comparison reveals potential anti-aging function of Dubosiella newyorkensis in mice.

Introduction: Previous study has indicated Dubosiella newyorkensis may act as a potential probiotic in age-related diseases. However, its detailed role in aging has not yet been promulgated. This study aimed to explore the potential anti-aging role of Dubosiella newyorkensis by comparing the anti-aging effect of resveratrol in young and old mice. Method: Measurement of intestinal aging-related factors in colon and serum, and vascular endothelial function-related factors in serum were performed by enzyme-linked immunosorbent assay (ELISA). Gut microbial analysis of intestinal contents were identified by 16S rRNA gene sequencing. Results: The effect of Dubosiella newyorkensis on reducing malondialdehyde (MDA) and increasing superoxide dismutase (SOD) in aged mice were greater than that of resveratrol. While the effect of Dubosiella newyorkensis on nitric oxide (NO) level was less than that of resveratrol, the reduction of vascular endothelial growth factor (VEGF) and pentosidine (PTD) was better than that of resveratrol in young mice. In young mice, Dubosiella newyorkensis promoted an increase in the beneficial genus Lactobacillus, Bifidobacterium and Ileibacterium less effectively as compared with resveratrol treatment. In aged mice, Dubosiella newyorkensis promoted the increase of Bifidobacterium, Ileibacterium less effectively than resveratrol, and promoted the increase of Akkermansia, Staphylococcus, Verrucomicrobiota expression better as compared with resveratrol treatment. Both young and old mice showed the same results for the remaining markers, including changes in gut microbial composition and predictions of function. Conclusion: Dubosiella newyorkensis has similar anti-aging functions with resveratrol. Dubosiella newyorkensis may even be more effective than resveratrol in reducing oxidative stress, improving vascular endothelial function, and redistributing gut microbiota. The research provides an innovative strategy of Dubosiella newyorkensis to improve aging.

Gut microbial evidence chain in high-salt diet exacerbates intestinal aging process.

Although excessive salt consumption appears to hasten intestinal aging and increases susceptibility to cardiovascular disease, the molecular mechanism is unknown. In this study, mutual validation of high salt (HS) and aging fecal microbiota transplantation (FMT) in C56BL/6 mice was used to clarify the molecular mechanism by which excessive salt consumption causes intestinal aging. Firstly, we observed HS causes vascular endothelial damage and can accelerate intestinal aging associated with decreased colon and serum expression of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and increased malondialdehyde (MDA); after transplantation with HS fecal microbiota in mice, vascular endothelial damage and intestinal aging can also occur. Secondly, we also found intestinal aging and vascular endothelial damage in older mice aged 14 months; and after transplantation of the older mice fecal microbiota, the same effect was observed in mice aged 6-8 weeks. Meanwhile, HS and aging significantly changed gut microbial diversity and composition, which was transferable by FMT. Eventually, based on the core genera both in HS and the aging gut microbiota network, a machine learning model was constructed which could predict HS susceptibility to intestinal aging. Further investigation revealed that the process of HS-related intestinal aging was highly linked to the signal transduction mediated by various bacteria. In conclusion, the present study provides an experimental basis of potential microbial evidence in the process of HS related intestinal aging. Even, avoiding excessive salt consumption and actively intervening in gut microbiota alteration may assist to delay the aging state that drives HS-related intestinal aging in clinical practice.

Correction: Systemic Delivery of MicroRNA-101 Potently Inhibits Hepatocellular Carcinoma In Vivo by Repressing Multiple Targets.

[This corrects the article DOI: 10.1371/journal.pgen.1004873.].

Network and 16S rRNA Sequencing-Combined Approach Provides Insightal Evidence of Vitamin K2 for Salt-Sensitive Hypertension.

Vitamin K2 (VK2), found to act to treat hypertension, has been widely used in the food and pharmaceutical industries nowadays. However, the potential targets and molecular mechanisms of VK2 for salt-sensitive hypertension have not been fully investigated. Therefore, the study aimed to investigate the potential molecular mechanisms of VK2 for salt-sensitive hypertension using network pharmacology and 16S rRNA sequencing strategy. The network pharmacology-based findings from KEGG enrichment analysis revealed that VK2-treated salt-sensitive hypertension was mechanically associated with the complement and coagulation cascades, calcium signaling pathway, renin-angiotensin system, etc. A total of 29 different bacteria in an animal experiment after VK2 supplementation were screened and functionally enriched using PICRUSt2. Additionally, 10 signaling pathways were identified in which the renin-angiotensin system was found to be the potential molecular mechanisms with the greatest change in multiple and statistical significance. Moreover, the results of the renin-angiotensin system-related protein expression exhibited VK2-inhibited renin-angiotensin system in salt-induced hypertensive mice, which significantly verified the previous biological and functional prediction analysis. Finally, spearman correlation analysis showed the different bacteria such as Dubosiella, Ileibacterium, etc., had a positive or negative correlation with renin-angiotensin system-related proteins in salt-induced mice. In conclusion, the potential molecular mechanisms of VK2 for salt-sensitive hypertension may be beneficially achieved by the specific inhibition of the renin-angiotensin system, contributing to the development for a new preventive strategy of salt-sensitive hypertension.

Gut Microbiota-Related Evidence Provides New Insights Into the Association Between Activating Transcription Factor 4 and Development of Salt-Induced Hypertension in Mice.

Activating transcription factor 4 (ATF4), which regulates genes associated with endoplasmic reticulum stress, apoptosis, autophagy, the gut microbiome, and metabolism, has been implicated in many diseases. However, its mechanistic role in hypertension remains unclear. In the present study, we investigated its role in salt-sensitive hypertensive mice. Wild-type (WT) C57BL/6J mice were used to establish Atf4 knockout (KO) and overexpression mice using CRISPR-Cas9 and lentiviral overexpression vectors. Then, fecal microbiota transplantation (FMT) from Atf4 ± mice and vitamin K2 (VK2) supplementation were separately carried out in high-salt-diet (8% NaCl)-induced mice for 4 weeks. We found that Atf4 KO inhibited and Atf4 overexpression enhanced the increase in blood pressure and endothelial dysfunction induced by high salt intake in mice, while regulating the gut microbiota composition and VK2 expression. It was further verified that ATF4 is involved in the regulation of salt-sensitive hypertension and vascular endothelial function, which is achieved through association with gut microbiota and may be related to VK2 and different bacteria such as Dubosiella. In addition, we found that VK2 supplementation prevents the development of salt-sensitive hypertension and maintains vascular endothelial function; moreover, VK2 supplementation increases the abundance of intestinal Dubosiella and downregulates the relative expression of Atf4 in the thoracic aorta of mice. We conclude that ATF4 plays an important role in regulating gut microbiota and VK2 production, providing new insights into the association between ATF4 and development of salt-induced hypertension in mice, meanwhile contributing to the development for a new preventive strategy of hypertension.

Verification of Resveratrol Inhibits Intestinal Aging by Downregulating ATF4/Chop/Bcl-2/Bax Signaling Pathway: Based on Network Pharmacology and Animal Experiment.

Resveratrol is one of the most well-known drugs used in the treatment of aging. However, the potential mechanisms of resveratrol on intestinal aging have not yet been fully investigated. Herein, we aimed to further explore the pharmacological mechanisms of resveratrol as a therapy for intestinal aging. We performed network construction and enrichment analysis via network pharmacology. Then a further animal experimental validation containing 20 female C57BL/6J (wild type, WT) and 16 female ATF4+/- (knock down, KD) naturally aging mice and oral supplementary resveratrol (44 mg/kg/day) for 30 days were conducted. The expression of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT), linear alkylethoxylate (AE), and malondialdehyde (MDA) were measured by ELISA, the observation of pathological changes and apoptosis in intestinal tissue were performed by HE, PAS, and TUNEL staining, the ATF4/Chop/Bcl-2/Bax signaling pathway-related proteins and mRNAs expression were measured by western blotting and real-time PCR. The network pharmacology showed 132 targets of resveratrol on aging. The enrichment analysis showed resveratrol antiaging involved mainly included protein heterodimerization activity, apoptosis, etc. Then ATF4/Chop/Bcl-2/Bax signaling pathway in biological process of apoptosis was selected to verify the potential mechanisms. Animal studies showed resveratrol upregulated the relative expression of SOD, GSH-Px, CAT, AE, whereas it downregulated the relative expression of MDA in intestine compared with the control group. There was also higher relative expression of SOD, GSH-Px, CAT, AE, and lower relative expression of MDA in KD mice than that in WT mice. Moreover, there was higher relative expression of SOD, GSH-Px, CAT, AE, and lower relative expression of MDA in KD mice than that in WT mice after resveratrol treatment. Decreased ATF4, Chop, Bax but increased Bcl-2 proteins and mRNAs expression were determined after resveratrol treatment compared with the control group; lower ATF4, Chop, Bax but higher Bcl-2 proteins and mRNAs expression were found in KD mice than that in WT mice. Additionally, lower relative proteins and mRNAs expression of ATF4, Chop, Bax and higher relative expression of Bcl-2 in KD mice than that in WT mice after resveratrol treatment. These findings demonstrated that resveratrol substantially inhibited intestinal aging via downregulating ATF4/Chop/Bcl-2/Bax signaling pathway.

Bacterial association and comparison between lung and intestine in rats.

The association between lung and intestine has already been reported, but the differences in community structures or functions between lung and intestine bacteria yet need to explore. To explore the differences in community structures or functions, the lung tissues and fecal contents in rats were collected and analyzed through 16S rRNA sequencing. It was found that intestine bacteria was more abundant and diverse than lung bacteria. In intestine bacteria, Firmicutes and Bacteroides were identified as major phyla while Lactobacillus was among the most abundant genus. However, in lung the major identified phylum was Proteobacteria and genus Pseudomonas was most prominent genus. On the other hand, in contrast the lung bacteria was more concentrated in cytoskeleton and function in energy production and conversion. While, intestine bacteria were enriched in RNA processing, modification chromatin structure, dynamics and amino acid metabolism. The study provides the basis for understanding the relationships between lung and intestine bacteria.

Network Pharmacology Identifies the Mechanisms of Action of TaohongSiwu Decoction Against Essential Hypertension.

BACKGROUND TaohongSiwu decoction (THSWT), a traditional herbal formula, has been used to treat cardiovascular and cerebrovascular diseases such as essential hypertension (EH) in China. However, the pharmacological mechanism is not clear. To investigate the mechanisms of THSWT in the treatment of EH, we performed compounds, targets prediction and network analysis using a network pharmacology method. MATERIAL AND METHODS We selected chemical constituents and targets of THSWT according to TCMSP and UniProtKB databases and collected therapeutic targets on EH from Online Mendelian Inheritance in Man (OMIM), Drugbank and DisGeNET databases. The protein-protein interaction (PPI) was analyzed by using String database. Then network was constructed by using Cytoscape_v3.7.1, and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment was performed by using Database for Annotation, Visualization and Integrated Discovery (DAVID) software. RESULTS The results of our network pharmacology research showed that the THSWT, composed of 6 Chinese herbs, contained 15 compounds, and 23 genes regulated the main signaling pathways related to EH. Moreover, the PPI network based on targets of THSWT on EH revealed the interaction relationship between targets. These core compounds were 6 of the 15 disease-related compounds in the network, kaempferol, quercetin, luteolin, Myricanone, beta-sitosterol, baicalein, and the core genes contained ADRB2, CALM1, HMOX1, JUN, PPARG, and VEGFA, which were regulated by more than 3 compounds and significantly associated with Calcium signaling pathway, cGMP-PKG signaling pathway, cAMP signaling pathway, PI3K-Akt signaling pathway, Rap1 signaling pathway, and Ras signaling pathway. CONCLUSIONS This network pharmacological study can reveal potential mechanisms of multi-target and multi-component THSWT in the treatment of EH, provide a scientific basis for studying the mechanism.

Bacterial Diversity in Roots, Stems, and Leaves of Chinese Medicinal Plant Paris polyphylla var. yunnanensis.

The root of Paris polyphylla var. yunnanensis, a famous and endangered traditional Chinese herb, has a significant medicinal value. The aim of this study was to analyze the composition and functional characteristics of bacterial endophytes in roots, stems, and leaves of P. polyphylla var. yunnanensis. The 16S rRNA gene sequencing and functional prediction of bacterial endophytes in roots, stems, and leaves of P. polyphylla var. yunnanensis were conducted. The Chao and Shannon indices of the bacteria in roots were significantly higher than those in stems and leaves. The dominant endophyte phyla were Cyanobacteria, Proteobacteria, Bacteroidetes, Firmicutes, and Actinobacteria. The main genera detected in roots were unclassified Cyanobacteria, Rhizobium, Flavobacterium, and Sphingobium; the main genera in stems were norank_c__Cyanobacteria, Bacillus, and Pseudomonas; the main genera in leaves were norank_c__Cyanobacteria and Rhizobium. The microbiota in roots was particularly enriched in functional categories "extracellular structures" and "cytoskeleton" compared with stems and leaves (p < 0.05). Our study reveals the structural and functional characteristics of the endophytic bacteria in roots, stems, and leaves of P. polyphylla var. yunnanensis, which aids in the scientific understanding of this plant.The root of Paris polyphylla var. yunnanensis, a famous and endangered traditional Chinese herb, has a significant medicinal value. The aim of this study was to analyze the composition and functional characteristics of bacterial endophytes in roots, stems, and leaves of P. polyphylla var. yunnanensis. The 16S rRNA gene sequencing and functional prediction of bacterial endophytes in roots, stems, and leaves of P. polyphylla var. yunnanensis were conducted. The Chao and Shannon indices of the bacteria in roots were significantly higher than those in stems and leaves. The dominant endophyte phyla were Cyanobacteria, Proteobacteria, Bacteroidetes, Firmicutes, and Actinobacteria. The main genera detected in roots were unclassified Cyanobacteria, Rhizobium, Flavobacterium, and Sphingobium; the main genera in stems were norank_c__Cyanobacteria, Bacillus, and Pseudomonas; the main genera in leaves were norank_c__Cyanobacteria and Rhizobium. The microbiota in roots was particularly enriched in functional categories "extracellular structures" and "cytoskeleton" compared with stems and leaves (p < 0.05). Our study reveals the structural and functional characteristics of the endophytic bacteria in roots, stems, and leaves of P. polyphylla var. yunnanensis, which aids in the scientific understanding of this plant.

Bioinformatics analysis of endophytic bacteria related to berberine in the Chinese medicinal plant Coptis teeta Wall.

Endophytic microorganisms absorb nutrients and prevent pathogen damage, supporting healthy plant growth. However, the relationship between endophytic bacteria and berberine synthesis in the medicinal plant Coptis teeta Wall. remains unclear. Herein, we explored the community composition of endophytic bacteria related to berberine in roots, stems, and leaves of wild-type and cultivated C. teeta. Endophytic bacterial communities were analyzed by 16S rRNA sequencing, and berberine content in roots was analyzed by high-performance liquid chromatography. Proteobacteria, Actinobacteria, and Bacteroidetes were the major phyla, and Mycobacterium, Salmonella, Nocardioides, Burkholderia-Paraburkholderia, and Rhizobium were the dominant genera in root, stem, and leaf tissues. Root berberine content was positively correlated with total N, total P, total K, and available K in rhizosphere soil. In addition, root berberine content was positively correlated with Microbacterium and norank_f_7B-8, whereas soil total K was positively correlated with Microbacterium and Burkholderia-Paraburkholderia in roots. Our results demonstrated a clear correlation between dominant endophytic bacteria and berberine synthesis in C. teeta. The findings are useful for the promotion of berberine production in C. teeta via manipulation of endophytic bacteria.

Role of the PERK-eIF2α-CHOP Signaling Pathway in the Effect of Needle Knife Therapy on Knee Joint Chondrocyte Apoptosis.

Needle knife therapy, a form of acupuncture and moxibustion, has been widely used in the clinical treatment of knee osteoarthritis (KOA). However, the mechanism is not clear. Therefore, we studied the mechanisms of action of needle knife intervention on KOA in rabbits, with the PERK-eIF2α-CHOP pathway as a starting point, in order to determine the mechanism underlying knee joint chondrocyte apoptosis. Apoptosis and ultrastructural changes in the articular cartilage were examined by pathological study and transmission electron microscopy, and PERK, eIF2α, and CHOP mRNA and protein levels were detected by qRT-PCR and western blot, respectively. PERK, eIF2α, and CHOP protein levels were significantly higher in the model group than in the normal group (P < 0.01) and were considerably downregulated in the needle knife and the medicine groups compared to the model group (P < 0.01). The eIF2α, p-eIF2α, and CHOP protein levels were not significantly different between the needle knife and medicine groups. The PERK, eIF2α, and CHOP mRNA levels in the drug group were higher than those in the needle knife group (P < 0.01). Needle knife therapy can regulate PERK-eIF2α-CHOP signaling pathway, which could be one of the mechanisms by which it affects chondrocyte apoptosis in KOA rabbits.

Curcumin and Curcumol Inhibit NF-κB and TGF-ß 1/Smads Signaling Pathways in CSE-Treated RAW246.7 Cells.

E-Zhu (Curcuma zedoaria) is known as a classical traditional Chinese medicine and widely used in the treatment of cancers, cardiovascular disease, inflammation, and other diseases. Its main components include curcumol and curcumin, which have anti-inflammatory and antifibrosis effects. Here we established an in vitro inflammatory injury model by stimulating RAW246.7 cells with cigarette smoke extract (CSE) and detected the intervention effects of curcumin and curcumol on CSE-treated Raw246.7 macrophage cells to explore whether the two compounds inhibited the expression of inflammatory cytokines by inhibiting the NF-κB signaling pathway. We detected the antifibrosis effects of curcumin and curcumol via TGF-ß 1/Smads signaling pathways. The model of macrophage damage group was established by CSE stimulation. Curcumol and curcumin were administered to Raw246.7 macrophage cells. The efficacy of curcumol and curcumin was evaluated by comparing the activation of proinflammatory factors, profibrotic factors, and NF-κB and TGF-ß 1/Smads signaling pathway. In addition, CSE-treated group was employed to detect whether the efficacy of curcumol and curcumin was dependent on the NF-κB signaling via the pretreatment with the inhibitor of NF-κB. Our findings demonstrated that curcumol and curcumin could reduce the release of intracellular ROS from macrophages, inhibit the NF-κB signaling pathway, and downregulate the release of proinflammatory factor. Curcumol and curcumin inhibited the TGF-ß 1/Smads signaling pathway and downregulated the release of fibrotic factors. Curcumin showed no anti-inflammatory effect in CSE-treated cells after the inhibition of NF-κB. Curcumol and curcumin showed an anti-inflammatory effect by inhibiting the NF-κB signaling pathway.

Regulation Effect of a Chinese Herbal Formula on Flora and Mucosal Immune Secretory Immunoglobulin A in Rats.

Flora and mucosal immunity are considered to be the barrier, which is associated with multiple respiratory diseases, including recurrent respiratory tract infection (RRTI). Fei-Xi-Tiao-Zhi-Fang (FTF) is a traditional Chinese herbal formula used in the treatment of RRTI. However, the mechanism is little known. This study aims to identify the function of FTF in flora and mucosal immune secretory immunoglobulin A (sIgA) in the model of RRTI rats. The samples of intestine and lung were collected to detect sIgA, short chain fatty acids (SCFAS), and flora with enzyme-linked immunosorbent assay (ELISA), gas chromatography, and 16S rDNA sequencing. The body weight and viscera index were increased dynamically in RRTI rats after the administration of FTF. Furthermore, the types and proportions of aboriginal flora were significantly changed in the model group, whereas the altered flora was rescued in the FTF administration group. Desulfovibrio increased in the intestinal microflora and Ralstonia and Blautia decreased in the pulmonary microflora at the genus level, similar to that in the normal group. In addition, the expressions of sIgA in pulmonary and intestinal tissues were significantly upregulated and the level of SCFAS was increased in FTF group compared to the RRTI model group. Our study suggests that FTF can alleviate the symptoms of RRTI by increasing sIgA and SCFAS, recovering flora, and improving the immunity.

The putative tumor activator ARHGEF3 promotes nasopharyngeal carcinoma cell pathogenesis by inhibiting cellular apoptosis.

Nasopharyngeal carcinoma (NPC) is one of the most prevalent forms of highly invasive malignancy in Southern China and Southeast Asia. The pathogenesis of NPC is a multistep process driven by the acquisition of numerous genetic abnormalities. We investigated the potential oncogenic role of the Rho-guanine nucleotide exchange factor 3 gene, ARHGEF3, in NPC pathogenesis. Expression levels of ARHGEF3 were frequently up-regulated in NPC cell lines and tissues. In a large cohort of clinical NPC tissues high expression of ARHGEF3 was positively associated with an increased T status, distant metastasis, and a more advanced clinical stage (P < 0.05). Survival analysis revealed that ARHGEF3 expression was a significant and independent prognosis factor for NPC patients. In NPC cell lines, knockdown of ARHGEF3 was sufficient to inhibit cell growth, motility, and invasion in vitro, whereas ectopic overexpression of ARHGEF3 substantially enhanced NPC cells tumorigenesis and metastasis in vivo. Depletion of ARHGEF3 in NPC cells dramatically promoted caspase-3 induced apoptosis and an anti-apoptosis factor, BIRC8, was identified as a critical downstream target of the ARHGEF3. Our findings suggest that increased expression of ARHGEF3 plays a critical oncogenic role in NPC pathogenesis by preventing cell apoptosis through the up-regulation of BIRC8, and ARHGEF3 might be employed as a novel prognostic marker and effective therapeutic target for human NPC.

Systemic delivery of microRNA-101 potently inhibits hepatocellular carcinoma in vivo by repressing multiple targets.

Targeted therapy based on adjustment of microRNA (miRNA)s activity takes great promise due to the ability of these small RNAs to modulate cellular behavior. However, the efficacy of miR-101 replacement therapy to hepatocellular carcinoma (HCC) remains unclear. In the current study, we first observed that plasma levels of miR-101 were significantly lower in distant metastatic HCC patients than in HCCs without distant metastasis, and down-regulation of plasma miR-101 predicted a worse disease-free survival (DFS, P<0.05). In an animal model of HCC, we demonstrated that systemic delivery of lentivirus-mediated miR-101 abrogated HCC growth in the liver, intrahepatic metastasis and distant metastasis to the lung and to the mediastinum, resulting in a dramatic suppression of HCC development and metastasis in mice without toxicity and extending life expectancy. Furthermore, enforced overexpression of miR-101 in HCC cells not only decreased EZH2, COX2 and STMN1, but also directly down-regulated a novel target ROCK2, inhibited Rho/Rac GTPase activation, and blocked HCC cells epithelial-mesenchymal transition (EMT) and angiogenesis, inducing a strong abrogation of HCC tumorigenesis and aggressiveness both in vitro and in vivo. These results provide proof-of-concept support for systemic delivery of lentivirus-mediated miR-101 as a powerful anti-HCC therapeutic modality by repressing multiple molecular targets.

The putative tumour suppressor microRNA-124 modulates hepatocellular carcinoma cell aggressiveness by repressing ROCK2 and EZH2.

BACKGROUND: Recent profile studies of microRNA (miRNA) expression have documented a deregulation of miRNA (miR-124) in hepatocellular carcinoma (HCC). OBJECTIVE: To determine the status of miR-124 expression and its underlying mechanisms in the pathogenesis of HCC. METHODS: The expression levels of miR-124 were first examined in HCC cell lines and tumour tissues by real-time PCR. The in vitro and in vivo functional effect of miR-124 was examined further. A luciferase reporter assay was conducted to confirm target associations. RESULTS: The expression levels of miR-124 were frequently reduced in HCC cells and tissues, and low-level expression of miR-124 was significantly associated with a more aggressive and/or poor prognostic phenotype of patients with HCC (p<0.05). In HCC cell lines, stable overexpression of miR-124 was sufficient to inhibit cell motility and invasion in vitro, and suppress intrahepatic and pulmonary metastasis in vivo. In addition, ectopic overexpression of miR-124 in HCC cells inhibited epithelial-mesenchymal cell transition, formation of stress fibres, filopodia and lamellipodia. Further studies showed that miR-124 could directly target the 3'-untranslated region (3'-UTR) of both ROCK2 and EZH2 mRNAs, and suppress their mRNA and protein expressions. These findings suggest that miR-124 plays a critical role in regulating cytoskeletal events and epithelial-mesenchymal cell transition and, ultimately, inhibits the invasive and/or metastatic potential of HCC, probably by its direct target on ROCK2 and EZH2 genes. These results provide functional and mechanistic links between the tumour suppressor miRNA-124 and the two oncogenes ROCK2 and EZH2 on the aggressive nature of HCC. CONCLUSION: These data highlight an important role for miR-124 in the regulation of invasion and metastasis in the molecular aetiology of HCC, and suggest a potential application of miR-124 in prognosis prediction and cancer treatment.

Silencing Fas-associated phosphatase 1 expression enhances efficiency of chemotherapy for colon carcinoma with oxaliplatin.

AIM: To investigate whether silencing Fas-associated phosphatase 1 (FAP-1) expression enhances the efficiency of chemotherapy for colon carcinoma with oxaliplatin. METHODS: Expression of FAP-1 in mRNA and protein was detected by reverse transcription polymerase chain reaction (RT-PCR) and flow cytometry. Small interfering RNA (siRNA) was designed according to the FAP-1 mRNA sequence. Cell proliferation was evaluated by methyl thiazolyl tetrazolium (MTT) assay. Anenxin V- and propidine iodine (PI) were assayed by flow cytometry for the detection of apoptosis. RESULTS: The expression of FAP-1 was increased in SW480 cells after chemotherapy with oxaliplatin. Transfection of FAP-1 siRNA into SW480 cells silenced the expression of FAP-1 and consequently abolished the inhibitory function of Fas/FasL-mediated apoptosis pathway, thus increasing the efficacy of chemotherapy for colon carcinoma with oxaliplatin. CONCLUSION: RNA interference combined with conventional chemotherapy is more effective against colon cancer.

[Serum proteomics in patients with RAEB myelodysplastic syndromes].

OBJECTIVE: To screen the molecular markers for refractory anemia with excess blasts in transformation (RAEB) in myelodysplastic syndromes (MDS) by serum proteome profiling. METHODS: The serum protein were isolated from patients with RAEB, acute myeloid leukemia or normal subjects by 2-dimensional electrophoresis (2-DE), and the electrophoresis gels were obtained to identify the differentially reacting protein spots. The replica gels of the differentially reacting proteins were analyzed to locate the matching protein spots, which were identified by peptide mass fingerprint based on matrix-assisted laser desorption/ionization time of-flight mass spectrometry (MALDI-TOF-MS) and database searching. RESULTS: Seven differentially expressed proteins in RAEB were found by 2-DE. Of the 7 proteins, 4 were identified by MALDI-TOF-MS to have significantly differential expression in RAEB, including dipeptidyl peptidase (DPP/CD26), polymerase (DNA directed) kappa, PRO2044 and an albumin-like protein. CONCLUSION: 2-DE-based serum proteome profiling helps identify serum proteomic biomarkers related to MDS. DDP/CD26 has increased expression in the serum in RAEB subtype MDS, suggesting its possible role in advanced MDS.