Electric pulse exposure reduces AAV8 dosage required to transduce HepG2 cells.

We demonstrate that applying electric field pulses to hepatocytes, in vitro, in the presence of enhanced green fluorescent protein (EGFP)-expressing adeno-associated virus (AAV8) vectors reduces the viral dosage required for a given transduction level by more than 50-fold, compared to hepatocytes exposed to AAV8-EGFP vectors without electric field pulse exposure. We conducted 48 experimental observations across 8 exposure conditions in standard well plates. The electric pulse exposures involved single 80-ms pulses with 375 V/cm field intensity. Our study suggests that electric pulse exposure results in enhanced EGFP expression in cells, indicative of increased transduction efficiency. The enhanced transduction observed in our study, if translated successfully to an in vivo setting, would be a promising indication of potential reduction in the required dose of AAV vectors. Understanding the effects of electric field pulses on AAV transduction in vitro is an important preliminary step.

Functional characterization of a TerC family protein of Riemerella anatipestifer in manganese detoxification and virulence.

Manganese (Mn) is an essential element for bacteria, but the overload of manganese is toxic. In a previous study, we showed that the cation diffusion facilitator protein MetA and the resistance-nodulation-division efflux pump MetB are responsible for Mn efflux in the bacterial pathogen Riemerella anatipestifer CH-1. However, whether this bacterium encodes additional manganese efflux proteins is unclear. In this study, we show that R. anatipestifer CH-1 encodes a tellurium resistance C (TerC) family protein with low similarity to other characterized TerC family proteins. Compared to the wild type (WT), the terC mutant of R. anatipestifer CH-1 (∆terC) is sensitive to Mn(II) intoxication. The ability of TerC to export manganese is higher than that of MetB but lower than that of MetA. Consistently, terC deletion (∆terC) led to intracellular accumulation of Mn2+ under excess manganese conditions. Further study showed that ∆terC was more sensitive than the WT to the oxidant hypoclorite but not to hydrogen peroxide. Mutagenesis studies showed that the mutant at amino acid sites of Glu116 (E116), Asp122 (D122), Glu245 (E245) Asp248 (D248), and Asp254 (D254) may be involved in the ability of TerC to export manganese. The transcription of terC was upregulated under excess manganese and downregulated under iron-limited conditions. However, this was not dependent on the manganese metabolism regulator MetR. In contrast to a strain lacking the manganese efflux pump MetA or MetB, the terC mutant is attenuated in virulence in a duckling model of infection due to increased sensitivity to duck serum. Finally, comparative analysis showed that homologs of TerC are distributed across the bacterial kingdom, suggesting that TerC exerts a conserved manganese efflux function.IMPORTANCERiemerella anatipestifer is a notorious bacterial pathogen of ducks and other birds. In R. anatipestifer, the genes involved in manganese efflux have not been completely identified, although MetA and MetB have been identified as two manganese exporters. Additionally, the function of TerC family proteins in manganese efflux is controversial. Here, we demonstrated that a TerC family protein helps prevent Mn(II) intoxication in R. anatipestifer and that the ability of TerC to export manganese is intermediate compared to that of MetA and MetB. Sequence analysis and mutagenesis studies showed that the conserved key amino sites of TerC are Glu116, Asp122, Glu245, Asp248, and Asp254. The transcription of terC was regulated by manganese excess and iron limitation. Finally, we show that TerC plays a role in the virulence of R. anatipestifer due to the increased sensitivity to duck serum, rather than the increased sensitivity to manganese. Taken together, these results expand our understanding of manganese efflux and the pathogenic mechanisms of R. anatipestifer.

High-fat diet promotes colitis-associated tumorigenesis by altering gut microbial butyrate metabolism.

Background: Dietary fat intake is associated with an increased risk of colitis associated cancer (CAC). A high-fat diet (HFD) leads to systemic low-grade inflammation. The colon is believed to be the first organ suffering from inflammation caused by the infiltration of pro-inflammatory macrophages, and promotes CAC progression. We explored the role of HFD in driving CAC by altering gut microbial butyrate metabolism. Methods: Changes in the gut microbiota caused by HFD were investigated via HFD treatment or fecal microbiota transplantation (FMT). The underlying mechanisms were further explored by analyzing the role of gut microbiota, microbial butyrate metabolism, and NLRP3 inflammasome in colon tissues in a CAC mouse model. Results: HFD accelerated CAC progression in mice, and it could be reversed by broad-spectrum antibiotics (ABX). 16S-rRNA sequencing revealed that HFD inhibited the abundance of butyrate-producing bacteria in the gut. The level of short-chain fatty acids (SCFAs), especially butyrate, in the gut of mice treated with HFD was significantly reduced. In addition, treatment with exogenous butyrate reversed the M1 polarization of proinflammatory macrophages, aggravation of intestinal inflammation, and accelerated tumor growth induced by HFD; the NLRP3/Caspase-1 pathway activated by HFD in the colon was also significantly inhibited. In vitro, macrophages were treated with lipopolysaccharide combined with butyrate to detect the M1 polarization level and NLRP3/Caspase-1 pathway expression, and the results were consistent with those of the in vivo experiments. Conclusion: HFD drives colitis-associated tumorigenesis by inducing gut microbial dysbiosis and inhibiting butyrate metabolism to skew macrophage polarization. Exogenous butyrate is a feasible new treatment strategy for CAC, and has good prospect for clinical application.

RFC5, regulated by circ_0038985/miR-3614-5p, functions as an oncogene in the progression of colorectal cancer.

Replication factor C 5 (RFC5) is involved in a variety of biological functions of cancer. However, the expression pattern of RFC5 and the underlying mechanisms in colorectal cancer (CRC) remain elusive. Here, we show that RFC5 is significantly upregulated in CRC tissues and cells. Patients with CRC and increased RFC5 levels have an unfavorable prognosis. RFC5 can promote the proliferation, migration, and invasion of CRC cells and inhibit the apoptosis of CRC cells. Additionally, upstream of RFC5, we constructed the competing endogenous RNA network and confirmed that RFC5 in this network was inhibited by miR-3614-5p by directly targeting its 3'-untranslated regions. We verified that circ_0038985, which is positively correlated with RFC5, directly targeted miR-3614-5p. Overexpression of circ_0038985 promoted CRC cell migration and invasion, and these effects were partially reversed by the reintroduction of miR-3614-5p. Moreover, we found that RFC5 may promote the vascular endothelial growth factor A (VEGFa)/vascular endothelial growth factor receptor 2 (VEGFR2)/extracellular signal-regulated protein kinase (ERK) pathway. The knockdown of RFC5 reduced CRC tumorigenesis in vivo. Collectively, these data demonstrate that the circ_0038985/miR-3614-5p/RFC5 axis plays a critical role in the progression of CRC, and RFC5 may promote CRC progression by affecting the VEGFa/VEGFR2/ERK pathway.

Hsa-circ-0052001 promotes gastric cancer cell proliferation and invasion via the MAPK pathway.

BACKGROUND: Gastric cancer (GC) ranks fourth among the causes of death from malignant tumors in the world. Studies have implicated the dysregulation of circRNAs with GC. However, the relationship between hsa-circ-0052001 and GC is unclear. METHODS: In our current study, we assessed the expression levels of hsa-circ-0052001 in GC cells and tissues using quantitative real-time PCR (qPCR). The role of hsa-circ-0052001 expression on the proliferation and invasion of GC cells was assessed using in vitro experiments. The role of hsa-circ-0052001 on the proliferation of GC cells was also analyzed using in vivo models. The pathways downstream of hsa-circ-0052001 were identified using bioinformatics analyses, western blot (WB) assays, and qRT-PCR. RESULTS: We found that compared with normal gastric mucosa epithelial cells and adjacent paracancer tissues, hsa-circ-0052001 was overexpressed in GC cells and tissues. Also, the hsa-circ-0052001 level was linked to patient clinicopathological characteristics of GC. Cell proliferation and metastatic ability were inhibited in gastric cancer cells when hsa-circ-0052001 was knocked down in vitro and cancer growth in vivo. Mechanistically, hsa-circ-0052001 promoted the carcinogenesis of GC cells via the MAPK signal pathway. CONCLUSION: Hsa-circ-0052001 functions as a tumor gene in promoting the progression of GC through MAPK pathway, which has provided a promising target for patients with GC.

STING promotes proliferation and induces drug resistance in colorectal cancer by regulating the AMPK-mTOR pathway.

Background: In recent years, reports regarding stimulator of interferon genes (STING) and the progression of colorectal cancer (CRC) have emerged rapidly, yet their association remains controversial. This research was aimed to provide an insight into the prognostic biomarker and therapeutic target significance of STING in CRC. Methods: CRC Cell lines of HCT116 and SW480, as well as 32 paired CRC specimens were chosen for this study. STING expressions were examined by immunohistochemistry to evaluate the correlation with clinicopathological factors. Data analysis of STING expressions in colon cancer and rectal cancer were performed using The Cancer Genome Atlas (TCGA) database. siRNA was transfected into cell lines for knocking down the expression of STING. Transwell assay was employed to evaluate cell migration and invasiveness. CCK-8 assay was used for assessing the change of cell proliferation. Drug sensitive test was involved to evaluate drug resistance of cell lines. Gene Set Enrichment Analysis (GSEA) was applied for exploring potential downstream mechanism of STING in CRC progression and Western blotting is used for mechanism validation. Results: In the thirty-two paired CRC and adjacent normal tissues, we found a significant up-regulated in STING expression with immunohistochemical staining in cancer tissues compared with adjacent normal tissues (P<0.01), which was correlated with the tumor-node-metastasis (TNM) stage of patients (P=0.028). Meanwhile, GESA enrichment analysis indicated a remarkable change in mTOR signaling following STING regulation. In HCT116 and SW480 cell lines of CRC, When STING was down-regulated, its biological behavior of cell viability, cell invasion and drug sensitivity to 5-fluorouracil were significantly reduced (P<0.05), we also observed the up-regulation of P-AMPK (P<0.05) and down-regulation of p-mTOR (P<0.05). Conclusions: STING expressions was significantly up-regulated in CRC tissues. Expression of STING was correlated with the TNM stage of patients. STING is found to promote cell proliferation, invasion ability and drug resistance mediating AMPK-mTOR signaling in CRC. STING could be a promising target for the sensitization of chemotherapy and inhibits CRC progression.

Roux-en-Y reconstruction alleviates radical gastrectomy-induced colitis via down-regulation of the butyrate/NLRP3 signaling pathway.

BACKGROUND: Different methods for digestive tract reconstruction have a complex impact on the nutritional status of gastric cancer (GC) patients after radical gastrectomy. Previous studies reported that Roux-en-Y (R-Y) reconstruction resulted in obvious weight reduction and improvement in type 2 diabetes in obese patients. We investigated the relationship between R-Y reconstruction, gut microbiota, and the NLRP3 inflammasome in GC patients with poor basic nutrition. METHODS: Changes in the gut microbiota after radical gastrectomy accomplished by different methods of digestive tract reconstruction were investigated via fecal microbiota transplantation. The underlying mechanisms were also explored by analyzing the role of the microbiota, butyrate, and the NLRP3 inflammasome in the colon tissues of colitis model mice and GC patients after radical gastrectomy. FINDINGS: R-Y reconstruction effectively relieved intestinal inflammation and facilitated nutrient absorption. 16S rRNA analysis revealed that gavage transplantation with the fecal microbiota of R-Y reconstruction patients could reverse dysbacteriosis triggered by radical gastrectomy and elevate the relative abundance of some short-chain fatty acid (SCFA)-producing bacteria. Subsequently, butyrate negatively regulated the NLRP3-mediated inflammatory signaling pathway to inhibit the activation of macrophages and the secretion of pro-inflammatory mediators such as caspase-1 and interleukin (IL)-1ß, decreasing the level of intestinal inflammation and promoting nutrient absorption. INTERPRETATION: R-Y reconstruction induced colonization with SCFA-producing bacteria to alleviate radical gastrectomy-induced colitis by down-regulating the NLRP3 signaling pathway. This can be a new strategy and theoretical basis for the management of the postoperative nutritional status of GC patients. FUNDING: This work was supported by the National Nature Science Foundation of China (81974375), the BoXi cultivation program (BXQN202130), and the Project of Youth Foundation in Science and Education of the Department of Public Health of Suzhou (KJXW2018001).

TBK1 Facilitates GLUT1-Dependent Glucose Consumption by suppressing mTORC1 Signaling in Colorectal Cancer Progression.

Intestinal inflammation is a vital precipitating factor of colorectal cancer (CRC), but the underlying mechanisms are still elusive. TANK-binding kinase 1 (TBK1) is a core enzyme downstream of several inflammatory signals. Recent studies brought the impacts of TBK1 in malignant disease to the forefront, we found aberrant TBK1 expression in CRC is correlated with CRC progression. TBK1 inhibition impaired CRC cell proliferation, migration, drug resistance and tumor growth. Bioinformatic analysis and experiments in vitro showed overexpressed TBK1 inhibited mTORC1 signaling activation in CRC along with elevated GLUT1 expression without inducing GLUT1 translation. TBK1 mediated mTORC1 inhibition induces intracellular autophagy, which in turn decreasing GLUT1 degradation. As a rescue, blocking of autophagosome and retromer respectively via autophagy-related gene 7 (ATG7) or TBC1 Domain Family Member 5 (TBC1D5) silence diminished the regulation of TBK1 to GLUT1. GLUT1 staining presented that TBK1 facilitated GLUT1 membrane translocation which subsequently enhanced glucose consumption. Inhibitor of TBK1 also decreased GLUT1 expression which potentiated drug-sensitivity of CRC cell. Collectively, TBK1 facilitates glucose consumption for supporting CRC progression via initiating mTORC1 inhibition induced autophagy which decreases GLUT1 degradation and increases GLUT1 membrane location. The adaptive signaling cascade between TBK1 and GLUT1 proposes a new strategy for CRC therapy.

Cortisol resistance in the degu (Octodon degus).

Cortisol resistance has also been reported in the degu, Octodon degus, a New World hystricomorph endemic to central Chile. The degu is used as a model for studies of stress and diurnal rhythms, parental behaviour and female masculinization. Another New World hystricomorph, the guinea pig, also exhibits glucocorticoid resistance, a result of amino acid sequences that differ from other mammalian glucocorticoid receptors (GR). Mutations in the ligand-binding domain of the human GR have been identified in familial or sporadic generalised cortisol resistance as have variants in the guinea pig. To address the possibility that the high levels of cortisol observed in the degu are a result of the same or similar sequence variations observed in the guinea pig GR, we have cloned, expressed and characterised the ligand-binding domain (LBD) of the degu GR. Somewhat unexpectedly, neither the amino acids nor the region involved in the resistance observed in the guinea pig GR are relevant in the degu GR. The relative resistance to cortisol observed in the degu GR is conferred by the substitution of two isoleucine residues, which are highly conserved in the GR across species, with a valine doublet. These amino acids lie in the region between helices 5 and 6 of the GR LBD, a region known to be important in determining the affinity of ligand-binding in steroid receptors.

CD36 regulates LPS-induced acute lung injury by promoting macrophages M1 polarization.

M1 polarization of macrophages works as a promoter in pathogenesis of acute lung injury / acute respiratory distress syndrome (ALI/ARDS) by the secretion of pro-inflammatory cytokines and recruiting other inflammatory cells. Lipopolysaccharide (LPS), a critical component of the wall of gram-negative bacteria, can induce M1 polarization and ALI. Recently, cluster of differentiation 36 (CD36) has been reported to be associated with inflammatory responses. However, it has not yet been clarified whether CD36 in macrophages is involved in LPS-induced ALI. Herein, we demonstrated that in macrophages, LPS-induced ALI was regulated by CD36. Loss of CD36 attenuated LPS-induced ALI by reducing M1 polarization. Mechanistically, CD36 promoted macrophage M1 polarization by regulating CD14 associated with TLR4 during LPS stimulation. The findings of this study, clarified the mechanism of LPS-induced ALI through CD36 in macrophages, which provides a potential target for the prevention and treatment of ALI.

Polluted lake restoration to promote sustainability in the Yangtze River Basin, China.

China has made a concerted effort to successfully improve water quality of rivers, but lake water quality has not improved. Lakes require controls on both catchment external nutrient loads and in-lake internal loads, where nature-based solutions are coupled with engineered systems to achieve the United Nations Sustainable Development Goals (SDGs).

ENO1 contributes to 5-fluorouracil resistance in colorectal cancer cells via EMT pathway.

Introduction: Chemoresistance is a major barrier in the treatment of colorectal cancer (CRC) and many other cancers. ENO1 has been associated with various biological characteristics of CRC. This study aimed to investigate the function of ENO1 in regulating 5-Fluorouracil (5-FU) resistance in CRC. Methods: ENO1 level in 120 pairs of tumor tissues and adjacent normal tissues was examined by immunohistochemistry, and the correlation between ENO1 expression and prognosis was explored by survival analysis. Its role and potential mechanisms in regulating 5-FU resistance in CRC were studied by Western blotting, MTT assay, colony formation assay and transwell invasion assay. Murine xenograft assay was implied to verify the results in vivo. Results: Our study indicated that ENO1 was elevated in CRC tissues and was associated with poor patient prognosis. High levels of ENO1 expression were detected as a significant influencing factor for overall survival. Furthermore, ENO1 expression was found to have increased in drug-resistant cells (HCT116/5-FU and SW620/5-FU) constructed by increasing concentrations of 5-FU. Knockdown of ENO1 markedly increased the drug susceptibility and inhibited the proliferation and migration ability of HCT116/5-FU and SW620/5-FU cells. It was found that down-regulation of ENO1 inhibited the epithelial-mesenchymal transformation (EMT) signaling process. Finally, a murine xenograft assay verified that the depletion of ENO1 alleviated 5-FU resistance. Conclusion: This study identified that ENO1 regulated 5-FU resistance via the EMT pathway and may be a novel target in the prevention and treatment of 5-FUresistant CRC.

Corrigendum: The Association of Aberrant Expression of FGF1 and mTOR-S6K1 in Colorectal Cancer.

[This corrects the article DOI: 10.3389/fonc.2021.706838.].

Bacteroides fragilis restricts colitis-associated cancer via negative regulation of the NLRP3 axis.

Patients with persistent ulcerative colitis (UC) are at a higher risk of developing colitis-associated cancer (CAC). Previous studies have reported that intestinal microbiota disturbance plays an important role in the process of CAC development in patients with UC, indicating that targeted intervention of intestinal microbiota and its metabolites may be a potential therapeutic strategy. Gut microbiota in the process of colorectal cancer development in UC patients was analyzed using the gutMEGA database and verified in fecal samples. The abundance of Bacteroides fragilis reduced significantly in the process of colitis associated cancer development. Broad-spectrum antibiotics (BSAB) intervene with the intestinal microbiota of mice and accelerate the process of colon cancer development. However, gavage transplantation with B. fragilis can effectively reverse the effects of BSAB. In the intestinal tract, B. fragilis promotes the secretion of short-chain fatty acids (SCFAs). Subsequently, SCFAs, especially butyrate, negatively regulate the inflammatory signaling pathway mediated by NLRP3 to inhibit the activation of macrophages and the secretion of proinflammatory mediators such as IL-18 and IL-1ß, reducing the level of intestinal inflammation and restricting CAC development. In conclusion, colonization with B. fragilis has been shown to be effective in ameliorating intestinal epithelial damage caused by chronic inflammation and preventing the development of colonic tumors. Thus, it can be a therapeutic intervention strategy with good clinical application prospects.

DAB2IP decreases cell growth and migration and increases sensitivity to chemotherapeutic drugs in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is one of the most common cancers worldwide with high rates of invasiveness and mortality. DAB2IP (DOC2/DAB2 interactive protein) is a member of the RAS-GTPase-activating protein (RAS-GAP) family that shows a suppressive effect on cancer progression, is downregulated in several cancers. However, the role of DAB2IP in CRC remains elusive. METHODS: Expression of DAB2IP was evaluated in human CRC tissues using immunohistochemistry (IHC), quantitative real-time reverse transcription PCR (qRT-PCR) and immunoblotting. Knockdown and overexpression of DAB2IP in CRC cells were achieved by transfecting siRNAs and DAB2IP expression vectors and assessed by qRT-PCR and immunoblotting. CCK-8, colony formation, wound-healing, and transwell assays were used to evaluate CRC cell growth, migration, and sensitivity to chemotherapeutic drugs. The cell cycle was analyzed by propidium iodide (PI) staining and flow cytometry. Cell apoptosis was evaluated by Annexin V-DAPI double staining and flow cytometry. The effect of DAB2IP overexpression on tumor formation was explored by an in vivo tumorigenesis assay. Finally, immunoblotting was performed to examine the molecules related to the action of DAB2IP in CRC. RESULTS: Compared with para-cancer tissues, there was a marked decrease of DAB2IP expression in surgically excised CRCs. In cultured CRC cells, enforced expression of DAB2IP inhibited cell growth and migration and sensitized the cells to DNA-acting cisplatin, oxaliplatin, and doxorubicin but not 5-fluorouracil (5-FU). In contrast, knockdown of DAB2IP produced the opposite effect. Moreover, DAB2IP overexpression hindered tumor growth in vivo. We further found that DAB2IP regulated the expression of cell growth, epithelial-mesenchymal transition (EMT), and apoptosis-related proteins in CRC cells and inhibited the phosphorylation of protein kinase B (AKT) and extracellular signal-regulated kinase (ERK). CONCLUSIONS: Expression of DAB2IP inhibited CRC cell growth and migration and sensitized CRC cells to chemotherapeutic drugs. Inhibition of the phosphorylation of AKT and ERK is associated with the effects of DAB2IP expression. Restoration of DAB2IP expression may be a novel target for treating CRC.

The Association of Aberrant Expression of FGF1 and mTOR-S6K1 in Colorectal Cancer.

Colorectal cancer (CRC) is one of the most frequent malignant neoplasms worldwide, and the effect of treatments is limited. Fibroblast growth factor 1 (FGF1) has been involved in a wide variety of several malignant diseases and takes part in the tumorigenesis of CRC. However, the function and mechanism of FGF1 in CRC remains elusive. In this study, the results indicated that FGF1 is elevated in CRC tissues and linked with poor prognosis (P < 0.001). In subgroup analysis of FGF1 in CRC, regardless of any clinic-factors except gender, high level FGF1 expression was associated with markedly shorter survival (P < 0.05). In addition, the expression of p-S6K1 and FGF1 was not associated in normal tissue (P = 0.781), but their expression was closely related in tumor tissue (P = 0.010). The oncogenic role of FGF1 was determined using in vitro and in vivo functional assays. FGF1 depletion inhibited the proliferation and migration of CRC cells in vitro and vivo. FGF1 was also significantly correlated with mTOR-S6K1 pathway on the gene and protein levels (P < 0.05). In conclusion, FGF1 acts as a tumor activator in CRC, and against FGF1 may provide a new visual field on treating CRC, especially for mTORC1-targeted resistant patients.

YAP promotes the proliferation and migration of colorectal cancer cells through the Glut3/AMPK signaling pathway.

Yes-associated protein (YAP), as a major downstream effector in the Hippo signaling pathway, is considered as an oncogene in cancer. The present study aimed to investigate the potential role of YAP in the development and progression of colorectal cancer (CRC). The mRNA and protein expression levels of YAP in human CRC tissue samples and adjacent normal tissue were analyzed using public databases, as well as clinical samples. The potential roles of YAP and the underlying mechanism regulating the proliferation and migration of CRC cells were examined using genetic manipulation in vitro. The correlation between the expression of the YAP gene and epithelial-to-mesenchymal transition (EMT) markers was investigated in order to determine the mechanism underlying the observed effects of YAP. YAP mRNA expression levels were significantly upregulated in CRC tissue compared with in normal tissue, as determined using datasets obtained from Oncomine. Similarly, in clinical samples, the protein expression levels of YAP were significantly upregulated in CRC tissue samples compared with in normal tissue samples. YAP knockdown inhibited the proliferation and migration of CRC cells in vitro, whereas its overexpression resulted in the opposite effect. The expression levels of the YAP gene were positively correlated with those of EMT markers (such as vimentin and N-cadherin) and EMT-inducing transcription factors (such as Snail1, Slug and zinc finger E-box binding homeobox 1 and 2) in CRC samples from Gene Expression Profiling Interactive Analysis. Furthermore, YAP silencing increased the protein expression of E-cadherin and decreased that of vimentin in CRC cells. By contrast, the overexpression of YAP had the opposite effect. YAP promoted the glucose transporter 3 (Glut3)/AMP-activated protein kinase (AMPK) signaling pathway in CRC cells. In conclusion, YAP promoted the proliferation and migration of CRC cells, as well as the expression of EMT markers, possibly by regulating the Glut3/AMPK signaling pathway.

A Broccoli aptamer chimera yields a fluorescent K+ sensor spanning physiological concentrations.

The RNA aptamer Broccoli accepts 2'fluorinated (2'F) pyrimidine nucleotide incorporation without perturbation of structure or fluorescence in the presence of potassium and DFHBI. However, the modification decreases Broccoli's apparent affinity for K+ >30-fold. A chimera of Broccoli RNAs with mixed chemistries displays linear fluorescent gain spanning physiological K+ concentrations, yielding an effective RNA-based fluorescent K+ sensor.

A New Biomarker of Fecal Bacteria for Non-Invasive Diagnosis of Colorectal Cancer.

Background: The intestinal flora is correlated with the occurrence of colorectal cancer. We evaluate a new predictive model for the non-invasive diagnosis of colorectal cancer based on intestinal flora to verify the clinical application prospects of the intestinal flora as a new biomarker in non-invasive screening of colorectal cancer. Methods: Subjects from two independent Asian cohorts (cohort I, consisting of 206 colorectal cancer and 112 healthy subjects; cohort II, consisting of 67 colorectal cancer and 54 healthy subjects) were included. A probe-based duplex quantitative PCR (qPCR) determination was established for the quantitative determination of candidate bacterial markers. Results: We screened through the gutMEGA database to identify potential non-invasive biomarkers for colorectal cancer, including Prevotella copri (Pc), Gemella morbillorum (Gm), Parvimonas micra (Pm), Cetobacterium somerae (Cs), and Pasteurella stomatis (Ps). A predictive model with good sensitivity and specificity was established as a new diagnostic tool for colorectal cancer. Under the best cutoff value that maximizes the sum of sensitivity and specificity, Gm and Pm had better specificity and sensitivity than other target bacteria. The combined detection model of five kinds of bacteria showed better diagnostic ability than Gm or Pm alone (AUC = 0.861, P < 0.001). These findings were further confirmed in the independent cohort II. Particularly, the combination of bacterial markers and fecal immunochemical test (FIT) improved the diagnostic ability of the five bacteria (sensitivity 67.96%, specificity 89.29%) for patients with colorectal cancer. Conclusion: Fecal-based colorectal cancer-related bacteria can be used as new non-invasive diagnostic biomarkers of colorectal cancer. Simultaneously, the molecular biomarkers in fecal samples are similar to FIT, have the applicability in combination with other detection methods, which is expected to improve the sensitivity of diagnosis for colorectal cancer, and have a promising prospect of clinical application.