Endogenously Gated DNA Walking Machine for Prescreened MicroRNA Detection in Extracellular Vesicles.

Tumor-derived extracellular vesicle (T-EV) microRNAs have been investigated as promising biomarkers in clinical diagnosis as well as disease progression monitoring. However, the expression profiles of microRNA in different tissues vary widely, the precise monitoring of microRNA levels in EVs originating from diseased tissues is susceptible to background interference, thus remains a challenge. Conventional assays require extensive processing, such as EV isolation and even sample lysis, which is both slow and laborious, and the cumbersome pretreatment could spoil the downstream analysis. To address this issue, we developed a generalizable strategy for T-EVs-selective activation and therefore specific amplified microRNA imaging. The conditional signal amplification is established by integrating a traditional DNA walker system with endogenously activated motif to achieve sensitized microRNA imaging in T-EVs. The preorganized endogenous activation with additional sensing criteria narrowed the scope against the complex specimens, and the amplified sensing with reduced off-target signals was supposed to be sensitive to monitor the tiny changes of microRNA expression during the disease course, which holds great potential for accurate diagnosis and prognosis.

Activating peroxymonosulfate with Fe-doped biochar for efficient removal of tetracycline: Dual action of reactive oxygen species and electron transfer.

If pharmaceutical wastewater is not managed effectively, the presence of residual antibiotics will result in significant environmental contamination. In addition, inadequate utilization of agricultural waste represents a squandering of resources. The objective of this research was to assess the efficacy of iron-doped biochar (Fe-BC) derived from peanut shells in degrading high concentrations of Tetracycline (TC) wastewater through activated peroxymonosulfate. Fe-BC demonstrated significant efficacy, achieving a removal efficiency of 87.5% for TC within 60 min without the need to adjust the initial pH (20 mg/L TC, 2 mM PMS, 0.5 g/L catalyst). The degradation mechanism of TC in this system involved a dual action, namely Reactive Oxygen Species (ROS) and electron transfer. The primary active sites were the Fe species, which facilitated the generation of SO4•-, •OH, O2•-, and 1O2. The presence of Fe species and the C=C structure in the Fe-BC catalyst support the electron transfer. Degradation pathways were elucidated through the identification of intermediate products and calculation of the Fukui index. The Toxicity Estimator Software Tool (T.E.S.T.) suggested that the intermediates exhibited lower levels of toxicity. Furthermore, the system exhibited exceptional capabilities in real water and circulation experiments, offering significant economic advantages. This investigation provides an efficient strategy for resource recycling and the treatment of high-concentration antibiotic wastewater.

Clinical Efficiency of Fixed-Bearing Unicompartmental Knee Arthroplasty Versus Total Knee Arthroplasty For Lateral Compartment Knee Osteoarthritis and the Effect on Recovery of Motor Function.

Objective: To evaluate the clinical efficiency of fixed-bearing unicompartmental knee arthroplasty (UKA) versus total knee arthroplasty (TKA) for lateral compartment knee osteoarthritis and the effect on the recovery of motor function. Methods: A total of 54 patients who underwent surgery for lateral compartment knee osteoarthritis and satisfied the inclusion criteria from September 2018 to February 2021 at our hospital were recruited and assigned to receive either UKA (UKA group, n=30) or TKA (TKA group, n=24) via random number table method. Among them, the randomization was carried out using an online web-based randomization tool (freely available at http://www.randomizer.org/). Inclusion criteria: 1) patients with lateral compartment knee osteoarthritis diagnosed by clinically relevant tests; 2) patients with structural and functional integrity of the knee ligaments; 3) all with a single knee lesion. Outcome measures included operative time, the reduction ratio of Hb 1d postoperatively, visual analog scale (VAS) score 7d postoperatively, length of hospital stay, postoperative Keen society score (KSS), Oxford knee score (OKS), range of motion (ROM), forgotten joint score (FJS), motor function recovery, and adverse events. Results: All patients were followed up postoperatively for 12-33 (21.71±7.45) months. Patients in the UKA group showed significantly shorter operative indices, a lower reduction ratio of Hb 1d postoperatively, and VAS scores of 7d postoperatively (P < .05). At 1 month and 6 months postoperatively, UKA resulted in significantly better KSS scores, OKS scores, ROM, and motor function recovery versus TKA (P < .05), while the difference of the above indices did not come up to the statistical standard at 1 year postoperatively (P < .05). At 1 year postoperatively, patients receiving UKA were associated with significantly higher FJS scores versus those given TKA. No documented thrombosis, knee, or prosthesis-related adverse events were observed during hospitalization and follow-up. Conclusion: In comparison to TKA, UKA resulted in smaller surgical incisions, improved postoperative healing, and greater restoration of knee function. Both arthroplasties are successful in alleviating pain and increasing knee function, although they are less effective in recovering patients' motor capabilities.

Quantification and Comparison of Risks Associated with Wastewater Use in Spray Irrigation.

In the U.S., spray irrigation is the most common method used in agriculture and supplementing with animal wastewater has the potential to reduce water demands. However, this could expose individuals to respiratory pathogens such as Legionella pneumophila and nontuberculosis Mycobacteria (NTM). Disinfection with methods like anaerobic digestion is an option but can increase concentrations of cytotoxic ammonia (personal communication). Our study aimed to model the annual risks of infection from these bacterial pathogens and the air concentrations of ammonia and determine if anaerobically digesting this wastewater is a safe option. Air dispersion modeling, conducted in AERMOD, generated air concentrations of water during the irrigation season (May-September) for the years 2013-2018. These values fed into the quantitative microbial risk assessments for the bacteria and allowed calculation of ammonia air concentrations. The outputs of these models were compared to the safety thresholds of 10-4 infections/year and 0.5 mg/m3 , respectively, to determine their potential for negative health outcomes. It was determined that infection from NTM was not a concern for individuals near active spray irrigators, but that infection with L. pneumophila could be a concern, with a maximum predicted annual risk of infection of 3.5 × 10-3 infections/year and 25.2% of parameter combinations exceeding the established threshold. Ammonia posed a minor risk, with 1.5% of parameter combinations surpassing the risk threshold of 0.5 mg/m3 . These findings suggest that animal wastewater should be anaerobically digested prior to use in irrigation to remove harmful pathogens.

TGF-ß/Smad and JAK/STAT pathways are involved in the anti-fibrotic effects of propylene glycol alginate sodium sulphate on hepatic fibrosis.

Liver fibrosis, a consequence of unhealthy modern lifestyles, has a growing impact on human health, particularly in developed countries. Here, we have explored the anti-fibrotic effects of propylene glycol alginate sodium sulphate (PSS), a natural extract from brown algae, in fibrotic mice and cell models. Thus, we established bile duct ligature and carbon tetrachloride mouse models and LX-2 cell models with or without PSS treatment. Liver pathological sections and the relevant indicators in serum and liver tissues were examined. PSS prevented hepatic injury and fibrosis to a significant extent, and induced up-regulation of matrix metalloproteinase-2 and down-regulation of tissue inhibitor of metalloproteinase-1 through suppressing the transforming growth factor ß1 (TGF-ß1)/Smad pathway. PSS additionally exerted an anti-autophagy effect through suppressing the Janus kinase (JAK) 2/transducer and activator of transcription 3 (STAT3) pathway. In conclusion, PSS prevents hepatic fibrosis by suppressing inflammation, promoting extracellular matrix (ECM) decomposition and inactivating hepatic stellate cells through mechanisms involving the TGF-ß1/Smad2/3 and JAK2/STAT3 pathways in vivo and in vitro.

The value of the atherogenic index of plasma in non-obese people with non-alcoholic fatty liver disease: a secondary analysis based on a cross-sectional study.

BACKGROUND AND OBJECTIVES: The atherogenic index of plasma (AIP) is elevated in fatty liver disease, but its value in non-obese people with non-alcoholic fatty liver disease (NAFLD) is unclear. This study aimed to investigate the relationship between AIP and NAFLD as well as to determine whether AIP might be used as an indicator of NAFLD in non-obese individuals. METHODS: The present study involved non-obese Chinese and Japanese participants. Risk factors are evaluated using univariate and multivariate analysis. The performance of risk factors was compared according to the area under the receiver operating characteristic curve. RESULTS: In the unadjusted model, the odds ratio (OR) for every 1 standard deviation (SD) increase in AIP was 52.30. In adjusted models I and II, the OR for every 1 SD increase in AIP was 36.57 and 50.84, respectively. The area under the receiver operating characteristic curve for AIP was 0.803 and 0.802 in the development and validation groups, respectively. The best cut-off value of AIP for discrimination between NAFLD and non-NAFLD was 0.005 in the Chinese group and - 0.220 in the Japanese group. CONCLUSIONS: AIP and NAFLD are positively correlated in Chinese and Japanese populations. Therefore, AIP can be used as a new screening indicator for non-obese people with NAFLD in different nations.

A Novel Detector Based on Convolution Neural Networks for Multiscale SAR Ship Detection in Complex Background.

Convolution neural network (CNN)-based detectors have shown great performance on ship detections of synthetic aperture radar (SAR) images. However, the performance of current models has not been satisfactory enough for detecting multiscale ships and small-size ones in front of complex backgrounds. To address the problem, we propose a novel SAR ship detector based on CNN, which consist of three subnetworks: the Fusion Feature Extractor Network (FFEN), Region Proposal Network (RPN), and Refine Detection Network (RDN). Instead of using a single feature map, we fuse feature maps in bottom-up and top-down ways and generate proposals from each fused feature map in FFEN. Furthermore, we further merge features generated by the region-of-interest (RoI) pooling layer in RDN. Based on the feature representation strategy, the CNN framework constructed can significantly enhance the location and semantics information for the multiscale ships, in particular for the small ships. On the other hand, the residual block is introduced to increase the network depth, through which the detection precision could be further improved. The public SAR ship dataset (SSDD) and China Gaofen-3 satellite SAR image are used to validate the proposed method. Our method shows excellent performance for detecting the multiscale and small-size ships with respect to some competitive models and exhibits high potential in practical application.

Long noncoding RNA HOTTIP mediates SRF expression through sponging miR-150 in hepatic stellate cells.

HOXA transcript at the distal tip (HOTTIP) has been shown to be up-regulated in a variety of cancers and is identified as an oncogenic long noncoding RNA. However, the biological role of HOTTIP in liver fibrosis is unclear. Here, we reported that HOTTIP was specifically overexpressed in activated hepatic stellate cells (HSCs). HOTTIP knockdown suppressed the activation and proliferation of HSCs. Luciferase reporter assay showed that HOTTIP and serum response factor (SRF) were targets of miR-150. RNA binding protein immunoprecipitation assay indicated the interaction between miR-150 and HOTTIP. Further study revealed that HOTTIP increased SRF expression as a competing endogenous RNA for miR-150, thus prompting HSC activation. Taken together, we provide a novel HOTTIP-miR-150-SRF signalling cascade in liver fibrosis.

Procyanidin B2 inhibits the activation of hepatic stellate cells and angiogenesis via the Hedgehog pathway during liver fibrosis.

BACKGROUND: Liver fibrosis is a wound-healing process of liver featured by the over-deposition of extracellular matrix (ECM) and angiogenesis. However, the effective treatment is lacking. Procyanidin B2 (PB2) is a flavonoid extract abundant in grape seeds with anti-oxidant, anti-inflammatory and anti-cancer properties. The present study aimed to determine effects of PB2 on liver fibrosis. METHOD: The CCl4-induced mouse liver fibrosis model and a human hepatic stellate cell (HSC) line (LX2 cells) were used to study the activation, ECM production and angiogenesis of HSCs through Western blotting analysis, immunohistochemistry, immunofluorescence staining, flow cytometry and tubulogenesis assay. A Hedgehog (Hh) pathway inhibitor (cyclopamine) and Smoothened agonist (SAG) were used to investigate the role of PB2 on Hh pathway. RESULTS: The results showed that PB2 could inhibit the proliferation and induce apoptosis of HSCs. PB2 could also down-regulate the expressions of VEGF-A, HIF-1α, α-SMA, Col-1 and TGF-ß1 of HSCs in vivo and in vitro. The application of SAG and cyclopamine proved that PB2 targets on Hh pathway. CONCLUSIONS: PB2 inhibited the Hh pathway to suppress the activation, ECM production and angiogenesis of HSCs, therefore reverses the progression of liver fibrosis in vivo and in vitro.

Alleviation of Hepatic Ischemia Reperfusion Injury by Oleanolic Acid Pretreating via Reducing HMGB1 Release and Inhibiting Apoptosis and Autophagy.

Hepatic ischemia reperfusion (IR) injury (IRI) occurs during liver transplantation, hepatectomy, and hemorrhagic shock. Oleanolic acid (OA) is a natural compound with antioxidant and anti-inflammatory activity that has been used to treat liver disorders in clinical practice for several years. Here, we investigated the effects and underlying mechanisms of OA in hepatic IRI. A 60-minute partial (70%) hepatic, warm, ischemic reperfusion model was established in BALB/c mice, and two doses (30 and 60 mg/kg) of OA were administered intragastrically for 7 consecutive days prior to hepatic IR. Orbital blood and liver specimens were collected at 2, 8, and 24 h after IR. The results showed that OA preconditioning significantly alleviated hepatic injury, as evidenced by decreased alanine aminotransferase and aspartate aminotransferase levels; improved histology, inhibition of JNK phosphorylation, and high mobility group box 1 (HMGB1); and tumor necrosis factor-α downregulation in hepatic IR mice. OA upregulated Bcl-2 and downregulated caspase-3, caspase-9, Bax, Beclin 1, and LC3, which play crucial roles in the regulation of apoptosis and autophagy. These findings highlighted the protective effects of OA against hepatic IRI mediated by the inhibition of apoptosis and autophagy and the release of HMGB1, which acted as a late inflammatory mediator in hepatic IRI.

Antinociceptive Effect of Ghrelin in a Rat Model of Irritable Bowel Syndrome Involves TRPV1/Opioid Systems.

BACKGROUND/AIMS: Irritable bowel syndrome (IBS), defined as recurrent abdominal pain and changes in bowel habits, seriously affects quality of life and ability to work. Ghrelin is a brain-gut hormone, which has been reported to show antinociceptive effects in peripheral pain. We investigated the effect of ghrelin on visceral hypersensitivity and pain in a rat model of IBS. METHODS: Maternal deprivation (MD) was used to provide a stress-induced model of IBS in Wistar rats. Colorectal distension (CRD) was used to detect visceral sensitivity, which was evaluated by abdominal withdrawal reflex (AWR) scores. Rats that were confirmed to have visceral hypersensitivity after MD were injected with ghrelin (10 µg/kg) subcutaneously twice a week from weeks 7 to 8. [D-Lys3]-GHRP-6 (100 nmol/L) and naloxone (100 nmol/L) were administered subcutaneously to block growth hormone secretagogue receptor 1α (GHS-R1α) and opioid receptors, respectively. Expression of transient receptor potential vanilloid type 1 (TRPV1) and µ and κ opioid receptors (MOR and KOR) in colon, dorsal root ganglion (DRG) and cerebral cortex tissues were detected by western blotting, quantitative real-time polymerase chain reaction (qRT-PCR), immunohistochemical analyses and immunofluorescence. RESULTS: Ghrelin treatment increased expression of opioid receptors and inhibited expression of TRPV1 in colon, dorsal root ganglion (DRG) and cerebral cortex. The antinociceptive effect of ghrelin in the rat model of IBS was partly blocked by both the ghrelin antagonist [D-Lys3]-GHRP-6 and the opioid receptor antagonist naloxone. CONCLUSION: The results indicate that ghrelin exerted an antinociceptive effect, which was mediated via TRPV1/opioid systems, in IBS-induced visceral hypersensitivity. Ghrelin might potentially be used as a new treatment for IBS.

Long Non-coding RNA Growth Arrest-specific Transcript 5 (GAS5) Inhibits Liver Fibrogenesis through a Mechanism of Competing Endogenous RNA.

Effective control of hepatic stellate cell (HSC) activation and proliferation is critical to the treatment of liver fibrosis. Long non-coding RNAs have been shown to play a pivotal role in the regulation of cellular processes. It has been reported that growth arrest-specific transcript 5 (GAS5) acts as a crucial mediator in the control of cell proliferation and growth. However, little is known about the role and underlying mechanism of GAS5 in liver fibrosis. In this study, our results indicated that GAS5 expression was reduced in mouse, rat, and human fibrotic liver samples and in activated HSCs. Overexpression of GAS5 suppressed the activation of primary HSCs in vitro and alleviated the accumulation of collagen in fibrotic liver tissues in vivo. We identified GAS5 as a target of microRNA-222 (miR-222) and showed that miR-222 could inhibit the expression of GAS5. Interestingly, GAS5 could also repress miR-222 expression. A pulldown assay further validated that GAS5 could directly bind to miR-222. As a competing endogenous RNAs, GAS5 had no effect on primary miR-222 expression. In addition, GAS5 was mainly localized in the cytoplasm. Quantitative RT-PCR further demonstrated that the copy numbers of GAS5 per cell are higher than those of miR-222. GAS5 increased the level of p27 protein by functioning as a competing endogenous RNA for miR-222, thereby inhibiting the activation and proliferation of HSCs. Taken together, a new regulatory circuitry in liver fibrosis has been identified in which RNAs cross-talk by competing for shared microRNAs. Our findings may provide a new therapeutic strategy for liver fibrosis.

Ghrelin Attenuated Lipotoxicity via Autophagy Induction and Nuclear Factor-κB Inhibition.

BACKGROUND/AIMS: Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide. Autophagy is associated with NAFLD. Ghrelin is a gut hormone with various functions including energy metabolism and inflammation inhibition. We investigated the therapeutic effect of ghrelin on NAFLD and its association with autophagy. METHODS: C57bl/6 mice were fed a high-fat diet for 8 weeks to induce a model of chronic NAFLD, with ghrelin (10 µg/kg) administrated subcutaneously twice weekly from weeks 6 to 8. LO2 cells were pretreated with ghrelin (10(-8) M) before stimulation with free fatty acid (palmitic and oleic acids; 1 mM). Lipid droplets were identified by hematoxylin and eosin and Red O staining and quantified by triglyceride test kits. LC3I/II, an important biomarker protein of autophagy was detected by western blotting, real-time polymerase chain reaction, immunohistochemistry and immunofluorescence. Tumor necrosis factor (TNF)-α and interleukin (IL)-6 were detected by ELISA and immunohistochemistry. Nuclear factor (NF)-κB p65 was detected by western blotting and immunofluorescence. AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) were detected by western blotting. RESULTS: Ghrelin reduced the triglyceride content in high fat diet (HFD) group in vivo and free fatty acid (FFA) group in vitro. TNF-α and IL-6 were significantly reduced in the ghrelin-treated mice compared with the control group. Autophagy induction was accompanied with intracellular lipid reduction in ghrelin-treated mice. Ghrelin upregulated autophagy via AMPK/mTOR restoration and inhibited translocation of NF-κB into the nucleus. CONCLUSIONS: The results indicate that ghrelin attenuates lipotoxicity by autophagy stimulation and NF-κB inhibition.

Long non-coding RNA APTR promotes the activation of hepatic stellate cells and the progression of liver fibrosis.

In this study, we aimed at assessing a role of Alu-mediated p21 transcriptional regulator (APTR) in hepatofibrogenesis. APTR was upregulated in fibrotic liver samples and activated hepatic stellate cells (HSCs). Knockdown of APTR inhibited the activation of HSCs in vitro and mitigated the accumulation of collagen in vivo. Importantly, APTR silencing could abrogate TGF-ß1-induced upregulation of α-SMA in HSCs. In addition, inhibition of cell cycle and cell proliferation by APTR knockdown was attenuated by p21 siRNA1 in primary HSCs. Finally, serum APTR levels were increased in patients with liver cirrhosis, indicating a potential biomarker for liver cirrhosis. Collectively, evidence is proposed for a new biological role of APTR in hepatofibrogenesis.

Ghrelin ameliorates intestinal barrier dysfunction in experimental colitis by inhibiting the activation of nuclear factor-kappa B.

AIM: This study aimed to investigate the effect and underlying mechanism of ghrelin on intestinal barrier dysfunction in dextran sulfate sodium (DSS)-induced colitis. METHODS AND RESULTS: Acute colitis was induced in C57BL/6J mice by administering 2.5% DSS. Saline or 25, 125, 250 µg/kg ghrelin was administrated intraperitoneally (IP) to mice 1 day before colitis induction and on days 4, 5, and 6 after DSS administration. IP injection of a ghrelin receptor antagonist, [D-lys(3)]-GHRP-6, was performed immediately prior to ghrelin injection. Ghrelin (125 or 250 µg/kg) could reduce the disease activity index, histological score, and myeloperoxidase activities in experimental colitis, and also prevented shortening of the colon. Ghrelin could prevent the reduction of transepithelial electrical resistance and tight junction expression, and bolstered tight junction structural integrity and regulated cytokine secretion. Ultimately, ghrelin inhibited nuclear factor kappa B (NF-κB), inhibitory κB-α, myosin light chain kinase, and phosphorylated myosin light chain 2 activation. CONCLUSIONS: Ghrelin prevented the breakdown of intestinal barrier function in DSS-induced colitis. The protective effects of ghrelin on intestinal barrier function were mediated by its receptor GHSR-1a. The inhibition of NF-κB activation might be part of the mechanism underlying the effects of ghrelin that protect against barrier dysfunction.

Ghrelin Attenuates Liver Fibrosis through Regulation of TGF-ß1 Expression and Autophagy.

Ghrelin is a stomach-derived growth hormone secretagogue that promotes various physiological effects, including energy metabolism and amelioration of inflammation. The purpose of this study was to investigate the protective mechanism of ghrelin against liver fibrosis. Liver fibrosis was induced in C57BL/6 mice by intraperitoneal injection of CCl4 (2.0 mL/kg of 10% CCl4 v/v solution in peanut oil) two times per week for eight weeks. Ghrelin (10 µg/kg) was intraperitoneally injected two times per week for eight weeks. A second murine liver fibrosis model was induced by bile duct ligation (BDL) and concurrent ghrelin administration for four weeks. Hematoxylin eosin (H&E), and Masson's trichrome were used to detect pathological changes to liver tissue. Western blotting was used to detect protein levels of transforming growth factor (TGF)-ß1, phosphorylated Smad3 (p-Smad3), I-collage, α-smooth muscle actin (α-SMA), matrix metalloproteinases (MMPs) 2, tissue inhibitor of matrix metalloproteinases (TIMPs) 1, phosphorylated NF-κB (p-NF-κB), and microtubule-associated protein light chain 3 (LC3). In addition, qRT-PCR was used to detect mRNA levels of TGF-ß1, I-collage, α-SMA, MMP2, TIMP1 and LC3, while levels of TGF-ß1, p-Smad3, I-collage, α-SMA, and LC3 were detected immunohistochemically. Levels of aspartate aminotransferase and alanine aminotransferase were significantly decreased by ghrelin treatment. Ghrelin administration also significantly reduced the extent of pathological changes in both murine liver fibrosis models. Expression levels of I-collage and α-SMA in both models were clearly reduced by ghrelin administration. Furthermore, ghrelin treatment decreased protein expression of TGF-ß1 and p-Smad3. The protein levels of NF-κB and LC3 were increased in the CCl4- and BDL-treatment groups but were significantly reduced following ghrelin treatment. In addition, ghrelin inhibited extracellular matrix formation by decreasing NF-κB expression and maintaining the balance between MMP2 and TIMP1. Our results demonstrated that ghrelin attenuates liver fibrosis via inhibition of the TGF-ß1/Smad3 and NF-κB signaling pathways, as well as autophagy suppression.