Construction of an Infectious DNA Clone of Grapevine Geminivirus A Isolate GN and Its Biological Activity in Plants Analyzed Using an Efficient and Simple Inoculation Method.

The pathogenicity of grapevine geminivirus A (GGVA), a recently identified DNA virus, to grapevine plants remains largely unclear. Here, we report a new GGVA isolate (named GGVAQN) obtained from grapevine 'Queen Nina' plants with severe disease symptoms. The infectious clone of GGVAQN (pXT-GGVAQN) was constructed to investigate its pathogenicity. Nicotiana benthamiana plants inoculated with GGVAQN by agroinfiltration displayed upward leaf curling and chlorotic mottling symptoms. A simple, quick, and efficient method for delivering DNA clones of GGVAQN into grapevine plants was developed, by which Agrobacterium tumefaciens cells carrying pXT-GGVAQN were introduced into the roots of in vitro-grown 'Red Globe' grape plantlets with a syringe. By this method, all 'Red Globe' grape plants were systemically infected with GGVAQN, and the plants exhibited chlorotic mottling symptoms on their upper leaves and downward curling, interveinal yellowing, and leaf-margin necrosis symptoms on their lower leaves. Our results provide insights into the pathogenicity of GGVA and a simple and efficient inoculation method to deliver infectious viral clones to woody perennial plants.

Mulberry fruit repairs alcoholic liver injury by modulating lipid metabolism and the expression of miR-155 and PPARα in rats.

As alcohol consumption increases, alcoholic liver disease (ALD) has become more popular and is threating our human life. In this study, we found mulberry fruit extract (MFE) repaired alcohol-caused liver diseases by regulating hepatic lipid biosynthesis pathway and oxidative singling in alcoholically liver injured (ALI) rats. MFE administration inhibited hepatic lipid accumulation and improved liver steatosis in ALI rats. MFE also enhanced the antioxidant capacity and alleviated the inflammatory response by increasing the activities of antioxidant enzymes and decreasing the contents of interleukin (IL)-1ß and tumor necrosis factor (TNF)-α. Additionally, MFE regulated the expression of miRNA-155 and lipid metabolism-related PPARα protein in rats. Both miR-155 and PPARα play important roles in liver function. The results indicate that MFE has hepatoprotective effects against ALI in rats.

Sesquiterpenoids from Artemisia argyi and their NO production inhibitory activity in RAW264.7 cells.

Investigation into the chemical diversity of Artemisia argyi led to the discovery of two new (1, 4) and four known (2-3, 5-6) sesquiterpenoids. The new structures were determined via extensive spectroscopic data, including IR, UV, MS, and NMR, and the absolute configurations of these compounds were elucidated by calculated ECD method. All isolates were tested for their inhibitory activity against NO production in RAW 264.7 macrophages, and the isolated sesquiterpenoids exhibited NO production inhibitory activity with IC50 values ranging from 1.91 to 36.52 µM.

Characteristics of peptic ulcer bleeding in cirrhotic patients with esophageal and gastric varices.

Upper gastrointestinal bleeding (UGIB) is common in liver cirrhosis. Although esophageal and gastric varices (EGV) is the main bleeding source, there were still a proportion of patients with peptic ulcer bleeding. Thus, this study aimed to analyze the characteristic of variceal bleeding and peptic ulcer bleeding in liver cirrhosis. Cirrhotic patients with confirmed UGIB by urgent endoscopy from July 2012 to June 2018 were enrolled, and classified into peptic ulcer bleeding group (n = 248) and variceal bleeding group (n = 402). Clinical and endoscopic characteristics, therapeutic efficacy and prognosis were evaluated, and independent risk factors for 42-day morality were determined. The mean age and gender ratio of peptic ulcer bleeding group were higher than those in variceal bleeding group (55.58 ± 11.37 vs. 52.87 ± 11.57, P < 0.01; 4.51:1 vs. 2.87:1, P = 0.023). Variceal bleeding group most commonly presented as red blood emesis and coffee grounds (67.16%), while peptic ulcer group primarily manifested as melena (62.10%). Hepatocellular carcinoma was more prevalent in peptic ulcer group (141 vs. 119, P < 0.01). Albumin level in variceal bleeding group was lower higher (P < 0.01), but serum bilirubin, creatinine and prothrombin time were significantly higher (all P < 0.01). Success rate of endoscopic hemostasis for variceal bleeding and peptic ulcer bleeding was 89.05% and 94.35% (P = 0.021). Univariate and multivariate analysis identified prothrombin time (P = 0.041, OR [95% CI] 0.884 [0.786-0.995]), MELD score (P = 0.000, OR [95% CI] 1.153 [1.073-1.240]), emergency intervention (P = 0.002, OR [95% CI] 8.656 [2.219-33.764]), hepatic encephalopathy before bleeding (P = 0.003, OR [95% CI] 8.119 [2.084-31.637]) and hepatic renal syndrome before bleeding (P = 0.029, OR [95% CI] 3.877 [1.152-13.045]) as the independent predictors for 42-day mortality. Peptic ulcer bleeding should be distinguished from variceal bleeding by clinical and endoscopic characteristics.

Kupffer cell activation by different microbial lysates: Toll-like receptor-2 plays pivotal role on thromboxane A2 production in mice and humans.

Thromboxane (TX) A2 has been identified as an important intrahepatic vasoconstrictor upon Kupffer cell (KC) activation during infections such as spontaneous bacterial peritonitis (SBP). The study aimed to investigate the role of TLRs in the TXA2 increase in liver nonparenchymal cells and their related mechanisms. Here, we identified TLR-2 as a common pathway for different microbials: microbial lysates including Gram-positive bacteria, Gram-negative bacteria, and fungi all increased TXA2 secretion via activation of TLR-2 in human KCs, accompanied by increased expression and phosphorylation of Myd88-related pathway. Of all TLR agonists, only TLR-1, -2, and -4 agonists increased TXA2 in human KCs. These results were further confirmed by mouse liver nonparenchymal cells. Comparing the effects of TLR-1, -2, and -4 antagonists, only TLR-2 antagonist showed inhibitory effects with all tested microbial lysates. Pretreatment with TLR-2 antagonist in human KCs blocked the secretion of IL-10, CXCL-10, TNF-α, and IL-6 induced by Gram-positive and Gram-negative bacterial stimulation. IL-23 and IL-1ß were only induced by Gram-negative bacteria. Thus, TLR-2 might be a potential marker and an attractive target for future treatment of patients with SBP. In addition, IL-23 and IL-1ß might distinguish early between Gram-positive and Gram-negative SBP.

CO2 or air cholangiography reduces the risk of post-ERCP cholangitis in patients with Bismuth type IV hilar biliary obstruction.

BACKGROUND: Endoscopic biliary stenting by endoscopic retrograde cholangiopancreatography (ERCP) is the most common form of palliation for malignant hilar obstruction. However, ERCP in such cases is associated with a risk of cholangitis. The incidence of post-ERCP cholangitis is particularly high in Bismuth type IV hilar obstruction, and this risk is further increased when the contrast injected for cholangiography is not drained. The present study aims to compare the incidence of cholangitis associated with the use of a contrast agent, air and CO2 for cholangiography in type IV hilar biliary lesions. METHODS: The clinical data of consecutive 70 patients with type IV hilar obstruction, who underwent ERCP from October 2013 to November 2017, were retrospectively analyzed. These patients were divided into three groups based on the agent used for cholangiography: group A, contrast (n = 22); group B, air (n = 18); group C, CO2 (n = 30). These three methods of cholangiography were chronologically separated. Prior to the ERCP, MRCP was obtained from all patients to guide the endoscopic intervention. RESULTS: At baseline, there was no significant difference in terms of the patient's age, gender, symptoms and liver function tests among the three groups (P > 0.05). The complication rates were significantly higher in group A than in groups B and C (63.6% vs. 26.7 and 27.8%, P < 0.05). The incidence of post-ERCP cholangitis was significantly higher in group A (P < 0.05), while the incidence of post-ERCP pancreatitis and bleeding were similar in the three groups. After the ERCP, the mean hospital stay was shorter in groups B and C, when compared to group A (P < 0.05). However, there was no significant difference in the 30-day mortality rate among the three groups (P > 0.05). Furthermore, there was no significant difference between groups B and C in terms of primary end points. CONCLUSION: CO2 or air cholangiography during ERCP for type IV hilar obstruction is associated with reduced risk of post-ERCP cholangitis, when compared to conventional contrast agents.

Early enteral feeding on esophageal cancer patients after esophageal resection and reconstruction.

BACKGROUND: In patients under esophagectomy, early postoperative oral feeding has traditionally been contraindicated to minimize the risk of anastomotic leaks. Because early oral feeding preserves the integrity and function of gut mucosa, the aim of this study was to investigate the impact of postoperative early oral feeding on esophagectomy. METHODS: Between Oct 2013 and Jan 2016, 41 consecutive patients with esophageal carcinoma (stages I- III), who had undergone minimally invasive Ivor-Lewis surgery, were enrolled in this study. The patients were randomly arranged into the early oral feeding (EOF) group (21 cases) and the simple tube feeding (STF) group (20 cases). The nutritional goal for both groups was 25 kcal/(kg·day). The patients in the EOF group were tube-fed with enteral nutrition and orally fed with 5% glucose in normal saline during the first 4 postoperative days, after that the patients were placed on a liquid diet. The patients in the STF group were postoperatively tube-fed with enteral nutrition for 7 days before being switched to liquid diet. The length of the postoperative hospital stay, rate of perioperative complications, and overall mortality were recorded. Preoperative and postoperative levels of serum albumin (ALB), prealbumin (PA), transferrin (TP), and C reactive protein (CRP) were also monitored. RESULTS: There were no significant differences of the incidence of postoperative complications and overall mortality between the EOF group and the STF group, but the duration of hospital stay, interval until the first liquid food, and time of ambulation in the EOF group were lower than those of the STF group (P<0.05). EOF patients also showed significantly lower CRP levels compared with the STF group (P<0.05). CONCLUSIONS: EOF might reduce the duration of hospital stays and CRP levels in esophageal cancer patients who underwent esophagectomy while the mortality and complications were not affected.

Second urgent endoscopy within 48-hour benefits cirrhosis patients with acute esophageal variceal bleeding.

Urgent endoscopy (UE) is important to the diagnosis and treatment of liver cirrhosis patients with esophageal variceal bleeding (EVB). It was reported that a second-look endoscopy may benefit acute upper gastrointestinal bleeding (UGIB) caused by peptic ulcer, while whether it could improve UGIB caused by liver cirrhosis associated EVB remains unclear. This study aimed to investigate the characteristics of second UE for liver cirrhosis with EVB and further examined the potential prognostic factors.Patients aged ≥18 years who underwent UE for EVB within 2 hours after the admission were included and divided into scheduled second-look group (n = 245) and uncontrolled bleeding group (n = 352) based on the indications for second UE within 48 hours after initial endoscopy. Demographic and clinical data were collected and analyzed. Univariate and multivariate analysis were used to identify the risk factors for prognosis. The value of different scoring system was compared.Statistical differences were found on history of bleeding and hepatocellular carcinoma, ascites, endoscopic type of bleeding, between scheduled second-look group and uncontrolled bleeding group. Univariate and multivariate logistic regression analysis confirmed that ascites, hemoglobin <60 g/L, AIMS65 score and failure to identify in initial UE were independent risk factors for bleeding uncontrolled after initial UE, and age, bilirubin level, initial unsatisfactory UE hemostasis, failure to identify bleeding on initial UE and tube/urgent TIPS suggested in initial UE were independent risk factors for 42-day mortality.A second-look UE could bring benefit for liver cirrhosis patients with EVB without increasing the complication rate.

Feasibility of non-intubated anesthesia and regional block for thoracoscopic surgery under spontaneous respiration: a prospective cohort study

Data about the feasibility and safety of thoracoscopic surgery under non-intubated anesthesia and regional block are limited. In this prospective study, 57 consecutive patients scheduled for thoracoscopic surgery were enrolled. Patients were sedated with dexmedetomidine and anesthetized with propofol and remifentanil. Ropivacaine was used for intercostal nerve and paravertebral block. Lidocaine was used for vagal block. The primary outcomes were mean arterial pressure (MAP), heart rate (HR), oxygen saturation, and end-tidal carbon dioxide partial pressure (ETCO2) at T0 (pre-anesthesia), T1 (immediately after laryngeal mask/nasopharyngeal airway placement), T2 (immediately after skin incision), T3 (10 min after opening the chest), T4 (end of surgery), and T5 (immediately after laryngeal mask/nasopharyngeal airway removal). One patient required conversion to intubation, 15 developed intraoperative hypotension, and two had hypoxemia. MAP at T0 and T5 was higher than at T1-T4; MAP at T3 was lower (P<0.05 vs other time points). HR at T0 and T5 was higher (P<0.05 vs other time points). ETCO2 at T2 and T3 was higher (P<0.05 vs other time points). Arterial pH, PCO2, and lactic acid at T1 differed from values at T0 and T2 (P<0.05). The Quality of Recovery-15 (QoR-15) score at 24 h was lower (P<0.05). One patient experienced dysphoria during recovery. Thoracoscopic surgery with regional block under direct thoracoscopic vision is a feasible and safe alternative to conventional surgery under general anesthesia, intubation, and one-lung ventilation.

Feasibility of non-intubated anesthesia and regional block for thoracoscopic surgery under spontaneous respiration: a prospective cohort study.

Data about the feasibility and safety of thoracoscopic surgery under non-intubated anesthesia and regional block are limited. In this prospective study, 57 consecutive patients scheduled for thoracoscopic surgery were enrolled. Patients were sedated with dexmedetomidine and anesthetized with propofol and remifentanil. Ropivacaine was used for intercostal nerve and paravertebral block. Lidocaine was used for vagal block. The primary outcomes were mean arterial pressure (MAP), heart rate (HR), oxygen saturation, and end-tidal carbon dioxide partial pressure (ETCO2) at T0 (pre-anesthesia), T1 (immediately after laryngeal mask/nasopharyngeal airway placement), T2 (immediately after skin incision), T3 (10 min after opening the chest), T4 (end of surgery), and T5 (immediately after laryngeal mask/nasopharyngeal airway removal). One patient required conversion to intubation, 15 developed intraoperative hypotension, and two had hypoxemia. MAP at T0 and T5 was higher than at T1-T4; MAP at T3 was lower (P<0.05 vs other time points). HR at T0 and T5 was higher (P<0.05 vs other time points). ETCO2 at T2 and T3 was higher (P<0.05 vs other time points). Arterial pH, PCO2, and lactic acid at T1 differed from values at T0 and T2 (P<0.05). The Quality of Recovery-15 (QoR-15) score at 24 h was lower (P<0.05). One patient experienced dysphoria during recovery. Thoracoscopic surgery with regional block under direct thoracoscopic vision is a feasible and safe alternative to conventional surgery under general anesthesia, intubation, and one-lung ventilation.

Effects of Portulaca Oleracea Extract on Acute Alcoholic Liver Injury of Rats.

The present study was envisaged to investigate the chemical constituents and the intervention effects of Portulaca oleracea extract (POE) on acute alcoholic liver injury of rats. The chemical composition of POE was detected by high performance liquid chromatography (HPLC). Sixty male Wistar rats were divided into 6 groups: Normal control (NC) group, acute alcoholic liver injury model group (ALI), low, medium and high dose of POE (25, 50, 100 mg/kg) groups and bifendate (BF, 3.75 mg/kg) group. Each group was given by intragastrical administration for 7 days. Alcoholic liver injury was induced in the experimental model by administering 50% ethanol at 8 mL/kg and repeated administration after 6 h, for a period of 7 days. The results showed that pretreatment with POE significantly reduced the ethanol-elevated serum level of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and triglyceride (TG). The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX) in liver were enhanced followed by administration of POE, while the content of nitric oxide (NO) and malondialdehyde (MDA) was found to decrease. Hepatic content of tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) was also reduced by POE treatment. These results indicated that POE could increase the antioxidant capacity and relieve the inflammatory injury of the liver cells induced by ethanol. Meanwhile, in our study, POE reduced the expression of miR-122, acetyl coenzyme A carboxylase (ACC) 1 mRNA and protein and increased the expression of lipoprotein lipase (LPL) mRNA and protein in liver, which indicated that POE could improve the lipid metabolism disorder induced by ethanol. Our findings suggested that POE had protective effects on acute alcoholic liver injury of rats.

Identification of plasma RGS18 and PPBP mRNAs as potential biomarkers for gastric cancer using transcriptome arrays.

Coding and noncoding RNAs serve a crucial role in tumorigenesis. Circulating RNAs have been recognized as a novel category of biomarkers for a variety of physiological and pathological conditions. To identify plasma RNA biomarkers for gastric cancer (GC), a genome-wide transcriptome analysis using GeneChip® Human Transcriptome Array, which contains probe sets covering exons of ~67500 coding and noncoding transcripts of annotated genes, was performed to screen for the RNAs that exhibited differential expression in the plasma samples of patients with GC and controls. The expression levels of 6 candidate RNAs, including regulator of G-protein signaling 18 (RGS18), integral membrane protein 2B, pro-platelet basic protein (PPBP), nucleosome assembly protein1-like 1, n324674 and ENST00000442382 were assessed in the plasma samples of 81 patients with GC and 77 healthy participants using reverse transcription-quantitative polymerase chain reaction. Furthermore, the expression levels of RGS18 and PPBP mRNAs were indicated to be significantly differentially expressed (P<0.0001) in an independent panel of plasma samples of 36 patients with GC compared with 34 healthy participants. The potential association of RGS18 and PPBP mRNA expression levels with clinicopathological features was subsequently analyzed. Receiver operating characteristic analysis indicated that the combination of these 2 mRNAs with an area under curve <0.812 was an improved indicator for gastric cancer compared with respective individual levels. The results of the present study indicate that RGS18 and PPBP mRNA expression was significantly downregulated in the plasma of patients with GC, and the combination of these 2 mRNAs may be a useful diagnostic or prognostic marker for GC.

Early steep decline of liver stiffness predicts histological reversal of fibrosis in chronic hepatitis B patients treated with entecavir.

It is unknown whether dynamic changes of liver stiffness measurement (LSM) can predict the reversibility of fibrosis. Therefore, we evaluated the utility of LSM changes in predicting histological changes of fibrosis in patients with chronic hepatitis B (CHB) on antiviral therapy. In a prospective cohort of CHB patients treated with entecavir, virological measurement and biochemical measurement along with LSM were measured at baseline and every 6 months. Liver biopsies were conducted at baseline and month 18 of treatment. Fibrosis regression was defined by the following two criteria: (a) Ishak score decrease ≥1 stage, (b) Ishak score decrease ≥1 stage or predominantly regressive by post-treatment PIR classification. The dynamic changes of LSM and its predictive value for histological reversibility were evaluated with piecewise linear mixed-effects model and ROC analysis. We found that at month 18 of antiviral therapy, liver fibrosis was reserved in 86 of 212 (40.6%) CHB patients by Ishak reversal criterion. Overall, a decline in LSM was associated with attenuation of Ishak score. The rate of LSM decline in the first 6 months was significantly faster in patients with fibrosis reversal (ΔLSM%Ishak  = -2.19%/month, P = 0.0025; ΔLSM%Ishak/PIR  = -2.56%/month, P = 0.0004). The predictive model based on baseline FIB-4 and Ishak score as well as baseline LSM, PLT, albumin and their changes during the first 6 months could predict histological reversal (AUROCIshak  = 0.74, 95% CI: 0.67-0.80; AUROCIshak/PIR  = 0.81, 95% CI: 0.74-0.87). We conclude that in CHB patients, changes in LSM during the first 6 months of entecavir therapy can predict histological reversibility of liver fibrosis at month 18 of antiviral therapy.

miR-183 and miR-141 in lesion tissues are potential risk factors for poor prognosis in patients with infected abdominal aortic aneurysm.

The expression levels of micro ribonucleic acid-183 (miR-183) and miR-141 in the lesion tissues of infected abdominal aortic aneurysm (IAAA) and their relationship with prognosis were investigated. Thirty-six patients with IAAA admitted and who underwent vascular surgery in People's Hospital of Shenzhen from June 2003 to June 2013 were selected. Reverse transcription polymerase chain reaction (RT-PCR) was utilized to detect the expression levels of miR-183 and miR-141 in lesion tissues and adjacent tissues 1 cm away from the aneurysm in 36 patients with IAAA. The relationship between the expression levels of miR-183 and miR-141 as well as the clinicopathological features of patients with IAAA were analyzed, and the factors influencing the prognosis of IAAA were analyzed by univariate and multiva-riate analysis. The expression levels of miR-183 and miR-141 were significantly downregulated in the lesions of patients with IAAA, and miR-183 and miR-141 levels in the lesion tissues of the IAAA patients were significantly lower than those in the adjacent tissues (P<0.05). The expression levels of miR-183 and miR-141 were not related to sex, age, history of hypertension, and alcoholism (P>0.05), but they were related to smoking history or aneurysm size (P<0.05). The overall survival rate of patients with IAAA was 41.6% (15/36). The multivariate analysis found that aneurysm size, low expression of miR-183, and low expression of miR-141 were independent factors affecting the prognosis of patients with IAAA. In conclusion, the expression levels of miR-183 and miR-141 in the lesion tissues of IAAA are low, and the lower the expression level is, the worse the prognosis gets. miR-183 and miR-141 can be used as predictors of prognosis in patients with IAAA.

Online Monitoring of Intraoperative Exhaled Propofol by Acetone-Assisted Negative Photoionization Ion Mobility Spectrometry Coupled with Time-Resolved Purge Introduction.

Online monitoring of exhaled propofol concentration is important for anesthetists to provide adequate anesthesia as propofol concentrations in plasma and breath are correlated reasonably well. Exhaled propofol could be detected by 63Ni ion mobility spectrometry in negative ion mode; however, the radioactivity of 63Ni source restricts its clinical application due to safety, environmental, and regulatory concerns. An acetone-assisted negative photoionization ion mobility spectrometer (AANP-IMS) using a side-mounted vacuum ultraviolet (VUV) lamp in the unidirectional (UD) flow mode was developed for sensitive measurement of exhaled propofol by producing a high percentage of O2-(H2O) n. An adsorption sampling and time-resolved purge introduction system was developed to eliminate the interference of residual inhaled anesthetic sevoflurane based on their different adsorptions between propofol and sevoflurane on the inwall of the fluorinated ethylene propylene (FEP) sample loop. The effects of the inner diameter and the length of the sample loop on the signal intensity of propofol and the time-resolution between propofol and sevoflurane were theoretically and experimentally investigated. A sample loop with 3 mm i.d. and 150 cm length allowed sensitive measurement of exhaled propofol with a response time of 4 s, a linear response range for propofol was achieved to be 0.2 to 14 ppbv with a limit of detection (LOD) of 60 pptv, and the quantification of propofol was not influenced by the change of the sevoflurane concentration. Finally, the performance of monitoring exhaled propofol during surgery was demonstrated on a patient undergoing laparoscopic distal pancreatectomy combined with cholecystectomy.

Preventive effect of hesperidin modulates inflammatory responses and antioxidant status following acute myocardial infarction through the expression of PPAR­Î³ and Bcl­2 in model mice.

Hesperetin is the main pharmacological ingredient of fruit of the citrus family, rutaceae. It is a flavanone, which has potent antioxidation and anti­inflammatory activities. The present study investigated the preventive effect of hesperidin in the modulation of acute myocardial infarction (AMI)­induced inflammatory responses and antioxidant status in a mouse model. The levels of creatine kinase­MB, tumor necrosis factor (TNF­α), interleukin (IL)­1ß, IL­6, monocyte chemoattractant protein 1 (MCP­1), intercellular adhesion molecule 1 (ICAM­1), malondialdehyde (MDA), catalase (CAT), superoxide dismutase (SOD) and caspase­3/9 were measured using ELISA kits. Western blot analysis analyzed p53 and B­cell lymphoma 2 (Bcl­2)­associated X protein/Bcl­2, and induced the expression of peroxisome proliferator­activated receptor­Î³ (PPAR­Î³). Hesperidin markedly decreased the myocardial infarction area, heart weight/body weight ratio and activity of creatine kinase­MB in AMI mice. Hesperidin treatment caused a significant decrease in the levels of TNF­α, IL­1ß, IL­6, MCP­1, ICAM­1, MDA, CAT, SOD and caspase­3/9 in mice with AMI. Hesperidin also significantly suppressed the protein expression levels of p53 and Bax/Bcl­2, and induced the expression of peroxisome proliferator­activated receptor­Î³ (PPAR­Î³) in mice with AMI. The preventive effect of hesperidin modulated the inflammatory response and antioxidant status following AMI through downregulation of the expression of PPAR­Î³ and Bcl­2 in the model mice.

DNA Modulates the Interaction of Genetically Engineered DNA-Binding Proteins and Gold Nanoparticles: Diagnosis of High-Risk HPV Infection.

Gene detection has an important role in diagnosing several serious diseases and genetic defects in modern clinical medicine. Herein, we report a fast and convenient gene detection method based on the modulation of the interaction between a heat-resistant double-stranded DNA (dsDNA)-binding protein (Sso7d) and gold nanoparticles (Au NPs). We prepared a recombinant Cys-Sso7d, which is Sso7d with an extra cysteine (Cys) residue in the N-terminus, through protein engineering to control the interaction between Sso7d and Au NPs. Cys-Sso7d exhibited a stronger affinity for Au NPs and more easily induced the aggregation of Au NPs than Sso7d. In addition, Cys-Sso7d retained its ability to bind with dsDNA. The aggregation of Au NPs induced by Cys-Sso7d was diminished in the presence of dsDNA, which could be utilized as a transduction mechanism for the detection of the polymerase chain reaction (PCR) products of human papillomavirus (HPV) gene fragments (HPV types 16 and 18). The Cys-Sso7d/Au NP probe could detect as few as 1 copy of the HPV gene. The sensitivity and specificity of the Cys-Sso7d/Au NP probe for Pap smear clinical specimens (n = 52) for HPV 16 and HPV 18 detection were 85.7%/100.0% and 85.7%/91.7%, respectively. Our results demonstrate that the Cys-Sso7d/Au NP probe can be used to diagnose high-risk HPV types in Pap smear samples with high sensitivity, specificity, and accuracy.

CRISPR/Cas9-Mediated CCR5 Ablation in Human Hematopoietic Stem/Progenitor Cells Confers HIV-1 Resistance In Vivo.

Transplantation of hematopoietic stem cells (HSCs) with a naturally occurring CCR5 mutation confers a loss of detectable HIV-1 in the patient, making ablation of the CCR5 gene in HSCs an ideal therapy for an HIV-1 cure. Although CCR5 disruption has been attempted in CD4+ T cells and hematopoietic stem/progenitor cells (HSPCs), efficient gene editing with high specificity and long-term therapeutic potential remains a major challenge for clinical translation. Here, we established a CRISPR/Cas9 gene editing system in human CD34+ HSPCs and achieved efficient CCR5 ablation evaluated in long-term reconstituted NOD/Prkdcscid/IL-2Rγnull mice. The CCR5 disruption efficiency in our system remained robust in secondary transplanted repopulating hematopoietic cells. More importantly, an HIV-1 resistance effect was observed as indicated by significant reduction of virus titration and enrichment of human CD4+ T cells. Hence, we successfully established a CRISPR/Cas9 mediated CCR5 ablating system in long-term HSCs, which confers HIV-1 resistance in vivo. Our study provides evidence for translating CCR5 gene-edited HSC transplantation for an HIV cure to the clinic.

The combination of circulating long noncoding RNAs AK001058, INHBA-AS1, MIR4435-2HG, and CEBPA-AS1 fragments in plasma serve as diagnostic markers for gastric cancer.

BACKGROUND: Suitable diagnostic markers for cancers are urgently required in clinical practice. Long non-coding RNAs, which have been reported in many cancer types, are a potential new class of biomarkers for tumor diagnosis. RESULTS: Five lncRNAs, including AK001058, INHBA-AS1, MIR4435-2HG, UCA1 and CEBPA-AS1 were validated to be increased in gastric cancer tissues. Furthermore, we found that plasma level of these five lncRNAs were significantly higher in gastric cancer patients compared with normal controls. By receiver operating characteristic analysis, we found that the combination of plasma lncRNAs with the area under the curve up to 0.921, including AK001058, INHBA-AS1, MIR4435-2HG, and CEBPA-AS1, is a better indicator of gastric cancer than their individual levels or other lncRNA combinations. Simultaneously, we found that the expression levels of a series of MIR4435-2HG fragments are different in gastric cancer plasma samples, but most of them higher than that in healthy control plasma samples. MATERIALS AND METHODS: LncRNA gene expression profiles were analyzed in two pairs of human gastric cancer and adjacent non-tumor tissues by microarray analysis. Nine gastric cancer-associated lncRNAs were selected and assessed by quantitative real-time polymerase chain reaction in gastric tissues, and 5 of them were further analyzed in gastric cancer patients' plasma. CONCLUSIONS: Our results demonstrate that certain lncRNAs, such as AK001058, INHBA-AS1, MIR4435-2HG, and CEBPA-AS1, are enriched in human gastric cancer tissues and significantly elevated in the plasma of patients with gastric cancer. These findings indicate that the combination of these four lncRNAs might be used as diagnostic or prognostic markers for gastric cancer patients.