Protective effect of apelin-13 on ventilator-induced acute lung injury.

BACKGROUND: Mechanical Ventilation (MV) is an essential mechanism of life support in the clinic. It may also lead to ventilator-induced acute lung injury (VILI) due to local alveolar overstretching and/or repeated alveolar collapse. However, the pathogenesis of VILI is not completely understood, and its occurrence and development may be related to physiological processes such as the inflammatory response, oxidative stress, and apoptosis. Some studies have found that the the apelin/APJ axis is an endogenous antagonistic mechanism activated during acute respiratory distress syndrome(ARDS), that can counteract the injury response and prevent uncontrolled lung injury. To indicate that apelin-13 plays a protective role in VILI, an animal model of VILI was established in this study to explore whether apelin-13 can alleviate VILI in rats by inhibiting inflammation, apoptosis and oxidative stress. METHODS: SD rats were divided into four groups: control, high tidal volume, high tidal volume + normal saline and high tidal volume + apelin-13. After tracheotomy, the rats in control maintained spontaneous breathing, and the other rats were connected to the small animal ventilator for 4 h to establish the rat VILI model. The mRNA expression of apelin was measured by real-time quantitative polymerase chain reaction(qRT-PCR), immunofluorescence and Western blotting(WB) were used to detect the expression level of APJ, and WB was used to detect the expression of the apoptotic proteins Bax and bcl-2. The degree of lung injury was evaluated by pathological staining of lung tissue,W/D ratio, and BALF total protein concentration. The expression of inflammatory factors(IL-1ß, IL-6, TNF-α) in alveolar lavage fluid was measured using ELISA. The activities of MPO and cat and the content of MDA, an oxidative product, in lung tissue were measured to evaluate the degree of oxidative stress in the lung. RESULTS: After treatment with apelin-13, the apelin/APJ axis in the lung tissue of VILI model rats was activated, and the effect was further enhanced. The pathological damage of lung tissue was alleviated, the expression of the antiapoptotic protein Bcl-2 and the proapoptotic protein Bax was reversed, and the levels of the inflammatory cytokines IL-1ß, IL-6, TNF-α levels were all decreased. MPO activity and MDA content decreased, while CAT activity increased. CONCLUSION: The apelin/apj axis is activated in VILI. Overexpression of apelin-13 further plays a protective role in VILI, mainly by including reducing pathological damage, the inflammatory response, apoptosis and antioxidant stress in lung tissue, thus delaying the occurrence and development of VILI.

Effect and mechanical mechanism of spontaneous breathing on oxygenation and lung injury in mild or moderate animal ARDS.

OBJECTIVE: The present study aimed to determine the effect and mechanical mechanism of spontaneous breathing during mechanical ventilation on oxygenation and lung injury using Beagles dogs mild or moderate acute respiratory distress syndrome (ARDS) model. METHODS: After inducing mild or moderate ARDS by infusion of oleic acid, Eighteen Beagles dogs were randomly split into Spontaneous breathing group (BIPAPSB, n = 6), and Complete muscle paralysis group (BIPAPPC, n = 6),Six Beagles without ventilator support comprised the control group. Both groups were ventilated for 8 h under BIPAP mode. High-pressure was titrated TV to 6 ml/kg. A multi-pair esophageal balloon electrode catheter was used to measure respiratory mechanics and electromyogram. End-expiratory lung volume (EELV), gas exchange and respiratory variables were recorded in the process of mechanical ventilation. The contents of Interleukin (IL)-6 and IL-8 in lung tissue were measure using qRT-PCR. Besides, lung injury score was calculated in the end of mechanical ventilation. RESULTS: Based on the comparable setting of ventilator, BIPAPSB group exhibited higher safety peak transpulmonary pressure, abdominal pressure, EELV and P/F(PaO2/FiO2) than BIPAPPC group, whereas mean transpulmonary pressure, the mRNA levels of the IL-6 and IL-8 in the lung tissues and lung injury score in BIPAPSB group were lower than those in BIPAPPC group. CONCLUSION: In mild to moderate ARDS animal models, during mechanical ventilation, SB may improve respiratory function and reduce ventilator-induced lung injury. The mechanism may be that spontaneous inspiration up-regulates peak transpulmonary pressure and EELV; Spontaneous expiration decreases mean transpulmonary pressure by up-regulating intra-abdominal pressure, thereby reducing stress and strain.

c-Fos is a mechanosensor that regulates inflammatory responses and lung barrier dysfunction during ventilator-induced acute lung injury.

BACKGROUND: As one of the basic treatments performed in the intensive care unit, mechanical ventilation can cause ventilator-induced acute lung injury (VILI). The typical features of VILI are an uncontrolled inflammatory response and impaired lung barrier function; however, its pathogenesis is not fully understood, and c-Fos protein is activated under mechanical stress. c-Fos/activating protein-1 (AP-1) plays a role by binding to AP-1 within the promoter region, which promotes inflammation and apoptosis. T-5224 is a specific inhibitor of c-Fos/AP-1, that controls the gene expression of many proinflammatory cytokines. This study investigated whether T-5224 attenuates VILI in rats by inhibiting inflammation and apoptosis. METHODS: The SD rats were divided into six groups: a control group, low tidal volume group, high tidal volume group, DMSO group, T-5224 group (low concentration), and T-5224 group (high concentration). After 3 h, the pathological damage, c-Fos protein expression, inflammatory reaction and apoptosis degree of lung tissue in each group were detected. RESULTS: c-Fos protein expression was increased within the lung tissue of VILI rats, and the pathological damage degree, inflammatory reaction and apoptosis in the lung tissue of VILI rats were significantly increased; T-5224 inhibited c-Fos protein expression in lung tissues, and T-5224 inhibit the inflammatory reaction and apoptosis of lung tissue by regulating the Fas/Fasl pathway. CONCLUSIONS: c-Fos is a regulatory factor during ventilator-induced acute lung injury, and the inhibition of its expression has a protective effect. Which is associated with the antiinflammatory and antiapoptotic effects of T-5224.

Biphasic positive airway pressure spontaneous breathing attenuates lung injury in an animal model of severe acute respiratory distress syndrome.

OBJECTIVE: To compare the effects of unassisted spontaneous breathing (SB) and complete muscle paralysis (PC) on early severe acute respiratory distress syndrome (ARDS) in an animal model, and to explore the possibility of biphasic positive airway pressure (BIPAP) as lung protective ventilation support for patients in the early stage of severe ARDS. METHODS: Twelve healthy beagle dogs between the ages of 10 and 15 months were randomly divided into two groups: the SB group (BIPAPSB) and the PC group (BIPAPPC). Arterial blood samples were drawn before modelling. Arterial blood gas analysis and mechanical tests were conducted. The animal model of severe ARDS was established using a deep intravenous injection of oleic acid, and BIPAP ventilation was performed for 8 hours. Lung tissue and blood were taken to detect lung function, inflammatory reactions and degree of pathological damage. RESULTS: At the beginning of the experiment, there was no significant difference in the arterial blood gas analysis between the two groups (p > 0.05). After successful modelling, the oxygenation index and the end-expiratory lung volume in the SB group were significantly higher than those in the PC group 8 hours after MV. Pathologically, the wet-dry ratio and pathological score of the PC group were higher than those of the SB group; the lung injury in the gravity-dependent area in the SB group was less than that in the PC group (p< 0.05). CONCLUSIONS: In the early stage of severe ARDS induced by oleic acid, compared with PC, retention of the BIPAP mode of SB can reduce the risk of lung injury and improve respiratory function.

Ultra-High Sensitivity Ultrasonic Sensor with an Extrinsic All-Polymer Cavity.

An ultra-high sensitivity ultrasonic sensor with an extrinsic all-polymer cavity is presented. The probe is constructed with a polymer ferrule and a polymer-based reflection diaphragm. A specially designed polymer cover is used to seal the cavity sensor head and apply pretension to the sensing diaphragm. It can be manufactured by a commercial 3D printer with good reproducibility. Due to its all-polymer structure and high coherence depth, the sensitivity of our proposed sensor is improved significantly compared with that of the other sensor structures. Its sensitivity is 189 times as great as that of the commercial standard ultrasonic sensor at the ultrasonic frequency of 50 KHz, and it has a good response to ultrasonic within the frequency range of 18.5 KHz-200 KHz.

Analysis of the Output Characteristics of a Novel Small-Angle Transducer Used in High-Precision Inertial Sensors.

This paper presents the design of a novel small-angle transducer characterized by a simple structure, fast response and very low reaction torque. A theoretical model is presented which describes the linear relationship between the output voltage and the angular displacement when the rotor rotates away from the null position. By analysis of the theoretical model, it is revealed that the small-angle transducer possesses a very high linearity within ±4° and a high sensitivity (approximately 0.34 V/°), and the parameters affecting output characteristics can be obtained. Furthermore, it is found that the transducer sensitivity can be improved by optimizing the load impedance and excitation frequency. These findings are verified by numerical evaluations. In addition, the established theoretical model and simulation analysis provide a quantitative method for analyzing the output characteristics of the novel small-angle transducer.

Hydrogeochemical and Isotopic Indicators of Hydraulic Fracturing Flowback Fluids in Shallow Groundwater and Stream Water, derived from Dameigou Shale Gas Extraction in the Northern Qaidam Basin.

Most of the shale gas production in northwest China is from continental shale. Identifying hydrogeochemical and isotopic indicators of toxic hydraulic fracturing flowback fluids (HFFF) has great significance in assessing the safety of drinking water from shallow groundwater and streamwater. Hydrogeochemical and isotopic data for HFFF from the Dameigou shale formations (Cl/Br ratio (1.81 × 10-4-6.52 × 10-4), Ba/Sr (>0.2), δ11B (-10-1‰), and εSWSr (56-65, where εSWSr is the deviation of the 87Sr/86Sr ratio from that of seawater in parts per 104)) were distinct from data for the background saline shallow groundwater and streamwater before fracturing. Mixing models indicated that inorganic elemental signatures (Br/Cl, Ba/Sr) and isotopic fingerprints (δ11B, εSWSr) can be used to distinguish between HFFF and conventional oil-field brine in shallow groundwater and streamwater. These diagnostic indicators were applied to identify potential releases of HFFF into shallow groundwater and streamwater during fracturing, flowback and storage. The monitored time series data for shallow groundwater and streamwater exhibit no clear trends along mixing curves toward the HFFF end member, indicating that there is no detectable release occurring at present.

Flexible Arc-Shaped Micro-Fiber Bragg Grating Array Three-Dimensional Tactile Sensor for Fingertip Signals Detection and Human Pulse Monitoring.

A flexible arc-shaped micro-Fiber Bragg Grating (mFBG) array three-dimensional tactile sensor for fingertip signal detection and human pulse monitoring is presented. It is based on a three mFBGs array which is embedded in an arc-shaped poly (dimethylsiloxane) (PDMS) elastomer, which can effectively discriminate the normal force, left force, and right force by monitoring the reflected intensity variation of the three mFBGs. Different from the traditional FBG sensors, this sensor measures force by detecting changes in light intensity, effectively avoiding the wavelength cross-sensitivity impact of temperature variations on the sensor performance. This design strategy simplifies the sensor structure, reduces the system complexity and signal interrogation cost, and enhances reliability and practicality. Through systematic experiments, we successfully validated the sensor's superior performance, achieving a minimum detection force of 0.01 N and providing robust data support for practical applications. In addition, the sensor has been used to monitor human pulse accurately. The successful fabrication and experimental validation of this sensor lay a foundation for its widespread application in fields such as robot perception and human vital signal detection.

Water quality assessment and hydrochemical characteristics of shallow groundwater in eastern Chancheng district, Foshan, China.

Groundwater quality is the critical factor that affects human health and the quality of industrial products in Foshan City, South China. Multivariate statistical techniques, including hierarchical cluster analysis (HCA) and principal component analysis (PCA), were applied to evaluate and interpret the complex groundwater quality in eastern Chancheng district, Foshan City. During the dry and wet seasons, 60% and 11% of the total groundwater samples (respectively) are suitable for drinking purposes; other samples can be used for drinking after being treated for pH, Fe, Mn, Al, NH4, and NO3. Similarly, during the dry and wet seasons, 75% and 33% of the total groundwater samples (respectively) are suitable for industrial purposes; other samples can be used for industrial purposes after being treated for NH4 and NO3. Five principal components are extracted from PCA and used to explain 81.78% of the variance in groundwater. The indicators to groundwater quality assessment are EC, Na, Cl, Fe, Mn, NH4, pH, Eh, PO4, HCO3, and K from PCA. HCA reveals that groundwater samples in the study area can be classified into three groups: one reflecting the interaction of groundwater and sediment medium along with the role of cation exchange; another reflecting the role of anion exchange between phosphate and carbonate; and the final reflecting the reducing environment.

Origins of groundwater nitrate in a typical alluvial-pluvial plain of North China plain: New insights from groundwater age-dating and isotopic fingerprinting.

Identifying nitrate sources and their temporal evolution in different land use is important for the sustainable management of groundwater resources. In this study, groundwater dating (3H-3He and time series of 3H) was combined with chemical and stable isotope analyses to resolve the evolution of nitrate sources and the driving mechanism of nitrate contamination. Approximately 75% of the groundwater samples (collected in 2014 and 2018) had nitrate concentrations exceeding World Health Organization's guideline for drinking water (50 mg/L), and 44% exceeded the groundwater quality standard of China (88.6 mg/L), indicating severe nitrate pollution. The shift of nitrate sources in different land use was identified using stable isotope composition of nitrate and groundwater age. The decreasing median value of δ15N from 10.6‰ to 7.5‰ of dated groundwater in farmland irrigated by clean water indicated the shift of nitrate sources from manure toward the mixing of fertilizer and manure due to the increased application of chemical fertilizers from intensive plant farming since 1980s. Comparably, the trend of increasing δ15N (the median value from 7‰ to 12‰) in farmland irrigated by wastewater might be attributed to the decreasing proportion of industrial wastewater since 2000s. The prevailing sources of nitrate in residential area were manure and sewage, and showed no obvious change along the recharge time. Driven by rapid urbanization, the nitrate sources of land use change area exhibited a marked shift from inorganic fertilizers toward manure and sewage. Principal component analysis (PCA) on nitrate concentrations with multiple parameters indicated nitrogen input in agricultural development and urbanization were the main controlling factors of nitrate contamination in the study area. The study results are a good reference for groundwater management in regions with nitrate source change during the process of rapid urbanization and agricultural intensification. The coupling of chemical, isotopic analyses and groundwater dating proved to be invaluable and should be applied in similar studies of nitrate contamination.

Short Carbon Nanotube Nano-Film-Based Polymer/SiO2 Hybrid Waveguide Micro-Ring Filter.

A polymer/SiO2 hybrid waveguide micro-ring filter based on a short carbon nanotube (CNT) nano-film is presented. The nano-film is coated on the top surface of the polymer waveguide, and both the nano-film and polymer waveguide serve as the core of the device. The film can effectively improve the coherent depth and quality factor values of the filter. Due to the excellent light absorption characteristic and large thermo-optic effect of the CNT nano-film, the filter shows good all-optical tunable performance. Experimental results show that its all-optical wavelength tuning efficiency reaches -0.030 nm/mW, and it is 1.67 times as much as that of the filter without the nano-film. With the CNT nano-film, its thermal sensitivity reaches 25.7 times as much as that of the filter without the film. The device shows the advantages of high tuning efficiency, small size, compact structure, and all-optical control, and it is expected to have potential applications in optical filters, optical switches, optical sensors, spectral analysis, and so on.

Using Compound Specific Isotope Analysis to decipher the 1,2,3-trichloropropane-to-Allyl chloride transformation by groundwater microbial communities.

1,2,3-trichloropropane (TCP), a refractory contaminant, can be reductive dehalogenated to allyl chloride (AC) by microorganisms, which has been shown a potential in situ bioremediation (ISB) strategy for TCP remediation in groundwater. In practice, however, it is hard to monitor the bioreduction extent because the TCP concentrations may also be decreased by non-biodegradation processes. Compound specific isotope analysis (CSIA) can be promising in determining the extent of degradation by quantifying the isotope enrichment factors (ε) of relevant degradation mechanisms. To date, no CSIA study has been reported on TCP degradation. In this study, a novel TCP-to-AC transformation enrichment culture (dominated by Azotobacter, Parabacteroides, Fusibacter, Hydrogenophaga, Trichococcus Desulfovibrio, etc) in the absence of the already identified TCP anaerobic reductive dechlorinating microorganisms (e.g., Dehalogenimonas) was derived from a chlorinated hydrocarbon-contaminated aquifer. A TCP degradation experiment was carried out by adding yeast extract to produce hydrogen as an electron donor. The TCP-to-AC transformation was found to conform to zero-order conversion kinetics with the rate constant 11 ± 0.34 µmol L-1 d-1 during the main biodegradation stage. The bulk carbon isotope enrichment factor (εbulk) of the TCP-to-AC transformation was firstly evaluated as -5.2 ± 0.1‰. This study for the first time characterized the carbon isotope fractionations during TCP biodegradation using a novel enrichment culture, which would provide a promising tool for the incorporation of ISB for TCP removal in the future.

The Wnt/ß-catenin pathway regulates inflammation and apoptosis in ventilator-induced lung injury.

Ventilator-induced lung injury (VILI) may be caused by incorrect mechanical ventilation (MV), and its progression is mainly related to inflammatory reaction, apoptosis, and oxidative stress. The Wnt/ß-catenin pathway can modulate inflammation and apoptosis; however, its role in VILI is unknown. This research aims to explore the role of the Wnt/ß-catenin pathway in VILI. VILI models were established using rats and type II alveolar epithelial (ATII) cells. Glycogen synthase kinase 3ß (GSK-3ß), ß-catenin, and cyclin D1 were determined using western blotting and immunofluorescence. Apoptosis of lung tissues was evaluated using TUNEL, flow cytometry, Bax, and Bcl2 protein. Interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) were detected via enzyme-linked immunosorbent assay (ELISA). Lung pathological injury was evaluated through hematoxylin and eosin (H&E) staining. Lung permeability was evaluated by the ratio of dry to wet weight of lung tissue and the total protein level of bronchoalveolar lavage fluid (BALF). The results showed that GSK-3ß expression was enhanced and ß-catenin expression was diminished in lung tissue under MV. SB216763 increased ß-catenin and cyclin D1 expression by inhibiting GSK-3ß expression and inhibited the inflammatory response and apoptosis of lung, alleviated pulmonary edema and lung tissue permeability, and significantly mitigated lung injury. However, inhibition of ß-catenin expression by MSAB attenuated the anti-inflammatory and antiapoptotic effects of SB216763 in VILI. Overall, the present study demonstrates that the Wnt/ß-catenin pathway activation in MV may play an anti-inflammatory and antiapoptotic role, thereby alleviating lung injury and delaying VILI progression, which may be a key point of intervention in VILI.

Experimental investigation on spontaneous combustion oxidation characteristics and stages of coal with different metamorphic degrees.

Coal spontaneous combustion (CSC) is a major disaster threatening coal mine safety; therefore, the investigation of coal spontaneous combustion and oxidation characteristics has been a hot topic in the long term. In this paper, the experimental temperature programmed system is used to carry out the simulation experiment of coal spontaneous combustion and oxidation of three kinds of coal with different metamorphic degrees under three oxygen concentrations (9%, 15%, 21%). The effects of metamorphic degree and oxygen concentration on coal oxidation characteristics were analyzed, and the variation laws of crossing point temperature, three characteristic point temperature, and apparent activation energy were qualitatively discussed. Finally, coal oxidation reaction stages were evaluated and divided. The results show that the concentrations of CO and C2H4 are negatively correlated with the degree of deterioration but increase with the increase of oxygen concentration. High metamorphic coal corresponds to high crossing point temperature (CPT). The average error between the CPT value calculated from the BM empirical correlation and the experimental data is very small, which is 6.42%. The higher the metamorphic degree of coal, the higher the three characteristic temperature points (critical temperature, xerochasy temperature, and activity temperature). The oxidation process of the three coal samples is divided into four stages: surface oxidation, oxidation self-heating, accelerated oxidation, and deep oxidation. The apparent activation energy of each stage exhibits significant variability, with varying patterns displayed with the degree of metamorphism.

MiR-9a-5p alleviates ventilator-induced lung injury in rats by inhibiting the activation of the MAPK signaling pathway via CXCR4 expression downregulation.

BACKGROUND: Globally, Mechanical ventilation is the most commonly used short-term life support technology. Ventilator-induced lung injury (VILI) is an inflammatory injury caused by mechanical ventilation. MicroRNAs (miRNAs) are considered as new gene regulators that play an important role in lung injury and inflammation. However, the role and mechanism of action of miR-9a-5p in VILI remain unclear. METHODS: Herein, a rat model of VILI was established. To determine the expression levels of miR-9a-5p and CXCR4 mRNA, real-time quantitative polymerase chain reactions (qRT-PCR) were conducted. As well as western blot (WB) and immunofluorescence analyses, we determined the expression of CXCR4, SDF-1 and MAPK signaling pathway-related kinases. Hematoxylin and eosin (H&E) staining and the wet-dry ratio of the lung tissue were used to evaluate organ injury. An enzyme-linked immunosorbent assay (Elisa) and myeloperoxidase (MPO) activity measurements were performed to evaluate the inflammatory response. In addition, double luciferase reporter assays were used to verify the association between miR-9a-5p and CXCR4. RESULTS: The expression of miR-9a-5p was low, whereas that of CXCR4 was high in the lung tissues of VILI rats. The overexpression of miR-9a-5p alleviated the degree of pathological injury in the lung tissues of rats with VILI, downregulating inflammatory cytokine expression and MPO activity. In the VILI rat model, miR-9a-5p targeted the negative regulation of CXCR4, and CXCR4 overexpression to reverse the lung-protective and anti-inflammatory effects of miR-9a-5p overexpression in VILI rats. miR-9a-5p also inhibited the phosphorylation of extracellular signal receptor-activated kinase (ERK), a protein related to the MAPK signaling pathway, by downregulating CXCR4 expression. CONCLUSION: miR-9a-5p can hinder the activation of the MAPK/ERK signaling pathway and reduce inflammatory reactions and lung injury in VILI rats through the targeted regulation of CXCR4 expression. Therefore, miR-9a-5p could serve as an intervention target to supply a new strategy for the care of VILI.

Groundwater is important for the geochemical cycling of phosphorus in rapidly urbanized areas: a case study in the Pearl River Delta.

The fate of phosphorus in groundwater needs to be understood because phosphorus-rich groundwater is discharged into surface water bodies, which causes eutrophication, especially in urbanized areas. The present study investigated the spatial distributions and driving forces related to the groundwater phosphate levels in various aquifers in the Pearl River Delta (PRD), which has undergone three decades of urbanization, as well as the relationship between groundwater phosphate and arsenic was also discussed. The results showed that most of the high-phosphate (>1.53 mg/L) groundwater occurred in granular aquifers. The proportion of high-phosphate groundwater in granular aquifers was more than four times that in fissured aquifers, whereas high-phosphate groundwater was not observed in karst aquifers in the PRD. High-phosphate groundwater primarily occurred in urbanized areas in the PRD, and the proportion of high-phosphate groundwater had a significant positive correlation with the urbanization level. In granular aquifers, reductive environment and alkalization led to enrichment of the groundwater with phosphate. Anthropogenic sources such as wastewater from township-village enterprises (TVE) and animal wastes were the main sources of high-phosphate groundwater in urbanized areas, and the external input of phosphate enriched the groundwater arsenic levels in urbanized areas. By contrast, geogenic sources such as the release of phosphate from the reduction of Fe/Mn (hydr)oxides and the seawater intrusion accompanied by the release of phosphate from secondary minerals were mainly responsible for the occurrence of high-phosphate groundwater in peri-urban and non-urbanized areas, respectively. The high concentrations of both phosphate and arsenic in groundwater in fissured aquifers were mainly attributed to the infiltration of wastewater from TVEs. In contrast to the granular aquifers, the groundwater Eh and pH conditions were not conductive to the occurrence of high-phosphate groundwater in fissured aquifers.

Overproduction of soluble recombinant transglutaminase from Streptomyces netropsis in Escherichia coli.

A novel microbial transglutaminase (TGase) from the cultural filtrate of Streptomyces netropsis BCRC 12429 (Sn) was purified. The specific activity of the purified TGase was 18.2 U/mg protein with an estimated molecular mass of 38 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis. The TGase gene of S. netropsis was cloned and an open reading frame of 1,242 bp encoding a protein of 413 amino acids was identified. The Sn TGase was synthesized as a precursor protein with a preproregion of 82 amino acid residues. The deduced amino acid sequence of the mature S. netropsis TGase shares 78.9-89.6% identities with TGases from Streptomyces spp. A high level of soluble Sn TGase with its N-terminal propeptide fused with thioredoxin was expressed in E. coli. A simple and efficient process was applied to convert the purified recombinant protein into an active enzyme and showed activity equivalent to the authentic mature TGase.

Messenger RNA as a source of transposase for sleeping beauty transposon-mediated correction of hereditary tyrosinemia type I.

The Sleeping Beauty (SB) transposon system mediates chromosomal integration and stable gene expression when an engineered SB transposon is delivered along with transposase. One concern in the therapeutic application of the SB system is that persistent expression of transposase could result in transposon instability and genotoxicity. Here, we tested the use of transposase-encoding RNA plus transposon DNA for correction of murine fumarylacetoacetate hydrolase (FAH) deficiency. A bi-functional transposon containing both mouse FAH and firefly luciferase sequences was used to track the growth of genetically corrected liver tissue by in vivo bioluminescence imaging after delivery of DNA or RNA as a source of transposase. Supplying SB transposase in the form of RNA resulted in selective repopulation of corrected hepatocytes with stable expression of FAH and luciferase. Plasma succinylacetone and amino acid levels were normalized, suggesting normal liver metabolism of catabolized protein products. Secondary FAH-deficient animals transplanted with hepatocytes (250,000) isolated from primary treated animals survived 2-(2-nitro-4-trifluoro-methylbenzoyl)-1,3-cyclohexanedione (NTBC) withdrawal, gained weight consistently, and demonstrated stable expression of luciferase. We conclude that transposase-encoding messenger RNA (mRNA) can be used to mediate stable non-viral gene therapy, resulting in complete phenotypic correction, and is thus an effective source of recombinase activity for use in human gene therapy.

Postpolypectomy Bleeding Prevention and More Complete Precancerous Colon Polyp Removal With Endoscopic Mucosal Stripping (EMS).

Background and Aims: Postpolypectomy bleeding and incomplete polyp removal are important complication and quality concerns of colonoscopy for colon cancer prevention. We investigated if endoscopic mucosal stripping (EMS) as a technical modification of traditional cold snare polypectomy to avoid submucosal injury during removal of non-pedunculated colon polyps could prevent postpolypectomy bleeding and facilitate complete polyp removal. Methods: This is an Internal Review Board exemption-granted retrospective analysis of 5,142 colonoscopies with snare polypectomy performed by one of the authors (ZJC) at Minnesota Gastroenterology ambulatory endoscopy centers during a 12-year period divided into pre-EMS era (2005-2012, n = 2,973) and EMS era (2013-2016, n = 2169) with systemic adoption of EMS starting 2013. Change in postpolypectomy bleeding rate before and after EMS adoption and EMS polypectomy completeness were evaluated. Results: Zero postpolypectomy bleeding case was found during EMS era (rate 0%) compared with 10 bleeding cases during pre-EMS era (rate 0.336%). This difference was statistically significant (P = 0.0055) and remained so after excluding 2 bleeding cases of pedunculated polyps (P = 0.012). All bleeding cases involved hot snare polypectomy. Histological examination of the involved polyps showed substantial submucosal vascular damage in contrast to a remarkable paucity of submucosa in comparable advanced polyps removed using EMS. Both biopsy and follow-up colonoscopy examination of the polypectomy sites confirmed that EMS more completely removed non-pedunculated advanced polyps. Conclusions: EMS polypectomy was effective in preventing postpolypectomy bleeding and facilitated complete polyp removal.