BMP7-Loaded Human Umbilical Cord Mesenchymal Stem Cell-Derived Small Extracellular Vesicles Ameliorate Liver Fibrosis by Targeting Activated Hepatic Stellate Cells.

Purpose: Human umbilical cord mesenchymal stem cell (hucMSC)-derived small extracellular vesicles (sEVs) are natural nanocarriers with promising potential in treating liver fibrosis and have widespread applications in the fields of nanomedicine and regenerative medicine. However, the therapeutic efficacy of natural hucMSC-sEVs is currently limited owing to their non-specific distribution in vivo and partial removal by mononuclear macrophages following systemic delivery. Thus, the therapeutic efficacy can be improved through the development of engineered hucMSC-sEVs capable to overcome these limitations. Patients and Methods: To improve the anti-liver fibrosis efficacy of hucMSC-sEVs, we genetically engineered hucMSC-sEVs to overexpress the anti-fibrotic gene bone morphogenic protein 7 (BMP7) in parental cells. This was achieved using lentiviral transfection, following which BMP7-loaded hucMSC-sEVs were isolated through ultracentrifugation. First, the liver fibrosis was induced in C57BL/6J mice by intraperitoneal injection of 50% carbon tetrachloride (CCL4) twice a week for 8 weeks. These mice were subsequently treated with BMP7+sEVs via tail vein injection, and the anti-liver fibrosis effect of BMP7+sEVs was validated using small animal in vivo imaging, immunohistochemistry (IHC), tissue immunofluorescence, and enzyme-linked immunosorbent assay (ELISA). Finally, cell function studies were performed to confirm the in vivo results. Results: Liver imaging and liver histopathology confirmed that the engineered hucMSC-sEVs could reach the liver of mice and aggregate around activated hepatic stellate cells (aHSCs) with a significantly stronger anti-liver fibrosis effect of BMP7-loaded hucMSC-sEVs compared to those of blank or negative control-transfected hucMSC-sEVs. In vitro, BMP7-loaded hucMSC-sEVs promoted the phenotypic reversal of aHSCs and inhibited their proliferation to enhance the anti-fibrotic effects. Conclusion: These engineered BMP7-loaded hucMSC-sEVs offer a novel and promising strategy for the clinical treatment of liver fibrosis.

In vivo and in vitro performances of chitosan-coated Mg-Zn-Zr-Gd-Ca alloys as bone biodegradable materials in rat models.

Mg alloys have attracted significant attention as promising biomedical materials, specifically as fixation materials for promoting fracture healing. However, their unsatisfactory corrosion resistances hinder further clinical applications and thus require attention. This study aims to determine the performance of novel chitosan-coated Mg-1Zn-0.3Zr-2Gd-1Ca alloy and its ability to promote the healing of osteoporotic fractures. Moreover, its corrosion resistance and biocompatibility were assessed. Performance degradations of the samples were measured via electrochemical tests, weight loss test and morphological analysis, and the uncoated and chitosan-coated fixations were compared based on their effects on biocompatibility via the cytotoxicity test, X-rays, and hematoxylin and eosin staining. The effect of bone growth and healing was investigated via immunohistochemical test. Results of the electrochemical tests indicated that compared with the bare body, chitosan-coated Mg-Zn-Ca-Zr-Gd alloys improved by one order of magnitude. Additionally, scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), and weight loss test demonstrated that the corrosion resistance of the chitosan-coated Mg alloy is better than that of the uncoated alloy. In addition, cytotoxicity analysis indicated that the viability and morphology of the chitosan-coated alloy groups were superior to the uncoated groups in vitro. During in vivo analysis, chitosan-coated and uncoated Mg-1Zn-0.3Zr-2Gd-1Ca alloys were implanted into ovariectomized SD female rats with osteoporotic fractures for 1, 2, and 3 weeks. No displacement and shedding were observed through X-rays, and pathological analyses proved that the material was not harmful for liver and kidney tissues. Immunohistochemistry revealed that the chitosan-coated Mg-Zn-Ca-Zr-Gd alloy material contributed to the healing of osteoporotic fractures in the SD rat models. In conclusion, this study demonstrated the chitosan-coated Mg-Zn-Ca-Zr-Gd alloys have improved corrosion resistance and biocompatibility. Moreover, the alloy was found to accelerate the healing of osteoporotic fractures in SD rat models. Therefore, it has significant potential as a fixation material for osteoporotic fractures.

Biodegradable magnesium alloy with eddy thermal effect for effective and accurate magnetic hyperthermia ablation of tumors.

Magnetic hyperthermia therapy (MHT) is able to ablate tumors using an alternating magnetic field (AMF) to heat up magnetocaloric agents (e.g. magnetic nanoparticles) administered into the tumors. For clinical applications, there is still a demand to find new magnetocaloric agents with strong AMF-induced heating performance and excellent biocompatibility. As a kind of biocompatible and biodegradable material, magnesium (Mg) and its alloys have been extensively used in the clinic as an implant metal. Herein, we discovered that the eddy thermal effect of the magnesium alloy (MgA) could be employed for MHT to effectively ablate tumors. Under low-field-intensity AMFs, MgA rods could be rapidly heated, resulting in a temperature increase in nearby tissues. Such AMF-induced eddy thermal heating of MgA could not only be used to kill tumor cells in vitro, but also be employed for effective and accurate ablation of tumors in vivo. In addition to killing tumors in mice, we further demonstrated that VX2 tumors of much larger sizes growing in rabbits after implantation of MgA rods could also be eliminated after exposure to an AMF, illustrating the ability of MgA-based MHT to kill large-sized tumors. Moreover, the implanted MgA rods showed excellent biocompatibility and ∼20% of their mass was degraded within three months. Our work thus discovered for the first time that non-magnetic biodegradable MgA, an extensively used implant metal in clinic, could be used for effective magnetic thermal ablation of tumors under a low-field-intensity AMF. Such a strategy could be readily translated into clinical use.

Corrosion and biocompatibility behaviours of microarc oxidation/phytic acid coated magnesium alloy clips for use in cholecystectomy in a rabbit model.

With the popularisation of laparoscopic cholecystectomy, ligation clips have been commonly used for ligating the cystic duct and cystic artery. However, non-degradable clips remain in the body long-term, which significantly increases the risk of the clip becoming detached. Thus, magnesium alloys have attracted tremendous attention owing to their biodegradability and good biocompatibility. However, the poor corrosion resistance hinders the clinical application of magnesium alloys with microarc oxidation/phytic acid (MAO/PA) composite coatings as protective coatings. Here, these alloys were used to hinder the rapid material degradation in aqueous solution. Electrochemical tests were conducted to evaluate the in vivo degradation behaviour in simulated body fluid (SBF) for Mg-Zn-Y-Nd alloys, and scanning electron microscopy (SEM) was used to observe the micromorphology of in vivo clip degradation. Cell toxicity, cell adhesion, and flow cytometry were performed in vitro to detect cytocompatibility. Biochemical detection of serum magnesium, serum creatinine (CREA), blood urea nitrogen (BUN), alanine transaminase (ALT), and alanine aminotransferase (AST), and haematoxylin-eosin (HE) staining of the heart, liver, and kidney tissues in vivo was conducted to determine the biocompatibility properties after surgery. Electrochemical measurements and SEM images revealed that the MAO/PA-coated magnesium alloy delayed corrosion in SBF. The apoptosis rate increased slightly with increased extract concentration. Nevertheless, MAO/PA-coated magnesium alloys still exhibited good cytocompatibility. No obvious abnormality was observed in the blood biochemical test or HE staining. Thus, MAO/PA-coated magnesium alloys exhibit better corrosion than bare magnesium. In addition, Mg-Zn-Y-Nd and MAO/PA-coated magnesium alloys exhibited no cytotoxicity, good adhesion, and biosafety.

Biological behavior exploration of a paclitaxel-eluting poly-l-lactide-coated Mg-Zn-Y-Nd alloy intestinal stent in vivo.

As a new type of intestinal stent, the MAO/PLLA/paclitaxel/Mg-Zn-Y-Nd alloy stent has shown good degradability, although its biocompatibility in vitro and in vivo has not been investigated in detail. In this study, its in vivo biocompatibility was evaluated by animal study. New Zealand white rabbits were implanted with degradable intestinal Mg-Zn-Y-Nd alloy stents that were exposed to different treatments. Stent degradation behavior was observed both macroscopically and using a scanning electron microscope (SEM). Energy dispersion spectrum (EDS) and histological observations were performed to investigate stent biological safety. Macroscopic analysis showed that the MAO/PLLA/paclitaxel/Mg-Zn-Y-Nd stents could not be located 12 days after implantation. SEM observations showed that corrosion degree of the MAO/PLLA/paclitaxel/Mg-Zn-Y-Nd stents implanted in rabbits was significantly lower than that in the PLLA/Mg-Zn-Y-Nd stent group. Both histopathological testing and serological analysis of in vivo biocompatibility demonstrated that the MAO/PLLA/paclitaxel/Mg-Zn-Y-Nd alloy stents could significantly inhibit intestinal tissue proliferation compared to the PLLA/Mg-Zn-Y-Nd alloy stents, thus providing the basis for designing excellent biodegradable drug stents.

Correction: Biological behavior exploration of a paclitaxel-eluting poly-l-lactide-coated Mg-Zn-Y-Nd alloy intestinal stent in vivo.

[This corrects the article DOI: 10.1039/C9RA10156J.].

Greenhouse gases emission from the sewage draining rivers.

Carbon dioxide (CO2), methane (CH4) and nitrous oxide (N2O) concentration, saturation and fluxes in rivers (Beitang drainage river, Dagu drainage rive, Duliujianhe river, Yongdingxinhe river and Nanyunhe river) of Tianjin city (Haihe watershed) were investigated during July and October in 2014, and January and April in 2015 by static headspace gas chromatography method and the two-layer model of diffusive gas exchange. The influence of environmental variables on greenhouse gases (GHGs) concentration under the disturbance of anthropogenic activities was discussed by Spearman correlative analysis and multiple stepwise regression analysis. The results showed that the concentration and fluxes of CO2, CH4 and N2O were seasonally variable with >winter>fall>summer, spring>summer>winter>fall and summer>spring>winter>fall for concentrations and spring>summer>fall>winter, spring>summer>winter>fall and summer>spring>fall>winter for fluxes respectively. The GHGs concentration and saturation were higher in comprehensively polluted river sites and lower in lightly polluted river sites. The three GHGs emission fluxes in two sewage draining rivers of Tianjin were clearly higher than those of other rivers (natural rivers) and the spatial variation of CH4 was more obvious than the others. CO2 and N2O air-water interface emission fluxes of the sewage draining rivers in four seasons were about 1.20-2.41 times and 1.13-3.12 times of those in the natural rivers. The CH4 emission fluxes of the sewage draining rivers were 3.09 times in fall to 10.87 times in spring of those in the natural rivers in different season. The wind speed, water temperature and air temperature were related to GHGs concentrations. Nitrate and nitrite (NO3-+NO2--N) and ammonia (NH4+-N) were positively correlated with CO2 concentration and CH4 concentration; and dissolved oxygen (DO) concentration was negatively correlated with CH4 concentration and N2O concentration. The effect of human activities on carbon and nitrogen cycling in river is great.

A novel approach to cloning and expression of human thymidylate synthase.

Thymidylate synthase (TS) catalyzes the transfer of a methyl group from methylenetetrahydrofolate to dUMP to form dTMP. It is a primary target in the chemotherapy of colorectal cancers and some other neoplasms. In order to obtain pure protein for analysis of structure and biological function, an expression vector TS-pET28b (+) was constructed by inserting wild-type human thymidylate synthase (hTS) cDNA into pET28b (+). Then an expression strain was selected after transformation of the recombined plasmid into Rosetta (DE3). Fusion protein with His-tag was efficiently expressed in the form of inclusion bodies after IPTG induction and the content was approximately 40.0% of total bacteria proteins after optimizing expression conditions. When inclusion bodies were washed, dissolved and purified by Ni-NTA under denatured conditions, the purity was up to 90%. On SDS-PAGE and West-blotting, the protein band was found to match well with the predicted relative molecular mass-36kDa. Bioactivity was 0.1 U/mg. The results indicated that high-level expression of wild-type hTS cDNA can be achieved in prokaryotes with our novel method, facilitating research into related chemotherapy.