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Mesenchymal stem cells (MSCs) represent a promising therapeutic candidate for treating many diseases. However, their proliferation and therapeutic abilities decline during the aging process and disease development. Therefore, fetal MSCs derived from the umbilical cord (UC) attract more attention. Storing and delivering the UC is one critical step for efficient MSC isolation. Although the culture medium-based solution is suitable for UC storage, it is not feasible for large-scale preparation because of its high price. Thus, we demonstrate here that a simple solution containing a pH buffering reagent, calcium, magnesium and glucose could be used as a cost-effective storage solution for UC delivery and efficient MSC isolation.

Objectives Mesenchymal stem cells (MSCs) have been intensively investigated as to their therapeutic potentials. However, the full chemical-defined medium supporting the isolation and expansion of human MSCs has not been developed yet. Materials and methods Here, we developed the full chemical-defined medium, NBVbe medium, via RNA sequencing, bioinformatic analysis, and growth factor screening. Results The NBVbe medium contains N2B27 medium with the BSA (bovine serum albumin) replaced by the recombinant human albumin, bFGF (basic fibroblast growth factor), vitamin C, and EGF (epidermal growth factor). The NBVbe medium could support the isolation and expansion of human MSCs from the umbilical cords. Conclusions The full chemical-defined medium supporting the isolation and expansion of human MSCs has been developed. This would be helpful for further optimization of the MSC medium, their clinical applications, and molecular characterization.

Background pWB980 derived from pUB110 is a promising expression vector in Bacillus for its high copy number and high stability. However, the low transformation rate of recombinant plasmids to the wild cells limited the application of it. On the basis of pWB980, constructing an E. coli–B. subtilis shuttle plasmid could facilitate the transformation rate to Bacillus cells. Because the insertion site for E. coli replication origin sequence (ori) is not unique in pWB980, in order to investigate the best insertion site, eight shuttle plasmids (pUC980-1 ~ pUC980-8) containing all possible insertion sites and directions were constructed. Results The results showed that all the selected insertion sites could be used to construct shuttle plasmid but some sites required a specific direction. And different insertion sites led to different properties of the shuttle plasmids. The best shuttle plasmids pUC980-1 and pUC980-2, which showed copies more than 450 per cell and segregational stabilities up to 98%, were selected for heterologous expressions of an alkaline pectate lyase gene pelN, an alkaline protease spro1 and a pullulanase gene pulA11, respectively. The highest extracellular activities of PelN, Spro1 and PulA11 were up to 5200 U/mL, 21,537 U/mL and 504 U/mL correspondingly after 54 h, 60 h and 48 h fermentation in a 10 L fermentor. Notably, PelN and Spro1 showed remarkably higher yields in Bacillus than previous reports. Conclusion The optimum ori insertion site was the upstream region of BA3-1 in pWB980 which resulted in shuttle plasmids with higher copy numbers and higher stabilities. The novel shuttle plasmids pUC980-1 and pUC980-2 will be promising expression vectors in B. subtilis. Moreover, the ori insertion mechanism revealed in this work could provide theoretical guidance for further studies of pWB980 and constructions of other shuttle plasmids.

Objectives Myocardial infarction (MI) is the most predominant type of cardiovascular diseases with high mortality and morbidity. Stem cell therapy, especially cardiac progenitor cell therapy, has been proposed as a promising approach for cardiac regeneration and MI treatment. Previously, we have successfully generated cardiac progenitor-like cells, induced cardiosphere (iCS), via somatic reprogramming. However, the genome integration characteristic of virus-based reprogramming approach hampered their therapeutic applications due to the risk of tumour formation. In the current study, we aim to establish a safer iCS generation strategy with transgene-free approaches. Materials and methods Four transgene-free approaches for somatic reprogramming, including episome, minicircle, self-replicative RNA, and sendai virus, were compared, from the perspective of cardiac progenitor marker expression, iCS formation, and cardiac differentiation. The therapeutic effects were assessed in the mouse model of MI, from the perspective of survival rate, cardiac function, and structural alterations. Results The self-replicative RNA approach produced more iCS, which had cardiomyocyte differentiation ability and therapeutic effects on the mouse model of MI with comparable levels with endogenous cardiospheres and iCS generated with retrovirus. In addition, the CXCR4 (C-X-C chemokine receptor 4) positive subpopulation of iCS derived cells (iCSDC) delivered by intravenous injection was found to have similar therapeutic effects with intramyocardial injection on the mouse model of MI, representing a safer delivery approach. Conclusion Thus, the optimized strategy for iCS generation is safer and has more therapeutic potentials.

Objectives Myocardial infarction (MI) is the most predominant type of cardiovascular diseases with high mortality and morbidity. Stem cell therapy, especially cardiac progenitor cell therapy, has been proposed as a promising approach for cardiac regeneration and MI treatment. Previously, we have successfully generated cardiac progenitor‐like cells, induced cardiosphere (iCS), via somatic reprogramming. However, the genome integration characteristic of virus‐based reprogramming approach hampered their therapeutic applications due to the risk of tumour formation. In the current study, we aim to establish a safer iCS generation strategy with transgene‐free approaches. Materials and Methods Four transgene‐free approaches for somatic reprogramming, including episome, minicircle, self‐replicative RNA, and sendai virus, were compared, from the perspective of cardiac progenitor marker expression, iCS formation, and cardiac differentiation. The therapeutic effects were assessed in the mouse model of MI, from the perspective of survival rate, cardiac function, and structural alterations. Results The self‐replicative RNA approach produced more iCS, which had cardiomyocyte differentiation ability and therapeutic effects on the mouse model of MI with comparable levels with endogenous cardiospheres and iCS generated with retrovirus. In addition, the CXCR4 (C‐X‐C chemokine receptor 4) positive subpopulation of iCS derived cells (iCSDC) delivered by intravenous injection was found to have similar therapeutic effects with intramyocardial injection on the mouse model of MI, representing a safer delivery approach. Conclusion Thus, the optimized strategy for iCS generation is safer and has more therapeutic potentials., Previously, we and others have successfully reprogrammed mouse fibroblast cells into cardiac progenitor‐like cells. However, several issues should be further addressed, including the reprogramming strategy and delivery approach for myocardial infarction (MI) treatment. In the current study, we screened several transgene‐free approaches for somatic reprogramming. Our data showed the self‐replicative RNA approach produced more iCS, which had cardiomyocyte differentiation ability and therapeutic effects on the mouse model of MI with comparable levels with endogenous cardiospheres and iCS generated with retrovirus. Then, the CXCR4+ (C‐X‐C chemokine receptor 4) subpopulation of iCS derived cells (iCSDC) delivered by intravenous injection was found to have similar therapeutic effects with intramyocardial injection on the mouse model of MI, representing a more safer delivery approach. Thus, the optimized strategy for iCS generation is much safer and has more therapeutic potentials.

Osteosarcoma (OS) is the most common primary malignant tumor of bone. It is a common phenomenon that osteosarcoma cells have a hypoxic microenvironment. Hypoxia can dedifferentiate cells of several malignant tumor types into stem cell-like phenotypes. However, the role of hypoxia in stemness induction and the expression of cancer stem cell (CSC) markers in human osteosarcoma cells has not been reported. The present study examined the effects of hypoxia on stem-like cells in the human osteosarcoma MNNG/HOS cells. Under the incubation with 1% oxygen, the expression of CSCs markers (Oct-4, Nanog and CD133) in MNNG/HOS cells were increased. Moreover, MNNG/HOS cells cultured under hypoxic conditions were more likely to proliferate into spheres and resulted in larger xenograft tumor. Hypoxia also increased the mRNA and protein levels of hypoxia-inducible factor (HIF)-1α. Then rapamycin was used, which has been shown to lower HIF-1α protein level, to inhibit the hypoxic response. Rapamycin suppressed the expression of HIF-1α protein and CSCs markers (Oct4, Nanog and CD133) in MNNG/HOS cells. In addition, pretreatment with rapamycin reduced the efficiency of MNNG/HOS cells in forming spheres and xenograft tumors. The results demonstrated that hypoxia (1% oxygen) can dedifferentiate some of the MNNG/HOS cells into stem cell-like phenotypes, and that the mTOR signaling pathway participates in this process via regulating the expression of HIF-1α protein.

International audience; Pearl River Mouth Basin (PRMB) is a typical episodically rifting basin, and its sediment records exhibit a multistage infilling process. To build the coupling relationship between the spatial variability of tectonic movement and sediment dispersal pattern, we character the source-to-sink system distribution within continental margin and analysis their dominated factors with quantitative statistical methods. First, we identify the faults and calculate the rifting rate by interpreting the two-dimensional (2D) seismic data and zircon or apatite fission-track data (ZFT/AFT). Accordingly, based on the various trending of the rifting rate evolution, we classified rifting process episodically into three patterns: accelerated rifting decelerated rifting, and uniform rifting. Then, we integrated the data available on detrital zircon U–Pb, rare earth element, and heavy mineral assemblage to identify the responding S2S components associated with the above various rifting processes. Results show that in the first tectonic stage during the early Eocene, the sediments were mainly sourced from an intrabasin provenance area, whereas in the late Eocene to the Oligocene, the paleo-Pearl River provided the sediments to the sink area. We restore the facies association of the sink area by studying the core samples, electric well logs, and three-dimensional (3D) seismic data. During the accelerated and uniform rifting processes, coarse-grained deposited along the boundary faults, which includes the fan delta and subaqueous fans. During decelerated rifting, the depositional environment was dominated by braided channels, meandering channels, and tide-influenced channel accretion. Based on the quantitative analysis the relationship between the grain size of the sediment in each S2S segment and the rifting rate, more fine-grained components were preserved in the decelerated rifting system, while coarse-grained sediments fill the accelerated rifting depression and uniform rifting components. Furthermore, the sediment supply and relative height can influence the volume of the S2S system in the uniform rifting system. In the decelerated rifting basin, the sediment supply is the major factor associated with the development of S2S, whereas the relative height is the major factor associated with the accelerated rifting basin.

A high heterologous expression of an alkaline pectate lyase (APL) pelNK93I in E. coli was obtained through optimizing the lactose feeding and fed-batch fermentation. The highest soluble APL activity produced by E. coli BL21 (pET22b-pelNK93I) was 10,181 U/mL which is the highest level so far. On this basis, to improve the extracellular yield of APL, optimized glycine feeding was used to achieve elevated extracellular production of pelNK93I. The highest extracellular APL activity produced by E. coli BL21 (pET22b-pelNK93I) was 6357 U/mL which was also relatively higher than that in previous reports. The final productivity of APL was 282.8 U/mL/h in the fermentation of E. coli BL21 (pET22b-pelNK93I) in a 10 L fermenter. Thus the current study has provided a cost-effective method for the over-expression and preparation of alkaline pectate lyase pelNK93I for its industrial applications. Moreover, pelNK93I (4 U/mL) used for bioscouring increased cottonseed husk removal and radial capillary effect of cotton fabric by 37.63% and 47.06%, respectively, making it a promising enzyme in green textile technology.

Direct ink writing promotes ink formulation design gradually become a hotspot in the application research of nanothermite. In order to obtain an energetic ink with a certain mechanical strength and formability, it is necessary to compound the energetic material with an inert binder. The inert component will not only reduce the sensitivity and energy release efficiency of energetic systems, but also affect its application in micro ignition devices. Based on the thermal gelation behavior of hydroxypropyl methylcellulose (HPMC), the nAl/pCuO/HPMC ink was prepared by HPMC, nano aluminum powder (nAl), and sheet-like porous CuO (pCuO). It has found out that the energetic ink has a favorable printing accuracy when the HPMC content is 7 wt%. To compare the influence from the size and morphology of copper oxide and aluminum on ink printing and reaction performances, the mAl/pCuO/HPMC, nAl/nCuO/HPMC and nAl/mCuO/HPMC inks have been prepared using the same way. Due to the large particle size of the components, nAl/mCuO/HPMC cannot be printed continuously during the printing process. For the mAl/pCuO/HPMC, it cannot form self-sustaining combustion on the glass substrate when the ink is a single layer. The activation energy results show that nAl/pCuO/HPMC has the smallest value (68.39 kJ/mol). The calculated burning rate of nAl/pCuO/HPMC (32.56 cm/s) is about 6 times than that of nAl/nCuO/HPMC system (5.28 cm/s). The ignition performance test shows that only the nAl/pCuO/HPMC system can be successfully ignited by a SCB micro igniter when the HPMC content is 7 wt%. Meanwhile, the pressure test indicates that the nAl/pCuO/HPMC system (7 wt% HPMC, Ф=1.4) has the shortest pressure rising time (46.7 μs), but the largest pressurization rate (1661.7 GPa/s).

Background: pWB980 derived from pUB110 is a promising expression vector in Bacillus for its high copy number and high stability. However, the low transformation rate of recombinant plasmids to the wild cells limited the application of it. On the basis of pWB980, constructing an E. coli - B. subtilis shuttle plasmid could facilitate the transformation rate to Bacillus cells. Because the insertion site for E. coli replication origin sequence ( ori ) is not unique in pWB980, in order to investigate the best insertion site, eight shuttle plasmids (pUC980-1~pUC980-8) containing all possible insertion sites and directions were constructed.Results: The results showed that all the selected insertion sites could be used to construct shuttle plasmid but some sites required a specific direction. And different insertion sites led to different properties of the shuttle plasmids. The best shuttle plasmids pUC980-1 and pUC980-2, which showed copies more than 450 per cell and segregational stabilities up to 98%, were selected for heterologous expressions of an alkaline pectate lyase gene pelN , an alkaline protease spro1 and a pullulanase gene pulA11 , respectively. The highest extracellular activities of PelN, Spro1 and PulA11 were upto 5200 U/mL, 21537 U/mL and 504 U/mL correspondingly after 54 h, 60 h and 48 h fermentation in a 10 L fermentor. Notably, PelN and Spro1 showed remarkably higher yields in Bacillus than previous reports.Conclusion: In this work we can conclude that the optimum ori insertion site was the upstream region of BA3-1 in pWB980 which resulted in shuttle plasmids with higher copy numbers and higher stabilities. The novel shuttle plasmids pUC980-1 and pUC980-2 will be promising expression vectors in B. subtilis . And the ori insertion mechanism revealed in this work could provide theoretical guidance for further studies of pWB980 and constructions of other shuttle plasmids.

Background/Aims: The target genome editing technology not only plays an important role in basic biology studies but also holds a great promise for potential clinical applications. The new generation of engineered nuclease RGEN (RNA Guided EndoNuclease) is much easier to construct and modify, and attracts more attentions. In the current study, we compared different plasmid construction strategies of Cas9-gRNA (guide RNA). Methods: Different plasmid construction strategies of Cas9-gRNA were compared. And more modifications were introduced into the plasmid construction strategy. Results: The plasmid construction efficiency of expressing the gRNA and Cas9 in one plasmid was lower than expressing them in two separate plasmids. However, they showed the similar genome editing efficiency. We further introduced the Golden-gate assembly and blue-white screening approaches into the Cas9-gRNA construction procedures, without the process of vector digestion and gel purification. Conclusions: Combing with the optimized gRNA structure (gRNA-BL) we identified before, we established one more cost-effective, time-saving and efficient plasmid construction strategy for Cas9-gRNA.

LNG bunkering pontoons are presented to solve the fueling difficulty of LNG ships on inland waterways with a great variation of water height and channel width by seasonal change. First, the background of this concept was analyzed as well as the demand of LNG bunkering pontoons, and the involved idea of levels management. Then, from the safety barrier perspective, safety design principles were discussed, the safety principles and implement measures of LNG storage were provided, LNG tanks for the pontoon were analyzed in detail in terms of their types, design pressure, material selection and leakage protection principles. Also, the safety concerns of LNG supply and bunkering process were raised, and an improved handy bunkering arm & hose, lighter and more flexible than ever before, was introduced. Berthing and unberthing are related to the survival of the pontoon so that the key points of such design were highlighted. It is concluded that (1) based on the levels management, the pontoon equipped with IMO Type C tanks, with reasonable berthing/unberthing and safety designs adopted, is proved to be safe and reliable by practices; (2) by 2025, nearly 30 such improved LNG bunkering pontoons with a fueling capacity of 500 m3 each will be required on Yangtze and Pearl Rivers; and (3) the pontoon's safety and reliability has been verified by practices and its fourth generation has been developed. Keywords: LNG bunkering pontoon, Floating fueling station, LNG fuel, Storage, IMO type C tanks, Bunkering arm & hose, Technical security, Levels management

Anti-inflammatory factors and regulatory T cells function as negative feedback circuit in immune responses. Interleukin-37 (IL-37) has been characterized as the anti-inflammatory factor recently and it is activated in inflammatory responses and autoimmune diseases. The activated IL-37 has high anti-inflammatory activities. In the IL-37 transgenic mice, there is a very low level of inflammatory factors when the mice are stimulated with antigens. IL-37 could bind to IL-18 receptor and activate downstream pathways, resulting in anti-inflammatory factors’ activation and immune suppression. In this review, we first discuss the regulatory T cell and cytokines involved in immune suppression and second discuss IL-37, including the history, mechanism of regulation and its role in the development of autoimmune diseases.

In-situ construction of energetics-on-a-chip by the microcharge technique has recently received much attention by researchers. In this study, a novel integrated CuN3/CL-20 explosive train film was fabricated in-situ on a Cu substrate. The porous CuN3 film, which was synthesized by facile electrochemical azidation of porous Cu, served as a skeleton for the integration of CL-20 by means of spin-coating and recrystallization. In this architecture, the introduction of CL-20 endowed the resultant explosive train film with a greater charge density and an enhanced energy output than a single CuN3 film. In particular, the tunability of exothermic capacity could be controlled by adjusting the loading amount of CL-20. Additionally, the embedded CL-20 and CuN3 film jointly ensured violent explosive behavior upon laser stimulation. Given these advantages, this CuN3/CL-20 explosive train film exhibits great potential for application in functional energetic chips.

The electric initiating explosive devices (EIEDs) are playing an increasingly important role in both military and civilian applications. With respect to good electrical conductivity and flexural strength, carbon paper can be endowed with primary explosive so that it serves as an independent module just like an ignition component in the EIEDs to satisfy the modular integration of EIEDs. But a traditional primary explosive cannot meet the need of sustainably developed devices. New energetic films need to be designed for EIEDs. The heat release and activation energy of the CA-CP film are 1402 J·g−1 and 104 kJ·mol−1, respectively. The electrostatic sensitivity of the CA film, whose E50 reaches up to 4.81 mJ, has been well improved due to its interweaved fibers. Moreover, the cuprous azide film could be successfully ignited under water after a hydrophobicity treatment. Herein, the CA-loaded carbon paper has been successfully integrated with the Ni-Cr bridge EIEDs for a greatly improved ignition performance, which will subsequently lead to a vast enhancement in the ability of EIEDs to ignite the secondary explosives or devices.

// Jianyong Xu 1, 2 , Wei Lian 1, 2 , Yuning Jia 1, 2 , Lingyun Li 1, 2 and Zhong Huang 1, 2 1 Institute of Biological Therapy, Shenzhen University, Shenzhen, P.R. China 2 Department of Pathogen Biology and Immunology, School of Medicine, Shenzhen University, Shenzhen, P.R. China Correspondence to: Jianyong Xu, email: xujianyong@szu.edu.cn Zhong Huang, email: zhuang809@126.com Keywords: RNA guided endonuclease, genome editing, Cas9, guide RNA, gRNA Received: August 09, 2017 Accepted: August 28, 2017 Published: October 07, 2017 ABSTRACT The genome editing tool Cas9-gRNA (guide RNA) has been successfully applied in different cell types and organisms with high efficiency. However, more efforts need to be made to enhance both efficiency and specificity. In the current study, we optimized the guide RNA structure of Streptococcus pyogenes CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas (CRISPR-associated) system to improve its genome editing efficiency. Comparing with the original functional structure of guide RNA, which is composed of crRNA and tracrRNA, the widely used chimeric gRNA has shorter crRNA and tracrRNA sequence. The deleted RNA sequence could form extra loop structure, which might enhance the stability of the guide RNA structure and subsequently the genome editing efficiency. Thus the genome editing efficiency of different forms of guide RNA was tested. And we found that the chimeric structure of gRNA with original full length of crRNA and tracrRNA showed higher genome editing efficiency than the conventional chimeric structure or other types of gRNA we tested. Therefore our data here uncovered the new type of gRNA structure with higher genome editing efficiency.

// Jianyong Xu 1,2,3 , Wei Lian 1,2,3 , Lingyun Li 1,2,3 and Zhong Huang 1,2,3 1 Institute of Biological Therapy, Shenzhen University, Shenzhen, China 2 Department of Pathogen Biology and Immunology, Shenzhen University School of Medicine, Shenzhen, China 3 Shenzhen City Shenzhen University Immunodiagnostic Technology Platform, Shenzhen, China Correspondence to: Jianyong Xu, email: // Zhong Huang, email: // Keywords : cardiac progenitor cell, CPC, somatic reprogramming, induced CPC Received : November 30, 2016 Accepted : January 24, 2017 Published : February 10, 2017 Abstract It has been demonstrated that cardiac progenitor cells (CPCs) represent a more effective cell-based therapy for treatment of myocardial infarction. Unfortunately, their therapeutic application is limited by low yield of cell harvesting, declining quality and quantity during the ageing process, and the need for highly invasive heart biopsy. Therefore, there is an emerging interest in generating CPC-like stem cells from somatic cells via somatic reprogramming. This novel approach would provide an unlimited source of stem cells with cardiac differentiation potential. Here we would firstly discuss the different types of CPC and their importance in stem cell therapy for treatment of myocardial infarction; secondly, the necessity of generating induced CPC from somatic cells via somatic reprogramming; and finally the current progress of somatic reprogramming in cardiac cells, especially induced CPC generation.

A high heterologous expression of an alkaline pectate lyase (APL) pelNK93I in E. coli was obtained through optimizing the lactose feeding and fed-batch fermentation. The highest soluble APL activity produced by E. coli BL21 (pET22b-pelNK93I) was 10,181 U/mL which is the highest level so far. On this basis, to improve the extracellular yield of APL, optimized glycine feeding was used to achieve elevated extracellular production of pelNK93I. The highest extracellular APL activity produced by E. coli BL21 (pET22b-pelNK93I) was 6357 U/mL which was also relatively higher than that in previous reports. The final productivity of APL was 282.8 U/mL/h in the fermentation of E. coli BL21 (pET22b-pelNK93I) in a 10 L fermenter. Thus the current study has provided a cost-effective method for the over-expression and preparation of alkaline pectate lyase pelNK93I for its industrial applications. Moreover, pelNK93I (4 U/mL) used for bioscouring increased cottonseed husk removal and radial capillary effect of cotton fabric by 37.63% and 47.06%, respectively, making it a promising enzyme in green textile technology.

Mesenchymal stem cells (MSCs) can be derived from various adult tissues with multipotent and self‐renewal abilities. The characteristics of presenting no major ethical concerns, having low immunogenicity and possessing immune modulation functions make MSCs promising candidates for stem cell therapies. MSCs could promote inflammation when the immune system is underactivated and restrain inflammation when the immune system is overactivated to avoid self‐overattack. These cells express many immune suppressors to switch them from a pro‐inflammatory phenotype to an anti‐inflammatory phenotype, resulting in immune effector cell suppression and immune suppressor cell activation. We would discuss the mechanisms governing the immune modulation function of these cells in this review, especially the immune‐suppressive effects of MSCs.

Background Although mesenchymal stem cells (MSCs) might offer a promising strategy for treating SLE, their immunoregulatory plasticity makes their therapeutic effects unpredictable. Whether overexpressing IL-37, an IL-1 family member with immunosuppressive activity, might enhance the therapeutic effects of these cells for SLE is unknown. Methods We genetically modified MSCs to overexpress IL-37 and assessed their effects on immune suppression in vitro. We also evaluated the effects of such cells versus effects of various controls after transplanting them into MRL/lpr mice (model of SLE). Results Stem cell characteristics did not appear altered in MSCs overexpressing IL-37. These cells had enhanced immunosuppression in vitro in terms of inhibiting splenocyte proliferation, reducing proinflammatory factors (IL-1β, TNF-α, IL-17, and IL-6), and suppressing autoantibodies (anti-dsDNA and anti-ANA). Compared with animals receiving control MSCs or IL-37 treatment alone, MRL/lpr mice transplanted with IL-37-overexpressing cells displayed improved survival and reduced signs of SLE (indicated by urine protein levels, spleen weight, and renal pathologic scores); they also had significantly lower expression of proinflammatory factors, lower total antibody levels in serum and urine, lower autoantibody production, and showed reduced T cell numbers in the serum and kidney. Expression of IL-37 by MSCs can maintain higher serum levels of IL-37, and MSCs had prolonged survival after transplantation, perhaps through IL-37 suppressing the inflammatory microenvironment. Conclusions Mutually reinforcing interaction between MSCs and IL-37 appears to underlie their additive therapeutic effects. Genetic modification to overexpress IL-37 might offer a way to enhance the stability and effectiveness of MSCs in treating SLE.

Mesenchymal stem cells (MSCs) have been intensively investigated and widely applied in regenerative medicine and immune modulation. However, their efficacy declines during the aging or disease process. Thus, genome-edited MSCs with over-expression or inhibition of specific genes hold a great deal of promise in terms of their therapeutic application. Here we optimized the direct PCR approach for rapid identification of genome-edited MSCs with only ten cells required, which reduces the time and labor to expand the MSC colonies. Combined with our previously optimized guide RNA structure and plasmid construction strategy for Cas9, we successfully identified MSC colonies over-expressing IL-10 in the AAVS1 locus.

In order to study the dynamic response and motion response characteristics of the river-sea direct ship’s structure under different conditions and waves during the navigation of the river section and the sea section, a container ship on the route from Wuhan to Yangshan was tested by using dynamic strain gauge, motion sensor and wave height gauge. The test points are arranged in the front and rear end walls of the cargo tank area, the middle section of the ship, the corner of the hatch, the intersection of the longitudinal bulkhead and the inner bottom plate and other areas that may produce high stress. The test results show that the wave environment conditions of the route area of the Yangtze River estuary are more complex, and the response components of the ship structure are more variable, which is quite different from the response of the structure in the Yangtze River Estuary; the predicted value of wave load is similar to the measured value, and the predicted bending moment is slightly larger than the measured bending moment.

Using the ultraviolet observation data of Huangshan Scenic area from March 2017 to February 2018, this paper analyzes the relationship between the daily, monthly and seasonal variation characteristics of ultraviolet radiation intensity and its related meteorological elements. The results show that the diurnal variation of ultraviolet radiation intensity in Huangshan Scenic Area is normal distribution at noon; the monthly average ultraviolet radiation intensity reaches the maximum in April and May; the ultraviolet radiation intensity in all seasons is above 4, and the largest in spring. Autumn and winter are second, and summer is the smallest. Cloud amount, relative humidity and air pressure are all significantly negatively correlated with UV radiation intensity, and cloud amount, especially low cloud amount, is the most critical factor affecting UV radiation intensity.

Carbohydrate binding module (CBM) as a kind of non-catalytic domain has significant effects on the substrate binding and catalytic properties of glycoside hydrolases. CBM68 of an Anoxybacillus sp. pullulanase (PulA) was identified as a new type of CBM in our previous studies. Then, four key substrate binding amino acid residues (Y14, V91, G92, and R96) were obtained by alanine substitutions in this work. Through kinetic analysis of the mutants, V91A and G92A showed significant reduction both in Km values and kcat values against pullulan. To further identify the changes of substrate affinities of V91A and G92A, devitalized mutants V91A-D413N and G92A-D413N were under measuring by surface plasmon resonance (SPR). Compared with that of PulA-D413N, the substrate affinities of V91A-D413N and G92A-D413N were improved by 1.6-fold and 2.2-fold, respectively. However, as to the product (maltotriose) binding force tested by the isothermal titration calorimetry (ITC), G92A showed higher binding force than that of V91A and PulA by 4.2-fold and 6.2-fold, respectively. That may cause G92A showing significantly lower catalytic efficiency than V91A and PulA. Moreover, four different kinds of amino acids (leucine, serine, glutamic acid and arginine) substitutions for V91 and G92 showed various changes both on the kinetic parameters and enzymatic properties, which demonstrated that V91 and G92 were the critical binding residues in the CBM68. The results of this study made contributed to the rational design for improving the catalytic efficiency of PulA.

Bacillus sp. strains as attractive hosts for the production of heterologous secretory proteins usually play important roles in bio-industry. However, low transformation efficiency of exogenous plasmids limited the application of Bacillus species. Here, a novel plasmid interspecific transfer system, with high transformation efficiency, high positive rate, and convenient manipulation, has been successfully constructed. A high electrotransformation efficiency strain Bacillus subtilis F-168 containing the counter-selectable marker mazF was used as the plasmid donor strain in this transfer method. A shuttled plasmid pBE980 and its recombinant plasmids pBE980::pulA and pBE980::HSPA were successfully transferred into the recipient Bacillus strains (Bacillus amyloliquefaciens 66, Bacillus licheniformis 124 and Bacillus megaterium 258) by this method. After co-culturing the donor cells (OD600nm = 1.3–1.7) and the recipient cells (OD600nm = 0.5–0.9) for 24 h in 22 °C, more than 1.0 × 105 positive transformants were obtained and a interspecific transformation efficiency of 1.0 × 10−3. It would provide a new approach for genetic manipulation in Bacillus strains and accelerate the research progress of the wild Bacillus strains in bio-industry.