MSC-derived exosomes attenuate hepatic fibrosis in primary sclerosing cholangitis through inhibition of Th17 differentiation

Primary sclerosing cholangitis(PSC)is an autoimmune cholangiopathy characterized by chronic inflammation of the biliary epithelium and periductal fibrosis,with no curative treatment available,and liver transplantation is inevitable for end-stage patients.Human placentalmesenchymal stem cell(hpMSC)-derived exosomes have demonstrated the ability to prevent fibrosis,inhibit collagen production and possess immunomodulatory properties in autoimmune liver disease.Here,we prepared hpMSC-derived exosomes(Exo^(MSC))and further investigated the anti-fibrotic effects and detailed mechanism on PSC based on Mdr2^(−/−)mice and multicellular organoids established from PSC patients.The results showed that Exo^(MSC) ameliorated liver fibrosis in Mdr2^(−/−)mice with significant collagen reduction in the preductal area where Th17 differentiation was inhibited as demonstrated by RNAseq analysis,and the percentage of CD4+IL-17A+T cells was reduced both in Exo^(MSC)-treated Mdr2^(−/−)mice(Mdr2^(−/−)-Exo)in vivo and Exo^(MSC)-treated Th17 differentiation progressed in vitro.Furthermore,Exo^(MSC) improved the hypersecretory phenotype and intercellular interactions in the hepatic Th17 microenvironment by regulating PERK/CHOP signaling as supported by multicellular organoids.Thus,our data demonstrate the antifibrosis effect of Exo^(MSC) in PSC disease by inhibiting Th17 differentiation,and ameliorating the Th17-induced microenvironment,indicating the promising potential therapeutic role of Exo^(MSC) in liver fibrosis of PSC or Th17-related diseases.

左乳腺癌保乳术后全乳和锁骨上下及内乳淋巴结区三种放疗方式的剂量学研究

目的:对左乳腺癌保乳术后全乳及锁骨上下、内乳淋巴结引流区三种放疗方式在靶区及危及器官(organs at risk,OARs)的剂量学进行研究。方法:选取2018年1月至2019年10月20例于昆明医科大学第一附属医院行左乳腺癌保乳术后放疗患者的定位CT资料,设计固定野滑窗调强放疗(dynamic multi-leaf collimator,dMLC)、容积弧形调强放疗(volumetric intensity modulated arctherapy,VMAT)和螺旋断层放疗(tomotherapy,HT)三种计划,并对剂量指标进行统计学分析。结果:HT在术后瘤床(plan target volume of tumor bed,PTVtb)的平均剂量(mean dose,Dmean)、D1、适形度(homogeneity index,HI)和全乳及淋巴引流区的计划靶区(plan target volume,PTV)的Dmean、均匀性(conformity index,CI)均优于dMLC和VMAT,并且降低心脏和患侧肺的Dmean、V5、V10、V30。HT在冠状动脉左前降支和右冠状动脉的Dmean以及D1比较中剂量最低,但相比VMAT及dMLC,HT增加健侧乳腺的D1和Dmean。结论:对左乳腺癌保乳术后患者,HT对降低心脏毒性和保护患侧肺最有优势,但对健侧肺低剂量控制不如VMAT和dMLC,dMLC对健侧肺及健侧乳腺低剂量控制最佳,但对心脏及患侧肺的高剂量控制最差。在临床中需根据实际情况选择合适的计划方式。

Enhanced production of glycyrrhetic acid 3-O-mono-β-D-glucuronide by fed-batch fermentation using p H and dissolved oxygen as feedback parameters

Glycyrrhetic acid 3-O-mono-β-D-glucuronide(GAMG), the major functional ingredient in licorice, has widespread applications in food, pharmacy and cosmetics industry. The production of GAMG through Penicillium purpurogenum Li-3 cultivation was for the first time performed through both batch and fed-batch processes in bioreactors. In batch process, under optimal conditions(p H 5.0, temperature 32 °C, agitation speed 100 r·min-1), 3.55 g·L-1GAMG was obtained in a 2.5 L fermentor. To further enhance GAMG production, a fine fed-batch process was developed by using p H and DO as feedback parameters. Starting from 48 h, 100 ml 90 g·L-1substrate Glycyrrhizin(GL) was fed each time when p H increased to above 5.0 and DO was increased to above 80%. This strategy can significantly enhance GAMG production: the achieved GL conversion was 95.34% with GAMG yield of 95.15%, and GAMG concentration was 16.62 g·L-1which was 5 times higher than that of batch. Then, a two-step separation strategy was established to separate GAMG from fermentation broth by crude extraction of 15 ml column packed with D101 resin followed by fine purification with preparative C18 chromatography. The obtained GAMG purity was 95.79%. This study provides a new insight into the industrial bioprocess of high-level GAMG production.

Predication of 3-D Viscous Flowfield of a Centrifugal Impeller

A three-dimensional viscous code has been developed to solve Reynolds-averaged Navier-Stokes equations. The governing equations in finite volume form are solved by two-step Runge-Kutta scheme with implicit residual smoothing. The eddy viscous is obtained using the Baldwin-Lomax model. A prediction of the 3-D turbulent flow and the performance in the “all-over controlled vortex distribution” centrifugal impeller with a vaneless diffuser has been made for the compressor at design and off-design condition. The predicted effi-ciency is a little higher than the experiment data. These results suggest that the present calculation code is able to determine the flow development in the impeller and also the turbulence model in the centrifugal im-peller should be improved.

Extramedullary skeletal muscle metastasis of glioblastoma:A case report and literature review

Objective The aim of the study was to explore the clinicopathologic, immunophenotypic, and diagnostic features of extramedullary metastases of glioblastoma. Methods One case of extramedullary skeletal muscle metastasis of glioblastoma was studied, including the clinical, histological, and immunohistochemical features. Results A 24-year-old man underwent surgical resection for glioblastoma(WHO grade IV) in the left temporal parietal region followed by radiotherapy and temozolomide therapy. One year and nine months later, he developed an extramedullary skeletal muscle metastasis in L4, and the histology was remarkably different from that of the primary glioblastoma specimen. The immunohistochemical analysis also showed changes. In the metastasis, the small cells were negative for GFAP; weakly positive for S-100; and positive for nestin, NSE, and CD56, with 60% of cells positive for p53 and 40% positive for Ki-67. The giant cells showed strong positivity for GFAP and S-100, and weak expression of p53, Ki-67, nestin, NSE, and CD56. The primary glioblastoma specimen showed strong positivity for GFAP and S-100 and was negative for NSE, nestin, and CD56, with around 25% of the tumor cells positive for p53 and a Ki-67 labeling index of 20%. Conclusion Extraneural metastasis(ENM) is a rare complication of glial tumors and glioma stem cells may be related to the metastasis. Since extraneural metastasis may occur in patients without central nervous symptoms, any unusual signs during the follow-up of patients diagnosed with glioblastoma should not be underestimated.

糖苷酶耐热性改造策略与应用

糖苷酶作为绿色温和的生物催化剂,能够水解包含糖苷、寡糖、多糖等在内的各种含糖化合物的糖苷键生成具有高生理和药理活性的衍生物,在食品、医药等工业领域应用广泛。而工业应用的糖苷酶经常需在高温条件下进行催化,以提高反应效率并减少污染,但大多数糖苷酶属于中温酶,在实际生产条件下的活性较低且损失较快,因此,提高糖苷酶在高温下的稳定性非常重要。本文综述了近年来利用定向进化、理性设计和半理性设计等策略改造糖苷酶耐热性的研究进展与应用,比较了不同策略之间的优势与不足,并对未来糖苷酶耐热性的改造方向进行了展望。

Enhanced production of β-glucuronidase from Penicillium purpurogenum Li-3 by optimizing fermentation and downstream processes

β-Glucuronidase from Penicillium purpuro- genum Li-3 （PGUS） can efficiently hydrolyze glycyrrhizin into the more valuable glycyrrhetic acid monoglucuronide. However, a low productivity of PGUS and the lack of an effective separation strategy have significantly limited its industrial applications. Therefore, the production of PGUS has been improved by optimizing both the fermentation and purification strategies. A two-stage fermentation strategy was developed where PGUS was first grown with glucose and then PGUS was produced in the presence of glycyrrhizin as an inducer. By using this strategy, the biomass was increased 1.5 times and the PGUS activity increased 5.4 times compared to that when glycyrrhizin was used as the sole carbon source. The amount of PGUS produced was increased another 16.6% when the fermentation was expanded to a 15-L fermenter. An effective protocol was also established to purify the PGUS using a sequential combination of hydrophobic, strong anionexchange and gel filtration chromatography. This protocol had a recovery yield of 6% and gave PGUS that was 39 times purer than the crude PGUS. The purified PGUS had a specific activity of 350 U. mg-1.

Rational synthetic combination genetic devices boosting high temperature ethanol fermentation

The growth and production of yeast in the industrial fermentation are seriously restrained by heat stress and exacerbated by heat induced oxidative stress.In this study,a novel synthetic biology approach was developed to globally boost the viability and production ability of S.cerevisiae at high temperature through rationally designing and combing heat shock protein(HSP)and superoxide dismutase(SOD)genetic devices to ultimately synergistically alleviate both heat stress and oxidative stress.HSP and SOD from extremophiles were constructed to be different genetic devices and they were preliminary screened by heat resistant experiments and anti-oxidative experiments,respectively.Then in order to customize and further improve thermotolerance of S.cerevisiae,the HSP genetic device and SOD genetic device were rationally combined.The results show the simply assemble of the same function genetic devices to solve heat stress or oxidative stress could not enhance the thermotolerance considerably.Only S.cerevisiae with the combination genetic device(FBA1p-sod-MB4-FBA1p-shsp-HB8)solving both stress showed 250%better thermotolerance than the control and displayed further 55%enhanced cell density compared with the strains with single FBA1p-sod-MB4 or FBA1p-shsp-HB8 at 42
C.Then the most excellent combination genetic device was introduced into lab S.cerevisiae and industrial S.cerevisiae for ethanol fermentation.The ethanol yields of the two strains were increased by 20.6%and 26.3%compared with the control under high temperature,respectively.These results indicate synergistically defensing both heat stress and oxidative stress is absolutely necessary to enhance the thermotolerance and production of S.cerevisiae.

Construction of a CaHP04-PGUSl hybrid nanoflower through protein-inorganic self-assembly, and its application in glycyrrhetinic acid 3-0-mono-β-D-glucuronide preparation

Glycyrrhetinic acid 3-0-mono-β-D-glucuronide (GAMG), an important pharmaceutical intermediate and functional sweetener, has broad applications in the food and medical industries. A green and cost-effective method for its preparation is highly desired. Using sitedirected mutagenesis, we previously obtained a variant of β-glucuronidase from Aspergillus oryzae Li-3 (PGUS1), which can specifically transform glycyrrhizin (GL) into GAMG. In this study, a facile method was established to prepare a CaHP04-PGUSl hybrid nanoflower for enzyme immobilization, based on protein-inorganic hybrid selfassembly. Under optimal conditions, 1.2 mg of a CaHP04- PGUS1 hybrid nanoflower precipitate with 71.2% immobilization efficiency, 35.60 mg·g^-1 loading capacity, and 118% relative activity was obtained. Confocal laser scanning microscope and scanning electron microscope results showed that the enzyme was encapsulated in the CaHP04-PGUSl hybrid nanoflower. Moreover, the thermostability of the CaHP04-PGUSl hybrid nanoflower at 55°C was improved, and its half-life increased by 1.3 folds. Additionally, the CaHP04-PGUSl hybrid nanoflower was used for the preparation of GAMG through GL hydrolysis, with the conversion rate of 92% in 8 h, and after eight consecutive runs, it had 60% of its original activity.