Effect of Huanglian Wendantang on Classical Pathway of Skeletal Muscle Pyroptosis in IGT Rats / 中国实验方剂学杂志

Objective:To explore the effects of Huanglian Wendantang （HLWDT） on pyroptosis of skeletal muscle in rats with impaired glucose tolerance （IGT） and to explain the mechanism based on NOD-like receptor protein 3 （NLRP3）/cysteine aspartate-specific protease-1 （Caspase-1）/gasdermin D （GSDMD）/interleukin-1<italic>β </italic>（IL-1<italic>β</italic>）/IL-18 signaling pathway. Method:The SD male rats were fed with 45% high-fat diet for 20 weeks to induce the IGT model. After modeling， the rats were randomly divided into a blank group， a model group， a positive control group （metformin hydrochloride， 0.05 g·kg^-1d^-1）， and an HLWDT （7.8 g·kg^-1d^-1） group based on the body weight of rats. The blank group and the model group were fed with the same volume of distilled water. The dose for each group was set as 10 mL·kg^-1d^-1. After four weeks of continuous gavage， blood was collected and serum was separated. The skeletal muscles of rats were stored in liquid nitrogen. Subsequently， serum IL-1<italic>β</italic> and IL-18 were detected by the enzyme-linked immunosorbent assay （ELISA）. The mRNA and protein expression levels of NLRP3， Caspase-1， and GSDMD were detected by real-time quantitative PCR （Real-time PCR） and Western blot， respectively. The expression of GSDMD， IL-1<italic>β</italic>， and IL-18 proteins in skeletal muscle tissues was detected by immunofluorescence. Hematoxylin-eosin （HE） staining was used to observe the pathological changes of skeletal muscles. Result:Compared with the blank group， the model group showed increased IL-1<italic>β</italic> and IL-18 in serum， and NLRP3， Caspase-1， and GSDMD gene and protein expression in skeletal muscle tissues （<italic>P</italic><0.01）. Immunofluorescence assay showed that GSDMD， IL-18， and IL-1<italic>β </italic>protein expression in skeletal muscle tissues of the model group was significantly elevated （<italic>P</italic><0.01）. HE staining showed obvious pathological changes in skeletal muscles. Compared with the model group， the HLWDT group and the positive control group could decrease IL-1<italic>β</italic> and IL-18 in serum and NLRP3， Caspase-1， and GSDMD gene and protein expression in skeletal muscle tissues （<italic>P</italic><0.01）. In addition， immunofluorescence assay revealed that HLWDT could reduce protein expression levels of GSDMD， IL-1<italic>β</italic>， and IL-18 in skeletal muscles of IGT rats （<italic>P</italic><0.01）. The results of HE staining showed that HLWDT could improve the pathological changes of skeletal muscles in IGT rats<bold>.</bold> Conclusion:HLWDT can inhibit skeletal muscle pyroptosis of IGT rats， and the mechanism may be closely related to NLRP3/Caspase-1/GSDMD/IL-18/IL-1<italic>β</italic> signaling pathway.

Mechanism of Huanglian Wendan Decoction in improving impaired glucose tolerance based on skeletal muscle NLRP3/caspase-1/IL-1β, IL-18 pathway / 中国中药杂志

This study investigated the mechanism of improving impaired glucose tolerance(IGT) of rats by Huanglian Wendan Decoction from the perspective of the skeletal muscle Nod-like receptor protein 3(NLRP3)/cysteinyl aspartate specific proteinase-1(caspase-1)/interleukin-1β(IL-1β), interleukin-18(IL-18) pathway. Healthy male SD rats were fed with the diet containing 45% fat for 20 weeks, accompanied by a high-temperature and high-humidity environment and an inactive lifestyle, for the establishment of the IGT rat model. The rats were divided into the blank control group, model control group, metformin hydrochloride group(positive drug group, 0.05 g·kg~(-1)·d~(-1)) and Huanglian Wendan Decoction group(7.8 g·kg~(-1)·d~(-1)). After continuous intragastric administration for 4 weeks, the obesity and glycemic indexes of all the rats were measured. The fasting serum insulin(FINS) level was determined by ELISA, with the insulin sensitivity index(ISI) and insulin resistance index(IRI) calculated. The mRNA and protein expression le-vels of nuclear factor kappaB(NF-κB), NLRP3, caspase-1, IL-1β and IL-18 in skeletal muscle tissue were detected by real-time polymerase chain reaction(PCR), Western blot and immunofluorescence. Compared with the blank control group, the model control group witnessed significantly increased mRNA and protein expression of NF-κB, NLRP3, caspase-1, IL-1β and IL-18. As revealed by the comparison with the model control group, Huanglian Wendan Decoction could effectively regulate the obesity status, reduce body weight, correct the abnormal levels of 2-hour plasma glucose(2 hPG), insulin resistance index(IRI), insulin sensitivity index(ISI), and lower the mRNA and protein expression of NF-κB, NLRP3, caspase-1, IL-1β and IL-18 in the skeletal muscle tissue of IGT rats. Combined with previous studies, the above results showed that the occurrence and development of IGT was closely related to inflammatory response and the classic pyroptosis pathway in skeletal muscle, such as NLRP3/caspase-1/IL-1β, IL-18. It is inferred that the mechanism of Huanglian Wendan Decoction was to alleviate insulin resistance(IR) and then reverse the course of IGT lies in the regulation of the abnormal insulin receptor signaling pathway based on the NLRP3 inflammasome pathway.

Discharge and Contents of Exosome from Leukocyte-Reduced Apheresis Platelet Stored for Different Storage Time / 中国实验血液学杂志

OBJECTIVE@#To explore the effect of storage time on discharge and content of exosome from leukocyte-reduced apheresis platelets (LRA-Plt).@*METHODS@#Exosome (EXO) from LRA-Plt were acquired by ExoQuick, and its' morphology, immunological marker and particle size distribution were detected by transmission electron microscopy (TEM), Western blotting and dynamic light scattering (DLS), respectively. The changes in particle size distribution of EXO from LRA-Plt with different storage time were detected by DLS. The changes in content of protein and RNA of EXO from LRA-Plt with different storage time were detected by Nanodrop® ND-2000.@*RESULTS@#EXO from LRA-Plt was acquired successfully, which was characterized by cup-like shape, CD63/TSG101 enriched and Calnexin negative, and the particle size of which ranged from 30 to 200 nm. At early stored stage (stored for 1 day and 2 days), particle size of EXO from LRA-Plt was small and ranges from 30 to 40 nm. Meanwhile, the contents of protein and RNA were low. The particle size distribution, contents of protein and RNA of EXO from LRA-Plt were not significanty different ammg groups (P＞0.05). At middle-late stored stage (stored for 3, 4 and 5 days), the particle size of EXO from LRA-Plt was larger than that of early stored stage, which ranges was from 130 to 200 nm. Meanwhile, the contents of protein and RNA were higher than those of early stored stage. Particle size distribution, contents of protein and RNA of EXO from LRA-Plt stored for middle-late stage were significant higher than those of early stored stage (P＜0.05).@*CONCLUSION@#Morphology of EXO from LRA-Plt stored for middle-late stage was different from that stored for early stored stage. Moreover, the particle size distribution, contents of protein and RNA of EXO from LRA-Plt stored for middle-late stage were higher than those of early stored stage. A large amount of protein and RNA contained in EXO from LRA-Plt may participate in the multiple functions caused by platelet transfusion.

Outcome mapping for decision-making in health system / 浙江大学学报·医学版

Outcome mapping (OM) is a participant-oriented new method for project planning,monitoring and evaluation.It is based on continuous learning and focuses on changes in behavior,relationship and activities of persons,groups and organizations.The use of OM can help to clarify the roles and assignments of the government,managers,health workers,and other relevant personnel,making full use of the limited social resources and improving the efficiency and quality of health services.We introduce the principle and operation steps of OM with case simulation in health resource integration,to show its application in decision-making.

Effect of Weifuchun on inhibiting inflammation of Helicobacter pylori-infected GES-1 cells and NF-kappaB signaling pathway / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To study the effect of Weifuchun on inflammation of Helicobacter pylori (Hp)-infected gastric epithelial cells (GES-1) and its correlation with NF-kappaB signaling pathway.</p><p><b>METHODS</b>Hp standard home-made strain (CagA +, VacA +) NCTCI 1637 infected GES-1 cells were used. Weifuchun was used as intervention. Weifuchun of different concentrations (5,10, and 20 microg/mL) were screened by MTT assay. A blank group and the model group were set up. Then the growth inhibition rate of drugs on gastric epithelial GES-1 cells was detected with MTT assay. Cell cycle was detected using flow cytometry. The supernatant liquid was separated to detect the contents of IL-8 and IL-4 by ELISA.The protein expression level of NF-kappaB was detected by Western blot analysis.</p><p><b>RESULTS</b>MTT assay indicated significantly inhibitory effect of Weifuchun on GES-1 cells [5% inhibiting concentration (IC5)] was 10 microg/ml in the Weifuchun group. After GES-1 and Hp were cultured together,the contents of IL-8 in the supernatant were more obviously higher in the model group than in the blank group (P < 0.05), and then gradually decreased. After treatment with different concentrations of Weifuchun, the levels of IL-8 in the supernatant were less when compared with the model group at 12, 24, 48, and 72 h (P < 0.05). The decrement was the most significant in the high dose Weifuchun group. The IL-4 level in the supernatant was obviously lower in the model group than in the blank group. It obviously increased in the high concentration Weifuchun group (P < 0.05). There was no statistical difference in the IL-4 level between middle, low concentration Weifuchun group and the blank group (P > 0.05). The protein expression of intranuclear P65 increased and that of IkBalpha decreased 60 min after Hp infection. But the protein expression of intranuclear P65 decreased and the protein expression of IkBalpha increased after intervention of Weifuchun.</p><p><b>CONCLUSIONS</b>Weifuchun adjusted H. pylori induced IL-8 and IL-4 production by gastric epithelial cells through blocking NF-kappaB pathways. Its mechanisms might possibly lie in inhibiting p65 from entry into nucleus and the degradation of IkBalpha. Weifuchun was an effective drug for treatment of Hp correlated chronic gastritis.</p>