microRNA (miRNA) is a class of endogenous non-coding small RNAs,they play an important role in post-transcriptional regulation of gene expression through combining the target mRNA that the target protein would not synthesis.The present studies have found that miRNA is involved in many kinds of physiological processes,as well as the pathological processes.The abnormal expression of miRNA in many diseases can be used in diagnosis,prognosis and treatment monitoring.But all of these studies depend on an ideal detection method of miRNA.A lot of detection methods of miRNA expression developed from qualitative analysis to quantitative analysis step by step and from single miRNA detection to high throughput screening in recent years.Various detection methods are improved constantly,the specificity and sensitivity has been improved at the same time,detection procedure become simple and practicable,detection time shorten considerably and cost also reduced ceaselessly,which make the application of miRNA in clinical possible.This review highlights the latest research progress of the common detection methods of miRNA.

Objective To investigate the dynamic changes of splenocyte cytokines in mice immunized with the transgenic alfalfa containing Eg95-EgA31 fusion gene of Echinococcus granulosus (Eg). Methods Eighty-eight Balb/c mice were divided into 2 groups randomly according to body weights,and immunized orally or intranasally with 100μl or 10μl extracted leaf protein from the transgenic alfalfa(20 g/L) respectively once per 3 days for 2 months. Four mice randomized from each group were killed to get splenocyte on week 0(control),2,4,6,8,10,12,14,16,18 and 20 after the last immunization. The splenocyte were cultured in medium for 48 hours with EgAg or concanavalin A (ConA) stimulation to induce the interleukin (IL)-12,interferon γ(IFN-γ) and IL-10,and cultured for 72 hours with EgAg or lipopolysaccharide (LPS) stimulus to induce the tumor necrosis factor α (TNF-α). Then the supernatant was collected to measure the level of IL-12,IFN-γ,TNF-α and IL-10 by ELISA. Results In the oral immunization group,the level of IL-12,IFN-γ,TNF-α and IL-10 increased significantly from week 4 to week 6,week 2 to week 8,week 2 to week 6 and week 4 to week 12,respectively,reaching the highest level(25.0±5.8)ng/L on week 4,(575.0±28.9)ng/L on week 2,(50.0±11.5)ng/L on week 2 and (42.5±2.9)ng/L on week 8,respectively,as compared with the values on week 0[(11.3±2.5),(125.0±28.9),(11.3±2.5),(12.5±2.9)ng/L,all P < 0.01]; in the intranasal immunization group,it was similar about the values of IL-12,IFN-γ,TNF-α and IL-10 could be seen from week 4 to week 6,week 2 to week 10,week 4 to week 10 and week 6 to week 16,respectively,reaching the highest level(25.0±5.8)ng/L on week 6,(725.0±28.9)ng/L on week 4,(27.5±2.9)ng/L on week 6 and (60.0±11.5)ng/L on week 6,respectively,as compared with the values on week 0[(11.3±2.5),(125.0±28.9),(11.3±2.5),(12.5±2.9)ng/L,all P < 0.01]. The cytokine levels in the groups with EgAg,ConA or LPS stimulus were significantly higher than those in the corresponding splenocyte suspension groups(P < 0.05 or < 0.01),and the cytokine levels in the groups with ConA or LPS stimulus were obviously higher than those in the corresponding groups with EgAg stimulation (P < 0.05 or < 0.01). Conclusion The mixed responses of Th1 and Th2 types can be induced in mice immunized with the transgenic alfalfa in the early period post immunization(2-10 weeks).

Objective To investigate the change of splenocyte subsets in Balb/c mice immunized with transgenic alfalfa(Medicago sativa)containing Eg95-EgA31 fusion gene of Echinococcus granulosus(Eg) and challenged with Eg protoscoleces.Methods Leaf protein was extracted from transgenic alfalfa containing Eg95-EgA31 fusion gene by heat-coagulation method,and concentration of 20 g/L was used in the study.Meanwhile,leaf protein extracted from the transgenic alfalfa containing blank vector(pBI121)and the normal alfalfa was served as control.Thirty-two female Balb/c mice were randomly divided into 4 groups,8 mice in each group.Oral group was immunized with the leaf protein containing Eg95-EgA31 fusion antigen intragastrically(100μl per mouse);intranasal group was immunized with the leaf protein containing Eg95-EgA31 fusion antigen intranasally(10 μl per mouse);blank vector group was vaccinated intranasally with 10μl leaf protein with blank vector(pBI121);and normal control group was given 100μl normal leaf protein intragastrically.All mice in the above mentioned groups were immunized every 3 days for 2 months.Then,the mice were challenged intraperitoneally with Eg protoscoleces(50 protoscoleces per mouse)8 weeks after last vaccination and sacrified 24 weeks pest infection to separate the splenocytes.Flow cytometry was used to measure the percentages of CD4+ and CD8+ T ceils subsets.Resuits Compared with the normal control group(0.166±0.018,0.083±0.006,2.019 ±0.369),the percentages of CD4+(0.286±0.009)and CD8+(0.102±0.004)T cell subsets and the ratio of CD4+/CD8+(2.814±0.014)in oral group increased significantly (P<0.01 or<0.05).The percentage of CD4+ subset(0.269±0.016)and the ratio of CD4+/CD8+(2.955±0.986) in intranasal group was significantly higher than that ofthe normal control group(all P<0.01).The percentage of CD4+ subset in oral group was significantly higher than that of the intranasal group(P<0.05).No significant difference was found in the percentages of CD4+ and CD8+ T cell subsets and the ratio of CD4+/CD8+ between the blank vector group(0.169±0.018,0.093±0.019,1.852±0.188)and the normal control group(all P>0.05).Conclusions CD4+ T cell may play an important role in the protection induced by transgenic alfalfa vaccine against the challenge of Eg protoscoleces.Intragastrical immunization may be a good route.

Objective To cultivate and identify the transgenic affalfa containing Echinococcus granulosus Eg95 gene. Methods The alfalfa plants were transformed by co-cultivating alfalfa cotyledons via recombinant Agrobacterium tumefaciens LBA4404 harboring pBI-Eg95. The transgenic alfalfa explants were selected by kanamyein after calli formation, shoots and roots regeneration in the selective medium, the seedlings of transgenic plants were obtained which were finally transplanted into pots containing nutrient soil. After 2-3 months growth, the complete transgenic alfalfa plants containing Echinococcus granulosus Eg95 gene were obtained. To identify the transgenic alfalfa plants, the total DNA, RNA and leaf protein were extracted from fresh leaf tissue of the transgenic alfalfa plants and confirmed by PCR, RT-PCR, SDS-PAGE and Western blot assay. Results A specific band around 471 bp was amplified by PCR with total DNA, and the same band was obtained by RT-PCR with total RNA, which confirmed that the Eg95 gene was stably integrated into the transformed alfalfa genome. SDS-PAGE analysis showed that the relative molecular mass(Mr) of the expressed protein was about 16.5×103, consistent with the Eg95 protein, and the level of Eg95 expression was up to 0.06% of total soluble leaf protein by Bio-Rad Quantity one assay. Western blot verified the expressed protein was reactive with the sera of mice infected with Echinococcus granulosus. Conclusion The transgenic alfalfa plants containing Echinococcus granulosus Eg95 gene are successfully cultivated.

0.05).Our results showed that,for recommended oral doses of ciprofloxacin and moxifloxacin,44.2% of E.coli isolates from the intestine of the health population,and 29.5%,18.7% and 10.7% of isolates from the three sterile sites of the patients,would be selected as resistant mutants.When ciprofloxacin and moxifloxacin were taken by injection route,the ratio of the selection of resistant mutants would be 9.3% for E.coli isolates from the intestine of the health population and 8.2%,6.2% and 8.9% for the three sterile sites of the patients,respectively.The maximum attainable concentration of levofloxacin in serum showed little distinction between the oral and the other routes.CONCLUSIONS The values of MIC and MPC of three fluoroquinolones in E.coli isolates from different populations and sites show no association.The values of MPC couldn't be predicted by the MIC.The values of MPC and MPC90 of three drugs show no significant discrepancy for tested isolates,these E.coli strains are isolated from the intestine of heath persons,and from the blood,ascites,bile of patients.

OBJECTIVE:To investigate the compatibility stability of Salvianolic acid for injection (SAFI) combined with 8 kinds of common solvents. METHODS:Referring to package inserts,SAFI was collected and combined with 0.9% Sodium chlo-ride injection,5% Glucose injection,10% Glucose injection,Compound sodium chloride injection,Mannitol injection,Glycerol fructose and sodium chloride injection,Sodium lactate Ringer's injection,Glucose and sodium chloride injection,250 mL each re-spectively,and then sealed in the dark at the room temperature. The appearance of mixtures were observed,pH value,the number of insoluble particles,maximal absorption wavelength and maximal absorbance were detected,and the contents of salvianolic acid B,rosmarinic acid,lithospermic acid and salvianolic acid Y in mixtures were determined by HPLC at 0,1,2,4,8 h after mix-ing. RESULTS:Under above condition,no obvious change was found in appearance or pH values of the mixtures within 8 h. Maxi-mal absorption wavelength ranged 284.5-286.0 nm. After mixed with Mannitol injection,the number of particles≥10μm(1-8 h af-ter mixing)and particles ≥25 μm(4-8 h after mixing)exceeded the scope of Chinese Pharmacopoeia(2015 edition);the maxi-mal absorbance changed significantly(RSD=9.17%,n=5);the relative content of salvianolic acid B,rosmarinic acid,lithosper-mic acid and salvianolic acid Y decreased by more than 10%(RSD=14.65%,6.45%,8.97%,12.49%,n=5);after mixed with Sodium lactate Ringer's injection,the relative content of rosmarinic acid and lithospermic acid changed greatly (RSD=14.57%, 7.28%,n=5);after mixed with 5% Glucose injection(4-8 h after mixing)and Glycerol fructose and sodium chloride injection(8 h after mixing),the relative content of rosmarinic acid were less than 90%(RSD=6.30%,4.86%,n=5);and the number of particles ≥25μm exceeded the scope of phamcopoeia after mixing with Glycerol fructose and sodium injection(0 h). The number of insoluble particles in other mixtures were in line with the standard of pharmacopoeia;maximal absorbance had no significant change(RSD<5%,n=5),and the relative content change of analytes were all less than 10%. CONCLUSIONS:Clinical appli-cation of SAFI combined with Mannitol injection,Sodium lactate Ringe's injection and Glycerol fructose and sodium injecrion should be avoided. After mixed with 5% Glucose injection,SAFI should be used within 4 h. SAFI can be compatible with 0.9%Sodium chloride injection,10% Glucose injection,Compound sodium chloride injection and Glucose and sodium chloride injec-tion.

Febuxostat, as a xanthine oxidase inhibitor, is a classic anti-gout drug with significant therapeutic effects and good tolerability. The structures of febuxostat and its derivatives can be divided into two parts: a substituted phenyl ring and a five-membered or six-membered heterocyclic ring with a carboxyl substitution. This paper reviewed the research progress of febuxostat derivatives in recent ten years and classified the structure-activity relationships of various febuxostat derivatives. Exploring the action mechanisms and structure-activity relationships of xanthine oxidase inhibitors might be significant for the rational design and development of new anti-gout chemical entities.

BACKGROUNDCompensatory sweating (CS) is one of the most common postoperative complications after thoracic sympathectomy, sympathicotomy or endoscopic sympathetic block (ESB) for palmar hyperhidrosis. This study was conducted to examine the relevance between CS and the sympathetic segment being transected in the surgical treatment of palmar hyperhidrosis, and thus to detect the potential mechanism of the occurrence of CS.

METHODSBetween October 2004 and June 2006, 163 patients with primary hyperhidrosis were randomly divided into two groups, T(3) sympathicotomy (78 patients) and T(4) sympathicotomy (85), who were operated upon under general anesthesia via single lumen intubation and intercostal video-mediastinoscopy (VM).

RESULTSNo morbidity or mortality occurred. Palmar hyperhidrosis was cured in all patients. Follow-up (mean (13.8 +/- 6.2) months) showed no recurrence of palmar hyperhidrosis. The difference of rates of mild CS in groups T(3) and T(4) was of no statistical significance. The rate of moderate CS was significantly lower in group T(4) than in group T(3). No severe CS occurred.

CONCLUSIONThe rates of occurrence and severity of CS are lowered with the lower sympathetic chain being transected.

Adolescent ; Adult ; Female ; Follow-Up Studies ; Humans ; Hyperhidrosis ; surgery ; Male ; Middle Aged ; Postoperative Complications ; etiology ; Prospective Studies ; Sweating ; Sympathectomy ; adverse effects ; methods ; Thoracic Surgery, Video-Assisted

[Objective] To observe the effect of fluoride foam on dental caries increment in primary teeth, and provide the basis for prevention and control of children's dental caries. [ Methods] Three hundred and thirty-five children were included randomly from junior classes of 4 kindergartens as experi-mental group , and another 268 children from junior classes of another 4 kindergartens as control group .The experimental group used 1.23%fluoride foam twice a year , and the control group did not have any inter-ventions.The two groups of children were followed up for 3 years, and examined for the situation of their caries detection rate and average risk of caries , and for evaluation of the effect of fluoride foam in prevention of children's dental caries . [ Results] Caries detection rate in the two groups increased with age .After 3 years, caries detection rate in control group increased by 34.05%and in experimental group increased by 22. 19%.The use of fluoride foam reduced 10.89%caries disease for experimental group .Caries detection rates were 55.62%in experimental group and 66.51%in control group .The difference in detection rate was sig-nificant between experimental group and control group (P<0.05).Average caries in experimental group was 3.4 ±4.38 after 3 years, and 4.7 ±4.80 in control group.There was a significant difference in average caries between the two groups (P<0.05). [Conclusion] Application of 1.23%fluoride foam was effective in reducing the increment of dental caries in primary teeth , and it is worth promoting in the kindergartens .

Non-alcoholic fatty liver disease (NAFLD) has become a major public health hazard threatening human health worldwide.Yet, due to its complex pathogenesis, new drug development is difficult, with still insufficient clinical medication.Palmitoylation is a universal posttranslational modification of proteins catalyzed by palmitoyltransferase, affecting their stability, membrane localization and function.Recent studies have shown that palmitoylation is closely associated with NAFLD.This review summarizes the mechanisms of palmitoylation in NAFLD and analyzes the expression levels of the palmitoyltransferase family in liver tissues of NAFLD patients from GEO database, aiming to provide important clues to explore new mechanisms for NAFLD.