Optimization of Extraction and Purification Process of Total Iridoids and Total Crocins in Gardeniae Fructus / 中国实验方剂学杂志

ObjectiveTo optimize the extraction and purification process of Gardeniae Fructus for industrial production， and to obtain the total iridoid and total crocin extracts. MethodOrthogonal test was used to optimize the water extraction process by taking contents of geniposide， genipin gentiobioside， gardenoside， crocin-1 and crocin-2 as indicators and the decocting time， decocting times and water amount as factors. The purification process was optimized by single factor test， and four different types of macroporous adsorption resins were screened. The process conditions such as resin type， maximum loading amount， water washing amount， ethanol concentration， ethanol dosage， and flow rate of sample loading were mainly investigated. In addition， the drying methods （vacuum drying and spray drying） of the extract were investigated， and a pilot scale-up verification test was carried out. ResultThe optimal water extraction process of Gardeniae Fructus was to add 15， 10 times the amount of water for decocting twice， 1 h each time. The optimal purification process was as follows：the water extract through SP825L macroporous resin column， the amount of crude drug-the amount of resin （1∶1.5）， the sample loading flow rate of 3 BV h-1， adding 2 BV of water to remove impurities， adding 4 BV of 30% ethanol to obtain the iridoid part， then adding 3 BV of 70% ethanol to obtain the crocin part， collecting the ethanol lotion， and drying at 70 ℃. Under these conditions， the extraction amount of total iridoids was 590.75 mg·g-1 with the transfer rate of 70.48%， and the yield of dry extract was 8.89%. The extraction amount of total crocins was 83.37 mg·g-1 with the transfer rate of 22.20%， and the dry extract yield was 2.60%. ConclusionThe optimized extraction and purification process is stable and feasible with high extraction rate of active components， which is suitable for the industrial extraction and purification of active parts of Gardeniae Fructus.

Research Status of Zexietang / 中国实验方剂学杂志

Zexietang is derived from Jingui Yaolue （《金匮要略》）， which is composed of Alismatis Rhizoma and Atractylodis Macrocephalae Rhizoma， and has the effect of inducing diuresis and invigorating the spleen to produce water. Compared with western medicine in the treatment of related diseases， Zexietang can not only improve the curative effect， but also reduce the occurrence of adverse reactions， so as to achieve long-term stable administration. The authors sorted out and analyzed the chemical composition， pharmacological effect and clinical application of Zexietang in recent years. It was found that the main active components of Zexietang were alismol A and B， 23-acetyl-alismol B and C， atractylenolides （atractylenolide Ⅰ， Ⅱ， Ⅲ） and polysaccharides. Pharmacological experiments showed that they had diuretic， hypolipidemic， anti-inflammatory and others. And it can be used in the treatment of hypertension， hyperlipidemia， vertigo， cerebral vascular insufficiency and other diseases combined with other Chinese materia medica， and the curative effect is obvious. By summarizing the research status of Zexietang in recent years， its active components and pharmacological mechanism can be further clarified， which provides the basis for the clinical application of Zexietang and guides the direction of its further research.

Research Progress of Targeted Drug Delivery Nanocarriers Through Blood-brain Barrier / 中国实验方剂学杂志

The small size， moderate drug loading， and targeting properties of nano-preparations make them can be excellent delivery tools for drugs， genes or proteins crossing the cell or blood-brain barrier （BBB）. Currently， facilitating drug crossing BBB with innovative nano-drug delivery systems is considered as a strategic approach for the prevention， diagnosis and treatment of central nervous system （CNS） diseases. However， with the deepening of the research， the adverse reactions and toxicity of nanocarriers have gradually attracted the attention of researchers. Based on this， this paper summarized the situation of BBB-penetrating targeted nano-preparations at home and abroad in recent years from the perspective of classification of types and properties of nanocarriers， and analyzed the advantages and disadvantages of each carrier. The results showed that nano-preparations with active ingredients of traditional Chinese medicine （TCM） as carriers have become a promising way of cancer treatment， but the complexity and diversity of TCM components limited its application to a certain extent. Further studies should be strengthened to lay a foundation for the application and development of TCM nano-preparations in the field of CNS diseases.

Preparation of paclitaxel-loaded and folic acid-modified poly (lactic-co-glycolic acid) nano-micelles and in vitro anticancer effect on cervical cancer HeLa cells / 中国中药杂志

The paclitaxel-loaded and folic acid-modified poly(lactic-co-glycolic acid) nano-micelles(PTX@FA-PLGA-NMs) were prepared by the emulsion solvent evaporation method, and the parameters of paclitaxel-loaded nano-micelles were optimized with the particle size and PDI as evaluation indexes. The morphology of the nano-micelles was observed by transmission electron microscopy(TEM), and the stability, drug loading and encapsulation efficiency were systematically investigated. In vitro experiments were performed to study the cytotoxic effects of nano-micelles, apoptosis, and cellular uptake. Under the optimal parameters, the nano-micelles showed the particle size of(125.3±1.2) nm, the PDI of 0.086±0.026, the zeta potential of(-20.0±3.8) mV, the drug loading of 7.2%±0.75%, and the encapsulation efficiency of 50.7%±1.0%. The nano-micelles were in regular spherical shape as observed by TEM. The blank FA-PLGA-NMs exhibited almost no inhibitory effect on the proliferation and growth of tumor cells, while the drug-loaded nano-micelles and free PTX exhibited significant inhibitory effects. The IC_(50) of PTX@FA-PLGA-NMs and PTX was 0.56 μg·mL~(-1) and 0.66 μg·mL~(-1), respectively. The paclitaxel-loaded nano-micelles were potent in inhibiting cell migration as assessed by the scratch assay. PTX@FA-PLGA-NMs had good pro-apoptotic effect on cervical cancer HeLa cells and significantly promoted the uptake of HeLa cells. The results of in vitro experiments suggested that PTX@FA-PLGA-NMs could target and treat cervical cancer HeLa cells. Therefore, as nanodrug carriers, PTX@FA-PLGA-NMs with anti-cancer activity are a promising nano-system for improving the-rapeutic effects on tumors.

Cancer-associated fibroblasts mediate differentiation of myeloid-derived suppressor cells in pancreatic cancer through IL-6/STAT3 pathway / 第二军医大学学报

Objective To explore the mechanism and biological significance of differentiation of myeloid-derived suppressor cells (MDSCs) mediated by cancer-associated fibroblasts (CAFs) in the pancreatic ductal adenocarcinoma (PDAC), so as to provide theoretical and experimental basis for revealing the roles of CAFs and MDSCs in promoting the progression of pancreatic cancer by remodeling the pancreatic cancer microenvironment. Methods We isolated and purified primary CAFs from PDAC tumor tissues, and screened the up-regulated cytokines in CAFs by quantitative real-time PCR and enzyme-like immunosorbent assay. The human foreskin fibroblasts (HFFs) were used as controls. Human peripheral blood mononuclear cells (PBMCs) were cultured with supernatant of CAFs and HFFs, respectively. The differentiation of PBMCs was observed and the mechanisms of the above cytokines in regulating the differentiation and recruitment of MDSCs were studied. Results The biomarkers (α-smooth muscle actin [α-SMA] and fibroblast activation protein a [FAPa]) were detected in the isolated primary CAFs, but not found in the HFFs. The expression levels of interleukin 6 (IL-6), stromal cell-derived factor 1 (SDF-1) and monocyte chemotactic protein 1 (MCP-1) in the culture supernatant were significantly gradually increased in the CAFs than those in the HFFs (all P0.01). Compared with culture supernatant of HFFs, the culture supernatant of CAFs promoted more PBMCs to differentiate into CD13-high expression neutrophil-like MDSCs (CD13hi-nMDSCs; P0.01). IL-6 human recombinant protein alone in the co-culture system could induce the differentiation of PBMCs into CD13hi-nMDSCs (P0.01). SDF-1 or MCP-1 human recombinant protein alone could not induce the increase of CD13hi-nMDSCs subpopulation. IL-6 neutralizing antibody or signal transducer and activator of transcription 3 (STAT3) blocker FLLL32 could significantly inhibit the differentiation induced by the culture supernatant of CAFs (P0.05). Conclusion CAFs can promote the differentiation of PBMCs into CD13hi-nMDSCs via the IL-6/STAT3 pathway.

Effect of magnesium-calcium alloy extract on matrix metalloproteinase-9 and matrix metalloproteinase inhibitor-3 in human colonic epithelial cells / 新乡医学院学报

Objective To investigate the effect of different concentrations of magnesium-calcium alloy extract on the expression of matrix metalloproteinase-9 (MMP9) and tissue inhibitor of metalloproteinase-3 (TIMP3) in human colonic epithelial NCM460 cells.Methods The different concentrations of extracts (the volume fraction was 10％,50％ and 100％ respectively) were made with magnesium-calcium alloy.The 5 × 106 L-1 NCM460 suspension was randomly divided into control group,experimental group 1,experimental group 2 and experimental group 3.The cells in the control group were cultured by 2 000 μL high glucose Dulbecco's modified Eagle's medium (containing 10％ volume fraction of fetal bovine serum).The cells in the experimental group 1,2 and 3 were cultured by 2 000 μL magnesium-calcium alloy extract with volume fraction of 10％,50％ and 100％ respectively.The expressions of MMP9 and TIMP3 mRNA in NCM460 cells was detected by real-time fluorescence quantitative polymerase chain reaction,and the expression of MMP9 and TIMP3 protein in NCM460 cells was detected by Western blot at after one,three and five days of cultivation respectively.Results The expression of MMP9 mRNA and TIMP3 mRNA in NCM460 cells of the experimental group 1,2 and 3 was significantly lower than that in the control group after one day of cultivation (P ＜ 0.05).After three and five days of cultivation,the expression of MMP9 mRNA in NCM460 cells of the experimental group 1 was significantly lower than that in the control group (P ＜ 0.05),but the expression of MMP9 mRNA in the NCM460 cells of the experimental group 2 and 3 was significantly higher than that in the control group and the experimental group 1 (P ＜ 0.05).The expression of MMP9 mRNA in NCM460 cells of the experimental group 3 was significantly higher than that in the experimental group 2 after five days of cultivation (P ＜ 0.05).The expression of MMP9 mRNA in NCM460 cells of the experimental group 1,2 and 3 after three and five days of cultivation was significantly higher than that after one day of cultivation(P ＜ 0.05).There was no significant difference in the expression of MMP9 mRNA in NCM460 cells of the experimental group 1 between three and five days of cultivation (P ＞ 0.05).The expression of MMP9 mRNA in NCM460 cells of the experimental group 2 and 3 after five days of cultivation was significantly higher than that after three days of cultivation(P ＜ 0.05).The expression of TIMP3 mRNA in NCM460 cells of the experimental group 2 and 3 was significantly higher than that in the experimental group 1 after one day of euhivation (P ＜ 0.05).After three days of cultivation,the expression of TIMP3 mRNA in NCM460 cells of the experimental group 1,2 and 3 was significantly lower than that in the control group (P ＜ 0.05);the expression of TIMP3 mRNA in NCM460 cells of the experimental group 2 was significantly lower than that in the experimental group 1 and 3 (P ＜ 0.05).After five days of cultivation,the expression of TIMP3 mRNA in NCM460 cells of the experimental group 1,2 and 3 was significantly higher than that in the control group (P ＜ 0.05).The expression of TIMP3 mRNA in NCM460 cells after three and five days of cultivation was significantly higher than that after one day of cultivation (P ＜ 0.05),and the expression of TIMP3 after five days of cultivation was significantly higher than that after three days of cultivation in the experimental group 1 (P ＜ 0.05).The expression of TIMP3 mRNA in NCM460 cells after three days of cultivation was significantly lower than that after one day of cultivation (P ＜ 0.05),and the expression of TIMP3 after five days of cultivation was significantly higher than that after one and three days of cultivation in the experimental group 2 (P ＜ 0.05).The expression of TIMP3 mRNA in NCM460 cells after five days of cultivation was significantly higher than that after one and three days of cultivation in the experimental group 3 (P ＜ 0.05).After five days of cultivation,there was no significant difference in the expression of MMP9 protein in NCM460 cells between the experimental group 1 and control group (P ＞ 0.05),the expression of MMP9 protein in NCM460 cells of the experimental group 2 and 3 was significantly higher than that in the control group and the experimental group 1 (P ＜ 0.05),but there was no significant difference in the expression of MMP9 protein in NCM460 cells between the experimental group 2 and 3 (P ＞ 0.05).After five days of cultivation,the expression of TIMP3 protein in NCM460 cells of the experimental group 1,2 and 3 was significantly higher than that in the control group (P ＜0.05);but there was no significant difference in the expression of TIMP3 protein in NCM460 cells among the experimental group 1,2and 3 (P ＞ 0.05).Conclusions The high concentration of magnesium-calcium alloy extract has certain influence on the expression of MMP9 and TIMP3 gene in NCM460 cells,which may lead to the early inflammatory reaction,and the mechanism may be related to the calcium ion concentration in the extract.

Case-control study on two osteotomy techniques for the treatment of distal radial malunion / 中国骨伤

<p><b>OBJECTIVE</b>Radial corrective osteotomy is an established but challenging treatment for distal radial malunion. There is an ongoing discussion about whether an opening or closing-wedge osteotomy between should employed. The purpose of the present study was to retrospectively compare the clinical and radio graphic results between conventional opening-wedge osteotomy and closing-wedge technique.</p><p><b>METHODS</b>From January 2004 and December 2012,42 patients with extra-articular distal radial malunion were managed with corrective osteotomy and were followed for a minimum of one year. Twenty-two patients (5 males and 17 females, ranging in age from 25 to 75 years old) were managed with radial opening-wedge osteotomy and implanting of interpositional bone graft or bone-graft substitute, and twenty patients (4 males and 16 females, ranging in age from 19 to 79 years) were managed with simultaneous radial closing-wedge and ulnar shortening osteotomy without bone graft. The selection of the surgical procedure was determined by the surgeon. Each patient was evaluated on the basis of objective radio graphic measurements, and functional outcomes were determined on the basis of clinical examinations, including range of wrist motion, grip strength, pain-rating score, Mayo wrist score, and Disabilities of the Arm, Shoulder and Hand (DASH) score.</p><p><b>RESULTS</b>The mean duration of follow-up was 36 months (ranged, 12 to 101 months) for the opening-wedge cohort and 28 months (ranged, 12 to 87 months) for the closing-wedge cohort. The two techniques were comparable in terms of complications. Post-operative volar tilt and ulnar variance were improved significantly in each cohort. The ulnar variance was more frequently restored to within defined criteria (22.5 to 0.5 mm) in the closing-wedge cohort than that in the opening-wedge cohort. The post-operative mean extension-flexion are of the wrist and Mayo wrist score were significantly better in the closing-wedge cohort. Differences in the pronation-supination arc, grip strength, pain-rating score, and DASH scores between these two cohorts were not significant.</p><p><b>CONCLUSION</b>The closing wedge osteotomy technique is an effective reconstructive procedure for the treatment of extra-articular distal radial malunion. It is significantly better than the opening-wedge osteotomy technique in terms of the restoration of ulnar variance, the extension-flexion arc of wrist motion, and the Mayo wrist score.</p>

Meta-analysis for the Association of Apolipoprotein E ε2/ε3/ε4 Polymorphism with Coronary Heart Disease / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Coronary heart disease (CHD) is a multifactorial disease and is thought to have a polygenic basis. Apolipoprotein E (APOE) gene is one such candidate with its common ε2/ε3/ε4 polymorphism in CHD. In recent years, numerous case-control studies have investigated the relationship of APOE polymorphism with CHD risk. However, the results are confusing.</p><p><b>METHODS</b>To clarify this point, we undertook a meta-analysis based on 14 published studies including 5746 CHD cases and 19,120 controls. Crude odds ratios (ORs) with 95% confidence intervals (CIs) were assessed for association using a random-effects or fixed-effects model using STATA version 10 (StataCorp LP, College Station, TX, USA).</p><p><b>RESULTS</b>Overall, the analysis showed that carriers of APOE ε2 allele decreased risk for CHD (ε2 allele vs. ε3 allele: OR = 0.82, 95% CI: 0.75-0.90, P < 0.001; ε2 carriers vs. ε3 carriers: OR = 0.81, 95% CI: 0.73-0.89, P < 0.001), compared with those carrying ε3 allele, especially in Caucasian population. However, those with ε4 allele had a significant increased risk for CHD (ε4 allele vs. ε3 allele: OR = 1.34, 95% CI: 1.15-1.57, P < 0.001), especially in Mongoloid population. Potential publication bias was observed in the genetic model of ε4 versus ε3, but the results might not be affected deeply by the publication bias. When we accounted for publication bias using the trim and fill method, the results were not materially alerted, suggesting the stability of our results.</p><p><b>CONCLUSIONS</b>Taken together, our meta-analysis supported a genetic association between APOE gene and CHD. ε4 increased the risk of CHD, whereas ε2 decreased the risk of CHD.</p>

Surgical treatment of refractory cerebrospinal fluid rhinorrhea with recurrent bacterial meningitis in 1 case and literature review / 中华实用儿科临床杂志

Objective To explore the surgical treatment of refractory cerebrospinal fluid rhinorrhea with recurrent bacterial meningitis.Methods The clinical data of 1 case of a 10-year-old boy who had 9 episodes of bacterial meningitis and underwent 4 surgical repair procedures for congenital cerebrospinal fluid rhinorrhea at the Children's Hospital Affiliated to Soochow University were analyzed,and the related literatures were reviewed.Results During the intervals of 9 episodes of bacterial meningitis,the patient experienced 4 neurosurgical repairs of cerebrospinal fluid rhinorrhea,including 2 endoscopic repairs via the lateral nasal cavity,a craniotomy approach repair via forehead epidural,and an endoscopic repair in combination with a ventriculo-peritoneal shunt.The first 3 surgeries were all failed,but the final surgery was successful,with no recurrence of cerebrospinal fluid rhinorrhea or bacterial meningitis in 3.5 years of follow-up.Conclusions For recurrent meningitis and refractory cerebrospinal fluid rhinorrhea,the ventriculo-peritoneal shunt can be considered in addition to conventional nasal endoscopic cerebrospinal fluid repair to eliminate the increased cerebrospinal fluid attributable to long-term chronic compensation,and effectively reduce postoperative intracranial hypertension to make the operation success.

Effect of MGF-Ct24E and mechanical stimulation on gene expression of osteoblasts / 医用生物力学

Objective To identify the differentially expressed genes of osteoblasts under the stimulation of mechano growth factor E peptide( MGF-Ct24E) and mechanical stress by microarray analysis. Methods Primary osteoblasts were cultured in vitro, which were subjected to mechanical stimulation(with the mechanical strain of 12% and frequency of 0.5 Hz) and MGF-Ct24E treatment(50 mg/L), respectively. The gene expression profiles were analysed by cDNA microarrys and quantitative PCR was used to validate the microarray data. ResultsCompared with the control group, 1 866 genes were found to have differentially expressed in the mechanical loading group, in which 1 113 genes were up-regulated, while 753 genes were down-regulated. 1 178 genes were found to have differentially expressed in the MGF-Ct24E group, in which 796 genes were up-regulated and 382 genes were down-regulated. GO analysis suggested that the gene expression profile of MGF-Ct24E group was consistent with that of the mechanical loading group and differentially expressed genes were mainly involved in cell proliferation and differentiation, response to mechanical stress and mechaotransduction. ConclusionsThe microarray analysis showed that MGF-Ct24E treatment had similar effects with the mechanical loading on the gene expression of osteoblasts, which might provide a novel approach to study the usage of MGF-Ct24E for treating bone repair in the absence of mechanical stimulation.