BACKGROUND: Current treatmemt strategy cannot efficiently relieve or reverse the disk degeneration, and neither surgical treatment nor nonsurgical treatment has gained satisfactory long-term effect. Therefore, more and more researches have focused on the cell therapy. OBJECTIVE: To explore the present states and application prospect of cell therapy, especially stem cells for degenerative intervertebral disc repair.METHODS: A computer-based search for related articles in PubMed database published between January 2011 and January 2016 using the English keywords of stem cell, intervertebral disk degeneration. Literatures addressing cell therapy for degenerative intervertebral disc repair were selected, and the articles published lately or original researches in authoritative journals were preferred.RESULTS AND CONCLUSION: A total of 205 articles were selected firstly, and 50 eligible articles were enrolled finally in accordance with the inclusion criteria. The mixtures of stem cells and carrier are injected into the degenerative intervertebral disk, to repair or displace abnormal cells. This strategy has been accepted by many researchers, and considered as a promising treatment. However, there is little evidence about the safety of clinical treatment with cell therapy, which still needs to be explored in depth.

The cis-epoxysuccinate hydrolase (CESH) from Rhizobium strain BK-20 is the key enzyme for L(+)-tartaric acid production. To establish a highly efficient and stable production process, we first optimized the enzyme production from Rhizobium strain BK-20, and then developed an immobilized cell-culture process for sustained production of L(+)-tartaric acid. The enzyme activity of free cells reached (3 498.0 +/- 142.6) U/g, and increased by 643% after optimization. The enzyme activity of immobilized cells reached (2 817.2 +/- 226.7) U/g, under the optimal condition with sodium alginate as carrier, cell concentration at 10% (W/V) and gel concentration at 1.5% (W/V). The immobilized cells preserved high enzyme activity and normal structure after 10 repeated batches. The conversion rate of the substrate was more than 98%, indicating its excellent production stability.
Alginates ; chemistry ; Cells, Immobilized ; Glucuronic Acid ; chemistry ; Hexuronic Acids ; chemistry ; Hydrolases ; metabolism ; Rhizobium ; enzymology ; metabolism ; Tartrates ; metabolism

Objective: To study the effects of different origins, collection and processing methods on the quality of Taraxacum mongolicum. Methods:The HPLC fingerprints of Taraxacum mongolicum were established. Totally 11 batches of Taraxacum mongoli-cum were analyzed by similarity evaluation and cluster analysis. Results:According to the results of HPLC, 11 batches of Taraxacum mongolicum had good baseline separation and showed 5 common peaks. Based on the established HPLC method, the quality of different batches of Taraxacum mongolicum showed difference according to the results of similarity evaluation and cluster analysis. The quality of batch 121231-1 was the best. Conclusion:The origin, collection and processing method show notable influence on the quality of Ta-raxacum mongolicum, and the comprehensive score method can be applied in the quality evaluation of Chinese herbs.

Gluconobacter oxydans converts glucose to gluconic acid and subsequently to 5-keto-D-gluconic acid (5-KGA), a precursor of industrially important L(+)-tartaric acid. To increase the yield of 5-KGA, fermentation conditions of 5-KGA production was optimized. Under the optimum medium and culture conditions in the shake flask, the highest 5-KGA production reached 19.7 g/L, increased by 43.8% after optimization. In a 5-L bioreactor, the pH was controlled at 5.5 and dissolved oxygen (DO) at 15%, 5-KGA production reached 46.0 g/L, raised at least 1.3 times than in the shake flask. Glucose feeding fermentation process was further developed, and the highest 5-KGA production of 75.5 g/L with 70% of yield was obtained, 32.0% higher than the highest reported value. Therefore, this newly developed fermentation process provided a practical and effective alternative for the commercial production of 5-KGA, and further of L(+)-tartaric acid.
Bioreactors ; Fermentation ; Gluconates ; metabolism ; Gluconobacter oxydans ; metabolism ; Glucose ; metabolism ; Industrial Microbiology ; Tartrates ; metabolism

Objective To compare the detecting results of rotavirus (RV)and adenovirus (AdV)antigens using auto stool pretreatment system (machine method)and manual method.Methods A total of 100 stools collectecd from diarrear patients admitted in gastroenterology outpatient department from September 2014 to Octorber 2014 in Peking University Medical College Hospital were detected to identify RV and AdV antigens using machine method and manual method respectively,and the nucletic acids of positive samples were detected by liquid chip method to verify the results.Results The RV,AdV and co-infection antigen positive detection rate using machine method were 17.0% (17/100),25.0% (25/100)and 12.0% (12/100)respectively,whereas those using the manual method were 4.0% (4/100),13.0% (13/100)and 2.0% (2/100),re-spectively.Taking the nucletic acids detection as the golden method,the false positive detection rate of RV,AdV and co-in-fection antigen using machine method were 23.5% (4/17),20.0% (5/25)and 33.3% (4/12)respectively,whereas those u-sing the manual method were 75.0% (3/4),69.2% (9/13)and 50.0% (1/2),respectively.χ2 test for paired data for RV and AdV positive detection rate,false positive detection rate of RV and false positive detection rate of AdV using two meth-ods were statistically significant (χ2 =15.0,52.8 and 47.5,P values <0.05).Two methods for detecting RV and AdV had poor consistency (Kappa value was 0.25,Kappa values <0.4).Conclusion Machine method has much more advantage on RV and AdV positive detection rate and false positive detection rate than manual method,which is good for clinical applica-tion.

Objective To investigate the clinical effects of minimally invasive vitrectomy combined with cataract surgery for myopic foveoschisis in young and middle-aged. Methods Sixty myopic eyes with myopic fo-veoschisis diagnosed from September 2013 to October 2015 in our hospital were included in the study. According to different surgical methods ,patients were divided into two groups:combined operation group(vitrectomy combined with lens replacement surgery,30 eyes)and staged operation group(simple vitrectomy group,30 eyes). Cataract surgery were performed according to the opacity of lens. All the patients were followed up for 6~12 months. Reop-eration rate of staged operation group ,the best corrected visual acuity(BCVA),the distant and near visual acuity and retinal reattachment rate of each group were observed. Every patient was interviewed with a questionnaire about visual quality at 6th month after surgery. Results 27 eyes developed lens opacity after operation(90%)in staged operation group,among them,21 eyes underwent secondary cataract surgery(70%). The BCVA recovery,postop-erative distant vision,and preoperative and postoperative distant vision difference in the combined operation group were significantly higher than those in the staged group(P<0.05). There were no significant differences in macu-lar retinal condition and retinal reattachment rate between the two groups(P > 0.05). Six months after operation, the visual quality of the combined operation group was significantly better than that of the staged group(t=-3.95, P = 0.00). In the combined surgery group,the scores of distant vision,stereopsis and visual fatigue were higher than those of staged operation group. The score of dry eye and night glare in staged operation group was higher than that in combined operation group(all P<0.05). Conclusions Vitrectomy combined with intraocular lens replace-ment for myopic foveoschisis in young and middle-aged people can avoid second operations and improves the visual function,proving to be a feasible operation in clinic.

Objective:To explore the efficient construction of HSF1 gene knockout mouse model using CRISPR/Cas9 gene editing technology, and to establish the early basis for the mouse model of primary osteosarcoma.Methods:According to exon 9 of HSF1 gene structure, the corresponding GRNA (guideRNA) was selected and screened. Then the transcription template of sgRNA (small guide RNA) was amplified by PCR, and four up stream primers were obtained. Subsequently, sgRNA was transcribed in vitro and screened by Tube Screen platform to screen the sgRNA with effective cutting, and the sgRNA with the highest cutting efficiency was selected from the screening results for subsequent experiments. The transcription template of SPCas9mRNA was amplified by PCR, and then Cas9mRNA was transcribed in vitro. The sgRNA transcribed in vitro and Cas9mRNA were injected into the fertilized eggs of healthy C57BL/6 mice, and the tissue was extracted from the tail of the born mice and identified by PCR sequencing. Heterozygous female mice of F0 generation were selected to mate with wild-type male mice too btain F1 generation off spring. The mutation of gene bases of F1 generation mice was detected by AGAR gel electrophoresis and gene sequencing. The heterozygous male mice of the F1 generation and female mice of the F0 generation were back crossed to obtain the F2 generation daughter mice. The tail tissues were cut and sequenced to obtain the F2 generation homozygous knockout mice. PCR was used to observe the cutting efficiency of sgRNA and the sequencing of rat tail tissue, and SNAPGene software was used for gene sequence alignment to determine the deletion of base fragments.Results:The up stream primers sgRNA-1 Primer-f, sgRNA-2 Primer-f, sgRNA-3 Primer-f, sgRNA-4 Primer-f and down stream primers sgRNA-4 Primer -r were obtained by PCR amplification. After in vitro tran scription and screening of sgrRNA, sgrRNA-1, sgrRNA-2 and sgrRNA-4 had high cleavage efficiency and were selected for subsequent experiments. T7 promoter was added to the 5 'end of Cas9 mRNA, and Cas9 mRNA was obtained by PCR and in vitro transcription kit. Mixed Cas9-sgRNA solution was injected into the fertilized eggs of mice and cultured. The cultured two-cell fertilized eggs were injected into the ampulla of the pseudo pregnant female mice, and the F0 generation mice were obtained successfully. A total of 8 heterozygous mice of F0 generation were obtained by Agar gel electrophoresis. Three heterozygous knockout mice of F1 generation were obtained by breeding the female heterozygous mice of F0 generation with healthy wild-type male mice and PCR and sequencing. Three heterozygous male mice of F1 generation were back crossed with female mice of F0 generation 3 to obtain F2 generation mice. Through the observation of electrophoresis and sequencing results of F2 generation mice, it was confirmed that 7 mice were missing HSF1 base sequence, and the electrophoresis results showed mutant bands and no wild-type bands, which were identified as homozygous. The F2 generation homozygous mice were able to breed stably. As eries of results proved that the HSF1 gene knockout mouse model was successfully established in this experiment.Conclusion:CRISPR/Cas9 technology was successfully used to construct HSF1 gene knockout mouse model, with strong stability and high reproducibility, which laida foundation for further study of HSF1 gene expression products and establishment of mouse model of primary osteosarcoma.

OBJECTIVETo synthesize novel aryl-substituent benzyl acid compounds targeting HIV gp41 and characterize their anti-HIV activities.

METHODSTwelve analogues of aryl-substituent benzyl acid were designed and synthesized by Suzuki- Miyaura cross-coupling and Knoevenagel condensation reactions using halo-benzyl acid or 3-carboxybenzeneboronic acid as the raw material. The inhibitory activities of these compounds on gp41 six-helix bundle formation were tested by ELISA, and their anti-HIV activities were determined using a luciferase assay.

RESULTSThe structures of the compounds were characterized by nuclear magnetic resonance and mass spectrography. Among the 12 compounds, 5 (7b, 7c, 7d, 7e, and 7g) could inhibit the gp41 six-helix bundle formation, and 7d showed the most potent effect, and could also inhibit the replication of HIV-1 SF33 strain with an IC(50) of 20 µmol/L.

CONCLUSIONThe synthesized aryl-substituent benzyl acid compound 7d could inhibit HIV replication by blocking the gp41 six-helix bundle formation.

Anti-HIV Agents ; chemical synthesis ; pharmacology ; Benzoates ; chemical synthesis ; pharmacology ; Drug Design ; HIV-1 ; drug effects ; Hydrocarbons, Aromatic ; chemical synthesis ; pharmacology ; Virus Replication ; drug effects

Evidence-based medicine (EBM) is a kind of clinic practice where clinicians use the best and the latest available evidence to diagnose and treat patients, and both evidence providers and users need to identify and control different kinds of biases in medical research.Directed acyclic graphsis is a tool to explore the causal relationship.The possible biases in the study can be revealed in a simple graphical language.The use of directed acyclic graphs could avoid the occurrence of bias and improve the quality of medical research and better guide clinical practice.

BACKGROUND:Quercetin plays an important role in the proliferation and differentiation of bone marrow mesenchymal stem cells,but less research has been done on its mechanism of promoting the migration of bone marrow mesenchymal stem cells. OBJECTIVE:To study the effect of quercetin on the migration of human bone marrow mesenchymal stem cells through in vitro experiments,and to explore the regulatory role of CCR1 and CXCR4. METHODS:Human bone marrow mesenchymal stem cells were selected as experimental subjects.CCK8 assay was used to detect the effect of quercetin on the proliferative activity of human bone marrow mesenchymal stem cells.Cell scratch assay and Transwell assay were used to detect the in vitro invasive and migratory abilities of human bone marrow mesenchymal stem cells after quercetin treatment,respectively.The role of quercetin in relation to CCR1 and CXCR4 was demonstrated with the help of molecular docking technology.Western blot assay and real-time fluorescence quantitative PCR were used to detect the migration-related chemokine expression after quercetin treatment. RESULTS AND CONCLUSION:(1)5 and 10 μmol/L quercetin could significantly promote the proliferation of human bone marrow mesenchymal stem cells,and the drug concentration of 10 μmol/L resulted in the highest cell proliferation efficiency.(2)To better explore the dose-effect relationship of quercetin affecting the migration of human bone marrow mesenchymal stem cells,5 and 10 μmol/L quercetin were selected for the subsequent experiments,and ligustrazine was used as the positive control drug,and the experiments were divided into blank control group,5 μmol/L quercetin group,10 μmol/L quercetin group,and 100 μmol/L ligustrazine group.(3)In vitro migration and invasion ability of human bone marrow mesenchymal stem cells were elevated in a concentration-dependent manner after quercetin treatment,and the migration effect of 10 μmol/L quercetin group was better than that of ligustrazine group.(4)The molecular docking results suggested that there was a strong interaction between quercetin and CCR1 and CXCR4.(5)Quercetin could up-regulate the expression of CCR1 and CXCR4 proteins and mRNA.(6)This study confirmed at the cellular level that quercetin could promote the migration of human bone marrow mesenchymal stem cells by targeting CCR1 and CXCR4.