Y-shaped DNA nanostructures assembled-spherical nucleic acids as target converters to activate CRISPR-Cas12a enabling sensitive ECL biosensing.

Considering the trans-cleavage capabilities, high-specificity and programmability, the CRISPR-Cas system has been recognized as a valuable platform to develop the next-generation diagnostic biosensors. However, due to the natural interaction with nucleic acids, current CRISPR-Cas-based detection mostly applies in nucleic acid analysis rather than non-nucleic acid analysis. By virtue of spherical nucleic acids (SNAs) with programmability and specificity, the Y-shaped DNA nanostructures assembled-SNAs (Y-SNAs) were rationally designed as target converters to achieve the quantitative activation of CRISPR-Cas12a, enabling a highly specific and sensitive electrochemiluminescence (ECL) determination of alpha-methylacyl-CoA racemase (AMACR), a high specific protein biomarker of prostate cancer. Significantly, the Y-shaped DNA nanostructures comprised of assisted DNA (A1), AMACR aptamer and DNA activator of CRISPR-Cas12a were loaded on Au nanoparticles modified Fe3O4 magnetic beads (Au@Fe3O4 MBs) to construct the robust Y-SNAs. In the presence of the target AMACR, the Y-SNAs as target converters could achieve quantitative activation of CRISPR-Cas12a by outputting the DNA activators with a linear relationship to the target. The amplified ECL signals were triggered by the release of the ferrocene-labeled quenching probes (QPs) on the electrode surface due to the trans-cleavage activity of CRISPR-Cas12a, thereby realizing the sensitive ECL determination of AMACR from 10 ng/mL to 100 µg/mL with the detection limit of 1.25 ng/mL. In general, this approach provides novel perspectives on how to design a universal ECL platform of the CRISPR-Cas system to detect the non-nucleic acid targets beyond the traditional methods.

Fire-needle acupuncture for upper limb spastic paralysis after stroke: Study protocol for a randomized controlled trial / 中西医结合学报

BACKGROUND@#Fire-needle acupuncture, an important kind of acupuncture therapy, has been clinically used to treat upper limb spastic paralysis (ULSP) after stroke. Clinical experience has indicated that fire-needle acupuncture treatment takes less time, requires fewer visits, and has more rapid results and fewer side effects compared to chemical medicine alternatives. This study will evaluate the effects of fire-needle acupuncture for ULSP in the context of standardized clinical research and provide high-quality data to inform clinical procedures and future study design.@*METHODS/DESIGN@#A randomized controlled trial will be carried out to evaluate the effects of fire-needle acupuncture therapy in patients with ULSP from stroke. ULSP patients (n = 120) will be recruited at Changhai Hospital in Shanghai, China. Patients will be randomly divided into three groups, including fire-needle acupuncture group (FAG), filiform-needle acupuncture group (FFAG) and rehabilitation treatment group (RTG). During the 3-week treatment, the FAG will be treated every two days, while FFAG and RTG will be treated 5 d in a row and then rest for 2 d. The Simplified Fugl-Meyer Motor Function Scale and Modified Ashworth Scale will be used as the primary outcome measures. Statistical analysis will be conducted by an independent statistician.@*DISCUSSION@#Through this study, the utility of fire-needle acupuncture in treating ULSP after stroke will be tested, and some specific claims of fire-needle acupuncture therapy will be evaluated, such as relieving spasm and muscular tension, improving activities of daily living, rapidity of response and less frequency of treatment compared with other treatments.@*TRIAL REGISTRATION@#Chinese Clinical Trial Registry (identifier: ChiCTR-IOR-17013875; registration date: 28 December 2016).

Identification of cytotoxic T-lymphocyte epitopes from hepatocellular car-cinoma antigen MAGEC2 / 中国病理生理杂志

AIM:To observe whether modified epitopes from hepatocellular carcinoma antigen MAGEC 2 have HLA-A2-restricted antitumor ability.METHODS:HLA-A2 epitopes from MAGEC2 protein were predicted by NetCTL 1. 2,SYFPEITHI and IEDB.The change of binding anchor motifs by replacing anchor residues created the modified peptides from MAGEC2.The binding affinity of the peptides to HLA-A*0201 molecule was evaluated by T2 cells binding assay. ELISPOT assay and intracellular cytokine staining were used to investigate the ability of the peptides inducing specific re -stricted cytotoxic T-lymphocytes(CTLs)to release interferon-γ(IFN-γ).The ability of the peptides to induce T-cell re-sponse was investigated by cytotoxicity assay in vitro.RESULTS:The candidate peptides P248, P248-1Y,P356,P356-1Y,P356-2L and P356-1Y2L showed moderate affinity toward HLA-A2 molecule.T2 binding assay showed that P248-1Y and P356-1Y2L showed significantly higher affinity for HLA-A2 than the native peptides.ELISPOT assay and intracellular cytokine staining showed P248, P248-1Y, P356 and P356-1Y2L were able to induce specific CTLs to release IFN-γ. ELISPOT assay showed that significantly higher levels of IFN-γrelease were induced by P248-1Y and P356-1Y2L than the native peptides.The CTLs induced by P248,P248-1Y,P356 and P356-1Y2L lysed HepG2 cells,and P248-1Y and P356-1Y2L peptide-specific CTLs showed higher cytotoxicity against HepG 2 cells than the native peptide-specific CTLs(P<0.05).CONCLUSION: Compared with the native peptides, modified epitopes P248-1Y and P356-1Y2L have higher binding affinity with HLA-A2 and retain immunogenecity.In addition, the antitumor immunity effects of modified epitope P248-1Y and P356-1Y2L are stronger than the native peptides.The peptides P248-1Y and P356-1Y2L are excellent HLA-A2-restricted CTL epitopes from tumor antigen MAGEC 2, which could serve as new candidates towards antitumor peptide vaccines.

Effect of ω-3 polyunsaturated fatty acid on oxidative stress production and apoptosis in hippocampus of neonatal rats with brain injury induced by lipopolysaccharide / 新乡医学院学报

Objective To investigate the effects of ω-3 polyunsaturated fatty acid(PUFA)on oxidative stress produc-tion and apoptosis in hippocampus of neonatal rats with brain injury induced by lipopolysaccharide(LPS). Methods Forty-eight neonatal Sprague Dawley rats(3 days)were randomly divided into control group,LPS group,ω-3 PUFA group and ω-6 PUFA group,with 12 rats in each group. The rats in the LPS group,ω-3 PUFA group and ω-6 PUFA group were given 0. 6 mg·kg - 1 LPS via intraperitoneal injection,then equal volume of saline,ω-3 PUFA and ω-6 PUFA was immediately given via intraperitoneal injection respectively;while the rats in the control group were all given equal volume of saline. The rats in each group were sacrificed at 24 hours after intraperitoneal injection with saline or fat emulsions to obtain the hippocampus. The levels of superoxide dismutase(SOD)and malondialdehyde(MDA),reduced glutathion(GSH),oxidized glutathione(GSSG) were detected and GSSG/ GSH was calculated. The apoptotic index was measured by terminal deoxynucleotidyl transferase-me-diated dUTP nick end labeling. Results Compared with the control group,the levels of SOD and GSH in hippocampus of rats in the LPS group,ω-6 PUFA group and ω-3 PUFA group were significantly decreased(P < 0. 05),and the levels of MDA, GSSG and the ratio of GSSG/ GSH were significantly increased(P < 0. 05). Compared with the LPS group,the levels of SOD and GSH in hippocampus of rats in ω-6 PUFA group were significantly decreased(P < 0. 05),the levels of MDA,GSSG and the ratio of GSSG/ GSH were significantly increased(P < 0. 05);the levels of SOD and GSH in hippocampus of rats in ω-3 PU-FA group were significantly increased(P < 0. 05),the levels of MDA,GSSG and the ratio of GSSG/ GSH were significantly de-creased(P < 0. 05). Compared with the ω-6 PUFA group,the levels of SOD and GSH in hippocampus of rats in ω-3 PUFA group were significantly decreased(P < 0. 05),the levels of MDA,GSSG and the ratio of GSSG/ GSH were significantly in-creased(P < 0. 05). The apoptotic index in the LPS group,ω-6 PUFA group and ω-3 PUFA group was higher than that in the control group(P < 0. 05). The apoptotic index in the ω-6 PUFA group was higher than that in the LPS group(P < 0. 05). The apoptotic index in the ω-3 PUFA group was lower than that in the LPS group and ω-6 PUFA group(P < 0. 05). Conclusion ω-3 PUFA can alleviate the oxidative stress,and decrease the apoptosis of hippocampus in neonatal rats with brain injury in-duced by LPS. So it has a neuroprotective effect in brain injury induced by LPS.

An HPLC-MS/MS method for the determination of trans-ferulic acid in human plasma / 药学学报

In this study, we developed a sensitive and rapid HPLC-MS/MS method for the determination of trans-ferulic acid (trans-FA) in plasma samples, and investigated the pharmacokinetics characteristics in healthy volunteers. The plasma samples were extracted with acetic ether, and then separated on a Hedera ODS-2 column with a mobile phase of methanol and 5 mmol·L-1 ammonium acetate buffer solution containing 0.05% acetic acid (34:66) at a flow rate of 0.4 mL·min-1. Electrospray ionization source was applied and operated in the positive ion mode using MRM. The method exhibited a good linearity over the concentration range of 0.1-5 ng·mL-1 (r ≥ 0.9992). The values on both the occasions (intra-and inter-day) were all within 9.2%, and the accuracy was 95.4%-111.4%. No matrix effect and carry-over effect were observed. Trans-FA was stable in human plasma under different storage conditions. The developed HPLC-MS/MS method is rapid, sensitive, accurate, and reproducible, and suitable for the pharmacokinetic study of trans-FA in healthy Chinese volunteers.

Professor He Pu-ren: the founder of Santong method of acupuncture and moxibustion / 中国针灸

Professor He Pu-ren, the founder of Santong method of acupuncture and moxibustion, is a well known acupuncturist at home and abroad. His main contributions include combined martial arts and Chinese medicine, showing obvious therapeutic effect; taking part in establishment of The Department of Acupuncture, Beijing Chinese Medicine Hospital; creating Santong method of acupuncture and moxibustion; advocating fire needle therapy; writing medical books and teaching students; advocating the culture of acupuncture; making the metal model of acupuncture and moxibustion, and others. His achievements have become an important part of acupuncture and moxibustion science.

Changes of cell adhesion molecules and proinflammatory cytokines during cardiopulmonary bypass and the effect of intervention / 中南大学学报(医学版)

OBJECTIVE@#To investigate the changes of the cell adhesion molecules and proinflammatory cytokines during cardiopulmonary bypass, and to observe the effect of intervention with ulinastatin.@*METHODS@#Twenty-two ASA II-III patients (9 males, 13 females), aged 20-60 years, undergoing cardiac operation with CPB were randomly divided into 2 groups: the control group (Group C, n=11) and the ulinastatin group (Group W, n=11). In Group W, patient received ulinastatin 1.2 x 10(4) U/kg, and half of the dose was given intravenously after the induction of anesthesia, while the same amount of ulinastatin added into the primary solution. And in Group C, normal saline was given instead of ulinastatin. Blood samples were taken from radial artery before the operation (T1), 20 min after the initiation of CPB (T2), 1 h (T3), 6 h (T4 ), 24 h (T5) after the CPB for the determination of plasma TNF-alpha, IL-6, sICAM-1 and sP-Selectin concentrations.@*RESULTS@#The concentrations of TNF-alpha, IL-6 increased significantly at T2-T4 in both groups compared with T1 (P < 0.05), and returned to the baseline level at Ts in Group W. The concentrations of TNF-alpha, IL-6 in Group C at T2-T5 were higher than that in Group W (P < 0.01). The concentrations of sICAM-1, sP-Selectin increased significantly at T3, T4 in both groups compared with that at T1 (P < 0.05). But at T5, the concentrations of sICAM-1, sP-Selectin decreased, especially in Group W the concentrations of sICAM-1, sP-Selectin returned to the baseline level. The sI-CAM-1, sP-Selectin concentrations in group C at T4, T5 were higher than that in group W (P < 0.05).@*CONCLUSION@#Ulinastatin can reduce the increase of the cell adhesion molecules and proinflammatory cytokines during cardiopulmonary bypass and effectively weaken the inflammatory response to CPB.

Effects of ulinastatin on cerebral inflammatory response during cardiopulmonary bypass / 中南大学学报(医学版)

OBJECTIVE@#To investigate the effects of ulinastatin (UTI) on cerebral inflammatory response during cardiopulmonary bypass (CPB).@*METHODS@#Twenty-four NYHA II-III patients (13 males and 11 females) aged 23-45 years, undergoing elective cardiac valve replacement under hypothermic CPB were randomly divided into 2 groups: ulinastatin group (Group U, n=12) and control group (Group C, n=12). In group U, UTI (1.2 x 10(4) U/kg) was given intravenously after the induction of anesthesia, 0.6 x 10(4) U/kg UTI was added to the priming solution, and 0.6 x 10(4) U/kg UTI was given about 5 min before the aortic decamping. In Group C, normal saline was given instead of UTI. Internal jugular vein was cannulated and the catheter was advanced retrogradely till jugular bulb. Blood samples were taken simultaneously from artery and jugular bulb after induction of anesthesia (T1), 60 min (T2) and 6 h (T3) after discontinuation of CPB for determination of TNFalpha, IL-6, IL-8 and IL-10. The juguloarterial gradients of these cytokines (deltaTNFalpha, deltaIL-6, deltaIL-8, and deltaIL-10) were calculated.@*RESULTS@#In Group C, arterial levels of TNFalpha, IL-6, IL-8, IL-10 at T2 and T3, deltaTNFalpha, deltaIL-8 and deltaIL-10 at T2, deltaTNFalpha, deltaIL-6 and deltaIL-10 at T3 significantly increased (P < 0.01). deltaIL-8 increased at T3 (P < 0.05). In Group U, arterial levels of IL-6, IL-8, IL-10 at T2, arterial levels of IL-6, IL-8,IL-L-10 and deltaTNFalpha, deltaIL-8 at T3 significantly increased (P < 0.01). Arterial levels of TNFalpha at T2 and T3, deltaTNFalpha, deltaIL-10 at T2, deltaIL-6 at T3 increased (P < 0.05). Arterial levels of TNFalpha, IL-6 and deltaTNFalpha, deltaIL-8 at T2, arterial levels of TNFalpha and deltaIL-6 at T3 in Group U were lower than those in Group C (P < 0.05). Arterial levels of IL-6 at T3, IL-8 at T2 and T3 in Group U were significantly lower than those in Group C (P < 0.01). Arterial levels of IL-10 and deltaIL-10 at T3 in Group U were higher than those in Group C (P < 0.05).@*CONCLUSION@#Systemic and cerebral activation of inflammatory response during CPB can be alleviated by ulinastatin.