An injectable, self-healing, electroconductive hydrogel loaded with neural stem cells and donepezil for enhancing local therapy effect of spinal cord injury.

BACKGROUND: Spinal cord injury (SCI) is a serious injury with high mortality and disability rates, and there is no effective treatment at present. It has been reported that some treatments, such as drug intervention and stem cell transplantation have positive effects in promoting neurological recovery. Although those treatments are effective for nerve regeneration, many drawbacks, such as low stem cell survival rates and side effects caused by systemic medication, have limited their development. In recent years, injectable hydrogel materials have been widely used in tissue engineering due to their good biocompatibility, biodegradability, controllable properties, and low invasiveness. The treatment strategy of injectable hydrogels combined with stem cells or drugs has made some progress in SCI repair, showing the potential to overcome the drawbacks of traditional drugs and stem cell therapy. METHODS: In this study, a novel injectable electroactive hydrogel (NGP) based on sodium hyaluronate oxide (SAO) and polyaniline-grafted gelatine (NH2-Gel-PANI) was developed as a material in which to load neural stem cells (NSCs) and donepezil (DPL) to facilitate nerve regeneration after SCI. To evaluate the potential of the prepared NGP hydrogel in SCI repair applications, the surface morphology, self-repairing properties, electrical conductivity and cytocompatibility of the resulting hydrogel were analysed. Meanwhile, we evaluated the neural repair ability of NGP hydrogels loaded with DPL and NSCs using a rat model of spinal cord injury. RESULTS: The NGP hydrogel has a suitable pore size, good biocompatibility, excellent conductivity, and injectable and self-repairing properties, and its degradation rate matches the repair cycle of spinal cord injury. In addition, DPL could be released continuously and slowly from the NGP hydrogel; thus, the NGP hydrogel could serve as an excellent carrier for drugs and cells. The results of in vitro cell experiments showed that the NGP hydrogel had good cytocompatibility and could significantly promote the neuronal differentiation and axon growth of NSCs, and loading the hydrogel with DPL could significantly enhance this effect. More importantly, the NGP hydrogel loaded with DPL showed a significant inhibitory effect on astrocytic differentiation of NSCs in vitro. Animal experiments showed that the combination of NGP hydrogel, DPL, and NSCs had the best therapeutic effect on the recovery of motor function and nerve conduction function in rats. NGP hydrogel loaded with NSCs and DPL not only significantly increased the myelin sheath area, number of new neurons and axon area but also minimized the area of the cystic cavity and glial scar and promoted neural circuit reconstruction. CONCLUSIONS: The DPL- and NSC-laden electroactive hydrogel developed in this study is an ideal biomaterial for the treatment of traumatic spinal cord injury.

A novel enzyme-linked immunostaining technique based on silk membrane for the prenatal detection of fetomaternal haemorrhage.

Objective: Developing a simple, rapid, reliable, sensitive, and cost-effective method for prenatal detection of fetomaternal haemorrhage by combining multi-aperture silk membrane with enzyme-linked immunosorbent assay (ELISA), which does not require any complicated instruments and can be visually colored, so as to provide a new method for clinical detection of fetomaternal haemorrhage. Methods: As a carrier, a chemically treated silk membrane was used to immobilize anti-A/anti-B antibody reagent. PBS washed slowly after vertically dropping red blood cells. After adding biotin-labeled anti-A/anti-B antibody reagent, PBS is slowly washed, enzyme-labeled avidin is added, and TMB is used for color development after washing. Results: When there were both anti-A and anti-B fetal erythrocytes in pregnant women's peripheral blood, the final color was dark brown. When there are no anti-A and anti-B fetal red blood cells in pregnant women's peripheral blood, the final color development results do not change, which corresponds to the color of chemically treated silk membrane. Conclusion: The new enzyme-linked immunosorbent assay (ELISA) based on a silk membrane can distinguish fetal red blood cells from maternal red blood cells prenatally and can be used for prenatal detection of fetomaternal haemorrhage.

Prenatal diagnosis of fetomaternal hemorrhage by a novel hydrogel fluoroimmunoassay that accurately quantifies fetal haemoglobin.

Objective: Fetomaternal hemorrhage (FMH) is an alloimmunization resulting caused by the incompatibility between fetal and maternal blood. For the prevention of newborn haemolytic disease (HDN), it is crucial to quantify the amount of fetomaternal hemorrhage. However, the classical Kleihauer-Betke test (K-B test) for detecting fetomaternal hemorrhage is limited by experimental tools and conditions and is not suitable for routine clinical use. Consequently, the method of prenatal diagnosis of fetomaternal hemorrhage applicable to the clinic is a topic worthy of further study. Therefore, it is worthwhile to further investigation on the clinically applicable prenatal diagnosis method for fetomaternal hemorrhage. Methods: This experiment demonstrates hydrogel's ability to separate sensitized red blood cells from soluble antibodies. Using flow cytometry the fluorescence values of sensitized red blood cells and fluorophore-labeled antibodies were measured, and the testing steps for the detection products of a novel technology were determined. The properties of a hydrogel fluoroimmunoassay were evaluated by distinguishing between the amounts of fetal and adult haemoglobin. The precision of this technology is evaluated using the Kleihauer-Betke test as a comparison. Results: This experiment compared the detection of haemoglobin fluorescence in adults (n = 2) and fetuses (n = 6). At the same time, the fluorescence intensity of different fetal haemoglobin (HbF) in adult haemoglobin (HbA) was calculated. The fluorescence value is 1.6% when the fetal hemoglobin concentration is 0.1%. Conclusion: The novel hydrogel fluoroimmunoassay can accurately determine the fluorescence intensity by flow cytometry to differentiate fetal haemoglobin from adult haemoglobin, quantitatively prenatally diagnose fetal haemoglobin, address the incompatibility between fetal and maternal blood types, and prevent alloimmunization.

The development and application of a novel reagent for fixing red blood cells with glutaraldehyde and paraformaldehyde.

OBJECTIVE: The currently employed red blood cell reagents have a short shelf life. Some hospitals with a small number of specimens will be unable to utilize them within the validity period, resulting in a substantial increase in the purchase price. Therefore, the method of developing long-term red blood cell reagents is a problem worthy of further study. METHODS: In this experiment, the type and concentration of the red blood cell reagent treatment solution were evaluated based on the red blood cell antigen concentration 24 h after treatment. In addition, the qualified glutaraldehyde/paraformaldehyde reagent was stored for six months, and five red blood cell indices were measured every month. At the same time, the detection indices of treated red blood cell reagents and untreated red blood cell reagents were compared. RESULTS: It was discovered that treated red blood cells containing 0.005% GA and 0.05% PFA were more suitable for the preservation of red blood cells than other treated concentrations, and the preservation time could reach six months. The test tube method (n = 24) and microcolumn gel card (n = 35) were used to determine the accuracy of the treated blood cells containing 0.005% glutaraldehyde +0.05% paraformaldehyde, with an accuracy of 100%. CONCLUSION: This experiment resulted in the development of a novel reagent for treating red blood cells with glutaraldehyde/paraformaldehyde fixed solution that can effectively prolong its storage time by two to three times that of red blood cell reagents currently on the market.

Transcription Factor MafB Suppresses Type I Interferon Production by CD14+ Monocytes in Patients With Chronic Hepatitis C.

Transcription factor MafB regulates differentiation and activity of monocytes/macrophage and is associated with the development of atherosclerosis and cancers. However, the role of MafB in modulation of CD14+ monocytes in chronic viral hepatitis was not fully elucidated. Thus, the aim of current study was to investigate the immunoregulatory function of MafB to type I interferon (IFN) secretion by CD14+ monocytes and its contribution to pathogenesis of chronic hepatitis C virus (HCV) infection. A total of 29 chronic hepatitis C patients and 21 healthy individuals were enrolled. Serum IFN-α1 and IFN-ß was measured by ELISA, while MafB mRNA and protein expression were assessed by real-time PCR and Western blot. MafB siRNA or MafB expression plasmid was transfected into purified CD14+ monocytes to suppress or increase MafB expression. The function of MafB siRNA transfected CD14+ monocytes to HCV in cell culture (HCVcc)-infected Huh7.5 cells or CD4+ T cells was also investigated in direct and indirect contact co-culture system. Serum IFN-α1 and IFN-ß was robustly reduced in chronic hepatitis C patients. By contrast, MafB was notably elevated in chronic hepatitis C patients and negatively correlated with serum IFN-α1. Overexpression of MafB reduced the IFN-α1 production by CD14+ monocytes from healthy individuals. However, MafB inhibition elevated IFN-α1 secretion by CD14+ monocytes and interferon regulatory factor 3 phosphorylation in chronic hepatitis C. MafB inhibition also promoted CD14+ monocytes-induced viral clearance in HCVcc-infected Huh7.5 cells by up-regulation of IFN-α1 and IFN-ß without increasingly destroying hepatocytes, however, did not affect CD14+ monocytes-induced CD4+ T cells differentiation in chronic hepatitis C patients. The current data revealed that overexpression of MafB in chronic hepatitis C patients might suppress type I IFN production by CD14+ monocytes, leading to the viral persistence. MafB might be a potential therapeutic target for treatment of chronic hepatitis C.

Gastrin promotes angiogenesis by activating HIF-1α/ß-catenin/VEGF signaling in gastric cancer.

Angiogenesis is recognized as a sign of cancer and facilitates cancer progression and metastasis. Suppression of angiogenesis is a desirable strategy for gastric cancer (GC) management. In this study, we showed a novel role of gastrin in angiogenesis of GC. We observed that treatment with gastrin 17 (G17) increased the proliferation of AGS cells and enhanced tube formation during normoxia and hypoxia. The expression level of VEGF were increased by G17 treatment as well. Experiments on the mechanism showed that G17 promoted HIF-1α expression, which subsequently enhanced ß-catenin nuclear localization and activation of TCF3 and LEF1 and finally resulted in angiogenesis by upregulating VEGF. An in vivo experiment confirmed that G17 enhanced GC cell proliferation and angiogenesis in the resultant tumor. In conclusion, our findings indicate that gastrin promotes angiogenesis via activating HIF-1α/ß-catenin/VEGF axis in GC.

Identification and functional characterization of intermediate-size non-coding RNAs in maize.

BACKGROUND: The majority of eukaryote genomes can be actively transcribed into non-coding RNAs (ncRNAs), which are functionally important in development and evolution. In the study of maize, an important crop for both humans and animals, aside from microRNAs and long non-coding RNAs, few studies have been conducted on intermediate-size ncRNAs. RESULTS: We constructed a homogenized cDNA library of 50-500 nt RNAs in the maize inbred line Chang 7-2. Sequencing revealed 169 ncRNAs, which contained 58 known and 111 novel ncRNAs (including 70 snoRNAs, 27 snRNAs, 13 unclassified ncRNAs and one tRNA). Forty of the novel ncRNAs were specific to the Panicoideae, and 24% of them are located on sense-strand of the 5' or 3' terminus of protein coding genes on chromosome. Target site analysis found that 22 snoRNAs can guide to 38 2'-O-methylation and pseudouridylation modification sites of ribosomal RNAs and small nuclear RNAs. Expression analysis showed that 43 ncRNAs exhibited significantly altered expression in different tissues or developmental stages of maize seedlings, eight ncRNAs had tissue-specific expression and five ncRNAs were strictly accumulated in the early stage of leaf development. Further analysis showed that 3 of the 5 stage-specific ncRNAs (Zm-3, Zm-18, and Zm-73) can be highly induced under drought and salt stress, while one snoRNA Zm-8 can be repressed under PEG-simulated drought condition. CONCLUSIONS: We provided a genome-wide identification and functional analysis of ncRNAs with a size range of 50-500 nt in maize. 111 novel ncRNAs were cloned and 40 ncRNAs were determined to be specific to Panicoideae. 43 ncRNAs changed significantly during maize development, three ncRNAs can be strongly induced under drought and salt stress, suggesting their roles in maize stress response. This work set a foundation for further study of intermediate-size ncRNAs in maize.

Neonatal alloimmune thrombocytopenia caused by anti-HPA antibodies in pregnant Chinese women: a study protocol for a multicentre, prospective cohort trial.

BACKGROUND: Neonatal alloimmune thrombocytopenia (NAIT), caused by maternal antibodies raised against alloantigens carried on foetal platelets, is a very common haematological abnormality in newborns worldwide. However, baseline data on NAIT in China are lacking. Therefore, this study seeks to explore the incidence of alloantibody against the human platelet antigen (HPA) in pregnant women and its associations with NAIT in China. METHODS: A multicentre, prospective cohort study design will be used, and 55,497 pregnant women will be recruited for the first screening of the anti-HPA antibody at 12 to 28 weeks of gestational age. Subjects who are positive in the first screening for the anti-HPA antibody will be included in the exposure group. Re-tests of the antibody titre, antigen-specificity and genotyping of HPA and HLA will be conducted during admission. A ratio of 1:1 paired individuals with the same ethnicity and parity but testing negative for the anti-HPA antibody will be randomly selected to be included in the non-exposure group. NAIT will be diagnosed in the newborns on day one of the birth. The HPA of the neonates in the exposure group will also be genotyped by sequencing. Associations of maternal HLA with the occurrence of the anti-HPA antibody and correlation of the severity of NAIT with the titre of the anti-HPA antibody will be further analysed. DISCUSSION: The study is expected to provide baseline data on NAIT in China. Besides, we hope to find out a population who expresses particular HLA molecules has significant higher risk of HPA alloimmunization in Chinese individuals. We also hope to find a Chinese-specific cut-off antibody titre for the prediction of the severity of NAIT and to provide a means to evaluate the necessity of antenatal treatment. TRIAL REGISTRATION: ClinicalTrials.gov: NCT02934906 (date registered: 13.10.2016).

Screening glioma stem cells in U251 cells based on the P1 promoter of the CD133 gene.

Cluster of differentiation (CD)133 is an important cell surface marker of glioma stem cells (GSCs). The transcription of the CD133 gene is controlled by five alternative promoters (P1, P2, P3, P4 and P5), which are expressed in a tissue-specific manner. In the present study, gene recombination technology was used to construct two types of gene expression vectors that contained the P1 promoter of the CD133 gene, which regulated either the neomycin-resistance gene or the herpes simplex virus thymidine kinase (HSV-TK) gene. Following the stable transfection of U251 glioblastoma cells with these two gene vectors, the cells expressing the P1 promoter that regulated the neomycin-resistance gene were named CD133 (+) cells, while the cells expressing the P1 promoter regulating the HSV-TK gene were called CD133 (-) cells. The expression of CD133 was detected by flow cytometry and reverse transcription-quantitative polymerase chain reaction. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was used to assess cell proliferation ability, while the cell cycle was analyzed by flow cytometry, and a clone formation test was performed to evaluate the invasive capability of the cells. The results demonstrated that, due to CD133 expression, the cell proliferation ability and the invasive capability of CD133 (+) cells were significantly higher than those of CD133 (-) cells. In conclusion, the present study successfully established a novel method of screening GSCs in U251 cells based on the P1 promoter of the CD133 gene.

(Z)-N-{(E)-10-[(2,6-Diisopropyl-phen-yl)-imino]-9,10-dihydro-phenanthren-9-yl-idene}-2,6-dimethyl-aniline.

The title compound, C(34)H(34)N(2), adopts a Z,E configuration with respect to the N=C-C=N backbone, with an N-C-C-N torsion angle of 41.1 (4)° The dihedral angle between the benzene rings in the 9,10-dihydro-phenanthrene moiety is 18.0 (1)°.

Effect of chitosan/type I collagen/gelatin composites in biocompatibility and nerve repair.

Chitosan, collagen I and gelatin were mixed in appropriate quantities to develop a new nerve repair material, with good arrangement and structure, as well as even aperture size. The composite material was sterilized by (60)Co irradiation for 24 hours prior to implantation in the right thigh of rats following sciatic nerve damage. Results showed that the material was nontoxic to the kidneys and the liver, and did not induce an inflammatory response in the muscles. The composite material enhanced the recovery of sciatic nerve damage in rats. These experimental findings indicate that the composite material offers good biocompatibility and has a positive effect on injured nerve rehabilitation.

[Simulation model of barley leaf area index].

To simulate leaf area index (LAI) accurately is the key for the prediction of crop growth and yield in a crop growth model. Based on the analysis of the dynamic changes in the LAI of high-yielding barley cultivars in Wuhan and Yangzhou, a simulation model of barley LAI was established, in which, the LAI was the function of expansion coefficient of LAI for cultivar genetic property, climatic factors such as daily air temperature difference, sunshine hours, and accumulation of photosynthetic available radiation after sowing (sigma PAR), and limitation indices of water and nutrients. It was indicated that the maximum LAI and optimal LAI at the stages of booting and heading were not the same conception, but differed significantly. The model was tested by the field experiments with different barley cultivars under different sowing dates and nitrogen application rates in Yangzhou, Nanjing, and Kunming. The results showed that this model gave the good predictions of LAI at different development stages, with the RMSE values ranged in 0.742 and 2.865, and averaged 1.348. The simulated and observed LAI values were significantly positively correlated, and the correlation coefficient from y = x regression analysis was between 0.511 and 0.954.

[Individual and interactive effects of aluminum, selenium, silicon and phosphorus on rice seedlings growth].

By using quadratic orthogonal rotation combinatorial design, and taking aluminum (Al), selenium (Se), silicon (Si), and phosphorus (P) as test factors, the regression models of test factors and the survival rate (SR), shoot mass per hundred plants (SMHP), root mass per one hundred plant seedlings (RMHP), and proline contents (PC) of rice seedlings were established, with the individual and interactive effects of the factors analyzed. The individual effects of test factors on the SR, SMHP, RMHP, and PC of rice seedlings decreased in the order of Al > P > Se > Si. Al showed negative effect, while Se, Si, and P showed positive effect. There were no significant interactive effects between the factors, except between Al and Si. Simulated optimization test showed that rice seedlings could grow better in an Al-containing solution when the Al was within the range of 0.587-0.913 mmol L(-1), Se was of 0.478-0.564 mg x L(-1), Si was of 0.613-1.069 mmol x L(-1), and P was of 2.252-2.657 mmol L(-1).

[Simulation model on barley yield formation].

By the method of yield component, a simulation model on barley yield formation was established, which could be available for different barley cultivars in different areas. The establishment process of the model was as the following. With the yield components of different cultivars under optimal conditions in Wuhan, Yangzhou, and Kunming as the bases, the regression equations of the relative values of ears per plant, kernels per ears, and thousand-grain weight with the accumulated photosynthetic effective radiation (SigmaPAR) were established, and the functional relations between the ears per plant, kernels per ears, and thousand-grain weight under actual conditions as well as their potential values under optimal conditions and the variables soil moisture regime and nutrient contents under actual conditions were formed. The model more comprehensively considered the internal and external factors of barley growth and development. The internal factors reflecting the genetic properties of cultivar included potential ears per plant, potential kernels per ear, potential thousand grain weight, and grain-filling duration; while the external factors included SigmaPAR and soil moisture regime and nutrient contents. Field experiments with different barley cultivars at different sowing dates were conducted in Wuhan, Kunming, and Yangzhou, and the data obtained were used to calibrate and validate the model. The results showed that the model better simulated the yield components and theoretical yield, with the average absolute values of relative error (RE) of ears per plant, kernels per ear, and thousand-grain weight being 1.96%, 1.88% and 1.67%, respectively. The correlation coefficient from y = x regression analysis on the simulated and observed values of the three yield components and theoretical yield was between 0.9464 and 0.9987, indicating that this model was more predictable and applicable.