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With the rapid development of the cultivated meat industry, large-scale cell culture technology applied to cultivated meat production has received extensive attention. In this study, two microcarriers, Cytodex 1 and 3D TableTrix™, were used for suspension culture of myoblasts in spinner flasks to select a more suitable one for large-scale culture of myoblasts, and then the effects of different culture conditions on the large-scale culture of cells were explored. The microscopic results showed that Cytodex 1 was spheres with a smooth surface and no macroporous structure, and 3D TableTrix™ was irregular spheres with a porous surface and large pores. The results of cell culture in spinner flasks showed that the cells grew on 3D TableTrix™ more efficiently, and after 10 days of culture, the cell yield was 8.97 × 105 cells/mL. The optimal conditions for cell attachment were intermittent stirring at 40 r/min for 10 min with intervals of 50 min. The best cell culture efficiency was obtained by using cell inoculation density of 1 × 105 cells/mL, microcarrier density of 2 mg/mL, agitation speed of 40 r/min, and initial culture serum concentration of 20% as well as replacing the medium with a new one containing 10% serum (V/V) after 24 h of culture, and replacing 50% of the medium with a new one with every other day. By bead to bead transfer of the microcarriers, the cells were transferred to a 2 L spinner flask. Finally, 1.07 × 109 cells were harvested with a survival rate of more than 95%. Microcarriers loaded with cells could be cryopreserved and resuscitated, and no significant differences in proliferation status and morphology were observed compared with thawed cells. The optimized microcarrier based culture process will have promising applications in the industrial scale production of cultivated meat.

BackgroundThere are daytime variability in pulmonary function indexes such as peak expiratory flow (PEF) and forced expiratory volume in 1 second (FEV1) in asthma patients. Studies evaluating the effects of drug therapy on lung function and airway hyperresponsiveness (AHR) in asthma patients all required patients to perform spirometry and bronchial challenge test in the same time point of the days. However, whether there is a daily diurnal AHR variability is still not clear.ObjectiveTo explore the characteristics and diurnal variability of AHR in asthma patients.MethodsThe data of 202 patients with asthma who consulted in respiratory department of the First Affiliated Hospital of Guangzhou Medical University from January 2018 to September 2020 were included for statistical analysis. All patients completed the methacholine bronchial provocation tests, they were divided into the morning detection group (morning group) with 81 cases and the afternoon detection group (afternoon group) with 121 cases; according to the disease course, 98 cases were divided into the initial diagnosis group if the disease course was ≤6 months, and 104 cases were divided into the follow-up group if the disease course was >6 months. The initial diagnosis group and the follow-up group were divided into the initial diagnosis morning group, the initial diagnosis afternoon group, the follow-up morning group, and the follow-up afternoon group according to the detection time; according to the AHR, the patients were divided into very mild, mild, moderate and severe groups. The characteristics of AHR and the main pulmonary function indexes including FVC%pred, FEV1%pred, PEF%pred, MMEF%pred, MEF50%pred, MEF25%pred, PD20-FEV1, PD20-PEF, PD20-MMEF, PD20-MEF25%, PD20-MEF50% of these groups were analyzed and compared.ResultsThere were no significant differences of the math pulmonary function indexes and PD20 between morning and afternoon groups (P0.05) . In follow-up group, MMEF%pred and MEF50%pred were higher in afternoon than in morning (P0.05) .ConclusionThe longer the duration of asthma, more serious impairment of lung function found in asthma, while the AHR had no significant difference between morning and afternoon.

Objective: To compare the clinical efficacy of treating different diseases with the same acupuncture comprehensive therapy and intramuscular injection of ranibizumab in the treatment of macular edema, and to explore an effective treatment., Methods: A retrospective study was conducted, ①Acupuncture combined with EA at Xinming one (Extra), Sizhukong (TE 23), Tongziliao (GB 1), once every other day; ②acupoint injection, alternation with compound anisodine and mecobalamine injection at Qiuhou (EX-HN 7), Taiyang (EX-HN 5), once every other day; ③auricular acupressure at yan (LO 5 ), gan (CO 12 ), shen (CO 10 ) and other points; ④plum-blossom needle at Zhengguang 1 (Extra), Zhengguang 2 (Extra), once every other day were given in the acupuncture group (20 cases, 24 affected eyes). Intramuscular injection of 0.5 mg ranibizumab was given in the ranibizumab group (22 cases, 23 affected eyes). The macular foveal thickness, early treatment diabetic retinopathy study of (ETDRS) visual acuity chart, self-evaluation scores of visual function impairment ophthalmopathy patient's quality of life scale were observed before treatment, after 3, 6, 9 and 12 months of treatment, and the clinical efficacy was evaluated., Results: ①At all the observation time points of the treatment, the macular thickness was lower than that before treatment in the two groups (all P <0.05), and there was no significant difference between the acupuncture group and the ranibizumab group (all P >0.05). ②Visual acuity was higher than that before treatment at all the time points in the two groups (all P <0.05). After 3-months treatment, there was no statistical significance between the two groups ( P >0.05). After 6, 9, and 12 months treatment, the visual acuity in the acupuncture group was better than that in the ranibizumab group ( P <0.05, P <0.01). ③At all the time points, the quality of life scores were lower than those before treatment in the two groups (all P <0.05). There was no statistical significance in the ranibizumab group compared with those before treatment (all P >0.05). In 3, 6, 9 and 12 months of treatment, the quality of life scores in the acupuncture group was better than those in the ranibizumab group ( P <0.05, P <0.01). ④The total effective rate of the acupuncture group was 79.2% (19/24), which was better than 30.4% (7/23) in the ranibizumab group ( P <0.05). ⑤The improvement of visual acuity before and after treatment was negatively correlated with the course of disease ( P <0.05), ie, the longer the disease course of the eyes, the worse the visual acuity and the worse the effect., Conclusion: Acupuncture comprehensive treatment can effectively treat macular edema, significantly improve the patient's vision, improve the subjective experience and the quality of life, and the shorter the course of the disease the more significant effect. Acupuncture comprehensive treatment is better than intramuscular injection of ranibizumab.

Transcription factor is a key trans-acting factor to mediate stress response by regulating gene expression. Plants have developed a series of mechanisms to modulate development, stress response, signaling and disease resistance at transcription level. DNA binding with one finger (DOF), containing one C₂-C₂ zinc finger domain, is a special plant transcription factor. Specifically, the conserved domain at N-terminus of DOF has multiple functions, including interacting with DNA and protein, which could be involved in plant development and stress response. Although many DOF family genes are characterized in plant stress response, it is not clear if DOF genes have functions in cereal plants. In the present paper, the role of DOF family genes on cereal plants were discussed based on a comprehensive phylogenetic relationship analysis, expression profiles in different tissues and various environmental conditions. The results obtained here will provide an important reference for further understanding the mechanism of gramineous crops in stress resistance.

Objective: To investigate whether cigarette smoke extract (CSE) induces the senescence of skeletal muscle cells by oxidative stress-induced down-regulation of histone deacetylase 2 (HDAC2)., Methods: C2C12 myoblasts were induced to differentiate into skeletal muscle cells. H2O2 and N-acetyl cysteine (NAC) were used to investigate the effects of CSE on oxidative stress, cell senescence and the expression of HDAC2 in skeletal muscle cells, and detect the changes of the activities of malondialdehyde (MDA), superoxide dismutase (SOD), reactive oxygen species (ROS) and glutathione peroxidase (GSH-Px). Cell senescence was identified by senescence-associated β-galactosidase staining. The expressions of HDAC2 mRNA and protein were measured by real-time quantitative PCR and Western blotting, respectively., Results: Compared with the control group, MDA concentration and ROS activity significantly increased, the activities of SOD and GSH-Px significantly decreased, the expression of β-galactosidase was up-regulated, and the expressions of HDAC2 mRNA and protein were down-regulated in the CSE group and the H2O2 group. However, the changes in the CSE group were reversed after the pretreatment of NAC., Conclusion: CSE may lead to the senescence of murine skeletal muscle cells by oxidative stress-induced down-regulation of HDAC2.

Objective: Analysis of normal young people multiple stimuli ASSR (m-ASSR), single stimuli ASSR (s-ASSR), C-ABR, PTA threshold, exploratory investigation of relevance of hearing test combination thresholds (C-ABR combines with 0.5, 1.0 kHz s-ASSR) predict behavioral thresholds, using a within-subjects design., Method: Forty-three normal hearing adults (86 ears) were treated with pure tone audiometry (PTA), m-ASSR, 0.5, 1.0 kHz s-ASSR, C-ABR. Data were analyzed by SPSS 18.0 for bivariate correlation analysis., Result: There was significant correlation between thresholds of hearing test combination and PTA,and the correlation coefficient (r) were 0.803, 0.650, 0.649, 0.734 at 0.5, 1.0, 2.0 and 4.0 kHz; while the correlation coefficient (r) between thresholds of m-ASSR and PTA were 0.48, 0.64, 0.57, 0.64., Conclusion: There is significant correlation between thresholds of hearing test combination and PTA, hearing test combination can be more accurate, more reliable assessment of adults with normal hearing threshold level.

We describe a genetic transformation method of secondary somatic embryogenesis in alfalfa through cotyledon-stage somatic embryos of alfalfa infected by Agrobacterium strain GV3101. The Agrobacterium strain GV3101 contained binary vector pCAMBIA2301 that had gus gene as reporter and npt II gene as selectable marker. The infected primary embryos were induced through series of medium under 75 mg/L kanamycin selection. We obtained the transgenic alfalfa plants. Then, GUS expression in different tissue of transgenic alfalfa was tested by GUS histochemical analysis. Further, the stable integration and transformation efficiency were tested by polymerase chain reaction and Southern blotting hybridization. The result showed that GUS expression was different in different organs of transgenic alfalfa; the copy number of integrated npt II gene was from 1 to 4; the transformation efficiency via primary somatic embryogenesis was 65.82%.