Objective To construct a recombinant poxvirus vector vaccine, rVTTδTK-RBD, and to evaluate its safety and immunogenicity. Methods The receptor-binding domain (RBD) gene was synthesized with reference to the gene sequence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and was inserted into the polyclonal site of the self-constructed recombinant plasmid pSTKE, to construct the recombinant poxvirus shuttle vector pSTKE-RBD. This was then transfected into BHK-21 cells pre-infected with the vaccinia virus Tiantan strain (VTT). The recombinant poxvirus rVTTδTK-RBD was successfully obtained after several rounds of fluorescence phage screening. The effect of rVTTδTK-RBD on the body mass of BALB/c mice was detected after immunizing mice by intra-nasal vaccination. The levels of specific and neutralizing antibodies produced by rVTTδTK-RBD on BALB/c mice were analyzed after immunizing mice intramuscularly. The effect of rVTTδTK-RBD on T cell subsets in BALB/c mice was detected by flow cytometry. Results Through homologous recombination, enhanced green fluorescent protein (EGFP) screening marker, and multiple rounds of fluorescent phosphorescence phage screening, a recombinant poxvirus rVTTδTK-RBD, expressing RBD with deletions in the thymidine kinase (TK) gene, was successfully obtained, which was validated by PCR. The in vivo experiments on BALB/c mice showed that rVTTδTK-RBD was highly immunogenic against SARS-CoV-2 and significantly reduced toxicity to the body compared to the parental strain VTT. Conclusion The recombinant poxvirus vaccine rVTTδTK-RBD against SARS-CoV-2 is successfully constructed and obtained, with its safety and immunogenicity confirmed through various experiments.
Animals ; Mice ; SARS-CoV-2/genetics* ; COVID-19 ; Vaccines, Synthetic/genetics* ; Genes, Reporter ; Bacteriophages ; Mice, Inbred BALB C

Lipopolysaccharides (LPS) are major outer membrane components of Gram-negative bacteria. Unlike most Gram-negative bacteria, Acinetobacter baumannii can still survive and acquire polymyxin resistance after complete loss of LPS. Previous studies of LPS-deficient Acinetobacter baumannii mostly focused on LPS-deficient strains induced by polymyxin, whose background was complex and unstable. To investigate LPS loss-mediated polymyxin resistant-Acinetobacter baumannii, this study constructed a stable and clear background LPS-deficient strain by knocking out lpxC gene in Acinetobacter baumannii ATCC 19606 with CIRSPR/Cas9, and then studied the phenotypic changes of the lpxC-deficient strain including morphology, growth rate, antibiotic susceptibility, virulence, membrane permeability, and membrane potential. Animal experiments were approved by the Animal Care and Welfare Committee Institute of Medicinal Biotechnology, CAMS and PUMC (approval number: IMB-20240119D₉). The results indicated that the lpxC gene of Acinetobacter baumannii ATCC 19606 was successfully knocked out. After losing the lpxC, the strain underwent the morphological change from rod-shaped to spherical. Furthermore, it leads to reduced growth rate, enhanced membrane permeability, decreased membrane potential, lower virulence, and increased antibiotic susceptibility to β-lactams, quinolones, aminoglycosides, macrolides, glycopeptides. The lpxC deletion results in significant changes in membrane homeostasis and adaptability of Acinetobacter baumannii. Understanding the phenotypic changes of colistin-resistant Acinetobacter baumannii mediated by LPS loss is useful for exploring the resistance mechanism of Acinetobacter baumannii and developing new therapeutic strategies.

Disharmony between nutritive qi(ying)and defensive qi(wei)is the core pathogenesis of insomnia.The normal function of ying-wei in the generation,dispersion,divergence and aggregation is the precondition for the realization of the coordination between ying and wei.The disordered function of ying-wei in the generation,dispersion,divergence and aggregation will cause the disharmony between ying and wei,and then the insomnia occurs.For the treatment of insomnia caused by the disordered function of ying-wei in the generation,Guizhi Decoction associated prescriptions are used for strengthening middle energizer and nourishing ying and wei.For the treatment of insomnia caused by the disordered function of ying-wei in the dispersion,Mahuang Decoction associated prescriptions are used to relieve the exterior and eliminate the pathogen for insomnia patients with the manifestations of the attack of exopathogens,and Xiao Chaihu Decoction associated prescriptions are used to dredge the triple energizer for insomnia patients with the dysfunction of the triple energizer.For the treatment of insomnia caused by the disordered function of ying-wei in the divergence,Rhei Radix et Rhizoma associated bitter-cold prescriptions are used to purge the interior heat for insomnia patients with abundant interior heat syndrome,Gypsum Fibrosum associated pungent-cold prescriptions are used to release muscles and clear heat for insomnia patients with the interior heat complicated by exterior syndrome,Natrii Sulfas Exsiccatus associated salty-cold prescriptions are used to clear heat,moisten dryness and dissipate the masses for insomnia patients with interior heat complicated by dryness syndrome,sour-cold medicines are used to clear heat and remove retained water,supplement deficiency and relieve exterior for insomnia patients with interior heat complicated by water-retention syndrome,deficiency syndrome and exterior syndrome,and Ophiopogonis Radix associated prescriptions and Lillli Bulbus associated prescriptions are used to clear heat and nourish ying for insomnia patients with the consumption of ying and yin.For the treatment of insomnia caused by the disordered function of ying-wei in the aggregation,the compatibility of Poria and Cinnamomi Ramulus is used for warming yang and resolving fluid retention in patients with fluid retention,Taohong Siwu Decoction associated prescriptions are used to activate blood and remove stasis in patients with predominance of blood stasis syndrome,the compatibility of Poria and Paeoniae Radix Alba are used to treat retained water and blood stasis in patients with water-blood co-morbidity.Treating insomnia caused by disharmony between ying and wei from the perspective of the function of ying-wei in the generation,dispersion,divergence and aggregation is aimed at the core pathogenesis of insomnia,which makes the treatment easy to be carried out,and can provide reference for clinical differentiation and treatment of insomnia.

Objective To investigate the incidence rate,clinical characteristics,and prognosis of neonatal stroke in Shenzhen,China.Methods Led by Shenzhen Children's Hospital,the Shenzhen Neonatal Data Collaboration Network organized 21 institutions to collect 36 cases of neonatal stroke from January 2020 to December 2022.The incidence,clinical characteristics,treatment,and prognosis of neonatal stroke in Shenzhen were analyzed.Results The incidence rate of neonatal stroke in 21 hospitals from 2020 to 2022 was 1/15 137,1/6 060,and 1/7 704,respectively.Ischemic stroke accounted for 75％(27/36);boys accounted for 64％(23/36).Among the 36 neonates,31(86％)had disease onset within 3 days after birth,and 19(53％)had convulsion as the initial presentation.Cerebral MRI showed that 22 neonates(61％)had left cerebral infarction and 13(36％)had basal ganglia infarction.Magnetic resonance angiography was performed for 12 neonates,among whom 9(75％)had involvement of the middle cerebral artery.Electroencephalography was performed for 29 neonates,with sharp waves in 21 neonates(72％)and seizures in 10 neonates(34％).Symptomatic/supportive treatment varied across different hospitals.Neonatal Behavioral Neurological Assessment was performed for 12 neonates(33％,12/36),with a mean score of(32±4)points.The prognosis of 27 neonates was followed up to around 12 months of age,with 44％(12/27)of the neonates having a good prognosis.Conclusions Ischemic stroke is the main type of neonatal stroke,often with convulsions as the initial presentation,involvement of the middle cerebral artery,sharp waves on electroencephalography,and a relatively low neurodevelopment score.Symptomatic/supportive treatment is the main treatment method,and some neonates tend to have a poor prognosis.

Objective We aim to analyse semaglutide related adverse events in real-world,overall and by gender and age subgroups and compare the differences of different gender and age patients in adverse events,in order to supply references for security usage in the clinic.Methods OpenVigil 2.1 was used to search FDA Adverse Event Reporting System for semaglutide related adverse events from the establishment of the databases to April 2023.According to age and gender,patients were divided into 18-64 years old group and≥65 years old group,male group and female group.We selected ten adverse events which we interested(nausea,diarrhoea,vomiting,pancreatitis,cholecystitis,cholelithiasis,hypoglycaemia,diabetic retinopathy,acute kidney injury and thyroid cancer/medullary thyroid cancer)and analyzed overall and each group of semaglutide adverse events.Results A total of 5 330 cases and 15 558 adverse events were collected.2 935 patients aged 18-64 old group years with 8 553 adverse events;2 395 patients aged≥65 years old group with 7 005 adverse events.2 231 male group patients with 6 195 adverse events;3 059 female group patients with 9 277 adverse events.The sex of 40 patients was unknown.Nausea(1 089 cases/7.00％),vomiting(775 cases/4.98％)and diarrhoea(545 cases/3.50％)remained the most common adverse events.The constituent ratio of pancreatitis was significantly higher in patients aged 18-64 years old group than in patients aged 65 years old group(P＜0.05);the constituent ratio of diarrhoeaand cholelithiasis was significantly lower in patients aged 18-64 years old group than patients aged≥65 years old group(P＜0.05).The constituent ratio of diarrhoea,vomiting,pancreatitis,cholecystitis,cholelithiasis,diabetic retinopathy,and acute kidney injurywas significantly higher in male group patients than in female group(P＜0.05).Conclusion Nausea,diarrhoea and vomiting remined the most common adverse events of semaglutide.Male should be more concerned about gastrointestinal,pancreatitis,gallbladder events,retinopathy and acute kidney injury.Elderly patients should be more alert diarrhoea and cholelithiasis.

Objective To explore the role of Panax notoginseng saponins(PNS)in regulating the expression of sortilin and ATP-binding cassette transporter A1(ABCA1)in macrophages,and the effect of PNS on inhibiting formation of foam cells and the potential mechanism of PNS adjusting sortilin expression and cholesterol metabolism.Methods The macrophages were divided into five groups as follows:group A(only added with cell culture),group B(transfected with negative control lentivirus),group C(transfected with lentivirus-mediated sortilin overexpression),group D(transfected with lentivirus-mediated sortilin overexpression+60 μg·mL-1PNS),group E(transfected with lentivirus-mediated sortilin overexpression+10 μmol·L-1 mitogen-activated protein kinase kinase(MEK)inhibitor PD98059+60 μg·mL-1 PNS).The protein contents of sortilin,ABCA1,extracellular signal-regulated kinase(ERK)and phosphorylated-ERK(p-ERK)were evaluated with Western blot.All the cells in five groups were cultured with 50 μg·mL-1ox-LDL to form foam cells.The lipid in macrophages was investigated with red O assay.Results The relative expression levels of sortilin protein were 1.00±0.08,0.91±0.15,2.28±0.13,1.62±0.09 and 2.01±0.08;the relative expression levels of ABCA1 protein were 1.00±0.01,0.92±0.07,0.29±0.04,0.66±0.09 and 0.44±0.07;the ratios of p-ERK/ERK protein were 1.00±0.09,0.92±0.05,1.03±0.12,2.00±0.12 and 1.64±0.14;the contents of lipid in macrophages were(4.82±2.19)％,(6.70±0.88)％,(44.56±4.15)％,(27.66±3.25)％and(41.67±5.45)％.Except the ratios of p-ERK/ERK,the other parameters between group C and group A were statistically significant difference(all P＜0.01).Meanwhile,there were also statistically significant difference between group D and group C as well as group D and group E(P＜0.05,P＜0.01).Conclusion PNS inhibits the lipid accumulation in macrophages through upregulating ABCA1 and downregulating sortilin,and ERK signaling pathway may be as one of important mechanisms influencing the expression of sortilin and ABCA1 mediated by PNS.

[摘 要] 目的：探讨新补骨脂异黄酮（NBIF）对肝细胞癌（HCC）Huh-7细胞焦亡的影响及其分子机制。方法：体外培养Huh-7细胞，用CCK-8法检测不同浓度的NBIF处理48 h时对细胞存活率的影响，光学显微镜下观察NBIF处理后Huh-7细胞的形态变化，乳酸脱氢酶（LDH）释放实验检测细胞的LDH释放量，WB实验检测细胞中GSDME、caspase-3的蛋白水平变化。采用siRNA干扰Huh-7细胞中caspase-3、GSDME表达后，CCK-8法检测NBIF处理对细胞存活率的影响，WB实验检测GSDME蛋白表达水平，观察NBIF处理对细胞形态的影响，并检测细胞LDH释放量。结果：60 μmol/L以上的NBIF均能显著抑制Huh-7细胞的增殖（均P<0.01），光学显微镜下观察到NBIF处理后的细胞出现肿胀、吐泡现象，且LDH释放增加（P<0.01）；WB实验结果表明，NBIF能够激活caspase-3蛋白并切割GSDME蛋白，增加GSDME-N的表达（均P<0.01）。干扰caspase-3、GSDME表达后，NBIF对细胞的抑制作用减弱（均P<0.01），GSDME-N蛋白表达受到抑制（P<0.01），显微镜下细胞肿胀、吐泡现象几乎消失，LDH释放明显减少（P<0.05）。结论：NBIF能够通过caspase-3/GSDME途径诱导Huh-7细胞发生焦亡，从而抑制HCC细胞的增殖，为HCC的治疗提供一种新思路。

We wished to establish an expert consensus on late stage of critical care (CC) management. The panel comprised 13 experts in CC medicine. Each statement was assessed based on the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) principle. Then, the Delphi method was adopted by 17 experts to reassess the following 28 statements. (1) ESCAPE has evolved from a strategy of delirium management to a strategy of late stage of CC management. (2) The new version of ESCAPE is a strategy for optimizing treatment and comprehensive care of critically ill patients (CIPs) after the rescue period, including early mobilization, early rehabilitation, nutritional support, sleep management, mental assessment, cognitive-function training, emotional support, and optimizing sedation and analgesia. (3) Disease assessment to determine the starting point of early mobilization, early rehabilitation, and early enteral nutrition. (4) Early mobilization has synergistic effects upon the recovery of organ function. (5) Early functional exercise and rehabilitation are important means to promote CIP recovery, and gives them a sense of future prospects. (6) Timely start of enteral nutrition is conducive to early mobilization and early rehabilitation. (7) The spontaneous breathing test should be started as soon as possible, and a weaning plan should be selected step-by-step. (8) The waking process of CIPs should be realized in a planned and purposeful way. (9) Establishment of a sleep-wake rhythm is the key to sleep management in post-CC management. (10) The spontaneous awakening trial, spontaneous breathing trial, and sleep management should be carried out together. (11) The depth of sedation should be adjusted dynamically in the late stage of CC period. (12) Standardized sedation assessment is the premise of rational sedation. (13) Appropriate sedative drugs should be selected according to the objectives of sedation and drug characteristics. (14) A goal-directed minimization strategy for sedation should be implemented. (15) The principle of analgesia must be mastered first. (16) Subjective assessment is preferred for analgesia assessment. (17) Opioid-based analgesic strategies should be selected step-by-step according to the characteristics of different drugs. (18) There must be rational use of non-opioid analgesics and non-drug-based analgesic measures. (19) Pay attention to evaluation of the psychological status of CIPs. (20) Cognitive function in CIPs cannot be ignored. (21) Delirium management should be based on non-drug-based measures and rational use of drugs. (22) Reset treatment can be considered for severe delirium. (23) Psychological assessment should be conducted as early as possible to screen-out high-risk groups with post-traumatic stress disorder. (24) Emotional support, flexible visiting, and environment management are important components of humanistic management in the intensive care unit (ICU). (25) Emotional support from medical teams and families should be promoted through"ICU diaries"and other forms. (26) Environmental management should be carried out by enriching environmental content, limiting environmental interference, and optimizing the environmental atmosphere. (27) Reasonable promotion of flexible visitation should be done on the basis of prevention of nosocomial infection. (28) ESCAPE is an excellent project for late stage of CC management.
Humans ; Consensus ; Critical Care/methods* ; Intensive Care Units ; Pain/drug therapy* ; Analgesics/therapeutic use* ; Delirium/therapy* ; Critical Illness

From the perspective of market classification of Cnidii Fructus, this paper revealed the scientific connotation of evaluating the quality grade of Cnidii Fructus by its appearance traits. Thirty batches of Cnidii Fructus in different grades were selected as the research objects. The canonical correlation analysis and principal component analysis(PCA) were used to explore the measurement values of 15 appearance traits and intrinsic content indexes. The results of correlation analysis showed that except the aspect ratio, the 5 appearance trait indexes(length, width, 1 000-grain weight, broken grain weight proportion, and chroma) and 9 internal content indexes(the content of moisture, total ash, acid insoluble ash, osthole, imperatorin, 5-methoxy psoralen, isopimpinellin, xanthotoxin, and xanthotol) showed significant correlation to varying degrees. In addition, there was a significant positive correlation between the first typical variable U_1 composed of appearance traits and the first typical variable V_1 composed of internal content indexes(CR_1=0.963, P<0.01). The results of PCA showed that the classification results of appearance traits for 30 batches of Cnidii Fructus were consistent with the actual information of the samples. Under the same analysis conditions, 30 batches of Cnidii Fructus were reclassified by 9 groups of internal content indexes, and the analysis results were consistent. From the classification standard of the appearance traits of the system study, the statistical results of 6 appearance traits of Cnidii Fructus showed a correlation with grades. There was a good correlation between the appearance and the internal content of Cnidii Fructus, and the appearance quality effectively predicted the level of the internal content. There is a certain scientific basis for the quality classification of Cnidii Fructus by main appearance traits. Appearance classification can replace quality grading to realize the "quality evaluation through morphological identification" of Cnidii Fructus.
Fruit ; Phenotype ; Principal Component Analysis ; Social Group

In this study, the effect of brassinosteroid(BR) on the physiological and biochemical conditions of 2-year-old Panax notoginseng under the cadmium stress was investigated by the pot experiments. The results showed that cadmium treatment at 10 mg·kg~(-1) inhibited the root viability of P. notoginseng, significantly increased the content of H_2O_2 and MDA in the leaves and roots of P. noto-ginseng, caused oxidative damage of P. notoginseng, and reduced the activities of SOD and CAT. Cadmium stress reduced the chlorophyll content of P. notoginseng, increased leaf F_o, reduced F_m, F_v/F_m, and PIABS, and damaged the photosynthesis system of P. notoginseng. Cadmium treatment increased the soluble sugar content of P. notoginseng leaves and roots, inhibited the synthesis of soluble proteins, reduced the fresh weight and dry weight, and inhibited the growth of P. notoginseng. External spray application of 0.1 mg·L~(-1) BR reduced the H_2O_2 and MDA content in P. notoginseng leaves and roots under the cadmium stress, alleviated cadmium-induced oxidative damage to P. notoginseng, improved the antioxidant enzyme activity and root activity of P. notoginseng, increased the content of chlorophyll, reduced the F_o of P. notoginseng leaves, increased F_m, F_v/F_m, and PIABS, alleviated the cadmium-induced damage to the photosynthesis system, and improved the synthesis ability of soluble proteins. In summary, BR can enhance the anti-cadmium stress ability of P. notoginseng by regulating the antioxidant enzyme system and photosynthesis system of P. notoginseng under the cadmium stress. In the context of 0.1 mg·L~(-1) BR, P. notoginseng can better absorb and utilize light energy and synthesize more nutrients, which is more suitable for the growth and development of P. notoginseng.
Cadmium/metabolism* ; Antioxidants/pharmacology* ; Panax notoginseng ; Brassinosteroids/pharmacology* ; Chlorophyll/metabolism* ; Plant Roots/metabolism* ; Stress, Physiological