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To establish and identify induced pluripotent stem cells (iPSCs) derived from patients with Aicardi-Goutières syndrome (AGS) with TREX1 gene 667G>A mutation, and obtain a specific induced pluripotent stem cell model for Aicardi-Goutières syndrome (AGS-iPSCs). A 3-year-old male child with Aicardi-Goutieres syndrome was admitted to Zhongshan People's Hospital in December 2020. After obtaining the informed consent of the patient's family members, 5 ml peripheral blood samples from the patient were collected, and mononuclear cells were isolated. Then,the peripheral blood mononuclear cells(PBMCs) were transduced with OCT3/4, SOX2, c-Myc and Klf4 by using Sendai virus, and PBMCs were reprogrammed into iPSCs. The pluripotency and differentiation ability of the cells were identified by cellular morphological analysis, real-time PCR, alkaline phosphatase staining (AP), immunofluorescence, teratoma formation experiments in mice. The results showed that the induced pluripotent stem cell line of Aicardi-Goutieres syndrome was successfully constructed and showed typical embryonic stem-like morphology after stable passage, RT-PCR showed mRNA expression of stem cell markers, AP staining was positive, OCT4, SOX2, NANOG, SSEA4, TRA-1-81 and TRA-1-60 pluripotency marker proteins were strongly expressed. In vivo teratoma formation experiments showed that iPSCs differentiate into the ectoderm (neural tube like tissue), mesoderm (vascular wall tissue) and endoderm (glandular tissue). Karyotype analysis also confirmed that iPSCs still maintained the original karyotype (46, XY). In conclusion, induced pluripotent stem cell line for Aicardi-Goutières syndrome was successfully established using Sendai virus, which provided an important model platform for studying the pathogenesis of the disease and for drug screening.
Subject(s)
Animals , Male , Mice , Child, Preschool , Cell Differentiation , Induced Pluripotent Stem Cells/pathology , Leukocytes, Mononuclear , Teratoma/pathologyABSTRACT
Objective:To evaluate the role of basolateral amygdala (BLA)-prelimbic cortex (PL) brain-derived neurotrophic factor (BDNF) in perioperative neurocognitive disorders (PND) and its relationship with tyrosine kinase B (TrkB) receptors in mice.Methods:Forty-eight clean-grade healthy C57BL/6J mice, aged 6 months, weighing 25-30 g, were divided into 4 groups ( n=12 each) using a random number table method: control group (group C), surgery group (group S), BDNF overexpression in BLA group (group B) and BDNF overexpression in BLA+ ANA-12 injection in PL group (group A). In group S, at 30 min after the end of training session of fear conditioning system, exploratory laparotomy was performed.In group B, recombinant adenovirus 0.3 μl was injected in BLA, fear conditioning test was performed 3 weeks later, and exploratory laparotomy was performed at 30 min after the end of the training session of fear conditioning system.In group A, recombinant adenovirus 0.3 μl was injected in BLA, catheterization was performed in PL, the fear conditioning test used performed 3 weeks later, TrkB receptor antagonist ANA-12 0.25 μg was given in PL starting from 30 min before training session, and exploratory laparotomy was performed at 30 min after training session.Test session was started at 24 h after the end of training session of fear conditioning system, and the percentage of time spent freezing was calculated in all groups.At 30 min after the end of the behavioral test, the expression of BDNF in brain areas, phosphorylated TrkB (p-TrkB) and phosphorylated extracellular signal-regulated kinase 1/2 (p-ERKl/2) was determined by Western blot, and the expression BDNF mRNA in BLA was detected using reverse transcription fluorescence quantitative polymerase chain reaction assay. Results:Compared with group C, the percentage of time spent freezing was significantly decreased, and expression of BNDF protein and its mRNA in BLA and BDNF, p-TrkB and p-ERK1/2 in PL was down-regulated in group S ( P<0.05). Compared with group S, the percentage of time spent freezing was significantly increased, and expression of BNDF protein and its mRNA in BLA and BDNF, p-TrkB and p-ERK1/2 in PL was up-regulated in group B ( P<0.05). Compared with group B, the percentage of time spent freezing was significantly decreased, and expression of p-TrkB and p-ERK1/2 in PL was down-regulated in group A ( P<0.05). Conclusion:The mechanism of PND is probably related to reduction of BDNF secretion from BLA to PL caused by down-regulation of BDNF expression in BLA, and decreasing of post-synaptic phosphorylation of TrkB receptors in mice.
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Alzheimer's disease is a common disease among the elderly.It seriously affects the quality of life of patients.There is still no effective treatment.The existence of blood-brain barrier further limits the efficacy of therapeutic drugs.As a highly safe nano-carrier,liposomes can not only carry drugs into the brain,but also have the advantages of reducing toxicity and side effects.Therefore,liposomes are promising in the treatment of Alzheimer's disease.This paper summarized the application of different types and functions of liposomes in various therapeutic drugs for Alzheimer's disease,aiming to provide an effective delivery strategy for the treatment of Alzheimer's disease.
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Objective To investigate the effect and mechanism of suberoylanilide hydroxamic acid (SAHA) on the fear extinction in mice with chronic social defeat stress (SD). Methods Fifty-six male C57BL/6J mice aged 7-8 weeks were randomly divided into control group,social defeat group,control-SAHA group and social defeat-SAHA group to investigate the effect of SAHA and social defeat group,social defeat-AAV BDNF group and social defeat-AAV blank group to investigate the effect of BDNF. Fear extinction in mice was evaluated by fear conditioning test (FC). The levels of BDNF and HDAC2 in mice hippocampus were detected by Western blot (WB). The expression of BDNF-overexpressing virus in hippocampus of mice was detected by immunofluorescence assay. Results (1) Compared with control group,fear extinction in the social defeat group was significantly decreased (P<0. 05). Compared with control group, the level of HDAC2(0. 50±0. 02) in the social defeat group was significantly increased (P<0. 001),while the level of BDNF(0. 16 ± 0. 03) was significantly decreased (P<0. 001) in the social defeat group. ( 2) After using SAHA,fear extinction of mice significantly improved (P<0. 05). Compared with control group,the level of HDAC2 (0. 26±0. 02) in the control-SAHA group was significantly decreased(P<0. 001),and the level of BDNF (0. 40±0. 03) was significantly increased (P<0. 001). Compared with social defeat group,the level of HDAC2 (0. 39±0. 03) in the social defeat-SAHA group was significantly decreased (P<0. 001),and the lev-el of BDNF (0. 28±0. 01) was significantly increased (P<0. 001). (3)After injection BDNF-overexpressing virus,fear extinction was significantly improved(P<0. 05). Conclusion SAHA can enhance fear extinction in mice with chronic social defeat stress and its mechanism may be related to the up-regulation of BDNF ex-pression in hippocampus by inhibiting HDAC2 in hippocampal.
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Objective To understand biofilm-forming ability of Acinetobacter baumannii(A.baumannii) in clinical isolates,and evaluate the effect of biofilm formation on antimicrobial resistance.Methods A total of 47 A.baumannii clinical isolates were collected,biofilm-forming ability was detected by crystal violet staining assay,the minimal inhibitory concentration(MIC) of 12 kinds of antimicrobial agents to biofilrn forming isolates were detected when the isolates were in planktonic or biofilm formatted condition;biofilm-forming ability of positive isolates at different culture condition (shaking or stable) and on different materials(glass tube or polypropylene plastic centrifugal tube) were detected.Results Thirteen isolates (27.66％) exhibited biofilm formation.In planktonic condition,antimicrobial resistance rates of these isolates were all ＜35％,no isolates shown resistance to polymyxin and tigecycline,in biofilm formatted condition,resistance rates to tigecycline was 15.38％,resistance rates to the other antimicrobial agents were all 100％.Of 13 tested isolates,≥10 isolates shown higher absorbance in the shaking condition than in the stable condition,≥10 tested isolates shown higher absorbance in polypropylene plastic centrifugal tube than in the glass tube.Conclusion The resistance of A.baumannii to antimicrobial agents is greatly changed after the formation of biofilm,which may results in the failure of antimicrobial therapy for infection.
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Objective To investigate the effect and mechanism of dexmedetomidine, mifepristone and dexmedetomidine plus mifepristone on the fear memory in rats with post?traumatic stress disorder ( PTSD) . Methods 40 male SD rats were randomly divided into five groups with 8 in each group:control group (group C),PTSD model group (group P),dexmedetomidine group (group D),mifepristone group ( group M) and dexmedetomidine plus mifepristone group ( group U) . Fear memory in rats was evaluated by fear conditioning test ( FC) . Anxiety?like behavior was assessed by the elevated plus?maze test ( EPM) . Ex?pressions of BDNF and its receptor TrkB in the hippocampus of rats after fear condition were detected using Western blot ( WB) and CORT level in the serum was detected using enzyme?linked immunosorbent assay (ELISA). Results Compared with group P,the freezing scores in the FC in group D((32.29±8.09) %), M((33.33±8.21) %),and U((9.38±3.31) %) were significantly decreased (P<0.05). The times and en?tries in the open arms of the EPM were significantly increased (P<0.05) . The expressions of BDNF in group D(0.65±0.04),M(0.71±0.04),U(0.79±0.07) and TrkB in group D(0.66±0.04),M(0.71±0.04),U (0.86±0.03) were obviously rescued in hippocampus of rats (P<0.05). The CORT level in serum in group D ((37.65±12.37)μg/L) and U((59.10±5.23)μg/L) was decreased (P<0.05). There was no difference be?tween group P and M. Conclusion These results suggest that dexmedetomidine, mifepristone and dexme?detomidine plus mifepristone can significantly enhance fear extinction and improve anxiety?like behaviors in rats with PTSD. The mechanism may be that dexmedetomidine and mifepristone could enhance the expres? sions of BDNF and TrkB in the hippocampus.
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To better understand the outcomes of small cell lung cancer (SCLC), we examined the clinical features and prognostic factors of SCLC in this study. A total of 148 patients who were diagnosed as having SCLC between January 2009 and December 2013 in Cancer Center of Union Hospital, Wuhan, China, were enrolled and their clinical features and prognostic factors were retrospectively analyzed. Log-rank test and Cox regression model were employed for analysis of prognostic factors. The 1- and 2-year overall survival (OS) rates were 59.7% and 25.7%, respectively, for limited disease (LD) patients whose median survival time (MST) was 16 months. The 1- and 2-year OS rates were 29.5% and 5.3%, respectively, for extensive disease (ED) patients whose MST was 10 months. The univariate analysis and multivariate analysis revealed that age, tumor stage, serum CEA and Ki-67 antigen were significantly correlated to the outcomes of SCLC, and they were significant prognostic factors for SCLC.