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Objective·To investigate the effect of C9ORF72 amyotrophic lateral sclerosis/frontotemporal dementia (ALS/FTD)-related poly-glycine-arginine (poly-GR) on mitochondrial morphology and function, and analyze the rescue effect of metformin on mitochondrial damage induced by poly-GR and its underlying mechanism.Methods·SK-N-SH cells stably overexpressing 50 repeated glycine-arginine sequences [(GR)50] or green fluorescent protein (GFP) were constructed by lentivirus infection, which were respectively named as (GR)50-SK cell line and GFP CTRL-SK cell line. (GR)50 expression in (GR)50-SK cells was verified by Western blotting. GFP expression in GFP GTRL-SK cells was observed by fluorescence microscope. Propidium iodide (PI) staining was used to detect the apoptosis levels of (GR)50-SK and GFP CTRL-SK cells. Immunofluorescence (IF) staining was performed to determine the subcellular location of (GR)50. Reactive oxygen species (ROS) level of mitochondria was evaluated by staining cells with MitoSOX Red followed by observing the intensity of red fluorescence under fluorescence microscope. The mitochondrial morphology of (GR)50-SK and GFP CTRL-SK cells was observed by transmission electron microscopy. Western blotting was used to detect protein kinase B (PKB, also known as AKT) and its phosphorylation levels in (GR)50-SK and GFP CTRL-SK cells. SC79 was used to activate AKT in (GR)50-SK cells, and MitoSOX Red staining and PI staining were used to analyze mitochondrial ROS and apoptosis levels after phosphorylated AKT increased. Metformin was used to treat (GR)50-SK and GFP CTRL-SK cells, respectively, and the apoptosis levels, mitochondrial ROS levels, mitochondrial morphology, AKT and its phosphorylation levels, and ATP concentrations of the two cells were detected by the above methods and ATP detection kit, respectively.Results·Western blotting showed that the construction of (GR)50-SK cells was successful, and fluorescence microscopy showed that the construction of GFP CTRL-SK cells was also successful. PI staining results showed that the apoptosis level of (GR)50-SK cells was higher than that of the GFP CTRL-SK cells (P=0.016). IF staining results showed that there was partial co-localization of (GR)50 in the mitochondria of (GR)50-SK cells. Compared with GFP CTRL-SK cells, the mitochondrial morphology and structure of (GR)50-SK cells were significant abnormalities, with a significantly increased ROS levels. The AKT levels in (GR)50-SK cells were similar to those in the GFP CTRL-SK cells, but there was a significant decrease in phosphorylated AKT levels. After (GR)50-SK cells were treated with SC79, the AKT phosphorylation level was significantly upregulated, and ROS level and apoptosis level were significantly downregulated. Metformin could significantly up-regulate the phosphorylated AKT levels in (GR)50-SK cells, but had no effect on AKT levels; it could reshape the morphology and structure of some mitochondria, reduce ROS levels, increase ATP production (P=0.000), and down-regulate the level of cell apoptosis (P=0.000).Conclusion·(GR)50 can cause mitochondrial morphology and function abnormalities by down-regulating AKT phosphorylation, and promote cell apoptosis. Metformin can effectively reduce the occurrence of the above pathological events induced by (GR)50.

Integrated stress response (ISR) is a cellular adaptive response induced by stress, which is strictly regulated by multiple phosphokinases, phosphatases and other proteins to maintain protein homeostasis. Studies have shown that ISR is abnormally activated in Alzheimer's disease, and targeted regulation of different proteins in ISR pathway inhibits the abnormal activation of ISR, leading to restoration of protein homeostasis and alleviation of the neuropathological changes and memory impairment in Alzheimer's disease models. These lines of evidence suggest that ISR has the potential to be a therapeutic target in Alzheimer's disease treatment. This paper reviews the abnormal activation and regulation mechanism of ISR in Alzheimer's disease and discusses the application of ISR as therapeutic targets to Alzheimer's disease models.

Hexanucleotide GGGGCC repeat expansion in C9ORF72 is the most prevalent genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). One pathogenic mechanism is the aberrant accumulation of dipeptide repeat (DPR) proteins produced by the unconventional translation of expanded RNA repeats. Here, we performed genome-wide CRISPR-Cas9 screens for modifiers of DPR protein production in human cells. We found that DDX3X, an RNA helicase, suppresses the repeat-associated non-AUG translation of GGGGCC repeats. DDX3X directly binds to (GGGGCC)n RNAs but not antisense (CCCCGG)n RNAs. Its helicase activity is essential for the translation repression. Reduction of DDX3X increases DPR levels in C9ORF72-ALS/FTD patient cells and enhances (GGGGCC)n-mediated toxicity in Drosophila. Elevating DDX3X expression is sufficient to decrease DPR levels, rescue nucleocytoplasmic transport abnormalities, and improve survival of patient iPSC-differentiated neurons. This work identifies genetic modifiers of DPR protein production and provides potential therapeutic targets for C9ORF72-ALS/FTD.

Alzheimer's disease is a common neurodegenerative disease characterized by progressive memory loss and overall cognitive decline. Two typical pathological features are senile plaques and neurofibrillary tangles detected in the brain, which consist of large amounts of precipitated amyloid peptide and hyperphosphorylated Tau protein, respectively. In addition, there are other pathological features such as neuroinflammatory response and extensive loss of neurons. In the past decades, a lot of research has been done on Alzheimer's disease, but its etiology and pathogenesis are still unclear, and there is no ideal treatment or radical cure. This review introduced the characteristics of various transgenic animal models and their current application in preclinical studies, in order to provide theoretical basis for future experimental studies.

Objective To explore the effect of preoperative high-resolution anorectal manometry-assisted pelvic floor muscle exercises on anal function recovery in children with tethered cord syndrome. Methods Totally 68 cases of patients with tethered cord syndrome were divided into a study group (n=38) and a control group(n=38). The high-resolution anorectal manometry was performed preoperatively for evaluation of functions of the anal canal. The control group received routine nursing and health education, and the study group received an intervention of exercise for strengthening levator ani muscles. The results of anorectal pressure measurement were analyzed during postoperative follow-up, and the recovery of anal function was compared between two groups. Results There was no significant difference in anal canal resting pressure and maximum anal squeeze pressure between two groups before surgery(P＞0. 05). After a three-month follow-up, better outcomes of anal canal resting pressure, rectal sensitivity test and maximum anal squeeze pressure were measured in the study group compared with that in the control group (P＜0. 05). Conclusion Based on the evaluation results of preoperative anorectal pressure measurement, targeted pelvic floor muscle exercises can help children with anal function recovery. (目的 探讨高分辨率肛门直肠测压技术指导下的提肛训练在儿童脊髓栓系综合征患儿术后肛管功能恢复中的应用效果。方法 68例脊髓栓系综合症行手术治疗的患儿分为试验组38例和对照组30例。采用高分辨率多通道胃肠功能检测系统对脊髓栓系患儿术前的肛管功能进行评估。对照组接受常规护理和健康宣教, 试验组由护士根据测压结果给予盆底肌肉功能锻炼为主的提肛训练。术后随访患儿肛门直肠测压结果并分析, 比较两组患儿术后肛管功能的恢复情况。结果 术前两组肛管静息压、肛管最大收缩压比较, 差异无统计学意义(P＞0. 05)。术后3个月后门诊随访, 试验组患儿肛管静息压力明显高于对照组, 直肠敏感性、肛管最大收缩压监测结果试验组要明显优于对照组, 差异有统计学意义(P＜0. 05)。结论 结合患儿术前肛门直肠测压的评估结果, 有针对性地开展术后提肛训练, 有助于患儿肛门功能的恢复。)

Abstract Background Fetal macrosomia is associated with an increased risk of several maternal and newborn complications. Antenatal predication of fetal macrosomia remains challenging. We aimed to develop a nomogram model for the prediction of macrosomia using real-world clinical data to improve the sensitivity and specificity of macrosomia prediction. Methods In the present study, we performed a retrospective, observational study based on 13,403 medical records of pregnant women who delivered singleton infants at a tertiary hospital in Shanghai from 1 January 2018 through 31 December 2019. We split the original dataset into a training set (n = 9382) and a validation set (n = 4021) at a 7:3 ratio to generate and validate our model. The candidate variables, including maternal characteristics, laboratory tests, and sonographic parameters were compared between the two groups. A univariate and multivariate logistic regression was carried out to explore the independent risk factors for macrosomia in pregnant women. Thus, the regression model was adopted to establish a nomogram to predict the risk of macrosomia. Nomogram performance was determined by discrimination and calibration metrics. All the statistical analysis was analyzed using R software. Results We compared the differences between the macrosomic and non-macrosomic groups within the training set and found 16 independent risk factors for macrosomia (P