Objective To investigate the effect of Fasudil on H2O2-induced apoptosis and synaptic plasticity in human neuroblastoma SY5Y cells and its mechanism. Methods The cells were divided into three groups: PBS control group, H2O2 model group (250 μmol/L H2O2 treatment) and Fasudil intervention group (250 μmol/L H2O2 combined with 15 μg/mL Fasudil treatment). MTT assay was applied to detect cell activity and TUNEL was performed to detect cell apoptosis respectively. Immunofluorescence cytochemical staining was used to determine the expression of neurite outgrowth inhibitor A (NogoA), Nogo receptor (NgR) and synaptophysin (Syn). Western blotting was then conducted to detect the expression of NogoA, NgR, p75 neurotrophin receptor (p75NTR), leucine-rich repeat Ig domain-containing Nogo-interacting protein 1 (LINGO-1), Syn and postsynaptic density protein-95 (PSD-95). Results Compared with the PBS group, the H2O2 group showed decreased cell viability and increased apoptosis rate while Fasudil treatment significantly increased the cell viability and reduced the apoptosis rate. Compared with the H2O2 model group, Fasudil intervention increased expressions of Syn and PSD-95. Compared with the PBS group, the expression of NogoA and its receptor complex NgR/p75NTR/LINGO-1 grew significantly in the H2O2 group, suggesting Fasudil treatment could inhibit the expression of NogoA and its receptor complex NgR/p75NTR/LINGO-1. Conclusion Fasudil may inhibit the activation of the NogoA/NgR signaling pathway, therefore reducing the apoptosis induced by H2O2 in SH-SY5Y cells and enhancing the plasticity of the synapses.
SignificanceAlzheimer's disease (AD) is a predominant form of dementia that can lead to a decline in the quality of life and mortality. The understanding of the pathological changes requires monitoring of multiple cerebral biomarkers simultaneously with high resolution. Photoacoustic microscopy resolves single capillaries, allowing investigations into the most affected types of vessels. Combined with confocal fluorescence microscopy, the relationship between plaque deposition and small vessel pathology could be better understood.AimWe aim to introduce a dual-modality imaging system combining photoacoustic microscopy (PAM) and confocal fluorescence microscopy (CFM) to provide a comprehensive view of both cerebral cortical vessels and amyloid-β (Aβ) plaque in AD mouse model in vivo and to identify the pathological changes of these two biomarkers.ApproachWe developed a dual-modality imaging system to image both cerebral vessel structure and Aβ plaque on groups of mice with different ages and phenotypes. Vessel imaging is enabled by PAM, whereas Aβ plaque is imaged by CFM with the aid of fluorescent dye.ResultsThe small vessel density in the AD group was significantly lower than in the control group, whereas the Aβ plaque density in the AD group was not only higher but also increased with age.ConclusionsThis dual-modality system provides a powerful platform for biomarker monitoring of AD expressing multi-dimensional pathological changes.
Abstract Background Early detection and intervention using hospital database were followed to provide accurate data support and effective intervention programs for elderly population with cognitive dysfunction. Materials and Methods A total of 3352 rural-urban residents aged ≥ 60 years in urban area, Guangling and Tianzhen counties were selected by random sampling method from the hospital database for demographic information, MoCA scale assessment and prevalence were assessed. We analyzed the influencing factors related with MCI prevalence by logistic regression test. Results The total prevalence of mild cognitive impairment (MCI) in urban and rural residents in Datong was 13.98%. There were no significant difference in the prevalence of MCI between urban and rural residents in gender and among same gender. The prevalence of MCI in rural were significantly higher as compared with urban residents in the age groups of 70–79 and 80–89, however 60–69 and above 90 years age group did not showed significant change. Elderly with the same residence and different educational levels, the prevalence of MCI gradually decreased with the increase of educational level. Also, elderly with chronic diseases in urban and rural residents was showed higher MCI prevalence as compared to elderly without chronic diseases. Conclusion MCI disparities among 70–90 age groups with different genders, ages, living conditions, educational levels and diseases condition. Old age, female, low education, living in rural areas and chronic disease condition were major risk factors for MCI. Seventy years old could be the vulnerable age for MCI prevalence in urban and rural residents. Our study suggests that attention should be paid to the rural elderly aged 60–69 with chronic diseases. Early screening strategy and targeted intervention of MCI could be best possible approarch to mitigate the disparity.
Abstract Alzheimer’s disease (AD) is characterized by cognitive impairment caused by the accumulation of beta-amyloid (Aβ) plaques and trans-synaptic spread of tau pathology. Exosome has emerged as key mediators for neuronal development, maintenance, and cellular communication. However, the molecular mechanism of exosomal miRNAs related to AD remains unknown. In the present study, APPswe/PSEN1dE9 transgenic (APP/PS1) mice (AD) were treated with vehicle (ADNS) and fasudil (ADF), whereas C57BL/6 (control) mice were treated with vehicle (WT). Cognitive function was assessed by Y-maze test and AD pathology was confirmed by immunostaining of Aβ plaque and phosphorylated tau. Exosomal RNAs from each mouse of the group were extracted, sequenced and analyzed. Our results showed amelioration of cognitive function, decreased Aβ plaque load, and phosphorylated tau protein after fasudil treatment. Exosomal miRNA analysis showed 3 miRNAs (mmu-let-7i-5p, mmu-miR-19a-3p, mmu-miR-451a) in the intersection of ADNS vs ADF and WT vs ADNS. GO annotation and KEGG pathway showed the target gene of miR-19a-3p are Pten and Tnf; mmu-miR-451a are Nsmaf, Gnai3 and Akt3. In conclusions, fasudil treatment improved cognitive function by regulating exosomal MicroRNAs (mmu-miR-451a and mmu-miR-19a-3p). These MicroRNAs could be potential biomarker of AD and therapeutic target for novel treatment for AD.
Ulcerative colitis (UC) and Crohn’s disease (CD) are chronic inflammatory conditions of the gastrointestinal (GI) tract that have been linked to intestinal microbial dysbiosis. While adherent-invasive Escherichia coli are associated with CD, recent studies have identified E. coli of phylogroup B2 as being frequently isolated from UC patients. B2 E. coli isolated from UC patients (such as p19A) harbour extra-intestinal pathogenic E. coli virulence factors including alpha hemolysin genes (hlyI, II). Correspondingly, the p19A strain causes cell death and barrier dysfunction in human epithelial colorectal cell lines, however its role in intestinal immunopathology is unclear because of the lack of a suitable animal model. The current study explores the potential to establish a mouse model of GI infection by the UC-associated E. coli strain p19A, as well as defines the mechanisms by which it promotes colitis. C57BL/6 and Sigirr (Single Ig IL-1R-related Receptor) deficient mice were orally administered with 5 mg vancomycin 6 hours prior to the oral infection with 108 colony forming units (CFU) of P19A. Sigirr-/- mice were examined because they show increased susceptibility to infection by enteric bacterial pathogens. Intestinal tissues as well as feces were homogenized and plated to enumerate CFU. An in vivo imaging system visualized the colonization of p19A in the intact intestines. To test the impact of pre-existing p19A colonization on experimental colitis, mice were colonized with DH10B (non-pathogenic E. coli control), p19A wild-type strain or p19A-ΔhlyI, II (a mutant strain lacking hemolysin genes). The next day, mice were given 3% (wt/vol) dextran sulfate sodium (DSS) in their drinking water for 4 days. Mice were monitored daily and euthanized to collect samples for further analysis. Vancomycin pretreatment led to persistent p19A colonization of the intestinal lumen of C57BL/6 mice, based on luciferase based imaging and fecal shedding data. Furthermore, p19A colonized the intestinal mucosal surface of Sigirr-/- mice. While p19A infection caused only minimal pathology on its own, it dramatically worsened the course of DSS colitis, in concert with deep penetration of the damaged colonic mucosa. Notably, a p19A strain deficient in hemolysin genes was severely attenuated in its ability to promote DSS colitis in Sigirr-/- mice. Our findings provide evidence that a UC E. coli pathobiont can readily and persistently colonize the intestines of susceptible hosts, and significantly worsen the course of colitis. This model thus facilitates research into the role played by UC associated E. coli pathobionts in the pathogenesis of IBD. CCC, CIHRNSERC, MITACs, and CHILD Foundation
To observe the effect of acupuncture on microglia polarization and inflammatory reaction in rats with cerebral ischemia-reperfusion injury (CIRI), so as to explore its mechanisms underlying improvement of CIRI.Thirty male SD rats were randomly divided into sham operation, model, and acupuncture groups, with 10 rats in each group. The CIRI model was established by occlusion of the middle cerebral artery (MCAO) for 1 h, followed by reperfusion. After modeling, rats in the acupuncture group received manual acupuncture stimulation of "Dazhui" (GV14), "Baihui"(GV20), "Shuigou" (GV26), bilateral "Zusanli" (ST36) and "Fengchi" (GB20) by twirling the needles rapidly for 10 s/acupoint every 10 min, with the needles retained for 20 min. The treatment was conducted once daily for successive 7 days. The neurological function was evaluated according to Longa's method. The state of CIRI was observed after Nissl staining, and the expression levels of Iba-1, iNOS, Arg1, BDNF, GDNF and NeuN in the ischemic cortex tissue were detected by immunofluorescence staining. The contents of TNF-α, IL-6 and IL-10 in the ischemic tissue were assayed by ELISA. The protein expression levels of BDNF, GDNF, TLR4, MyD88 and NF-κB in the ischemic tissues were detected by Western blot.The neurological deficit score on the 24 h and 7th day was considerably higher in the model group than in the sham operation group (P<0.01), and evidently lower on the 7th day in the acupuncture group than in the model group (P<0.01). The number of NeuN positive cells，the area of immunofluorescence dual labelling of Arg1, BDNF and GDNF positive staining, IL-10 content, BDNF and GDNF protein expressions were significantly decreased (P<0.01), and the immunofluorescence dual labelling area of Iba-1 and iNOS, TNF-α and IL-6 contents, the pretein expression levels of TLR4, MyD88 and NF-κB considerably increased (P<0.01) in the model group relevant to the sham operation group. In contrast to the model group, the acupuncture group had a significant increase in the number of NeuN positive cells, the immunofluorescence dual labelling area of Arg1, BDNF and GDNF positive staining, IL-10 content, and BDNF and GDNF protein expressions (P<0.05, P<0.01), and an evident decrease in Iba-1 and iNOS positive staining, contents of TNF-α and IL-6, and the protein expression levels of TLR4, MyD88 and NF-κB (P<0.01, P<0.05). Nissl staining showed a marked reduction in the number of neurons, the nucleus pyknosis and nissl bodies and loose arrangement of the neuronal cells in the model group, which was relatively milder in the acupuncture group.Acupuncture intervention can improve neurological function in CIRI rats, which may be related to its effects in regulating the polarization of microglia, reducing inflammatory reaction and increasing the secretion of neurotrophic factors in the brain, inhibiting TLR4/MyD88/NF-κB signaling pathway.目的：观察针刺对脑缺血再灌注(I/R)损伤大鼠小胶质细胞极化和炎性反应的影响，探讨其潜在的作用机制。方法：雄性SD大鼠随机分为假手术组、模型组、针刺组，每组10只。采用大脑中动脉栓塞法制备大鼠脑I/R损伤模型。针刺组大鼠在造模后于“大椎”“百会”“水沟”及双侧“足三里”“风池”进行针刺，20 min/次，1次/d，连续7 d。采用神经功能评分法评估各组大鼠神经功能；尼氏染色法观察脑梗死侧皮质区神经元损伤情况；免疫荧光染色法检测缺血侧皮质区神经元特异性核蛋白(NeuN)、离子钙结合蛋白1(Iba-1)、诱导型一氧化氮合酶(iNOS)、精氨酸酶1(Arg1)、脑源性神经营养因子(BDNF)和胶质细胞源性神经营养因子(GDNF)的表达；ELISA法检测缺血脑组织中肿瘤坏死因子-α（TNF-α）、白细胞介素（IL）-6和IL-10的含量；Western blot法检测缺血脑组织中BDNF、GDNF、Toll样受体4(TLR4)、髓样分化因子88(MyD88)和核因子κB(NF-κB)的表达水平。结果：与假手术组比较，模型组大鼠神经功能评分升高(P<0.01)，脑缺血侧皮质区神经元数量及尼氏体减少，NeuN阳性细胞数、Arg1阳性表达减少(P<0.01)，Iba-1、iNOS阳性表达增加(P<0.01)，BDNF、GDNF阳性表达和蛋白表达水平及IL-10含量降低(P<0.01)，TNF-α、IL-6含量升高(P<0.01)，TLR4、MyD88和NF-κB蛋白表达水平升高(P<0.01)。与模型组比较，针刺组大鼠神经功能评分降低(P<0.01)，缺血侧皮质区神经元数量及尼氏体增多，NeuN阳性细胞数、Arg1阳性表达增多(P<0.05)，Iba-1、iNOS阳性表达减少(P<0.01)，BDNF、GDNF阳性表达和蛋白表达水平及IL-10含量升高(P<0.01，P<0.05)，TNF-α、IL-6含量降低(P<0.01)，TLR4、MyD88和NF-κB蛋白表达水平降低(P<0.05，P<0.01)。结论：针刺干预可以通过调节脑I/R后小胶质细胞极化、减轻炎性反应和增加神经营养因子的分泌，对I/R后脑组织起神经保护作用，其机制可能与抑制TLR4/MyD88/NF-κB信号通路有关。.
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