Neural tube defects (NTDs) are complex congenital malformations resulting from incomplete neurulation in embryo. Despite surgical repair of the defect, most of the patients who survive with NTDs have a multiple system handicap due to neuron deficiency of the defective spinal cord. In this study, we successfully devised a prenatal surgical approach and transplanted mesenchymal stem cells (MSCs) to foetal rat spinal column to treat retinoic acid induced NTDs in rat. Transplanted MSCs survived, grew and expressed markers of neurons, glia and myoblasts in the defective spinal cord. MSCs expressed and perhaps induced the surrounding spinal tissue to express neurotrophic factors. In addition, MSC reduced spinal tissue apoptosis in NTD. Our results suggested that prenatal MSC transplantation could treat spinal neuron deficiency in NTDs by the regeneration of neurons and reduced spinal neuron death in the defective spinal cord.
Background: The function of IκB kinase α (IKKα) in the brain is largely unknown. This study examined the effects of IKKα on autophagy after cerebral ischemia.Methods: Permanent distal middle cerebral artery occlusion (dMCAO) was conducted in C57/BL6 mice.Oxygen-glucose deprivation/reperfusion (OGD/R) was performed to mimic ischemia injury in neuro-2A (N2A) cells in vitro. Autophagy activation was assessed by detecting the ratio of microtubule-associated protein 1 light chain 3β (LC3B)-II/LC3B-I and Cyto-ID autophagic fluorescence. The infarct volume was verified by 2,3,5-triphenyltetrazolium chloride (TTC) staining and magnetic resonance imaging (MRI).Neurological functions were evaluated using the modified Garcia test. Cell death after dMCAO was confirmed with terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay. To determine the role of IKKα, small interfering RNA (siRNA) was transfected into N2A cells or injected intracerebroventricularly.Results: IKKα and LC3B II/I expression levels were increased both in OGD/R treated N2A cells and dMCAO mice. Under the same conditions, IKKβ expression was not altered. IKKα siRNA significantly decreased the infarct volume and the apparent diffusion coefficient (ADC) related to brain edema, and promoted the neurological outcomes after dMCAO. Furthermore, inhibition of IKKα attenuated ischemiainduced the conversion of LC3B Ⅰ to LC3B II both in vitro and in vivo. In addition, IKKα siRNA alleviated the formation of autophagic vacuoles and LC3 positive puncta after cerebral ischemia.Conclusions: These findings indicate that IKKα, but not IKKβ, plays a critical role in ischemia-induced autophagy. Inhibition of IKKα protects the brain from ischemia injury and this may have potential benefits in stroke therapy.
We investigated the influence of signal transducer and activator of transcription-3 (STAT3) on the spinal cord tissue grafts of rat fetuses with spina bifida aperta. In particular, we hoped to identify whether transfection of the STAT3 overexpression plasmid increases the survival of spinal cord transplantation in order to improve therapeutic efficacy. The fetal rat model of spina bifida aperta was established using retinoic acid and treated with a microsurgical injection of bone marrow mesenchymal stem cells (BMSCs). The animals were divided into either the blank control group, negative control group or the experimental group. The optical density (OD) value of BMSCs viability was determined using the Cell Counting Kit-8 (CCK-8). The expression of STAT3, phosphorylated STAT3 (pSTAT3), neural markers and apoptosis-related factors were evaluated using real-time PCR and Western blot. The OD value in the experimental group was highest at eight hours after transplantation using CCK-8. The expression of pSTAT3, glial fibrillary acidic protein, neuron-specific enolase, neurofilament and nestin in the experimental group was significantly higher compared to the blank control group and negative control group ( P <0.05). However, STAT3 expression in the experimental group was statistically significantly decreased ( P< 0.05). The relative expression of caspase-8 and bcl-2 in the experimental group were significantly lower compared to the blank control group and negative control group ( P <0.05). Transfection of the recombinant lentivirus-mediated STAT3 overexpression plasmid with BMSCs can help improve the efficiency of transforming into neural cells and provide new seed cells for the treatment of congenital spina bifida aperta.
Background Long noncoding RNAs (LncRNAs) are regulatory molecules that play important roles in various biological and pathological processes. Herein, we aimed to explore whether maternally expressed gene 8 (MEG8) promotes M1 macrophage polarization among Henoch-Schonlein purpura (HSP) rats, and to investigate the underlying mechanism. Methods Relative mRNA expression of MEG8, miR-181a-5p and suppressor of SH2 domain-containing tyrosine phosphatase 2 (SHP2) were examined using quantitative reverse transcription polymerase chain reaction. Furthermore, expression of SHP2 and the Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) pathway-related proteins was identified using western blot. Luciferase activity assay was conducted to evaluate whether miR-181a-5p could bind to MEG8 or SHP2. The macrophage phenotype was determined using flow cytometry and enzyme-linked immunosorbent assay. Results We observed macrophage polarization towards the M2 phenotype in the peripheral blood of HSP rats. Furthermore, MEG8 and SHP2 expression were down-regulated, while miR-181a-5p was up-regulated in monocyte-derived macrophages from the HSP rats compared to the control group. Furthermore, MEG8 functioned as a sponge for miR-181a-5p in order to facilitate SHP2 expression. Moreover, miR-181a-5p mimic and SHP2 knockdown significantly reversed the MEG8 overexpression-mediated suppression of JAK2/STAT3 signalling, and promotion of M1 polarization. Conclusions The lncRNA MEG8 sponged miR-181a-5p, which contributes to M1 macrophage polarization by regulating SHP2 expression in HSP rats. Key Messages LncRNA MEG8 downregulation and M2 polarization in Henoch Schonlein purpura rats. MEG8 upregulation enhances M1 polarization and suppresses JAK2/STAT3 pathway. MEG8 sponges miRNA-181a-5p to regulate SHP2 expression. MiRNA-181a-5p upregulation reverses lncRNA MEG8-mediated enhancement of M1 polarization and inhibition of JAK2/STAT3 pathway. SHP2 downregulation reverses lncRNA MEG8-mediated enhancement of M1 polarization and inhibition of JAK2/STAT3 pathway.
Background: Long noncoding RNAs (lncRNAs) are important regulatory molecules in various biological and pathological processes. We herein aimed to explore whether maternally expressed gene 8 (MEG8) promotes M1 macrophage polarization in Henoch-Schonlein purpura (HSP) rats and to investigate the underlying mechanism.Methods: Relative MEG8 and miR-181a-5p expression and suppressor of SH2 domain-containing tyrosine phosphatase 2 (SHP2) RNA level were examined using quantitative reverse transcription polymerase chain reaction. Expression of SHP2 and the Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) pathway-related proteins was detected using western blot. Luciferase activity assay was performed to test whether miR-181a-5p could bind to MEG8 or SHP2. The macrophage phenotype was determined using flow cytometry analysis and enzyme-linked immunosorbent assay.Results: The macrophage polarization toward the M2 phenotype was observed inperipheral blood from HSP rats. Furthermore, MEG8 and SHP2 expression were down-regulated but miR-181a-5p was up-regulated in monocyte-derived macrophages from HSP rats compared with the control group. Furthermore, MEG8 acted as a sponge for miR-181a-5p to facilitate SHP2 expression. Moreover, miR-181a-5p mimic and SHP2 knockdown significantly reversed the MEG8 overexpression-mediated suppression of the JAK2/STAT3 signaling and promotion of M1 polarization.Conclusion: IncRNA MEG8 sponging miR-181a-5p contributes to M1 macrophage polarization by regulating SHP2 expression in Henoch Schonlein purpura rats.
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