Objective : To investigate the effect of paederosidic acid methyl ester (PAME) on postoperative learning and memory impairment in mice and its mechanism. Methods : C57BL/6J male mice were randomly divided into Sham group , operation group , operation + PAME group ( PAME group) , operation + NADPH oxidase 4(Nox4) adeno⁃associated virus overexpression group (Nox4 overexpression group) , operation + Nox4 adeno⁃associated virus no⁃laden group ( AAV no⁃load group) , and operation + PAME + Nox4 overexpression group ( PN group) . Exploratory laparotomy was performed. PAME(20 mg/kg) was administered by continuous gavage for 7 days after operation , and adeno⁃associated virus was injected into the hippocampus 28 days before operation. Morris water maze test and conditioned fear test were used to detect the learning and memory ability of mice. The expression of Nox4 protein was observed by immunofluorescence. The protein expressions of Nox4 , long chain acyl CoA synthetase 4 (ACSL4) and glutathione peroxidase 4 (GPX4) were detected by Western blot. Reactive oxygen species (ROS) and iron content were determined by spectrophotometry. Results : Compared with the Sham group , the learning and memory ability of the operation group , the Nox4 overexpression group and the AAV no⁃load group decreased , the protein expression of Nox4 and ACSL4 increased , the protein expression of GPX4 decreased , and the ROS and iron content increased. After PAME treatment , the postoperative learning and memory ability of mice was improved , and Nox4 and ferroptosis in hippocampal neurons were alleviated. Conclusion Conclusion PAME treatment can improve the learning and memory ability of postoperative mice , which may be related to the inhibition of hippocampal Nox4⁃mediated ferroptosis.

Parvalbumin interneurons belong to the major types of GABAergic interneurons. Although the distribution and pathological alterations of parvalbumin interneuron somata have been widely studied, the distribution and vulnerability of the neurites and fibers extending from parvalbumin interneurons have not been detailly interrogated. Through the Cre recombinase-reporter system, we visualized parvalbumin-positive fibers and thoroughly investigated their spatial distribution in the mouse brain. We found that parvalbumin fibers are widely distributed in the brain with specific morphological characteristics in different regions, among which the cortex and thalamus exhibited the most intense parvalbumin signals. In regions such as the striatum and optic tract, even long-range thick parvalbumin projections were detected. Furthermore, in mouse models of temporal lobe epilepsy and Parkinson's disease, parvalbumin fibers suffered both massive and subtle morphological alterations. Our study provides an overview of parvalbumin fibers in the brain and emphasizes the potential pathological implications of parvalbumin fiber alterations.
Mice ; Animals ; Epilepsy, Temporal Lobe/pathology* ; Parvalbumins/metabolism* ; Parkinson Disease/pathology* ; Neurons/metabolism* ; Interneurons/physiology* ; Disease Models, Animal ; Brain/pathology*

Objective: To investigate the effects of adipose-derived stem cell released exosomes (ADSC-Exos) on wound healing in diabetic mice. Methods: The ADSCs were isolated from the adipose tissue donated by the patients and cultured by enzymatic digestion. The supernatant of the 3rd generation ADSCs was used to extract Exos (ADSC-Exos). The morphology of ADSC-Exos was observed by transmission electron microscopy. The membrane-labeled proteins (Alix and CD63) were detected by Western blot, and the particle size distribution was detected by nanoparticle tracking analyzer. The fibroblasts were isolated from the skin tissue donated by the patients and cultured by enzymatic digestion. The 5th generation fibroblasts were cultured with PKH26-labeled ADSC-Exos, and observed by confocal fluorescence microscopy. The effects of ADSC-Exos on proliferation and migration of fibroblasts were observed with cell counting kit 8 (CCK-8) and scratch method. Twenty-four 8-week-old Balb/c male mice were used to prepare a diabetic model. A full-thickness skin defect of 8 mm in diameter was prepared on the back. And 0.2 mL of ADSC-Exos and PBS were injected into the dermis of the experimental group ( n=12) and the control group ( n=12), respectively. On the 1st, 4th, 7th, 11th, 16th, and 21st days, the wound healing was observed and the wound healing rate was calculated. On the 7th, 14th, and 21st days, the histology (HE and Masson) and CD31 immunohistochemical staining were performed to observe the wound structure, collagen fibers, and neovascularization. Results: ADSC-Exos were the membranous vesicles with clear edges and uniform size; the particle size was 40-200 nm with an average of 102.1 nm; the membrane-labeled proteins (Alix and CD63) were positive. The composite culture observation showed that ADSC-Exos could enter the fibroblasts and promote the proliferation and migration of fibroblasts. Animal experiments showed that the wound healing of the experimental group was significantly faster than that of the control group, and the wound healing rate was significantly different at each time point ( P<0.05). Compared with the control group, the wound healing of the experimental group was better. There were more microvessels in the early healing stage, and more deposited collagen fibers in the late healing stage. There were significant differences in the length of wound on the 7th, 14th, and 21st days, the number of microvessels on the 7th and 14th days, and the rate of deposited collagen fibers on the 14th and 21st days between the two groups ( P<0.05). Conclusion: ADSC-Exos can promote the wound healing in diabetic mice by promoting angiogenesis and proliferation and migration of fibroblasts and collagen synthesis.

Objective:This study aims to explore the potential effects of adipose-derived stem cell-extracellular vesicles(ASC-EVs) on TGFβ-Smad signaling pathway during myofibroblast trans-differentiation in vitro. Methods:ASCs were isolated from liposuction and flow cytometry was used to detect the surface protein markers. ASC-EVs were isolated from the supernatant of the third to fifth generation ASCs, and the microscopic morphology was observed by transmission electron microscope. The particle size distribution was detected by nano-particle tracking analyzer NanoSight and the membrane surface marker proteins CD63, Alix and TSG101 were detected by flow cytometry. The uptake of EVs by dermal fibroblasts co-cultured with PKH67 fluorescence labeled ASC-EVs was observed by confocal microscope. Dermal fibroblasts were continuously induced by TGFβ1 for five days, and ASC-EVs at the dose of 50 and 100 μg/ml were added. The expression of α-SMA and Smad-2/3/4 were detected by immunofluorescence staining, RT-PCR and Western Blot.Results:The results of flow cytometry showed that the surface markers CD73, CD49d, CD90 and CD105 of the third generation ASCs, were positive, and CD34 and CD45 were negative. Under transmission electron microscope, ASC-EVs was a round membranous vesicle with clear edge and surrounded by bilayer phospholipid membrane. The particle size of more than 95% of the ASC-EVs was distributed between 30 nm to 261 nm, with an average of (166.0±86.1)nm. The specific marker proteins of extracellular vesicle, CD63, Alix and TSG101 were highly expressed. Under confocal microscope, ASC-EVs with green fluorescence were uptake by dermal fibroblasts and distributed in the cytoplasm, and part of ASC-EVs was distributed around the nucleus.TGF-β1 induced a significant increase in the expression of α-SMA in dermal fibroblasts, and the addition of 50 or 100 μg/ml ASC-EVs reduced the expression of α-SMA genes and proteins, but did not show a dose-dependent manner. The gene and protein expression changes of Smad-2/3/4 were consistent with α-SMA. In addition, ASC-EVs could significantly reduced the content of type Ⅰ collagen in the supernatant, but had no significant effect on the secretion of type Ⅲ collagen.Conclusions:The mechanism of ASC-EVs inhibiting scar hyperplasia may be closely related to the suppression of TGF-β-Smad signaling pathway in dermal fibroblasts, inhibition of myofibroblast trans-differentiation and reduction of type Ⅰ collagen secretion.

Objective:This study aims to explore the potential effects of adipose-derived stem cell-extracellular vesicles(ASC-EVs) on TGFβ-Smad signaling pathway during myofibroblast trans-differentiation in vitro. Methods:ASCs were isolated from liposuction and flow cytometry was used to detect the surface protein markers. ASC-EVs were isolated from the supernatant of the third to fifth generation ASCs, and the microscopic morphology was observed by transmission electron microscope. The particle size distribution was detected by nano-particle tracking analyzer NanoSight and the membrane surface marker proteins CD63, Alix and TSG101 were detected by flow cytometry. The uptake of EVs by dermal fibroblasts co-cultured with PKH67 fluorescence labeled ASC-EVs was observed by confocal microscope. Dermal fibroblasts were continuously induced by TGFβ1 for five days, and ASC-EVs at the dose of 50 and 100 μg/ml were added. The expression of α-SMA and Smad-2/3/4 were detected by immunofluorescence staining, RT-PCR and Western Blot.Results:The results of flow cytometry showed that the surface markers CD73, CD49d, CD90 and CD105 of the third generation ASCs, were positive, and CD34 and CD45 were negative. Under transmission electron microscope, ASC-EVs was a round membranous vesicle with clear edge and surrounded by bilayer phospholipid membrane. The particle size of more than 95% of the ASC-EVs was distributed between 30 nm to 261 nm, with an average of (166.0±86.1)nm. The specific marker proteins of extracellular vesicle, CD63, Alix and TSG101 were highly expressed. Under confocal microscope, ASC-EVs with green fluorescence were uptake by dermal fibroblasts and distributed in the cytoplasm, and part of ASC-EVs was distributed around the nucleus.TGF-β1 induced a significant increase in the expression of α-SMA in dermal fibroblasts, and the addition of 50 or 100 μg/ml ASC-EVs reduced the expression of α-SMA genes and proteins, but did not show a dose-dependent manner. The gene and protein expression changes of Smad-2/3/4 were consistent with α-SMA. In addition, ASC-EVs could significantly reduced the content of type Ⅰ collagen in the supernatant, but had no significant effect on the secretion of type Ⅲ collagen.Conclusions:The mechanism of ASC-EVs inhibiting scar hyperplasia may be closely related to the suppression of TGF-β-Smad signaling pathway in dermal fibroblasts, inhibition of myofibroblast trans-differentiation and reduction of type Ⅰ collagen secretion.

Objective To investigate whether Wnt/β-catenin signaling pathway mediating the neuroprotection of isoflurane post-conditioning in hippocampal neurons damage induced by ischemia/reperfusion injury in rats.Methods According to the randomized principle, 60 male Sprague-Dawley rats were randomly divided into five groups (12 rats in each group):sham group (group S), model group (group M), ISO+model group (group MI), ISO+model+DKK-1 group (group MDI) and model+DKK-1 group (group MD).A rat model of middle cerebral artery occlusion (MCAO) was established with 90 min ischemia followed by 24 hreperfusion.Group S was only exposed to one side of the internal carotid artery without fishing line.Isoflurane post-conditioning groups (group MI, MDI) were immediately treated with 1.5％isoflurane for 60 min at the onset of reperfusion.DKK-1 (5μg/kg) was injected intracerebroventricularly 30 min before the model established in group MDI and group MD.After reperfusion for 24 h, Longa score method was used for neurological deficit score.HE staining and Tunel fluorescence was employed to observe the morphological changes of neurons.Immunohistochemistry and Western Blot were applied to detect the expression of target protein in CA1 region.Results Compared with group S, the neurobehavioral score, the number of apoptosis and the expression of Bax and GSK-3βprotein in group M all increased (P<0.05), while the expression ofβ-catenin and Bcl-2/Bax ratio decreased (P<0.05) ;Compared with group M, the neurobehavioral score, the number of apoptosis and the expression of Bax protein were significantly decreased (P<0.05), while the expression of Bcl-2, β-catenin protein and the Bcl-2/Bax ratio were significantly increased (P<0.05) in group MI.Compared with group MI, the neurobehavioral score, the number of apoptosis, Bax and GSK-3βprotein in group MDI were significantly increased (P<0.05), while the Bcl-2, β-catenin protein expression, and Bcl-2/Bax ratio were significantly decreased (P<0.05).Conclusion Isoflurane post-conditioning may protect the hippocampus neurons against cerebral ischemic reperfusion-induced damage via the way that the Wnt/β-catenin signaling pathway regulates the expression levels of Bcl-2 and Bax proteins in rats.

Objective To evaluate the effects of isoflurane postconditioning on angiogenesis during cerebral ischemia-reperfusion ( I∕R) in rats and the role of Shh signaling pathway. Methods Thirty-two clean-grade healthy male Sprague-Dawley rats, aged 6-8 weeks, weighing 220-280 g, were divided into 4 groups ( n=8 each) by a random number table method:sham operation group ( Sham group) , I∕R group, isoflurane postconditioning group ( ISO group) , and isoflurane postconditioning plus Shh signaling pathway specific inhibitor cyclopamine group ( ISO+CYC group) . Cerebral ischemia was produced by inserting a 3-0 nylon thread with a rounded tip into the internal jugular vein. The nylon thread was threaded cranially until resistance was met. Occlusion was maintained for 1. 5 h followed by 24 h reperfusion. Neurological deficit was scored at 24 h of reperfusion. Rats were then sacrificed, and brains were removed for determination of cerebral infarct volume ( by TTC) and expression of glioma-associated oncogene homolog 1 ( Gli1) , vascu-lar endothelial growth factor ( VEGF) and transmembrane phosphoglycoprotein protein ( CD34) in cerebral cortex (by Western blot) and for examination of the pathological changes (by Nissl staining). Results Compared with Sham group, the neurological deficit score and cerebral infarct volume were significantly in-creased, and the expression of Gli1, VEGF and CD34 in the cerebral cortex was up-regulated in I∕R and ISO groups ( P <0. 05) . Compared with I∕R group, the neurological deficit score and cerebral infarct vol-ume were significantly decreased, and the expression of Gli1, VEGF and CD34 in the cerebral cortex was up-regulated ( P<0. 05) , and the pathological changes of brain tissues were significantly attenuated in ISO group, and no significant change was found in the parameters mentioned above in ISO + CYC group ( P>0. 05) . Compared with ISO group, the neurological deficit score and cerebral infarct volume were signifi-cantly increased, and the expression of Gli1, VEGF and CD34 in the cerebral cortex was down-regulated in ISO+CYC group ( P<0. 05) . Conclusion The mechanism by which isoflurane post-conditioning attenuates cerebral I∕R injury is related to activating Shh signaling pathway and promoting angiogenesis in rats.

Objective: To summarize the research progress of mesenchymal stem cells derived exosomes (MSCs-EXOs) in wound repair in recent years. Methods: The literature about the role of MSCs-EXOs in wound repair at home and abroad was extensively consulted. The mechanism of MSCs-EXOs in wound repair and its clinical application prospects were summarized and analyzed. Results: MSCs-EXOs can inhibit early inflammatory reaction, promote angiogenesis, proliferation, and migration of epithelial cells, regulate collagen synthesis, and inhibit scar proliferation in the later stage of wound healing. Compared with MSCs, MSCs-EXOs have many advantages, such as high stability, easy storage, non-tumorigenicity, no proliferation, easy quantitative use, and so on. It has broad clinical application prospects. Conclusion: MSCs-EXOs can promote wound repair and hopefully develop into a clinical product to promote the repair of acute or chronic wounds.

OBJECTIVE To explore the clinical features of patients carrying deletions of the rod domain of the dystrophin gene. METHODS Clinical data of 12 Chinese patients with Becker muscular dystrophy (BMD) and such deletions was reviewed. RESULTS Most patients complained of muscle weakness of lower limbs. Two patients had muscle cramps, one had increased creatine kinase (CK) level, and one had dilated cardiomyopathy. CONCLUSION Compared with DMD, the clinical features of BMD are much more variable, particularly for those carrying deletions of the rod domain of the dystrophin gene. Muscular weakness may not be the sole complaint of BMD. The diagnosis of BMD cannot be excluded by moderately elevated CK. For male patients with dilated cardiomyopathy, the possibility of BMD should be considered.