This study explored the protective effect of astragaloside Ⅳ(AS-Ⅳ) on oxygen-glucose deprivation(OGD)-induced autophagic injury in PC12 cells and its underlying mechanism. An OGD-induced autophagic injury model in vitro was established in PC12 cells. The cells were divided into a normal group, an OGD group, low-, medium-, and high-dose AS-Ⅳ groups, and a positive drug dexmedetomidine(DEX) group. Cell viability was measured using the MTT assay. Transmission electron microscopy was used to observe autophagosomes and autolysosomes, and the MDC staining method was used to assess the fluorescence intensity of autophagosomes. Western blot was conducted to determine the relative expression levels of functional proteins LC3-Ⅱ/LC3-Ⅰ, Beclin1, p-Akt/Akt, p-mTOR/mTOR, and HIF-1α. Compared with the normal group, the OGD group exhibited a significant decrease in cell viability(P<0.01), an increase in autophagosomes(P<0.01), enhanced fluorescence intensity of autophagosomes(P<0.01), up-regulated Beclin1, LC3-Ⅱ/LC3-Ⅰ, and HIF-1α(P<0.05 or P<0.01), and down-regulated p-Akt/Akt and p-mTOR/mTOR(P<0.05 or P<0.01). Compared with the OGD group, the low-and medium-dose AS-Ⅳ groups and the DEX group showed a significant increase in cell viability(P<0.01), decreased autophagosomes(P<0.01), weakened fluorescence intensity of autophagosomes(P<0.01), down-regulated Beclin1, LC3-Ⅱ/LC3-Ⅰ, and HIF-1α(P<0.05 or P<0.01), and up-regulated p-Akt/Akt and p-mTOR/mTOR(P<0.01). AS-Ⅳ at low and medium doses exerted a protective effect against OGD-induced autophagic injury in PC12 cells by activating the Akt/mTOR pathway, subsequently influencing HIF-1α. The high-dose AS-Ⅳ group did not show a statistically significant difference compared with the OGD group. This study provides a certain target reference for the prevention and treatment of OGD-induced cellular autophagic injury by AS-Ⅳ and accumulates laboratory data for the secondary development of Astragali Radix and AS-Ⅳ.
Rats ; Animals ; PC12 Cells ; Proto-Oncogene Proteins c-akt/genetics* ; Glucose/therapeutic use* ; Oxygen/metabolism* ; Beclin-1/pharmacology* ; TOR Serine-Threonine Kinases/metabolism* ; Autophagy ; Apoptosis ; Reperfusion Injury/drug therapy*

ObjectiveThe mechanism of cerebral ischemia-reperfusion injury is highly associated with the inflammatory response. MiRNA-126 plays a key role in vascular inflammation. This study aims to investigate the effect of miRNA-126 on the inflammatory response in mice accompanying cerebral ischemia-reperfusion injury through the NLRP3/NF-κB signal axis, and to explore the mechanisms involved.Methods A total of forty-eight male mice were randomly divided into four groups: the sham-operated group, cerebral ischemia-reperfusion model group, miRNA negative control group (miRNA-126 agomir NC group) and miRNA-126 overexpression group (miRNA-126 agomir group), and each group included twelve mice. The neurobehavioral score was recorded. The left-brain of the mice was sacrificed after anesthesia, and the water content of the brain tissue was measured. HE staining and light microscopy were used to identify the histopathological changes of the cerebral of the mice. The expression levels of inflammatory cytokines IL-1β and IL-6 in brain tissue and serum of mice were detected by ELISA. Western Blot method was used to determine the protein expression levels of NLRP3, ASC, caspase-1, NF-κB p65, NF-κB P50, p-NF-κB 65 and p-NF-κB 50 in brain tissues of mice in each group. RT-PCR was used to test the expression levels of miRNA126, NLRP3, ASC, caspase-1, NF-κB p65 and NF-κB P50 in brain tissue and serum of mice.ResultsIn the sham-operated group, the morphology, and structure of cerebral cortex were normal as healthy mice, being with the dense and orderly arrangement of nerve fibers, with no occurrence of impaired nerve function, and the neurobehavioral score was zero. In both of model group and the miRNA-126 agomir NC group, the ruptured cerebral cortex could be observed visually being with necrotic and disordered cells. The blurred pyknosis and interstitial edema occurred with increased water content of brain tissue. The nerve damage was observed with a significantly increased neurobehavioral score (P<0.05). Compared to the model group, the pathological morphology of the cerebral cortex in the miRNA-126 agomir group was significantly improved, and the number of necrotic cells was decreased, the arrangement of which was denser and more orderly. Reduced interstitial edema and the neurobehavioral score were identified. The significantly improved nerve injury and the decreased water content of brain tissue were observed as well (P<0.05). Compared to the sham-operated group A, the expression level of miRNA-126 mRNA in the model group and the miRNA-126 agomir NC group decreased significantly. The expression level of IL-1β and IL-6 increased, while the expression levels of NLRP3, ASC, caspase-1, NF-κB p65, NF-κB p50, p-NF-κB p65, p-NF-κB p50, and NLRP3, ASC, caspase-1, NF-κB p65, NF-κB p50 mRNA increased generally (P<0.05). Compared to the model group and the miRNA-126 agomir NC group, the expression level of miRNA-126 mRNA in the miRNA-126 agomir group increased. However, the expression level of IL-1β and IL-6 decreased, and the expression level of NLRP3/NF-κB signal axis related gene protein and mRNA decreased (P<0.05).ConclusionOverexpression of miRNA-126 can inhibit the expression of NLRP3/NF-κB signal axis related genes and the level of inflammation in brain tissue, and improve the neurological injury of cerebral ischemia-reperfusion in mice.

Objective To study the biological properties of microRNA (miRNA) -21 expression in esophageal squamous cell carcinoma. Methods MiRNA-21 sense expression vector, miRNA-21 empty vector and miRNA-21 inhibitor were transfected and detected by fluorescence Real-time PCR. The normal cells of esophageal squamous cell carcinoma TE-1 were used as the control group, and the transfection result were detected. The cell proliferation activity was detected by cell counting kit-8(CCK-8), and the apoptosis ability was detected by flow cytometry. The invasion ability and migration ability of TE-1 cells were observed by Transwell chamber and scratch test. Changes in the sensitivity of TE-1 cells to radiotherapy were observed by colony formation experiments. The data were analyzed using SPSS 19. 0 software. Results The result of Real-time PCR showed that the expression level of miRNA-21 in TE-1 cells was significantly decreased after transfected with miRNA-21 inhibitor (P<0. 05). Compared with the normal cell growth group, positive control group and negative control group, the proliferation of TE-1 cells was reduced and apoptosis was accelerated after transfection with miRNA-21 inhibitor (P<0. 05). In addition, the ability of cell invasion and migration in the experimental inhibition group was decreased. Clone formation assays showed that inhibition of miRNA-21 expression significantly increased the sensitivity of TE-1 cells to radiation. Conclusion Down-regulation of miRNA-21 reduces the proliferation, invasion, migration and sensitivity of radiotherapy in esophageal squamous cell carcinoma TE-1 cells, and is a potential target for the future treatment of esophageal cancer.

A high-content GABA was found in Sojae Semen Praeparatum(SSP), which is a famous traditional Chinese medicine and officially listed in Chinese Pharmacopoeia. To screen out and identify GABA-producing microbes from samples at different time points during the fermenting process of SSP, traditional microbiological methods combined with molecular biological methods were used to study the predominant GABA-producing microorganisms existing in the fermenting process of SSP. This study would lay a foundation for further studying the processing mechanism of SSP. The fermenting process of SSP was based on Chinese Pharmacopoeia(2010 edition), and samples were taken at different time points during the fermenting process of SSP. The bacteria and fungi from samples at different time points in the fermenting process of SSP were cultured, isolated and purified by selective medium, and dominant strains were selected. The dominant bacteria were cultured in the designated liquid medium to prepare the fermentation broths, and GABA in the fermentation broth was qualitatively screened out by thin-layer chromatography. The microbial fermentation broth with GABA spots in the primary screening was quantitatively detected by online pre-column derivatization and high performance liquid chromatography established in our laboratory. GABA-producing microorganisms were screened out from predominant strains, and their GABA contents in fermentation broth were determined. The DNA sequences of GABA-producing bacteria and fungi were amplified using 16S rDNA and 18S rDNA sequences by PCR respectively. The amplified products were sequenced, and the sequencing results were identified through NCBI homology comparison. Molecular biological identification was made by phylogenetic tree constructed by MEGA 7.0 software. Through the homology comparison of NCBI and the construction of phylogenetic tree by MEGA 7.0 software, nine GABA-producing microorganisms were screened out and identified in this study. They were Bacillus subtilis, Enterococcus faecium, E. avium, Aspergillus tamarii, A. flavus, A. niger, Cladosporium tenuissimum, Penicillium citrinum and Phanerochaete sordida respectively. For the first time, nine GABA-producing microorganisms were screened out and identified in the samples at different time points during the fermenting process of SSP in this study. The results indicated that multiple predominant GABA-producing microorganisms exist in the fermenting process of SSP and may play an important role in the formation of GABA.
Bacteria ; classification ; metabolism ; Chromatography, High Pressure Liquid ; Fermentation ; Fungi ; classification ; metabolism ; Phylogeny ; Seeds ; microbiology ; Soybeans ; microbiology ; gamma-Aminobutyric Acid ; biosynthesis

Objective Gastrokines 2(GKN2) protein expression is detected in gastric cancer tissues, the distal gastric mucosa tissues and the adjacent normal gastric tissues; the effect of GKN2 overexpression is displayed on proliferation,migration and invasion in human gastric cancer cell line MKN28. Methods Immunohistochemistry was performed to observe GKN2 expression in gastric cancinoma tissues,adjacent gastric mucosa tissues and distal gastric mucosa tissues. Then,the GKN2 gene eukaryotic expression vector Xhol GKN3SP-hGKN2-TEV-SBP Xhol was transfected into human gastric cancinoma MKN28 cells. Western blot was used to evaluate the expression of GKN2 in MKN28 cells. Cell viability,migration and invasion ability were investigated by MTT,Transwell migra-tion assay and Transwell invasion assay. Results GKN2 expression in gastric cancinoma tissues is lower in com-parison with those in the adjacent normal gastric tissues or distal gastric mucosa tissues respectively (P< 0.05).The viability of MKN28 cells was significantly inhibited after transfected by GKN2 vs untransfected cells (P<0.05). GKN2 overexpression decreased the cells' number passing through both the membrane and matrigel, compared to those of untransfected cells(P < 0.05). Conclusions Downregulation of GKN2 protein expression is associated with the occurrence of gastric cancer; GKN2 overexpression can suppress the prolifera-tion, migration and invasion of MKN28 cells.

Adenine ; Animals ; Gene Editing ; Mice ; Zygote ; metabolism

OBJECTIVETo investigate the impact of cement distribution index on the occurrence of refracture in the adjacent segments after percutaneous vertebroplasty.

METHODSThis retrospective analysis was conducted among 143 patients who received percutaneous vertebroplasty for osteoporotic vertebral compression fracture between April, 2011 and April, 2014. Of the 134 patients with complete follow-up data, 18 had adjacent segment fracture within 1 year following the surgeries (re-fracture group), and 116 patients without new fracture served as the control group. All the patients underwent X-ray examinations after the surgery and according to the position and shape, the cement in the vertebrae were classified into 5 types (I to V), and the volume-cubage index was computed based on the cement volume and vertebral cubage. Age, gender, bone mineral density (BMD), cement distribution index, volume-cubage index, and cement leakage were evaluated in the 2 groups, and the variables with significant differences between the 2 groups were analyzed in Logistic regression analysis.

RESULTSBMD was significantly lower and the rate of cement leakage was significantly higher in the re-fracture group than in the control group (P<0.05). Significant difference was found in cement distribution index between the 2 groups (P<0.05) but not in age, gender, cement volume or volume-cubage index (P>0.05). Logistic regression analysis indicated that BMD, cement leakage and cement distribution index all significantly affected the occurrence of adjacent vertebral fractures following percutaneous vertebroplasty.

CONCLUSIONA low BMD, cement leakage and a low cement distribution index are all risks factor of adjacent vertebral fracture after percutaneous vertebroplasty.

Objective To develop an online organ doses reporting software VirtualDose-IR, which can compute the radiation doses and provide an easy access to evaluation and control of patients ′radiation doses.Methods Monte Carlo method was applied to simulating various interventional radiology ( IR) processes , which included various parameters such as different patient models at different ages and with different weights , different projection angles and regions of interest , and other parameters .All of the dose data was acquired and then integrated into a database , and displayed with hyper text markup language (HTML), so only a web browser was necessary for users .Results A web-based software that reports organ doses for patients under IR progress was developed .The organ doses assessed with VirtualDose-IR were compared with other experiment and simulation data , and the results were basically consistent with each other .Conclusions VirtualDose-IR is a easy and efficient method to assess patients′radiation doses of IR.

Objective To develop an emergency clinical laboratory system for civilian and military uses to enhance medical support of filed medical unit for mass casualties.Methods The missions of field medical unit were analyzed in the actual confrontation,and an emergency clinical laboratory system for civilian and military uses was built to fulfill clinical laboratory support.The factors were explored for clinical laboratory examination,and some measures were taken accordingly.Results The requirements of field medical unit were met by the developed system.Conclusion The system can be applied in largescale military exercises to enhance clinical laboratory support ability of field medical unit.

OBJECTIVETo investigate the epidemiological and clinical features of calicivirus-associated diarrhea in hospitalized children in Chengdu, China in recent years.

METHODSThe clinical data of 267 children with calicivirus-associated diarrhea aged <5 years who were hospitalized in Chengdu Women and Children's Central Hospital (the only sentinel hospital for sample collection of pediatric viral diarrhea in Chengdu, Sichuan) between January 2012 and December 2014 were retrospectively studied.

RESULTSAmong the 267 children, 200 (74.9%) were aged less than 1 year. The infection rate of calicivirus was 28.4%, 21.6%, and 27.1% in 2012, 2013, and 2014, respectively. Calicivirus was prevalent in summer and autumn (August to October). The detection rate of Norovirus II was 85.8% (229/267), and 244 children (91.4%) experienced an acute clinical course. Watery stool was the most common change in stool properties (82.0%, 219 children), and some specimens showed mucus and/or blood. Most children had moderate to severe fever. One hundred and thirty-eight children (53.9%) experienced a reduced serum prealbumin level. One hundred and fifty-nine children (59.6%) experienced flora imbalance.

CONCLUSIONSCalicivirus has become one of the major pathogens for diarrhea in children aged <5 years in Chengdu, with Norovirus II as the dominant strain. Calicivirus is prevalent in summer and autumn. Infants aged <1 year are the main population affected by calicivirus-associated diarrhea, with watery stool as the most common manifestation.

Adolescent ; Caliciviridae Infections ; epidemiology ; Child ; Child, Hospitalized ; Child, Preschool ; China ; epidemiology ; Diarrhea ; epidemiology ; Female ; Humans ; Infant ; Male ; Time Factors