Objective To explore the expressions of Toll-like receptors (TLRs) 2 and 4 in mouse skin during early immune responses against Sporothrix.Methods A total of 60 BALB/c mice were randomly and equally divided into an experimental group and a control group to be intracutaneously injected with Sporothrix conidium suspensions at a concentration of 1 × 106 cfu/ml and sodium chloride physiological solution respectively.Five mice were sacrificed before the injection,and at 6,12,24,48,and 96 hours after the injection in each group,blood samples were obtained from the mice before sacrifice,and skin tissue specimens were resected from the area around the injection sites after sacrifice.Realtime fluorescence-based quantitative PCR was performed to quantify the mRNA expressions of TLR2 and TLR4,and immunohistochemical staining to observe the protein expressions of TLR2 and TLR4 in mouse skin specimens.Enzymelinked immunosorbent assay (ELISA) was conducted to determine the levels of interleukin 12 (IL-12) and tumor necrosis factor α (TNFα) in serum samples from the mice.Results After injection of Sporothrix conidium suspensions,the mRNA expression level of TLR2 gradually increased and peaked at 24 hours,which was 18.8 times that in the control group at 6 hours and 34 times at 24 hours.In addition,the mRNA expression level of TLR4 in the experiment group reached a peak,and was 56.7 times that in the control group at 6 hours after injection,then gradually decreased and reached the nadir at 96 hours.As immunohistochemical staining revealed,TLR2 and TLR4 were apparently expressed in both keratinocytes and macrophages in skin specimens from the experimental group,but not obviously in those from the control group.No significant differences were observed between the experimental group and control group in serum levels of IL-12 or TNF-α at any of the sampling time points.Conclusion TLR2 and TLR4 may play a favoring role in immunological defense by participating in the recognition of Sporothrix by keratinocytes and macrophages in mouse skin.

Objective To investigate the signaling pathway and the key signal molecules of protein kinase (RIPK)3 in SH-SY5Y cells. Methods SH-SY5Y cells were transfected with RIPK3 expression plasmid vector to upregulate intracellular RIPK3, while the SH-SY5Y cells were transfected with empty vector plasmid, which was considered as control group. Western blot assay was used to check the expression of exogenous RIPK3 in cells. The proliferation rate of SH-SY5Y cells was determined by MTT assay at designated time to detect exogenous RIPK3 activity. Whole transcriptome sequencing (RNAseq) was used to detect the transcription of genes. Whole-transcriptomic gene transcription was measured by following Ingenuity Pathway Analysis (IPA) to obtain downstream signaling pathways and the key molecule, which were partly confirmed by following droplet digital PCR (ddPCR). Results Exogenous RIPK3 showed biological activity in SH-SY5Y, which inhibited the proliferation of cells. IPA showed that znic finger protein 36 (ZFP36) was significantly up-regulated as compared with that of the control group. The tran?scription levels of ZFP36 downstream genes such as tumor necrosis factor (TNF), brain derived neurotrophic factor (BDNF), vascular endothelial growth factor (VEGF) and mRNA-decapping enzyme 2 (DCP2) were affected at the same time. Conclusion Within the limitations of this study, it seems that RIPK3 is notable for the development, inflammation and tumorigenesis of the nervous system as an independent regulator of ZFP36 gene and downstream effectors.

Objective To investigate the effects of synuclein-γ (SNCG) gene silencing on the proliferation and apoptosis of glioma U87-MG cells.Methods Five small hairpin RNA templates targeting SNCG and a negative control were synthesized and cloned into the lentiviral vector system and all the constructs were sequenced.Then the recombinant lentiviral vectors were used to infect U87-MG cells.The lentiviruses which can effectively inhibit protein expression levels of SNCG were selected by RT-PCR for further study.Colony formation and flow cytometry assay were used to investigate the effects of SNCG downregulation by RNA interference on the clony formation,proliferation,and apoptosis of U87-MG cells,respectively.Results The lentiviral vectors carrying 5 shRNAs targeting the SNCG gene were successfully constructed,and SNCG siRNA3 and siRNA5 showed higher interfering efficiency than other vectors.In comparison with the group of negative control,SNCG siRNA3 and siRNA5 were observed to significantly inhibit SNCG expression at the mRNA levels (the relative mRNA levels:siRNA3 (0.17± 0.01)％,siRNA5 (0.13±0.01)％ vs (1.00±0.10)％,P＜0.05).Also,SNCG suppression mediated by RNAi significantly inhibited the clone formation (colony number:siRNA3 (66± 12),siRNA5 (1 ± 1) vs (80± 5),P＜0.05),and the proliferation (ratio of cells in S phase:siRNA3 (41.2±0.7) ％,siRNA5 (39.9±0.5) ％ vs (47.6±2.2) ％,P ＜0.05),but promoted the apoptosis (cell apoptosis:siRNA3 (22.9± 0.4) ％,siRNA5 (28.6± 0.9) ％ vs (1.1 ± 0.1) ％,P＜0.01) of transfected U87-MG cells.Conclusion SNCG suppression at the mRNA level mediated by RNAi can inhibit the proliferation and the clony formation,but induce the apoptosis of glioma U87-MG cells in vitro,suggesting that SNCG suppression mediated by an RNAi strategy may become a novel approach for treating human gliomas.

We evaluated the differential expression and function of chitinase 3‐like‐1 in macrophage stimulated by Sporothrix schenckii and Candida albicans fungicidal ability of macrophage after stimulation with Sporothrix schenckii and Candida albi‐cans separately was detected .The expression of CHI3L1 gene in macrophage stimulated by Sporothrix Schenckii and Candida albicans was evaluated with real‐time PCR .The function of CHI3L1 protein in macrophages against the reproduction of Sporo‐thrix schenckii and Candida albicans was detected in vitro .Results showed that macrophages could engulf and kill Sporothrix Schenckii and Candida albicans in vitro .The expression of CHI3L1 gene in macrophage stimulated by Candida albicans was increased obviously .At the same time ,CHI3L1 protein can damper the reproduction of Candida albicans .However ,the ex‐pression of CHI3L1 gene was not elevated when macrophage was stimulated by Sporothrix schenckii and CHI3L1 protein played little role in reproduction of Sporothrix schenckii .The expression of CHI3L1 gene in macrophage was elevated after stimulation with Candida albicans ,but was not elevated with Sporothrix Schenckii .In correspondence with differential ex‐pression ,CHI3L1 in macrophages could impair the reproduction of Candida albicans but had a weak function on Sporothrix schenckii which might contribute to the pathogenesis of spo‐rotricosis .

Objective To screen altered proteins of hippocampus in the stress-induced depression (STRID) rat model, and explore the potential molecular mechanism. Methods Twenty Sprague-Dawley rats were randomly divided into the control group and STRID group, 10 rats in each group. Chronic unpredictable mild stress (CUMS) methods including fasting for solids and liquids, electric foot-shock, reversing day and night, cold water swimming, cage tilt, scare stimulation and tail pinch were conducted on STRID rats with no repeats for 28 days to make up the depression animal model. The control group was normally fed during this period. After the stress stimulation, the hippocampus protein samples were used for two dimensional electrophoresis to screen the differentially expressed protein, and then mass spectrum identification and function analyze were conducted. Results Compared with the control group, 34 proteins were altered in STRID group. Among which, 18 were up-regulated, and 16 were down-regulated. The differentially expressed proteins mainly located in cytoplasm, mitochondrion, extracellular exosome and myelin sheath. The involved signaling pathways included metabolic pathway, oxidative phosphorylation pathway, and Alzheimer's disease, Parkinson's disease and Huntington's disease pathways. Conclusion The altered proteins and dysfunction of nerve signaling, and the excess of oxidative phosphorylation in hippocampus of STRID rats may be one of the pathogenesises.

The stable isotope labeling by amino acids in culture (SILAC) based quantitative proteomics serves as a gold standard because of the high accuracy and throughput for protein identifications and quantification. In this study, we discussed the application of SILAC technology in mammal model, and developed quantitative internal standard for comparative proteomics of disease model. The C57BL/6J mice fed by special diet containing the 13C6-Lysine and bred F2 generation. We identified and analyzed total proteins of 9 mice tissues of F2 generation, including brain, lung, heart, stomach, intestine, liver, spleen, kidney, and muscle. Quantitative analysis information could evaluate the mice and different tissues' labeling efficiency. Liver was the most efficient, brain the least, and the labeling efficiency were 96.34%±0.90% and 92.62%±1.98% respectively. The average of the labeling efficiency of F2 generation was 95.80%±0.64%, which met the international standard (≥ 95%) for SILAC quantitative proteomics effective study. SILAC technology was successfully extended to mammalian model system, which will provide powerful tools for the mechanism study of the pathophysiology process with mouse model.
Amino Acids ; chemistry ; Animals ; Diet ; veterinary ; Isotope Labeling ; Lysine ; chemistry ; Mice ; Mice, Inbred C57BL ; Proteins ; chemistry ; Proteomics ; methods

Extracorporeal membrane oxygenation (ECMO) could pump the blood from human veins to the outside of the body, oxygenate the red blood cells in an artificial environment and then return them back into the body. ECMO could replace the heart and lungs to complete gas exchange and systemic blood perfusion in patients with severe cardiopulmonary insufficiency, which also plays an important role in the field of heart transplantation. Besides circulatory support treatment after heart transplantation, ECMO may also be used to prolong the waiting time for heart transplantation in patients with respiratory and circulatory failure before operation, as a bridging therapy for heart transplantation. However, at present, the application of ECMO in pediatric heart transplantation still exist challenges, such as high perioperative mortality and difficulty in determining the timing of treatment, etc. In this article, the development history of ECMO application in pediatric heart transplantation, use of ECMO before and after pediatric heart transplantation, ECMO-related complications in children, and application of ventricular assist device (VAD) in pediatric heart transplantation were briefly reviewed, aiming to provide reference for promoting the application of ECMO in pediatric heart transplantation.

Atrial fibrillation is a common arrhythmia associated with high mortality and morbidity, and the current treatment of atrial fibrillation is still limited. Histone deacetylase (HDAC) plays an important role in the pathophysiology of cardiovascular disease and promotes the occurrence of atrial fibrillation. Inhibition of HDAC may be a new therapeutic strategy through the regulation of atrial remodeling. Therefore, we reviewed the research progress of the HDAC and atrial fibrillation.

Objective To explore the effects of different doses of dexmedetomidine on acute brain edema in mice with traumatic brain injury (TBI).Methods A total of 132 male C57BL/6J mice were randomly divided into six groups:control group (group C),sham-operation group (group Sham),traumatic brain injury group (group TBI),Dex 20 μg/kg (group D20),40 μg/kg (group D40),and 60 μg/kg (group D60),n=22 in each group.The TBI animal model was established by electric controlled cortical impactor (eCCI),then intraperitoneal injected by the administration of different doses of dexmedetomidine at 0,2 and 4 h after TBI.Twenty-four hours post-TBI,brain water content was measured by the dry-wet method,histological observation was performed using HE staining,and aquaporin 4 (AQP4) and NF-κB expression were detected using Western blot assay,respectively.Then,the modified neurological scale scores (mNSS) on 1,2,3,and 7 d and Morris water maze (MWM) test on 4,5,6 and 7 d post-TBI were used to evaluate the neurologic deficit of TBI mice.Results After traumatic brain injury,the mNSS scores,the escape latency,the brain water content and the expression of AQP4 and NF-κB increased significantly in group TBI (P＜0.01).Different doses of dexmedetomidine significantly reduced the mNSS scores,the escape latency,the brain water content and the expression of AQP4 and NF-κB (P ＜ 0.05 or P ＜ 0.01).And meanwhile dexrnedetomidine can lessen neuronal degeneration,and inflammation response.Additionally,the effect was remarkably in group D60 compared with group D20 (P ＜ 0.05 or P ＜ 0.01).Conclusion Dexmedetomidine can lessen brain edema and cognition impairment induced with traumatic brain injury,which is a dose-effect relationship within 20-60 μg/kg,and this effect may be related to the downregulation of AQP4 and NF-κB expression.

Objective To assess the safety of the removal of pericardial and mediastinal drain within different drainage volume after cardiac valvular replacement surgery. Methods Between July 2013 and July 2017, 201 patients with rheumatic heart disease (CHD) were treated with valve replacement in our hospital, including 57 males and 144 females, aged 15 to 72 years. They were divided into two groups according to the amount of 24-h drainage before the drain removal: a group one with 24-h drainage volume≤50 ml (n=127) and a group two with 24-h drainage volume>50 ml (n=74). The postoperative hospital stay and the incidence of severe complications between the two groups were compared. Results There was no difference between the two groups in the baseline information or the incidence of severe pericardial effusion and tamponade, while the group two tended to have a shorter length of hospital stay after surgery (8.0 dvs.7.5 d, P=0.013). Conclusion In CHD patients undergoing valvular surgery, compared with a relatively low amount of drainage before the drain removal, drawing the tube at a greater amount of drainage (24-h drainage volume>50 ml) will shorten the length of hospital stay after cardiac surgery while incidence of severe complications remains the same.