BACKGROUND: Serious scalding leads to dysfunction of each aspect in immune system, and activated macrophage can secret many bioactive transmitters. The relationship between macrophage dysfunction and signal conduction after scalding is unclear at present.OBJECTIVE: To observe the alternation of tumor necrosis factor- alpha(TNF-α) at different time points after scalding and the activity of nuclear factor κB(NF-κB) and alternation of protein kinase C (PKC) after the application of specific modulator H-7 to explore whether PKC participates in the modulation of TNF-α in macrophage on signal conduction level for the clarification of some mechanisms of macrophage dysfunction.DESIGN: A randomized controlled experimental study by employing experimental animals as subjectsSETTING: An institute of burn research of a military medical university MATERIALS: The study was conducted in the Laboratory (state) of the Institute of Burn Research, Third Military Medical University of Chinese PLA between January and December 1999. Experimental animals were 32 healthy clean inbreeding Kunming white mice.METHODS: 15% Ⅲ scalding was created in mice for the establishment of routine scalding model. Mice were randomly divided into 6 groups according to different time points before or after scalding, I.e. 0(normal control group), 2, 6, 12, 24, or 48 hours group. Celiac macrophages were collected for the detection of TNF-α content by radioimmunoassay, NF-κB activity by electrophoretic mobility shift analysis (EMSA), and membrane or plasma PKC activity by isotope analysis.MAIN OUTCOME MEASURES: ① TNF-α content; ② NF-κB activity; ③Membrane or plasma PKC activity RESULTS: After scalding, macrophage excessively secreted TNF-α and reached its peak of (1 085.65 ± 122.99) ng/L at 12 hours, which was significantly higher than that of control group( t = 14.92, P ＜ 0.01 ).Compared with control group, membrane PKC activity increased after scalding, which significantly heightened to(231.80 ± 31.66) nmol/min · g at 2hours( t = 7. 930, P ＜ 0.01 ), slightly decreased to close to normal level of (174.29±16.80) nmol/min· gat 6hours(t=2.531, P ＜0.05), and rapidly elevated at 12 hours [512. 10 ±33.42) nmol/min · g] and 24 hours [ (454.70 ± 21.06) nmol/min · g] to reach its peak of(530.49 ± 28.54)nmol/min. G at 48 hours( t = 29.42, 28.03, 30. 19, P ＜ 0. 01 ). Correlation analysis of the alternation between TNF-α and membrane PKC indicated a significant positive correlation( r = 0. 796 4, P ＜ 0. 05) . As indicated by EMSA image, NF-κB activity significantly elevated after scalding. Twelve hours after scalding was set as modulation point, NF-κB activity was significantly inhibited by the application of H-7.CONCLUSION: The secretion of TNF-α and the activities of PKC and NF-κB are significantly activated in celiac macrophage after scalding, and PKC-NF-κB signal pathway participates in the modulation of TNF-α expression, which provide experimental data for the modulation of immune function and rehabilitative intervention during scalding.serious scalding.

BACKGROUND: Severe scald injury leads to a variety of disorders in the immune system. Activated macrophages are known to secrete many kinds of biologically active transmitter, but the relation between the functional disorder of the macrophages and signal transduction after burn injury has not been fully understood.OBJECTIVE: To observe the changes in nuclear factor(NF) -κκB activity and expressions of IκκB-α and tumor necrosis factor(TNF) -α in peritoneal macrophage of mice at different time points after severe scald injury and after the application of specific NF-κκB inhibitor pyrrolidine dithiocarbamate (PDTC), thereby to explore the mechanism of macrophage dysfunction in light of signal transduction.DESIGN: A randomized controlled experimental research.SETTING: State Key Laboratory of Trauma, Burn and Combined Injury.MATERIALS: The experiment was carried out in the State Key Laboratory of Institute of Burn Research, Third Military Medical University during the period from January to June, 1999, using 30 healthy clean-grade Kunming mice of inbred strain.INTERVENTIONS: Common scald injury models(with third degree burn of 15% total body surface area) were established in the mice, which were randomized into 6 groups according to different time points after the injury for observation, namely 0 hour(normal control group) and postburn 2, 6, 12,24 and 48 hours. Peritoneal macrophages were collected at these time points for examining TNF-α content using radio-immunoassay and NF-κκB activity by means of electrophoretic mobility shift assay(EMSA). The expressions of IκκB-α and TNF-α mRNA were determined by immunoblotting method and reverse transcription-PCR, respectively.MAIN OUTCOME MEASURES:Examinations of ① the content of TNF-α, ② NF-κκB activity,③ expression of IκB-α, and ④ expression of TNF-α mRNA.RESULTS: Macrophage secretion of TNF-α was enhanced postburn, reaching the peak level at 12 hours[(1085.65 ± 122.99) ng/L], which was significantly higher than that in the normal control group( t = 14.92, P ＜ 0.01) .Postburn NF-κκB activity significantly increased after the injury, peaking at 2 hours[ (56. 8 ± 7.3)RDU], which occurred much earlier than the peak of TNF-α secretion( t=13. 31, P ＜ 0.01 ). Compared with that in the normal control group, IκB-α expression decreased significantly 2 hours postburn ( t =4. 23, P ＜ 0. 01) to 0. 632 ±0. 086, followed by gradual increase to the peak level to 1. 161 ± 0. 097 24 hours after the burn injury( t = 7.06, P＜ 0. 01) and then by slight decrease to 1. 149 ±0. 167 till 48 hours(t = 4. 82, P ＜ 0.01) . Twelve hours after injury was the time point for intervention with PDTC application, when NF-κκB activity and TNF-α mRNA expression both decreased significantly( P ＜ 0.01 ).CONCLUSION: NF-κB activity and TNF-αmRNA expression decrease significantly after severe scald. At high levels, IκB-α and NF-κκB maintain an interaction for their restriction. After burn injury, NF-κκB signal transduction pathway is involved in the modulation of TNF-α expression in mouse peritoneal macrophage.

Methods: A total of 10 specific pathogen free (SPF) grade female DBA/2J mice were randomly divided into model group and QGA II group (n = 5 for each group), while additional 5 SPF-grade female C57BL/6J mice were assigned to control group. Mice presented spontaneous high IOP and showed elevated approximately at the age of seven months. The high IOP was maintained until week 38, when gavage was initiated. Mice in control group underwent the same intragastric treatment, while those in QGA II group were gavaged with QGA II (9.67 g/kg), once a day for four weeks. Retinal morphology was examined using hematoxylin and eosin (HE) staining, with the number of retinal ganglion cells (RGCs) counted. The expression level of Brn3a protein, a specific marker for RGCs, was detected by immunofluorescence, with the mean optical density (OD) measured for quantitative analysis. In addition, 16S rDNA sequencing was leveraged to analyze changes in the diversity of gut microbiota, including their α-diversity (Chao1, Shannon, Pielou’s evenness, and observed species index) and β-diversity. Venn diagrams and linear discriminant analysis effect size (LEfSe) analysis was employed to investigate the number of amplicon sequence variants (ASVs), the abundance of differential gut microbiota species, and the classification of species at both the phylum and genus levels within the three groups of mice. Results: HE staining revealed that compared with control group, model group showed significant reduction in the number of RGCs (P < 0.01), with intracellular vacuolar degeneration and nuclear pyknosis. After QGA II treatment, the number of RGCs was significantly increased compared with model group (P < 0.01), with notable improvements in intracellular vacuolar degeneration. Immunofluorescence analysis showed that the mean OD of Brn3a protein was significantly decreased in model group compared with control group (P < 0.01), while QGA II treatment significantly elevated its expression level (P < 0.01). Analysis of α-diversity showed that after QGA II intervention, the Chao1, Shannon, and Pielou’s evenness indices were significantly increased (P < 0.01), and the observed species index was elevated (P < 0.05). β-Diversity analysis demonstrated distinct clustering among the three groups, indicating relatively low similarity in bacterial community structures. ASV clustering identified a total of 14 061 ASVs across all groups, with 9 514 ASVs shared between model and QGA II groups. At the phylum level, the abundance of Bacteroidetes was significantly decreased in model group compared with control group (P < 0.01), while Firmicutes and the Firmicutes/Bacteroidetes (F/B) ratio were significantly increased (P < 0.01). QGA II treatment significantly reduced both Firmicutes abundance and the F/B ratio (P < 0.01). At the genus level, Lactobacillus was dominant across all groups, with its abundance significantly increased in model group (P < 0.01) and subsequently decreased following QGA II intervention (P < 0.05). Conclusion QGA II restructured the gut microbiota of DBA/2J mice with chronic high IOP, bringing changes in their diversity and abundance of components. Firmicutes, Bacteroidetes, Lactobacillus, along with their associated microorganisms, are likely critical components of the gut microbiota that contribute to the optic neuroprotective effects of QGA II on chronic high IOP mice.

Objective To evaluate the ability of contrast-enhanced ultrasonography (CEUS)in differentiate reactive lymph node,metastatic lymph node and lymphoma.Methods In a prospective study CEUS was performed in 129 patients with cervical lymph node enlargement.The entire process were recorded and preserved in DICOM format.The results were registered with Sonoliver.The receiver operating characteristic curve (ROC curve)analysis was performed to find the corresponding cutoff values. The selected node was removed surgically and submitted for histology.Results Of all the nodes,26 were reactive nodes,85 were metastases and 1 8 were lymphoma.Enhancement pattern was the most accurate way to characterize lymph nodes.The enhancement pattern of reactive lymph nodes was homogeneous and most of them were enhanced by lymphatic type while the metastatic lymph nodes were inhomogeneously enhanced or weakly enhanced by peripheral type.Lymph node lymphoma usually had no fixed enhancement pattern. Arrive time (AT),rise time (RT),time to peak (TTP),mean transit time (mTT),maximum intensity (IMAX),under the curve (AUC),rising slope(Kup),semi descending slope(Kdown)and perfusion index (PI)were significantly different in the three groups(P <0.05).RT,TTP and mTT of reactive lymph nodes were the shortest,which had significant difference compared with those of metastatic lymph nodes and lymphoma (P < 0.05 ).Kup,Kdown,IMAX%,AUC,PI in the reactive lymph nodes were significantly decreased compared with the metastatic group (P <0.05),but there was no significant difference compared with those in lymphoma (P >0.05 ).When TTP≥ 7.74 s,mTT≥26.54 s,metastatic lymph nodes were considered.When RT≥4.62 s,TTP ≥ 7.74 s,mTT ≥ 28.32 s,reactive lymph nodes were not considered. Conclusions Dynamic contrast-enhanced ultrasound image and enhancement pattern can distinguish neck lymph nodes while the optimal cut-off point time of the time-intensity curve parameters can further contribute to the identification of lymph nodes.

Objective To preliminarily observe the clinical efficacy of hippocampal-sparing prophylactic cranial irradiation (HS-PCI) using helical tomotherapy (HT) in patients with limited-stage small-cell lung cancer (LS-SCLC) after chemoradiotherapy,and compare HT with intensity-modulated radiotherapy (IMRT) and volumetric modulated arc therapy (VMAT) in dose distribution.Methods From April to June,2014,six patients with LS-SCLC who had achieved a complete remission after chemoradiotherapy were assigned to HS-PCI using HT within a month after brain metastasis was ruled out using brain magnetic resonance imaging (MRI).After fusing CT images and MRI images,the hippocampus was contoured in the fusion images and hippocampal avoidance regions were created using a volumetric expansion of 3 mm around the hippocampus.A dose of 25 Gy in 10 fractions to 95％ of planning target volume (PTV) was prescribed in HT,IMRT,and VMAT.The clinical efficacy,adverse reactions,neurocognitive function,and brain metastasis were evaluated for HT.The dose distribution in PTV and hippocampus were compared between HT,IMRT,and VMAT.Results There were one patient with abdominal wall and abdominal lymph node metastases,one patient with local recurrence,and no patient with brain metastasis during the observation period.The numbers of patients with grade 1 and grade 2 headache,dizziness,and hair loss reactions were 3 and 1,3 and 1,and 4 and 2,respectively.There were no significant differences in the average score of the Mini-Mental State Examination before treatment and at 3 and 6 months after treatment (29.7,29.2,and 29.3 ; P =0.083,0.317,and 0.157).The mean dose to the hippocampus was 16.85 Gy for IMRT and 17.59 Gy for VMAT.For HT,the mean doses to the hippocampus and avoidance regions were reduced to 5.26 Gy and 6.21 Gy,respectively.The prescribed dose for HT was reduced by 79％ and 71％ compared with IMRT and VMAT,respectively.The average coverage rate of the prescribed dose was 94.48％ for HT.Conclusions HT achieves promising dose distribution and target coverage in sparing of the hippocampus.Moreover,HT dose not increase the incidence of adverse reactions.The change in neurocognitive function needs to be further studied with longterm observation and large-scale sampling.

A plan of constructing a platform of excavating and protecting characteristic technology of diagnosis and treatment in research-based hospital of TCM was put forward by Shuguang Hospital Affiliated with Shanghai University of Traditional Chinese Medicine and Research Institution of Characteristic Technology of Diagnosis and Treatment Affiliated with Shanghai Academy of Traditional Chinese Medicine. The platform has begun to take shape on the basis of previous theoretical thought and preliminary practice. It has its own management system and working standards and contents with a clear and definite developing direction. Its working contents include excavating and protecting clinical experience of famous and aged practitioner of Chinese medicine, folk proved recipe and technology of diagnosis and treatment of TCM, intangible cultural heritage of TCM. Some problems appeared during the work, such as the evaluation criterion of the folk proved recipe and technology of diagnosis and treatment, the management of clinical proving projects, the issue of intellectual property protection and so on, which must be further solved.

Our previous studies demonstrated that CD151 gene promoted neovascularization in ischemic heart model. To improve the delivery efficacy and target specificity of CD151 gene to ischemic heart, we generated an adeno-associated virus (AAV) vector in which CD151 expression was controlled by the myosin light chain (MLC-2v) promoter to achieve the cardiac-specific expression of CD151 gene in ischemic myocardium and to limit unwanted CD151 expression in extracardiac organs. The function of this vector was examined in rat ischemic myocardium model. The protein expression of CD151 in the ischemic myocardium areas, liver and kidney was confirmed by using Western blot, while the microvessels within ischemic myocardium areas were detected by using immunohistochemistry. The results showed that MLC-2v significantly enhanced the expression of CD151 in ischemic myocardium, but attenuated its expression in other organs. The forced CD151 expression could increase the number of microvessels in the ischemic myocardium. This study demonstrates the AAV-mediated and MLC-2v regulated CD151 gene is highly expressed in the ischemic myocardium and cardiac-specific delivery that is more efficiently targets CD151 to the ischemia myocardium after myocardial infarction.

Objective To analyze our clinical results of head and neck reconstruction using microsur- gical free flap transfer techniques.Methods The free flap donor sites with long vascular pedicle and large diameter of vessel were routinely chosed,and chose receipt vessels with large diameter and proper position, and perform vessel ananstomosis under surgical loups instead of microscope.The un-buried free flap with a mo- nitoring window were harvest,and do double venous anastomoses in some flaps to ensure adequate venous out- flow.Results From May 1999 to March 2005,1066 consecutive free flap transfers were used to reconstruct head and neck defects.The overall success rate of free flap was 98.3%.The vessel thrombosis rate was 3.1%,and the flap salvage rate was 45.5%.Conclusion Head and neck reconstruetion using microsurgi- cal free flap transfer technique is safe and reliable,and good clinical results can be obtained.

BACKGROUND: Induced pluripotent stem cells (iPSCs) are a special type of cells with self-renewal and multi-differentiation potential, which can differentiate into intestinal organoids under certain conditions. OBJECTIVE: To explore whether iPSCs can differentiate into intestinal organoids under specific conditions in vitro.METHODS: iPSCs from B6J mice were recovered and cultured for 3 days until clone units covered about 80％ of the culture dish, and then the cells were cultured in the medium containing Activin A for 3 days until the deterministic endoderm formed. Further, the culture medium was replaced by the medium with fibroblast growth factor 4 and Wnt3A for 4 days to differentiate into the spheroids with CDX2+. After that, spheroids were collected and mixed with Matrigel,and then the mixture was dropped into the 4-well plate and cultured with Rspondin1, Noggin, epidermal growth factor, B27 and other growth factors to differentiate into intestinal organoids. Cell morphology was observed, FoxA2 and Sox17 expresson in the deterministic endoderm was detected, and CDX2, Sox9, CGA, MMP7 were measured.RESULTS AND CONCLUSION: iPSCs were cultured with Activin A for 3 days with higher cell fusion, initial differentiation and FoxA2/Sox17 expression (P < 0.05) than those of non-induced iPSCs. Spheroids began to appear at the 3rd day after culture with fibroblast growth factor 4 and WNT3A, and formed a lot at the 4th day. And CDX2 expression in spheroids was significantly increased compared with that in the deterministic endoderm (P < 0.05). Organoids gradually formed after 3 days culture, which contained all cell types of intestinal organoids, and expressions of specific markers, Sox9, CGA, MMP7, were significantly higher than those in spheroids (P < 0.05). To conclude, iPSCs can be induced to differentiate into intestinal organoids in three-dimensional niche in vitro.

Objective To observe the application value and therapeutic efficacy of wide-field digital pediatric retinal imaging system (Retcam Ⅲ) fundus fluorescein angiograms (FFA) assisted photocoagulation on familial exudative vitreoretinopathy (FEVR).Methods The study included 46 eyes of 34 patients with staging 2 FEVR.All patients received color fundus photography and FFA under general anesthesia.The blood vessel reliability of color fundus photography and FFA was comparatively determined.Binocular indirect ophthalmoscope laser photocoagulation was applied to peripheral retina with abnormal leakage as indicated by FFA,the wavelength was 532nm,the duration was 0.25 s and the energy was 200-280 mW.After laser photocoagulation,fundus imaging and FFA was repeated.Further laser photocoagulation was immediately added to areas with vessel leakage but missing the photocoagulation.After treatment,the mean follow-up duration was 14.4 months.The follow up focused on neovascularization,exudative lesions,vitreous traction and merging of photocoagulation spots within 3 months,and on fibrosis membrane resulting in macular traction,tractional retinal detachment,vitreous hemorrhage or Coats disease-like retinal exudates after 3 months.Results It was hard to identify the blood vessels based on the color fundus images and some avascular zone maybe missed.Neovascularization can't be determined by shape of the blood vessels.On the other hand,those new blood vessels can be easily recognized by FFA as leakage sites at the boundary of avascular zone.The surgeon could quickly and accurately locate the FEVR area guided by the color fundus images and FFA from same angle under binocular indirect ophthalmoscope.During the treatment,there was no retinal FEVR area missed laser photocoagulation for all patients.There was no neovascularization,exudative lesions,vitreous traction within 3 months,and no fibrosis membrane,tractional retinal detachment,vitreous hemorrhage or Coats disease-like retinal exudates after 3 months.There were no ocular and systemic complications during and after the FFA and laser photocoagulation.Conclusion Wide-field Retcam Ⅲ FFA can help retinal specialists to identify abnormal neovascularization,locate the lesion area,and thus increase the success rate of laser photocoagulation,reduce the ocular and systemic complications for FEVR.