Background C57BL/6andB10R Ⅲareroutinemurinespeciesusedinexperimental autoimmune uveitis (EAU).The inflammation is light for mouse after immunization whereas it is prominent for B10R Ⅲ.ObjectiveThis study was to observe the killing effect of interphotoreceptor retinoid binding protein (IRBP) 1-20-specific T cells on mouse retinal astrocyte.Th1 and Th17 cells effect in the EAU mechanism was discussed.MethodsB10RllⅢ mice and C57BL/6 mice were immunized with IRBP 161-180 and IRBP 1-20 in complete Freund adjuvant (CFA).The infiltrating cells of diseased B10R Ⅲ eyes were analyzed by flow cytometry.IRBP 1-20-specific T cells were isolated from the drainage lymph node and spleen and cultured in IL-2 or IL-23 for Th1 and Th17 cells polarization,respectively.Th1 and Th17 cells cultured for 5 days were seeded on the mouse retinal astrocyte monolayer pretreated with gamma interferon.Cell interaction was observed and the quantity of TNF-α was tested by ELISA.Every test was repeated 6 times and the mean was calculated.The maintenance of experimental animals complied with the Statement of ARVO.ResultsThere were lots of infiltrating cells in the eyes of B10Rm mice after immunization,including 9.5％ IFNγ+ cells,5.1％ IL-17+cells and 41.4％ CD45+ cells.Six days after IRBP1-20 stimulation and cultured by IL-2 and IL-23,44.0％ and 8.0％ cells were IFNγ+,and 1.0％ and 26.0％ cells were IL17+.Twentyfour hours after the interaction between Th1 or Th17 and retinal astrocyte,retinal astrocyte died and detached.The killing effect of Th17 was stronger than Th1.48 hours after co-culture of Th1 or Th17T cells with astrocytes,the concentrations of TNF-α were ( 500± 10 ) and ( 801 ±24 μg/L) μg/L,respectively,with a significant statistical difference (t =-20.36,P =0.00).ConclusionsBoth Th1 and Th17 can kill retinal astrocyte,but Th17 plays a key role in the EAU pathogenesis process.The killing effect is caused by intercellular contact and interaction under the induction of cytokines.

Bacterial Infections ; transmission ; Drug Resistance, Microbial

Background Proliferative vitreoretinopathy (PVR) is one of the major causes of retinal detachment surgery failure.Based on proteomic studies of PVR vitreous,the insulin-like growth factor binding protein-6 (IGFBP-6) protein was specifically expressed in the vitreous and serum of PVR patients.Furthermore,its expression level is higher in the vitreous and serum in severe PVR patients than that in mild PVR patients.Objective This experiment was to detect the expression of IGFBP-6 in a PVR rat model.Methods Seventy 7-week old male SPF Wistar rats were included and were randomized into the PVR model group and control group.A mixture of RPE-J cell suspension(5 μl) and platelet-rich plasma (5 μl) was intravitreally injected in the left eyes of adult Wistar rats to establish the PVR model,and normal saline solution was administered in the same way in the control group.The rat eyes were clinically examined 1 week,2,3 and 4 weeks after injection,and PVR was graded based on the criteria of Francine.The animals were sacrificed after 1 week,2,4 or 8 weeks for the preparation of retinal sections and liver extraction.Expression levels of IGFBP-6 mRNA in the rat retina and liver were assayed by real-time Q-PCR.The expression of IGFBP-6 protein in the rat serum and vitreous was detected by ELISA.The use of animals complied with the Regulations for the Administration of Affairs Concerning Experimental Animals by State Science and Technology Commission.Results Purified IGFBP-6 RNA was extracted from the liver and retina of Wistar rat and quantified by real-time Q-PCR.The expression level of IGFBP-6 mRNA in retina was (3.79± 1.33) × 10-4 in the PVR model rats,showing a significant decline in comparison with the control rats with a level of(8.32±2.96) × 10 4,4 weeks after injection (t =3.42,P＜0.01).The expression of IGFBP-6 mRNA in the 4th week was significantly lower than that of 1 week,2 or 8 weeks after the establishment of the PVR model(P＜0.05).No significant difference was found in the IGFBP-6 mRNA level in the liver between the PVR group and control group(27.60± 14.01 × 10 4 vs.25.01 ± 12.04 ×10-4,respectively),as well as among the different time points(P＞0.05).IGFBP-6 mRNA content in the retina was significantly reduced in grades 1,2 or 3 of the PVR groups compared with the control group(P＞0.05),but there was no significant difference among the different grades of PVR groups (P＞0.05).Concentrations of IGFBP-6 protein in grades 1,2 and 3 of the PVR model group were (221.00 ± 19.32),(229.63 ± 18.89) and (225.70 ± 26.71) μg/L,with a significant elevation in comparison with (173.25 ±21.11) μg/L of the control group (t =2.14,P＜0.05).However,there was no significant change among the different grades of PVR groups(t=1.24,1.46,P＞0.05).The concentrations of IGFBP-6 protein in the vitreous and serum were higher in PVR rat samples (vitreous:225.44±19.36 μg/L;serum:108.48 ± 15.78 μg/L) than in control rats (vitreous:173.25 ± 21.11 μg/L,serum:95.96 ±17.40 μg/L)(P＜0.05).Conclusions The concentrations of IGFBP-6 protein in the vitreous and serum increase in PVR rats.The results indicate that the increased IGFBP-6 in the vitreous might be a localized autocrine secretion of the eye.

Humans ; Laparoscopy ; Proctectomy ; Rectal Neoplasms/surgery* ; Rectum/surgery* ; Transanal Endoscopic Surgery

OBJECTIVETo study the effect of Buyang Huanwu decoction (BYHWD) inducing angiogenesis on the neuroblast migration from the subventricular zone and its mechanisms after focal cerebral ischemia.

METHODThe middle cerebral artery occlusion (MCAO) was performed to mice for 30 minutes to establish the model. The rats were divided into sham group, model group, BYHWD group and endostatin group. BYHWD (20 g x kg(-1), ig) and endostatin (10 μg, sc) were administered 24 h after ischemia once a day for consecutively 14 days. At 14 d after ischemia, the density of micro-vessel and the number of neuroblasts in the ischemia border zone were determined by immunofluorescence staining. The mRNA and protein expression of cell-derived factor-1 (SDF-1) and brain-derived neurotrophic (BDNF) were examined by real-time PCR and Western blot.

RESULTCompared with the model group, BYHWD significantly increased the density of micro-vessel and the number of DCX positive cells in the ischemia border zone (P < 0.01), and significantly increased the SDF-1 and BDNF mRNA and protein expression (P < 0.01). Compared with BYHWD group, endostatin significantly reduced the density of micro-vessel and the number of DCX positive cells in the ischemia border zone (P < 0.01), as well as the SDF-1, BDNF mRNA and protein expression (P < 0.01).

CONCLUSIONBYHWD could promote the neuroblast migration from the subventricular zone via inducing angiogenesis after cerebral ischemia, the mechanism may be correlated with up-regulating the expression of SDF-1 and BDNF.

Angiogenesis Inducing Agents ; pharmacology ; Animals ; Brain Ischemia ; pathology ; physiopathology ; Brain-Derived Neurotrophic Factor ; analysis ; genetics ; Cell Movement ; drug effects ; Cerebral Ventricles ; pathology ; Chemokine CXCL12 ; analysis ; genetics ; Drugs, Chinese Herbal ; pharmacology ; Male ; Mice ; Mice, Inbred ICR ; Neurons ; drug effects ; physiology

OBJECTIVETo investigate the therapeutic mechanism of three-step sequential method (TSSM) on patients with corticosteroid-dependent asthma (SDA).

METHODSForty patients with SDA were randomly assigned according to the randomizing number table to two groups equally, the treated group treated with three-step sequential recipes plus inhalation of Pulmicort Turbuhaler 200 microg, twice a day, and the control group treated with Pulmicort Turbuhaler alone. The therapeutic course for both groups was 12 - 14 weeks. Changes of the symptom score of asthma, the corticosteroid dosage used and the lung function were observed and the positive expression rate of IFN-gamma and IL-4 in peripheral CD4+ T cells were determined by flow cytometry before and after treatment.

RESULTSThere was significant difference in the asthma symptom score, the oral corticosteroid dosage and the lung function between the treated group and the control group after treatment (P < 0.01). The expression rate of Th2 reduced, the ratio of Th1/Th2 increased significantly after treatment in both groups (P < 0.01, P < 0.05), but the changes were more remarkable in the treated group than those in the control group, showing significant difference between them (P < 0.01), while the expression rate of Th1 had no obvious change after treatment with no significant difference shown between the two groups (P > 0.05).

CONCLUSIONTSSM can regulate imbalance of Th1/Th2, inhibit generation of inflammatory cytokines, decrease airway hyper-response, and therefore improve the pulmonary function, alleviate the asthmatic symptoms and reduce the patients' dependence on corticosteroid.

Adult ; Asthma ; drug therapy ; Drugs, Chinese Herbal ; administration & dosage ; therapeutic use ; Female ; Glucocorticoids ; adverse effects ; therapeutic use ; Humans ; Male ; Middle Aged ; Phytotherapy ; methods ; Substance-Related Disorders ; drug therapy ; etiology ; T-Lymphocyte Subsets ; drug effects ; T-Lymphocytes, Helper-Inducer ; drug effects

Previous studies showed that three methods for regulating and invigorating lung and kidney (lung invigorating and spleen strengthening, lung invigorating and kidney tonifying, and Qi supplementing and kidney nourishing) could regulate inflammatory signaling pathways of chronic obstructive pulmonary disease (COPD) in rats, so as to alleviate inflammation. In the present study, R-value comprehensive evaluation method was used to evaluate the comprehensive effect of three methods for regulating and invigorating lung and kidney on inflammatory signaling pathways. Rats were randomly divided into control, model, lung invigorating and spleen strengthening, lung invigorating and kidney tonifying, Qi supplementing and kidney nourishing and aminophylline groups. The COPD rat models were established by cigarette smoking combined with bacterial infection, and orally administered with drugs between the 9th and 20th week. Afterwards, efforts were made to observe the long-term effects between the drug withdrawal and the 32rd week and detect indicators in two batches in the 20th week and 32th week. Specifically, (1) Linking JAK/STAT signaling pathway: JAK2 mRNA, and protein expressions of STAT-1, STAT-3, STAT-5, JAK-2; (2) NF-kappaB signaling pathway: Smad2 mRNA and protein expressions of I-kappaB, NF-kappaB, TGF-beta1; (3) PPARgamma and antioxidant signaling pathway: SOD, PGE mRNA, PPARgamma protein. According to the results, 5 indicators in JAK/STAT pathway, 4 indicators in NF-kappaB pathway, and 3 indicators in PPARgamma pathway were significantly rectified by three methods for regulating and invigorating lung and kidney in between the 20th week and 32nd week. Between the 20th and 32nd week, the recipes for rectifying JAK/STAT pathway with intensity from high to low were recipes for lung invigorating and spleen strengthening, Qi supplementing and kidney nourishing, lung invigorating and kidney tonifying, aminophylline, particularly those for lung invigorating and spleen strengthening; The recipes for rectifying NF-kappaB pathway with intensity from high to low were recipes for lung invigorating and spleen strengthening, lung invigorating and kidney tonifying, Qi supplementing and kidney nourishing and aminophylline, particularly the first three types of drugs. The recipes for rectifying PPARgamma and antioxidant signaling pathway with intensity from high to low were recipes for lung invigorating and kidney tonifying, Qi supplementing and kidney nourishing, lung invigorating and spleen strengthening and aminophylline. Therefore, three methods for regulating and invigorating lung and kidney showed better long-term effects in regulating COPD lung inflammation signaling pathways. Specifically, recipe for lung invigorating and spleen strengthening showed a better effect in JAK/STAT and NF-kappaB pathways, while recipe for lung invigorating and kidney tonifying and Qi supplementing and kidney nourishing showed better effects in PPARgamma and antioxidant signaling pathways. In conclusion, R-value comprehensive evaluation method can evaluate the comprehensive effect of medicines and define the ranking of multiple drugs and their main targets.
Animals ; Disease Models, Animal ; Drugs, Chinese Herbal ; administration & dosage ; Humans ; Kidney ; drug effects ; physiopathology ; Lung ; drug effects ; immunology ; metabolism ; physiopathology ; NF-kappa B ; immunology ; Pulmonary Disease, Chronic Obstructive ; drug therapy ; immunology ; metabolism ; physiopathology ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Smad2 Protein ; metabolism ; Transforming Growth Factor beta1 ; metabolism

Background C57BL/6(B6) is a kind of routine mouse specie used in experimental autoimmune uveitis (EAU) research.Previous studies showed that the pathogenesis of uveitis related to inflammatory cytokines secreted by different helper T(Th) cells.However,the interaction of different Th cells in EAU is unclear.Objective This study was to investigate the dynamic changes of inflammatory factors in the spleen and serum after immunization in EAU mice.Methods Forty-four SPF B6 mice were immunized by injection of interphotoreceptor retinoid-binding protein (IRBP) and complete Freund adjuvant (CFA) emulsion via caudal vein and footpad.Indirect ophthalmoscope was used to examine the eyes 3 times per week and the inflammatory response was scored based on Thurau's criteria.In the thirty day after injection,20 model eyes were extracted and the sections of eye tissue were prepared for histopathological examination.The spleens of model mice were enucleated before injection and 2,5,10,15,20,25,30 days after injection,and reverse transcriptase PCR (RT-PCR) was used to detect the contents of interleukin-17 (IL-17) mRNA,interferon-γ (IFN-γ) mRNA,tumor necrosis factor-α (TNF-α) mRNA and IL-10 mRNA,and the contents of IL-17,IFN-γ,TNF-α and IL-10 in model serum were assayed by ELISA in 24 model mice.The experimental protocol and use of the animals were approved by Ethic Committee for Care and Use of Laboratory Animals of Shandong University of Traditional Chinese Medicine.Results Mild inflammatory response was seen in 12 days under the indirect ophthalmoscope with the scores of 0.5.The inflammatory scores peaked in 13-15 days with the scores of 1.0 and alleviated after that with the inflammatory scores of 0.5 in 30 days after injection.The histopathological score was consistent with the clinical score in the models on the 30 days.The serum IL-17 content of model mice was (0.98±0.05) ng/L before injection and increased to (51.85 ±2.42) ng/L on the fifth day,and decreased to (4.01±0.06)ng/L on the fifteen day.But,the serum IL-17 level increased to (25.00±0.94)ng/L again on the 25th day,and then lowed to (6.01 ±0.21)ng/L 30 days after injection,showing a significant elevation in comparison with that of before injection (P=0.000).The serum IFN-γ content of the model mice was (1.02±0.09)ng/L before injection and increased to (50.54±0.48) ng/L on the fifth day,and (73.21±0.12) ng/L on the tenth day,and then it declined gradually until (5.15±0.18)ng/L in the 30th day,which was still higher than that of before injection (P=0.000).After injection of IRBP+CFA,the serum TNF-α level upregulated from the second day to fifth day with the peak values (134.25±0.59)ng/L,and declined to valley on 15th day.A repeat elevation of serum TNF-α level was found on the 20th day with the values (60.54±0.62)ng/L and followed by decrease till the 30th day,which was higher than that of before injection (P=0.660).Serum IL-10 was detectable in the tenth day and peaked on the 15th day.Then a slight decrease was seen till the 30th day,compared with before injection(P =0.000).The contents of IL-10 mRNA,IL-17 mRNA,TNF-α mRNA,IFN-γmRNA in mice spleens followed the same pattern with serum levels of their proteins.Conclusions IL-17,IFN-γ,TNF-α and IL-10 are key inflammatory factors of Th1,Th2 and Th17,they present with specific changes during EAU,it confirming that IFN-γ probably play a pathogenic role in EAU,IL-17 and TNF-α levels probably associated with the chronic and recurrent procedure of uveitis,IL-10 plays an inhibit role in EAU.

Objective To retrospectively study the risk factors of aortic arch calcificationand its influence on the survival prognosis of maintenance peritoneal dialysis patients. Methods One hundred seventy-seven cases of maintenance peritoneal dialysis patients were enrolled, including 66 cases of aortic arch calcification cases. Their general dialysis data were collected for the evaluation of dialysis adequacy and residual renal function, and their chest X-rays were recorded to assess the degree of aortic arch calcification. The two variables Logistics regression was used to analyze independent risk factors of aortic arch calcification; Kaplan-Meier analysis was used to analyze the influence on prognosis of dialysis patients; and multivariate COX regression was employed to analyze independent risk factors of death in dialysis patients. Results Among the 177 selected cases of peritoneal dialysis patients, 66 cases (37.29%) presented with aortic arch calcification. Elevated serum phosphorus was an independent risk factor of aortic arch calcification (OR=54.69 ,95%CI:10.01-298.65, P<0.01). The probability of survival in patients with mild and moderate (severe) calcification of aortic arch was less than those without calcification. Moderate (severe) calcification of aortic arch was the independent risk factor of all-cause mortality and cardiovascular disease mortality, whose hazard ratios in patients with calcification were 3.779 times and 5.636 times of those in patients without calcification respectively. Conclusions Hyperphosphatemia is an independent risk factor promoting the development of calcification of aortic arch. The probability of survival in patients with mild and moderate (severe) calcification of aortic arch is less than those without calcification; moderate (severe) calcification of aortic arch is the independent risk factor of all-cause mortality and cardiovascular disease mortality.

OBJECTIVETo explore the relationship between quinone oxidoreductase1 (NQO1) gene nonsynonymous cSNP and the genetic susceptibility of esophageal cancer.

METHODSPolymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and Allele-Specific PCR (AS-PCR) were employed to assess the polymorphism of NQO1 genes both in 106 patients with esophageal cancer and control subjects matched by age, gender and origin.

RESULTSIt was shown that no C/C genotype was found at 406 of NQO1. The allelic frequency of NQO1 609T was significantly higher in patients with esophageal cancer than in the control subjects (P < 0.005) and the individuals with 609T allelic genotype of NQO1 gene were at greater risk to develop esophageal cancer (OR = 4.76, 95% CI = 1.064 - 3.397). But Individuals with mutant allele of NQO1 465 genotype did not show the rising risk of esophageal cancer.

CONCLUSIONSThe NQO1 C609T polymorphisms should likely be associated with the genetic susceptibility of esophageal cancer.

Alleles ; China ; Esophageal Neoplasms ; ethnology ; genetics ; Gene Frequency ; Genetic Predisposition to Disease ; Genotype ; Humans ; NAD(P)H Dehydrogenase (Quinone) ; genetics ; Polymorphism, Single Nucleotide