AIM: To investigate atypical protein kinase C(aPKC), extracellular signal regulated kinnase (ERK) regulating NF-E2-related factor 2 (NRF2)-?-gutamylcysteine synthetase (?-GCS) and the effect on lung of rats with chronic obstructive pulmonary disease (COPD). METHODS: The rat COPD model was established by intratracheal instillation of lipopolysaccharide twice and exposure to cigarette smoke daily. The ?-GCS activity was measured. The expression of ?-GCS mRNA in lung tissue was examined by in situ hybridization (ISH) and reverse transcription-polymerase chain reaction (RT-PCR). The protein expressions of p-aPKC?/?, p-ERK, NRF2 and ?-GCS in lung tissue were detected by immunohistochemistry (IH) and Western blotting, respectively. RESULTS: (1) The ?-GCS activity was higher in COPD group than that in control group. (2) The expression of ?-GCS mRNA in the COPD group was stronger than that in control group. ISH showed that the ?-GCS mRNA was expressed in alveolar epithelium and bronchiolar smooth muscle cell in the COPD group. (3) The protein expressions of p-aPKC, p-ERK, NRF2, ?-GCS were significantly higher than those in control group. IH showed that p-aPKC, p-ERK, NRF2, ?-GCS proteins were expressed in alveolar and bronchiolar epithelium in the COPD group. (4) There was a positive correlation between NRF2 and ?-GCS. ?-GCS mRNA, p-aPKC?/?, p-ERK were also positively correlated with NRF2. CONCLUSION: By upregulating the signal transduction of NRF2-?-GCS, the ERK and aPKC?/? may play an important role in the mechanism of COPD formation.

Nasopharyngeal carcinoma (NPC), a common head and neck neoplasm in southern China,is mostly treated with radiotherapy, however, it can not be completely cured by chemoradiotherapy. Recent investigation on NPC suggests that the nasopharyngeal carcinoma stem cells can be differentiated to heterogeneous cancer cells that have a strong ability for self renovation and further contribute to the development and progression of tumor itself which is closely related to drug resistance, recurrence and metastasis of NPC. Wnt/β-catenin, Notch, Hedgehog, NF-κB and mTOR signaling pathways play important roles in cancer stem cells. Study on nasopharyngeal carcinoma stem cells targeted therapies and its related pathways provides a new strategy for the clinical treatment of NPC.

Objective To evaluate three different Chlamydophila pneumoniae recombinant antigens for use in Chlamydophila pneumoniae serodiagnosis. Methods The recombinant plasmids pGEX6p-2/ Cpn0146,Cpn0147 and Cpn0308 were constructed and expressed as GST fusion proteins. The immunogenicity and the immunocompetence of these recombinant protein were analyzed by Western-blot and indirect ELISA. A total of 183 sera samples of patients with respiratory tract infection and 32 sera samples of patients with Chlamydia trachomatis infection were detected with indirect ELISA coated microwell plates with the purified recombinant proteins comparing with SeroCP-TM IgG ELISA kits. The positive recognition rate, sensitivity and specificity of each method were analyzed. Results GST-Cpn0146, Cpn0147 and Cpn0308 were obtained after expression and purification. The titers of the specific IgG antibodies against Cpn0146, Cpn0147 and Cpn0308 were higher than 1:6 400, 1:128 00 and 1:128 00, respectively. When the indirect ELISA was developed to detect the IgG antibody against Chlamydophila pneumoniae in 183 samples, the concordance rate between the indirect ELISA test and SeroCP-TM IgG ELISA kits were 92. 3% (Cpn0146) , 94.5% (Cpn0147) and 96.7% (Cpn0308), respectively. The recombinant Cpn0146, Cpn0147 and Cpn0308 were recognized by 71 (38.8% positive recognition rate), 75 (40.9%), and 82 (44.8%) samples, respectively. The recombinant antigen-based detection assays displayed > 97% of detection specificity and>87%of sensitivity.Condusion GST-Cpn0308 shows a better sensitivity and specificity,which suggests it could be used for developing serodiagnosis kits of Chlamydophila pneumoniae infection.

Objective To establish a new model of cardiac arrest (CA) in rats by transcutaneous electrical epicardium stimulation. Method Two acupuncture needles connected to the anode and cathode of a stimulator were transcutaneously inserted into the epicardium as electrodes. The stimulating current was steered to the epicardium and the stimulation was maintained for 3 minutes to induce CA. Cardiopulmonary resuscitation (CPR) was performed at 6 minutes after a period of nonintervention. Results The success rate of induction was 12/20 at the current intensity of 1 mA; and reached 20/20 when the current intensity was increased to 2 mA. The average time from the electrical stimulation to CA induction was (5. 10 ± 2. 81) seconds. When the electrical stimulation stopped, 18/20 rats had ventricular fibrillation and 2/20 rats had pulseless electrical activity. CPR was performed for averagely 207.4 ( ± 148.8) seconds. The restoration of spontaneous circulation was 20/20. The death rate within 4 hours after CA was 5/20, and the 72-hour survival rate was 10/20. There were only two cases of complications, a minor muscle contraction and a minor lung lobe injury. Conclusions The model of CA in rats induced by transcutaneous electrical epicardium stimulation is a stable model that requires low-intensity current and has fewer complications.

Objective:To study the effects of compound preparation of Fanshiliu on lipid metabolism and pancreatic pathological changes in type 2 diabetes mellitus(T2DM)rats with insulin resistance. Methods:After fed with high-calorie food for 8 weeks,the rats received intraperitoneal injection with low dose of streptozotocin(STZ)to induce T2DM animal model,and then the rats were randomly divided into the model group,metformin group,high and low dose group of compound preparation of Fanshiliu. The normal group was given ordinary feed. The rats were sacrified after intragastric administration for six weeks,and fasted for 12 hours after the last administration. The lipid and lipoprotein indices were detected,and pathological morphology of pancreatic tissue was observed. Results:Compared with those in the model group,TC,TG,LDL-C and FFA in each treatment group were reduced,and HDL-C was increased(P < 0. 05). Fanshiliu high dose group and metformin group had obvious hypoglycemic and lipid-lowering effects. The results of pathological morphology of pancreatic tissue under a light microscope showed that the damage was the most obvious in the model group. Compound preparation of Fanshiliu groups and metformin group showed damage in varying degrees. Conclusion:Fanshiliu preparation can effectively adjust blood lipid levels,enhance the insulin sensitivity and reduce damage in pancreas in T2DM rats.

Objective To observe the effects of Xifeng Tongnao capsules on the content of TNF-α and IL-1β in brain tissue of rats with focal cerebral ischemia-reperfusion injury. Methods A total of 120 adult SD rats were randomized into 6 groups: Xifeng Tongnao low-, middle- and high-dose groups, model control group, Buchang Naoxintong group and sham-operated group.Buchang Naoxintong group were treated with Buchang Naoxintong capsules at 0.864 g??kg-1.Xifeng Tongnao high-, middle-and low-dose groups were treated with 3.456, 1.728, and 0.864 g??kg-1 Xifeng Tongnao capsules, respectively;sham-operated group and model control group were treated with equal volume of purified water. Medications were administered intragastrically once daily for 7 days. The acute transient focal cerebral ischemia-reperfusion model was established by middle cerebral artery occlusion ( MCAO) 1 h after the final dose, and rats in the sham-operated group only received anesthesia and stripping without occlusion.All rats were sacrificed after reperfusion for 24 h, and expression levels of TNF-α and IL-1β in the brain tissue were detected by ELISA. Results TNF-αcontent in Xifeng Tongnao capsutes low-, middle-and high-dose groups were (35.34±8.95), (33.75±6.92), and (40.95±5.39) ng??L-1, respectively.IL-1β content were (1.44±0.47), (1.45± 0.23), and (1.61±0.33) ng??L-1 in low-, middle- and high-dose groups, respectively.TNF-α and IL-1β were (38.96±9.84) and (1.56±0.31) ng??L-1, respectively in Buchang Naoxintong group, (52.74±6.76) and (2.79±0.45) ng??L-1in the model control group, and (32.54±4.00) and (1.32±0.22) ng??L-1 in sham-operated group.TNF-αand IL-1βcontent were significantly higher in Buchang Naoxintong group than in sham-operated group ( P<0. 05 ) . TNF-α and IL-1β content were significantly decreased in Xifeng Tongnao high-, middle- and low-dose groups (all P<0.05). Conclusion Xifeng Tongnao capsules can protect brain tissue by supressing TNF-α and IL-1β and alleviating inflammatory injury from ischemia.

AIMTo investigate the expression and relationship of gamma-glutamylcysteine synthetase (gamma-GCS) and NF-E2-related factor2 (NRR2) in lung of rat with chronic obstructive pulmonary disease (COPD)in order to elucidate the possible important role of gamma-GCS and NRF2 in pathogenesis of COPD.

METHODSThe rat COPD model was established by intratracheal instillation of lipopolysaccharide twice and exposed to cigarette smoke daily. The gamma-GCS activity was measured, the expression of gamma-GCS mRNA in lung was examined by in situ hybridization (ISH) and reverse transcription-polymerase chain reaction (RT-PCR), the protein expressions of NRF2, gamma-GCS in lung were detected by immunohistochemical (IH) and Western blot respectively.

RESULTSThe gamma-GCS activity was higher in COPD group than that in control group. The expressions of gamma-GCS mRNA in COPD group was stronger than those in control group. ISH showed that the gamma-GCS mRNA was expressed in alveolar epithelium and bronchial smooth muscle cell in COPD. The protein expressions of NRF2, gamma-GCS were significantly higher than the control group. IH showed that NRF2, gamma-GCS proteins were expressed in alveolar and bronchial epithelium in the COPD group. There was a positive correlation between NRF2 and gamma-GCS and gamma-GCS mRNA.

CONCLUSIONNRF2 may play an important role in the mechanism of COPD oxidative stress vis up-regulation of gamma-GCS.

Animals ; Glutamate-Cysteine Ligase ; metabolism ; Lung ; metabolism ; Male ; NF-E2-Related Factor 2 ; metabolism ; Oxidative Stress ; Pulmonary Disease, Chronic Obstructive ; metabolism ; physiopathology ; Random Allocation ; Rats ; Rats, Wistar

BackgroundThe quest to look for seed cells is a hot spot of cornea transplant research in solving the problem of the lack of donor. Bone marrow mesenchymal stem cells(BMSCs) have been successfully induced into retinal ganglion cells(RGCs) in vivo,but the successful induction of BMSCs into corneal endothelial cells has not been reported.Objective This experiment was to study the transplantation of BMSCs on corneal endothelial surface using the splitting Descemet's membrane. MethodsFour healthy adult rhesus monkeys were divided into the experimental group ( 3 monkeys) and control group ( 1 monkey). Mesenchymal stem cells (MSCs) were isolated from bone marrow by density gradient centrifugation combined with adhering means. The cultured cells were identified by flow cytometry and its ability to differentiate was determined by allowing them to differentiate into adipocytes in vitro and labeled by 5-bromodeoxyuridine ( BrdU ) for subsequent identification. Corneal grafts of 7 mm in size with tearing of the Descemet' s membrane were prepared in the experimental group and control group. After labeling by 5-bromodeoxyuridine( BrdU ) ,cultured cells were transplanted onto the endothelial surface of cornea grafts in the experimental group, but no cultured cells were seeded in the graft of the control group. The corneal grafts were then sutured in situ, and were removed 1,2 or 3 months after operation to examine the distribution and connection between transplanted cells and their morphologic changes under the electron microscope. Results High purity MSCs were harvested by density gradient centrifugation combined with adhering method. Cultured cells reached confluency after 12 to 16 days, presenting with a spindle shape and parallel or swirling arrangement. Flow cytometry analysis showed that 94.26％ of cells were positive for CD29,7. 51％ for CD34 and 4. 02％ for CD45. Larger nuclei filled with plastosomes, golgiosomes and rough endoplasmic reticula were found on the graft under the transmission electron microscope( TEM ). After 3 weeks, MSCs were differentiated into adipocytes where Oil Red O staining resulted in an orange-red staining in the cytoplasm and blue staining in the nuclei. The transplanted cells attached loosely on the endothelial surface of the corneal graft and came in contact with each other in one month. The shape of the cells appeared as spindle-shaped and polygonal after 2 months and became tightly packed after 3 months. The positive cells retained the BrdU label and presented with brown nuclei. No endothelia cells grew in the cornea graft in the control group, with an absence of BrdU labeling. Conclusions Mesenchymal stem cells can be transplanted onto the corneal endothelial surface successfully and form a monolayer using the centrifugation method, and present with good survival and proliferation ability.

Objective To determine the effecting factors for diagnosis of binign or malignant in cystic lesions of pancreas(CLP).Methods One hundred twenty-six patients undergoing operations for CLP or suspected CLP in this hospital from January 1984 to June 2008 were reviewed.Patients were divided into two groups according to lesion's histological features after operation.The predictive effect of various preoperative factors on the malignant potential of CLP was evaluated.Results One hundred twenty-six patients underwent operations for suspected pancreatic cystic neoplasms.There were 89 benign and 37 malignant CLPs.The univariate analysis showed that gender, clinical symptoms(jaundice and weight loss), elevated serum CA199, and presence of one or more of three morphologic features such as solid component, nodule or septation were significantly different between benign and malignant cystic neoplasm of pancreas.The multivariate analysis indicated that imaging features and gender were independent predictors of malignancy.Conclusion In patients with suspected pancreatic cystic neoplasms, elevated serum CA199, clinical symptoms(jaundice and weight loss)and presence of suspicious morphologic features on imaging are predictors of malignant potential of CLP.Patients with a high likelihood of a potentially malignant or malignant lesion based on these three factors should undergo operation without additional investigations.

Objective To assess the clinical significance of fetal pyelectasis and its changing in utero. Methods One hundred and ninty-seven isolated pyelectasis cases were retrospective reviewed from Jan 2012 to Jul 2014.Isolated pyelectasis was defined as a renal pelvis anteroposterior diameter (RPAPD)of ≥5 mm without other fetal anomaly in second trimester.Persistent or progressive pyelectasis was defined as a RPAPD of ≥10 mm before delivery.They were divided into two groups according to the size of renal pelvis in second trimester:group A (RPAPD 5 - 10 mm)and group B (RPAPD ≥ 10 mm).As the same,there were two groups after 32 weeks of gestation:group C (RPAPD < 10 mm)and group D (RPAPD ≥ 10 mm).Results Totally 1 54 cases were followed up.There were 1 88 cases (95.4%)in group A,with 41 cases lost,141 cases (95.9%)RPAPD <10 mm,6 cases (4.1 %)RPAPD ≥10 mm before delivery.There were 9 cases (4.6%)in group B,with 2 cases lost,remained 7 cases RPAPD ≥ 10 mm before delivery. Conclusions Although most of the fetuses with RPAPD 5 - 10 mm in second trimester will remain the same or resolved before delivery,those with RPAPD ≥ 10 mm may persistent or progress.Prenatal assessment of fetal renal pelvis may provide properly consultation.