Objective To investigate the construction of compound membrane with corneal stromal cells and collagen-chitosan by tissue engineering technique and its biocompatibility.Methods Rabbit and human corneal stromal cells were separated and seeded into collagen-chitosan membrane.The compound membrane was transplanted into rabbit corneal stroma.Then the growth condition of keratocytes,the effect on normal keratocytes and degradation of compound membrane were detected by corneal confocal microscope,anterior OCT and histological and immunohistochemical methods ex vivo 1,2,4 weeks after grafting.Results The rabbit and human corneal stromal cells grown well in collagen-chitosan scaffold.The compound membrane degradated gradually after grafting.There was no necrosis and dissolvation.Corneal epithelium,stroma and endothelial cells were all normal.Conclusion Collagen-chitosan can be used as a biological scaffold for construction of corneal stroma.Corneal confocal microscopy and anterior OCT are new methods to observe the biological activity of constructed corneal stroma.

OBJECTIVE:To study the effects and mechanism of rapamycin on invasion and metastasis of cervical cancer HeLa cell. METHODS:HeLa cells were divided into control group and rapamycin low-dose,medium-dose and high-dose groups (10, 30,100 nmol/L). After treated for 48 h,cell viability was measured by MTT assay,and inhibitory rate was calculated;migration and invasion of cell was tested by Transwell assay. The expression of matrix metalloproteinase 2(MMP-2),MMP-9,Vimentin and E-cadherin,and phosphorylation of protein kinase B (Akt),mammalian target of rapamycin (mTOR) were detected by Western blot. RESULTS:Compared with control group,the inhibition rate of cell viability was increased in rapamycin groups(P<0.01);the number of invasion and metastasis cells decreased(P<0.01);the expression of MMP-2,MMP-9 and Vimentin were decreased (P<0.01 or P<0.05);the expression of E-cadherin was enhanced(P<0.01 or P<0.05);the phosphorylation of Akt and mTOR were reduced (P<0.01). CONCLUSIONS:Rapamycin could inhibit invasion and metastasis of HeLa cell via Akt/mTOR signal pathway.

Objective To investigate effect of folic acid combined with xin funing on CRP, HGF, IL-2,TNF-αof patients with cervical cancer caused by human papillomavirus.Methods 80 cases of cervical cancer patients were randomly divided into control group, 40 cases in the control group were given conventional chemotherapy, 40 cases in the experimental group were on the base of the control with folic acid combined with xin funing.CRP, HGF, TNF-α, IL-2 and T lymphocyte subsets were compared before and after the treatment.Results Compared with the control group, the serum CRP, HGF and TNF-αof the experiment group were lower(P<0.05), IL-2 levels was higher (P<0.05), CD4 +and CD4 +/CD8 +level were higher(P<0.05), level of CD8 +was lower(P<0.05) and the clinical effective rate were higher(P<0.05).Conclusion Folic acid combined with Xin Funing has important significance for the treatment of patients with cervical cancer.It is speculated that the mechanism may be to reduce the level of serum CRP and HGF in patients with cervical cancer, and to increase the level of IL-2, and to regulate immune cells.

Objective To establish the fingerprints analysis of the methanol extracts of nutmeg,and study quality uniformity of nutmeg in different areas.Methods A ZORBAX EclipseXDB-C18 (4.6 mm?150 mm,5 ?m) column was used.The mobile phase consisted of methanol-acetonitrile-water (25∶35∶40),the flow rate was 1 mL/min,the column temperature was 30 ℃,the detective wavelength was 270 nm.Dehydrodiisoeugenol was used as reference compound.Results Fingerprint of nutmeg was established,consisted of l7 common peaks.The similarity of fingerprints was over 0.9.Conclusion The fingerprints of nutmeg in different areas have no differences.This method is accurate,reliable and provides a scientific basis for the quality control of nutmeg.

Objective To investigate the effect of hydrogen-rich water on cerebral edema and aquaporin 1 (AQP1) expression in rats with traumatic brain injury (TBI).Methods Ninety male Sprague-Dawley (SD) rats were randomly divided into sham operation group,TBI model group,hydrogen-rich water treatment group (H group),with 30 rats in each group.TBI model was reproduced by weight dropping method.The skulls of rats in sham operation group underwent only craniotomy without direct hit and with bone wax sealed suture.5 mL/kg of hydrogen-rich water injection was given intraperitoneally after model reproduction in H group,and equal amount of normal saline was given in sham and TBI groups,once a day for both groups for 5 days.Six rats from each group were sacrificed at 6,12,24,48 hours and 5 days after evaluating neurological severity scores (NSS).The cerebral cortex was harvested,and the pathological changes in morphology of brain tissue were observed with light microscope.The positive expression of AQP1 in cerebral cortex was observed with immunohistochemistry by light microscopy,the AQP1 mRNA expression in cerebral cortex was determined by real-time fluorescent quantization reverse transcription-polymerase chain reaction (RT-PCR),and the AQP1 protein expression in cerebral cortex was determined by Western Blot.Results ① All rats in sham operation group had a NSS of zero at each time point.NSS of TBI group was obviously raised with time prolongation,and peaked at 24 hours followed by a lower tendency,while the score in H group was significantly lower than that of TBI group,and the difference was the most obvious at 24 hours as compared with TBI group (9.83 ± 2.78 vs.13.50± 2.42,P ＜ 0.05).② It was shown by light microscope that in the TBI group there were pathological changes in cerebral cortex,including obvious irregular arrangement of nerve cells,cerebral edema,obvious bleeding,especially at 24 hours,then the cerebral edema became vanished gradually;and the positive expression of AQP1 in the pia mater at all the time points in the TBI group was significantly increased,and it was most obvious at 24 hours.Compared with TBI group,the pathological changes at time points of 12 hours to 5 days in H group was significantly lessened,and the positive expression of AQP1 in the cerebral pia mater was reduced obviously.③ Compared with sham operation group,the mRNA and protein expressions of AQP1 in cerebral cortex in TBI group were significantly elevated,peaked at 24 hours [AQP1 mRNA (2-△△Ct):7.50±0.26 vs.1,AQP1 protein (gray value):1.986±0.110 vs.0.336±0.034,both P ＜ 0.05],then they gradually declined.The mRNA and protein expressions of AQP1 in cerebral cortex were significantly decreased after hydrogen-rich water treatment [24-hour AQP1 mRNA (2-△△Ct):5.40±0.21 vs.7.50±0.26,24-hour AQP1 protein (gray value):1.246±0.137 vs.1.986±0.110,both P ＜ 0.05].Conclusions The up-regulation of AQP1 mRNA and protein in ratst cerebral cortex after TBI perhaps participates in edema formation which might be involved in the pathophysiology of cerebral edema in TBI.Early treatment with an intraperitoneally injection of hydrogen-rich water is capable of attenuating the extent of TBI-induced up-regulation of AQP1 mRNA and protein,alleviating cerebral edema,and achieving its protective effects.

Objective: To assess the values of conventional vascular ultrasound (US) and superb micro vascular imaging (SMI) for diagnosing carotid artery stenosis in relevant patients. Methods: A total of 37 patients of extra cranial carotid stenosis (with 70 blood vessels) treated in our hospital from 2014-08 to 2015-03 were retrospectively studied. Digital subtraction angiography (DAS) examination was used as golden standard, the diagnostic efifcacies for carotid artery stenosis by US and by SMI were compared. Results: The accuracy, sensitivity and specificity for diagnosing carotid stenosis by US were 81.42%, 83.33% and 80.95%; by SMI were 91.43%, 92.16% and 89.47% respectively. Conclusion: US and SMI showed good agreement for diagnosing carotid artery stenosis, while SMI was superior to US for accurately assess the degree of carotid stenosis, it might be used as a more reliable method for evaluating carotid plaque and stenosis in relevant patients.

Background The pathogenic mechanism of thyroid associated ophthalmopathy (TAO) is still unclear,which is considered to be an autoimmune disease.It is confirmed that interleukin-17A (IL-17A) plays an important role in the occurrence and development of many autoimmune diseases.It is unclear that whether IL-17A participates in the pathogenesis of TAO.Objective This study was to explore whether IL-17A secreted by coculture system of peripheral blood mononuclear cells (PBMCs) and orbital fibroblasts (OFs) participates in the pathogenesis of TAO and its possible mechanism.Methods Periphery blood and orbital connective tissue were obtained from 12 patients with TAO and 8 patients who received prosthesis implantation for eyeball atrophy in Xiangya Hospital during April to December 2014.PBMCs were isolated by density gradient centrifugation,and OFs were cultured by explant culture method.The purity of T leukomonocyte in PBMCs was tested by flow cytometry,and OFs were identified by Giemsa staining and immunochemistry.OFs and PMBCs were incubated into 96-well plate in a 1:20 proportion to establish co-culture system.Different concentrations of phytagglutinin (PHA) (0,1.0,2.5,5.0,10.0 μg/ml) was added for 72 hours,and IL-6,IL-17A levels in the co-culture system supernatant and IL-17A receptor (IL-17RA) of the total cell membranes in the co-culture system were assayed by ELISA.The differences of IL-6,IL-17A,IL-17RA levels in co-culture system were compared between the TAO group and control group.Results The mean purity of T leukomonocyte in PBMCs was (81.10±0.21)％ in the TAO group and (80.05 ±0.38)％ in the control group respectively,with no significant difference between them(t =0.923,P＞0.05).Cultured OFs showed the positive response for Vimentin expression and Giemsa staining.After stimulated by 1.0 μg/ml PHA,the proliferation of both PBMCs and OFs were increased in the co-culture system.Apoptosis exist in PBMCs and the number of OFs decreased when PHA was higher than 1.0 μg/ml.The growth of PBMCs and OFs was faster in the TAO group than that in the control group in the same concentration of PHA.The contents of IL-6,IL-17A and IL-17RA in co-culture system were significantly different among various concentrations of PHA subgroups (IL-6:Fgroup =12.561,P=0.000;F ion =23.356,P =0.001.IL-17A:Fgroup =12.037,P =0.000;Fconcentration =19.206,P=0.000.IL-17RA:Fgroup =16.216,P=0.000;Fconcentraction =4.627,P=0.018).The production of IL-6,IL-17A and IL-17RA reached peak in both TAO group and the control group after 1.0 μg/ml PHA stimulated.However,the concentrations of IL-6,IL-17A and IL-17RA reduced with the increase of PHA concentration.The concentrations of IL-6,IL-17A and IL-17RA in co-culture system were significantly higher in the TAO group than those in the control group under the stimulation of the same concentration of PHA (all at P＜0.05).Conclusions The co-culture system of PBMCs and OFs stimulated with PHA can be the imitation of TAO pathogenesis in vitro,and PHA can amplify its immune reaction to imitate TAO pathogenic processes intuitively.The IL-6,IL-17A and IL-17RA secreted by PBMCs and induced by PHA are increased in TAO patients,implying that IL-17A participates in the pathogenesis of TAO through magnifying cellular immune response and inflammatory reaction.

Objective To investigate the effects of pioglitazone on angiotensin Ⅱ(AngⅡ) -induced proliferation and collagen typeⅠ expression of adventitial fibroblasts (AF). Methods The AFs were isolated from rat thoracic aorta. MTT colorimetry and flow cytometry were used to study the effects of pioglitazone on proliferation and cell cycles of AF. The expression of collagen typeⅠ regulated by pioglitazone was examined by the method of Western blot. Results Pioglitazone inhibited the proliferation of AF in a dose-dependent manner, and the most marked effect could be observed at the concentration of 10?10-6 mol/L for pioglitazone (P

Neurolymphomatosis(NL) is characterized by lymphomatous infiltration of the peripheral nervous system. We report a case of neurolymphomatosis(NL) which was confirmed by sural nerve biopsy. Sural nerve specimen from a 49-year-old female patient with weakness of limbs was examined with routine histochemical and immunohistochemistry staining, in which the first antibodies against CD3, CD20, CD45, CD45RO and CD68 were used. Numerous T-lymphoma cells invaded in the adipose tissue of epineurium of sural nerve. The nerve biopsy showed marked axonal degeneration of myelinated fibers. The clinical and histopathologic findings confirmed the diagnosis of neurolymphomatosis.

Objective To analyze the clinical features of systemic lupus erythematosus (SLE) com-plicated by noncirrhotic portal hypertention (NCPH),and improve the recognition of NCPH.Methods Clinical data from SLE complicated by NCPH in our hospital were retrospectively analyzed and summarized,while the related literatures were reviewed.Results Four patients diagnosed as SLE complicated by NCPH were all women.NCPH presented with the clinical features of portal hypertension with normal or slightly elevated transaminase.Anticardiolipin (ACL) antibodies were positive in 2 patients.Two patients underwent liver needle biopsy,showing nodular regenerative hyperplasia,of which,one with liver portal fibrosis.The treatment strategy was managing the primary disorder and controling of portal hypertention in four patients.Twenty-two cases of SLE complicated by NCPH were reviewed and analyzed,including 18 cases from related literatures and our 4 cases.Among the 22 cases,the mean time between the diagnosis of SLE and NCPH was eight years,of which one patient with NCPH before SLE,one diagnosed at the same time and the rest with NCPH after SLE.19％ (4/21) of patients presented with Raynaud's phenomenon and 18％ (4/22) complicated by pulmonary hypertension.In serological tests,patients presented with positive ACL anti-bodies [33％(7/21)] and anti-dsDNA [48％(10/21)],as well as increased IgG and γ-Globulin [38％(8/21)].Liver needle biopsy showed nodular regenerative hyperplasia or liver portal fibrosis with the prevalence of 80％ (16/20) and 25％ (5/20),respectively.Conclusion SLE complicated by NCPH is very rare clinically and is easily being misdiagnosed without obvious symptoms and signs in the early stage.Positive ACL antibodies and Raynaud's phenomenon maybe be closely related to SLE complicated by NCPH.