Objective To establish an effective method of isolating and culturing circulating fibrocytes from human peripheral blood and study the relationship between the expression specific molecule markers expression and the morphological characteristics. Methods Total peripheral blood leukocytes were isolated from human peripheral blood by being centrifuged over Ficoll-Paque and cultured in DMEM supplemented with 20% fetal calf serum.Adhered cells were detected with immunocytochemistry,FCM and electron microscopy.The collagen synthesis was studied by measurement of the hydroxyproline concentration in the medium with chemical method. Results After 9 days cultue in vitro,CD34,CD45 and collagen Ⅰ staining was positive and 83.5% of these cells secreted collagen Ⅰ detected by FCM.Electron microscopy of circulating fibrocytes showed morphological characteristics of fibroblasts.The hydroxyproline concentration in the medium was 11.17%mg/L,which was statistically and significantly different compared with 8.07mg/L in the control medium(P

All kinds of primary and secondary kidney disease finally result in renal fibrosis.This review summarized cytological mechanism and the advancement in TCM research on some essential cells(such as mesangial cells,glomerular capillary endothelial cells,podocyte,renal tubular epithelial cells and fibroblasts)in renal fibrosis from the perspective of cytology.

The bioactive principles contained in blueberries (Vaccinium) are various kind of anthocyanins (anthocyanidins, or phenolic aglycone, conjugated with sugar), chlorogenic acid, flavonids, alpha-linolenic acid, pterostilbene, resveratrol, and vitamins. After oral administration, anthocyanins can pass through blood-brain barrier and thus appear in various organs and brain. Improve visual function by increasing rhodopsin regeneration and ocular health is the earliest reported bioactivities of anthocyanin. Recent studies demonstrated the benefit of blueberries to prevent the age-related chronic diseases such as cancer, diabeties, hyperlipidemia, hypertension, neurodegeneration, obesity, and osteoporosis through its apoptosis, antioxidant, antiinflammation, and antiangiogenesis effects. Blueberries can eradicate microorganisms for the prevention of symptomatic urinary tract infections in women. Thus, blueberries are recognized as one of the most nutritious foods and cultivated worldwide. However, how to prolong the shelving time of fresh fruit, well utilize the leaf and stem to isolate the bioactive chemicals, improve quality consistency of juicy and dry products, all should be further concerned.

Objective: To explore the effects of 17 β-estradiol (E2) and 2-methoxyestradiol (2ME) on endothelium-1/nitric oxide (ET-1/NO) cascade in experimental rats with hypoxic pulmonary hypertension. Methods: A total of 48 female SD rats were randomly divided into 6 groups:①Sham operation group,②Ovariectomy (OVX) group,③Hypoxia group,④OVX+hypoxia group,⑤OVX+hypoxia+E2 group, the rats received subcutaneous E2 at 20μg/(kg?d) and⑥OVX+hypoxia+2ME group, the rats received subcutaneous 2ME at 240μg/(kg?d).n=8 in each group. Blood levels of ET-1, NO, eNOS activity and the expressions of pulmonary tissue endothelium A receptor (ETAR), ETBR and eNOS were compared among different groups. Results: Compared with Sham operation group, Hypoxia and OVX+hypoxia groups showed small pulmonary artery thickening with lumen narrowing, increased mean pulmonary arterial pressure (mPAP), allP<0.01; the above morphological and mPAP changes were reduced by E2 and 2ME intervention. Compared with Sham operation group, OVX and Hypoxia groups had increased blood ET-1 and pulmonary mRNA, protein expressions of ETAR, decreased pulmonary ETBR, all P<0.01; the above changes were more obvious in OVX+hypoxia group; E2 and 2ME intervention reduced blood ET-1 and pulmonary ETAR expression, but they were still higher than Sham operation group, meanwhile, ETBR expression was elevated, but it was still lower than Sham operation group, allP<0.01; blood ET-1 was lower in OVX+hypoxia+2ME group than OVX+hypoxia+E2 group,P<0.05. Compared with Sham operation group, OVX group had decreased pulmonary eNOS protein expression,P<0.01; Hypoxia group had decreased blood NO and pulmonary eNOS protein expression,P<0.05 orP<0.01; OVX+hypoxia group had decreased blood NO, eNOS activity and decreased pulmonary mRNA and protein expressions of eNOS, allP<0.01; E2 and 2ME intervention elevated the above indexes,P<0.05 orP<0.01, but they were still lower than Sham operation group, allP<0.05. Conclusion: E2 and 2ME could decrease blood ET-1 and pulmonary ETAR expression, increase pulmonary ETBR expression; elevate blood NO, eNOS activity and pulmonary eNOS expression. E2 and 2ME may partially reverse pulmonary hypertension via improving ET-1/NO cascade in experimental rats.

Objective To investigate the pharmacokinetic difference between the combination of cyclophosphamide and cisplatin and single medication.Methods High performance liquid chromatography (HPLC) was used to detect the pharmacokinetic parameters and distribution parameters in tissue of cisplatin and cyclophosphamide.Results The pharmacokinetic curve of the CP regimen showed two-compartment model,and there was no significant difference between the absorption half life of the combination of the two drugs (0.56±0.02,0.72±0.02) and that of single medication (0.92±0.02,1.16±0.12) (t =2.091,t =2.379,both P ＞ 0.05).While the elimination half life of the combination (0.56±0.02,0.72±0.02) was significantly longer compared with that of single medication (0.92±0.02,1.16±0.12) and there were significant differences (t =5.796,t =6.213,both P ＜ 0.05).There was no significant difference of MRT in heart between the disturbution of the combination of the two drugs and that of single medication (t =2.231,t =2.096,both P ＞ 0.05).While there was significantly longer compared with that of single medication and there were significant differences in lung and liver (t =5.976,t =6.270,both P ＜ 0.05).Conclusion When used in combination,the elimination of the drugs is slower,which suggests that the interval and dosage of the drugs should be adjusted to their pharmacokinetic parameters in clinical use to improve effects and lower side effects.

Objective To express and purify pET-p53 fusion protein and investigate the effects of the protein on the proliferation of human leukemia cell line K562. Methods The fragment of human wild type p53 cDNA was amplified by PCR and the expression plasmid of pET-p53 was constructed. Recombinant plasmids were transformed into E.coli BL21,then induced by IPTG at 1mmol/L. The protein was purified by the column of Ni-NAT and analyzed by SDS-PAGE. After treated with purified P53 protein with different concentrations,the proliferation of K562 was tested by MTT assay, clone formation and FCM. Results Prokaryotic expression vectors of pET-p53 were constructed correctly. pET-p53 fusion protein was successfully expressed and purified. With its increasing concentrations, P53 protein reversed its effect on K562 significantly. The ratio of inhibition had linear relation to the concentration of P53 protein when the concentration of the protein was between 0.1?g/ml and 100?g/ml. And its IC50 was 6.74?g/ml. Conclusion The obtained pET-p53 fusion protein might induce the leukemia cell line K562 apoptosis.

Objective To investigate the influence of Wnt-3a signaling on aggregative growth of dermal papilla cells.Methods Recombinant adenovirus pAdEasy-1/Wnt-3a was constructed at first,and used to transfect cultured dermal papilla cells(DPCs).Then,the growth pattem of DPCs was observed by inverse microscopy;laser confocal microscopy and Western blot were utilized to detect the expressions of Wnt-3a,β-catenin and versican in low-passage DPCs,high-passage DPCs and transfected high-passage DPCs.Results The over-expression of Wnt-3a protein could effectively induce the aggregative growth of DPCs.Laser confocal microscopy showed that the fluorescence intensity of β-catenin and versican increased from 34.16±9.62 and 18.81±9.59 in high-passage DPCs to 87.35±17.28 and 92.18±18.39 in transfected high-passage DPCs(t=11.98,17.88,both P＜0.0 1),respectively.Also,Western blot proved a significant increase of expression intensity of β-catenin(15.27±2.17 vs 60.09±7.24,t=10.10,P＜0.01)and versican(19.32±2.46 vs 58.46±2.78,t=20.63,P＜0.01)in transfected high-passage DPCs compared with untransfected high-passage DPCS.ConclusionsWnt-3a signaling plays an important role in modulating aggregative growth of DPCs,which is mainly through the classical pathway with β-catenin as the key protein.As an important downstream gene of Wnt signaling,vesican is an indispensable part for the modulation of aggregative growth of DPCs.

Objective:To investigate the effect and mechanism of transcription factor Foxp 3 on the proliferation and cell cycle of human lung adenocarcinoma cell line A 549.Methods:We knocked down the expression of Foxp 3 using siRNA.Foxp3 inhibition was detected by RT-PCR.Cell proliferation was detected by MTT.Cell cycle of A549 cells were detected by flow cytometry after the transfection of siRNA.Cell cycle-related checkpoint genes were filtered by RT-PCR.The regulation of Foxp3 on cell cycle-related checkpoint genes were detected by immunofluorescence and dual -luciferase reporter assay system.Results: The proliferation of A549 cells were inhibited after silencing Foxp3,and A549 cells were arrested in G0/G1 cycle.G1/S cycle checkpoint gene CCND1 was down regulated.Mechanism research show that Foxp 3 can regulate the expression of CCND 1 directly.Conclusion: Foxp3 can promote the proliferation of A549 cell line by up regulating G 1/S cycle checkpoint gene CCND 1.This provides a new target for the therapeutic targets of lung adenocarcinoma.

Objective To compare 3 kinds of preoperative scoring systems used to predict 30-day mortality in a hip fracture population,Physiological and Operative Severity Score for Enumeration of Mortality and Morbidity (POSSUM),Portsmouth modified POSSUM (P-POSSUM) and Estimation of Physiologic Ability and Surgical Stress (E-PASS).Methods A retrospective study was conducted to analyze the aged 654 patients who had undergone surgery for femoral intertrochanteric fracture or femoral neck fracture from January 2010 through June 2014 at our hospital.They were 225 men and 429 women,60 to 103 years of age (average,71.7 years).There were 363 femoral intertrochanteric fractures and 291 femoral neck fractures.Closed reduction and intramedullary nailing was performed in 363 cases,artificial dipolar replacement in 242 cases,and total hip replacement in 49 cases.POSSUM,P-POSSUM and E-PASS scoring systems were used to predict the 30-day mortality.The discrepancy between the predictive risk and the actual observation was analyzed.Results Of the 654 patients,25 died within 30 days after operation.According to POSSUM scoring system,the predictability (observed deaths/POSSUM deaths) was 0.30,showing a significant difference between the predictive deaths and the actual deaths(x2=34.840,P=0.009).According to P-POSSUM and E-PASS scoring systems,the predictability was respectively 0.83 and 1.04,demonstrating no significant differences between the predictive deaths and the actual deaths(P ＞ 0.05).Conclusion P-POSSUM and E-PASS scoring systems may predict 30-day mortality more accurately than POSSUM scoring system in elderly patients undergoing hip fracture surgery.

The tuberous sclerosis complex (TSC) in children is also named Bourneville disease,the 3 main symptoms namely epilepsy,mental retardation and facial angiofibromas.It is an autosomal dominant disease.It is an important cause of epilepsy,skin disease,and renal and pulmonary disease in children and adults.The appropriate therapy and prognosis for TSC patients are often different than that for individuals with epilepsy,renal tumors,or interstitial lung disease from other causes.In recent years,certain progress has been made in management of tuberous sclerosis,inhibitors of the mammalian target of rapamycin (mTOR) have demonstrated regression of astrocytomas,angiofibromas,and angiomyoliomas,as well as improved pulmonary function in persons with TSC.This article reviews the current therapeutic recommendations for medical and surgical management of neurologic,renal,and pulmonary manifestations of TSC.