1.Study of bone marrow srt omal cells on a three-dimensional nano-zirconia porous scaffold for bone tissue engineering
Yinglan ZHU ; Ruiqiao ZHU ; Xiaolei SHI
Journal of Medical Postgraduates 2014;(6):564-567
Objective Nowadays, in the field of bone tissue engineering , searching for a kind of scaffold , which is both strong enough and biocompatible , is still a hot topic .Objective of this study is to evaluate the three-dimensional nano-zirconia porous scaffold′s effect on bone marrow stromal cells early attachment and proliferation . Methods Preparing nano-zirconia porous scaffold and testing its porosity and its mechanical strength .Hemolysis experiment was taken to evaluate its biosafety .Canine bone marrow stromal cells ( BM-SCs) were extracted from an adult beagle and cultured in vitro.The third generation cells were seeded onto the scaffold and induced to os-teoblasts under condition medium .Cells early attachment on scaffold was observed by SEM .MTT and ALP tests were taken to evaluate cells proliferation and osteogenesis within two weeks . Results Nano-zirconia porous scaffold in our study presented a porous structure with a well connected inner .Its porosity and compressive strength were (92.667 ±0.324)%and 4.38 MPa, respectively.Hemolysis rate was 2.19%, which was demonstrated to be safe to human body .Canine BMSCs could colonize the surface of the materials , bridge macro-poros-ity and proliferate on the scaffold .A values taken by the method of MTT showed that cells could proliferate quickly on the scaffold after 3, 6, 9, 12 d (A values were 0.604 ±0.059, 1.215 ±0.019, 1.385 ± 0.254, and 1.506 ±0.050), comparing with that on the first day (A value was 0.261 ±0.034).Statistical difference was found (P <0.01) .The ALP level raised up significantly on day 7 and 14 ( A val-ues were 0.032 ±0.002 and 0.062 ±0.007), comparing with that on the first day (A value was 0.016 ±0.003).Statistical difference was found (P<0.01). Conclusion The three-dimensional nano-ZrO2 porous scaffold in our study presented satisfactory biocompati-bility.It could offer a microenvironment for osteoblasts growth and secreted osteogenesis marker in vitro .
2.The expression of NOD2 and TLR9 in gastric carcinoma and precancerous lesions
Bai WEI ; Youqing ZHU ; Xiaoyan TANG ; Wei HOU ; Yinglan ZHAO
Chinese Journal of General Surgery 2008;23(11):866-868
Objective To investigate the expression of NOD2 and TLR9 in gastric carcinoma and precancerous lesions and their clinical significance. Methods The expression of NOD2 and TLR9 was measured by immunohistochemical staining (S-P method) in 84 patients with atrophic gastritis, 48 eases with gastric ulcer, 80 cases with gastric adenocarcinoma and 40 eases with superficial gastritis. Results NOD2 and TLR9 expression was up-regnlated in superficial gastritis, atrophic gastritis, gastric ulcer and gastric adenocarcinoma. The positive expression rate of NOD2 was 35% ,21% ,33% ,40% respectively, and the positive expression rate of TLR9 were 15%, 12% ,21% ,22% respectively. The expression of NOD2 and TLR9 in Hp complicated atrophic gastritis, gastric carcinoma was significantly higher than that of lip not complicated disease entities ( P < 0. 05 ). Conclusions The expression of NOD2 and TLR9 may be valuable index for predicting the development of gastric mucosal damage from superficial to atrophic gastritis, gastric ulcer and gastric carcinoma.
3.Polyvinyl butyral improves the properties of 3-dimensional nano-zirconia porous scaffolds for bone tissue engineering
Ruiqiao ZHU ; Juan MA ; Yinglan ZHU ; Xiaolei SHI ; Zhen DONG ; Lei JIN
Journal of Medical Postgraduates 2015;(5):521-525
Objective The nano-zirconia scaffolds we previously prepared had a good 3-dimensional ( 3D ) connectivity but did not achieve the ideal sintering rate and compressive strength .The objective of this study was to explore the enhancing effect of polyvinyl butyral ( PVB) as a dispersant on the compressive strength of 3D nano-zirconia porous scaffolds for bone tissue engineering . Methods We prepared the slurry containing different concentrations of PVB and ana-lyzed the improving effect of PVB on the mechanical properties of the scaffolds by sediment experiment , compressive strength test and scan-ning electron microscopy . Results The sediment experiment showed
no significant stratification in the slurry with 0.2wt%PVB, white suspension in the upper layer and white precipitate in the lower layer , with a significantly higher compressive strength of the scaffold ([0.324 ±0.030] MPa) than that of the scaffold prepared by adding other concentrations of PVB to the slurry (P <0.01).And the compressive strength of the scaffold constructed by adding no dispersant ([0.109 ±0.021] MPa) was remarkably lower than that of the scaffold constructed by adding PVB to the slurry (P<0.05).Scanning electron microscopy demonstrated that the scaffold prepared by adding 0.2wt%PVB to the slurry had a complete porous structure with the fewest and most sparsely distributed surface cracks as compared with other PVB concentration groups . Conclusion PVB can signifi-cantly improve the stability of zirconia slurry , enhance the compressive strength of the nano-zirconia porous scaffold , and make the scaf-fold more applicable to bone tissue engineering .
4.Construction and identification of a lentiviral vector of RNA interference containing human Notch-1 gene.
Qing-qing ZHANG ; Senlin ZHANG ; Yinglan ZHU ; Zhen DONG ; Gang CAO ; Wei CHEN
West China Journal of Stomatology 2014;32(3):267-272
OBJECTIVETo construct and identify a lentiviral vector of RNA interference targeting human Notch-1 gene.
METHODSTo determine the Notch-1 gene sequences, three RNAi target sequences (shRNA1-3) were designed in accordance with the RNAi sequence design principles and cloned into the lentiviral vector pLenOR-THM by endonuclease BamH I restriction, EcoR I double digestion, and T4 DNA-ligase ligation. After the transformation into competent DH5alpha bacteria, the candidate clones were identified by Kpn I and EcoR I double digestion and DNA sequencing. The recombinant and three packaging plasmids were co-transfected into human embryonic kidney cell line 293T cells by lipofectamine to produce the lentiviral particles. The viral titer was determined. The 293T cells were infected by the lentiviral particles obtained, and transfection efficiency was assessed using a fluorescent microscope. The lentiviral vector particles were also transfected into ACC-M cells. The Notch-1 expression in the transfected cells was assayed by quantitative reverse transcription polymerase chain reaction (QRT-PCR) and Western blot analysis.
RESULTSThe lentiviral RNAi vector pLenOR-THM-Notchl for Notch-1 gene was constructed successfully. Strong green fluorescence was observed in the 293T cells under fluorescent microscope after co-transfection of the cells with the four plasmids of lentiviral vector. The virus in the supernatant reached a titer of 5.8 x 10(8) TU x mL(-1). The transfection efficiency of the collected virus exceeded 90% in 293T cells with 1 as a multiplicity of infection. The third lentiviral vector was found to significantly inhibit the Notch-1 expression at the mRNA and protein levels.
CONCLUSIONThe lentiviral RNAi vector of Notch-1 has been successfully constructed and identified.
Cell Line ; Genetic Vectors ; Humans ; Lentivirus ; Plasmids ; RNA Interference ; RNA, Messenger ; RNA, Small Interfering ; Receptor, Notch1 ; Transfection
5.Changes in serum homocysteine and its correlation with altitude, folacin and high-sensitivity C-reactive protein in Tibetan patients with mild-to-moderate Alzheimer's disease at different altitudes
Xin ZHONG ; Yinglan LI ; Can DU ; Guofeng LI ; Hongjuan LI ; Aiqin ZHU
Chinese Journal of Geriatrics 2016;35(9):934-938
Objective To study changes in serum homocysteine(Hcy) and its correlation with serum levels of folic acid and high sensitivity C-reactive protein(hs-CRP)in Tibetan patients with mildto-moderate Alzheimer's disease at various high altitude areas,so as to direct the clinical diagnosis and treatment of AD in plateau hypoxia environment Method 108 cases were divided into four groups:23 AD Tibetan patients at middle altitude(AD/middle altitude group)and 23 healthy Tibetan subjects (healthy/middle altitude group) in Qinghai-Tibet Plateau Xining region,altitude at 2,260 m,31 AD Tibetan patients (AD/high altitude group)and 31 healthy Tibetan elderly subjects (healthy/high altitude group)in Yushu region at altitude of 3,800 m.Among the total study subjects,half are males,aged from 60 to 85 years.The levels of serum Hcy,Vitamin B12 and folic acid(FA)were measured by the Fluorescence Polarization Immunoassay(FPIA).Serum hs-CRP,triglyceride(TG)and low density lipoprotein cholesterol(LDL-C)were measured by automatic biochemistry analyzer.Correlation of Hcy with FA and hs-CRP was analyzed.Result Both high altitude and middle altitude group showed the levels of Hcy and hs-CRP were significantly higher in AD Tibetan patients than in healthy control at the same altitude(allP<0.05).The levels of Hcy,LDL-C and hs-CRP of subjects were higherat high altitude than at middle altitude(P<0.05).In contrast,folic acid levels in AD and control groups were lower at the high altitude than at middle altitude(P<0.05).The levels of vitamin B12 and TG were not significantly different among all four groups.Multiple logistic regression analysis showed that altitude,folacin and hs-CRP were the risk factors for Hcy in patients with AD at plateau(OR =0.351,2.794,3.021,P=0.045,0.037,0.016).Conclusion Along with increased altitude,serum level of Hcy is significantly increased in AD Tibetan patients living in high altitude area.High altitude,high hs-CRP and lower folacin may be the risk factors for hyper-homocysteine in AD Tibetan patients with high altitude,and their combined effects are involved in the occurrence and development of AD.