Objective To compare the effects of three nickel-titanium file systems,ProTaper Next,TruNatomy,and VDW.RO-TATE,on the morphology of the Danger Zone of the mesial buccal root of mandibular first molar after root canal preparation using Mi-cro-CT and 3D printing technology.Methods 3D-printed mandibular first molars were selected and designed for purpose.They were randomly divided into three groups according to the used NiTi files(n=20).Micro-CT was used to scan the cross-sectional images of the Danger Zone 2 mm below the furcation of the mesial buccal root before and after root canal preparation.The changes in the root ca-nal wall thickness,root canal volume,surface area,cross-sectional area,and root canal transportation in the Danger Zone were ob-served.Statistical analysis was performed using one-way ANOVA(P＜0.05).Results Statistically significant differences were observed before and after root canal preparation in the Danger Zone among the three groups(P＜0.05).Among the three groups,the PTN files caused the largest change before and after preparation,followed by VDW files,and the TRU files had the smallest change.Conclusion The study highlights TruNatomy's conservative shaping capacity,advocating its use in minimally invasive endodontics,whereas Pro-Taper Next may be reserved for cases requiring aggressive canal preparation.

Objective To compare the effects of three nickel-titanium file systems,ProTaper Next,TruNatomy,and VDW.RO-TATE,on the morphology of the Danger Zone of the mesial buccal root of mandibular first molar after root canal preparation using Mi-cro-CT and 3D printing technology.Methods 3D-printed mandibular first molars were selected and designed for purpose.They were randomly divided into three groups according to the used NiTi files(n=20).Micro-CT was used to scan the cross-sectional images of the Danger Zone 2 mm below the furcation of the mesial buccal root before and after root canal preparation.The changes in the root ca-nal wall thickness,root canal volume,surface area,cross-sectional area,and root canal transportation in the Danger Zone were ob-served.Statistical analysis was performed using one-way ANOVA(P＜0.05).Results Statistically significant differences were observed before and after root canal preparation in the Danger Zone among the three groups(P＜0.05).Among the three groups,the PTN files caused the largest change before and after preparation,followed by VDW files,and the TRU files had the smallest change.Conclusion The study highlights TruNatomy's conservative shaping capacity,advocating its use in minimally invasive endodontics,whereas Pro-Taper Next may be reserved for cases requiring aggressive canal preparation.

Objective To determine the function of conserved amino acids in the fusion-promoting domain of Newcastle disease virus (NDV) hemagglutinin-neuraminidase (HN) protein,clearly understanding mechanism of cell fusion.MethodsUsing a PCR-based site-directed mutagenesis method and the method of homology recombination occurred in vivo to change six conservative amino acids into alanine respectively.Wild type (WT) and all mutant HN proteins were exepressed in BHK-21 cells by the vacciniaT7 RNA polymerase expression system.The amount of each HN protein at the cell surface was determined by fluorescence-activated cell sorter (FACS).Cell fusion efficiency,hemadsorption activity (or receptor binding activity) and neuraminidase activity were determined.Results There was no statistic difference of cell surface expression among WT and each mutant HN protein ( P＜0.05 ).Cell fusion efficiency of each mutant protein decreased to some extent,especially 1103A decreased to 14.2％ in head.Hemadsorption activity of mutant proteins were reduced in different extent,the maximum reduction of which was also 1103A,28.2％ of wt NDV HN.There was different neuraminidase activity among each mutant HN protein.L74A increased slightly to 118.6％.L110A decreased most to 5.2％.I103A decreased second most to 5.7％.Conclusion Conserved amino acids in fusion-promoting domain of NDV HN played an important role in cell fusion.I103 was identified as a key amino acid in this domain.

Objective To study the influence of 4-aminopyridine(4-AP)on proliferation,production,and apoptosis through inhibiting voltage-gated K+channel(Kv)in ovarian luteinized granulosa cells.nethods Ovarian luteinized granulosa cells were recovered from 25 women with regular menses who underwent in vitro fertilization programme.The cultured granulosa cells were divided into 4 groups:blank group,4-AP treated group,human chorionic gonadotropin(hCG)-induced group and hCG+4-AP cotreated group.The final concentrations of hCG and 4-AP were 1250 U/L and 5 nmol/L respectively.The progesterone production WaS detected by the chemoluminescence method.The expression of Kv mRNA on human ovarian luteinized granulosa cell was detected by RT.PCR The influence on the early apoptosis of gTanulosa cells bv 4-AP was observed by flow eytometry.Cellular caSpage-3 activities were observed with colorimetric method and the inhibition of the cell proliferation was studied using methyl thiazolyl tetrazolium(MTT)method.Results(1)Kv mRNA wag expressed in granulosa cell.(2)The progesterone production64),(206±32),(1991±172)and(763±79)nmol/L,respectively after24 hours culture.Exposure of the(3)The flow cytometry analysis and the cellular caapase-3 A405 showed that 4-AP increased the percentage ofearly phase apoptosis(P<0.01):4-AP treated group VS blank group[(40±5)%and 0.049 ±0.009]VS[(17±4)% and 0.029±0.008],hCG+4-AP CO-treated group VS hCG-induced group[(25±4)%and0.039 ±0.0081 VS[(15±3)%and 0.022 ±0.007].(4)24 hours after treated with 4-AP and hCG,theinhibitory rate of cultured granulosa cells of 4-AP treated group was higher than the blank group(19.7%VS0).and that of hCG+4-AP co-treated group was obviously higher than hCG-induced group(34.6% VS O,P<0.01).Conclusions The voltage-gated K+ channels expressed by ovarian luteinized granulosa cellplay an important role in cell proliferation,production,and apoptosis.4-AP may inhibit differentiation ofprogesterone in granulosa ceHs through the inhibition of proliferation and induction of apoptosis.

Objective To investigate the effect of Desert Hedgehog on direct differentiation of neural progenitor cells(NPCs) cultured from embryonic mesencephalon in the rats.Methods We infected DHH into COS7,NIH/3T3 and 9L cells,and detected the expression of DHH in the cells with immunofluorescence,real-time PCR and Western blot.All of the three cells were co-cultured with NPCs isolated from ventral mesencephalon in embryonic SD rats(E13.5) respectively.Immunoreactivities of tyrosine hydroxylase(TH) was detected by immunocytochemistry after 10 days.Results The expression of DHH in COS7,NIH/3T3 and 9L cells was remarkably detected,but few TH-positive cells were found in the three co-cultral systems at the 10th day.Conclusion The protein derived from DHH itself does not show any inductive effect on the differentiation of NPCs to the dopaminergic neurons in vitro.

Objective To observe whether the expression of Annexin A5(ANXA5) changes in human uterine cervical squamous cell carcinomas(UCSCC).Methods 25 UCSCC tissues and 15 normal human uterine cervical tissues were collected.Each sample was lysed in lysis buffer.Whole protein of the supernatant was estimated by BCA-100 Protein Quantitative Analysis Kit.The expressions of ANXA5 in UCSCCs and normal uterine cervical tissues were detected respectively with Western blotting.To further ensure the expression of ANXA5 in UCSCCs,another 45 UCSCC specimens and 20 normal cervical tissues were collected.ANXA5 expression was detected by in situ hybridization and immunohistochemistry respectively.Results The staining of ANXA5 band with Western blotting in UCSCCs was much heavier than that in normal uterine cervical tissues and the expression of ANXA5 was found much higher in UCSCCs by immunohistochemistry and in situ hybridization.Conclusion Expression of ANXA5 was up-regulated in human UCSCCs.There's some relationship between uterine cervical squamous cell carcinoma and ANXA5 protein.

Objective In order to investigate the roles of hIGF\|1 in treatment of diabetes mellitus and diabetic syndromes,the gene of human insulin like growth factor type Ⅰ(IGF\|1) was cloned and constructed into eukaryotic expression vector,then the expression and activity were determined. Methods Total cellular RNA of human fetal liver was abstracted and the RT\|PCR amplification of the cDNA fragment was performed.The fragment was cloned into pUCM\|T vector and sequenced.The eukaryiotic expression vector was recombined and transfected into fibroblast cell line,COS\|7.The expression of hIGF\|1 was examined by in situ hybridization and immunohistochemistry.The effect of hIGF\|1 on cultured islet cells was observed by glucose\|stimulated insulin release assay. Results The cDNA fragment of 710bp with additional Kozak sequence was amplified by RT\|PCR.Eukaryiotic expression vector pCI\|neo/hIGF\|1 was constructed and IGF\|1 gene expressed in COS\|7.The biological activity of hIGF\|1 was proved by increasing inslin secretion from islet cells.Conclusion\ The newly constructed vector,pCI\|neo/hIGF\|1 could be transfected into COS7 cells and its expressed product showed to have the biology activity of hIGF\|1.\;[

Objective In order to investigate the effect of nutrition and induction of Sertoli cell on cultured neural stem cells(NSCs) from embryonic mesemcephalon in the rat. Methods Sertoli cells from the testis and the neural precursors(NPs) from the ventral mesencephalon in embryonic SD rat(E13) were co-cultured.The detections of immunoreactivities of nestin,?-Ⅲ tubulin and tyrosine hydroxylase(TH) were made by immunocytochemistry after 5, 7, 10 and 14 days in vitro. Results The number of NPs with nestin-positive was decreasing along the time of the co-culture,while the ?-Ⅲ tubulin-positive cells and TH-positive cells were increasing.The number of ?-Ⅲ tubulin-positive cells was quite high at the 7th day and the TH-positive cells were at the highest level at the 10th day in vitro.At any stage,the numbers of TH-positive cells were higher in the co-culture,compared with those in the mono-culture.Conclusion The neural precursors from the ventral mesencephalon in embryonic rat can be directly induced to differentiate into the dopaminergic progenitors by co-cultured with the Sertoli cells.