5.The study of biological characteristics of myoblasts co-cultured with Schwann cells
Hua LIAO ; Lei YU ; Shao-Hu XIONG ; Al ET
Chinese Journal of Orthopaedics 2000;0(11):-
Objective In order to understand the relationship between myoblast and Schwann cell,our purpose was to investigate the effects of biological characters of myoblasts co-cul tured with Schwann cells and pro vide the basic theory for constructing artificial muscle involving artificial nerve.Methods After sterilizing with iodine tincture and alcohol,the brachial plexus,sciatic nerve and triceps surae muscle of neonatal SD rat were harvested and peeled off their membranes,blood vessels and fat tissues under operating microscope thor oughly.The nerve and muscle tissues were cut in pieces by microscis sors,and then digested and isolated by collagenase and pancreatin in20and15minutes respectively.DMEM medium was employed to culture my oblasts and Schwann cells.After co-culturing myoblasts of rats with Schwann cells in vitro,the morphological characteristics and the growth condition of two cells were observed under inverted phase contrast mi croscope,the effects of Schwann cells secretion for proliferation of myoblasts were detected by 3 HTdR iso topic tracing and expressed by disintegration per minute(DPM),formation rate of myotubes was counted under micro scope and statistic data was analyzed,the functional differentiation degree of my oblasts affected by Schwann cells was analysed by?-sarcomeric actin immunohistochemistry(SABC)and imaging analysis tech nique.Results Co-cultured myoblasts proliferated,and myotubes ap peared earlier.Comparing with sole-cultured my oblasts,the shape of myobutes from co-cultured myoblasts tended to be elongating and robust.The value of DPM far-exceeded the control group,and reached its peak of 2500(just800for con-trol group).The positive cells of ?-sarcomeric actin appeared in brown red color.However,syn thesis and excre tion of a-sarcome ric actin in co-cultured myoblasts were much greater than control group,and the gray ash value between two groups was of a significant difference.Conclusion Primary rat myoblasts co-cultured with Schwann cells in vitro is beneficial in regulating its the growth,proliferation and the differentiation.
7.Correlation between invasion and metastasis of gastric cancer and serum angiopoietin-2
Shao-ping LIU ; Chun-hua FANG ; Li-qun YU ; Xian-feng ZHANG ; Ya-hua HU
Chinese Journal of Postgraduates of Medicine 2011;34(28):31-34
Objective To study the clinical significance of the serum angiopoietin-2(Ang-2) in the diagnosis,recurrence,invasion and metastasis of gastric cancer.Methods The serum Ang-2 and carcino-embryonic antigen (CEA) levels in 158 patients with gastric cancer (gastric cancer group) and 30 normal controls(control group) were measured by enzyme linked immunosorbent assay(ELISA) technique respectively.The serum Ang-2 and CEA levels were also measured 2 weeks after operation in gastric cancer group and reexamined in the recurred gastric cancer patients in 2 years after operation (recurred and metastasis group).The correlation between the serum Ang-2 level and pathologic c haracterization of gastric cancer was evaluated.Results The serum Ang-2 and CEA levels in gastric cancer group [ (331.8 ±64.3),(42.6 ±37.3)μg/L] and recurred and metastasis group [(318.7 ±72.9),(40.5 ±36.7)μg/L] were significantly higher than those in control group [ (187.4 ± 32.7),(4.2 ± 3.1 )μ g/L] (P < 0.01 ),and the serum Ang-2 level 2 weeks after operation [ (211.6 ± 75.1 ) μ g/L ] was significantly decreased to the control group (P > 0.05 ),while the serum CEA level [ (33.4 ± 30.6) μ g/L ] was still significantly higher than the control group (P < 0.01 ).The sensitivity of the serum Ang-2 for diagnosis of gastric cancer was markedly higher than that of the serum CEA (P < 0.01 ).There was correlation between serum Ang-2 and degree of tumor differentiation,TNM pathological staging,lymphatic metastasis,invasion depth and tumor size (p <0.01 ),but there was no correlation between serum Ang-2 and tissue classification and location of gastric cancer (P> 0.05).Conclusion The serum Ang-2 level is suggested to be a valuable gastric cancer marker and conduce to the diagnosis of gastric cancer,the monitoring of recurrence after operation and evaluation of prognosis.
9.Establishment and optimization of real-time PCR with SYBR Green Ⅰ for quantification of survivin
Li-Hua HU ; Zheng-Jiang CHENG ; Yi-Rong LI ; Shao-Jun HUANG ;
Chinese Journal of Laboratory Medicine 2003;0(12):-
Objective To establish real-time PCR with SYBR Green Ⅰ for quantification the gene of survivin.Methods The components and conditions of PCR system were optimally determined by fluorescence intensity,cycle threshold(Ct),melting curve,coefficiency and slope of the standard curve.The means to eliminate the contaminating fluorescence of primer-dimers and the mode for Ct value determination were also optimized.Using the developed PCR system,we quantificated the survivin gene in 43 patients with gastric carcinoma.Results The optimized condition for PCR amplification of survivin were 2 mmol/L of MgCl_2,2.5 U/100 ?l of Taq DNA polymerase,0.2 ?mol/L of primers,and the optimized annealing temperatures for PCR were 58℃.The influence of primer dimmer can be eliminated by setting the fluorescence collecting temperature below the Tm of the specific amplicon by 2℃.The second derivative maximum mode,instead of fit point mode,was a feasible method to determine the Ct value for quantification.The sensitivity of this method was 10 copies/?l,and a good linearity was found from 10~1 to 10~4 copies/?l(r = 0.999 7).The inter-experimental coefficient of variation was 1.13%-1.91%,whereas the coefficient of variation between runs was 3.31%-4.50%.Using the optimized PCR system,we quantificated the gene of survivin,the result indicated that survivin gene was amplified in 13.9% of gastric carcinomas.Conclusions The optimal real-time PCR with SYBR Green Ⅰ,as a cost-effective and feasible DNA quantitative method,is fit for quantification of the survivin with satisfactory repeatability and high sensitivity.
10.Identification and Characterization of a Lactococcus lactis Strain with the Distinctive Antimicrobial Activity Against Gram-negative Bacteria and a Yeast
Xin-Feng HUANG ; Meng-Tian JIANG ; Xiao-Hu SHAO ; Xiao-Hua LI ; Lin LI ;
Microbiology 2008;0(09):-
Using the Gram-positive Staphylococcus aureus as the indictor bacterium,fourteen antibacterial strains were initially obtained by the bilayer-media screening method from the raw milk samples,and one isolate was found to exhibit the higher antibacterial activity against the indicator. This isolate was further studied on its individual and cultural morphology features,partial physiological and biochemical reaction activities,G+C content,the sequence features of the 16S rDNA and the species-specific N-acetylmuraminidase gene(acmA) ,consequently,it was identified as the Lactococcus lactis subsp. lactis strain,named as MB191. An evaluation of the antimicrobial spectra of MB191 was subsequently performed,it showed the remarkable activities against not only the tested Gram-positive bacteria,but also several Gram-negative bacteria including Pseudomonas syringae and P. fluorescens,as well as the yeast Debaryomyces hansenii,which was a distinctive feature that was not reported prior to this study.