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Objective:To observe the difference of the human telomeres RNA component (hTERC) genes’amplification in the cervical tissue by applying the environment-friendly fixative poly hydroxy acrylic acid and the transparent dewaxing solution Van-clear separately or jointly to replace the traditional fixative 4% (volume fraction)neutral buffered formalin and the conventional transparent dewaxing solu-tion xylene in the use of fluorescence in situ hybridization (FISH)for detection.Methods:In the study, 255 cases of cervical tissue specimens submitted by the Department of Gynecology in Zhongshan Boai Ho-sipital were collected from Mar.2013 to Apr.2015.Four samples were taken from the same lesion site. All the cases were divided into 4 groups and named group A,B,C,and D.Group A used 4% neutral buffered formalin fixed and xylene dewaxing to make slices.Group B used poly hydroxy acrylic fixed and xylene dewaxing to make slices.Group C used 4% neutral buffered formalin fixed and Van-clear trans-parent to make slices.Group D used poly hydroxy acrylic fixed and Van-clear transparent dewaxing to make slices.The amplification of hTERC genes in the four groups of cervical specimens was also detected by FISH technique.Results:When the hTERC genes were detected by FISH method under the fluore-scence microscope,it was obvious that the tissue profile and the background of group A,B,C and D were all clear.The probe was fixed in the accurate position so that the bright red or green fluorescence signals were easily found in these four groups.Compared with the positive rate of group A,there was no statistical significance in that of group B,C and D (P>0.05).At the same time,the coincidence rate of the FISH results was high,which showed that the new environment-friendly reagent had no significant difference in the detection of cervical hTERC genes by FISH technique.Conclusion:It is possible for the environment-friendly reagent poly hydroxy acrylic acid and Van-clear to replace 4%neutral buffered for-malin and xylene separately or jointly to detect the cervical hTERC genes by adopting FISH technique.
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OBJECTIVE@#To observe the effect of Van-Clear on vamplification of human telomerase RNA component (hTERC) gene in cervical tissues by fluorescence in situ hybridization, and to determine the potential for Van-Clear to replace xylene. @*METHODS@#A total of 278 specimens of cervix uteri were collected from inpatients of Department of Gynaecology in Boai Hospital of Zhongshan from January to February, 2015, with 81 cases of normal specimens, 68 cases of cervical intraepithelial neoplasia (CIN) I, 57cases of CIN2, 42 cases of CIN3 and 30 cases of cervical invasive cancer. Double samples were collected from the same region. Fluorescence in situ hybridization was applied to detect the changes in the amplification of hTERC gene in 2 groups of specimens from the cervical biopsy. @*RESULTS@#Differences in the positive expression rate of hTERC gene between the 2 groups of cervical lesions at all levels were not statistically significant (P>0.05). @*CONCLUSION@#There is no significant difference in the positive rate of hTERC gene expression between the slices made by Van-clear and xylene. As an environmental-friend product, Van-Clear possesses certain value in detection of cervical hTERC gene by fluorescence in situ hybridization.
Assuntos
Feminino , Humanos , Displasia do Colo do Útero , Genética , Amplificação de Genes , Hibridização in Situ Fluorescente , RNA , Genética , Telomerase , Genética , Neoplasias do Colo do Útero , Genética , Xilenos , QuímicaRESUMO
Background and purpose: Adequate tissue ifxation, transparent dewaxing is an important step of hematoxylin eosin (HE) staining and lfuorescence in situ hybridization (FISH) in detection of breast cancerHER-2 gene. The purpose of this study was to make a comparison between poly hydroxyl acrylic acid which is an environmen-tally friendly ifxation liquid and 4% neutral buffered formaldehyde in tissue ifxation for HE staining and FISH to detect theHER-2 gene in the breast cancer tissue sections. The study aimed to evaluate the feasibility of replacing 4% buffered formaldehyde, a traditional ifxation liquid, with the poly hydroxyl acrylic acid, an environmentally friendly ifxation lfuid.Methods:This project was performed on tissue samples collected from 69 cases of breast cancer, 41 cases of breast ifbroadenoma, 40 cases of uterine leiomyoma, 25 cases of cervical tissue, 25 cases of placenta obtained from the outpatient and inpatient departments of Zhongshan Boai Hospital from Mar. 2011 to Jan. 2015, from each of which two samples were drawn and two blocks of each specimen were divided into two groups randomly. Then one group was ifxed with 4% neutral buffered formaldehyde and made into 200 sections by HE while the other group was ifxed with poly hydroxyl acrylic acid and made into another 200 sections. The slice level of the two groups was determined by the staining condition of the sections, and SPSS 19.0 was employed to compare the excellent and good rate of HE staining. Additional 69 sections were produced with two groups of breast invasive ductal carcinoma tissues, and SPSS 19.0 was used to detect the ampliifcation ofHER-2 gene by FISH.Results:First, the number of best-quality slices stained with HE ifxed separately by poly hydroxyl acrylic acid and 4% neutral buffered formaldehyde was 155 and 166, respectively. The number of excellent pieces was 41 and 33, respectively, while the number of mediocre pieces was 3 and 1 with bad pieces being 1 and 0, respectively. The excellent and good rates of HE staining were 98% and 99.5%, respectively. There was no significant difference between the two groups (χ2=1.33,P>0.05).Second, the positive rates of the tis-sue slices by FISH ifxed separately by poly hydroxyl acrylic acid and 4% neutral buffered were 26.09% and 23.19%, respectively. There was no signiifcant difference between the two groups (χ2=0.50,P>0.05).Conclusion:The results obtained with HE staining and FISH using poly hydroxyl acrylic acid as a ifxation liquid are not signiifcantly different from those using 10% neutral buffered formaldehyde. Therefore, poly hydroxyl acrylic acid meets the requirements of environmental protection, and thus has the potential to be promoted and widely used.
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Objective To investigate the expressions and effects of EGR-1, p53, p16, Cyclin D1 in Nasopharyngeal Carcinoma (NPC). Methods Expressions of EGR-1, p53, p16, and Cyclin D1 protein in paraffin-embedded specimens of 29 chronic nasopharyngitis, 45 NPC and 24 lymph-node metastasis were studied by immunohistochemical method (SABC). Results The positive rates of EGR-1, p16, and Cyclin D1 protein expressions in chronic nasopharyngitis were 44.83 %(13/29), 89.66 %(26/29) and 24.14 %(7/29) respectively, but no p53 protein expression. In NPC, the positive expression rates of the four proteins were 22.22 %(10/45), 91.11 %(41/45), 53.33 %(24/45) and 66.67 %(30/45), with significant difference compared with chronic nasopharyngitis respectively; and in lymph nodes metastasis were 16.67 %(4/24), 79.17 %(19/24), 70.83 %(17/24) and 75.00 %(18/24) with significant difference compared with those of chronic nasopharyngitis and NPC respectively except for p16 protein. Spearman correlation analysis showed that there were no significant correlation between the expression of EGR-1 and p53, p16, Cyclin D1 respectively both in NPC and in lymph nodes metastasis. In NPC, the expression of p53 showed a positive correlation with the expression of p16; and in lymph nodes metastasis the expression of Cyclin D1 showed positive correlation with the expressions of p16 and p53. Conclusions The expression of EGR-1 protein was significantly decreased both in NPC and in lymph-node metastasis, which suggested that Egr-1 gene may be a tumor suppressor gene and have an important effect in the pathogenesis and the progression of NPC. The overexpression of p53 and Cyclin D1 also have an important role in NPC, but p16 probably have no biological significance in NPC.
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Objective To investigate the effects of H_2O_2 on growth and proliferation in CNE-2Z cells.Methods To identify the effects of H_2O_2 on the growth and proliferation in CNE-2Z cells,Cell growth curves,MTT assay and plate colony formation rate were used;Meanwhile,Cell cycle distribution,proliferation index and apoptosis analysis were performed by flow cytometry(FCM) in human NPC cells CNE-2Z in vitro,before and after the action of H_2O_2;the protein expressions of Egr-1,p53,p16,Cyclin D1 were detected by Western-Blotting analysis.Results There were no EGR-1 and P16 proteins expressions in CNE-2Z cells by Western-Blotting analysis before the action of H2O2;the growth and proliferation of CNE-2Z cells were inhibited significantly by H_2O_2 compared with the control cells,and the apoptosis cells were increased.The increased expressions of EGR-1,P53,and Cyclin D1 protein were also shown in CNE-2Z cells by means of Western-Blotting analysis after the action of H_2O_2;but no P16 protein expression;The expression of EGR-1 protein was significant correlation with P53 protein expression,but no significant correlation with Cyclin D1 protein expression.Conclusion There was no EGR-1 protein expression in CNE-2Z cells,and EGR-1 protein can be induced expression rapidly and transiently by H_2O_2,which probably have an important!role in the growth inhibition and apoptosis.The promotor of Egr-1 gene also can be induced activation by radiation(ROS,reactive oxygen species),there is an important application in therapy of NPC and other tumors.