Objective:To quantify the tryptase and T cell immunoglobulin mucin domain-3(TIM-3)double positive mast cells in hu-man chronic periodontitis tissue using double immunofluorescence staining.Methods:25 healthy controls,28 chronic mild periodontitis and 30 chronic advanced periodontitis patients were included.The gingival specimens were stained with HE for histology,and with double immunofluorescence staining for the identification of tryptase and TIM-3 double positive mast cells in gingival tissue.Results:In chronic periodontitis tissue the degree of gingival inflammation was significantly increased,the densities(cells/mm2 )of tryptase and TIM-3 double positive mast cells were significantly increased(P<0.05),in addition,that in chronic advanced periodontitis group was significantly higher than in the chronic mild periodontitis group(P<0.05).Conclusion:Tryptase and TIM-3 double positive mast cells has the similar tendency as the severity of periodontitis inflammation in human periodontitis tissue.Tryptase and TIM-3 double positive mast cells may play an important role in human chronic periodontitis.

AIM:To identify and quantify the expression of IL-1βand IL-17 in mast cells ( MCs) in different types of human pericapical diseases using double immunofluorescence staining .METHODS: The specimens ( n=102 ) , including healthy control ( n=35 ) , periapical cyst ( n=35 ) and periapical granuloma ( n=32 ) , were involved in the present study .The tissue samples were fixed in 10% buffered formalin for at least 48 h and then embedded in paraffin . Serial 5-μm-thick sections were deposited onto SuperFrost/Plus microscope glasses .Routine staining of the sections using hematoxylin&eosin ( HE) was performed for morphological evaluation .The number of IL-1βand IL-17 positive MCs was identified by double immunofluorescence staining .RESULTS:Compared with the healthy controls , the inflammation score of periapical lesions was significantly increased in the periapical patients (P<0.01).The density of IL-1βand IL-17 posi-tive MCs in the periapical lesions were obviously higher than that in the healthy controls (P<0.01).However, no signifi-cant difference between periapical cyst and periapical granuloma was observed .The Pearson correlation analysis showed that there was a positive correlation between the density of IL-1βand IL-17 double positive MCs and inflammation score in dif-ferent groups of specimens (P<0.01).CONCLUSION:There is significantly increased number of MCs , along with in-creased density of IL-1βand IL-17 positive MCs in human periapical lesions .The increased density of IL-1βand IL-17 positive MCs has the similar tendency as the severity of tissue inflammation in human periapical lesions , suggesting that IL-1βand IL-17 positive MCs may play an important role in the pathogenesis of human periapical diseases .

AIM: To investigate the expression of stem cell factor (SCF) in tryptase -positive mast cells ( MCs) in different types of human periapical diseases for determining the role of SCF and MCs in the pathogenesis of peria-pical diseases.METHODS: A total 50 cases of specimens were involved in this study, including healthy control (n=20), periapical cyst (n=15) and periapical granuloma (n=15).The tissue material was fixed in 10%formalin for at least 48 h, stained with hematoxylin and eosin for the observation of histopathology, stained with immunohistochemistry for identifying MCs and MCs degranulation, and stained with double immunofluorescence for identification of tryptase-SCF double positive MCs.RESULTS:Compared with healthy control, significantly higher densities of both total and degranu-lated MCs in human periapical lesions were observed.The densities of both total and degranulated MCs in the periapical cyst were significantly higher than that in the periapical granuloma.The density of tryptase-SCF double positive MCs in the periapical lesions was significantly higher than that in the healthy controls.The density oftryptase-SCF double positive MCs in the periapical cyst was significantly higher than that in periapical granuloma.No significant difference in the density of MCs between immunohistochemistry staining and double immunofluorescence staining was observed.CONCLUSION:The tryptase-SCF double positive MCs play an active role in the pathogenesis of the periapical inflammatory lesions, particularly in the formation of fibrous tissue in periapical cyst.The potential role of the tryptase-SCF double positive MCs relates with the initiation, development, and persistence of the periapical inflammatory process.

Objective To observe the effect of oxaliplatin on expression of drug resistant gene in DNA damage repair in human colon cancer cells.Methods The human colon cancer cell line HCT116 was processed with oxaliplatin for 48 hours.Western blot was used to examine DNA damage induced by oxaliplatin treatment.CCK-8 assay was used to explore the effect of oxaliplatin on growth of HCT116.The real-time PCR was used to detect the expression of DNA repair genes after oxaliplatin treatment.Results After the treatment of oxaliplatin,the expression of the DNA-damage-marker gene γ-H2AX increased significantly,and DNA damage excision repair related gene XPC increased significantly as well.The mRNA expression of other DNA damage repair genes had no significant increase.Conclusion The gene XPC might play a key role in generating drug resistance when treating colon cancer with oxaliplatin in clinic.

Objective To observe the effect of oxaliplatin on expression of drug resistant gene in DNA damage repair in human colon cancer cells.Methods The human colon cancer cell line HCT116 was processed with oxaliplatin for 48 hours.Western blot was used to examine DNA damage induced by oxaliplatin treatment.CCK-8 assay was used to explore the effect of oxaliplatin on growth of HCT116.The real-time PCR was used to detect the expression of DNA repair genes after oxaliplatin treatment.Results After the treatment of oxaliplatin,the expression of the DNA-damage-marker gene γ-H2AX increased significantly,and DNA damage excision repair related gene XPC increased significantly as well.The mRNA expression of other DNA damage repair genes had no significant increase.Conclusion The gene XPC might play a key role in generating drug resistance when treating colon cancer with oxaliplatin in clinic.

Objective To investigate the role of the activated rhuPAa-melittin in ovarian cancer cells and to study the inhibitory effect of rhuPAa-melittinon on ovarian cancer cells. Methods rhuPAa-melittin was used to treat the ovarian cancer cells at different concentrations for 48 hrs. Then flow cytometery was applied to detect the cell cycle and cell apoptosis. rhuPAa-melittin protein was delivered to the mouse mode to investigate the effect of rhuPAa-melittin on the growth of the xenotransplanted tumor. Results rhuPAa-melittin was used to treat ovarian cancer cells at the concentration of 0 ,4 ,8 and 16 μg/mL for 48 hrs ,respectively. The results of cell apoptosis assay was 1.16%,3.83%,6.51% and 10.2%,respectively. Moreover,different concentrations of rhuPAa-melittin had no effects on the cells at G0/G1 phase,rhuPAa-melittin inhibited S phase cells to process into G2/M phase, contributing to suppressing the growth of ovarian cancer cells. Conclusion The in vivo and in vitro studies revealed that rhuPAa-melittin inhibits the growth of ovarian cancer.

Objective Recent evidence has implicated Larp1,an RNA-binding protein,in cell motility and EMT therefore prompting the study of Larp1 in EOC.This project aims to examine the potential role of Larp1 in cell invasion.Methods Ovarian cancer cells SKOV-3 and OVCAR-3 were transfected with DNA constructs to overexpress Larp1 and transfect cells were used to assess the overexpression of Larp1 in cell invasion and metastasis using several functional assay.Results Larp1 overexpression in both OVCAR-3 and SKOV-3 cells appear to enhance the invasive ability of cells and the accumulation of Larp 1 in these chemoinvasive protrusions strongly suggests the potential role of Larp1 in invadopodia formation.Cell spot chemoinvasion assay demonstrated increased chemoinvasion of Larp1 overexpressing SKOV-3 and OVCAR-3 cells towards a gradient of TGF-β,EGF and bFGF.Conclusions With its potential role in EMT and cell invasion,a deeper understanding of the physiological role of Larp1 in cancer cell motility will be potentially beneficial in the diagnosis and treatments of ovarian cancer.

Objective:To explore the expression of long-chain noncoding RNA (lncRNA) and myocardial infarction-associated transcription (MIAT) in Leukocyte differentiation antigen (CD)4+T cells in peripheral blood of gastric cancer patients and its value of clinical application.Methods:Peripheral blood CD4+T cells were collected from 124 patients with gastric cancer, 90 benign gastric diseases patients and 80 healthy controls enrolled in Taizhou People′s Hospital from January 2019 to April 2021. The expression levels of MIAT and N6-methyladenosine(m6A) binding to MIAT promoter in CD4+T cells were detected by real-time fluorescent quantitative polymerase chain reaction (qPCR) and Chromatin immunoprecipitation (ChIP)-qPCR, respectively. Spearman test was used to analyze the correlation between MIAT and clinicopathological features, as well as between MIAT and regulatory T cell levels. The receivor operating characteristic curve (ROC) of the subjects was used to evaluate the MIAT expression level in the auxiliary diagnostic value of gastric cancer.Results:The relative expression levels of MIAT in the gastric cancer patients, the benign gastric diseases patients, and the healthy controls were 2.849 (2.131, 4.062), 1.511 (0.916, 1.855) and 0.963 (0.729, 1.432), respectively. The difference among the three groups was statistically significant ( H=158.25, P<0.001). The relative expression level of MIAT in the gastric cancer patients was significantly higher than the levels in the benign gastric diseases patients and healthy controls. The difference was statistically significant ( Z=100.63, 145.14, P<0.001). The binding activity of m6A to MIAT promoter in patients with early stage (stage Ⅰ and Ⅱ) and end stage (stage Ⅲ and Ⅳ) gastric cancer was 8.590±1.483 and 4.274±0.425, respectively. The difference was statistically significant ( t=6.255, P=0.002). Furthermore, the binding activity of m6A to MIAT promoter in the gastric cancer patients was significantly lower than that in patients with benign gastric diseases (17.267±3.106) and healthy controls (27.637±3.945) ( t=-7.331,-12.832, P<0.001). The relative expression of MIAT in CD4+T cells in peripheral blood of the gastric cancer patients had no significant difference in age(χ2=0.000, P=1.000), gender(χ2=0.000, P=1.000), CEA (χ2=0.648, P=0.421) and CA199(χ2=1.554, P=0.213), but had significant difference with tumor size expression(χ2=9.443, P<0.01), TNM stage(χ2=23.571, P=0.002) and lymph node metastasis (χ2=45.248, P<0.01). In addition, there was a significant positive correlation between the relative expression of MIAT in CD4+T cells and Treg level ( r2=0.76, P<0.001). The diagnostic efficacies of MIAT in CD4+T cells, CEA and CA199 in the gastric cancer patients were analyzed by ROC curve. When compared with patients with benign gastric diseases, the areas under the curve were 0.879, 0.635 and 0.611, respectively. When compared with healthy patients, the areas under the curve were 0.953, 0.784 and 0.598, respectively. Conclusions:The level of MIAT in CD4+T cells in peripheral blood of patients with gastric cancer is significantly higher than the levels in patients with benign gastric diseases and the healthy controls, which may be related with the decreased activity of m6A binding to the promoter of MIAT. The level of MIAT in CD4+T cells may be a relevant biomarker for the diagnosis and prognosis of gastric cancer.

Objective RhuPAa-melittin protein with high-purity was analyzed to investigate its biologic effect on normal and ovarian cancer cells.Methods Preliminary purified rhuPAa-melittin was added in medium normal cells and SKOV3 human ovarian cancer cells cultured for 48 hours,and then were measured at 490 nm wavelength absorbance to measure the number of living cells and evalu-ate the inhibitory effect of rhuPAa-melittin on cultured cells.Results The inhibitory rate of rhuPAa-melittin at 4 μg/mL was 66.59% in SKOV3 cancer cell progression.At a concentration of 16 μg/mL, rhuPAa-melittin killed all cancer cells (P <0.05).In normal adult fibrous cells,the inhibitory rate of rhuPAa-melittin at 4 μg /mL was only 7.3%,and 12.3% inhibition rate was obtained by using rhu-PAa-melittin (8 μg/mL).Moreover,inhibition rate topped at 22.0% when used rhuPAa-melittin at 16 μg/mL.Conclusion RhuPAa-melittin has a strong anti-tumor effect on SKOV3 cancer cells.

Objective RhuPAa-melittin protein with high-purity was analyzed to investigate its biologic effect on normal and ovarian cancer cells.Methods Preliminary purified rhuPAa-melittin was added in medium normal cells and SKOV3 human ovarian cancer cells cultured for 48 hours,and then were measured at 490 nm wavelength absorbance to measure the number of living cells and evalu-ate the inhibitory effect of rhuPAa-melittin on cultured cells.Results The inhibitory rate of rhuPAa-melittin at 4 μg/mL was 66.59% in SKOV3 cancer cell progression.At a concentration of 16 μg/mL, rhuPAa-melittin killed all cancer cells (P <0.05).In normal adult fibrous cells,the inhibitory rate of rhuPAa-melittin at 4 μg /mL was only 7.3%,and 12.3% inhibition rate was obtained by using rhu-PAa-melittin (8 μg/mL).Moreover,inhibition rate topped at 22.0% when used rhuPAa-melittin at 16 μg/mL.Conclusion RhuPAa-melittin has a strong anti-tumor effect on SKOV3 cancer cells.