1.Clinical study on the treatment of OSAHS in children with different surgical approaches based on H-uvu-lopalatopharyngoplasty
Zhitai CHEN ; Shengguo WANG ; Benzhong ZHOU ; Xianping SHI ; Feifeng WU ; Longqiao LI ; Zesheng ZHANG ; Guangjie ZHOU ; Yang LIU
Journal of Practical Stomatology 2024;40(5):687-690
Objective:To study the clinical effects of modified surgical method with H-uvulopalatopharyngoplasty(H-UPPP)and traditional surgical method in the treatment of obstructive sleep apnea hypopnea syndrome(OSAHS)in children.Methods:The clini-cal data of 364 pediatric patients with OSAHS were retrospectively analyzed.The patients were divided into 2 groups and treated by low-temperature plasma knife radio frequency ablation for bilateral tonsil removal combined with H-UPPP(group A,n=204)and low-temperature plasma knife radio frequency ablation only(group B,n=160)respectively.The operative time,intraoperative bleed-ing,postoperative secondary bleeding,VAS of pain following operation,postoperative traumatic white film shedding time,OAH1,LSaO2,ESS scores and complications of the 2 groups were compared.Results:In group A the operative time,intraoperative bleed-ing,VAS,OAHI and ESS scores were lower than those in group B(P<0.05),wile postoperative traumatic white film detachment time and LSaO2 were higher(P<0.05).No statistically significant difference in terms of the number of cases of postoperative seconda-ry bleeding and complications was observed between the 2 groups(P>0.05).Conclusion:The combination of low-temperature plas-ma radio frequency ablation and H-UPPP for the treatment of OSAHS may provide clear intraoperative view,and may improve the treatment effects.
2.Expression of Tim-3 in the peripheral blood of pancreatic cancer patients and the diagnostic value in combination with serum glucose chain antigen 19-9
Ke WANG ; Lijuan LIU ; Xianping CUI ; Wenying JIANG ; Changtao YU ; Jinxin SHI
Chinese Journal of Endocrine Surgery 2023;17(3):278-281
Objective:To investigate the expression of soluble T cell immunoglobulin and mucin domain-3 (Tim-3) in peripheral blood of patients with pancreatic cancer and its diagnostic value in combination with serum Carbohydrate antigen 19-9 (CA19-9) .Methods:106 newly diagnosed pancreatic cancer patients and 65 age and sex matched healthy individuals were enrolled. Tim-3 concentration was quantitatively determined by enzyme-linked immunosorbent assay (ELISA). According to the expression levels of soluble Tim-3 and serum CA19-9, a binary logistic regression model of receiver operating characteristic (ROC) curve was established to compare the diagnostic effects of serum CA19-9 and soluble Tim-3 alone or combined with the two tests.Results:The levels of soluble Tim-3 in the pancreatic cancer group were significantly higher than those in the healthy control group ( P<0.001). The expression level of soluble Tim-3 was significantly higher in patients with stage III-IV pancreatic cancer than in patients with stage I-II ( P=0.003). The AUC of soluble Tim-3 diagnosis for stage I-II pancreatic cancer was 0.856 (95%CI: 0.765 to 0.992 P<0.001), Serum CA19-9 The AUC used for the stage I-II pancreatic cancer diagnosis was 0.862 (95%CI: 0.772 to 0.926 P<0.001), The AUC for the combined diagnosis was 0.949 (95%CI: 0.880 - 0.985 P<0.001) ; In a healthy population and in patients with stage III-IV pancreatic cancer, the AUC of soluble T I I-IV pancreatic cancer in stage III was 0.927 (95%CI: 0.873 to 0.963 P<0.001), the AUC of serum CA19-9 used for the diagnosis of stage III-IV pancreatic cancer was 0.933 (95%CI: 0.881 to 0.968 P<0.001), the AUC for the combined diagnosis was 0.989 (95%CI: 0.956 to 0.999 P<0.001) . Conclusions:The combination of soluble Tim-3 and serum CA19-9 can improve the diagnostic rate of pancreatic cancer patients.
3. Impact and related mechanism on the improvement of hyperglycemia-induced pyroptosis in H9c2 cells by mircoRNA-214
Yan WANG ; Ranzun ZHAO ; Panke CHEN ; Guanxue XU ; Zhijiang LIU ; Xianping LONG ; Zhimei QIU ; Bei SHI
Chinese Journal of Cardiology 2019;47(10):820-828
Objective:
To investigate whether microRNA(miR)-214 can improve hyperglycemia induced pyroptosis in H9c2 cells through targeting caspase-1.
Methods:
H9c2 cells of rats those in good growth condition were selected and incubated into the T25 culture bottle after digestion and passage. Cells were cultured in an incubator at 37 ℃ with 5%CO2, repeat passage was made after cell density reached about 80%, The 5th to 8th generations of cells were selected for the subsequent experiments. To observe the effect of overexpression of miR-214 on pyroptosis and caspase-1 expression in H9c2 cells induced by hyperglycemia, the cells were divided into 4 groups: Control group(H9c2 cells cultured normally), Hyperglycemia group (HG group, 50 mmol/L glucose was used to intervene H9c2 cells for 24 hours), miR-214 mimics+hyperglycosis group (mimics+HG group, H9c2 cells were transfected with miR-214 mimics for 24 hours and then treated with 50 mmol/L hyperglycosis for 24 hours), miR-214 mimic-negative control+hyperglycaemic group(MNC+HG group, H9c2 cells were transfected with miR-214 mimic-negative control for 24 hours and then treated with 50 mmol/L hyperglycaemic for 24 hours). In order to further verify the anti-pyroptosis effect of miR-214 was mediated by targeted inhibition on caspase-1, cells overexpressing caspase-1 were used in the rescue experiment. The cells overexpressing caspase-1 were divided into 4 groups: Hyperglycemia group (HG group, 50 mmol/L glucose was used to intervene H9c2 cells for 24 hours), miR-214 mimics+hyperglycosis group (mimics+HG group, H9c2 cells were transfected with miR-214 mimics for 24 hours and then treated with 50 mmol/L hyperglycosis for 24 hours), miR-214 mimics+hyperglycosis+recombinant adenovirus (Ad-caspase-1-EGFP) group with caspase-1 gene and EGFP green fluorescent protein expression (mimics+HG+Ad-caspase-1-EGFP group, H9c2 cells were transfected with caspase-1-green fluorescent protein-carrying adenovirus for 48 hours, followed by transfection of miR-214 mimics for 24 hours, and then treated with 50 mmol/L hyperglycaemia for 24 hours), miR-214 mimics+HG+Ad-EGFP empty virus group (mimics+HG+Ad-EGFP group, H9c2 cells were transfected with empty adenovirus containing green fluorescent protein for 48 hours, followed by transfection with miR-214 mimics for 24 hours, and then treated with 50 mmol/L hyperglycosis for 24 hours). The mRNA expression levels of miRNA-214 and caspase-1 in cells were detected by real-time quantitative PCR. The expression and localization of caspase-1 protein were detected by immunofluorescence assay. Western blot was used to detect protein expression levels of procaspase-1, cleaved caspase-1, NLRP3 and ACS with β-actin as internal reference. The secretion of IL-1β and IL-18 in cell culture medium was detected by ELISA. The correlation between miR-214 and caspase-1 was detected by double luciferase reporter gene.
Results:
(1) The mRNA expression levels of miR-214 and caspase-1 in each group: the mRNA expressions of miR-214 in HG group and MNC+HG group were significantly lower than that in control group(
4.Impact and related mechanism on the improvement of hyperglycemia?induced pyroptosis in H9c2 cells by mircoRNA?214
Yan WANG ; Ranzun ZHAO ; Panke CHEN ; Guanxue XU ; Zhijiang LIU ; Xianping LONG ; Zhimei QIU ; Bei SHI
Chinese Journal of Cardiology 2019;47(10):820-828
Objective To investigate whether microRNA(miR)?214 can improve hyperglycemia induced pyroptosis in H9c2 cells through targeting caspase?1. Methods H9c2 cells of rats those in good growth condition were selected and incubated into the T25 culture bottle after digestion and passage. Cells were cultured in an incubator at 37℃with 5%CO2, repeat passage was made after cell density reached about 80%, The 5th to 8th generations of cells were selected for the subsequent experiments. To observe the effect of overexpression of miR?214 on pyroptosis and caspase?1 expression in H9c2 cells induced by hyperglycemia, the cells were divided into 4 groups: Control group(H9c2 cells cultured normally), Hyperglycemia group (HG group, 50 mmol/L glucose was used to intervene H9c2 cells for 24 hours), miR?214 mimics+hyperglycosis group (mimics+HG group, H9c2 cells were transfected with miR?214 mimics for 24 hours and then treated with 50 mmol/L hyperglycosis for 24 hours), miR?214 mimic?negative control+hyperglycaemic group(MNC+HG group, H9c2 cells were transfected with miR?214 mimic?negative control for 24 hours and then treated with 50 mmol/L hyperglycaemic for 24 hours). In order to further verify the anti?pyroptosis effect of miR?214 was mediated by targeted inhibition on caspase?1, cells overexpressing caspase?1 were used in the rescue experiment. The cells overexpressing caspase?1 were divided into 4 groups: Hyperglycemia group (HG group, 50 mmol/L glucose was used to intervene H9c2 cells for 24 hours), miR?214 mimics+hyperglycosis group (mimics+HG group, H9c2 cells were transfected with miR?214 mimics for 24 hours and then treated with 50 mmol/L hyperglycosis for 24 hours), miR?214 mimics+hyperglycosis+recombinant adenovirus (Ad?caspase?1?EGFP) group with caspase?1 gene and EGFP green fluorescent protein expression (mimics+HG + Ad?caspase?1?EGFP group, H9c2 cells were transfected with caspase?1?green fluorescent protein?carrying adenovirus for 48 hours, followed by transfection of miR?214 mimics for 24 hours, and then treated with 50 mmol/L hyperglycaemia for 24 hours), miR?214 mimics+HG+Ad?EGFP empty virus group (mimics+HG+Ad?EGFP group, H9c2 cells were transfected with empty adenovirus containing green fluorescent protein for 48 hours, followed by transfection with miR?214 mimics for 24 hours, and then treated with 50 mmol/L hyperglycosis for 24 hours). The mRNA expression levels of miRNA?214 and caspase?1 in cells were detected by real?time quantitative PCR. The expression and localization of caspase?1 protein were detected by immunofluorescence assay. Western blot was used to detect protein expression levels of procaspase?1, cleaved caspase?1, NLRP3 and ACS with β?actin as internal reference. The secretion of IL?1β and IL?18 in cell culture medium was detected by ELISA. The correlation between miR?214 and caspase?1 was detected by double luciferase reporter gene. Results (1) The mRNA expression levels of miR?214 and caspase?1 in each group: the mRNA expressions of miR?214 in HG group and MNC+HG group were significantly lower than that in control group(P<0.05). The mRNA expression of miR?214 in mimics+HG group was significantly higher than that in control group (P<0.05). The mRNA expression levels of caspase?1 in HG group and MNC+HG group were significantly higher than that in control group(P<0.05). The mRNA expression level of caspase?1 in mimics+HG group was lower than that in control group(P<0.05). (2) The expression of caspase?1 in each group: the green fluorescence intensity in the control group was weak, which was strong in the HG group and MNC+HG group. The green fluorescence expression was weaker in mimics+HG group than in HG group. (3) ASC and NLRP3 protein expression levels in each group: ASC and NLRP3 protein expression levels in HG group and MNC+HG group were higher than those in control group(P<0.05). ASC and NLRP3 protein expression levels were significantly lower in mimics+HG group than in mimics+HG group (P<0.05). (4) The secretion of IL?1β and IL?18 in the cell culture medium of each group: the content of IL?1β and IL?18 in the cell culture medium of HG group and MNC+HG group was significantly higher than that of control group (P<0.05). The content of IL?1β and IL?18 in the cell culture medium of mimics+HG group was significantly lower than that of the HG group (P<0.05). (5) Correlation between miR?214 and caspase?1: miR?214 specifically binds to caspase?1 3 'UTR. Meanwhile, Western blot results showed that cleaved caspase?1 protein expression levels were significantly higher in both HG group and MNC+HG group than in control group (P<0.05). The levels of cleaved caspase?1 were significantly lower in mimics+HG group than in HG group (P<0.05). There was no significant difference in procaspase?1 expression among groups (P>0.05). (6) The expression levels of procaspase?1, cleaved caspase?1, ASC and NLRP3 in each group in rescue experiment: there was no significant difference in the expression of procaspase?1 in each group (P>0.05). Cleaved caspase?1 , ASC and NLRP3 protein expressions were significantly lower in mimics+HG group than in HG group (P<0.05). However, cleaved caspase?1, ASC and NLRP3 protein expressions were significantly higher in mimics+HG+Ad?caspase?1?EGFP group than in mimics+HG group (P<0.05). (7) The expression of IL?1β and IL?18 in rescue experiment: the secretions of IL?1β and IL?18 in the cell culture medium of the mimics+HG group were significantly lower than that of HG group (P<0.05), which were significantly higher in mimics+HG+Ad?caspase?1?EGFP group than in mimics+HG group (P<0.05). Conclusion miR?214 can improve the hyperglycemia induced pyroptosis in H9c2 cells by targeted inhibition of the caspase?1.
5. Effect and related mechanism of miRNA-21 on hydrogen peroxide-induced C-kit+ cardiac stem cells apoptosis
Ranzun ZHAO ; Yan WANG ; Wenwen DENG ; Bei SHI ; Xianping LONG ; Zhenglong WANG ; Wenming CHEN
Chinese Journal of Cardiology 2017;45(11):971-977
Objective:
To explored the effect and related mechanism of miRNA-21 on hydrogen peroxide-induced C-kit+ cardiac stem cells apoptosis.
Methods:
C-kit+ cardiac stem cells were isolated from SD rats by the methods of enzyme digestion and magnetic bead. Cells were divided into the following experimental groups: (1) negative control mimics (NCM)group: cells were transfected with negative control miRNA-21 mimics for 48 hours; (2)mimics group: cells were transfected with miRNA-21 mimics for 48 hours; (3) NCM+ H2O2 group: negative control miRNA-21 mimics were transfected into cells for 48 hours and then treated with 100 μmol H2O2 for 2 hours; (4)mimics+ H2O2 group: miRNA-21 mimics were transfected into cells for 48 hours and then treated with 100 μmol H2O2 for 2 hours. mRNA of miRNA-21 was detected by RT-PCR. The apoptosis rate of C-kit+ cardiac stem cells was determined using the annexin V-FITC/PI staining assay. Western blot was employed to measure the expression of apoptosis related proteins(Caspase-3, Bax, and Bcl-2).
Results:
(1) Compared with the NCM group, the mRNA expression level of miRNA-21 was significantly up-regulated in mimics group, while obviously down-regulated in NCM+ H2O2 group(all
6.Observation for the Effect of Vacuum Sealing Drainage in Treating the Patients With Pacemaker Pocket Infection
Dongmei WANG ; Bei SHI ; Xianping LONG ; Jidong RONG
Chinese Circulation Journal 2016;31(10):1006-1008
Objective: Pocket infection in patients after total removal of implanted pacemaker has many problems for their electronic system; our research intends to explore the feasibility of conservatively treating such infection and retain the electronic system. Methods: A total of 4 patients with pacemaker pocket infection in our hospital from 2015-01 to 2016-02 were studied. Thorough debridement and disinfection were conducted in infected pockets and devices, meanwhile vacuum sealing drainage was applied. Electrode wire was kept and intravenous antibiotics were given for (7-10) days after the operation in all patients. Results: The average time of infection occurred at 14.75 months after operation with the type of isolated pacemaker pocket infection. Pocket vacuum suction drainage was performed, with the mean of 7.25 (5-10) months follow-up observation, infection was disappeared and the patients had good wound healing. Conclusion: With thorough debridement of infected pocket, rational treatment of residual electronic system and vacuum sealing drainage, the infection might be effectively controlled for complete recovery without lead extraction in relevant patients.
7.Transfection of calcitonin gene-related peptide mediated by lentivirus vec-tor in vitro and its effects on cardiac stem cell viability
Jidong RONG ; Ling LI ; Xianping LONG ; Wenwen DENG ; Bei SHI
Chinese Journal of Pathophysiology 2016;32(8):1445-1449,1456
AIM:To investigate the effect of calcitonin gene-related peptide (CGRP)transfection into c-kitpos cardiac stem cells (c-kit+CSCs) on the cell viability.METHODS: Under the sterile condition, the auricles of SD rats were taken out , and then c-kit +CSCs were collected through enzyme digestion and immunomagnetic bead separation (MACS).The cells were identified by flow cytometry .c-kit +CSCs were transfected with enhanced green fluorescent pro-tein CGRP lentiviral vector ( Lv-EGFP-CGRP) or enhanced green fluorescent protein lentiviral vector ( Lv-EGFP).The cells were randomly divided into Lv-EGFP-CGRP-CSCs group , Lv-EGFP-CSCs group and CSCs group .The transfection was observed under the fluorescence microscope .The transfection efficiency was detected by flow cytometry .The CGRP protein secretion in the cell culture supernatants was detected by ELISA .The viability of c-kit+CSCs transfected with Lv-EGFP-CGRP or Lv-EGFP was measured by CCK-8 assay.RESULTS:c-kit+CSCs were isolated and cultured successfully .The expression positive rate of c-kit was 91.0%and the expression positive rates of CD 45 and CD34 were 4.5% and 4.0%, respectively.After transfected with lentivirus for 48 h, the stable fluorescence in c-kit +CSCs was observed under fluores-cence microscope .The transfection efficiency were 80%when MOI was 20.The level of CGRP was significantly increased in Lv-ECFP-CGRP-CSCs group compared with Lv-EGFP-CSCs group and CSCs group (P<0.05).Meanwhile, transfection with lentiviral vector in each group did not affect the viability of c-kit+CSCs.CONCLUSION:Transfection of Lv-EGFP-CGRP into c-kit+CSCs was successful .The secretion of CGRP was found in the transfected c-kit+CSCs and the viability was not changed after transfection .CGRP-modified c-kit+CSCs may play a role in treating myocardial infarction .
8.Effect of calcitonin gene-related peptide on myocardin expression and phenotypic switch in vascular smooth muscle cells
Xianping LONG ; Wenwen DENG ; Song WANG ; Dongmei WANG ; Jin SHENG ; Bei SHI ; Ranzun ZHAO
Chinese Journal of Pathophysiology 2015;(8):1360-1364
AIM:Toinvestigatetheeffectsofcalcitoningene-relatedpeptide(CGRP)onmyocardinexpres-sion and phenotypic switch in vascular smooth muscle cells ( VSMCs) .METHODS:VSMCs were obtained by aortic tissue adherent culture and treated with angiotensin Ⅱ( AngⅡ) , AngⅡ+CGRP or AngⅡ+CGRP +CGRP8-37 .The protein expression of myocardin and the phenotypic proteins of the VSMCs was detected by Western blot .RESULTS:The expres-sion of myocardin in cultured VSMCs showed downregulation along with time expansion .The protein level of myocardin was higher at 48 h and 72 h than that at baseline in the cultured VSMCs (P<0.05).However, the myocardin was lower at 48 h and 72 h than that at baseline after treatment with CGRP in cultured VSMCs (P<0.05).Furthermore, at 48 h in cul-tured VSMCs, the myocardin decreased along with α-smooth muscle actin (α-SMA) (P<0.05), and osteopontin (OPN) increased (P<0.05) in AngⅡ group compared with control group .After treatment with CGRP, the levels of myocardin andα-SMA become higher ( P <0.05 ) but OPN was lower ( P <0.05 ) in CGRP group than those in AngⅡ group. CGRP8-37 abrogated CGRP-induced increase in myocardin and α-SMA and decrease in OPN in CGRP 8-37 group compared with CGRP group .CONCLUSION: CGRP may regulate the phenotypic switch of the VSMCs and maintain the cells in contractile phenotype through the upregulation of myocardin protein , which may be accomplished by the combination of CGRP and its receptor .
9.Experimental research on the effects of calcitonin gene related peptide on biological characteristics of bone marrow stem cells
Xianping LONG ; Panke CHEN ; Can CUI ; Song WANG ; Bei SHI ; Xiaojian YAO
Chinese Journal of Geriatrics 2015;34(6):671-675
Objective To study the effect of the recombinant Lentivirus containing calcitonin gene related peptide (CGRP) gene on cells biological activity and differentiation of rat bone marrow stem cells(MSCs).Methods Rat MSCs were isolated and cultured by granulocytes adherent.MSCs were transfected with Lenti-EGFP CGRP(MSCsCGRP+/+ group),While MSCs were transfected with Lenti-EGFP as control group.Cell transfection rate was detected by flow cytometry,protein secretion in the above-mentioned MSCsCGRP+ + supernatant was detected using ELISA method.Cells surface markers weare detected by flow cytometry and immunohistochemistry.Trypan blue was used to examin the survive rate,β galactosidase staining was used to examin aging of MSCs transfection,and MTT was used to examine cell vitality.Results At first day after transfecting with Lenti-EGFP-CGRP,fluorescence was not observed by fluorescence microscope,but a small amount of CGRP protein was detected by ELISA in MSCsCGRP+/+ group,at 3 days and 4 days after transfecting with MSCs,strong fluorescence was observed by fluorescence microscope (the cell transfection rates were 77.87% and 79.58%).The CGRP expression was significantly higher in MSCsCGRP+ + group than in control group [(19.53±0.50) pg/ml vs.(3.12±0.00) pg/ml,t=48.964,P<0.01].At three days after transfection with MSCs,CD29 and CD90 expression were significantly higher,as compared with control group,CD31 expression was increased in MSCsCGRP+ /+ group.Seven days after transfection with MSCs,CD31 expression was significantly increased in MSCsCGRP+ + group,vWF expression was significantly increased in MSCsCGRP+ + group after MSCs were transfected with LentiEGFP CGRP for 14 days,but a SMA expression was decreased in MSCsCGRP+ +group.At 3 days and 7 days after transfection with Lenti-EGFP-CGRP,the proliferation,survive and aging showed no difference in MSCsCGRP+/+group and in control group (the proliferation of cell:t=0.253,0.290the survive of cell t=-0.307,0.690,all P>0.05).At 14 days after transfection with Lenti-EGFP-CGRP,aging of cell were decreased in MSCsCGRP+ + group as compared with control group (t=2.446,P< 0.05).Conclusions After MSCs are transfected with Lenti EGFP-CGRP,biological characteristics of MSCs has no significant effects,there is still proliferation and differentiation activity.Cell secretion of CGRP can promote the endothelial cell differentiation,and inhibit the differentiation to smooth muscle cells.The CGRP modification of MSCs may play a role in the regulation of angiogenesis.
10.Lentiviral vector mediated CGRP gene in vitro transfection and its effects on biological properties of MSC
Panke CHEN ; Bei SHI ; Guanxue XU ; Zhijiang LIU ; Xianping LONG ; Wei ZHANG ; Shuai MA
Chongqing Medicine 2015;(14):1873-1875,1878
Objective To explore in vitro mesenchymal stem cell (MSC) transfection of lentiviral vector mediated calcitonin gene‐related peptide(CGRP) gene and its effects on biological properties of MSC .Methods MSC were isolated ,cultured and identi‐fied .MSC were infected by lentivirus encoding recombinant enhanced green fluorescent protein (EGFP) gene and CGRP (Lv‐EG‐FP‐CGRP) .The transfection efficiency was determined by the inverted fluorescence microscope and flow cytometry .The expression levels of CGRP were detected in CGRP‐modified MSC by using real‐time PCR ,immunocytochemistry and enzyme‐linked immu‐nosorbent assay (ELISA) .The proliferation ,aging and differentiation ability of MSC were evaluated by MTT ,β‐galactosidase stai‐ning and inducing differentiation respectively .Results After 48 h of MSC transfection by Lv‐EGFP‐CGRP ,EGFP/CGRP could be expressed stably .When multiplicity of infection (MOI) was 30 ,the transfection efficiency reached more than 80% .Compared with the MSC group and the MSC‐EGFP group ,the mRNA and protein expression levels of CGRP in CGRP‐modified MSC(MSC‐CGRP group) were markedly increased(all P<0 .01) .The results of MTT ,β‐galactosidase staining and inducing differentiation assay dem‐onstrated that the transfected CGRP basically had no effect on the proliferation ,aging and endotheliocyte differerntiation of MSC . Conclusion MSC is a kind of ideal genetic vector cell ,which can serve as the target cell of CGRP gene transduction for the applica‐tion of gene therapy and lays the foundation for follow‐up in vitro and vivo experiments .

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