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Objective:To investigate the relationship between the mechanism of ulinastatin reducing perioperative myocardial injury and ferroptosis in peripheral blood mononuclear cells (PBMCs) in pediatric patients undergoing heart surgery under cardiopulmonary bypass (CPB).Methods:A total of 60 pediatric patients of either sex, aged 4-8 yr, of American Association of Anesthesiologists physical status Ⅱ or Ⅲ, undergoing elective repair of ventricular septal defect under CPB, were divided into 2 groups by a random number table method: control group (C group) and ulinastatin group (UTI group), with 30 cases in each group.Combined intravenous-inhalational anesthesia was used.In UTI group, ulinastatin 20 000 U/kg was diluted to 100 ml in normal saline, 50 ml was infused through the central vein over 15 min starting from 20 min before skin incision, and the remaining 50 ml was instilled through the CPB pipeline over 15 min starting from 10 min of CPB.The equal volume of normal saline was given instead in C group.Blood samples from the internal jugular vein were collected after anesthesia induction and before skin incision (T 1), at 30 min after start of CPB (T 2), immediately after termination of CPB (T 3) and at 24 h after termination of CPB (T 4) for determination of the levels of amino-terminal B-type pro-brain natriuretic peptide (NT-proBNP), cardiac troponin I (cTnI) and creatine kinase isoenzymes (CK-MB) in plasma by enzyme-linked immunosorbent assay.PBMCs were extracted by modified Ficoll density gradient centrifugation method for determination of the concentrations of Fe 2+ and malondialdehyde (MDA) and activity of superoxide dismutase (SOD) in PBMCs (by colorimetric method) and expression of long-chain acyl-CoA synthase 4 (ACSL4) and glutathione peroxidase 4 (GPX4) in PBMCs (by Western blot). Results:Compared with the baseline at T 1, the levels of NT-proBNP, cTnI and CK-MB in plasma were significantly increased, the concentrations of Fe 2+ and MDA in PBMCs were increased, the expression of ACSL4 in PBMCs was up-regulated, and the activity of SOD was decreased, and the expression of GPX4 was down-regulated at T 2-4 in two groups ( P<0.05). Compared with C group, the plasma levels of NT-proBNP, cTnI and CK-MB were significantly decreased, the concentrations of Fe 2+ and MDA in PBMCs were decreased, the expression of ACSL4 in PBMCs was down-regulated, the activity of SOD was increased, and the expression of GPX4 was up-regulated at T 2-4 in UTI group ( P<0.05). Conclusion:The mechanism by which ulinastatin reduces perioperative myocardial injury may be related to inhibition of ferroptosis in PBMCs in the pediatric patients undergoing open heart surgery under CPB.
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Objective@#To study the effects of patient-controlled intravenous analgesia(PCIA) with Dexmedetomidine, Ropivacaine by local incision infiltration and combined analgesia on inflammatory factors and rapid rehabilitation in infants aged 3-6 months after radical resection of megacolon.@*Methods@#From June 2016 to March 2018, 90 infants aged 3-6 months underwent radical resection of megacolon in Henan Provincial Children′s Hospital as the subjects.According to the postoperative analgesia mode applied to the children, they were divided into Dexmedetomidine group, Ropivacaine group and combined group (Dexmedetomidine PCIA combined with local infiltration of Ropivacaine), 30 cases in each group.The serum levels of interleukin(IL)-6 and IL-10 were measured and recorded 2 hours before operation and 24 hours after operation, and the analgesic scores of 4, 8, 12 and 24 hours after operation were recorded.The rapid recovery index of each group, standard and the number of complications were compared.@*Results@#Compared with 2 hours before operation, the serum levels of IL-6 in the Dexmedetomidine group, Ropivacaine group and combined group [(24.61±1.44) ng/L vs.(13.84±0.65) ng/L, (26.39±1.29) ng/L vs.(13.85±0.67) ng/L, (20.58±2.06) ng/L vs.(13.87±0.63) ng/L], IL-10[(27.63±1.52) ng/L vs.(15.79±1.48) ng/L, (28.29±2.34) ng/L vs.(15.41±1.37) ng/L, (23.21±2.71) ng/L vs.(15.47±1.52) ng/L] were significantly higher (all P<0.05), while the levels of IL-6, IL-10 in combined group were lower than those in Dexmedetomidine group and Ropivacaine group (ta=9.172, 9.835; tb=10.221, 11.034, all P<0.05). At 4 h, 8 h, 12 h, 24 h after operation.The analgesic scores in combined group were(1.89±0.23) scores, (1.87±0.15) scores, (1.95±0.17) scores, (2.08±0.18) scores, those of dexmedetomidine group were(2.06±0.24) scores, (2.08±0.18) scores, (2.76±0.29) scores, (2.55±0.31) scores, those of Ropivacaine group were(2.10±0.26) scores, (2.15±0.26) scores, (2.32±0.19) scores, (3.00±0.28) scores, and the analgesic scores in combined group were significantly lower than those in Dexmedetomidine group and Ropivacaine group(ta=8.526, 9.364, 10.287, 9.521; tb=8.729, 9.514, 11.037, 9.184, all P<0.05). In comparison with the first exhaust time, anal extubation time, hospitalization days, hospitalization expenses and antibiotic use time in the three groups, it was found that combined group was significantly better than Dexmedetomidine group and Ropivacaine group(ta=8.315, 8.294, 7.883, 9.261, 10.487; tb=8.582, 8.329, 7.916, 9.348, 10.862, all P<0.05). The incidence of complications in the combined group, Dexmedetomidine group, and Ropivacaine group were not significantly different in 3 groups(χ2=0.577, P=0.749).@*Conclusions@#The continuous anesthesia of Dexmedetomidine combined with local infiltration of Ropivacaine incision can alleviate the postoperative inflammatory reaction and promote the rapid recovery of children.
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Objective@#To evaluate the role of necroptosis in hyperoxia-induced acute lung injury (ALI) in preadolescent rats.@*Methods@#A total of 72 clean-grade healthy male Sprague-Dawley rats, aged 14 days, weighing 40-50 g, were divided into 3 groups (n=24 each) by using a random number table method: control group (group C), hyperoxia-induced ALI group (group ALI) and hyperoxia-induced ALI and necrostatin-1 group (group ALI+ N). The rats of group ALI+ N was intraperitoneally injected with necrostatin-1 1.0 mg/kg once a day for 3 consecutive days.The rats were intraperitoneally injected with dimethyl sulfoxide 0.2 ml/kg once a day for 3 consecutive days in C and ALI groups.The animals were sacrificed at 72 h after inhaling oxygen, and bronchoalveolar lavage fluid (BALF) was collected for determination of interleukin-6 (IL-6) and IL-8 concentrations (by enzyme-linked immunosorbent assay), superoxide dismutase (SOD) activity (by xanthine oxidase method), and malondialdehyde (MDA) concentration (by thiobarbituric acid method). Lung tissues were taken for measurement of wet/dry weight ratio (W/D ratio) and for examination of the pathological changes (with a light microscope) and ultrastructure of lung tissues (with an electron microscope). The injured alveolus rate (IAR) was calculated.The expression of receptor-interacting protein kinase 1 (RIPK1), RIPK3 and mixed-lineage kinase domain-like protein (MLKL) in lung tissues was detected by Western blot.@*Results@#Compared with group C, the concentrations of IL-6, IL-8 and MDA in BALF were significantly increased, the activity of SOD in BALF was decreased, the W/D ratio and IAR of lung tissues were increased, the expression of RIPK1, RIPK3 and MLKL in lung tissues was up-regulated (P<0.05), and the pathological damage was accentuated in group ALI.Compared with group ALI, the concentrations of IL-6, IL-8 and MDA in BALF were significantly deceased, the activity of SOD in BALF was increased, the W/D ratio and IAR of lung tissues were decreased, the expression of RIPK1, RIPK3 and MLKL in lung tissues was down-regulated (P<0.05), and the pathological damage was significantly attenuated in group ALI+ N.@*Conclusion@#Necroptosis is involved in the pathophysiological process of hyperoxia-induced ALI in preadolescent rats.
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Objective To study the effects of patient-controlled intravenous analgesia( PCIA)with Dexme-detomidine,Ropivacaine by local incision infiltration and combined analgesia on inflammatory factors and rapid rehabili-tation in infants aged 3-6 months after radical resection of megacolon. Methods From June 2016 to March 2018,90 infants aged 3-6 months underwent radical resection of megacolon in Henan Provincial Children＇s Hospital as the sub-jects. According to the postoperative analgesia mode applied to the children,they were divided into Dexmedetomidine group,Ropivacaine group and combined group( Dexmedetomidine PCIA combined with local infiltration of Ropiva-caine),30 cases in each group. The serum levels of interleukin( IL)-6 and IL-10 were measured and recorded 2 hours before operation and 24 hours after operation,and the analgesic scores of 4,8,12 and 24 hours after operation were recorded. The rapid recovery index of each group,standard and the number of complications were compared. Results Compared with 2 hours before operation,the serum levels of IL-6 in the Dexmedetomidine group,Ropiva-caine group and combined group[(24. 61 ± 1. 44)ng/L υs.(13. 84 ± 0. 65)ng/L,(26. 39 ± 1. 29)ng/L υs.(13. 85 ± 0. 67)ng/L,(20. 58 ± 2. 06)ng/L υs.(13. 87 ± 0. 63)ng/L],IL-10[(27. 63 ± 1. 52)ng/L υs.(15. 79 ± 1. 48) ng/L,(28. 29 ± 2. 34)ng/L υs.(15. 41 ± 1. 37)ng/L,(23. 21 ± 2. 71)ng/L υs.(15. 47 ± 1. 52)ng/L]were signifi-cantly higher(all P<0. 05),while the levels of IL-6,IL-10 in combined group were lower than those in Dexmedeto-midine group and Ropivacaine group(ta =9. 172,9. 835;tb =10. 221,11. 034,all P<0. 05). At 4 h,8 h,12 h,24 h after operation. The analgesic scores in combined group were(1. 89 ± 0. 23)scores,(1. 87 ± 0. 15)scores,(1. 95 ± 0. 17)scores,(2. 08 ± 0. 18)scores,those of dexmedetomidine group were(2. 06 ± 0. 24)scores,(2. 08 ± 0. 18) scores,(2. 76 ± 0. 29)scores,(2. 55 ± 0. 31)scores,those of Ropivacaine group were(2. 10 ± 0. 26)scores,(2. 15 ± 0. 26)scores,(2. 32 ± 0. 19)scores,(3. 00 ± 0. 28)scores,and the analgesic scores in combined group were signifi-cantly lower than those in Dexmedetomidine group and Ropivacaine group( ta =8. 526,9. 364,10. 287,9. 521;tb =8. 729,9. 514,11. 037,9. 184,all P<0. 05). In comparison with the first exhaust time,anal extubation time,hospitali-zation days,hospitalization expenses and antibiotic use time in the three groups,it was found that combined group was significantly better than Dexmedetomidine group and Ropivacaine group(ta =8. 315,8. 294,7. 883,9. 261,10. 487;tb =8. 582,8. 329,7. 916,9. 348,10. 862,all P<0. 05). The incidence of complications in the combined group,Dexme-detomidine group,and Ropivacaine group were not significantly different in 3 groups( χ2 = 0. 577,P = 0. 749 ). Conclusions The continuous anesthesia of Dexmedetomidine combined with local infiltration of Ropivacaine incision can alleviate the postoperative inflammatory reaction and promote the rapid recovery of children.
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Objective To provide experimental materials for exploring the function of breast cancer susceptibility gene 2(BRCA2) gene by establishing of an mouse immortalized mammary gland epithelial cell line as a cell model system.Methods In this study,the primary mouse mammary epithelial cells were isolated and purified by enzyme digestion and differential centrifugation.The resulting primary mouse mammary gland epithelial cells were transducted with lentivirus expressing human telomerase (hTERT) gene,and screened by hygromycin B.The surviving epithelial cells were further characterized by morphology observations and cytokeratin marker detection.Results Immortalized mouse mammary epithelial cells were obtained by infection of MMECs with lentivirus expressing human telomerase(hTERT) gene,screened by hygromycin B.Morphology observations and detection of the cytokeratin marker showed the immortalized MMECs had a typical morphology of luminal epithelial cells with strong expression of cytokeratin 14.Conclusion An immortalized mouse luminal epithelial cell line is successfully established with an uniform morphology,which provides a cell model system for the future exploration on BRCA2-related tumorigenesis.
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Objective To evaluate the role of necroptosis in hyperoxia-induced acute lung injury(ALI)in preadolescent rats.Methods A total of 72 clean-grade healthy male Sprague-Dawley rats,aged 14 days,weighing 40-50 g,were divided into 3 groups(n＝24 each)by using a random number table method: control group(group C),hyperoxia-induced ALI group(group ALI)and hyperoxia-induced ALI and necrostatin-1 group(group ALI+N).The rats of group ALI+N was intraperitoneally injected with ne-crostatin-1 1.0 mg/kg once a day for 3 consecutive days.The rats were intraperitoneally injected with dime-thyl sulfoxide 0.2 ml/kg once a day for 3 consecutive days in C and ALI groups.The animals were sacrificed at 72 h after inhaling oxygen,and bronchoalveolar lavage fluid(BALF)was collected for determination of interleukin-6(IL-6)and IL-8 concentrations(by enzyme-linked immunosorbent assay),superoxide dis-mutase(SOD)activity(by xanthine oxidase method),and malondialdehyde(MDA)concentration(by thiobarbituric acid method).Lung tissues were taken for measurement of wet/dry weight ratio(W/D ratio)and for examination of the pathological changes(with a light microscope)and ultrastructure of lung tissues(with an electron microscope).The injured alveolus rate(IAR)was calculated.The expression of recep-tor-interacting protein kinase 1(RIPK1),RIPK3 and mixed-lineage kinase domain-like protein(MLKL)in lung tissues was detected by Western blot.Results Compared with group C,the concentrations of IL-6,IL-8 and MDA in BALF were significantly increased,the activity of SOD in BALF was decreased,the W/D ratio and IAR of lung tissues were increased,the expression of RIPK1,RIPK3 and MLKL in lung tis-sues was up-regulated(P＜0.05),and the pathological damage was accentuated in group ALI.Compared with group ALI,the concentrations of IL-6,IL-8 and MDA in BALF were significantly deceased,the ac-tivity of SOD in BALF was increased,the W/D ratio and IAR of lung tissues were decreased,the expres-sion of RIPK1,RIPK3 and MLKL in lung tissues was down-regulated(P＜0.05),and the pathological damage was significantly attenuated in group ALI+N.Conclusion Necroptosis is involved in the patho-physiological process of hyperoxia-induced ALI in preadolescent rats.
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VP1 gene encoded by the newly identified caprine enterovirus CEV-JL 14 was amplified and cloned to prokaryotic expression vector pGEX-4T-1.Recombinant GST-VP1 fusion protein was expressed,purified,emulsified with Freund's complete adjuvant and used to immunize the BALB/c mice following a standard procedure.The spleen cells from immunized mice were collected and fused with myeloma cells after its antibody titer reached over 104 times detected by indirect ELISA.Hybridoma cell clones secreting monoclonal antibodies against VP1 were screened and their stability and specificity were further determined.The identified hybridoma cells were injected to mice intraperitoneally and ascites were collected at 7 DPI.Isotypes of the monoclonal antibodies against the recombinant VP1 protein were characterized to be either IgG1 or IgG2b,which showed a high specificity for detection of caprine enterovirus antigens by immunoperoxidase monolayer assay,thus laying a solid basis for future study related to viral pathogenesis,detection and diagnostics for caprine enterovirus infection.