Objective To investigate the radiation response and proteomic profiling of esophageal epithelial cells cultured under physioxia and normoxia.Methods The human immortalized normal esophageal epithelial cell line SHEE c...Objective To investigate the radiation response and proteomic profiling of esophageal epithelial cells cultured under physioxia and normoxia.Methods The human immortalized normal esophageal epithelial cell line SHEE cells were cultured under normoxia(21%)and physioxia(4%),respectively.A clonogenic assay was performed to evaluate the radiation response of SHEE cells.Cellular proteomic profiling of SHEE cells maintained under physioxia and normoxia was conducted to determine the differentially expressed proteins.Then,the identified differentially expressed proteins were validated by Western blot.Results SHEE cells exposed to normoxia showed an increased radiation response compared to physioxia(irradiation dose≥10Gy,P<0.05).Over 1200 non-redundant proteins were identified in the collected samples.Protein expression was compared between physioxia and normoxia,42 proteins were downregulated and 45 proteins upregulated,in which oxidative phosphorylation was the most significantly enriched pathway.When cells were cultured under normoxia conditions,the induction of antioxidant genes appeared to contribute to form a phenotype adapted to the environment with high oxygen-content.Further analysis validated NRF2,BIP,VCP,SOD1,and YAP1 were the key regulators of this phenotype.Conclusions Compared with physioxia,normoxic cell culture condition can enhance the radiation response.This study could stimulate in vivo microenvironment,and provide a basis for radiation-induced normal tissue damage.展开更多
Background: We demonstrated previously that serum IgG from periodontitis patients interacting with the second extracellular loop of the human cardiac β1 adrenoreceptors triggers the production of second messengers. I...Background: We demonstrated previously that serum IgG from periodontitis patients interacting with the second extracellular loop of the human cardiac β1 adrenoreceptors triggers the production of second messengers. In this paper we quantified the production of nitrates/nitrites and nitric oxide (NO), which in turn induces nitric oxide synthase (NOS) mRNA expression. Methods: We determined using β1 IgG, NOS activity isoforms, NOS expression, nitrate/nitrite assay, cGMP accumulation and PKC activity. Results: We established that serum β1 IgG autoantibodies and NO might be considered as early markers in normoxia/hypoxia system in rat isolated atria. The β1 IgG autoantibodies from periodontitis patients, while stimulating myocardial atria β1 adrenoreceptors, exert an increase on NO levels indirectly quantified as nitrite/nitrate, which acts as NO-storage molecules with significant increase in neuronal NOS (nNOS) and inducible NOS (iNOS) mRNA levels in hypoxia conditions. The significant increase in nNOS/iNOS mRNA and NOS activity as well as in NO levels after short-time hypoxia in rat isolated atria was detected. The expression of these genes are related with the increase in atria dF/dt, cyclic GMP (cGMP) and protein kinase C (PKC) activity and resemble the results obtained by Isoproterenol, an β1 adrenoreceptor agonist. Conclusion: These findings indicate that short-term hypoxia up-regulated rat atria NO/NOS system in the presence of β1 IgG autoantibodies shows that an antibody interacting with rat atria β1 adrenoreceptor can act as expression inducer of proinflammatory NO and its metabolites and that it might be useful and helps to maintain heart function and to prevent necrosis and subsequent loss of heart function during hypoxia.展开更多
基金supported by the Medical Science and Technology Research Program of Henan province (No. LHGJ20210171)Bethune Cancer Radiotherapy Translational Medicine Research Program (No. flzh202115), China.
文摘Objective To investigate the radiation response and proteomic profiling of esophageal epithelial cells cultured under physioxia and normoxia.Methods The human immortalized normal esophageal epithelial cell line SHEE cells were cultured under normoxia(21%)and physioxia(4%),respectively.A clonogenic assay was performed to evaluate the radiation response of SHEE cells.Cellular proteomic profiling of SHEE cells maintained under physioxia and normoxia was conducted to determine the differentially expressed proteins.Then,the identified differentially expressed proteins were validated by Western blot.Results SHEE cells exposed to normoxia showed an increased radiation response compared to physioxia(irradiation dose≥10Gy,P<0.05).Over 1200 non-redundant proteins were identified in the collected samples.Protein expression was compared between physioxia and normoxia,42 proteins were downregulated and 45 proteins upregulated,in which oxidative phosphorylation was the most significantly enriched pathway.When cells were cultured under normoxia conditions,the induction of antioxidant genes appeared to contribute to form a phenotype adapted to the environment with high oxygen-content.Further analysis validated NRF2,BIP,VCP,SOD1,and YAP1 were the key regulators of this phenotype.Conclusions Compared with physioxia,normoxic cell culture condition can enhance the radiation response.This study could stimulate in vivo microenvironment,and provide a basis for radiation-induced normal tissue damage.
文摘Background: We demonstrated previously that serum IgG from periodontitis patients interacting with the second extracellular loop of the human cardiac β1 adrenoreceptors triggers the production of second messengers. In this paper we quantified the production of nitrates/nitrites and nitric oxide (NO), which in turn induces nitric oxide synthase (NOS) mRNA expression. Methods: We determined using β1 IgG, NOS activity isoforms, NOS expression, nitrate/nitrite assay, cGMP accumulation and PKC activity. Results: We established that serum β1 IgG autoantibodies and NO might be considered as early markers in normoxia/hypoxia system in rat isolated atria. The β1 IgG autoantibodies from periodontitis patients, while stimulating myocardial atria β1 adrenoreceptors, exert an increase on NO levels indirectly quantified as nitrite/nitrate, which acts as NO-storage molecules with significant increase in neuronal NOS (nNOS) and inducible NOS (iNOS) mRNA levels in hypoxia conditions. The significant increase in nNOS/iNOS mRNA and NOS activity as well as in NO levels after short-time hypoxia in rat isolated atria was detected. The expression of these genes are related with the increase in atria dF/dt, cyclic GMP (cGMP) and protein kinase C (PKC) activity and resemble the results obtained by Isoproterenol, an β1 adrenoreceptor agonist. Conclusion: These findings indicate that short-term hypoxia up-regulated rat atria NO/NOS system in the presence of β1 IgG autoantibodies shows that an antibody interacting with rat atria β1 adrenoreceptor can act as expression inducer of proinflammatory NO and its metabolites and that it might be useful and helps to maintain heart function and to prevent necrosis and subsequent loss of heart function during hypoxia.
