Apoptosis And Mitochondrial Functions In Children's Circularing Lymphocytes And Neutrophils Undergoing Surgery And General Anesthesia

Surgery and anesthesia cause immune surpression showing a reduction in the numbers of circulating lymphocytes and the dysfunctions of PMN.The recent studies have demonstrated that an unregulated activation of the process of apoptosis is connected with the cellular immune surpression and mitochondrias play an important role in the events of apoptosis. We choose children who underwent elective surgery and general anesthesia to investigate the apoptosis and mitochondrial membrane potential in children's circulating lymphocytes and neutrophils.Part one: Apoptosis and mitochondrial functions in children's circulating lymphocytes undergoing surgery and general anesthesiaObjective: To investigate the apoptosis and mitochondrial functions in children's circulating lymphocytes undergoing surgery and general anesthesia.Methods: 16 children who underwent elective surgery and general anesthesia were studied.Blood sampling for assessment of lymphocytes apoptosis and mitochondrial membrane potential were performed on the day before surgery (t0) and at 24 and 72 hours after operation (t| and t2 respectively).After isolated, the lymphocytes were maintained in RPMI- 1640 (10% heat-inactivated fetal calf serum, 1 mmol/L of glutamine, and 1%penicillin with streptomycin) at a concentration of 1 * 105 cells/ml for 15 hours. Then,the short-termcultured cells were stained with 7-amino-actinomycin D (7-AAD) to quantify the apoptosis and the cationic lipophilic fluorochrome 3,3'-dihexyloxacarbo-cyanine iodide [DioC6(3)] to analy mitochondrial membrane potential by a FACScan flow cytometer.Results: A significantly increased frequency of apoptotic CD4+ and CD8+ cells was observed at tj measurement (1.97 0.45 and 2.47 0.27 for CD4+ and CD8+ respectively),compared with preoperative values (3.50 0.42 and 4.26 0.53,P<0.05).At t2 measurement, they recovered to the preoperative levels (1.67 0.22,2.70 0.38,P>0.05),which incorporated higher levels of 3,3'-dihexyloxacarbo-cyanine iodide compared with preoperative values (503.31+ 59.43 and 381.67 + 61.15,vs 700.41+ 59.12,645.86 + 69.58.P<0.05). At t2 measurement, they reduced to the preoperative levels (521.62 + 68.05 and 431.07 +79.26, P>0.05) .Part two: Apoptosis and mitochondrial functions in children's circulating neutrophils undergoing surgery and general anesthesiaObjective: To investigate the apoptosis and mitochondrial functions in children's circulating neutrophils undergoing surgery and general anesthesia.Methods: 16 children who underwent elective surgery and general anesthesia were studied.Blood sampling for assessment of neutrophils apoptosis and mitochondrial membrane potential were performed on the day before surgery (to) and at 24 and 72 hours after operation (t1 and t2)respectively.After isolated, the neutrophils were maintained in RPMI- 1640 (10% heat-inactivated fetal calf serum, Immol/L of glutamine, and 1% penicillin with streptomycin) at a concentration of IxlO5 cells/ml for 15 hours.Then ,the short-termcultured cells were stained with 7-amino-actinomycin D (7-AAD) to quantify the apoptosis and the cationic lipophilic fluorochrome 3,3'-dihexyloxacarbocy-anine iodide [DioC6(3)] to analy mitochondrial membrane potential by a FACScan flow cytometer.Results: A significantly decreased frequency of apoptotic cells was observed at t, measurement (13.27 1.90 vs 7.55 1.26, P<0.05).At 72 h following surgery it was still decreased compared to that preoperative values (5.47 0.71,P<0.05),which incorporated lower levels of 3,3'-dihexyloxacarbo-cyanine iodide compared with preoperative values (46.08 7.80 vs 30.72 4.23, P<0.05). At 72 h following surgery it was still decreased compared to that preoperatively.( 29.63 5.98, P<0.05).Conclusions: 1) Surgical trauma is associated with a significant but transient increase in lymphocyte commitment to apoptosis with higher levels of 3,3'-dihexyloxacarbo-cyanine iodide which indicate more cells' mitochondrial membrane potential reduced .This induce a remarkable alteration in the human immune defense sys...