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目的:研究脂多糖(LPS)对胰腺腺泡细胞钙稳态的影响,及胞浆内钙超载时钙离子的来源,以探讨LPS致胰腺腺泡细胞损伤的机制。方法:胶原酶法分离胰腺腺泡细胞,Fluo-3/AM负载后,在无钙或含生理钙离子浓度的培养液中加入不同浓度LPS,采用激光共聚焦显微镜观察单个胰腺腺泡细胞[Ca~(2+)]_i。另外采用MTT法检测LPS作用的不同时间点胰腺腺泡细胞的活性。结果:LPS可致胰腺腺泡细胞损伤、细胞内[Ca~(2+)]_i显著升高,且呈浓度依赖性(P<0.05):在含1mmol/L依他酸的培养液中,LPS致腺泡细胞损伤程度明显减轻(P<0.05)、仅引起胰腺腺泡细胞[Ca~(2+)]_i缓慢、微弱的升高,而再次恢复培养液中Ca~(2+)至生理浓度,引发快速、幅度更高而持久的[Ca~(2+)]_i变化。腺泡细胞内[Ca~(2+)]_i的变化明显先于腺泡细胞的损伤。结论:LPS导致胰腺腺泡细胞内钙超载,主要源于胞外Ca~(2+)内流。钙超载作为早期的病理事件参与腺泡细胞损伤的发生,钙稳态失衡是LPS致胰腺细胞损伤的主要因素之一。
AIM: To investigate the effect of lipopolysaccharide (LPS) on the calcium homeostasis of pancreatic acinar cells and the source of calcium ions during cytoplasmic calcium overload in order to investigate the mechanism of LPS-induced pancreatic acinar cell injury. METHODS: Pancreatic acinar cells were isolated by collagenase method and loaded with Fluo-3 / AM after different concentrations of LPS in medium without calcium or physiological calcium concentration. Laser confocal microscopy was used to observe the expression of [Ca ~ (2 +)] _ i. In addition, MTT assay was used to detect the activity of pancreatic acinar cells at different time points. Results: LPS induced the injury of acinar cells in the pancreas and significantly increased [Ca ~ (2 +)] i in a concentration-dependent manner (P <0.05). In the medium containing 1 mmol / L etidronate, LPS-induced acinar cell damage was significantly reduced (P <0.05), only caused a slow and weak increase of [Ca ~ (2 +)] _i in the pancreatic acinar cells, and again restored the Ca ~ (2+) Physiological concentration, triggering rapid, higher amplitude and lasting [Ca ~ (2 +)] _i change. The change of [Ca ~ (2 +)] _i in acinar cells was obviously earlier than that of acinar cells. CONCLUSION: LPS leads to overload of calcium in pancreatic acinar cells, mainly due to extracellular Ca ~ (2+) influx. Calcium overload as an early pathological event involved in the occurrence of acinar cell damage, calcium homeostasis is one of the main factors of LPS-induced pancreatic cell damage.