Effect of Qingyitang on activity of intracellular Ca2+-Mg2+-ATPase in rats with acute pancreatitis

Abstract

Aim: To study the change of intracellular calcium-magnesium ATPase (Ca(2+)-Mg(2+)-ATPase) activity in pancreas, liver and kidney tissues of rats with acute pancreatitis (AP), and to investigate the effects of Qingyitang (QYT) (Decoction for clearing the pancreas) and tetrandrine (Tet) and vitamin E (VitE) on the activity of Ca(2+)-Mg(2+)-ATPase.

Methods: One hundred and five Sprague-Dawley rats were randomly divided into: normal control group, AP group, treatment group with QYT (1 ml/100 g) or Tet (0.4 ml/100 g) or VitE (100 mg/kg). AP model was prepared by a retrograde injection of sodium taurocholate into the pancreatic duct. Tissues of pancreas, liver and kidney of the animals were taken at 1 h, 5 h, 10 h respectively after AP induction, and the activity of Ca(2+)-Mg(2+)-ATPase was studied using enzyme-histochemistry staining. Meanwhile, the expression of Ca(2+)-Mg(2+)-ATPase of the tissues was studied by RT-PCR.

Results: The results showed that the positive rate of Ca(2+)-Mg(2+)-ATPase in AP group (8.3%, 25%, 29.2%) was lower than that in normal control group (100%) in all tissues (P<0.01), the positive rate of Ca(2+)-Mg(2+)-ATPase in treatment group with QYT (58.3%, 83.3%, 83.3%), Tet (50.0%, 70.8%, 75.0%) and VitE (54.2%, 75.0%, 79.2%) was higher than that in AP group (8.3%, 25.0%, 29.2%) in all tissues (P<0.01). RT-PCR results demonstrated that in treatment groups Ca(2+)-Mg(2+)-ATPase gene expression in pancreas tissue was higher than that in AP group at the observing time points, and the expression at 5 h was higher than that at 1 h. The expression of Ca(2+)-Mg(2+)-ATPase in liver tissue was positive, but without significant difference between different groups.

Conclusion: The activity and expression of intracellular Ca(2+)-Mg(2+)-ATPase decreased in rats with AP, suggesting that Ca(2+)-Mg(2+)-ATPase may contribute to the occurrence and development of cellular calcium overload in AP. QYT, Tet and VitE can increase the activity and expression of Ca(2+)-Mg(2+)-ATPase and may relieve intracellular calcium overload to protect the tissue and cells from injuries.

Figure 1

Histopathological findings in AP rats before and after treatment, HE stains. A: Morphological changes in pancreatic tissue of AP rats before treatment, × 200. B: Morphological changes in pancreatic tissue of AP rats after treatment with QYT, × 200. C: Morphological changes in hepatic tissue of AP rats before treatment, × 200. D: Morphological changes in hepatic tissue of AP rats after treatment with VitE, × 200. E: Morphological changes in renal tissue of AP rats before treatment, × 400. F: Morphological changes in hepatic tissue of AP rats after treatment with Tet, × 400.

Figure 2

Enzyme histochemistry staining for intracellular Ca²⁺ -Mg²⁺ -ATPase in AP rats before and after treatment. A: Pancreatic tissue in AP rats before treatment, × 400. B: Pancreatic tissue in AP rats after treatment with QYT, × 400. C: Hepatic tissue in AP rats before treatment, × 200. D: Hepatic tissue in AP rats after treatment with VitE, × 200. E: Renal tissue in AP rats before treatment, × 400. F: Renal tissue in AP rats after treatment with Tet, × 400.

Figure 3

Expression of Ca²⁺ - Mg²⁺ - ATPase in AP rats before and after treatment analyzed by RT-PCR. A: Expression of Ca²⁺ -Mg²⁺ - ATPase mRNA in pancreatic tissue of different groups. B: Expression of Ca²⁺ -Mg²⁺ -ATPase mRNA in hepatic tissue of different groups. C: amplification product of GAPDH gene in pancreatic tissue of different groups. D: amplification product of GAPDH geneinhepatic tissue of different groups.Note: 1: AP 1 h, 2: AP 5 h, 3: QYT + AP 1 h, 4: QYT + AP 5 h, 5: Normal control group, 6: Tet + AP1 h, 7: Tet + AP 5 h, 8: VitE + AP 1 h, 9:VitE + AP 5 h, M: PCR marker.