Department of Gastroenterology and Shanghai Key Laboratory of Pancreatic Disease, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Division of Pediatric Gastroenterology, University of Pittsburgh School of Medicine and the Children's Hospital of Pittsburgh of University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania.
Gastroenterology. 2020 Sep;159(3):1036-1050.e8. doi: 10.1053/j.gastro.2020.05.051. Epub 2020 May 20.
BACKGROUND & AIMS: Calcineurin is a ubiquitously expressed central Ca-responsive signaling molecule that mediates acute pancreatitis, but little is known about its effects. We compared the effects of calcineurin expression by hematopoietic cells vs pancreas in mouse models of pancreatitis and pancreatitis-associated lung inflammation.
We performed studies with mice with hematopoietic-specific or pancreas-specific deletion of protein phosphatase 3, regulatory subunit B, alpha isoform (PPP3R1, also called CNB1), in mice with deletion of CNB1 (Cnb1) and in the corresponding controls for each deletion of CNB1. Acute pancreatitis was induced in mice by administration of caerulein or high-pressure infusion of radiocontrast into biliopancreatic ducts; some mice were also given intraductal infusions of an adeno-associated virus vector that expressed nuclear factor of activated T -cells (NFAT)-luciferase into pancreas. Pancreas, bone marrow, liver, kidney, heart, and lung were collected and analyzed by histopathology, immunohistochemistry, and immunoblots; levels of cytokines were measured in serum. Mouse and human primary pancreatic acinar cells were transfected with a vector that expressed NFAT-luciferase and incubated with an agent that blocks interaction of NFAT with calcineurin; cells were analyzed by immunofluorescence. Calcineurin-mediated neutrophil chemotaxis and reactive oxygen species production were measured in neutrophils from mice.
Mice with hematopoietic-specific deletion of CNB1 developed the same level of local pancreatic inflammation as control mice after administration of caerulein or infusion of radiocontrast into biliopancreatic ducts. Cnb1 mice or mice with pancreas-specific deletion of CNB1 developed less severe pancreatitis and reduced pancreatic inflammation after administration of caerulein or infusion of radiocontrast into biliopancreatic ducts compared with control mice. NFAT was activated in pancreas of Swiss Webster mice given caerulein or infusions of radiocontrast into biliopancreatic ducts. Blocking the interaction between calcineurin and NFAT did not reduce pancreatic acinar cell necrosis in response to caerulein or infusions of radiocontrast. Mice with hematopoietic-specific deletion of CNB1 (but not mice with pancreas-specific deletion of CNB1) had reduced infiltration of lung tissues by neutrophils. Neutrophil chemotaxis and production of reactive oxygen species were decreased after incubation with a calcineurin inhibitor.
Hematopoietic and neutrophil expression of calcineurin promotes pancreatitis-associated lung inflammation, whereas pancreatic calcineurin promotes local pancreatic inflammation. The findings indicate that the protective effects of blocking or deleting calcineurin on pancreatitis are mediated by the source of its expression. This information should be used in the development of strategies to inhibit calcineurin for the prevention of pancreatitis and pancreatitis-associated lung inflammation.
钙调神经磷酸酶是一种广泛表达的中央 Ca 反应信号分子，介导急性胰腺炎，但对其作用知之甚少。我们比较了钙调神经磷酸酶在胰腺炎和胰腺炎相关肺炎症的小鼠模型中通过造血细胞与胰腺表达的作用。
我们对具有造血特异性或胰腺特异性蛋白磷酸酶 3、调节亚基 B、α 异构体（PPP3R1，也称为 CNB1）缺失的小鼠、CNB1（Cnb1）缺失的小鼠以及每种 Cnb1 缺失的相应对照小鼠进行了研究。通过给予蛙皮素或放射性对比剂高压输注到胆胰管内，在小鼠中诱导急性胰腺炎；一些小鼠还给予腺相关病毒载体的胰腺内导管内输注，该载体表达核因子活化 T 细胞（NFAT）-荧光素酶。收集胰腺、骨髓、肝脏、肾脏、心脏和肺组织，进行组织病理学、免疫组织化学和免疫印迹分析；测量血清细胞因子水平。用表达 NFAT-荧光素酶的载体转染小鼠和人原代胰腺腺泡细胞，并与一种阻断 NFAT 与钙调神经磷酸酶相互作用的试剂孵育；通过免疫荧光分析细胞。测量从小鼠中性粒细胞中钙调神经磷酸酶介导的中性粒细胞趋化和活性氧产生。
给予蛙皮素或放射性对比剂输注到胆胰管内后，具有造血特异性 CNB1 缺失的小鼠与对照小鼠一样，胰腺局部炎症程度相同。与对照小鼠相比，Cnb1 小鼠或胰腺特异性 CNB1 缺失的小鼠在给予蛙皮素或放射性对比剂输注到胆胰管内后，胰腺炎程度较轻，胰腺炎症减轻。给予蛙皮素或放射性对比剂输注到胆胰管内后，瑞士 Webster 小鼠的 NFAT 在胰腺中被激活。阻断钙调神经磷酸酶与 NFAT 之间的相互作用并没有减少对蛙皮素或放射性对比剂输注的胰腺腺泡细胞坏死的反应。具有造血特异性 CNB1 缺失的小鼠（但不是胰腺特异性 CNB1 缺失的小鼠）肺组织中中性粒细胞浸润减少。用钙调神经磷酸酶抑制剂孵育后，中性粒细胞趋化和活性氧的产生减少。
造血细胞和中性粒细胞表达的钙调神经磷酸酶促进胰腺炎相关的肺炎症，而胰腺钙调神经磷酸酶促进局部胰腺炎症。这些发现表明，阻断或删除钙调神经磷酸酶对胰腺炎的保护作用是由其表达的来源介导的。这些信息应在开发抑制钙调神经磷酸酶预防胰腺炎和胰腺炎相关肺炎症的策略中加以利用。
