Sharma Deepika, Malik Ankit, Guy Clifford S, Karki Rajendra, Vogel Peter, Kanneganti Thirumala-Devi
Department of Immunology, St. Jude Children's Research Hospital, Memphis, Tennessee.
Animal Resources Center and the Veterinary Pathology Core, St. Jude Children's Research Hospital, Memphis, Tennessee.
Gastroenterology. 2018 Mar;154(4):948-964.e8. doi: 10.1053/j.gastro.2017.11.276. Epub 2017 Dec 2.
BACKGROUND & AIMS: Inflammatory bowel diseases (IBD) increase risk for colorectal cancer. Mutations in the Mediterranean fever gene (MEFV or pyrin) are associated with hereditary autoinflammatory disease and severe IBD. Expression of MEFV, a sensor protein that the initiates assembly of the inflammasome complex, is increased in colon biopsies from patients with IBD. We investigated the role of pyrin in intestinal homeostasis in mice.
Mefv mice and C57/BL6 mice (controls) were given azoxymethane followed by multiple rounds of dextran sodium sulfate (DSS) to induce colitis and tumorigenesis. In some experiments, Mefv mice were given injections of recombinant interleukin 18 (rIL18) or saline (control) during DSS administration. Colon tissues were collected at different time points during colitis development and analyzed by histology, immunohistochemistry, immunoblots, or ELISAs (to measure cytokines). Spleen and mesenteric lymph node were collected, processed, and analyzed by flow cytometry. Colon epithelial permeability was measured in mice with colitis by gavage of fluorescent dextran and quantification of serum levels.
MEFV was expressed in colons of control mice and expression increased during chronic and acute inflammation; high levels were detected in colon tumor and adjacent non-tumor tissues. Mefv-/- mice developed more severe colitis than control mice, with a greater extent of epithelial hyperplasia and a larger tumor burden. Levels of inflammatory cytokines (IL6) and chemokines were significantly higher in colons of Mefv-/- mice than control mice following colitis induction, whereas the level IL18, which depends on the inflammasome for maturation and release, was significantly lower in colons of Mefv-/- mice. Mefv-/- mice had increased epithelial permeability following administration of DSS than control mice, and loss of the tight junction proteins occludin and claudin-2 from intercellular junctions. STAT3 was activated (phosphorylated) in inflamed colon tissues from Mefv-/-, which also had increased expression of stem cell markers (OLFM4, BMI1, and MSI1) compared with colons from control mice. Administration of rIL18 to Mefv-/- mice reduced epithelial permeability, intestinal inflammation, the severity of colitis, and colon tumorigenesis.
In studies with DSS-induced colitis, we found that pyrin (MEFV) is required for inflammasome activation and IL18 maturation, which promote intestinal barrier integrity and prevent colon inflammation and tumorigenesis. Strategies to increase activity of MEFV or IL18 might be developed for the treatment of IBD and prevention of colitis-associated tumorigenesis.
炎症性肠病（IBD）会增加患结直肠癌的风险。地中海热基因（MEFV或吡啉）突变与遗传性自身炎症性疾病和严重IBD相关。MEFV是一种启动炎性小体复合物组装的传感蛋白，在IBD患者的结肠活检组织中表达增加。我们研究了吡啉在小鼠肠道稳态中的作用。
给Mefv小鼠和C57/BL6小鼠（对照）注射氧化偶氮甲烷，随后进行多轮葡聚糖硫酸钠（DSS）处理以诱导结肠炎和肿瘤发生。在一些实验中，在给予DSS期间给Mefv小鼠注射重组白细胞介素18（rIL18）或生理盐水（对照）。在结肠炎发展的不同时间点收集结肠组织，并通过组织学、免疫组织化学、免疫印迹或酶联免疫吸附测定（ELISA）（用于测量细胞因子）进行分析。收集脾脏和肠系膜淋巴结，进行处理并通过流式细胞术分析。通过灌胃荧光葡聚糖并定量血清水平来测量结肠炎小鼠的结肠上皮通透性。
MEFV在对照小鼠的结肠中表达，并且在慢性和急性炎症期间表达增加；在结肠肿瘤和相邻的非肿瘤组织中检测到高水平表达。Mefv-/-小鼠比对照小鼠发生更严重的结肠炎，上皮增生程度更大，肿瘤负荷更大。诱导结肠炎后，Mefv-/-小鼠结肠中的炎性细胞因子（IL6）和趋化因子水平显著高于对照小鼠，而依赖炎性小体成熟和释放的IL18水平在Mefv-/-小鼠结肠中显著降低。给予DSS后，Mefv-/-小鼠的上皮通透性比对照小鼠增加，并且细胞间连接处的紧密连接蛋白occludin和claudin-2丢失。STAT-3在Mefv-/-小鼠发炎的结肠组织中被激活（磷酸化），与对照小鼠的结肠相比，其干细胞标志物（OLFM4、BMI1和MSI1）的表达也增加。给Mefv-/-小鼠注射rIL18可降低上皮通透性、肠道炎症、结肠炎的严重程度和结肠肿瘤发生。
在DSS诱导的结肠炎研究中，我们发现炎性小体激活和IL18成熟需要吡啉（MEFV），其可促进肠道屏障完整性并预防结肠炎症和肿瘤发生。或许可以开发提高MEFV或IL18活性的策略来治疗IBD和预防结肠炎相关的肿瘤发生。
