Foong Daphne, Mikhael Meena, Zhou Jerry, Zarrouk Ali, Liu Xiaodong, Schröder Jan, Polo Jose M, Ho Vincent, O'Connor Michael D
Regenerative Medicine Laboratory, School of Medicine, Western Sydney University, Campbelltown, NSW, Australia.
South West Surgery, Campbelltown Private Hospital, Campbelltown, NSW, Australia.
J Neurogastroenterol Motil. 2023 Apr 30;29(2):238-249. doi: 10.5056/jnm22078.
BACKGROUND/AIMS: Interstitial cells of Cajal (ICC) are specialized gastrointestinal (GI) pacemaker cells required for normal GI motility. Dysfunctions in ICC have been reported in patients with GI motility disorders, such as gastroparesis, who exhibit debilitating symptoms and greatly reduced quality of life. While the proteins, calcium-activated chloride channel anoctamin-1 (ANO1) and the receptor tyrosine kinase (KIT), are known to be expressed by human ICC, relatively little is known about the broad molecular circuitry underpinning human ICC functions. The present study therefore investigates the transcriptome and proteome of ANO1-expressing, KIT/CD45/CD11B ICC obtained from primary human gastric tissue.
Excess human gastric tissue resections were obtained from sleeve gastrectomy patients. ICC were purified using fluorescence-activated cell sorting (FACSorting). Then, ICC were characterized by using immunofluorescence, real-time polymerase chain reaction, RNA-sequencing and mass spectrometry.
Compared to unsorted cells, real-time polymerase chain reaction showed the KIT/CD45/CD11B ICC had: a 9-fold ( < 0.05) increase in ANO1 expression; unchanged KIT expression; and reduced expression for genes associated with hematopoietic cells (CD68, > 10-fold, < 0.001) and smooth muscle cells (DES, > 4-fold, < 0.05). RNA-sequencing and gene ontology analyses of the KIT/CD45/CD11B cells revealed a transcriptional profile consistent with ICC function. Similarly, mass spectrometry analyses of the KIT/CD45/CD11B cells presented a proteomic profile consistent with ICC activities. STRING-based protein interaction analyses using the RNA-sequencing and proteomic datasets predicted protein networks consistent with ICC-associated pacemaker activity and ion transport.
These new and complementary datasets provide a valuable molecular framework for further understanding how ICC pacemaker activity regulates smooth muscle contraction in both normal GI tissue and GI motility disorders.
背景/目的： Cajal间质细胞（ICC）是正常胃肠（GI）蠕动所需的特殊胃肠起搏器细胞。据报道，胃肠蠕动障碍患者（如胃轻瘫患者）的ICC功能存在异常，这些患者表现出使人衰弱的症状，生活质量大幅下降。虽然已知蛋白质、钙激活氯通道anoctamin-1（ANO1）和受体酪氨酸激酶（KIT）由人ICC表达，但对于支撑人ICC功能的广泛分子机制了解相对较少。因此，本研究调查了从原发性人胃组织中获得的表达ANO1、KIT/CD45/CD11B的ICC的转录组和蛋白质组。
从袖状胃切除术患者中获取多余的人胃组织切除标本。使用荧光激活细胞分选（FACSorting）纯化ICC。然后，通过免疫荧光、实时聚合酶链反应、RNA测序和质谱对ICC进行表征。
与未分选的细胞相比，实时聚合酶链反应显示KIT/CD45/CD11B ICC具有：ANO1表达增加9倍（<0.05）；KIT表达不变；与造血细胞相关的基因（CD68，>10倍，<0.001）和平滑肌细胞相关的基因（DES，>4倍，<0.05）表达降低。对KIT/CD45/CD11B细胞进行RNA测序和基因本体分析，揭示了与ICC功能一致的转录谱。同样，对KIT/CD45/CD11B细胞进行质谱分析，呈现了与ICC活性一致的蛋白质组学图谱。使用RNA测序和蛋白质组学数据集进行的基于STRING的蛋白质相互作用分析预测了与ICC相关的起搏器活性和离子转运一致的蛋白质网络。
这些新的互补数据集为进一步了解ICC起搏器活性如何调节正常胃肠组织和胃肠蠕动障碍中的平滑肌收缩提供了有价值的分子框架。
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