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首页> 外文期刊>American Journal of Physiology >Calcium signaling in cultured human and rat duodenal enterocytes.
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Calcium signaling in cultured human and rat duodenal enterocytes.

机译：培养的人和大鼠十二指肠肠上皮细胞中的钙信号传导。

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摘要

Vagal stimuli increase duodenal mucosal HCO-3 secretion and may provide anticipatory protection against acid injury, but duodenal enterocyte (duodenocyte) responses and cholinoceptor selectivity have not been defined. We therefore developed a stable primary culture model of duodenocytes from rats and humans. Brief digestion of scraped rat duodenal mucosa or human biopsies with collagenase/dispase yielded cells that attached to the extracellular matrix Matrigel within a few hours of plating. Columnar cells with villus enterocyte morphology that exhibited spontaneous active movement were evident between 1 and 3 days of culture. Rat duodenocytes loaded with fura 2 responded to carbachol with a transient increase in intracellular calcium concentration ([Ca2+]i), with an apparent EC50 of approximately 3 microM. In a first type of signaling pattern, [Ca2+]i returned to basal or near basal values within 3-5 min. In a second type, observed in cells with enlarged vacuoles characteristic of crypt cell morphology, the initial transient increase was followed by rhythmic oscillations. Human duodenocytes responded with a more sustained increase in [Ca2+]i, and oscillations were not observed. Rat as well as human duodenocytes also responded to CCK-octapeptide but not to vasoactive intestinal polypeptide. Equimolar concentrations (100 nM) of the subtype-independent muscarinic antagonist atropine and the M3 antagonist 4-diphenylacetoxy-N-methylpiperidine methiodide prevented the response to 10 microM carbachol, whereas the M1 antagonist pirenzepine and the M2 antagonists methoctramine and AF-DX 116BS had no effect at similar concentrations. Responses in rat and human duodenocytes were similar. A new agonist-sensitive primary culture model for rat and human duodenocytes has thus been established and the presence of enterocyte CCK and muscarinic M3 receptors demonstrated.
机译：迷走神经刺激可增加十二指肠粘膜HCO-3的分泌，并可能提供针对酸损伤的预期保护，但尚未定义十二指肠肠上皮细胞（十二指肠细胞）的反应和胆碱受体选择性。因此，我们开发了一种稳定的来自大鼠和人类十二指肠细胞的原代培养模型。用胶原酶/分散酶对刮下的大鼠十二指肠粘膜或人活检组织进行简要消化，可产生在铺板后数小时内附着于细胞外基质基质胶的细胞。在培养的1至3天之间，具有绒毛肠细胞形态的柱状细胞表现出自发的主动运动。装有呋喃2的大鼠十二指肠细胞对卡巴胆碱的反应是细胞内钙浓度（[Ca2 +] i）短暂增加，表观EC50约为3 microM。在第一类信号模式中，[Ca 2+] 1在3-5分钟内恢复至基础值或接近基础值。在第二种类型中，在隐窝细胞形态特征的液泡增大的细胞中观察到，最初的瞬时增加之后是有节奏的振荡。人类十二指肠细胞的[Ca2 +] i持续增加，并且未观察到振荡。大鼠和人十二指肠细胞对CCK-八肽也有反应，但对血管活性肠多肽无反应。亚摩尔浓度的毒蕈碱拮抗剂阿托品和M3拮抗剂4-二苯基乙酰氧基-N-甲基哌啶甲硫醇的等摩尔浓度（100 nM）阻止了对10 microM卡巴胆碱的反应，而M1拮抗剂哌仑西平和M2拮抗剂甲辛辛和AF-DX 116BS具有在相似浓度下无影响。大鼠和人十二指肠细胞的反应相似。因此，已经建立了一种新的对大鼠和人十二指肠细胞激动剂敏感的原代培养模型，并证明了肠上皮细胞CCK和毒蕈碱M3受体的存在。

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