Parish W E
Environmental Safety Laboratory, Unilever Research, Colworth House, Sharnbrook, Bedford MK44 1LQ, UK.
Toxicol In Vitro. 1990;4(4-5):231-41. doi: 10.1016/0887-2333(90)90056-y.
Release of mediators is a manifestation of tissue stimulation or damage applicable to in vitro toxicological investigations. Intercell mediators, or cytokines, are released on tissue stimulation or as a result of mild damage. Intracell mediators, or enzymes of respiration and metabolism, released from damaged cells, participate in inflammatory change and are the usual mediator determinants in in vitro tests. Cells examined for in vitro mediator release are mast cells, which are applicable to anaphylactic allergy and acute inflammation, macrophages, which are particularly applicable to toxicity of inhalable dusts, and fibroblasts/fibrocytes, which are widely used as a general model for cytotoxicity. In tests on organs, lungs may be perfused to identify cytokines released by test substances, which may be used to distinguish between effects due to allergy and those resulting from chemically-induced inflammation. Skin, both human and animal, as full depth or keratome slices of epidermis, has an application in the detection of irritant and corrosive substances. Weak irritants are identified by release of enzymes, for example acid phosphatase, neutral protease and lactate dehydrogenase. Strong irritants and corrosive substances inhibit release or destroy the enzymes. There are inconsistencies in the identification of moderate irritants, both chemicals and bacterial toxins, in which the rank order of severity of effect does not correspond to other in vitro features or in vivo potency. Detection of cytokines (prostaglandins, leukotrienes and interleukin-1) is possible with appropriate conditions of culture, but has been little applied to toxicology. Studies on two-organ/cell systems are briefly reported in which biological mediator release is detectable without specific identification. Cultured skin preparations were treated in vitro with irritants. The culture media were transferred to cultures of normal skin (examined for stimulus to DNA synthesis), to fibroblasts (examined for DNA and polysaccharide/heparan sulphate synthesis) and to suspensions of neutrophils in Boyden chambers (for chemotaxis). The media were also examined for histamine (none found), acid protease cathepsins, acid phosphatase and lactate dehydrogenase. As before, weak chemical irritants were detectable by stimulation of normal skin and fibroblasts to DNA synthesis, fibroblasts to increased heparan sulphate-like synthesis, and analytically by release of enzymes. Moderate irritants and Clostridium perfringens type C toxins depressed these features. Reference is made to the possibility to identify substances likely to induce progressive pulmonary fibrosis, by transfer to fibrocyte cultures of media from macrophages ingesting the dusts or co-culturing macrophages and fibrocytes, to detect a sustained stimulus to fibrogenesis.
介质的释放是组织刺激或损伤的一种表现，适用于体外毒理学研究。细胞间介质或细胞因子在组织受到刺激时或因轻度损伤而释放。细胞内介质或呼吸与代谢酶从受损细胞中释放出来，参与炎症变化，并且是体外试验中常见的介质决定因素。用于体外介质释放检测的细胞有肥大细胞（适用于过敏性过敏和急性炎症）、巨噬细胞（特别适用于可吸入粉尘的毒性研究）和成纤维细胞/纤维细胞（广泛用作细胞毒性的通用模型）。在器官试验中，可对肺进行灌注以鉴定受试物质释放的细胞因子，这些细胞因子可用于区分过敏效应和化学诱导炎症所产生的效应。人及动物的皮肤，无论是全层皮肤还是表皮角质层切片，都可用于检测刺激性和腐蚀性物质。弱刺激物可通过酶的释放来鉴定，例如酸性磷酸酶、中性蛋白酶和乳酸脱氢酶。强刺激物和腐蚀性物质会抑制酶的释放或破坏这些酶。在鉴定中度刺激物（包括化学物质和细菌毒素）方面存在不一致情况，其中效应的严重程度排序与其他体外特征或体内效力不对应。在适当的培养条件下可以检测细胞因子（前列腺素、白三烯和白细胞介素 -1），但在毒理学中的应用很少。简要报道了对双器官/细胞系统的研究，其中可检测到生物介质的释放但未进行特异性鉴定。用刺激物对培养的皮肤制剂进行体外处理。将培养基转移至正常皮肤培养物（检测对DNA合成的刺激）、成纤维细胞（检测DNA和多糖/硫酸乙酰肝素合成）以及博伊登小室中的中性粒细胞悬液（用于趋化性检测）。还对培养基进行组胺（未检测到）、酸性蛋白酶组织蛋白酶、酸性磷酸酶和乳酸脱氢酶的检测。如前所述，弱化学刺激物可通过刺激正常皮肤和成纤维细胞的DNA合成、刺激成纤维细胞增加硫酸乙酰肝素样合成以及通过酶的释放分析来检测。中度刺激物和C型产气荚膜梭菌毒素会抑制这些特征。文中提到了通过将摄取粉尘的巨噬细胞培养基转移至纤维细胞培养物中或使巨噬细胞与纤维细胞共培养来鉴定可能诱导进行性肺纤维化的物质的可能性，以检测对纤维生成的持续刺激。
