Paoletti F, Mocali A
Istituto di Patologia Generale, Università di Firenze, Italy.
J Neurol Sci. 1991 Oct;105(2):211-6. doi: 10.1016/0022-510x(91)90147-y.
Characteristic alterations of transketolase (TK) in extracts from cultured Alzheimer fibroblasts have previously been reported (Paoletti et al. (1990) Biochem. Biophys. Res. Commun., 172: 396-401). These abnormalities, encountered in 9 out of 13 Alzheimer patients, were revealed following isoelectric focusing and consisted of enzyme forms having unusually high alkaline pI values (alkaline bands). The present work has shown that immunologically detected alkaline bands were progressively expressed when Alzheimer fibroblasts were incubated for three weeks without medium changes. Full expression of the altered enzyme pattern was not linked to relative cell density in the petri dish; rather, it appeared to be dependent directly on the time elapsed since cell confluence was reached. Alkaline bands could artificially be induced also in both crude and pure TK preparations from normal cells by a treatment with commercial proteases, particularly chymotrypsin. Moreover, specific inhibitors of endogenous cysteine-proteases were capable of abolishing TK alkaline bands in Alzheimer fibroblasts thus turning a pathological into a normal enzyme pattern. Results obtained suggest that Alzheimer fibroblasts contain enhanced Ca(2+)-independent cysteine-proteolytic activities as compared to normal and other pathological cells. These enzymes, exhibiting chymotrypsin-like activity, might exert their degradative effects at the time of cell extraction using TK and probably other cell components as potential substrates. However, peculiar TK abnormalities represent so far an useful biochemical marker detectable in fibroblasts of living Alzheimer patients and closely associated to this neurological disorder.
先前已有报道（Paoletti等人，（1990年）《生物化学与生物物理研究通讯》，172: 396 - 401），培养的阿尔茨海默病成纤维细胞提取物中转酮醇酶（TK）有特征性改变。在13例阿尔茨海默病患者中的9例中发现了这些异常，这些异常在等电聚焦后显现出来，由具有异常高碱性pI值的酶形式（碱性条带）组成。目前的研究表明，当阿尔茨海默病成纤维细胞在不更换培养基的情况下培养三周时，免疫检测到的碱性条带会逐渐表达。改变后的酶模式的完全表达与培养皿中的相对细胞密度无关；相反，它似乎直接取决于细胞达到汇合后所经过的时间。通过用商业蛋白酶，特别是胰凝乳蛋白酶处理，也可以在来自正常细胞的粗制和纯制TK制剂中人工诱导出碱性条带。此外，内源性半胱氨酸蛋白酶的特异性抑制剂能够消除阿尔茨海默病成纤维细胞中的TK碱性条带，从而将病理酶模式转变为正常酶模式。所获得的结果表明，与正常细胞和其他病理细胞相比，阿尔茨海默病成纤维细胞含有增强的不依赖Ca(2+)的半胱氨酸蛋白水解活性。这些具有胰凝乳蛋白酶样活性的酶，可能在使用TK和可能其他细胞成分作为潜在底物进行细胞提取时发挥其降解作用。然而，到目前为止，独特的TK异常是一种有用的生化标志物，可在活体阿尔茨海默病患者的成纤维细胞中检测到，并且与这种神经疾病密切相关。
