Localization of bullous pemphigoid antigen (BPA) in isolated human keratinocytes.
Localization of bullous pemphigoid antigen (BPA) in isolated human keratinocytes.
复制标题
大疱性类天疱疮抗原 (BPA) 在分离的人角质形成细胞中的定位。
DOI：
10.1111/1523-1747.ep12276656
复制
发表时间：
1985
期刊：
影响因子：
--
通讯作者：
M. Prunieras
中科院分区：
文献类型：
--
作者：
M. Régnier;P. Vaigot;S. Michel;M. Prunieras
在早期的研究中，大疱性类天疱疮抗原（BPA）定位于基底膜区的透明板细胞外。然而，胰蛋白酶解离的基底细胞可以用大疱性类天疱疮血清（BPS）标记。通过免疫荧光，BPA出现在基底细胞（BC）的真皮极。这可能表明，当BC与底层基质分子分离时，BPA的块仍然附着在它们上面。在本研究中，新鲜的原油初始悬浮液（CIS）的表皮细胞制备胰蛋白酶-EDTA解离。将细胞涂片并风干。极性荧光细胞（即，BC）为42% +/-7%。然后使CIS通过荧光激活细胞分选仪（FACS）。在通过FACS选择的荧光阳性级分中，仅存在34% +/-7%的BC。将FACS阴性细胞部分涂抹在载玻片上，风干，并用BPS +异硫氰酸荧光素再染色;这些部分中存在66% +/- 10%的BC。这证明胰蛋白酶分离的BC包括两个亚群：一个具有BPA直接可及性，另一个不具有BPA直接可及性。活力试验和组织培养研究表明，FACS阳性细胞部分没有活力。从CIS、FACS阳性和FACS阴性组分中提取BPA，并针对BPS进行免疫印迹。用肝素酶、亚硝酸、甲醇-氯仿或EDTA处理FACS阴性细胞组分，而不改变反应细胞的数量。当用Triton X-100处理BC或通过连续冷冻和解冻进行透化时，阳性细胞的数量变得与通过空气干燥涂片获得的阳性细胞的数量相当。最后，BPA定位于细胞内部分的半桥粒的BC的免疫电镜。为了观察BPA是否也存在于细胞外，在小型猪中产生抽吸水泡，并将BPS注射到水泡腔中。发现BPA附着在细胞顶部的所有细胞上，但不附着在水泡的真皮基底上。当将在冷胰蛋白酶中过夜的皮肤片与BPS反应时，在裂缝的两侧（表皮和真皮）都发现了BPA。它的结论是，BPA有两个定位：一个细胞外，基本上不稳定的积累在真皮-表皮交界处;其他基本上是稳定的，保持在细胞内的一部分，基底细胞半桥粒，并可以检测后透化的细胞。
In early studies, the bullous pemphigoid antigen (BPA) has been localized extracellularly in the lamina lucida in the basement membrane zone. However, trypsin-dissociated basal cells can be tagged with bullous pemphigoid sera (BPS). By immunofluorescence, BPA appears located at the dermal pole of basal cells (BC). This may indicate that when BC are separated from the underlying matrix molecules, chunks of BPA remain attached to them. In the present study, fresh crude initial suspensions (CIS) of epidermal cells were prepared by trypsin-EDTA dissociation. The cells were smeared and air-dried. Polar fluorescent cells (i.e., BC) amounted to 42% +/- 7%. CIS were then passed through a fluorescence-activated cell sorter (FACS). In the fluorescent-positive fractions selected by FACS, 34% +/- 7% only of the BC were present. FACS-negative cell fractions were smeared on glass slides, air-dried, and restained with BPS + fluorescein isothiocyanate; 66% +/- 10% of BC were present in these fractions. This is evidence that trypsin-isolated BC comprise two subpopulations: one with BPA directly accessible, the other not. Viability tests and tissue culture studies indicated that the FACS-positive cell fractions were not viable. BPA was extracted from CIS, FACS-positive, and FACS-negative fractions and immunoblotted against BPS. Identical blots were found. FACS-negative cell fractions were treated with heparitinase, nitrous acid, methanol-chloroform, or EDTA without modifying the number of reacting cells. When BC were treated with Triton X-100 or permeabilized by successive freezings and thawings, the number of positive cells became comparable to those obtained by air-drying smears. Finally, BPA was localized on the intracellular part of hemidesmosomes of BC by immunoelectron microscopy. To see whether BPA was also present extracellularly, suction blisters were raised in minipigs and BPS injected into the blister cavity. BPA was found attached to all cells of the cellular roof but not to the dermal base of the blisters. When pieces of skin kept overnight in cold trypsin were reacted with BPS, BPA was found on both sides (epidermal and dermal) of the split. It is concluded that BPA has two localizations: one extracellular, essentially labile which accumulates at the dermal-epidermal junction; the other essentially stable which remains on the intracellular part of basal cell hemidesmosomes and which can be detected after permeabilization of the cells.
影响因子：
6.5
作者：
GAMMON, WR;BRIGGAMAN, RA;WHEELER, CE
通讯作者：
WHEELER, CE