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- Material and methods
Background: In nonimmediate cutaneous reactions to drugs, the skin is the organ most frequently involved, and T cells may play a relevant role. T cells related to skin immune responses express the cutaneous lymphocyte-associated antigen (CLA), the skin-homing receptor.
Methods: We studied the expression of the CLA in peripheral blood T cells from nine subjects with exanthematous reactions induced by β-lactams ( 4), phenytoin ( 2), propyphenazone ( 1), spiramycin plus metronidazol ( 1), and captopril plus tiazide ( 1). The cutaneous symptoms appeared at least 6 h after drug intake. CLA expression was evaluated by flow cytometry at the time of the reaction (T1) and 1 month later (T2). HLA-DR activation marker expression was also evaluated at T1. In four patients, it was necessary to readminister the culprit drug to establish a causal relationship, and sequential estimation of the markers was performed. Two control groups were included: healthy controls and subjects exposed to the culprit drugs with good tolerance. Values were compared by nonparametric statistics.
Results: The expression of circulating CLA+ T cells at T1 was increased compared to healthy controls (median=20.4 vs 9.4) (P<0.001), and the patients also expressed increased levels of HLA-DR (median=3.8) (P<0.005). Comparison between T1 and T2 (median=11.2) also showed differences in levels of CLA+ T cells (P<0.01). The patients re-exposed to the culprit drug showed an increase followed by a decrease of circulating CLA+ T cells (P<0.05) and CLA+ HLA-DR+(P<0.05) paralleling the symptoms.
Conclusions: These data support the immunologic nature of delayed skin reactions to drugs, and suggest that these CLA+ T cells parallel the disease evolution and may participate in the pathophysiologic mechanisms.
One of the most frequent causes of adverse reactions to drugs is an allergic response in which different immunologic mechanisms may be involved. Among the clinical manifestations reported are anaphylaxis, urticaria, exanthema, desquamative lesions, and Steven-Johnson syndrome ( 1). The appearance of symptoms may vary from minutes to days or even weeks after drug intake, depending on the mechanisms involved. In general, the anaphylactic reactions occurring immediately after drug intake are due to activation of mast cells and basophils by specific IgE antibodies that recognize the drug as a hapten ( 2). However, the great majority of drug reactions with skin involvement occur hours or days after drug intake, and IgE mechanisms do not seem to take part ( 3). Although the mechanisms that induce delayed cutaneous reactions to drugs are complex, the participation of activated T cells has been suggested ( 4, 5). In some drug-allergic patients, peripheral lymphocytes proliferate in the presence of the specific drug ( 6), and activated T cells are the primary component of the skin inflammatory infiltrate in these reactions ( 7).
T cells related to the cutaneous immune response can be identified because most skin-infiltrating T cells express the cutaneous lymphocyte-associated antigen (CLA) ( 8). Moreover, 10–15% of circulating CD3+ cells are CLA+, and CLA+ T cells bind and migrate through endothelial cells by adhesive interaction with E-selectin ( 9, 10). Leukocyte extravasation into tissue is a multistep process ( 11), and the molecules involved in the migration of CLA+ T cells include VCAM-1, ICAM-1, and the interleukin-8 receptor B ( 12, 13). However, the exact extravasation mechanism has not been completely characterized ( 14, 15). A recent study using a hu-SCID mouse grafted with human skin and human T cells provided direct evidence of the role of the CLA receptor in directing lymphocyte traffic into the skin ( 16). Moreover, the response to skin-associated antigens has been shown to be restricted to the circulating CLA+ T-cell population. In atopic dermatitis, CLA+ T cells preferentially respond to extract of house-dust mite; in contact dermatitis, the skin-homing population responds to nickel ( 17). Since these and other reports have underlined the importance of the CLA antigen as the human skin-homing receptor, we studied the CLA+ T-cell population in patients affected by delayed cutaneous allergic reactions to drugs. Our results indicate the involvement of CLA+ T cells in this type of reaction.
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- Material and methods
T cells have been shown to be involved in delayed-type skin reactions induced by drugs ( 7, 23), and, although their role in the pathophysiologic mechanism is relevant, the precise process by which they participate needs to be clarified. In recent years, there has been an increased interest in the evaluation of immunologic reactions to drugs, including allergy, but most of the data come from in vitro studies, and extrapolation to in vivo models still needs to be confirmed. Generation of T-cell lines and clones in vitro from skin-infiltrating lymphocytes has enabled the study of their antigen specificity, cytokine production, and antigen-presentation requirements with different drugs ( 6, 24). The in vivo monitoring of the acute response in peripheral blood from allergic patients has been carried out by several groups ( 25–27), but few data have been provided in studies of the allergic response to drugs ( 6). Therefore, for a better understanding of the immunologic mechanisms implicated in delayed-type, drug-induced cutaneous reactions, we studied in vivo the population of circulating skin-homing T cells, its activation state, and its association with the evolution of the clinical manifestations of this type of reaction.
Preliminary data indicated that subjects with allergic drug reactions expressed increased levels of CD3+ CLA+ lymphocytes, and in some their activated state paralleled the clinical symptoms ( 28–30). We have now compared the percentage of circulating CLA+ T cells in patients with delayed-type reactions induced by drugs, and our results indicate a clear increase in the percentage of circulating CLA+ T cells compared with healthy controls. In addition, the presence of the activation marker HLA-DR in the CLA+ population was also increased. Circulating CLA+ T cells decreased in parallel with the improvement of the clinical symptoms. These data indicate a close relationship between skin symptoms and the expansion and activation status of circulating CLA+ T cells.
In milk-induced eczema, it has been suggested that oral administration of antigen may lead to its deposition in the skin, where it can induce a T-cell-mediated immune response ( 31). It has been shown that casein can induce in vitro the expansion of specific CLA+ T cells from patients with milk-induced eczema ( 31). In our study, there was also an oral administration of antigen inducing cutaneous symptoms and CLA+ T-cell expansion in vivo. We think that these cells play an important role not only in those allergic reactions in which the antigen penetrates the skin ( 17) but also in those induced by the oral route.
To clarify the relationship between circulating CLA+ T cells and drug-induced cutaneous symptoms, we studied four patients who developed the same reaction after a controlled re-exposure to the original drug involved. The symptoms appeared within an interval of 12–30 h after drug intake, and there was a significant increase in the percentage of circulating CLA+ T cells with an increase in DR expression also, as had occurred in the initial reaction reported in each patient. These data support the hypothesis of the immunologic nature of the adverse drug reaction, since an increase of activated CLA+ lymphocytes followed the administration of the drug.
It has been previously shown that in antigen-induced, delayed-type hypersensitivity reactions CLA+ T cells are localized in the skin blisters ( 19, 32), and our results concerning the CLA+ T-cell population may help to clarify the mechanism involved in this process. We hypothesize that an immunologic stimulus originating in the skin induces and amplifies an immunologic response with the increase in peripheral blood of lymphocytes expressing the CLA and homing to the skin, contributing in this way to the inflammatory response. Since the circulating CLA+ T-cell subset is easily accessible and can be recognized by a specific marker, this will enable further information to be obtained about antigen specificity and cytokine production in drug-allergic reactions.