Study on the mechanism of EBV reactivation and NK cell oncogenesis in hypersensitivity to mosquito bites

蚊虫叮咬过敏(HMB)的特征是局部皮肤症状强烈，不仅包括红斑或大疱性红斑，还包括溃疡或疤痕，以及全身症状，如高烧、淋巴结病和肝脾肿大。最近的研究表明，在HMB患者中，EB病毒(EBV)DNA阳性的自然杀伤(NK)细胞，包括NK细胞白血病/淋巴瘤，具有淋巴增殖作用。我们先前证实，HMB患者的CD4^+T细胞在蚊子唾液腺提取物(SGE)的刺激下增殖良好，并在EB病毒在NK细胞中的潜伏状态重新激活过程中发挥关键作用。蚊虫叮咬可通过蚊虫抗原特异性的CD4^+T细胞在NK细胞中诱导病毒癌基因LMP1的表达，参与体内NK细胞的致癌过程。在这里，我们建立了来自HMB患者的NK细胞的长期培养。培养的NK细胞与蚊虫SGE刺激的自体CD4^+T细胞共培养后，LMP1表达增强，NK细胞生长增强。经蚊虫刺激后，CD4+T细胞中IL-4、IL-10和转化生长因子-β的基因和蛋白表达上调。加入抗IL-2中和抗体的NK和蚊虫SGE刺激的CD_4~++T细胞可抑制LMP1mRNA的表达，而加入抗转化生长因子-β的中和抗体则可增强LMP1mRNA的表达并抑制EB病毒的再激活。IL-4和IL-10中和抗体对LMP1表达和NK细胞生长无影响。此外，我们调查了一例氨苄西林引起的皮疹患者，发现氨苄西林可能在EBV感染的重新激活中起关键作用，这意味着EBV的重新激活在该患者皮疹的发病机制中起着重要作用。提示EBV的重新激活不仅与HMB有关，还与其他过敏性疾病有关。

Hypersensitivity to mosquito bites (HMB) is characterized by intense local skin symptoms, which consist of not only erythema or bulla but also ulceration or scarring, and systemic symptoms such as high fever, lymphadenopathy, and hepatosplenomegaly. Recent studies demonstrated the lymphoproliferation of EB virus (EBV) DNA-positive natural killer (NK) cells, including NK cell leukemia/lymphoma, in HMB patients. We previously demonstrated that CD4^+T cells of a patient with HMB proliferated well in response to mosquito salivary gland extracts (SGE) and played a key role in the reactivation of EBV from a latent state in NK cells. We also showed that mosquito bites can induce expression of the viral oncogene LMP1 in NK cells via mosquito antigen-specific CD4^+T cells, which is involved in the oncogenesis of NK cells in vivo. Here, we established long-term cultures of NK cells from the patients with HMB. When these cultured NK cells were cocultured with autologous CD4^+T cells stimulated by mosquito SGE, the expression of LMP1 and NK cell growth were enhanced. It was proved that mRNA and protein expressions of IL-4, IL-10, and TGF-β were upregulated in the CD4^+T cells by stimulation with the mosquito SGE. Adding a neutralizing antibody to IL-2 into the coculture of NK and mosquito SGE-stimulated CD4^+T inhibited the LMP1 mRNA expression, while adding a neutralizing antibody to TGF-β enhanced the LMP1 mRNA expression and suppressed the reactivation of EBV. On the other hand, neutralizing antibodies to IL-4 and IL-10 had no effects on LMP1 expression and NK cell growth. Furthermore, we investigated a patient with skin rash induced by ampicillin, and found that ampicillin might have played a key role in reactivation of EBV infection, implying that EBV reactivation contributed to the pathogenesis of skin rash in this patient. It suggests that the reactivation of EBV is related not only to HMB but also to other allergic diseases.