Graduate Program in Basic and Applied Immunology, Ribeirão Preto Medical School, University of São Paulo, Ribeirão Preto, São Paulo, Brazil.
Federal Institute of Education, Science and Technology of Mato Grosso, Campus Sorriso, Sorriso, State of Mato Grosso, Brazil.
PLoS One. 2024 Mar 21;19(3):e0300364. doi: 10.1371/journal.pone.0300364. eCollection 2024.
Paracoccidioides fungi are thermodimorphic microorganisms that cause paracoccidioidomycosis (PCM), an autochthonous disease from Latin America, with most cases in Brazil. Humans become infected by inhaling conidia or mycelial fragments that transform into yeast at body temperature. These fungi cause chronic-granulomatous inflammation, which may promote fibrosis and parenchyma destruction in the lungs. In response to stress imposed by the host, fungi Paracoccidioides spp. increase the expression of heat shock proteins (HSP), which protect them by sustaining cellular proteostasis. Our group has studied the role of HSP60 in PCM, and previous data show that the recombinant HSP60 (rHSP60) has a deleterious effect when used in a single dose as therapy for experimental PCM. Here, we investigated the mechanism by which rHSP60 could worsen the disease. We found that rHSP60 caused the viability loss of splenic or lymph node cells from both immunized and non-immunized mice, including in splenic T lymphocytes under polyclonal stimulation with concanavalin A, probably by undergoing apoptosis. Among analyzed splenic cells, lymphocytes were indeed the main cells to die. When we investigated the death mechanisms, remarkably, we found that there was no viability loss in rHSP60-stimulated splenic cells from mice deficient in Toll-like receptor 4, TRIF adapter protein, and TNF receptor 1(TNFR1), as well as rHSP60-stimulated WT cells incubated with anti-TNF antibody. Besides, caspase-8 inhibitor IETD-CHO blocked the rHSP60 effect on splenic cells, suggesting that rHSP60 induces the extrinsic apoptosis pathway dependent on signaling via TLR4/TRIF and TNFR1.
泊拉氏球腔菌是一种温度双相微生物，可引起泊拉氏球腔菌病（PCM），这是一种源自拉丁美洲的地方性疾病，在巴西最为常见。人类通过吸入分生孢子或菌丝片段感染，这些孢子或片段在体温下转化为酵母。这些真菌引起慢性肉芽肿性炎症，可能导致肺部纤维化和实质破坏。为了应对宿主施加的压力，泊拉氏球腔菌等真菌会增加热休克蛋白（HSP）的表达，这些 HSP 通过维持细胞蛋白平衡来保护它们。我们的研究小组研究了 HSP60 在 PCM 中的作用，先前的数据表明，重组 HSP60（rHSP60）在单次剂量作为实验性 PCM 治疗时具有有害作用。在这里，我们研究了 rHSP60 加重疾病的机制。我们发现，rHSP60 导致免疫和非免疫小鼠的脾或淋巴结细胞，包括在用刀豆蛋白 A 进行多克隆刺激下的脾 T 淋巴细胞的活力丧失，可能通过凋亡发生。在分析的脾细胞中，淋巴细胞确实是主要死亡的细胞。当我们研究死亡机制时，令人惊讶的是，我们发现 TLR4、TRIF 接头蛋白和 TNF 受体 1（TNFR1）缺陷型小鼠的 rHSP60 刺激脾细胞以及与抗 TNF 抗体孵育的 WT 细胞中 rHSP60 刺激均未导致活力丧失。此外，半胱氨酸天冬氨酸蛋白酶-8 抑制剂 IETD-CHO 阻断了 rHSP60 对脾细胞的作用，表明 rHSP60 诱导依赖于 TLR4/TRIF 和 TNFR1 信号的外在凋亡途径。
