Lactic acid bacteria have been studied as a vehicle for the delivery of plasmid DNA to the gastrointestinal tract. However, low levels of gene expression in vivo limit their practical use. Furthermore, it is still unclear how the orally administrated bacteria transfer their harbored plasmid DNA to host intestinal cells. To more easily track the delivery of plasmid DNA for eukaryotic expression in the intestine, we constructed an L. lactis-E. coli shuttle plasmid (pLEC) that allowed significantly elevated expression of the target protein of interest in eukaryotic cells. We first demonstrated its usefulness for delivery from L. lactis to Caco-2 cells in vitro. We then investigated the cellular target for the L. lactis DNA delivery system in vivo. Mice were orally administrated with LL/pLEC:EGFP, an L. lactis strain carrying pLEC for EGFP expression, and immunofluorescent analyses of frozen sections prepared from their small intestines identified a number of EGFP-expressing cells in the lamina propria and some in the sub-epithelial dome of the Peyer’s patches. Flow cytometric analysis revealed that these EGFP-expressing cells were both CD11c- and F4/80-positive but CX₃CR1-negative, suggesting that they are eosinophils. Immunostaining of the sections with an antibody against Siglec-F, a marker protein of eosinophils, confirmed the flow cytometric findings. Thus, the target cells of DNA delivery from L. lactis in the intestines are mainly eosinophils in the lamina propria and Peyer’s patches. This finding may open a new approach to the development of DNA vaccines for oral administration.

已经研究了乳酸细菌作为将质粒DNA递送至胃肠道的载体。然而，体内低水平的基因表达限制了它们的实际应用。此外，尚不清楚口服细菌如何将其携带的质粒DNA转移至宿主肠道细胞。为了更轻松地跟踪质粒DNA在肠中用于真核表达的递送，我们构建了乳酸乳球菌-E。大肠杆菌穿梭质粒（pLEC），可在真核细胞中显着提高目标靶蛋白的表达。我们首先证明了其在体外从乳酸乳球菌向Caco-2细胞传递的有用性。然后，我们研究了乳酸乳球菌的细胞靶标体内DNA传递系统。给小鼠口服LL / pLEC：EGFP，这是一种带有pLEC的乳酸乳球菌菌株，用于表达EGFP，对从小肠制备的冰冻切片进行免疫荧光分析，发现在固有层中有许多表达EGFP的细胞，在亚层中也有一些-Peyer斑块的上皮圆顶。流式细胞仪分析表明，这些表达EGFP的细胞均为CD11c和F4 / 80阳性，而CX ₃ CR1为阴性，表明它们是嗜酸性粒细胞。用针对嗜酸性粒细胞标记蛋白Siglec-F的抗体对切片进行免疫染色，证实了流式细胞仪的发现。因此，从乳酸乳球菌递送DNA的靶细胞肠道中的主要是固有层和Peyer斑中的嗜酸性粒细胞。这一发现可能为口服DNA疫苗的开发开辟新途径。