The University of Texas MD Anderson Cancer Center, Department of Immunology, Houston, TX, United States of America.
The University of Texas MD Anderson Cancer Center, Department of Veterinary Sciences, Bastrop, TX, United States of America.
PLoS One. 2017 Dec 8;12(12):e0188807. doi: 10.1371/journal.pone.0188807. eCollection 2017.
Studies in nonhuman primates (NHP) for prospective immune cell monitoring subsequent to infection and/or vaccination usually rely on periodic sampling of the blood samples with only occasional collections of biopsies from mucosal tissues because of safety concerns and practical constraints. Here we present evidence in support of cytobrush sampling of oral, rectal, and genital mucosal tissues as a minimally invasive approach for the phenotypic analyses of different T cells subsets de novo as well as prospectively after intranasal immunization in rhesus macaques. Significant percentages of viable lymphocytes were obtained consistently from both naïve and chronically SIV-infected rhesus macaques. The percentages of CD3+ T cells in the blood were significantly higher compared to those in the mucosal tissues analyzed in the naïve animals, while in the SIV+ animals the CD3+ T cells were significantly elevated in the rectal tissues, relative to all other sites analyzed. In the naïve, but not SIV+ macaques, the rectal and vaginal mucosal tissues, compared to oral mucosa and blood, showed higher diversity and percentages of CD4+ T cells expressing the HIV entry co-receptor CCR5 and mucosal specific adhesion (CD103) as well as activation (HLA-DR) and proliferation (Ki67) markers. Sequential daily cytobrush sampling from the oral, rectal, and genital mucosal tissues was performed in SIV+ animals from an ongoing study where they were administered intranasal immunization with adenoviral vectored vaccines incorporating the green fluorescent protein (GFP) reporter gene. We detected a transient increase in GFP+ CD4 T cells in only oral mucosa suggesting limited mucosal trafficking. In general, CD4+ and CD8+ T cells expressing Ki67 transiently increased in all mucosal tissues, but those expressing the CCR5, HLA-DR, and CD103 markers exhibited minor changes. We propose the minimally invasive cytobrush sampling as a practical approach for effective and prospective immune monitoring of the oral-genital mucosal tissues in NHP.
在非人灵长类动物（NHP）中进行前瞻性免疫细胞监测，以研究感染和/或接种疫苗后的情况，通常依赖于定期采集血液样本，仅偶尔从黏膜组织采集活检样本，这主要是出于安全性和实际限制的考虑。在这里，我们提供了证据支持使用细胞刷对口腔、直肠和生殖器黏膜组织进行采样，作为一种微创方法，可用于对不同 T 细胞亚群进行表型分析，无论是在恒河猴经鼻腔免疫接种前还是接种后进行前瞻性分析。从恒河猴的未感染和慢性 SIV 感染个体中，都能稳定地获得大量有活力的淋巴细胞。与分析未感染动物的黏膜组织相比，血液中 CD3+ T 细胞的比例明显更高，而在 SIV+动物中，直肠组织中的 CD3+ T 细胞比例明显高于分析的其他所有部位。在未感染 SIV 的动物中，与口腔黏膜和血液相比，直肠和阴道黏膜组织的 CD4+ T 细胞具有更高的多样性和表达 HIV 进入共受体 CCR5 和黏膜特异性粘附（CD103）以及激活（HLA-DR）和增殖（Ki67）标志物的比例。在正在进行的研究中，对 SIV+ 动物进行了每日顺序细胞刷采样，这些动物接受了鼻腔免疫接种，使用了包含绿色荧光蛋白（GFP）报告基因的腺病毒载体疫苗。我们仅在口腔黏膜中检测到 GFP+ CD4 T 细胞的短暂增加，这表明黏膜的转运有限。一般来说，所有黏膜组织中表达 Ki67 的 CD4+和 CD8+ T 细胞都会短暂增加，但表达 CCR5、HLA-DR 和 CD103 标志物的细胞变化较小。我们提出这种微创细胞刷采样方法是对 NHP 口腔-生殖器黏膜组织进行有效和前瞻性免疫监测的实用方法。
