Table_1_Mycobacterium leprae and host immune transcriptomic signatures for reactional states in leprosy.xlsx

BackgroundMycobacterium leprae transcriptomic and human host immune gene expression signatures that demonstrate a plausible association with type I (T1R) and type II reactions (T2R) aid in early diagnosis, prevention of nerve damage and consequent demyelinating neuropathy in leprosy. The aim of the study is to identify M. leprae and host-associated gene-expression signatures that are associated with reactional states in leprosy.
MethodsThe differentially expressed genes from the whole transcriptome of M. leprae were determined using genome-wide hybridization arrays with RNA extracted from skin biopsies of 20 T1R, 20 T2R and 20 non reactional controls (NR). Additionally, human immune gene-expressions were profiled using RT2-PCR profiler arrays and real-time qPCRs.
ResultsThe RNA quality was optimal in 16 NR, 18 T1R and 19 T2R samples. Whole transcriptome expression array of these samples revealed significant upregulation of the genes that encode integral and intrinsic membrane proteins, hydrolases and oxidoreductases. In T1R lesional skin biopsy specimens, the top 10 significantly upregulated genes are ML2064, ML1271, ML1960, ML1220, ML2498, ML1996, ML2388, ML0429, ML2030 and ML0224 in comparison to NR. In T2R, genes ML2498, ML1526, ML0394, ML1960, ML2388, ML0429, ML0281, ML1847, ML1618 and ML1271 were significantly upregulated. We noted ML2664 was significantly upregulated in T1R and repressed in T2R. Conversely, we have not noted any genes upregulated in T2R and repressed in T1R. In both T1R and T2R, ML2388 was significantly upregulated. This gene encodes a probable membrane protein and epitope prediction using Bepipred-2.0 revealed a distinct B-cell epitope. Overexpression of ML2388 was noted consistently across the reaction samples. From the host immune gene expression profiles, genes for CXCL9, CXCL10, CXCL2, CD40LG, IL17A and CXCL11 were upregulated in T1R when compared to the NR. In T2R, CXCL10, CXCL11, CXCL9, CXCL2 and CD40LG were upregulated when compared to the NR group.
ConclusionA gene set signature involving bacterial genes ML2388, ML2664, and host immune genes CXCL10 and IL-17A can be transcriptomic markers for reactional states in leprosy.

背景 麻风分枝杆菌（Mycobacterium leprae）转录组及人类宿主免疫基因表达特征若能证实与I型反应（T1R）、II型反应（T2R）存在可靠关联，将有助于麻风病的早期诊断、神经损伤预防及由此引发的脱髓鞘性神经病防控。本研究旨在筛选与麻风病反应状态相关的麻风分枝杆菌及宿主相关基因表达特征。
方法 本研究从20例T1R患者、20例T2R患者及20例非反应性对照（NR）的皮肤活检组织中提取RNA，通过全基因组杂交芯片检测麻风分枝杆菌全转录组的差异表达基因。此外，采用RT2-PCR基因表达谱芯片与实时定量聚合酶链反应（real-time qPCR）对人类免疫基因表达谱进行分析。
结果 16例非反应性对照、18例T1R患者及19例T2R患者的样本RNA质量达标。对上述样本开展全转录组表达芯片检测发现，编码整合膜蛋白、内在膜蛋白、水解酶及氧化还原酶的基因显著上调。与非反应性对照相比，T1R病变皮肤活检标本中前10位显著上调的基因为ML2064、ML1271、ML1960、ML1220、ML2498、ML1996、ML2388、ML0429、ML2030及ML0224。T2R组中，ML2498、ML1526、ML0394、ML1960、ML2388、ML0429、ML0281、ML1847、ML1618及ML1271基因显著上调。研究发现ML2664在T1R组中显著上调，而在T2R组中表达受抑制；反之，未发现存在于T2R组上调但在T1R组受抑制的基因。T1R与T2R组中ML2388均显著上调，该基因编码一种潜在膜蛋白，采用Bepipred-2.0进行表位预测显示其存在明确的B细胞表位，且在所有反应性样本中均持续呈现过表达状态。从宿主免疫基因表达谱来看，与非反应性对照组相比，T1R组中CXCL9、CXCL10、CXCL2、CD40LG、IL17A及CXCL11基因均上调；T2R组中CXCL10、CXCL11、CXCL9、CXCL2及CD40LG基因则显著上调。
结论 包含麻风分枝杆菌基因ML2388、ML2664以及宿主免疫基因CXCL10、IL17A的基因特征集，可作为麻风病反应状态的转录组学标志物。