RT-qPCR and GEO validation
The transcript expression levels of three genes, Retn, Sftpd, and Amy2a,
in 10 TB patients and 15 healthy volunteers were analyzed by RT-qPCR
assay (Figure 5). Compared with the healthy control group, the relative
expression of Retn gene in TB patients showed a downward trend, and
Amy2a gene in TB patients showed an upward trend, which is consistent
with the results of the gene expression profile of the mouse TB model.
However, the relative expression of Sftpd gene in initial treatment TB
patients exhibited an upward trend, which is inconsistent with the
results of the gene expression profile of the mouse TB model.
Through GEO2R, we verified four GSE datasets. The results were shown as
follows: (1) Of the top 20 DE genes in the TB model vs. normal group, 6
DE genes also showed differential expression in GSE89389 and GSE140943
datasets (Table 6), in which the differential expression trend of Sftpd,
Mgp, and Retnla was consistent with our results. (2) The expression
level of DE gene Ccdc65 decreased significantly after the immunotherapy
with ag85a/b DNA IM or EP and M72/AS01 (1.535 down-regulated in
GSE102459 datasets). (3) The differential expression trend of six DE
genes in GSE89403 after 4 and 24 weeks of anti-TB treatment is
consistent with that after effective treatment with ag85a/b DNA
IM or EP (Table 7). The higher expression levels of Cldn5 and Sftpd have
positive correlations respectively with 4 weeks or 24 weeks of treatment
response. In contrast, the expression level of Mgp was negatively
correlated with 4 weeks of treatment response.