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. 2022 Jul 1;26(4):313-23.
doi: 10.52547/ibj.3586.

Strain-Specific Behavior of Mycobacterium tuberculosis in Interruption of Autophagy Pathway in Human Alveolar Type II Epithelial A549 Cells

Nasim Ebrahimifard  1   2 , Shima Hadifar  1   2 , Mansour Kargarpour Kamakoli  1   2 , Ava Behrouzi  1   3 , Sharareh Khanipour  1   2 , Abolfazl Fateh  1   2 , Seyed Davar Siadat  1   2 , Farzam Vaziri  1   2
Affiliations

Strain-Specific Behavior of Mycobacterium tuberculosis in Interruption of Autophagy Pathway in Human Alveolar Type II Epithelial A549 Cells

Nasim Ebrahimifard et al. Iran Biomed J. .

Abstract

Background: Autophagy induction has been shown to differ in magnitude depending on the mycobacterial species. However, few studies have investigated the specific autophagic capacity of different Mycobacterium tuberculosis (Mtb) strains in alveolar epithelial cells (ATs). This study aimed to elucidate the host autophagic response to different Mtb strains in ATs responsible for TB in the capital of Iran, Tehran.

Methods: A549 cells were infected with three different Mtb clinical isolates (Beijing, NEW1, and CAS1/Delhi) and the reference strain H37Rv. Following RNA extraction, the expression of eight ATG genes, four mycobacterial genes, and three miRNAs was evaluated using quantitative RT-PCR.

Results: The results revealed that all four strains influenced the autophagy pathway in various ways at different magnitudes. The Beijing and H37Rv strains could inhibit autophagosome formation, whereas the CAS and NEW1 strains induced autophagosome formation. The expression of genes involved in the fusion of autophagosomes to lysosomes (LAMP1) indicated that all the studied strains impaired the autophagolysosomal fusion; this result is not unexpected as Mtb can block the autophagolysomal fusion. In addition, the Beijing and H37RV strains prevented the formation of autophagic vacuoles, besides mycobacterial targeting of lysosomes and protease activity.

Conclusion: This preliminary study improved our understanding of how Mtb manages to overcome the host immune system, such as autophagy, and evaluated the genes used by specific strains during this process. Further studies with a large number of Mtb strains, encompassing the other main Mtb lineages, are inevitable.

Keywords: A549 cells; Autophagy; MicroRNA; Mycobacterium tuberculosis.

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Conflict of interest statement

The authors declare that they have no conflicting interests.

Figures

Fig. 1
Fig. 1
Intracellular growth of the studied Mtb genotypes in the A549 cell line at 72 h post infection. (***p < 0.001).
Fig. 2
Fig. 2
Effects of different Mtb genotypes on the mRNA expression of genes involved in autophagy in A549 cell line. Data are normalized using GAPDH as a control gene. ∗p < 0.05, ∗∗p < 0.01, and ***p < 0.001 are considered statistically significant compared to the mock cell.
Fig. 3
Fig. 3
Expression of eis, pknG, esat6, and zmp1 genes in the studied Mtb genotypes. Data are normalized using secA as the control gene. ∗p < 0.05, ∗∗p < 0.01, and ***p < 0.001 are considered statistically significant compared to the mock cell.
Fig. 4
Fig. 4
Expression of miR-155, miR-142-3p, and miR-34a in the infected A549 cells by the studied Mtb genotypes; Data are normalized using U6 gene as a control gene (∗p < 0.05, ∗∗p < 0.01, and ***p < 0.001 are considered statistically significant compared to the mock cell, respectively).
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