Fig 2: Schematic figure of the current study. Increased TUFM expression in hypoxia-stimulated pulmonary arterial hypertension cells causes an increasing mtDNA translation, leading to dysfunction of the mitochondrial respiratory chain. Mitochondrial dysfunction induces cellular stress and then activates AMPK. On the one hand, activated AMPK decreases the phosphorylation level of mTOR, inhibits the activity of mTOR, and then disassociates from ULK1. Thus, phosphorylation of specific sites of ULK1 and Atg13 is released. Meanwhile, the ULK1 complex is activated through autophosphorylation at thr180 and phosphorylates Atg13, FIP200, atg101, and other Atg proteins. The activated ULK1 complex then translocates to the isolation membrane of the endoplasmic reticulum, where autophagy is initiated. On the other hand, activated AMPK will directly stimulate ULK1 and BECN1, initiating autophagy.

Fig 3: Expressions of TUFM and the autophagy indicators in different treated groups. (a) Western blot of TUFM, LC3II/I, Bax, Bcl2, BECN1, P62, and Apaf expression in each group as indicated. (b–h) Densitometry data represent the intensity of each group. Ctrl: normal control; PAH: pulmonary arterial hypertension; OE-NC: overexpression negative control; OE: overexpression of TUFM; Sh-NC: Sh-TUFM negative control; Sh: Sh-TUFM group. Data is presented as mean ± SD. n = 6 each group. *P < 0.05 vs. Ctrl group, **P < 0.01, ***P < 0.001, and ****P < 0.0001; #P < 0.05 vs. PAH group, ##P < 0.01, ###P < 0.001, and ####P < 0.0001.

Fig 4: Absence of TUFM improved the apoptosis and reduced the proliferation of PASMCs. (a) TUFM, Bax, and Bcl2 expression under hypoxia condition. (b–d) Densitometry data represent the intensity of each group. (e) Cell viability tested by CCK-8 under hypoxia condition. (f) Quantification of proliferation level in smooth muscle cell by EdU assay kit. (g) Cell proliferation activity of each group under hypoxia. Scale bar: 50 µm. Data is presented as mean ± SD. *P < 0.05, **P < 0.01, and ***P < 0.001.

Fig 5: Monocrotaline-induced pulmonary hypertension and the expression of TUFM in the pulmonary hypertension and normal control rats. (a) 28 days after monocrotaline induction of the model, the pulmonary arterial systolic pressure measured using the system. (b) Hematoxylin and eosin (HE) staining for the pulmonary arterioles of normal control and monocrotaline-induced PAH model. Scale bar: 50 µm. (c) Differential expression of TUFM from GSE15197 database analysis. (d) The initial test for TUFM, mitophagy marker P62, and apoptosis indicator Bax and Bcl2 expression in the normal control and monocrotaline-induced PAH rat. (e–h) Densitometry data represent the intensity of the quantitative expression of P62, TUFM, Bcl2, and Bax proteins. (i) Immunofluorescence chemistry examination indicated the location profile of TUFM in pulmonary arteries. Scale bar: 50 µm. Ctrl: normal control; PAH: pulmonary arterial hypertension. Data is presented as mean ± SD. n = 6 each group. *P < 0.05 vs. Ctrl group, **P < 0.01, and ***P < 0.001.