Intervention of hydrogen sulfide on FDX1-cuproptosis pathway in rats with hypoxemic pulmonary hypertension

doi:10.12092/j.issn.1009-2501.2026.07.002

Abstract

Abstract:

AIM: To investigate whether hydrogen sulfide (H₂S) improves hypoxic pulmonary hypertension (HPH) in rats through the FDX1-cuproptosis pathway. METHODS: Male Sprague-Dawley (SD) rats were randomly assigned to the Control group; Hypoxia group and Hypoxia+NaHS group, with 10 rats in each group. Rats in the Control group were housed in a normoxic environment (21% O₂) and received a daily intraperitoneal injection of an equal volume of normal saline; rats in the Hypoxia group and Hypoxia+NaHS group were placed in hypoxic chambers and received a daily intraperitoneal injection of 14 μmol/kg NaHS. After 21 days of model establishment, the mean pulmonary arterial pressure (mPAP) of each group was measured by right heart catheterization and the right ventricular hypertrophy index (RVHI) was calculated after weighing. The levels of endothelin-1 (ET-1), nitric oxide (NO) and prostacyclin (PGI₂) were measured by ELISA. Copper ion concentration was detected by inductively coupled plasma mass spectrometry (ICP-MS). The protein levels of copper transporter 1 (CTR1), copper-transporting ATPase α (ATP7A), copper-transporting ATPase β (ATP7B), ferredoxin 1 (FDX1), lipoic acid synthase (LIAS), dihydrolipoamide S-acetyltransferase (DLAT), dihydrolipoamide S-acetyltransferase dimer (DLAT Dimer) and dihydrolipoamide S-succinyltransferase (DLST) were detected by Western blot. The mRNA expression levels of FDX1, LIAS, DLAT and dihydrolipoamide dehydrogenase (DLD) were detected by RT-qPCR. Mitochondrial membrane potential was assessed using the JC-1 kit. ATP content was measured by chemiluminescence. The level of pyruvate dehydrogenase E1 subunit alpha 1 (PDHA1) was measured by ELISA. Citric acid (CA) content was measured by the sulfosalicylic acid method. RESULTS: Compared with the Control group, the Hypoxia group showed significantly increased mPAP and RVHI; significantly elevated ET-1 concentration while NO and PGI₂ concentrations were significantly decreased; significantly increased copper ion concentration; significantly elevated expression of copper transporter 1 while the expressions of copper efflux proteins ATP7A and ATP7B were significantly decreased; the expressions of cuproptosis-related proteins FDX1, LIAS, DLAT and DLST were all significantly decreased, whereas the expression of DLAT dimer is significantly increased; the mRNA expression levels of FDX1, LIAS, DLAT and DLD were significantly decreased; the mitochondrial function indicators JC-1 and ATP levels were significantly decreased and the tricarboxylic acid cycle indicators PDHA1 and CA levels were significantly decreased (all P<0.05). Compared with the Hypoxia group, all the above changes in the Hypoxia+NaHS group were significantly reversed by the therapeutic agent NaHS. CONCLUSION: H₂S may regulate the FDX1-cuproptosis pathway, thereby affecting mitochondrial function and metabolism in lung tissue cells, alleviating pulmonary vasoconstriction and ameliorating pulmonary hypertension.

Key words: copper ion, pulmonary arterial hypertension, cuproptosis, hydrogen sulfide

CLC Number:

Shuyuan WANG, Man HUANG, Sian CHEN, Congmei HAN, Yuan CHENG, Linbo YUAN, Huan YE, Wantie WANG. Intervention of hydrogen sulfide on FDX1-cuproptosis pathway in rats with hypoxemic pulmonary hypertension[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2026, 31(7): 875-884.

Figures/Tables 8

Table 1 Sequences of the primers for RT-qPCR

Name	Primer sequence (5'-3')
β-actin	Forward: GAGAGGGAAATCGTGCGTA
	Reverse: GGAGGAAGAGGATGCGG
FDX1	Forward: CAGTCCACTTCAAGAACCGAGATG
	Reverse: GATGACAGGTAGAGCAAGCCAAAG
LIAS	Forward: ACAGTGTCATGCTTGAAGGAAAGG
	Reverse: ATTCTGGAACCGTCTCTACATTGTG
DLAT	Forward: GGTGTTCGTTAGCCCTCTTGC
	Reverse: TGTCCTTCTTGATGATTCTGCCTTC
DLD	Forward: CCAGGTGCTGGAGAAATGGT
	Reverse: TGGTTTGCCAAAAGACGCAG

Fig.1 Effect of H2S on hemodynamic indexes in hypoxia pulmonary hypertension (HPH) rats ($ \overline{x} $±s, n=6) A: mean pulmonary artery pressure; B: mean pulmonary artery pressure waveform diagram; C: right ventricular hypertrophy index. cP<0.01, compared with control group; fP<0.01, compared with Hypoxia group.

Fig.2 Effect of H2S on ET-1, NO and PGI2 levels in the lung tissues of rats in each group ($ \overline{x} $±s, n=5) A: statistical chart of ET-1 concentration; B: statistical chart of NO concentration; C: statistical chart of PGI2 concentration. cP<0.01, compared with control group; eP<0.05, fP<0.01, compared with Hypoxia group.

Fig.3 Effect of H2S on copper ions in lung tissues of rats in different groups and on the expression of copper transport proteins ($ \overline{x} $±s, n=6) A: copper ion content in lung tissue of rats; B: protein expression levels of copper transporters proteins in lung tissue of rats; C: copper transporters proteins Western blot statistical chart. bP<0.05, cP<0.01, compared with control group; eP<0.05, fP<0.01, compared with Hypoxia group.

Fig.4 Effect of H2S on the expression of cuproptosis related proteins in the lung tissues of rats in each group（$ \overline{x} $±s，n=6) A: protein expression levels of cuproptosis related proteins in lung tissue of rats; B: cuproptosis related proteins Western blot statistical chart. cP<0.01, compared with control group; eP<0.05, fP<0.01, compared with Hypoxia group.

Fig.5 mRNA levels of FDX1, LIAS, DLAT and DLD in lung tissues of each group ($ \overline{x} $±s, n=6) A: relative mRNA level of FDX1; B: relative mRNA level of LIAS; C: relative mRNA level of DLAT; D: relative mRNA level of DLD. cP<0.01, compared with control group; eP<0.05, fP<0.01, compared with Hypoxia group.

Fig.6 Expression levels of JC-1 and ATP content in lung tissues of eachgroup ($ \overline{x} $±s, n=5) A: relative JC-1 level of each group; B: ATP concentration in each group. cP<0.01, compared with control group; fP<0.01, compared with Hypoxia group.

Fig.7 Expression levels of PDHA1 and CA concentrations in lung tissues of each group ($ \overline{x} $±s,n=5) A: expression level of PDHA1; B: expression level of CA. cP<0.01, comparedwith control group; eP<0.05, fP<0.01, compared with Hypoxia group.

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