Molecular Mechanisms Underlying TNFα-Induced Mitochondrial Biogenesis in Human Airway Smooth Muscle
Proinflammatory cytokines such as TNFα mediate airway inflammation. Previously, we showed that TNFα increases mitochondrial biogenesis in human ASM (hASM) cells, which is associated with increased PGC1α expression. We hypothesized that TNFα induces CREB and ATF1 phosphorylation (pCREB<sup>S133...
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MDPI AG
2023-03-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/24/6/5788 |
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| author | Debanjali Dasgupta Sanjana Mahadev Bhat Alexis L. Price Philippe Delmotte Gary C. Sieck |
| author_facet | Debanjali Dasgupta Sanjana Mahadev Bhat Alexis L. Price Philippe Delmotte Gary C. Sieck |
| author_sort | Debanjali Dasgupta |
| collection | DOAJ |
| description | Proinflammatory cytokines such as TNFα mediate airway inflammation. Previously, we showed that TNFα increases mitochondrial biogenesis in human ASM (hASM) cells, which is associated with increased PGC1α expression. We hypothesized that TNFα induces CREB and ATF1 phosphorylation (pCREB<sup>S133</sup> and pATF1<sup>S63</sup>), which transcriptionally co-activate PGC1α expression. Primary hASM cells were dissociated from bronchiolar tissue obtained from patients undergoing lung resection, cultured (one–three passages), and then differentiated by serum deprivation (48 h). hASM cells from the same patient were divided into two groups: TNFα (20 ng/mL) treated for 6 h and untreated controls. Mitochondria were labeled using MitoTracker green and imaged using 3D confocal microscopy to determine mitochondrial volume density. Mitochondrial biogenesis was assessed based on relative mitochondrial DNA (mtDNA) copy number determined by quantitative real-time PCR (qPCR). Gene and/or protein expression of pCREB<sup>S133</sup>, pATF1<sup>S63</sup>, PCG1α, and downstream signaling molecules (NRFs, TFAM) that regulate transcription and replication of the mitochondrial genome, were determined by qPCR and/or Western blot. TNFα increased mitochondrial volume density and mitochondrial biogenesis in hASM cells, which was associated with an increase in pCREB<sup>S133</sup>, pATF1<sup>S63</sup> and PCG1α expression, with downstream transcriptional activation of <i>NRF1, NRF2</i>, and <i>TFAM.</i> We conclude that TNFα increases mitochondrial volume density in hASM cells via a pCREB<sup>S133</sup>/pATF1<sup>S63</sup>/PCG1α-mediated pathway. |
| first_indexed | 2024-03-11T06:25:08Z |
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| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-11T06:25:08Z |
| publishDate | 2023-03-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-8614addf129547f9b633481918f9a1012023-11-17T11:39:01ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672023-03-01246578810.3390/ijms24065788Molecular Mechanisms Underlying TNFα-Induced Mitochondrial Biogenesis in Human Airway Smooth MuscleDebanjali Dasgupta0Sanjana Mahadev Bhat1Alexis L. Price2Philippe Delmotte3Gary C. Sieck4Department of Physiology & Biomedical Engineering, Mayo Clinic, Rochester, MN 55905, USADepartment of Physiology & Biomedical Engineering, Mayo Clinic, Rochester, MN 55905, USADepartment of Physiology & Biomedical Engineering, Mayo Clinic, Rochester, MN 55905, USADepartment of Physiology & Biomedical Engineering, Mayo Clinic, Rochester, MN 55905, USADepartment of Physiology & Biomedical Engineering, Mayo Clinic, Rochester, MN 55905, USAProinflammatory cytokines such as TNFα mediate airway inflammation. Previously, we showed that TNFα increases mitochondrial biogenesis in human ASM (hASM) cells, which is associated with increased PGC1α expression. We hypothesized that TNFα induces CREB and ATF1 phosphorylation (pCREB<sup>S133</sup> and pATF1<sup>S63</sup>), which transcriptionally co-activate PGC1α expression. Primary hASM cells were dissociated from bronchiolar tissue obtained from patients undergoing lung resection, cultured (one–three passages), and then differentiated by serum deprivation (48 h). hASM cells from the same patient were divided into two groups: TNFα (20 ng/mL) treated for 6 h and untreated controls. Mitochondria were labeled using MitoTracker green and imaged using 3D confocal microscopy to determine mitochondrial volume density. Mitochondrial biogenesis was assessed based on relative mitochondrial DNA (mtDNA) copy number determined by quantitative real-time PCR (qPCR). Gene and/or protein expression of pCREB<sup>S133</sup>, pATF1<sup>S63</sup>, PCG1α, and downstream signaling molecules (NRFs, TFAM) that regulate transcription and replication of the mitochondrial genome, were determined by qPCR and/or Western blot. TNFα increased mitochondrial volume density and mitochondrial biogenesis in hASM cells, which was associated with an increase in pCREB<sup>S133</sup>, pATF1<sup>S63</sup> and PCG1α expression, with downstream transcriptional activation of <i>NRF1, NRF2</i>, and <i>TFAM.</i> We conclude that TNFα increases mitochondrial volume density in hASM cells via a pCREB<sup>S133</sup>/pATF1<sup>S63</sup>/PCG1α-mediated pathway.https://www.mdpi.com/1422-0067/24/6/5788TNFαairway inflammationmitochondrial biogenesisCREBATF1PGC1α |
| spellingShingle | Debanjali Dasgupta Sanjana Mahadev Bhat Alexis L. Price Philippe Delmotte Gary C. Sieck Molecular Mechanisms Underlying TNFα-Induced Mitochondrial Biogenesis in Human Airway Smooth Muscle International Journal of Molecular Sciences TNFα airway inflammation mitochondrial biogenesis CREB ATF1 PGC1α |
| title | Molecular Mechanisms Underlying TNFα-Induced Mitochondrial Biogenesis in Human Airway Smooth Muscle |
| title_full | Molecular Mechanisms Underlying TNFα-Induced Mitochondrial Biogenesis in Human Airway Smooth Muscle |
| title_fullStr | Molecular Mechanisms Underlying TNFα-Induced Mitochondrial Biogenesis in Human Airway Smooth Muscle |
| title_full_unstemmed | Molecular Mechanisms Underlying TNFα-Induced Mitochondrial Biogenesis in Human Airway Smooth Muscle |
| title_short | Molecular Mechanisms Underlying TNFα-Induced Mitochondrial Biogenesis in Human Airway Smooth Muscle |
| title_sort | molecular mechanisms underlying tnfα induced mitochondrial biogenesis in human airway smooth muscle |
| topic | TNFα airway inflammation mitochondrial biogenesis CREB ATF1 PGC1α |
| url | https://www.mdpi.com/1422-0067/24/6/5788 |
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