TRPV1 Activation Antagonizes High-Fat Diet-Induced Obesity at Thermoneutrality and Enhances UCP-1 Transcription via PRDM-16
"> Figure 1
<p>TRPV1 expression and activity in WAT and BAT. (<b>A</b>). Western blot showing TRPV1 expression in mouse tissues. HEK<sup>TRPV1</sup> is the positive control. (<b>B</b>). TRPV1 mRNA expression normalized to the 18s RNA in these tissues (<span class="html-italic">n</span> = 5). (<b>C</b>). Representative micrograph showing the immunohistochemical detection of TRPV1 expression in the iWAT preadipocytes of WT and TRPV1<sup>−/−</sup> mice fed various diets. (<b>D</b>). Quantification of the fluorescence intensity (arbitrary units). (<b>E</b>). Representative traces of CAP-stimulated TRPV1 currents in the primary brown preadipocytes of WT and TRPV1<sup>−/−</sup> mice at −60 mV. (<b>F</b>). Average currents ± S.E.M. in these cells (<span class="html-italic">n</span> = 6 to 8). (<b>G</b>). CAP-stimulated TRPV1 currents in NCD- or HFD (±CAP)-fed primary brown preadipocytes from WT mice. (<b>H</b>). Average currents ± S.E.M. in these cells (<span class="html-italic">n</span> = 9 to 11). ** <span class="html-italic">p</span> < 0.01, significantly different.</p> "> Figure 2
<p>Rectal temperatures of WT mice at ambient temperature (<b>A</b>) and in the TNZ (<b>B</b>). Rectal temperatures of WT mice fed the NCD or HFD or HFD + CAP at ambient temperature and in the TNZ measured every week from 6 weeks till 38 weeks of age using a thermometer. Average mean temperatures ± S.E.Ms. of these mice (<span class="html-italic">n</span> = 8 for each condition). ** <span class="html-italic">p</span> < 0.01, significantly different.</p> "> Figure 3
<p>CAP counters HFD-induced weight gain in WT mice at ambient temperature and at thermoneutrality. Weight gain plotted against the feeding week for NCD- or HFD (±CAP, 0.01% in diet)-fed WT and UCP-1<sup>−/−</sup> mice at ambient temperature (<b>A</b>,<b>B</b>) and at thermoneutrality (<b>E</b>,<b>F</b>). Mean energy and water intake (±S.E.M.) of these mice at ambient temperature (<b>C</b>,<b>D</b>) and at thermoneutrality (<b>G</b>,<b>H</b>). (<b>I</b>). Weight gain plotted against the feeding week for NCD- or HFD (±CAP, 0.01% in diet)-fed WT mice that received the same number of calories that WT mice received at ambient temperature.</p> "> Figure 4
<p>CAP feeding increases the respiratory quotient (respiratory exchange ratio, RER= VCO2/VO2) and energy expenditure (EE) in WT mice maintained at thermoneutrality. RER (<b>A</b>,<b>C</b>), VCO2 (<b>E</b>,<b>G</b>), VO2 (<b>F</b>,<b>H</b>), EE (<b>I</b>,<b>K</b>), and locomotor activity (<b>M</b>,<b>O</b>) of NCD- or HFD (± CAP, 0.01% in diet)-fed WT and UCP-1<sup>−/−</sup> mice at thermoneutrality. Means ± S.E.Ms. for the RER (<b>B</b>,<b>D</b>), EE (<b>J</b>,<b>L</b>), and locomotor activities (<b>N</b>,<b>P</b>) of these mice. ** represents statistical significance at <span class="html-italic">p</span> < 0.01 for <span class="html-italic">n</span> = 8 mice for each condition.</p> "> Figure 5
<p>Effect of CAP feeding on the mRNA levels of adipogenic and thermogenic genes in the BAT of NCD- or HFD (±CAP)-fed WT and UCP-1<sup>−/−</sup> mice at thermoneutrality. Mean mRNA levels ± S.E.Ms. for Bmp4 (<b>A</b>), Bmp8a (<b>B</b>), Bmp8b (<b>C</b>), Cidea (<b>D</b>), CoxII (<b>E</b>), Dio2 (<b>F</b>), Foxc2 (<b>G</b>), Pgc-1α (<b>H</b>), and Sirt-1 (<b>I</b>) in the BAT of these mice. For quantitative RT-PCR experiments, 18s ribosomal RNA was used as control. ** represents statistical significance at <span class="html-italic">p</span> < 0.01 for <span class="html-italic">n</span> = 4 experiments.</p> "> Figure 6
<p>Effect of CAP feeding on the Ucp-1 (<b>A</b>) and Prdm-16 (<b>B</b>) mRNAs normalized to 18s RNA in NCD- or HFD (± CAP)-fed WT and UCP-1<sup>−/−</sup> mice. Mean Ucp-1 and Prdm-16 mRNA levels normalized to 18s RNA ± S.E.Ms in the BAT of these mice. ** represents statistical significance at <span class="html-italic">p</span> < 0.01 for <span class="html-italic">n</span> = 4 experiments.</p> "> Figure 7
<p>CAP does not counter obesity in <sup>AD</sup>PRDM-16<sup>−/−</sup> mice. (<b>A</b>) Mean body weight gain in NCD- or HFD (± CAP, 0.01% in diet)-fed <sup>AT</sup>PRDM-16<sup>−/−</sup> mice (<span class="html-italic">n</span> = 4). (<b>B</b>) Average daily energy and water intake in these mice (<span class="html-italic">n</span> = 6). (<b>C</b>) UCP-1 mRNA levels in the sWAT and BAT of these mice (<span class="html-italic">n</span> = 6 experiments). ** <span class="html-italic">p</span> < 0.01.</p> "> Figure 8
<p>PRDM-16 overexpression increases UCP-1 transcription and CAP treatment enhances it further. (<b>A</b>) Micrographs showing UCP-1-GFP expression in HEK TRPV1 cells from the control (basal; 1), PRDM-16 (2), PRDM-16 + CAP (1 μM; 3), PRDM-16, CPZ (10 μM; TRPV1 inhibitor) + CAP (1 μM), or CAP (1 μM) treatment groups. The magnification is 10x, and the scale bar is 100 μm. (<b>B</b>) Mean intensity of UCP-1-GFP normalized to the control (basal) group ± S.E.M. for <span class="html-italic">n</span> = 3 independent experiments. ** <span class="html-italic">p</span> < 0.01.</p> "> Figure 9
<p>Model describing the effect of TRPV1 activation by CAP on PRDM-16-dependent UCP-1 expression and therefore thermogenesis. (<b>A</b>). The HFD inhibits TRPV1 [<a href="#B10-pharmaceuticals-17-01098" class="html-bibr">10</a>], SiRT-1, and PRDM-16 [<a href="#B9-pharmaceuticals-17-01098" class="html-bibr">9</a>]. Inhibition of SiRT-1 suppresses the deacetylation of PRDM-16 [<a href="#B9-pharmaceuticals-17-01098" class="html-bibr">9</a>] and PRDM-16-dependent UCP-1 transcription. (<b>B</b>). CAP counters the effect of the HFD. (<b>C</b>). In UCP-1 KO mice, CAP activates the TRPV1-SIRT-1-PRDM-16-dependent signaling axis, but it fails to stimulate thermogenesis.</p> ">
Abstract
:1. Introduction
2. Results
2.1. TRPV1 Expression and Activity in BAT and iWAT
2.2. Decreased Body Temperature in Diet-Induced Obese WT Mice at Ambient Temperature and in the Thermoneutral Zone
2.3. CAP Inhibited Weight Gain in WT and UCP1−/− Mice at Ambient Temperature but Inhibited HFD-Induced Weight Gain Only in WT Mice and Not in UCP−/− Mice in the TNZ
2.4. Metabolic Activity
2.5. White Adipogenic and Thermogenic Gene Expression
2.6. UCP-1 and PRDM-16 Gene Expression
2.7. CAP Fails to Counter HFD-Induced Weight Gain in Adipose Tissue-Specific PRDM16−/− Mice
2.8. PRDM-16 Overexpression Upregulates UCP-1 Transcription in HEK TRPV1 Cells
3. Discussion
4. Materials and Methods
4.1. Mouse Model of High-Fat Diet-Induced Obesity
4.2. Adipose Tissue-Specific PRDM-16 Knockout Mice
4.3. Measurement of Rectal Temperature in Mice
4.4. Metabolic Activity Measurement
4.5. Fat Tissue Isolation [43]
4.6. Isolation and Primary Culture of Brown Adipose Tissue Preadipocytes
4.7. Immunohistochemistry
4.8. Whole-Cell Patch Clamp of Primary Brown Adipocytes
4.9. Quantitative RT-PCR Measurements (qRT-PCR)
4.10. Immunoblotting
4.11. PRDM16 and GFP-UCP1 Overexpression and Stimulation in the HEK TRPV1 Cell Line
4.12. Drugs and Chemicals
4.13. Statistical Analyses
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Gene (Accession Number) | Forward Primer | Reverse Primer |
---|---|---|
18s (X00686) | accgcagctaggaataatgga | gcctcagttccgaaaacca |
Gapdh (NM_001411844) | cgtgccgcctggagaaacc | tggaagagtgggagttgctgttg |
mtrpv1 (NM_001001445) | caacaagaaggggcttacacc | tctggagaatgtaggccaagac |
sirt-1 (NM_019812) | tcgtggagacatttttaatcagg | gcttcatgatggcaagtgg |
prdm-16 (XM_03616442) | cagcacggtgaagccattc | gcgtgcatccgcttgtg |
ucp-1 (NM_009463) | cgactcagtccaagagtacttctcttc | gccggctgagatcttgtttc |
pgc-1α (NR_175325) | agagaggcagaagcagaaagcaat | attctgtccgcgttgtgtcagg |
Cidea (NM_007702) | atcacaactggcctggttacg | tactacccggtgtccatttct |
cox2 (AF378830) | ataaccgagtcgttctgccaat | tttcagagcattggccatagaa |
foxc2 (NM_013519) | gcaacccaacagcaaactttc | gacggcgtagctcgatagg |
dio2 (NM_010050) | gttgcttctgagccgctc | gctctgcactggcaaagtc |
bmp4(NM_007554) | ctccagtctggggaggag | gatgaggtgcccaggcac |
bmp8a (NM_007558) | aaccatgccatcttgcagtct | cagaggtggcactcagtttgg |
bmp8b (NM_007559) | tccaccaaccacgccactat | cagtaggcacacagcacacct |
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Baskaran, P.; Gustafson, N.; Chavez, N. TRPV1 Activation Antagonizes High-Fat Diet-Induced Obesity at Thermoneutrality and Enhances UCP-1 Transcription via PRDM-16. Pharmaceuticals 2024, 17, 1098. https://doi.org/10.3390/ph17081098
Baskaran P, Gustafson N, Chavez N. TRPV1 Activation Antagonizes High-Fat Diet-Induced Obesity at Thermoneutrality and Enhances UCP-1 Transcription via PRDM-16. Pharmaceuticals. 2024; 17(8):1098. https://doi.org/10.3390/ph17081098
Chicago/Turabian StyleBaskaran, Padmamalini, Noah Gustafson, and Nicolas Chavez. 2024. "TRPV1 Activation Antagonizes High-Fat Diet-Induced Obesity at Thermoneutrality and Enhances UCP-1 Transcription via PRDM-16" Pharmaceuticals 17, no. 8: 1098. https://doi.org/10.3390/ph17081098