mouse-mpc2  (Merck & Co)

Journal: Cellular and molecular gastroenterology and hepatology

Article Title: A Critical Role for the Mitochondrial Pyruvate Carrier in Hepatic Stellate Cell Activation.

doi: 10.1016/j.jcmgh.2025.101517

Figure Lengend Snippet: Figure 3. Lrat-MPC2-/-

Article Snippet: Antibodies used included: collagen 1 (Cell Signaling Technology; 72026), collagen 3 (ProteinTech; 22734-1-AP), HIF1-a (Cell Signaling Technology; 36169), MPC1 (Cell Signaling Technology; D2L9I), MPC2 (Cell Signaling Technology; D4I7G), smooth muscle 3 May 2025 6:52 pm ce JO 17 2241 2242 2243 2244 2245 2246 2247 2248 2249 2250 2251 2252 2253 2254 2255 2256 2257 2258 2259 2260 2261 2262 2263 2264 2265 2266 2267 2268 2269 2270 2271 2272 2273 2274 2275 2276 2277 2278 2279 2280 2281 2282 2283 2284 2285 2286 2287 2288 2289 2290 2291 2292 2293 2294 2295 2296 2297 2298 2299 2300 2301 2302 2303 2304 2305 2306 2307 2308 2309 2310 2311 2312 2313 2314 2315 2316 2317 2318 2319 2320 2321 2322 2323 2324 2325 2326 2327 2328 2329 2330 2331 2332 2333 2334 2335 2336 2337 2338 2339 2340 2341 2342 2343 2344 2345 2346 2347 2348 2349 2350 2351 2352 2353 2354 2355 2356 2357 2358 actin (SMA; Sigma-Aldrich; CBL171), a-tubulin (SigmaAldrich; T5168), b-ACTIN (Cell Signaling Technology; 3700), and COXIV (Cell Signaling Technology; 11967).

Techniques:

Journal: Cellular and molecular gastroenterology and hepatology

Article Title: A Critical Role for the Mitochondrial Pyruvate Carrier in Hepatic Stellate Cell Activation.

doi: 10.1016/j.jcmgh.2025.101517

Figure Lengend Snippet: Figure 4. MPC deletion in hepatic stellate cells protects mice from MASH-inducing diet (CDAA). At about 10 weeks of age, male (blue data points) and female (pink data points) Lrat-Mpc2-/- knockout (KO) and WT littermates received a CDAA diet for a period of 10 weeks. (A) Body weight change in experimental period reported as percentage relative to initial body weight. (B) Plasma levels of ALT at sacrifice. (C) Liver weight measured at sacrifice normalized to final body weight. (D) Hepatic gene expression measured by RT-qPCR and expressed relative to WT chow control group. All data expressed as mean ± SEM (n ¼ 3 [chow] or 10 [CDAA]/group). *P< .05; **P< .01; ***P< .001 vs WT mice on the same diet as determined by 2-way ANOVA, followed by Sidak’s test for multiple comparisons. (E) Representative liver sections with H&E and Sirius red staining in knockout (KO) and WT mice fed a CDAA diet with quantification ****P < .001.

Article Snippet: Antibodies used included: collagen 1 (Cell Signaling Technology; 72026), collagen 3 (ProteinTech; 22734-1-AP), HIF1-a (Cell Signaling Technology; 36169), MPC1 (Cell Signaling Technology; D2L9I), MPC2 (Cell Signaling Technology; D4I7G), smooth muscle 3 May 2025 6:52 pm ce JO 17 2241 2242 2243 2244 2245 2246 2247 2248 2249 2250 2251 2252 2253 2254 2255 2256 2257 2258 2259 2260 2261 2262 2263 2264 2265 2266 2267 2268 2269 2270 2271 2272 2273 2274 2275 2276 2277 2278 2279 2280 2281 2282 2283 2284 2285 2286 2287 2288 2289 2290 2291 2292 2293 2294 2295 2296 2297 2298 2299 2300 2301 2302 2303 2304 2305 2306 2307 2308 2309 2310 2311 2312 2313 2314 2315 2316 2317 2318 2319 2320 2321 2322 2323 2324 2325 2326 2327 2328 2329 2330 2331 2332 2333 2334 2335 2336 2337 2338 2339 2340 2341 2342 2343 2344 2345 2346 2347 2348 2349 2350 2351 2352 2353 2354 2355 2356 2357 2358 actin (SMA; Sigma-Aldrich; CBL171), a-tubulin (SigmaAldrich; T5168), b-ACTIN (Cell Signaling Technology; 3700), and COXIV (Cell Signaling Technology; 11967).

Techniques: Knock-Out, Clinical Proteomics, Gene Expression, Quantitative RT-PCR, Control, Staining

Journal: Cellular and molecular gastroenterology and hepatology

Article Title: A Critical Role for the Mitochondrial Pyruvate Carrier in Hepatic Stellate Cell Activation.

doi: 10.1016/j.jcmgh.2025.101517

Figure Lengend Snippet: Figure 5. Lrat-Mpc2-/- mice are protected from MASH-inducing diets (HFC). (A–C) Volcano plots of differentially expressed genes with P< .05 comparing (A) HFC vs LFD in WT mice, (B) knockout (KO) vs WT mice on LFD, and (C) KO vs WT mice on HFC diet. DEGs with Log fold change (LogFC) less than 0.5, or greater than 0.5, were highlighted in either blue or red, respectively. (D–E) GSEA of perturbations in Hallmark gene set collections when comparing (D) HFC vs LFD in WT mice and (E) KO vs WT mice on HFC diet. The arrows highlight hypoxia signaling pathways. Differential expression analysis was then performed to analyze for differences between conditions. RNA-seq was performed on liver tissue from WT and Lrat-Mpc2-/- knockout (KO) mice on either a LFD or HFC diet (n ¼ 5/group).

Article Snippet: Antibodies used included: collagen 1 (Cell Signaling Technology; 72026), collagen 3 (ProteinTech; 22734-1-AP), HIF1-a (Cell Signaling Technology; 36169), MPC1 (Cell Signaling Technology; D2L9I), MPC2 (Cell Signaling Technology; D4I7G), smooth muscle 3 May 2025 6:52 pm ce JO 17 2241 2242 2243 2244 2245 2246 2247 2248 2249 2250 2251 2252 2253 2254 2255 2256 2257 2258 2259 2260 2261 2262 2263 2264 2265 2266 2267 2268 2269 2270 2271 2272 2273 2274 2275 2276 2277 2278 2279 2280 2281 2282 2283 2284 2285 2286 2287 2288 2289 2290 2291 2292 2293 2294 2295 2296 2297 2298 2299 2300 2301 2302 2303 2304 2305 2306 2307 2308 2309 2310 2311 2312 2313 2314 2315 2316 2317 2318 2319 2320 2321 2322 2323 2324 2325 2326 2327 2328 2329 2330 2331 2332 2333 2334 2335 2336 2337 2338 2339 2340 2341 2342 2343 2344 2345 2346 2347 2348 2349 2350 2351 2352 2353 2354 2355 2356 2357 2358 actin (SMA; Sigma-Aldrich; CBL171), a-tubulin (SigmaAldrich; T5168), b-ACTIN (Cell Signaling Technology; 3700), and COXIV (Cell Signaling Technology; 11967).

Techniques: Knock-Out, Protein-Protein interactions, Quantitative Proteomics, RNA Sequencing

Journal: Cellular and molecular gastroenterology and hepatology

Article Title: A Critical Role for the Mitochondrial Pyruvate Carrier in Hepatic Stellate Cell Activation.

doi: 10.1016/j.jcmgh.2025.101517

Figure Lengend Snippet: Figure 8. MPC suppression markedly reduces several TCA cycle intermediates in LX2 cells treated with uniformly labeled 13C-glucose. (A) Schematic of glucose metabolism and the TCA cycle tracing the route of incorporation of 13C- glucose. (B–C) TGF-b-1 (5 ng/mL) was added into media and LX2 cells expressing either Scr control shRNA or shRNA against MPC2. Cells were then cultured overnight in the presence of uniformly labeled 13C-glucose. (B) Incorporation of 13C-glucose into glycolytic intermediates. (C) Incorporation of 13C-glucose into TCA cycle intermediates and amino acids; M2, M3, indicate 2 or 3 carbon atoms are derived from labeled glucose, respectively. (D) Relative abundance of intermediates in cells expressing MPC2 shRNA or Scr control shRNA. Data are expressed as mean ± SEM (n ¼ 3) from a representative experiment. *P < .05 vs Scr shRNA as determined by unpaired Student t-test. Schematics created with BioRender.com

Article Snippet: Antibodies used included: collagen 1 (Cell Signaling Technology; 72026), collagen 3 (ProteinTech; 22734-1-AP), HIF1-a (Cell Signaling Technology; 36169), MPC1 (Cell Signaling Technology; D2L9I), MPC2 (Cell Signaling Technology; D4I7G), smooth muscle 3 May 2025 6:52 pm ce JO 17 2241 2242 2243 2244 2245 2246 2247 2248 2249 2250 2251 2252 2253 2254 2255 2256 2257 2258 2259 2260 2261 2262 2263 2264 2265 2266 2267 2268 2269 2270 2271 2272 2273 2274 2275 2276 2277 2278 2279 2280 2281 2282 2283 2284 2285 2286 2287 2288 2289 2290 2291 2292 2293 2294 2295 2296 2297 2298 2299 2300 2301 2302 2303 2304 2305 2306 2307 2308 2309 2310 2311 2312 2313 2314 2315 2316 2317 2318 2319 2320 2321 2322 2323 2324 2325 2326 2327 2328 2329 2330 2331 2332 2333 2334 2335 2336 2337 2338 2339 2340 2341 2342 2343 2344 2345 2346 2347 2348 2349 2350 2351 2352 2353 2354 2355 2356 2357 2358 actin (SMA; Sigma-Aldrich; CBL171), a-tubulin (SigmaAldrich; T5168), b-ACTIN (Cell Signaling Technology; 3700), and COXIV (Cell Signaling Technology; 11967).

Techniques: Labeling, Expressing, Control, shRNA, Cell Culture, Derivative Assay

Journal: Cellular and molecular gastroenterology and hepatology

Article Title: A Critical Role for the Mitochondrial Pyruvate Carrier in Hepatic Stellate Cell Activation.

doi: 10.1016/j.jcmgh.2025.101517

Figure Lengend Snippet: Figure 11. MPC deficiency leads to impaired HIF1a activation by a metabolic mechanism. (A) The abundance of indicated proteins in LX2 cells treated with or without TGF-b-1 (5 ng/mL), glutamine (Gln, 2 mM), dm-aKG (5 mM), or D- or L- 2-hydroxyglutarate (5 mM). (B) Protein abundance of collagen 1, collagen 3, and HIF1a in LX2 cells expressing Scr or MPC2 shRNA treated with TGF- b-1 (5 ng/mL) and with or without DMOG (1 mM), a cell-permeable HIF1a stabilizer. Quantified protein fold change in LX2 cells expressing Scr or shMPC2 treated in the same way. Data expressed as mean ± SEM (n ¼ 3/group) and analyzed by 2-way ANOVA followed by uncorrected Fisher’s LSD test for multiple comparisons; *P < .05. (C) Selected HIF1a target genes from RNAseq data expressed as counts per million (CPM) and represented as mean ± SEM (n ¼ 5/group). *P < .05 as determined by 2-way ANOVA, followed by Sidak’s test for multiple comparisons. RNA-seq was performed on liver tissue from WT and Lrat- Mpc2-/- (KO) mice and placed on either an LFD or HFC diet (n ¼ 5/group).

Article Snippet: Antibodies used included: collagen 1 (Cell Signaling Technology; 72026), collagen 3 (ProteinTech; 22734-1-AP), HIF1-a (Cell Signaling Technology; 36169), MPC1 (Cell Signaling Technology; D2L9I), MPC2 (Cell Signaling Technology; D4I7G), smooth muscle 3 May 2025 6:52 pm ce JO 17 2241 2242 2243 2244 2245 2246 2247 2248 2249 2250 2251 2252 2253 2254 2255 2256 2257 2258 2259 2260 2261 2262 2263 2264 2265 2266 2267 2268 2269 2270 2271 2272 2273 2274 2275 2276 2277 2278 2279 2280 2281 2282 2283 2284 2285 2286 2287 2288 2289 2290 2291 2292 2293 2294 2295 2296 2297 2298 2299 2300 2301 2302 2303 2304 2305 2306 2307 2308 2309 2310 2311 2312 2313 2314 2315 2316 2317 2318 2319 2320 2321 2322 2323 2324 2325 2326 2327 2328 2329 2330 2331 2332 2333 2334 2335 2336 2337 2338 2339 2340 2341 2342 2343 2344 2345 2346 2347 2348 2349 2350 2351 2352 2353 2354 2355 2356 2357 2358 actin (SMA; Sigma-Aldrich; CBL171), a-tubulin (SigmaAldrich; T5168), b-ACTIN (Cell Signaling Technology; 3700), and COXIV (Cell Signaling Technology; 11967).

Techniques: Activation Assay, Quantitative Proteomics, Expressing, shRNA, RNA Sequencing

Journal: eLife

Article Title: Paradoxical neuronal hyperexcitability in a mouse model of mitochondrial pyruvate import deficiency

doi: 10.7554/eLife.72595

Figure Lengend Snippet:

Article Snippet: Antibody , anti-MPC2 (Mouse) , Millipore (MABS1914) , MABS1914 , 1/500 (WB).

Techniques: Recombinant, Plasmid Preparation, TUNEL Assay, Software, Microscopy

Journal: The Journal of Biological Chemistry

Article Title: Decreased Mitochondrial Pyruvate Transport Activity in the Diabetic Heart

doi: 10.1074/jbc.M116.753509

Figure Lengend Snippet: Akita heart mitochondria have impaired pyruvate supported respiration, PDH activity, and pyruvate transport. A, mitochondria were isolated from control and Akita hearts. Respiration was measured by a fiber optic oxygen measurement system with 10 mm malate and either 30 μm PC or the indicated amounts of pyruvate. State 3 was initiated by the addition of 0.5 mm ADP. Representative oxygen traces are shown. B, state 3 respiration rates were quantified and are shown either as specific activities (left) or as the percentage of Akita relative to control rates compared on a day-by-day basis (right; n = 5–6). C, mitochondria were incubated with the indicated amounts of pyruvate for 2.0 min at room temperature. 0.5 mm ADP was added as indicated, and samples were incubated an additional minute. PDH activity was then measured as described under “Experimental Procedures” (n = 4). D, pyruvate uptake was measured in isolated mitochondria as described under “Experimental Procedures” (n = 3). E, MPC1 and MPC2 levels were measured by Western blotting (WB) analysis as described under “Experimental Procedures” (n = 5). The MPC1 and MPC2 Western blots reveal single bands at the expected molecular masses and are cropped for clarity. LA, lipoic acid. Experimental points are from unique mitochondrial preparations, and error bars are the standard deviation. *, p < 0.05, unpaired Student's t test; NS, not significant.

Article Snippet: Point mutations were engineered into the mouse cDNA of MPC2-Myc (OriGene, MR200690) using site-directed mutagenesis to generate K19R, K19Q, K26R, and K26Q and K19R/K26R and K19Q/K26Q double mutations.

Techniques: Activity Assay, Isolation, Incubation, Western Blot, Standard Deviation

Journal: The Journal of Biological Chemistry

Article Title: Decreased Mitochondrial Pyruvate Transport Activity in the Diabetic Heart

doi: 10.1074/jbc.M116.753509

Figure Lengend Snippet: Acetylation of MPC2 at Lys-19 and Lys-26 is significantly increased in Akita heart mitochondria. A, representative Western blots (WB), on duplicate samples, of MPC2 immunoprecipitated (IP) from wild type or Akita heart mitochondria. B, a selected reaction monitoring technique demonstrated that lysines 19 and 26 are significantly more acetylated in Akita heart mitochondria than in wild types (n = 3). *, p < 0.05, unpaired Student's t test. Error bars are the standard deviation.

Article Snippet: Point mutations were engineered into the mouse cDNA of MPC2-Myc (OriGene, MR200690) using site-directed mutagenesis to generate K19R, K19Q, K26R, and K26Q and K19R/K26R and K19Q/K26Q double mutations.

Techniques: Western Blot, Immunoprecipitation, Standard Deviation

Journal: The Journal of Biological Chemistry

Article Title: Decreased Mitochondrial Pyruvate Transport Activity in the Diabetic Heart

doi: 10.1074/jbc.M116.753509

Figure Lengend Snippet: The double acetylation mimetic of Lys-19 and Lys-26 (K19Q/K26Q) decreases the pyruvate-dependent cellular oxygen consumption rate. A, H9c2 cells were placed in medium containing pyruvate as the sole nutrient source, and the OCR was measured as described under “Experimental Procedures.” CHC (0.1 mm) was added just prior to the beginning of the experiment as indicated. OCR measurements taken were basal (1), post-oligomycin (2), post-FCCP (3), and post-antimycin A (4). B, the maximal OCR was calculated as the post-FCCP OCR minus the post-oligomycin OCR (n = 4). C, wild type Myc-MPC2 and RR and QQ Myc-tagged MPC2 were expressed in H9c2 cells. A representative Western blot (WB) is shown indicating protein expression levels. D and E, the effects of exogenously expressed Myc-MPC2 on OCR (D) and maximal (Max) OCR (E) were evaluated. F, a schematic representation of the proposed transmembrane domains and topology of MPC2 (adapted from Ref. 36) and relative positions of Lys-19 and Lys-26. IMM, inner mitochondrial membrane; IMS, intermembrane space. n = 4; *, p < 0.05, unpaired Student's t test. Error bars are the standard deviation.

Article Snippet: Point mutations were engineered into the mouse cDNA of MPC2-Myc (OriGene, MR200690) using site-directed mutagenesis to generate K19R, K19Q, K26R, and K26Q and K19R/K26R and K19Q/K26Q double mutations.

Techniques: Western Blot, Expressing, Standard Deviation