Image_1_ATP-citrate lyase inhibitor improves ectopic lipid accumulation in the kidney in a db/db mouse model.tif

AimWe evaluated a novel treatment for obesity-related renal, an ATP-citrate lyase (ACL) inhibitor, to attenuate ectopic lipid accumulation (ELA) in the kidney and the ensuing inflammation.Materials and methodsAn ACL inhibitor was administered intragastrically to 12-week-old db/db mice for 30 days. The appearance of ELA was observed by staining kidney sections with Oil Red O, and the differences in tissue lipid metabolites were assessed by mass spectrometry. The anti-obesity and renoprotection effects of ACL inhibitors were observed by histological examination and multiple biochemical assays.ResultsUsing the AutoDock Vina application, we determined that among the four known ACL inhibitors (SB-204990, ETC-1002, NDI-091143, and BMS-303141), BMS-303141 had the highest affinity for ACL and reduced ACL expression in the kidneys of db/db mice. We reported that BMS-303141 administration could decrease the levels of serum lipid and renal lipogenic enzymes acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), HMG-CoA reductase (HMGCR), and diminish renal ELA in db/db mice. In addition, we found that reducing ELA improved renal injuries, inflammation, and tubulointerstitial fibrosis.ConclusionACL inhibitor BMS-303141 protects against obesity-related renal injuries.</p

Table_1_ATP-citrate lyase inhibitor improves ectopic lipid accumulation in the kidney in a db/db mouse model.docx

AimWe evaluated a novel treatment for obesity-related renal, an ATP-citrate lyase (ACL) inhibitor, to attenuate ectopic lipid accumulation (ELA) in the kidney and the ensuing inflammation.Materials and methodsAn ACL inhibitor was administered intragastrically to 12-week-old db/db mice for 30 days. The appearance of ELA was observed by staining kidney sections with Oil Red O, and the differences in tissue lipid metabolites were assessed by mass spectrometry. The anti-obesity and renoprotection effects of ACL inhibitors were observed by histological examination and multiple biochemical assays.ResultsUsing the AutoDock Vina application, we determined that among the four known ACL inhibitors (SB-204990, ETC-1002, NDI-091143, and BMS-303141), BMS-303141 had the highest affinity for ACL and reduced ACL expression in the kidneys of db/db mice. We reported that BMS-303141 administration could decrease the levels of serum lipid and renal lipogenic enzymes acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), HMG-CoA reductase (HMGCR), and diminish renal ELA in db/db mice. In addition, we found that reducing ELA improved renal injuries, inflammation, and tubulointerstitial fibrosis.ConclusionACL inhibitor BMS-303141 protects against obesity-related renal injuries.</p

Image_2_ATP-citrate lyase inhibitor improves ectopic lipid accumulation in the kidney in a db/db mouse model.tif

AimWe evaluated a novel treatment for obesity-related renal, an ATP-citrate lyase (ACL) inhibitor, to attenuate ectopic lipid accumulation (ELA) in the kidney and the ensuing inflammation.Materials and methodsAn ACL inhibitor was administered intragastrically to 12-week-old db/db mice for 30 days. The appearance of ELA was observed by staining kidney sections with Oil Red O, and the differences in tissue lipid metabolites were assessed by mass spectrometry. The anti-obesity and renoprotection effects of ACL inhibitors were observed by histological examination and multiple biochemical assays.ResultsUsing the AutoDock Vina application, we determined that among the four known ACL inhibitors (SB-204990, ETC-1002, NDI-091143, and BMS-303141), BMS-303141 had the highest affinity for ACL and reduced ACL expression in the kidneys of db/db mice. We reported that BMS-303141 administration could decrease the levels of serum lipid and renal lipogenic enzymes acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), HMG-CoA reductase (HMGCR), and diminish renal ELA in db/db mice. In addition, we found that reducing ELA improved renal injuries, inflammation, and tubulointerstitial fibrosis.ConclusionACL inhibitor BMS-303141 protects against obesity-related renal injuries.</p

The primer sequences for the preparation of promoter-reporter plasmids.

<p>Restriction endonuclease sequences are shown in bold.</p

Functional analysis of the human <i>FTO</i> promoter.

<p>Different sizes of 5′deletion fragments of the human <i>FTO</i> promoter were cloned into pGL3-Basic luciferase reporter plasmids. The reporter construct was transfected into HEK 293 and Hela cell lines and the reporter activity was measured. Relative firefly luciferase activities were averages of three independent transfections normalized to renilla control activities. Data are presented a mean±S.D. (*<i>P</i><0.05).</p

Putative binding sites in the <i>FTO</i> gene promoter region.

<p>The underlined sequences are putative binding sites for transcription factors in the <i>FTO</i> promoter region, based upon database searches using the TFsearch and Alibaba programs. The transcription start site (TSS) is indicated in the figure.</p

ChIP assays to test the binding of Foxa2 to the promoter of <i>FTO in vivo</i>.

<p>Formaldehyde cross-linked chromatin prepared from HEK 293 cells was immunoprecipitated with a Foxa2 antibody. PCR was performed with special primers to amplify a 156 bp DNA fragment containing Foxa2 binding sites. Sonicated decrosslinked DNA was used as positive input control for PCR. ChIP analysis reveals that Foxa2 could bind the sequence about 100 bp away from the transcription start site of <i>FTO</i>.</p

Characterization of the transcription factor binding elements by mutagenesis and luciferase assay.

<p>HEK 293 cells were transfected with the <i>FTO</i> promoter (PGL3-100), three mutant variants (mutated at the <i>Foxa2</i> site, USF site or both of them), pHD vector, pHD-Foxa2, negative control siRNA (NTC siRNA) or Foxa2 siRNA, and the promoter-less PGL3-basic construct. Relative luciferase activity (RLU), expressed as the fold induction relative to PGL3-basic vector, was measured. Results are presented as mean RLU±SE of three independent experiments (*<i>P</i><0.05).</p

Constitutive expression of miR-122 in cervical epithelial carcinoma cells.

<p>The constitutive expression of miR-122 were separately detected in several cell lines by RT-qPCR. U6 was employed as control.</p

Promotion of type I IFN signal pathway by miR-122.

<p>(A) The expression of OAS-1 and MxA mRNAs in SiHa cells after transfected with miR-122 or miR-NC for 24 h, 48 h and 72 h, evaluated by RT-qPCR. (B) The phosphorylation level of STAT1 after transfected with miR-122 or miR-NC plasmid for 24 h, 48 h and 72 h, evaluated by western blotting. (C) The expression of OAS-1 and MxA mRNAs in SiHa cells after transfected with AMO-122 and AMO-NC for 24 h, 48 h and 72 h, evaluated by RT-qPCR. (D) The phosphorylation level of STAT1 after transfected with AMO-122 and AMO-NC for 24 h, 48 h and 72 h, evaluated by western blotting. (*<i>p</i><0.05, **<i>p</i><0.01).</p