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1.
Physiol Genomics ; 54(6): 206-219, 2022 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-35467982

RESUMO

Transcriptomic analysis in metabolically active tissues allows a systems genetics approach to identify causal genes and networks involved in metabolic disease. Outbred heterogeneous stock (HS) rats are used for genetic mapping of complex traits, but to-date, a systems genetics analysis of metabolic tissues has not been done. We investigated whether adiposity-associated genes and gene coexpression networks in outbred heterogeneous stock (HS) rats overlap those found in humans. We analyzed RNAseq data from adipose tissue of 415 male HS rats, correlated these transcripts with body weight (BW) and compared transcriptome signatures to two human cohorts: the "African American Genetics of Metabolism and Expression" and "Metabolic Syndrome in Men." We used weighted gene coexpression network analysis to identify adiposity-associated gene networks and mediation analysis to identify genes under genetic control whose expression drives adiposity. We identified 554 orthologous "consensus genes" whose expression correlates with BW in the rat and with body mass index (BMI) in both human cohorts. Consensus genes fell within eight coexpressed networks and were enriched for genes involved in immune system function, cell growth, extracellular matrix organization, and lipid metabolic processes. We identified 19 consensus genes for which genetic variation may influence BW via their expression, including those involved in lipolysis (e.g., Hcar1), inflammation (e.g., Rgs1), adipogenesis (e.g., Tmem120b), or no previously known role in obesity (e.g., St14 and Ms4a6a). Strong concordance between HS rat and human BW/BMI associated transcripts demonstrates translational utility of the rat model, while identification of novel genes expands our knowledge of the genetics underlying obesity.


Assuntos
Redes Reguladoras de Genes , Obesidade , Transcriptoma , Tecido Adiposo/metabolismo , Adiposidade/genética , Animais , Perfilação da Expressão Gênica , Humanos , Masculino , Obesidade/genética , Ratos
2.
Cytogenet Genome Res ; 160(1): 2-10, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31865307

RESUMO

Strumae ovarii are neoplasms composed of normal-appearing thyroid tissue that occur within the ovary and rarely spread to extraovarian sites. A unique case of struma ovarii with widespread dissemination detected 48 years after removal of a pelvic dermoid provided the opportunity to reexamine the molecular nature of this form of neoplasm. One tumor, from the heart, consisting of benign thyroid tissue was found to have whole-genome homozygosity. Another tumor from the right mandible composed of malignant-appearing thyroid tissue showed whole-genome homozygosity and a deletion of 7p, presumably the second hit that transformed it into a cancerous tumor. Specimens from 2 other cases of extraovarian struma confined to the abdomen and 8 of 9 cases of intraovarian struma showed genome-wide segmental homozygosity. These findings confirm errors in meiosis as the origin of struma ovarii. The histological and molecular findings further demonstrate that even when outside the ovary, strumae ovarii can behave nonaggressively until they receive a second hit, thereafter behaving like cancer.


Assuntos
Carcinoma/genética , Genoma Humano , Meiose , Neoplasias Ovarianas/genética , Estruma Ovariano/genética , Teratoma/genética , Adulto , Idoso , Carcinoma/diagnóstico , Feminino , Deleção de Genes , Neoplasias Cardíacas/genética , Neoplasias Cardíacas/secundário , Homozigoto , Humanos , Neoplasias Mandibulares/genética , Neoplasias Mandibulares/secundário , Pessoa de Meia-Idade , Recidiva Local de Neoplasia , Análise de Sequência com Séries de Oligonucleotídeos , Neoplasias Ovarianas/diagnóstico , Análise de Sequência de RNA , Estruma Ovariano/diagnóstico , Teratoma/diagnóstico , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/patologia
3.
J Am Soc Nephrol ; 28(4): 1093-1105, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-27821631

RESUMO

APOL1 G1 and G2 variants facilitate kidney disease in blacks. To elucidate the pathways whereby these variants contribute to disease pathogenesis, we established HEK293 cell lines stably expressing doxycycline-inducible (Tet-on) reference APOL1 G0 or the G1 and G2 renal-risk variants, and used Illumina human HT-12 v4 arrays and Affymetrix HTA 2.0 arrays to generate global gene expression data with doxycycline induction. Significantly altered pathways identified through bioinformatics analyses involved mitochondrial function; results from immunoblotting, immunofluorescence, and functional assays validated these findings. Overexpression of APOL1 by doxycycline induction in HEK293 Tet-on G1 and G2 cells led to impaired mitochondrial function, with markedly reduced maximum respiration rate, reserve respiration capacity, and mitochondrial membrane potential. Impaired mitochondrial function occurred before intracellular potassium depletion or reduced cell viability occurred. Analysis of global gene expression profiles in nondiseased primary proximal tubule cells from black patients revealed that the nicotinate phosphoribosyltransferase gene, responsible for NAD biosynthesis, was among the top downregulated transcripts in cells with two APOL1 renal-risk variants compared with those without renal-risk variants; nicotinate phosphoribosyltransferase also displayed gene expression patterns linked to mitochondrial dysfunction in HEK293 Tet-on APOL1 cell pathway analyses. These results suggest a pivotal role for mitochondrial dysfunction in APOL1-associated kidney disease.


Assuntos
Apolipoproteínas/genética , Nefropatias/genética , Lipoproteínas HDL/genética , Doenças Mitocondriais/genética , Apolipoproteína L1 , População Negra , Células Cultivadas , Feminino , Regulação da Expressão Gênica , Células HEK293 , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Risco
4.
Diabetes ; 72(1): 135-148, 2023 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-36219827

RESUMO

Despite the successes of human genome-wide association studies, the causal genes underlying most metabolic traits remain unclear. We used outbred heterogeneous stock (HS) rats, coupled with expression data and mediation analysis, to identify quantitative trait loci (QTLs) and candidate gene mediators for adiposity, glucose tolerance, serum lipids, and other metabolic traits. Physiological traits were measured in 1,519 male HS rats, with liver and adipose transcriptomes measured in >410 rats. Genotypes were imputed from low-coverage whole-genome sequencing. Linear mixed models were used to detect physiological and expression QTLs (pQTLs and eQTLs, respectively), using both single nucleotide polymorphism (SNP)- and haplotype-based models for pQTL mapping. Genes with cis-eQTLs that overlapped pQTLs were assessed as causal candidates through mediation analysis. We identified 14 SNP-based pQTLs and 19 haplotype-based pQTLs, of which 10 were in common. Using mediation, we identified the following genes as candidate mediators of pQTLs: Grk5 for fat pad weight and serum triglyceride pQTLs on Chr1, Krtcap3 for fat pad weight and serum triglyceride pQTLs on Chr6, Ilrun for a fat pad weight pQTL on Chr20, and Rfx6 for a whole pancreatic insulin content pQTL on Chr20. Furthermore, we verified Grk5 and Ktrcap3 using gene knockdown/out models, thereby shedding light on novel regulators of obesity.


Assuntos
Adiposidade , Insulinas , Ratos , Masculino , Humanos , Animais , Adiposidade/genética , Estudo de Associação Genômica Ampla , Obesidade/genética , Triglicerídeos , Insulinas/genética , Lipídeos , Polimorfismo de Nucleotídeo Único
5.
Proc Natl Acad Sci U S A ; 105(10): 3891-6, 2008 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-18292224

RESUMO

The apical sodium-dependent bile acid transporter (Asbt) is responsible for transport across the intestinal brush border membrane; however, the carrier(s) responsible for basolateral bile acid export into the portal circulation remains to be determined. Although the heteromeric organic solute transporter Ostalpha-Ostbeta exhibits many properties predicted for a candidate intestinal basolateral bile acid transporter, the in vivo functions of Ostalpha-Ostbeta have not been investigated. To determine the role of Ostalpha-Ostbeta in intestinal bile acid absorption, the Ostalpha gene was disrupted by homologous recombination in mice. Ostalpha(-/-) mice were physically indistinguishable from wild-type mice. In everted gut sac experiments, transileal transport of taurocholate was reduced by >80% in Ostalpha(-/-) vs. wild-type mice; the residual taurocholate transport was further reduced to near-background levels in gut sacs prepared from Ostalpha(-/-)Mrp3(-/-) mice. The bile acid pool size was significantly reduced (>65%) in Ostalpha(-/-) mice, but fecal bile acid excretion was not elevated. The decreased pool size in Ostalpha(-/-) mice resulted from reduced hepatic Cyp7a1 expression that was inversely correlated with ileal expression of fibroblast growth factor 15 (FGF15). These data indicate that Ostalpha-Ostbeta is essential for intestinal bile acid transport in mice. Unlike a block in intestinal apical bile acid uptake, genetic ablation of basolateral bile acid export disrupts the classical homeostatic control of hepatic bile acid biosynthesis.


Assuntos
Ácidos e Sais Biliares/metabolismo , Homeostase , Mucosa Intestinal/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Colesterol 7-alfa-Hidroxilase/metabolismo , Ácido Cólico/administração & dosagem , Ácido Cólico/farmacologia , Fezes/química , Regulação da Expressão Gênica/efeitos dos fármacos , Marcação de Genes , Homeostase/efeitos dos fármacos , Absorção Intestinal/efeitos dos fármacos , Mucosa Intestinal/efeitos dos fármacos , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/metabolismo , Intestinos/efeitos dos fármacos , Lipídeos/isolamento & purificação , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Proteínas de Membrana Transportadoras/deficiência , Camundongos , Camundongos Knockout , Modelos Biológicos , Fenótipo , Membrana Serosa/efeitos dos fármacos , Membrana Serosa/metabolismo
6.
Kidney Int Rep ; 5(6): 891-904, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32518871

RESUMO

INTRODUCTION: APOL1 G1 and G2 nephropathy-risk variants cause mitochondrial dysfunction and contribute to kidney disease. Analyses were performed to determine the genetic regulation of APOL1 and elucidate potential mechanisms in APOL1-nephropathy. METHODS: A global gene expression analysis was performed in human primary renal tubule cell lines derived from 50 African American individuals. Follow-up gene knock out, cell-based rescue, and microscopy experiments were performed. RESULTS: APOL1 genotypes did not alter APOL1 expression levels in the global gene expression analysis. Expression quantitative trait locus (eQTL) analysis in polyinosinic-polycytidylic acid (poly IC)-stimulated renal tubule cells revealed that single nucleotide polymorphism (SNP) rs513349 adjacent to BAK1 was a trans eQTL for APOL1 and a cis eQTL for BAK1; APOL1 and BAK1 were co-expressed in cells. BAK1 knockout in a human podocyte cell line resulted in diminished APOL1 protein, supporting a pivotal effect for BAK1 on APOL1 expression. Because BAK1 is involved in mitochondrial dynamics, mitochondrial morphology was examined in primary renal tubule cells and HEK293 Tet-on cells of various APOL1 genotypes. Mitochondria in APOL1 wild-type (G0G0) tubule cells maintained elongated morphology when stimulated by low-dose poly IC, whereas those with G1G1, G2G2, and G1G2 genotypes appeared to fragment. HEK293 Tet-on cells overexpressing APOL1 G0, G1, and G2 were created; G0 cells appeared to promote mitochondrial fusion, whereas G1 and G2 induced mitochondrial fission. The mitochondrial dynamic regulator Mdivi-1 significantly preserved cell viability and mitochondrial cristae structure and reversed mitochondrial fission induced by overexpression of G1 and G2. CONCLUSION: Results suggest the mitochondrial fusion/fission pathway may be a therapeutic target in APOL1-nephropathy.

7.
Anticancer Res ; 36(8): 4007-11, 2016 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-27466506

RESUMO

AIM: To identify differentially expressed genes (DEGs) between perineural invasion-positive (PP) and -negative (PN) cutaneous squamous cell cancers (CSCC). MATERIALS/METHODS: Forty CSCC samples with and without perineural invasion were processed for RNA isolation and hybridization to Affymetrix-U219 DNA microarrays. Raw gene expression data were normalized by Robust Multi-array Averaging (RMA) and log2 transformed. Gene expression-based classification models were created and accuracies evaluated using leave-one-out cross-validation. RESULTS: At a stringent limma p-value (p<0.001), 24 genes were differentially expressed between PP and PN samples. The cross-validated performance of the eight classification models exhibited a mean accuracy of 85-95%. Diagonal linear discriminant was most accurate at 95%, followed by Bayesian compound covariate at 94%. The poorest accuracy (85%) was observed for 1-Nearest neighbor and Support vector machines. CONCLUSION: Gene expression may distinguish between PP and PN CSCC. Understanding these gene patterns may potentiate more timely diagnosis of perineural invasion and guide comprehensive therapies.


Assuntos
Proteínas de Neoplasias/biossíntese , Neoplasias de Células Escamosas/genética , Neoplasias de Bainha Neural/genética , Neoplasias Cutâneas/genética , Teorema de Bayes , Biomarcadores Tumorais/biossíntese , Biomarcadores Tumorais/genética , Células Epiteliais/patologia , Regulação Neoplásica da Expressão Gênica , Humanos , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Proteínas de Neoplasias/genética , Neoplasias de Células Escamosas/complicações , Neoplasias de Células Escamosas/patologia , Neoplasias de Bainha Neural/complicações , Neoplasias de Bainha Neural/patologia , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Neoplasias Cutâneas/complicações , Neoplasias Cutâneas/patologia , Máquina de Vetores de Suporte
9.
J Biol Chem ; 280(8): 6960-8, 2005 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-15563450

RESUMO

Bile acids are transported across the ileal enterocyte brush border membrane by the well characterized apical sodium-dependent bile acid transporter (Asbt) Slc10a2; however, the carrier(s) responsible for transporting bile acids across the ileocyte basolateral membrane into the portal circulation have not been fully identified. Transcriptional profiling of wild type and Slc10a2 null mice was employed to identify a new candidate basolateral bile acid carrier, the heteromeric organic solute transporter (Ost)alpha-Ostbeta. By Northern blot analysis, Ostalpha and Ostbeta mRNA was detected only in mouse kidney and intestine, mirroring the horizontal gradient of expression of Asbt in the gastrointestinal tract. Analysis of Ostalpha and Ostbeta protein expression by immunohistochemistry localized both subunits to the basolateral surface of the mouse ileal enterocyte. The transport properties of Ostalpha-Ostbeta were analyzed in stably transfected Madin-Darby canine kidney cells. Co-expression of mouse Ostalpha-Ostbeta, but not the individual subunits, stimulated Na(+)-independent bile acid uptake and the apical-to-basolateral transport of taurocholate. In contrast, basolateral-to-apical transport was not affected by Ostalpha-Ostbeta expression. Co-expression of Ostalpha and Ostbeta was required to convert the Ostalpha subunit to a mature glycosylated endoglycosidase H-resistant form, suggesting that co-expression facilitates the trafficking of Ostalpha through the Golgi apparatus. Immunolocalization studies showed that co-expression was necessary for plasma membrane expression of both Ostalpha and Ostbeta. These results demonstrate that the mouse Ostalpha-Ostbeta heteromeric transporter is a basolateral bile acid carrier and may be responsible for bile acid efflux in ileum and other ASBT-expressing tissues.


Assuntos
Proteínas de Transporte , Íleo/química , Glicoproteínas de Membrana , Proteínas de Membrana Transportadoras/fisiologia , Animais , Ácidos e Sais Biliares/metabolismo , Linhagem Celular , Perfilação da Expressão Gênica , Humanos , Proteínas de Membrana Transportadoras/genética , Camundongos , Camundongos Knockout , Transportadores de Ânions Orgânicos Dependentes de Sódio/genética , Transporte Proteico , RNA Mensageiro/análise , Simportadores/genética , Ácido Taurocólico/metabolismo , Distribuição Tecidual
10.
J Biol Chem ; 278(36): 33920-7, 2003 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-12819193

RESUMO

The ileal apical sodium bile acid cotransporter participates in the enterohepatic circulation of bile acids. In patients with primary bile acid malabsorption, mutations in the ileal bile acid transporter gene (Slc10a2) lead to congenital diarrhea, steatorrhea, and reduced plasma cholesterol levels. To elucidate the quantitative role of Slc10a2 in intestinal bile acid absorption, the Slc10a2 gene was disrupted by homologous recombination in mice. Animals heterozygous (Slc10a2+/-) and homozygous (Slc10a2-/-) for this mutation were physically indistinguishable from wild type mice. In the Slc10a2-/- mice, fecal bile acid excretion was elevated 10- to 20-fold and was not further increased by feeding a bile acid binding resin. Despite increased bile acid synthesis, the bile acid pool size was decreased by 80% and selectively enriched in cholic acid in the Slc10a2-/- mice. On a low fat diet, the Slc10a2-/- mice did not have steatorrhea. Fecal neutral sterol excretion was increased only 3-fold, and intestinal cholesterol absorption was reduced only 20%, indicating that the smaller cholic acid-enriched bile acid pool was sufficient to facilitate intestinal lipid absorption. Liver cholesteryl ester content was reduced by 50% in Slc10a2-/- mice, and unexpectedly plasma high density lipoprotein cholesterol levels were slightly elevated. These data indicate that Slc10a2 is essential for efficient intestinal absorption of bile acids and that alternative absorptive mechanisms are unable to compensate for loss of Slc10a2 function.


Assuntos
Ácidos e Sais Biliares/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/fisiologia , Deleção de Genes , Fígado/metabolismo , Transportadores de Ânions Orgânicos Dependentes de Sódio , Simportadores , Alelos , Animais , Transporte Biológico , Northern Blotting , Colesterol/sangue , Colesterol/metabolismo , Colesterol 7-alfa-Hidroxilase/metabolismo , Ésteres do Colesterol/metabolismo , DNA Complementar/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Dados de Sequência Molecular , Mutação , RNA/metabolismo , Recombinação Genética , Fatores Sexuais , Sódio/metabolismo
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