Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 132
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Artigo em Inglês | MEDLINE | ID: mdl-31778829

RESUMO

BACKGROUND & AIMS: Self-renewal and multipotent differentiation are cardinal properties of intestinal stem cells (ISCs), mediated in part by WNT and NOTCH signaling. Although these pathways are well characterized, the molecular mechanisms that control the 'stemness' of ISCs are still not well defined. Here, we investigated the role of Krüppel-like factor 5 (KLF5) in regulating ISC functions. METHODS: We performed studies in adult Lgr5EGFP-IRES-creERT2;Rosa26LSLtdTomato (Lgr5Ctrl) and Lgr5EGFP-IRES-creERT2;Klf5fl/fl;Rosa26LSLtdTomato (Lgr5ΔKlf5) mice. Mice were injected with tamoxifen to activate Cre recombinase, which deletes Klf5 from the intestinal epithelium in Lgr5ΔKlf5 but not Lgr5Crtl mice. In experiments involving irradiation, mice were subjected to 12 Gy total body irradiation (TBI). Tissues were collected for immunofluorescence (IF) analysis and next generation sequencing. Oganoids were derived from fluoresecence activated cell sorted- (FACS-) single cells from tamoxifen-treated Lgr5ΔKlf5 or Lgr5Crtl mice and examined by immunofluorescence stain. RESULTS: Lgr5+ ISCs lacking KLF5 proliferate faster than control ISCs but fail to self-renew, resulting in a depleted ISC compartment. Transcriptome analysis revealed that Klf5-null Lgr5+ cells lose ISC identity and prematurely differentiate. Following irradiation injury, which depletes Lgr5+ ISCs, reserve Klf5-null progenitor cells fail to dedifferentiate and regenerate the epithelium. Absence of KLF5 inactivates numerous selected enhancer elements and direct transcriptional targets including canonical WNT-and NOTCH-responsive genes. Analysis of human intestinal tissues showed increased levels of KLF5 in the regenerating epithelium as compared to those of healthy controls. CONCLUSION: 2We conclude that ISC self-renewal, lineage specification, and precursor dedifferentiation require KLF5, by its ability to regulate epigenetic and transcriptional activities of ISC-specific gene sets. These findings have the potential for modulating ISC functions by targeting KLF5 in the intestinal epithelium.

2.
Gastroenterology ; 157(5): 1413-1428.e11, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31352001

RESUMO

BACKGROUND & AIMS: Obesity is a risk factor for pancreatic cancer. In mice, a high-fat diet (HFD) and expression of oncogenic KRAS lead to development of invasive pancreatic ductal adenocarcinoma (PDAC) by unknown mechanisms. We investigated how oncogenic KRAS regulates the expression of fibroblast growth factor 21, FGF21, a metabolic regulator that prevents obesity, and the effects of recombinant human FGF21 (rhFGF21) on pancreatic tumorigenesis. METHODS: We performed immunohistochemical analyses of FGF21 levels in human pancreatic tissue arrays, comprising 59 PDAC specimens and 45 nontumor tissues. We also studied mice with tamoxifen-inducible expression of oncogenic KRAS in acinar cells (KrasG12D/+ mice) and fElasCreERT mice (controls). KrasG12D/+ mice were placed on an HFD or regular chow diet (control) and given injections of rhFGF21 or vehicle; pancreata were collected and analyzed by histology, immunoblots, quantitative polymerase chain reaction, and immunohistochemistry. We measured markers of inflammation in the pancreas, liver, and adipose tissue. Activity of RAS was measured based on the amount of bound guanosine triphosphate. RESULTS: Pancreatic tissues of mice expressed high levels of FGF21 compared with liver tissues. FGF21 and its receptor proteins were expressed by acinar cells. Acinar cells that expressed KrasG12D/+ had significantly lower expression of Fgf21 messenger RNA compared with acinar cells from control mice, partly due to down-regulation of PPARG expression-a transcription factor that activates Fgf21 transcription. Pancreata from KrasG12D/+ mice on a control diet and given injections of rhFGF21 had reduced pancreatic inflammation, infiltration by immune cells, and acinar-to-ductal metaplasia compared with mice given injections of vehicle. HFD-fed KrasG12D/+ mice given injections of vehicle accumulated abdominal fat, developed extensive inflammation, pancreatic cysts, and high-grade pancreatic intraepithelial neoplasias (PanINs); half the mice developed PDAC with liver metastases. HFD-fed KrasG12D/+ mice given injections of rhFGF21 had reduced accumulation of abdominal fat and pancreatic triglycerides, fewer pancreatic cysts, reduced systemic and pancreatic markers of inflammation, fewer PanINs, and longer survival-only approximately 12% of the mice developed PDACs, and none of the mice had metastases. Pancreata from HFD-fed KrasG12D/+ mice given injections of rhFGF21 had lower levels of active RAS than from mice given vehicle. CONCLUSIONS: Normal acinar cells from mice and humans express high levels of FGF21. In mice, acinar expression of oncogenic KRAS significantly reduces FGF21 expression. When these mice are placed on an HFD, they develop extensive inflammation, pancreatic cysts, PanINs, and PDACs, which are reduced by injection of FGF21. FGF21 also reduces the guanosine triphosphate binding capacity of RAS. FGF21 might be used in the prevention or treatment of pancreatic cancer.


Assuntos
Células Acinares/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Transformação Celular Neoplásica/metabolismo , Dieta Hiperlipídica , Fatores de Crescimento de Fibroblastos/metabolismo , Neoplasias Intraductais Pancreáticas/metabolismo , Neoplasias Pancreáticas/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Células Acinares/patologia , Animais , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Carcinoma Ductal Pancreático/prevenção & controle , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Regulação para Baixo , Fatores de Crescimento de Fibroblastos/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos Transgênicos , Mutação , PPAR gama/genética , PPAR gama/metabolismo , Cisto Pancreático/genética , Cisto Pancreático/metabolismo , Cisto Pancreático/patologia , Neoplasias Intraductais Pancreáticas/genética , Neoplasias Intraductais Pancreáticas/patologia , Neoplasias Intraductais Pancreáticas/prevenção & controle , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/prevenção & controle , Pancreatite/genética , Pancreatite/metabolismo , Pancreatite/patologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Transdução de Sinais , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
3.
Mol Cancer Ther ; 18(11): 1973-1984, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31358661

RESUMO

Krüppel-like factor 5 (KLF5), a member of the SP/KLF family of zinc finger transcription factors, is overexpressed in human colorectal cancer specimens, and this overabundance is associated with aggressive cancer development and progression. We demonstrated that mice haploinsufficient for Klf5 had reduced intestinal tumor burden in the background of germline mutation in Apc, a gatekeeper of intestinal tumorigenesis. Based on a high-throughput screening strategy, we developed ML264, a small-molecule compound that inhibits KLF5, and showed that it inhibits growth of colorectal cancer in vitro and in vivo Through optimization efforts based on the structure of ML264, we have now identified a new lead compound, SR18662. We find that treatment with SR18662 significantly reduces growth and proliferation of colorectal cancer cells as compared with treatment with vehicle control, ML264, or SR15006 (a less optimized analogue from SAR efforts leading to SR18662). SR18662 showed improved efficacy in reducing the viability of multiple colorectal cancer cell lines. Flow cytometry analysis following SR18662 treatment showed an increase in cells captured in either S or G2-M phases of the cell cycle and a significant increase in the number of apoptotic cells, the latter a unique property compared with ML264 or SR15006. SR18662 treatment also reduces the expression of cyclins and components of the MAPK and WNT signaling pathways. Importantly, we observed a significant dose-dependent inhibition of xenograft growth in mice following SR18662 treatment that exceeded the effect of ML264 at equivalent doses. These findings support further development of SR18662 and its analogues for colorectal cancer therapy.

4.
Cell Commun Signal ; 17(1): 19, 2019 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-30819189

RESUMO

Oncogenic KRAS plays a vital role in controlling tumor metabolism by enhancing aerobic glycolysis. Obesity driven by chronic consumption of high-fat diet (HFD) is a major risk factor for oncogenic KRAS-mediated pancreatic ductal adenocarcinoma (PDAC). However, the role of HFD in KRAS-mediated metabolic reprogramming has been obscure. Here, by using genetically engineered mouse models expressing an endogenous level of KRASG12D in pancreatic acinar cells, we demonstrate that hyperactivation of KRASG12D by obesogenic HFD, as compared to carbohydrate-rich diet, is responsible for enhanced aerobic glycolysis that associates with critical pathogenic responses in the path towards PDAC. Ablation of Cox-2 attenuates KRAS hyperactivation leading to the reversal of both aggravated aerobic glycolysis and high-grade dysplasia under HFD challenge. Our data highlight a pivotal role of the cooperative interaction between obesity-ensuing HFD and oncogenic KRAS in driving the heightened aerobic glycolysis during pancreatic tumorigenesis and suggest that in addition to directly targeting KRAS and aerobic glycolysis pathway, strategies to target the upstream of KRAS hyperactivation may bear important therapeutic value.


Assuntos
Dieta Hiperlipídica , Glicólise , Obesidade/metabolismo , Oncogenes , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Aerobiose , Animais , Ciclo-Oxigenase 2/metabolismo , Carboidratos da Dieta , Camundongos , Modelos Biológicos , Obesidade/patologia , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia
5.
Mol Cancer Res ; 17(1): 165-176, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30108164

RESUMO

Krüppel-like factor 4 (KLF4), a zinc finger transcription factor, regulates homeostasis of the intestinal epithelium. Previously, it was reported that KLF4 functions as a tumor suppressor in colorectal cancer. Here, evidence demonstrates that KLF4 mitigates the development and progression of colitis-associated colorectal cancer (CAC) in a murine model. Mice with intestinal epithelium-specific deletion of Klf4 (Klf4ΔIS ) and control mice (Klf4fl/fl ) were used to explore the role of KLF4 in the development of azoxymethane (AOM) and dextran sodium sulfate (DSS)-induced CAC. Upon AOM and DSS treatment, KLF4 expression was progressively lost in colonic tissues of Klf4fl/fl mice during tumor development. Klf4ΔIS mice treated with AOM/DSS developed significantly more adenomatous polyps and carcinomas in situ in comparison with treated Klf4fl/fl mice. Adenomatous polyps, but not normal-appearing mucosa, from colonic tissues of treated Klf4ΔIS mice contained a significantly increased number of mitotic cells with more than 2 centrosomes relative to treated control mice. KLF4 and p53 colocalize to the centrosomes in mouse embryonic fibroblasts (MEF). Absence of KLF4 in Klf4-/- MEFs inhibits and its overexpression restores p53 localization to the centrosomes in Klf4-/- MEFs. IMPLICATIONS: Taken together, these results indicate that KLF4 plays a protective role against progression of CAC by guarding against genetic instability.


Assuntos
Colite/genética , Colite/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Mucosa Intestinal/metabolismo , Fatores de Transcrição Kruppel-Like/deficiência , Animais , Carcinogênese , Proliferação de Células/fisiologia , Centrossomo/metabolismo , Colite/patologia , Neoplasias Colorretais/patologia , Deleção de Genes , Mucosa Intestinal/patologia , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Sistema de Sinalização das MAP Quinases , Camundongos , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
6.
Diabetes ; 67(11): 2420-2433, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30115650

RESUMO

Mitochondrial injury is uniformly observed in several murine models as well as in individuals with diabetic kidney disease (DKD). Although emerging evidence has highlighted the role of key transcriptional regulators in mitochondrial biogenesis, little is known about the regulation of mitochondrial cytochrome c oxidase assembly in the podocyte under diabetic conditions. We recently reported a critical role of the zinc finger Krüppel-like factor 6 (KLF6) in maintaining mitochondrial function and preventing apoptosis in a proteinuric murine model. In this study, we report that podocyte-specific knockdown of Klf6 increased the susceptibility to streptozotocin-induced DKD in the resistant C57BL/6 mouse strain. We observed that the loss of KLF6 in podocytes reduced the expression of synthesis of cytochrome c oxidase 2 with resultant increased mitochondrial injury, leading to activation of the intrinsic apoptotic pathway under diabetic conditions. Conversely, mitochondrial injury and apoptosis were significantly attenuated with overexpression of KLF6 in cultured human podocytes under hyperglycemic conditions. Finally, we observed a significant reduction in glomerular and podocyte-specific expression of KLF6 in human kidney biopsies with progression of DKD. Collectively, these data suggest that podocyte-specific KLF6 is critical to preventing mitochondrial injury and apoptosis under diabetic conditions.


Assuntos
Diabetes Mellitus Experimental/metabolismo , Nefropatias Diabéticas/metabolismo , Fator 6 Semelhante a Kruppel/metabolismo , Mitocôndrias/metabolismo , Podócitos/metabolismo , Proteinúria/metabolismo , Animais , Apoptose/fisiologia , Pressão Sanguínea/fisiologia , Diabetes Mellitus Experimental/patologia , Nefropatias Diabéticas/patologia , Taxa de Filtração Glomerular/fisiologia , Humanos , Rim/metabolismo , Rim/patologia , Fator 6 Semelhante a Kruppel/genética , Camundongos , Mitocôndrias/patologia , Podócitos/patologia , Proteinúria/patologia
7.
J Am Soc Nephrol ; 29(10): 2529-2545, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30143559

RESUMO

BACKGROUND: Podocyte injury is the hallmark of proteinuric kidney diseases, such as FSGS and minimal change disease, and destabilization of the podocyte's actin cytoskeleton contributes to podocyte dysfunction in many of these conditions. Although agents, such as glucocorticoids and cyclosporin, stabilize the actin cytoskeleton, systemic toxicity hinders chronic use. We previously showed that loss of the kidney-enriched zinc finger transcription factor Krüppel-like factor 15 (KLF15) increases susceptibility to proteinuric kidney disease and attenuates the salutary effects of retinoic acid and glucocorticoids in the podocyte. METHODS: We induced podocyte-specific KLF15 in two proteinuric murine models, HIV-1 transgenic (Tg26) mice and adriamycin (ADR)-induced nephropathy, and used RNA sequencing of isolated glomeruli and subsequent enrichment analysis to investigate pathways mediated by podocyte-specific KLF15 in Tg26 mice. We also explored in cultured human podocytes the potential mediating role of Wilms Tumor 1 (WT1), a transcription factor critical for podocyte differentiation. RESULTS: In Tg26 mice, inducing podocyte-specific KLF15 attenuated podocyte injury, glomerulosclerosis, tubulointerstitial fibrosis, and inflammation, while improving renal function and overall survival; it also attenuated podocyte injury in ADR-treated mice. Enrichment analysis of RNA sequencing from the Tg26 mouse model shows that KLF15 induction activates pathways involved in stabilization of actin cytoskeleton, focal adhesion, and podocyte differentiation. Transcription factor enrichment analysis, with further experimental validation, suggests that KLF15 activity is in part mediated by WT1. CONCLUSIONS: Inducing podocyte-specific KLF15 attenuates kidney injury by directly and indirectly upregulating genes critical for podocyte differentiation, suggesting that KLF15 induction might be a potential strategy for treating proteinuric kidney disease.


Assuntos
Proteínas de Ligação a DNA/biossíntese , Nefropatias/metabolismo , Podócitos/metabolismo , Proteinúria/metabolismo , Fatores de Transcrição/biossíntese , Citoesqueleto de Actina/metabolismo , Animais , Diferenciação Celular , Células Cultivadas , Proteínas de Ligação a DNA/genética , Modelos Animais de Doenças , Adesões Focais , Técnicas de Silenciamento de Genes , Glomerulosclerose Segmentar e Focal/genética , Glomerulosclerose Segmentar e Focal/metabolismo , Glomerulosclerose Segmentar e Focal/patologia , Humanos , Nefropatias/genética , Nefropatias/patologia , Fatores de Transcrição Kruppel-Like/antagonistas & inibidores , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Camundongos , Camundongos Transgênicos , Nefrose Lipoide/genética , Nefrose Lipoide/metabolismo , Nefrose Lipoide/patologia , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Podócitos/patologia , Proteinúria/genética , Proteinúria/patologia , Fatores de Transcrição/genética , Regulação para Cima , Proteínas WT1/antagonistas & inibidores , Proteínas WT1/genética , Proteínas WT1/metabolismo
8.
JCI Insight ; 3(12)2018 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-29925693

RESUMO

Pathologic glomerular epithelial cell (GEC) hyperplasia is characteristic of both rapidly progressive glomerulonephritis (RPGN) and subtypes of focal segmental glomerulosclerosis (FSGS). Although initial podocyte injury resulting in activation of STAT3 signals GEC proliferation in both diseases, mechanisms regulating this are unknown. Here, we show that the loss of Krüppel-like factor 4 (KLF4), a zinc-finger transcription factor, enhances GEC proliferation in both RPGN and FSGS due to dysregulated STAT3 signaling. We observed that podocyte-specific knockdown of Klf4 (C57BL/6J) increased STAT3 signaling and exacerbated crescent formation after nephrotoxic serum treatment. Interestingly, podocyte-specific knockdown of Klf4 in the FVB/N background alone was sufficient to activate STAT3 signaling, resulting in FSGS with extracapillary proliferation, as well as renal failure and reduced survival. In cultured podocytes, loss of KLF4 resulted in STAT3 activation and cell-cycle reentry, leading to mitotic catastrophe. This triggered IL-6 release into the supernatant, which activated STAT3 signaling in parietal epithelial cells. Conversely, either restoration of KLF4 expression or inhibition of STAT3 signaling improved survival in KLF4-knockdown podocytes. Finally, human kidney biopsy specimens with RPGN exhibited reduced KLF4 expression with a concomitant increase in phospho-STAT3 expression as compared with controls. Collectively, these results suggest the essential role of KLF4/STAT3 signaling in podocyte injury and its regulation of aberrant GEC proliferation.


Assuntos
Células Epiteliais/metabolismo , Glomérulos Renais/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Fator de Transcrição STAT3/metabolismo , Animais , Ciclo Celular , Proliferação de Células , Modelos Animais de Doenças , Células Epiteliais/patologia , Glomerulosclerose Segmentar e Focal , Humanos , Interleucina-6 , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Glomérulos Renais/patologia , Fatores de Transcrição Kruppel-Like/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Nefrite/sangue , Podócitos/patologia , Fatores de Transcrição
9.
Am J Cancer Res ; 8(4): 723-731, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29736317

RESUMO

Oncogenic K-Ras activation is a common mutational event in colorectal cancer. We previously showed that transcription factor, Krüppel-like factor 5 (KLF5), contributes to intestinal polyposis in mice with K-Ras activation. At 14 months of age, Villin-Cre/LSL-K-RasG12D mice developed small intestinal and colonic hyperplastic polyps while LSL-K-RasG12D had none. The intestinal crypts of Villin-Cre/LSL-K-RasG12D mice contained a higher number of mitotic figures and increased crypt heights compared to controls. The intestinal epithelium of Villin-Cre/LSL-K-RasG12D mice showed prolific KLF5 expression throughout and above the elongated crypts. In contrast, KLF5 expression was limited to the upper crypt region in the controls. The levels of K-Ras effectors were significantly increased in Villin-Cre/LSL-K-RasG12D as compared to controls. The Villin-Cre/LSL-K-RasG12D mice showed decreased survival upon treatment with azoxymethane (AOM) as compared to controls. Furthermore, loss of one of Klf5 alleles reduced levels of K-Ras effector proteins and prevented mortality of Villin-Cre/LSL-K-RasG12D mice upon AOM treatment. The Villin-Cre/LSL-K-RasG12D mice spontaneously develop hyperplastic intestinal polyps and display a hyper-proliferative intestinal phenotype with elongated crypts, increased numbers of mitotic figures, elevated expression of KLF5, and other pro-proliferative targets. Induction of colonic tumorigenesis with AOM is detrimental to Villin-Cre/LSL-K-RasG12D mice that is in part dependent of KLF5.

10.
Gastroenterology ; 154(5): 1494-1508.e13, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29248441

RESUMO

BACKGROUND & AIMS: Activating mutations in KRAS are detected in most pancreatic ductal adenocarcinomas (PDACs). Expression of an activated form of KRAS (KrasG12D) in pancreata of mice is sufficient to induce formation of pancreatic intraepithelial neoplasia (PanINs)-a precursor of PDAC. Pancreatitis increases formation of PanINs in mice that express KrasG12D by promoting acinar-to-ductal metaplasia (ADM). We investigated the role of the transcription factor Krüppel-like factor 5 (KLF5) in ADM and KRAS-mediated formation of PanINs. METHODS: We performed studies in adult mice with conditional disruption of Klf5 (Klf5fl/fl) and/or expression of KrasG12D (LSL-KrasG12D) via CreERTM recombinase regulated by an acinar cell-specific promoter (Ptf1a). Activation of KrasG12D and loss of KLF5 was achieved by administration of tamoxifen. Pancreatitis was induced in mice by administration of cerulein; pancreatic tissues were collected, analyzed by histology and immunohistochemistry, and transcriptomes were compared between mice that did or did not express KLF5. We performed immunohistochemical analyses of human tissue microarrays, comparing levels of KLF5 among 96 human samples of PDAC. UN-KC-6141 cells (pancreatic cancer cells derived from Pdx1-Cre;LSL-KrasG12D mice) were incubated with inhibitors of different kinases and analyzed in proliferation assays and by immunoblots. Expression of KLF5 was knocked down with small hairpin RNAs or CRISPR/Cas9 strategies; cells were analyzed in proliferation and gene expression assays, and compared with cells expressing control vectors. Cells were subcutaneously injected into flanks of syngeneic mice and tumor growth was assessed. RESULTS: Of the 96 PDAC samples analyzed, 73% were positive for KLF5 (defined as nuclear staining in more than 5% of tumor cells). Pancreata from Ptf1a-CreERTM;LSL-KrasG12D mice contained ADM and PanIN lesions, which contained high levels of nuclear KLF5 within these structures. In contrast, Ptf1a-CreERTM;LSL-KrasG12D;Klf5fl/fl mice formed fewer PanINs. After cerulein administration, Ptf1a-CreERTM;LSL-KrasG12D mice formed more extensive ADM than Ptf1a-CreERTM;LSL-KrasG12D;Klf5fl/fl mice. Pancreata from Ptf1a-CreERTM;LSL-KrasG12D;Klf5fl/fl mice had increased expression of the tumor suppressor NDRG2 and reduced phosphorylation (activation) of STAT3, compared with Ptf1a-CreERTM;LSL-KrasG12D mice. In UN-KC-6141 cells, PI3K and MEK signaling increased expression of KLF5; a high level of KLF5 increased proliferation. Cells with knockdown of Klf5 had reduced proliferation, compared with control cells, had reduced expression of ductal markers, and formed smaller tumors (71.61 ± 30.79 mm3 vs 121.44 ± 34.90 mm3 from control cells) in flanks of mice. CONCLUSION: Levels of KLF5 are increased in human PDAC samples and in PanINs of Ptf1a-CreERTM;LSL-KrasG12D mice, compared with controls. KLF5 disruption increases expression of NDRG2 and reduces activation of STAT3 and reduces ADM and PanINs formation in mice. Strategies to reduce KLF5 activity might reduce progression of acinar cells from ADM to PanIN and pancreatic tumorigenesis.


Assuntos
Carcinoma in Situ/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Proliferação de Células , Transformação Celular Neoplásica/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Neoplasias Pancreáticas/metabolismo , Animais , Carcinoma in Situ/genética , Carcinoma in Situ/patologia , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Ceruletídeo , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica , Genes ras , Humanos , Fatores de Transcrição Kruppel-Like/deficiência , Fatores de Transcrição Kruppel-Like/genética , Metaplasia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mutação , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Pancreatite/induzido quimicamente , Pancreatite/genética , Pancreatite/metabolismo , Pancreatite/patologia , Interferência de RNA , Transdução de Sinais , Fatores de Tempo , Transfecção , Carga Tumoral
11.
Curr Stem Cell Rep ; 4(1): 95, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-31329798

RESUMO

[This corrects the article DOI: 10.1007/s40778-017-0103-7.].

12.
Cell Tissue Res ; 370(3): 441-449, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28856432

RESUMO

Kruppel-like factor 4 (KLF4) is a zinc finger transcription factor that plays crucial roles during the development and maintenance of multiple organs. We and others have previously shown that KLF4 is involved in bone modeling and remodeling but roles played by KLF4 during skeletogenesis are still not fully understood. Here, we show that KLF4 is expressed in the epiphyseal growth plate and articular chondrocytes. Most articular chondrocytes expressed KLF4 in embryos but it localized only in a subset of superficial zone cells in postnatal mice. When KLF4 was overexpressed in chondrocytes in vitro, it severely repressed chondrocytic gene expressions. Global gene expression profiling of KLF4-transduced chondrocytes revealed matrix degrading proteinases of the matrix metalloproteinase and disintegrin and metalloproteinase with thrombospondin-1 domain families within the group of upregulated genes. Proteinase induction by KLF4 was alleviated by Trichostatin A treatment suggesting the possible involvement of epigenetic mechanisms on proteinase induction by KLF4. These results indicate the possible involvement of KLF4 in physiological and pathological aspects during cartilage development and maintenance.


Assuntos
Cartilagem Articular/metabolismo , Condrócitos/metabolismo , Endopeptidases/biossíntese , Fatores de Transcrição Kruppel-Like/metabolismo , Metaloproteinases da Matriz/biossíntese , Trombospondina 1/biossíntese , Animais , Células Cultivadas , Endopeptidases/genética , Regulação da Expressão Gênica no Desenvolvimento , Ácidos Hidroxâmicos/farmacologia , Masculino , Metaloproteinases da Matriz/genética , Camundongos , Camundongos Endogâmicos ICR , Inibidores da Síntese de Proteínas/farmacologia , Trombospondina 1/genética
13.
Am J Physiol Gastrointest Liver Physiol ; 313(5): G478-G491, 2017 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-28864500

RESUMO

Krüppel-like factor 5 (KLF5) is a member of the zinc finger family of transcription factors that regulates homeostasis of the intestinal epithelium. Previous studies suggested an indispensable role of KLF5 in maintaining intestinal barrier function. In the current study, we investigated the mechanisms by which KLF5 regulates colonic barrier function in vivo and in vitro. We used an inducible and a constitutive intestine-specific Klf5 knockout mouse models (Villin-CreERT2;Klf5fl/fl designated as Klf5ΔIND and Villin-Cre;Klf5fl/fl as Klf5ΔIS ) and studied an inducible KLF5 knockdown in Caco-2 BBe cells using a lentiviral Tet-on system (Caco-2 BBe KLF5ΔIND). Specific knockout of Klf5 in colonic tissues, either inducible or constitutive, resulted in increased intestinal permeability. The phenotype was accompanied by a significant reduction in Dsg2, which encodes desmoglein-2, a desmosomal cadherin, at both mRNA and protein levels. Transmission electron microscopy showed alterations of desmosomal morphology in both KLF5 knockdown Caco-2 BBe cells and Klf5 knockout mouse colonic tissues. Inducible knockdown of KLF5 in Caco-2BBe cells grown on Transwell plates led to impaired barrier function as evidenced by decreased transepithelial electrical resistance and increased paracellular permeability to fluorescein isothiocyanate-4 kDa dextran. Furthermore, DSG2 was significantly decreased in KLF5 knockdown cells, and DSG2 overexpression partially rescued the impaired barrier function caused by KLF5 knockdown. Electron microscopy studies demonstrated altered desmosomal morphology after KLF5 knockdown. In combination with chromatin immunoprecipitation analysis and promoter study, our data show that KLF5 regulates intestinal barrier function by mediating the transcription of DSG2, a gene encoding a major component of desmosome structures.NEW & NOTEWORTHY The study is original research on the direct function of a Krüppel-like factor on intestinal barrier function, which is commonly exerted by cell junctions, including tight junctions, adherens junctions, and desmosomes. Numerous previous studies were focused on tight junctions and adherens junctions. However, this study provided a new perspective on how the intestinal barrier function is regulated by KLF5 through DSG2, a component of desmosome complexes.


Assuntos
Colo/fisiologia , Fatores de Transcrição Kruppel-Like/fisiologia , Animais , Células CACO-2 , Desmogleína 2/biossíntese , Desmogleína 2/genética , Desmossomos/ultraestrutura , Impedância Elétrica , Regulação da Expressão Gênica/genética , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/ultraestrutura , Glicoproteínas de Membrana/biossíntese , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Permeabilidade , RNA Mensageiro/biossíntese , RNA Mensageiro/genética
14.
Stem Cell Reports ; 9(1): 247-263, 2017 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-28602611

RESUMO

The irreversibility of developmental processes in mammalian cells has been challenged by rising evidence that de-differentiation of hepatocytes occurs in adult liver. However, whether reversibility exists in mesenchymal stromal cell (MSC)-derived hepatocytes (dHeps) remains elusive. In this study, we find that hepatogenic differentiation (HD) of MSCs is a reversible process and is modulated by DNA methyltransferases (DNMTs). DNMTs are regulated by transforming growth factor ß1 (TGFß1), which in turn controls hepatogenic differentiation and de-differentiation. In addition, a stepwise reduction in TGFß1 concentrations in culture media increases DNMT1 and decreases DNMT3 in primary hepatocytes (Heps) and confers Heps with multi-differentiation potentials similarly to MSCs. Hepatic lineage reversibility of MSCs and lineage conversion of Heps are regulated by DNMTs in response to TGFß1. This previously unrecognized TGFß1-DNMTs-MSC-HD axis may further increase the understanding the normal and pathological processes in the liver, as well as functions of MSCs after transplantation to treat liver diseases.


Assuntos
Diferenciação Celular , Metilases de Modificação do DNA/metabolismo , Hepatócitos/citologia , Células-Tronco Mesenquimais/citologia , Animais , Linhagem da Célula , Células Cultivadas , DNA (Citosina-5-)-Metiltransferase 1/metabolismo , DNA (Citosina-5-)-Metiltransferases/metabolismo , Humanos , Fígado/citologia , Fígado/metabolismo , Células-Tronco Mesenquimais/metabolismo , Camundongos , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo
15.
Kidney Int ; 92(5): 1178-1193, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-28651950

RESUMO

Large epidemiological studies clearly demonstrate that multiple episodes of acute kidney injury contribute to the development and progression of kidney fibrosis. Although our understanding of kidney fibrosis has improved in the past two decades, we have limited therapeutic strategies to halt its progression. Myofibroblast differentiation and proliferation remain critical to the progression of kidney fibrosis. Although canonical Wnt signaling can trigger the activation of myofibroblasts in the kidney, mediators of Wnt inhibition in the resident progenitor cells are unclear. Recent studies demonstrate that the loss of a Krüppel-like factor 15 (KLF15), a kidney-enriched zinc-finger transcription factor, exacerbates kidney fibrosis in murine models. Here, we tested whether Klf15 mRNA and protein expression are reduced in late stages of fibrosis in mice that underwent unilateral ureteric obstruction, a model of progressive renal fibrosis. Knockdown of Klf15 in Foxd1-expressing cells (Foxd1-Cre Klf15fl/fl) increased extracellular matrix deposition and myofibroblast proliferation as compared to wildtype (Foxd1-Cre Klf15+/+) mice after three and seven days of ureteral obstruction. This was validated in mice receiving angiotensin II treatment for six weeks. In both these murine models, the increase in renal fibrosis was found in Foxd1-Cre Klf15fl/fl mice and accompanied by the activation of Wnt/ß-catenin signaling. Furthermore, knockdown of Klf15 in cultured mouse embryonic fibroblasts activated canonical Wnt/ß-catenin signaling, increased profibrotic transcripts, and increased proliferation after treatment with a Wnt1 ligand. Conversely, the overexpression of KLF15 inhibited phospho-ß-catenin (Ser552) expression in Wnt1-treated cells. Thus, KLF15 has a critical role in attenuating kidney fibrosis by inhibiting the canonical Wnt/ß-catenin pathway.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Nefropatias/patologia , Rim/patologia , Miofibroblastos/patologia , Fatores de Transcrição/metabolismo , Via de Sinalização Wnt , Angiotensina II/toxicidade , Animais , Proliferação de Células , Células Cultivadas , Proteínas de Ligação a DNA/genética , Modelos Animais de Doenças , Progressão da Doença , Fibrose , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Rim/citologia , Nefropatias/etiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Miofibroblastos/metabolismo , Fosforilação , RNA Mensageiro/metabolismo , Células Estromais/metabolismo , Células Estromais/patologia , Fatores de Transcrição/genética , Proteína Wnt1/metabolismo , beta Catenina/metabolismo
16.
Gastroenterology ; 152(8): 1845-1875, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28366734

RESUMO

Specificity proteins (SPs) and Krüppel-like factors (KLFs) belong to the family of transcription factors that contain conserved zinc finger domains involved in binding to target DNA sequences. Many of these proteins are expressed in different tissues and have distinct tissue-specific activities and functions. Studies have shown that SPs and KLFs regulate not only physiological processes such as growth, development, differentiation, proliferation, and embryogenesis, but pathogenesis of many diseases, including cancer and inflammatory disorders. Consistently, these proteins have been shown to regulate normal functions and pathobiology in the digestive system. We review recent findings on the tissue- and organ-specific functions of SPs and KLFs in the digestive system including the oral cavity, esophagus, stomach, small and large intestines, pancreas, and liver. We provide a list of agents under development to target these proteins.


Assuntos
Doenças do Sistema Digestório/metabolismo , Sistema Digestório/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Fatores de Transcrição Sp/metabolismo , Animais , Sistema Digestório/patologia , Sistema Digestório/fisiopatologia , Doenças do Sistema Digestório/genética , Doenças do Sistema Digestório/patologia , Doenças do Sistema Digestório/fisiopatologia , Regulação da Expressão Gênica , Humanos , Fatores de Transcrição Kruppel-Like/genética , Transdução de Sinais , Fatores de Transcrição Sp/genética
17.
Development ; 144(5): 737-754, 2017 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-28246209

RESUMO

Krüppel-like factors (KLFs) are a family of zinc-finger transcription factors that are found in many species. Recent studies have shown that KLFs play a fundamental role in regulating diverse biological processes such as cell proliferation, differentiation, development and regeneration. Of note, several KLFs are also crucial for maintaining pluripotency and, hence, have been linked to reprogramming and regenerative medicine approaches. Here, we review the crucial functions of KLFs in mammalian embryogenesis, stem cell biology and regeneration, as revealed by studies of animal models. We also highlight how KLFs have been implicated in human diseases and outline potential avenues for future research.


Assuntos
Desenvolvimento Embrionário , Fatores de Transcrição Kruppel-Like/metabolismo , Pulmão/embriologia , Células-Tronco/citologia , Animais , Diferenciação Celular , Linhagem da Célula , Feminino , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Filogenia , Regeneração , Fatores de Transcrição/metabolismo , Dedos de Zinco
18.
Gene ; 611: 27-37, 2017 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-28237823

RESUMO

Krüppel-like factor 4 (KLF4) is an evolutionarily conserved zinc finger-containing transcription factor that regulates diverse cellular processes such as cell growth, proliferation, and differentiation. Since its discovery in 1996, KLF4 has been gaining a lot of attention, particularly after it was shown in 2006 as one of four factors involved in the induction of pluripotent stem cells (iPSCs). Here we review the current knowledge about the different functions and roles of KLF4 in various tissue and organ systems.


Assuntos
Diferenciação Celular/genética , Proliferação de Células/genética , Regulação da Expressão Gênica , Células-Tronco Pluripotentes Induzidas/metabolismo , Fatores de Transcrição Kruppel-Like/genética , Animais , Sequência de Bases , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Homologia de Sequência de Aminoácidos
19.
Curr Stem Cell Rep ; 3(4): 320-332, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29497599

RESUMO

Purpose of Review: Intestinal epithelial cells show remarkable plasticity in regenerating the epithelium following radiation injury. In this review, we explore the regenerative capacity and mechanisms of various populations of intestinal stem cells (ISCs) in response to ionizing radiation. Recent Findings: Ionizing radiation targets mitotic cells that include "active" ISCs and progenitor cells. Lineage-tracing experiments showed that several different cell types identified by a single or combination of markers are capable of regenerating the epithelium, confirming that ISCs exhibit a high degree of plasticity. However, the identities of the contributing cells marked by various markers require further validation. Summary: Following radiation injury, quiescent and/or radioresistant cells become active stem cells to regenerate the epithelium. Looking forward, understanding the mechanisms by which ISCs govern tissue regeneration is crucial to determine therapeutic approaches to promote intestinal epithelial regeneration following injury.

20.
J Am Soc Nephrol ; 28(1): 166-184, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27288011

RESUMO

Podocyte injury is the inciting event in primary glomerulopathies, such as minimal change disease and primary FSGS, and glucocorticoids remain the initial and often, the primary treatment of choice for these glomerulopathies. Because inflammation is not readily apparent in these diseases, understanding the direct effects of glucocorticoids on the podocyte, independent of the immunomodulatory effects, may lead to the identification of targets downstream of glucocorticoids that minimize toxicity without compromising efficacy. Several studies showed that treatment with glucocorticoids restores podocyte differentiation markers and normal ultrastructure and improves cell survival in murine podocytes. We previously determined that Krüppel-like factor 15 (KLF15), a kidney-enriched zinc finger transcription factor, is required for restoring podocyte differentiation markers in mice and human podocytes under cell stress. Here, we show that in vitro treatment with dexamethasone induced a rapid increase of KLF15 expression in human and murine podocytes and enhanced the affinity of glucocorticoid receptor binding to the promoter region of KLF15 In three independent proteinuric murine models, podocyte-specific loss of Klf15 abrogated dexamethasone-induced podocyte recovery. Furthermore, knockdown of KLF15 reduced cell survival and destabilized the actin cytoskeleton in differentiated human podocytes. Conversely, overexpression of KLF15 stabilized the actin cytoskeleton under cell stress in human podocytes. Finally, the level of KLF15 expression in the podocytes and glomeruli from human biopsy specimens correlated with glucocorticoid responsiveness in 35 patients with minimal change disease or primary FSGS. Thus, these studies identify the critical role of KLF15 in mediating the salutary effects of glucocorticoids in the podocyte.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Proteínas de Ligação a DNA/fisiologia , Glucocorticoides/farmacologia , Podócitos/citologia , Podócitos/efeitos dos fármacos , Fatores de Transcrição/fisiologia , Adolescente , Adulto , Animais , Antígenos de Diferenciação/efeitos dos fármacos , Criança , Dexametasona/farmacologia , Feminino , Glomerulosclerose Segmentar e Focal/imunologia , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Nefrose Lipoide/imunologia , Adulto Jovem
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA