Clinical relevance of cholesterol homeostasis genes in colorectal cancer.

Accumulation of cholesterol is a well-known feature in cancer. Preclinical studies suggest the contribution of various cholesterol regulators in CRC. However, their clinical relevance remains poorly understood. The aim of the present study is to evaluate the expression of these modulators in CRC and elucidate their diagnostic and prognostic value. mRNA levels of HMGCR, SREBF2, NR1H3 and NR1H2 were downregulated in tumors in local and TCGA cohort. The expression of LDLR, ABCA1 and SCARB1 was not consistent in the two cohorts. Western Blot analysis showed the increased levels of LDLR and reduced levels of LXR in early stage patients. Tumoral SREBP2 levels were enhanced in early stage whereas decreased in late stage. The individual expression of HMGCR, SREBF2, NR1H3 and NR1H2 did not have the potential to be used as independent prognostic marker, however, the combined expression of these genes associated with poor clinical outcome independent of lymph node metastasis, distant metastasis and advanced stage. This work sheds light on deregulation of cholesterol uptake and efflux pathways and provides novel leads in the development of biomarkers and therapeutic regimens that can detect and target CRC at initial stages.

Duodenal Niemann-Pick C1-like 1 expression was negatively correlated with liver X receptor expression in nonalcoholic fatty liver disease.

BACKGROUND/AIMS: Intestinal cholesterol absorption includes intestinal Niemann-Pick C1-like 1 (NPC1L1) and is an important target pathway in nonalcoholic fatty liver disease (NAFLD). We investigated the expression of NPC1L1 and its correlation with liver X receptor (LXR) expression in peripheral mononuclear (PMN) cells in patients with NAFLD. METHODS: We evaluated intestinal expression of NPC1L1 in 25 NAFLD patients and 28 healthy controls. We calculated the mRNA expression levels of LXR and farnesoid X receptor (FXR), which are master players of cholesterol metabolism in PMN cells. The protein expression of ABCA1, ABCG5/8, NPC1L1, SREBP, LXR, FXR, and CD36 was measured on tissue samples from the duodenum and ileum. RESULTS: The expression of LXR (p = 0.01) and FXR (p = 0.03) in PMN cells was increased in the NAFLD group compared to the control group. Duodenal NPC1L1 decreased in the NAFLD group compared to the healthy controls (3.38 ± 1.4 vs. 2.42 ± 1.2, p = 0.05). NPC1L1 expression in the duodenum was negatively correlated with LXR expression in PMN cells. Expression of LXR and FXR in the ileum was also negatively correlated with the expression of LXR in PMN cells. CONCLUSION: Duodenal NPC1L1 expression was decreased in NAFLD and was negatively correlated with LXR expression in PMN cells.

Tumor LXR Expression is a Prognostic Marker for Patients with Hepatocellular Carcinoma.

Liver X receptor (LXR) activation exerts an anti-tumor effect. However, whether the tumor LXR expression has prognostic significance in hepatocellular carcinoma (HCC) patient has not been addressed yet. Primary HCC and the adjacent non-tumor tissues were obtained from 169 patients who underwent routine curative surgical treatment. All patients were followed for prognosis analyses. Tumor LXR was detected by immunohistochemical analysis. In in vitro study, several HCC cell lines were cultured for cellular protein detection of LXR and other cytokines, including nuclear factor kappa (NFκB), Matrix metalloproteinases 2 and 9 (MMP-2 and -9). Meanwhile, the invasion ability of cultured HCC cell lines was performed. We found that LXR expression status in tumor samples is associated with the clinical characteristics, such as tumor stage and metastasis, of HCC patients. Prognosis analysis shows that tumor LXR expression status is closely related to the post-operative outcome in HCC patients who underwent surgical treatment. Patients with low LXR expression have a significantly lower mean 5-year overall survival rate and mean overall survival period than those with high LXR level. Our in vitro data reveal that HCC cell lines had increased NF-κB, MMP2, MMP9 and invasive ability than normal cell line, which are suppressed by LXR activation via NFκB pathway. Our data suggest that LXR could be used as a biomarker for HCC prognosis. Further study is warranted to explore the molecular mechanism under which LXR regulates tumor behaves.

Krüppel-Like Factor 4 Regulation of Cholesterol-25-Hydroxylase and Liver X Receptor Mitigates Atherosclerosis Susceptibility.

BACKGROUND: Atherosclerosis is a multifaceted inflammatory disease involving cells in the vascular wall (eg, endothelial cells [ECs]), as well as circulating and resident immunogenic cells (eg, monocytes/macrophages). Acting as a ligand for liver X receptor (LXR), but an inhibitor of SREBP2 (sterol regulatory element-binding protein 2), 25-hydroxycholesterol, and its catalyzing enzyme cholesterol-25-hydroxylase (Ch25h) are important in regulating cellular inflammatory status and cholesterol biosynthesis in both ECs and monocytes/macrophages. METHODS: Bioinformatic analyses were used to investigate RNA-sequencing data to identify cholesterol oxidation and efflux genes regulated by Krüppel-like factor 4 (KLF4). In vitro experiments involving cultured ECs and macrophages and in vivo methods involving mice with Ch25h ablation were then used to explore the atheroprotective role of KLF4-Ch25h/LXR. RESULTS: Vasoprotective stimuli increased the expression of Ch25h and LXR via KLF4. The KLF4-Ch25h/LXR homeostatic axis functions through suppressing inflammation, evidenced by the reduction of inflammasome activity in ECs and the promotion of M1 to M2 phenotypic transition in macrophages. The increased atherosclerosis in apolipoprotein E-/-/Ch25h-/- mice further demonstrates the beneficial role of the KLF4-Ch25h/LXR axis in vascular function and disease. CONCLUSIONS: KLF4 transactivates Ch25h and LXR, thereby promoting the synergistic effects between ECs and macrophages to protect against atherosclerosis susceptibility.

28-Homocastasterone down regulates blood glucose, cholesterol, triglycerides, SREBP1c and activates LxR expression in normal & diabetic male rat.

Plant steroids are being recognized as influential secondary bio factors, assimilating in animal tissues through diet and affecting their cellular metabolic function to varying degree. They modulate catalytic and signaling functions in mammalian cells, affecting cellular homeostasis. The effect of phyto brassinosteroid ketoisoform 28-homocastasterone (28-HC), was assessed for its influence on blood glucose, plasma lipid and selective signal marker levels in normal and diabetic male wistar rat models. A 15 day oral feed regimen employing the experimental rat, noted that circulating blood glucose, cholesterol and triglyceride level in diabetic rat were markedly reduced by this compound. This study confirmed that the keto form had anti-hyperglycemic and anti-lipidemic potency associated with it and was available to man and animals in their diet. Western blots of marker protein, PCR amplicons of marker mRNA expressions and In Silico studies suggested that 28-HCeffect is being mediated through LxR molecular operatives in the rat cell.

Relationship of Liver X Receptors α and Endoglin Levels in Serum and Placenta with Preeclampsia.

BACKGROUND: Liver X receptor alpha (LXRα) and endoglin have been postulated to play roles in trophoblast invasion and lipid metabolic disturbances. However, the relationship between LXRα and endoglin levels in serum and placenta of patients with preeclampsia remains poorly understood. The objective of this study was to identify correlations between LXRα, endoglin and preeclampsia and provide new feasible methods of clinical prediction and treatment for preeclampsia. METHODS: We enrolled 45 patients with preeclampsia (24 with moderate preeclampsia and 21 with severe preeclampsia) and 15 normal pregnant women (control group) who were admitted to the Department of Obstetrics of the General Hospital of Beijing Command between October 2012 and July 2013 in this study. Serum and placental LXRα and endoglin levels were analyzed by enzyme-linked immunosorbent assay, real-time quantitative PCR, tissue microarray and immunohistochemistry. RESULTS: Serum and placental LXRα and endoglin levels were significantly higher in patients with preeclampsia than those in control group (P<0.05, each). Moreover, patients with severe preeclampsia displayed significantly higher LXRα and endoglin levels than those with moderate preeclampsia (P<0.05, each). The LXRα sensitivity, specificity and positive and negative predictive values were 66.00%, 80.00%, 89.19% and 48.48%, respectively, while those of endoglin levels were 62.00%, 85.00%, 91.18% and 47.22%, respectively. LXRα and endoglin levels in serum and placenta from patients with preeclampsia were positively correlated (serum: r = 0.486, P<0.01; placenta: r = 0.569, P<0.01). CONCLUSIONS: Elevated LXRα and endoglin levels may be associated with preeclampsia pathogenesis and development and could be used as potential predictors for this disorder.

Propriedades antiaterogênicas de novas tiazolidino-2, 4-dionas / Antiatherogenic properties of new thiazolidin-2, 4-diones

Tiazolidinadionas (TZDs) são agentes sensibilizadores de insulina que agem por ligação ao receptor gama ativado por proliferador de peroxissomos (PPARγ). Elas têm apresentado efeitos cardioprotetores em humanos e propriedades anti-aterogênicas em modelos animais. Estudos in vitro têm sugerido que esses efeitos anti-aterogênicos da ativação de PPARγ ocorrem por inibição da expressão de genes pro-inflamatórios e por aumentar o efluxo de colesterol via ativação dos receptores LXR-ABCA1. Entretanto, vários efeitos colaterais são associados ao tratamento com as TZDs, tornando necessária a pesquisa por novos compostos desta classe. Neste estudo, 14 novas tiazolidina-2,4- dionas, que são TZDs modificadas por bioisosterismo, foram avaliadas quanto à expressão de fatores aterogênicos e inflamatórios em linhagens de macrófagos J774 e RAW 264.7 e em camundongos com deleção genética para o receptor de LDL (LDLr-/-). Após a avaliação da citotoxicidade em macrófagos, foram eleitas cinco TZDs, denominadas de GQ-11, GQ-97, GQ-177, GQ-145 e LYSO-7. Três destas TZDs (GQ- 145, GQ-177 e LYSO-7) aumentaram significativamente a expressão de RNAm dos fatores de transcrição PPARγ1, PPARγ2 e do receptor CD36, assim como também aumentaram a expressão gênica de ABCA1 em 2.9, 3.5 e 6.7 vezes, respectivamente. Em adição, estas TZDs diminuíram a expressão gênica de iNOS, COX2, VCAM e IL-6 associado a redução na produção de nitritos, mas apenas a LYSO-7 reduziu significativamente a expressão desses genes quando comparada à rosiglitazona (RSG), além de diminuir a expressão da proteína-1 quimiotática para monócitos (MCP-1). No estudo experimental, os camundongos LDLr-/- machos foram alimentados com dieta hipercolesterolêmica por 16 semanas e quatro semanas antes da eutanásia receberam os derivados tiazolidínicos (20 mg/kg/dia) por gavagem. GQ-177 inibiu a progressão da placa aterosclerótica associada à aumento nas concentrações plasmáticas de HDL-C, com elevação na expressão de ABCA1, e redução da via inflamatória CD40-CD40L. LYSO-7 também mostrou inibição da aterogênese associada à redução das concentrações plasmáticas de colesterol total e triacilgliceróis, com diminuição na interação entre CD40-CD40L e expressão de citocinas inflamatórias. A GQ-145 não alterou os níveis plasmáticos dos lipídeos, mas aumentou a expressão de todos os genes pró-aterogênicos e pró-inflamatórios. Adicionalmente, as vias de ativação destas novas TZDs também foram estudadas por ensaio de luciferase, como gene repórter. A GQ-177 induziu ativação de PPARγ e ligação ao seu domínio, enquanto a LYSO-7 estimulou ativação de PPARα e PPARδ. Portanto, conclui-se que as novas TZDs, especialmente a GQ-177 e a LYSO-7, podem apresentar propriedades ateroprotetoras associadas ao transporte reverso de colesterol e aos efeitos antiinflamatórios, e poderiam ser uma alternativa promissora para o tratamento da aterosclerose. Porém, estudos complementares são requeridos para caracterizar as vias de sinalização intracelular, visto que as duas demonstraram ativar diferentes isotipos do fator de transcrição PPAR

Thiazolidinediones (TZDs) are insulin-sensitizing agents that act by binding to peroxisome proliferator-activated receptor-γ (PPARγ). They have been demonstrated to possess cardioprotective effects in humans and antiatherogenic properties in animal models. In vitro studies have also suggested that these antiatherogenic effects of PPARγ activation occur by inhibiting the inflammatory gene expression and by increasing cholesterol efflux via LXR-ABCA1 activation. However, several side effects are associated with TZDs treatment making necessary the search for new compounds. In this study, 14 new thiazolidine-2,4-diones, modified TZDs by bioisosterism, were tested for aterogenic and inflammtary factors in RAW 264.7 macrophages and in low-density lipoprotein receptor-deficient mice. After the citotoxicity evaluation in RAW 264.7 macrophages the TZDs named GQ-11, GQ-97, GQ-177, GQ-145 e LYSO-7 were selected for this study. Three of these TZDs (GQ-177, GQ-145 and LYSO-7) significantly increased the expression of PPARγ1, PPARγ2 and CD36 mRNA, and enhanced the expression of ABCA1 mRNA in 2.9, 3.5 and 6.7 fold, respectively. Moreover, they also significantly decreased the expression of iNOS, COX2, VCAM and IL-6 mRNA in relation to control, and these results are associated to reduction on nitrits concentration. In addition, LYSO-7 significantly reduced the expression of these genes when compared to rosiglitazone, and decreased expression of MCP1 mRNA. In the experimental study, male LDLr-/- mice were fed an atherogenic diet containing 0.5% cholesterol for 16 weeks, and 4 weeks before euthanasia they received TZDs (20mg/kg/ per day) by gavage. GQ-177 treatment inhibited progression of atherosclerotic plaque associated to increased plasma concentrations of HDL-C, with enhance of ABCA1 expression and reduction on CD40-CD40L interaction. LYSO-7 treatment also showed inhibition of the atherogenesis associated to decreased plasma concentrations of total cholesterol and TAG, with reduction on CD40-CD40L pathway and inflammatory cytokines expression.GQ-145 did not alter the lipid plasma levels and increased the expression of all pro-atherogenic and pro-inflammatory genes. Furthermore, the activation of PPARs has also been studied, by luciferase assay as reporter gene. GQ-177 induced activation of PPARγ, whereas LYSO-7 stimulated activation of PPARα and PPARß/δ. Altogether, our data suggest that the new TZDs derivatives, specially GQ- 177 and LYSO-7, may have atheroprotective properties associated with the reverse cholesterol transport and anti-inflammatory effects, and could be a promising alternative for the treatment of atherosclerosis. However, further studies are warranted in order to characterize the pathways of intracellular signaling since both have demonstrated to activate different isotypes of PPAR