Acção anti-cancerígena da Quercetina no Carcinoma Hepatocelular: o papel do GLUT-1

Hepatocellular Carcinoma (HCC) is one of the most fatal cancers, with rising incidence. Without specific treatment, the prognosis is very poor and diminished survival. The most effective therapy is liver transplantation and complete surgical resection, however, since only 15% of patients are candidates for such therapies, a wide range of patients are subjected to treatment with conventional therapies, and the rate success is greatly diminished. It is thought that the expression of glucose transporter 1 (GLUT-1) may be altered in HCC. A recent study showed that suppression of GLUT-1 expression, using siRNA (small interfering RNA) could significantly reduce tumorigenesis in HCC cell lines, suggesting that GLUT-1 may be a therapeutic target for this highly aggressive tumor. Thus, this project aims to evaluate the anticancer effect of quercetin, a possible inhibitor of GLUT-1, in a human HCC cell line HepG2, as well as check the effect of theis compound on 18F-FDG (a glucose radiolabelled analogue) uptake in this cell line. These results shown that quercetin have anti-proliferative effect on HCC cell line studied. This compound also have shown ability to decrease the 18F-FDG uptake. However, using flow cytometry it was found that HepG2 cells remain viable after treatment with quercetin, and this compound doesn’t inhibit the GLUT-1 protein expression. These results indicate that quercetin inhibits the GLUT-1 function, but doesn’t inhibit the expression of this transporter.Keywords: Quercetin, Hepatocellular Carcinoma, GLUT-1 O Carcinoma Hepatocelular (CHC) é um dos cancros mais letais, com uma crescente incidência em diversas regiões por todo o mundo. Sem tratamento específico, o prognóstico é muito pobre e a sobrevida diminuta. A terapia mais eficaz consiste no transplante hepático e na ressecção cirúrgica, no entanto, e uma vez que apenas 15% dos doentes são candidatos a tratamento cirúrgico, torna-se urgente a procura de novas opções terapêuticas para este tipo de tumor. Alguns estudos demonstraram que a expressão do transportador de glucose-1 (GLUT-1) pode estar alterada neste tipo de tumor. Um estudo recente demonstrou que a supressão da expressão de GLUT-1, recorrendo a siRNA (small interfering RNA) conseguiu reduzir significativamente a tumorigénese em culturas celulares de CHC, sugerindo que o GLUT-1 pode ser um alvo terapêutico para este tipo de tumor altamente agressivo. Assim, o objectivo deste trabalho experimental foi avaliar o efeito anti-cancerígeno da quercetina, um possível inibidor do GLUT-1, numa linha celular humana de CHC (HepG2, ATCC), assim como avaliar o seu efeito na captação de 18F-FDG, um análogo da glucose radiomarcado com Flúor-18. Com os resultados obtidos verificou-se que a quercetina possui a capacidade de inibir a proliferação da linha celular em estudo e, para além disso, parece ter influência na captação de 18F-FDG já que conseguiu diminuir a percentagem de captação do radiofármaco nesta linha celular. No entanto, através da técnica de citometria de fluxo verificou-se que as células permanecem viáveis, e que este composto não inibe a expressão proteica do GLUT-1. Estes resultados indicam que a quercetina inibe este transportador de glucose quanto à função, mas não quanto à expressão. Palavras-chave: Quercetina, Carcinoma Hepatocelular, GLUT-1

Cinética da regeneração hepática e sua correlação com a função hepática após hepatectomia

Resumo da comunicação apresentado ao XII Congresso Nacional de Medicina Nuclear, 12-14 Novembro 2009, Mealhad

Estudo da cinética da regeneração hepática no homem pós-hepatectomia por métodos radioisotopicos

Resumo da comunicação apresentado ao XII Congresso Nacional de Medicina Nuclear, 12-14 Novembro 2009, Mealhad

Bariatric Surgery Induces Alterations in the Immune Profile of Peripheral Blood T Cells

Low-grade inflammation is closely linked to obesity and obesity-related comorbidities; therefore, immune cells have become an important topic in obesity research. Here, we performed a deep phenotypic characterization of circulating T cells in people with obesity, using flow cytometry. Forty-one individuals with obesity (OB) and clinical criteria for bariatric surgery were enrolled in this study. We identified and quantified 44 different circulating T cell subsets and assessed their activation status and the expression of immune-checkpoint molecules, immediately before (T1) and 7–18 months after (T2) the bariatric surgery. Twelve age- and sex-matched healthy individuals (nOB) were also recruited. The OB participants showed higher leukocyte counts and a higher percentage of neutrophils. The percentage of circulating Th1 cells were negatively correlated to HbA1c and insulin levels. OB Th1 cells displayed a higher activation status and lower PD-1 expression. The percentage of Th17 and Th1/17 cells were increased in OB, whereas the CD4+ Tregs’ percentage was decreased. Interestingly, a higher proportion of OB CD4+ Tregs were polarized toward Th1- and Th1/17-like cells and expressed higher levels of CCR5. Bariatric surgery induced the recovery of CD4+ Treg cell levels and the expansion and activation of Tfh and B cells. Our results show alterations in the distribution and phenotype of circulating T cells from OB people, including activation markers and immune-checkpoint proteins, demonstrating that different metabolic profiles are associated to distinct immune profiles, and both are modulated by bariatric surgery

Estudo do efeito da clampagem peroperatória selectivo da veia porta na função hepatocelular no modelo animal com fígado normal

Resumo do poster apresentado ao XII Congresso Nacional de Medicina Nuclear, 12-14 Novembro 2009, Mealhad

Elevated soluble TNFα levels and upregulated TNFα mRNA expression in purified peripheral blood monocyte subsets associated with high-grade hepatocellular carcinoma

Chronic inflammation is involved in the initiation and progression of various cancers, including liver cancer. The current study focuses on the characterization of the peripheral immune response in hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA) patients, before and after surgical procedure, in order to assess the effect of tumor resection in the immune system homeostasis and to determine possible prognostic factors associated with high-grade tumors. We developed a whole-blood assay to monitor immune alterations and functional competence of peripheral monocytes in a group of 10 healthy individuals (HG), in 20 HCC patients and 8 CCA patients, by multi-color flow cytometry, qRT-PCR, and ELISA techniques. Results: The qRT-PCR analysis showed an upregulation of TNFα expression by classical and intermediate monocytes purified from HCC patients presenting tumors in grade G3-G4 as compared to G1-G2 HCC patients. Moreover, ELISA assay confirmed elevated serum levels of TNFα in G3-G4 compared to G1-G2 HCC patients. A significant decrease of circulating non-classical monocytes was detected in both CCA and HCC patients before and after surgical procedure. In addition, a functional defect in circulating classical and intermediate monocytes was observed in both groups of cancer patients when compared to the HG, with partial recovery after the surgical intervention. Conclusions: This integrated analysis permitted the identification of altered functional competence of monocyte subsets in CCA and HCC patients. In addition, our results point to a potential role of TNFα as a prognostic peripheral biomarker in HCC patients, indicating the presence of high-grade tumors that should be further validated

Functional and Phenotypic Characterization of Tumor-Infiltrating Leukocyte Subsets and Their Contribution to the Pathogenesis of Hepatocellular Carcinoma and Cholangiocarcinoma

Hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA) represent the most common primary liver malignancies whose outcome is influenced by the immune response. In the present study, we evaluated the tumor-infiltrating leukocyte (TIL) populations in 21 HCC patients and 8 CCA patients by flow cytometry immediately after the surgical procedure. Moreover, CD4+ T cells, CD8+ T cells, monocytes, and macrophages were purified by cell sorting for further analysis of gene expression by quantitative reverse-transcription polymerase chain reaction. Regarding tumor-infiltrating macrophages, we observed a significantly higher expression of markers associated with M2 phenotype and a higher expression of PD-L1 in patients with HCC in comparison to CCA. In addition, for HCC patients, we found a significant increase in the expression of CD200R in macrophages from tumors that were in grade G3-G4 as compared to tumors in grade G1-G2. Besides, a significantly higher frequency of tumor-infiltrating lymphocytes, CD8+CD56+ T cells, and natural killer cells was detected in HCC biopsies in comparison to CCA. In summary, this study has revealed functional and phenotypic differences in TIL cell subpopulations between CCA and HCC, as well as among different histopathological grades and tumor aggressiveness degrees, and it has provided evidence to better understand the tumor immune microenvironment of CCA and HCC