Da-Yong Cao

Intestinal Absorption and First-Pass Metabolism of Polyphenol Compounds in Rat and Their Transport Dynamics in Caco-2 Cells

Background Polyphenols, a group of complex naturally occurring compounds, are widely distributed throughout the plant kingdom and are therefore readily consumed by humans. The relationship between their chemical structure and intestinal absorption, transport, and first-pass metabolism remains unresolved, however. Methods Here, we investigated the intestinal absorption and first-pass metabolism of four polyphenol compounds, apigenin, resveratrol, emodin and chrysophanol, using the in vitro Caco-2 cell monolayer model system and in situ intestinal perfusion and in vivo pharmacokinetic studies in rats, so as to better understand the relationship between the chemical structure and biological fate of the dietary polyphenols. Conclusion After oral administration, emodin and chrysophanol exhibited different absorptive and metabolic behaviours compared to apigenin and resveratrol. The differences in their chemical structures presumably resulted in differing affinities for drug-metabolizing enzymes, such as glucuronidase and sulphatase, and transporters, such as MRP2, SGLT1, and P-glycoprotein, which are found in intestinal epithelial cells.

Tolerance Induction by Exosomes from Immature Dendritic Cells and Rapamycin in a Mouse Cardiac Allograft Model

Background Dendritic cells (DCs) release bioactive exosomes that play an important role in immune regulation. Because they express low levels of class I major histocompatibility complex (MHC) and co-stimulatory molecules, exosomes derived from donor immature DCs (imDex) prolong allograft survival by inhibiting T-cell activation. However, this effect is limited and does not induce immunological tolerance when imDex are administered alone. Thus, we tested the effect of combined treatment with donor imDex and low-dose rapamycin on inducing tolerance in a mouse cardiac transplantation model. Methods ImDex were obtained from the culture supernatant of immature DCs derived from donor mouse (C57BL/6) bone marrow and were injected with suboptimal doses of rapamycin into recipient mouse (BALB/c) before and after transplantation. The capacity of this treatment to induce immune tolerance was analyzed in vitro and in vivo using the mouse cardiac transplantation model. Results Donor imDex expressed moderate levels of MHC class II and low levels of MHC class I and co-stimulatory molecules, but neither imDex nor subtherapeutic rapamycin dose alone induced cardiac allograft tolerance. Combined treatment with imDex and rapamycin, however, led to donor specific cardiac allograft tolerance. This effect was accompanied by decreased anti-donor antigen cellular response and an increased percentage of spleen CD4+CD25+ T cells in recipients. Furthermore, this donor specific tolerance could be further transferred to naïve allograft recipients through injection of splenocytes, but not serum, from tolerant recipients. Conclusion Combined with immunosuppressive treatment, donor imDex can prolong cardiac allograft survival and induce donor specific allograft tolerance.

Annexin A2 Promotes the Migration and Invasion of Human Hepatocellular Carcinoma Cells In Vitro by Regulating the Shedding of CD147-Harboring Microvesicles from Tumor Cells

It has been reported that Annexin A2 (ANXA2) is up-regulated in hepatocellular carcinoma (HCC), but the roles of ANXA2 in the migration and invasion of HCC cells have not been determined. In this study, we found that ANXA2-specific siRNA (si-ANXA2) significantly inhibited the migration and invasion of HCC cells co-cultured with fibroblasts in vitro. In addition, the production of MMP-2 by fibroblasts cultured in supernatant collected from si-ANXA2-transfected HCC cells was notably down-regulated. ANXA2 was also found to be co-localized and co-immunoprecipitated with CD147. Further investigation revealed that the expression of ANXA2 in HCC cells affected the shedding of CD147-harboring membrane microvesicles, acting as a vehicle for CD147 in tumor-stromal interactions and thereby regulating the production of MMP-2 by fibroblasts. Together, these results suggest that ANXA2 enhances the migration and invasion potential of HCC cells in vitro by regulating the trafficking of CD147-harboring membrane microvesicles.

Altered expression levels of miRNAs in serum as sensitive biomarkers for early diagnosis of traumatic injury

MicroRNAs (miRNAs) are small non‐coding RNAs of approximately 22 nucleotides in length which regulate gene expression negatively and play important roles in many pathological processes. It has been demonstrated that circulating miRNAs hold promise to serve as practicable molecular markers for diverse physiological and pathological conditions. In this investigation, we chose partial hepatectomy (PH) as traumatic injury model. There were significantly differential expression of miRNAs in rat serum post‐traumatic injury (21 miRNAs were more than twofold up‐regulated). Especially, the expression of miR‐9 showed the highest up‐regulated (>70‐fold), and it possessed the characteristics of biomarker that was more sensitive than aspartate aminotransferase and alanine aminotransferase and C‐reactive protein for traumatic liver injury. There was also a prominent increase in the expression levels of miR‐9 in different brain areas after traumatic injury. Our data suggest that serum miR‐9 may serve as promising biomarker for traumatic injury with high sensitivity. Furthermore, these findings may help to elucidate the complex network which mediates stress response to traumatic injury. J. Cell. Biochem. 112: 2435–2442, 2011.

Increased expression of a disintegrin and metalloprotease-9 in hepatocellular carcinoma: implications for tumor progression and prognosis.

OBJECTIVE A disintegrin and metalloprotease-9 has been involved in the carcinogenesis of various solid tumors. However, its role in hepatocellular carcinoma remains unknown. The aim of this study was to investigate the clinicopathological and prognostic relevance of a disintegrin and metalloprotease-9 by immunohistochemistry. METHODS The expression profile of a disintegrin and metalloprotease-9 in association with the clinicopathological factors was determined by immunohistochemical analysis in hepatocellular carcinoma patients, and its potential prognostic value was investigated by comparing the survival rate between a disintegrin and metalloprotease-9-positive and a disintegrin and metalloprotease-9-negative patients. RESULTS Hepatocellular carcinoma tissues with positive a disintegrin and metalloprotease-9 expression were larger and less differentiated than those with negative expression (P = 0.02 and 0.008, respectively). Portal venous invasion, hepatic venous invasion, bile duct invasion and intrahepatic metastasis were detected significantly more frequently in a disintegrin and metalloprotease-9-positive group (P = 0.009, 0.01, 0.03 and 0.02, respectively). In addition, high alpha-fetoprotein levels were significantly associated with the expression of a disintegrin and metalloprotease-9 in hepatocellular carcinoma (P = 0.01). Moreover, a disintegrin and metalloprotease-9-positive group had significantly poorer outcomes than a disintegrin and metalloprotease-9-negative group (P = 0.01) and was an independent prognostic factor for overall survival. CONCLUSIONS A disintegrin and metalloprotease-9 is over-expressed in hepatocellular carcinoma tissues, consistent with findings in other tumor entities, and is an independent prognostic marker of overall survival following hepatectomy. Further studies are needed to investigate the precise function of a disintegrin and metalloprotease-9 in the progression of hepatocellular carcinoma.

Comparative analysis of DC fused with allogeneic hepatocellular carcinoma cell line HepG2 and autologous tumor cells as potential cancer vaccines against hepatocellular carcinoma

Fusions of patient-derived dendritic cells (DCs) and autologous tumor cells induce T-cell responses against autologous tumors in animal models and human clinical trials. These fusion cells require patient-derived tumor cells, which are not, however, always available. Here we fused autologous DCs from patients with hepatocellular carcinoma (HCC) to an allogeneic HCC cell line (HepG2). These fusion cells co-expressed tumor-associated antigens (TAAs) and DC-derived costimulatory and MHC molecules. Both CD4(+) and CD8(+) T cells were activated by the fusion cells. Cytotoxic T lymphocytes (CTLs) induced by the fusion cells were able to kill autologous HCC by HLA-A2- and/or HLA-A24-restricted mechanisms. CTL activity against shared TAAs indicates that the presence of alloantigens does not prevent the development of CTLs with activity against autologous HCC cells. These fusion cells may have applications in anti-tumor immunotherapy through cross-priming against shared tumor antigens and may provide a platform for adoptive immunotherapy.

Dendritic cells fused with allogeneic hepatocellular carcinoma cell line compared with fused autologous tumor cells as hepatocellular carcinoma vaccines

Purpose:  To investigate the specific antitumor responses against autologous hepatocellular carcinoma (HCC) cells of dendritic cells (DCs) fused with allogeneic HCC cell line, and evaluated the feasibility of BEL7402 as an alternative strategy to deliver shared HCC antigens to DCs.

Improvement of dendritic-based vaccine efficacy against hepatitis B virus-related hepatocellular carcinoma by two tumor-associated antigen gene-infected dendritic cells.

Recently, studies on dendritic cell (DC) vaccine have focused on the development of more effective DC vaccine regimen, such as the application of multiple tumor-associated antigen-targeted DC vaccine. This approach could be used to enhance efficacy of DC-based vaccine against tumors and infectious diseases. In this study, we analyzed whether DC from patients with hepatocellular carcinoma can be infected with the alpha-fetoprotein (AFP) gene and/or HBsAg gene (hepatocellular carcinoma-related antigen). Further, it was examined whether vaccination using these genetically engineered DC can induce stronger therapeutic antitumor immunity. Results revealed that DC infected with AdAFP (adenovirus AFP)/HBsAg can express AFP and HBsAg by reverse transcription-polymerase chain reaction and Western blot techniques. Compared with those before transfection, the expressions of membrane molecules increased dramatically. Specific T cells generated by DCs infected with AdAFP/HBsAg specifically recognized human leukocyte antigen-matched HepG2.2.15 cell lines. Moreover, the cytotoxic activity of cytotoxic T lymphocytes against HepG2.2.15 with DCs expressing AFP was significantly augmented by coinfection with the HBsAg gene. Administration with such vaccine also significantly increased the production of interleukin-12p70 and interferon-gamma. Most importantly, in vivo results suggested that inhibitors of tumor growth were most significant in severe combined immunodeficiency mice model, which was treated with induced cytotoxic T lymphocyte by the AFP/HBsAg-DC vaccine. These results indicate that a vaccination therapy using DCs coinfected with the two tumor-associated antigen genes is an effective strategy for immunotherapy in the activation of DCs, CD4(+) T cells, and CD8(+) T cells, and may be useful in the clinical application of cancer vaccine therapy.

Antigen-specific T cell response from dendritic cell vaccination using side population cell-associated antigens targets hepatocellular carcinoma

Dendritic cell (DC) vaccination targeting cancer stem cells is an effective way to suppress tumor progression and reduce the metastasis and recurrence. In the present study, we explored the suitability of side population (SP) cells as source of antigens for DC vaccination against hepatocellular carcinoma (HCC) in a mouse model. In this study, we identified the “stem-like” characteristics of SP cells in the MHCC97 and Hepa 1-6 HCC cell lines. We found that SP cells express high levels of tumor-associated antigens and MHC class I molecules. Although loading with cell lysates did not change the characteristics of DCs, SP cell lysate-pulsed DCs induced antigen-specific T cell responses, including T cell proliferation and increased IFN-γ production by stimulated CD8+ T cells. We investigated the cytotoxicity of T cells stimulated by SP cell lysate-pulsed DCs in nude mice co-injected with MHCC97 cells. To mimic the in vivo environment, we also confirmed the result in mouse HCC cell line Hepa 1-6 induced tumor-bearing C57/BL6 immune competent mice, and we demonstrated that vaccination with DCs loaded with Hepa 1-6 SP cell lysates could induce a T cell response in vivo and suppress the tumor growth. Our results may have applications for anti-HCC immunotherapy by targeting the cancer stem cells and may provide new insight for cancer vaccines.