Abstract
BackgroundMany studies have suggested that the immune response may play a crucial role in the progression of hepatocellular carcinoma (HCC). Therefore, our aim was to establish a (i) functional culture of primary human tumor hepatocytes and non-tumor from patients with hepatocellular carcinoma (HCC) and (ii) a co-culture system of HCC and non-HCC hepatocytes with autologous peripheral blood mononuclear cells (PBMCs) in order to study in vitro cell-to-cell interactions.MethodsTumor (HCC) and non-tumor (non-HCC) hepatocytes were isolated from the liver resection specimens of 11 patients operated for HCC, while PBMCs were retrieved immediately prior to surgery. Four biopsies were obtained from patients with no liver disease who had surgery for non malignant tumor (normal hepatocytes). Hepatocytes were either cultured alone (monoculture) or co-cultured with PBMCs. Flow cytometry measurements for MHC class II expression, apoptosis, necrosis and viability (7AAD) were performed 24 h, 48 h and 72 h in co-culture and monocultures.ResultsHCC and non-HCC hepatocytes exhibited increased MHC-II expression at 48h and 72h in co-culture with PBMCs as compared to monoculture, with MHC II-expressing HCC hepatocytes showing increased viability at 72 h. PBMCs showed increased MHC-II expression (activation) in co-culture with HCC as compared to non-HCC hepatocytes at all time points. Moreover, CD8+ T cells had significantly increased apoptosis and necrosis at 48h in co-culture with HCC hepatocytes as compared to monocultures. Interestingly, MHC-II expression on both HCC and non-HCC hepatocytes in co-culture was positively correlated with the respective activated CD8+ T cells.ConclusionsWe have established an in vitro co-culture model to study interactions between autologous PBMCs and primary HCC and non-HCC hepatocytes. This direct interaction leads to increased antigen presenting ability of HCC hepatocytes, activation of PBMCs with a concomitant apoptosis of activated CD8+ T cells. Although, a partially effective immune response against HCC exists, still tumor hepatocytes manage to escape.
Highlights
Many studies have suggested that the immune response may play a crucial role in the progression of hepatocellular carcinoma (HCC)
We examined the correlation between the MHCII expression on HCC and non-HCC hepatocytes and activation of CD8+ T cells and peripheral blood mononuclear cells (PBMCs)
With the respect to the effect of hepatocytes on PBMCs our experiments showed that the expression of MHC II on total PBMCs, which indicates an activation status, was statistically higher in the presence of HCC than non-HCC cells, after 24 h and 48 h of incubation
Summary
Many studies have suggested that the immune response may play a crucial role in the progression of hepatocellular carcinoma (HCC). The effective immune response against hepatocellular carcinoma is of great importance in the progression and clearance of the tumor. Many studies have shown that the immune response against HCC is restrained and not effective [1,2,3]. Cytotoxic T lymphocytes (CTLs) play a central role in induction of anti-tumor immunity. The progression of tumors despite the presence of infiltrating or peripheral cytotoxic CD8+ T cells suggests that immunological tolerance is induced, at least in part, by tumors. Tumor-specific immune responses were observed in a significant proportion of patients with HCC. Spontaneous clearance of established tumors by endogenous immune mechanisms is rare [5]. The pressure of the host immune system may lead to the development of various mechanisms by which the tumor cells escape from immune attack [6]
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