In the present study, we evaluated circulating and intra-hepatic IL34 in HBV related liver diseases. Circulating IL34 of HBVHCC patients was significantly higher than that of CHB, HBVcirrhosis and HCs. The highest AUC was detected from AFP plus serum IL-34, suggesting the combination boosts sensitivity and specificity than AFP alone. Furthermore, circulating IL34 was suppressed with antitumor TACE treatment in HBVHCC, further confirmed the potential role of IL34 during the development of HBV-HCC. Consistent with circulating IL34, it was also detected that upregulated intrahepatic IL34 from HBV-HCC, compared to that of CHB, HBVcirrhosis and HCs. Intrahepatic IL34 was associated with high HBVDNA, HBeAg-, poor tumor differentiation and small tumor size of HBV-HCC patients. Intrahepatic CD68+ TAMs were upregulated in HBVHCC compared to that from CHB and HBVcirrhosis. Intrahepatic CD68+ TAMs were associated with high HBVDNA, high tumor differentiation, small tumor size, abnormal AFP and more tumor number. Our current data suggests that IL-34 probably contributes to the development of HBV-HCC, i.e. promoting the disease progression from CHB, HBV cirrhosis and then HBV-HCC. The current observation is supported by the findings from the Zhou group, showing that IL-34 is a key regulator for the growth of HCC in nude mice, probably is via miR-28-5p mediated activation of TAM [20]. Thus our finding is an extension and validation of the important role of IL-34 during the development of HBV-HCC in human tissues.
In our current study, circulating IL34 from HBVHCC patients was significantly higher than that of CHB, HBVcirrhosis, and HCs, suggesting that IL34 may contribute to tumorigenesis of HCC, enhancing progression from CHB patients to cirrhosis and finally HCC. Furthermore, intra-hepatic IL34 expression was consistent with circulating IL-34, which is in line with previous studies, showing that high IL34 in autoimmune diseases [15, 16]. More specifically, IL34 is overexpressed in the inflamed synovium of rheumatoid arthritis patients, where it perhaps acting synergistic with TNF and IL1β, induces osteoclastogenesis and contributes to tissue inflammation and bone erosion [27]. In addition, upregulated circulating and intra-hepatic IL-34 in HBV-HCC from the current study is supported by the others, showing that the circulating IL34 markedly increased in HBVHCC patients, compared to those in CHB and HBV-negative HCC patients [28], and the HBX gene of HBV upregulates IL-34 expression in hepatoma [28]. However, our previous study demonstrates that IL-34 is inversely correlated with differentiation, metastasis and invasion of gastric cancer [14], which is rather controversial with our current discovery in HBV-HCC disease. Our explanation for such discrepancy between HBV-HCC and gastric cancer is more likely related to the different carcinogenesis between gastric cancer and HBV-HCC, as well as completely different microenvironments, despite of gastric cancer and HBV-HCC are all belonging to gastrointestinal system.
The development of tumor is closely related to the microenvironment, including tumor cells, monocytes/macrophages, cytokines and neovasculisation. TAMs are mixed phenotype, expressing M1 or M2 markers [5], and may be influenced by different microenvironments in different regions and/or in different individuals. In our current study, intrahepatic CD68+ TAMs were increased gradually with the order from CHB, HBVcirrhosis to HBVHCC patients. Our current observation invites speculation that the increased infiltrating CD68+ TAMs may be M2 dominant, contributing to stimulate tumor growth activity [5]. This is in line with Zhou et. al. showing that IL-34 induced TAMs can inhibit miR-28-5p, which is one of the miRNAs that decrease IL-34 production, in HCC cells in vitro via TGFβ1, suggesting a paracrine factor fashion among miR-28-5p, IL-34 and macrophage. In clinical HCC study, lower miR-28-5p is correlated with high IL-34 and TAMs in HCC patients with a poor overall survival and recurrence [20].
IL34 is upregulated in HCV infection and inhibited the production of IFNγ [12], and IL34 may also be associated with inflammatory activity and liver fibrosis in CHB [21]. Moreover, baseline circulating IL34 seems to serve as a prognostic factor for progression in such patients. The high serum IL-34 level is associated with poor prognosis in non-viral HCC patients compared with the patients with low serum IL-34 level [29]. In our current study, there was only circulating IL34 significantly decreased in postantitumor treatment compared to that of pre-treatment of HBVHCC, suggesting IL-34 is closely related the weight of HBV-HCC or partial source of IL-34 is coming from HCC cells. Thus, serum IL34 was significantly correlated with the incidence of HBVHCC. The sensitivity and specificity of AFP plus circulating IL-34 seems to be greatly improved, compared to that of AFP alone, suggesting that AFP plus serum IL-34 could improve the diagnostic accuracy of AFP for detecting HBV-HCC, as well as IL34 could be used as a diagnostic biomarker for HBVHCC, which will be further clarified in vitro and in vivo.
IL34 induces differentiation of leukemia cells into monocytelike, macrophagelike cells and mature macrophages [30], may also enhances differentiation of other cancers [14, 31]. These reports are consistent with our current finding that IL34 was correlated with the differentiation and tumor size of HBVHCC. In addition, intrahepatic IL34 was associated with HBVDNA, HBeAg, tumor differentiation and tumor size of HBVHCC patients. Our data may provide an explanation for the possible role of IL34 in the development of HBVHCC, i.e. IL34 also regulates differentiation of HBVHCC, which would have potential clinical relevance regarding IL34 as a therapeutic target for malignancy. Such speculation is in line with others, showing that IL-34 inhibits HBV replication in vivo and in vitro [32], and IL-34 is beneficially to the HBV-HCC patients for potential therapeutic target.
There are limitations in the current study. There is no kinetics of intrahepatic or circulating IL34 during the development and management of HCC, and no correlation between IL34 and prognosis is detected. These two interesting points will be determined in our future study. We also acknowledge that the sample size in the current study is relatively small, which may not perfectly reflect the real cases. However, the current experiment is just a proof of concept. We will explore the underlying mechanism with large sample size and multiple center studies in future.
In conclusion, the current study improves our understanding of the role of IL34 in HBV related liver disease. Increased IL34 may contribute to the transformation of HBV HCC, which is a potential predictor of HBVHCC. The underlying mechanism of IL34 in HBVHCC is being currently investigated.