The existence of OBI makes the management of HBV infection more difficult, and is a serious threat to the transfusion safety [8, 9]. To improve the screening and a better understanding of the pathogenesis of OBI is therefore desirable [1, 4, 6, 29]. Recent studies have identified many lncRNAs associated with HBV infection, and an understanding of their functional roles in the pathogenesis of HBV infectious diseases is just emerging [20, 21]. However, the lncRNA expression in OBI is still unclear.
In the present study, among 15 HBV infection-related lncRNAs, only plasma AP000253 was consistently differently expressed between OBI and HCs. Despite there was a rising trend from CHB to OBI, ASC and HCs, no significant difference in plasma level of AP000253 was found among OBI, ASC and CHB groups. Subsequently, We respectively evaluated the correlation between plasma AP000253 and clinical characteristics in OBI, ASC and CHB groups, and no association with demographic characteristics or serologic viral biomarkers such as antigen, antibody and nucleic acid, was found, which indicated that plasma AP000253 might serve as an independent biomarker of infection regardless of the progression status of HBV infection. Unfortunately, due to the scarcity and limited volume of OBI samples, the associations between AP000253 and overall clinical parameters could not be systematically and comprehensively assessed.
Currently, the gold standard for the diagnosis of OBI was based on the detection of replication-competent viral DNA (i.e. cccDNA and rcDNA), not the presence of integrated viral DNA fragment in the liver [31,32,33]. Regrettably, a standardized assay with internal and external validity is still nonexistent yet. In clinical practice, due to the always unavailable liver tissues, the diagnosis of OBI is commonly based on the detection of HBV DNA in the blood instead. While, unfortunately, because of limited sensitivity of the existing commercial assays, OBI is often missed . As reported, to prevent HBV transmission by transfusion, the sensitivity of nucleic acid assays would need to be lowered from the current 3.4 IU/ml to a new lower limit of detection of 0.15 IU/ml . Hence, to identify novel biomarkers or approaches to improve the OBI screening is urgent. Recently, increasing evidence has demonstrated that lncRNAs had great potential as a diagnostic biomarker for many diseases [34, 35]. Then, the potential of AP000253 as a novel blood-based biomarker in OBI screening was evaluated. Consequently, plasma AP000253 could moderately differentiate OBI from HCs, but fail to discriminate OBI from ASC as well as CHB. When differentiating patients with HBV infection from HCs, similar performance was also observed. These results suggested that AP000253 might be useful in identifying patients with HBV infection, but fail to specifically separate the stage of chronic HBV infection. In the setting of blood donations, in order to reduce the residual risk of transfusion-transmitted HBV infection, the screening strategy adopted by Chinese blood centers is as following: (1) HBsAg rapid screening test in pre-donation, (2) if nonreactive (HBsAg negative), and then two rounds of HBsAg screening with different ELISA assays combined with nucleic acid testing in post-donation [36, 37]. In our blood center, only when HBsAg screening is nonreactive, nucleic acid testing for HBV DNA is available. Hence, in this circumstance, plasma AP000253 would be more useful and practical in discriminating OBI from HCs in HBsAg-negative populations, despite the diagnostic performance is moderate.
Like miRNAs, the lncRNA secretion may be also a selective process, and the peripheral blood level of lncRNAs is not always a true reflection of the intracellular level . Therefore, we further investigated the AP000253 expression in liver tissues with CHB instead, because of the liver tissues with OBI were unavailable. Similarly, in the public GSE81348 dataset, the level of AP000253 was also significantly decreased. However, in HepG2.2.15 and HBV-transfected Huh7 cell lines, the expression level was opposite. One possible explanation for this discrepancy might be the interference of interstitial cells in liver tissue. Regrettably, owing to the limited resources of the liver tissues from CHB patients and healthy people, we failed to further experimentally confirm the AP000253 expression in liver tissues, which deserve further confirmation, especially in liver tissues with OBI.
Finally, accumulating evidence has indicated that lncRNAs can participate in diverse physiological and pathological processes and affect disparate cellular functions . To the best of our knowledge, this is the first report to elaborate AP000253. AP000253 (LOC102724449), a neighbor lncRNA gene of human SOD1 gene, is located at chromosome 21q22.11 with a length of 4282 bp, containing three exons. According to the RNA-Seq expression data from GTEx database, AP000253 shows tissue-specific distributions, and is highly expressed in the liver and testis. Moreover, it has no coding probabilities predicted by the Coding Potential Calculator. The subcellular localization of lncRNAs always determines their possible mechanisms. From the lncLocator database, AP000253 transcript (XR_430363.3) is predominantly located in cytoplasm, suggesting that it may exert biological functions at post-transcriptional level. Due to the cell or animal models of OBI are not available, we then try to preliminarily explore the possible biological functions of AP000253 using HepG2.2.15 and HBV-transfected Huh7 cell lines. ELISA and qPCR assays indicated that AP000253 could significantly enhance the levels of HBsAg, HBeAg and HBV DNA, suggesting that AP000253 might be able to modulate HBV transcription and replication in hepatoma cells. However, the molecular mechanisms of AP000253 in modulating HBV replication need to be further clarified, which is what we are now working on.