Compared with other organ transplantations, VCA has a higher acute rejection rate, but most patients can achieve long-term survival through timely treatment [12]. However, VCA mild rejection can damage many normal tissues, leading to dysfunction and even graft loss. Therefore, early diagnosis and timely treatment are very important for acute rejection. At present, rejection is mainly evaluated by subjective examination and blood indexes. These methods are not enough to directly reflect the current graft state and can not accurately predict the future development trend.
Moreover, miRNAs are non-coding RNAs that can be used as biomarkers of immune responses, in vivo index regulation, and tumor development [13, 14]. Also, it has been reported that there is a difference in the expression of plasma miRNAs in acute rejection induced by VCA in rats [15], and the TaqMan miRNA reverse transcription kit has been used to detect the expression of known miRNAs in the skin and other tissues [11]. However, the miRNAs detected by these methods are very limited, and whether other miRNAs are important has not been fully clarified. Therefore, we used miRNA-seq to detect miRNAs that are important during VCA acute immune rejection.
The expression of special miRNA in VCA immune rejection was previously summarized in a review [16]. The roles of rno-mir-142-5p and rno-mir-146a-5p were further confirmed, consistent with the results described in our current research. In addition to these miRNAs, we also found that many new miRNAs are important during acute rejection, such as rno-miR-150-5p, rno-miR-16-5p, rno-miR-21-5p, rno-miR-3068-3p, rno-miR-340-5p, rno-miR-425-5p, rno-miR-451-5p, rno-let-7b-5p, rno-miR-100-5p, rno-miR-125a-5p, rno-miR-125b-2-3p, rno-miR-125b-5p, rno-miR-127-3p, rno-miR-145-3p, rno-miR-145-5p, rno-miR-152-3p, rno-miR-195-5p, rno-miR-196a-5p, rno-miR-196b-5p, and rno-miR-379-5p. We screened four miRNA according to their differential expression and predicted the number of target genes. The RT-qPCR results verified that rno-mir-340-5p and rno-mir-21-5p were upregulated, and rno-mir-145-5p and rno-mir-195-5p were downregulated. Previously, it was reported that rno-mir-340-5p was significantly upregulated in the plasma of a mouse allogeneic transplantation model [17]. Besides, rno-mir-145-5p can help in the diagnose of renal transplantation rejection [18]. The results of our allograft rat models further confirmed their performance in animal models, which might lead to reliable biomarkers for human organ allotransplantation detection.
Also, rno-mir-340-5p and rno-mir-21-5p have many target genes enriched in the PI3K-Akt signaling pathway, which is closely related to corneal transplantation rejection. Additionally, rno-mir-340-5p plays a role in the progression of osteosarcoma through the PI3K Akt pathway [19]. However, the mechanism of rno-mir-340-5p/PI3K-Akt is rarely reported. Whether rno-mir-340-5p and its most abundant target gene regulate VCA acute immune rejection via the PI3K-Akt signaling pathway needs to be further explored. Significantly downregulated rno-mir-145-5p and rno-mir-195-5p were enriched in the Hippo signaling pathway. However, little is known about the role of the Hippo signaling pathway in immune rejection. Loss of Hippo tumor suppressor activity and hyperactivation of Yap are commonly observed in cancers [20]. Tumor and liver-related Hippo's research has been very in-depth. In the future, we can focus on immunity mechanisms.
This study also has some limitations. First, we detected the expression of only four miRNAs. More miRNAs should be detected and the combination of different miRNAs should be analyzed to improve the specificity and sensitivity of rejection detection. Second, only rat miRNAs were used in this study and, although rats share many genes with humans, they are not the same. Hence, it would be better to have clinical specimens to detect human miRNAs to further assist clinical diagnosis and treatment.