In this research, we aimed to explore the crosstalk target genes, related pathways and possible TFs between IgAN and SS through the integrative bioinformatic analyses of transcriptomes and IHC technique. First of all, we computed and combined the shared genes in the WGCNA modules and the common DEGs of IgAN and SS. The GO and KEGG enrichment analyses indicated that these genes were mainly involved in the biological processes of response to virus and antigen processing and presentation. Afterwards, in the PPI and cytoHubba analyses, we acquired 24 crosstalk genes between IgAN and SS, and then identified 5 hub genes (PSMB8, PSMB9, IFI44, ISG15, and CD53) in the validation datasets. The immune cell infiltration demonstrated that the EM CD8 T and Tfh cells were obviously activated in the IgAN and SS, and the corresponding proportions exhibited positively correlations with the expressions of the 5 hub genes. Additionally, we predicted and verified the STAT1 as the possible TF of the ISG15 and CD53 in the two autoimmune diseases.
As we known, the infections activate the mucosal immune system, which has been recognized to contribute to the onset of IgAN, causing it to produce excess IgA, leading to the deposition [1, 2, 13]. In the renal specimens of IgAN patients, the antigens of Human Cytomegalovirus, Adeno and Herpes simplex virus, Epstein-Barr virus, and even Staphylococcus have been detected, accompanied by the IgA deposits[13, 14]. Likewise, numerous epidemiological and experimental investigations provide evidence of an association between prior viral infection and subsequent development of SS, including Epstein-Barr virus, Cytomegalovirus, hepatitis C virus, etc[7, 15, 16]. The abnormal interference of exogenetic virus or endogenetic viral elements lead to the activation of mucosal epithelial cells and immune systems with the auto-antibody secretion[5]. These auto-antibodies are aggregated into immune complexes that induce the production of interferon (IFN) α, resulting in a cycle of immune-system activation that leads to tissue damage[5]. In our research, the shared genes between IgAN and SS were dramatically enriched in the biological processes of response to virus and the pathways of Epstein-Barr virus, Influenza A, and Human T-cell leukemia virus 1 infection, Staphylococcus aureus infection, and intestinal immune network for IgA production.
The PSMB8 and PSMB9 genes code for low molecular weight protein 7 (LMP7) and LMP2, respectively, involved in the proteasome switch and antigen processing to generate major histocompatibility complex class I binding peptides, and then contribute to the activation of lymphocyte in the IgAN[17]. In a genome-wide association research, rs9357155 at the PSMB8/9 locus was related to an increased risk of IgAN[18]. In the SS, PSMB8 has been reported to be up-regulated and hypo-methylated in the salivary glands[19, 20], and PSMB9 appears to be a good predictor for the classification with a fold change of 2.81 and an adjusted P value of 9.29e − 07[21]. Consistent with these findings, we recognized and verified the PSMB8 and PSMB9 as the hub genes, and the enrichment analyses revealed the key role of antigen processing and presentation in the pathogenesis of IgAN and SS.
Another three hub genes between the IgAN and SS that we screened were CD53, IFI44, and ISG15. The CD53 encodes a tetraspanin, which is known to be expressed by different immune cells, and exerts important and non-redundant roles in the immune cell adhesion and migration, and regulation of immune cell signaling[22]. A bioinformatics study showed that the CD53 could be a meaningful classifier for the SS with a fold change of 3.11 and an adjusted P of 2.52E − 06[21]. Our data also demonstrated that the expressions of CD53 in the glomeruli and tobulointerstitium of IgAN were negatively correlated with the GFR, respectively. It would be of great benefit to explore the pathophysiological functions of CD53 in the antigen-presenting cells of IgAN and SS, and to further extend findings to primary cells and in vivo models. The IFN systems are unusually activated in the peripheral blood and impaired tissue of SS and IgAN patients resulting in the activation of autoimmune response and tissue damage which was mentioned above. IFI44 is a type I IFN signature gene, which may participate in the pathogenesis of autoimmune diseases[23]. As one of the earliest IFN stimulated genes (ISGs), ISG15 is abnormally up- or down-regulated in multiple types of cancer and infectious diseases[24].
In the Immune cell infiltration analyses, we observed that the EM CD8 + T cell and Tfh cell showed significant positive correlations with the five hub genes, respectively. Tfh cell is a CD4 + T cell subset that promotes B cell maturation and differentiation, antibody production, and formation of germinal center in lymphoid follicles, and plays critical roles in the pathogenesis of many autoimmune diseases, such as systemic lupus erythematosus, rheumatoid arthritis, SS, systemic sclerosis etc[25]. Significantly increased proportion of activated cTfh cells and cTfh/follicular regulatory T cell ratio are found in SS patients[25]. Higher frequency of PD-1+ICOS+ Tfh cells is positively correlated with higher level of autoantibodies, ESR, IgG, cytokines and disease activity as well[27]. In the IgAN, recent studies showed that Tfh cells in tertiary lymphoid structure contributed to the renal fibrosis by IL-21 and the renal progression by activating B cells cell-cell interactions[28, 29]. Additionally, several studies indicated that the EM CD8 + T cell subset, in the SS, for example, had high cytolytic activity and were more efficient in migrating to inflamed peripheral tissues during the effector phase of immune response than other EM subsets and took part in pathogens clearance[30, 31]. More valuable studies are needed to uncover the pathophysiological role of EM CD8 + T cells in the various phases of the immune response in the IgAN.
The study has several strengths. We used the systematic bioinformatics analysis as a new approach to explore the relationship between the IgAN and SS. Reasonable internal and external validation, and IHC methods further improved the robustness of the conclusions. The application of immune infiltration analysis made it possible to explore the common immune microenvironment of the two diseases. There are also some limitations in our study. In the selected cohorts, some clinical information was not considered for analysis, such as, age, sex, medication, comorbidities of patients, etc. The influence of these characteristics on the results cannot be ignored. Additionally, further experimental study is needed to confirm our findings in this work, which provides novel insights for further studies on molecular biological mechanisms of IgAN and SS.
In conclusion, our work depicted the common molecule, pathway and immune cell features of the IgAN and SS using the bioinformatics analysis and experimental validation. A better understanding of the pathogenesis of each disease plays a critical role in identifying new targets for early decision making and intervention from the perspective of PPPM.