Exosomal microRNA in autoimmunity

Exosomes are small extracellular vesicles that contain a potpourri of nucleic acids, including microRNAs (miRNAs). Importantly, exosomes and these miRNAs are actively secreted from multiple cell populations and play a critical role in intercellular communication. The exosomal miRNA profile reflects the cellular pathophysiological status and modulates multiple biological processes. Accumulating evidence indicates that exosomes and miRNAs are biomarkers for disease diagnosis and may have therapeutic targets. Research studies on exosomal miRNAs have brought new insights into understanding autoimmune diseases. This article summarizes the detection methodologies and updates the potential applications of exosomal miRNAs in autoimmune diseases.

Exosomes released from immune and nonimmune cells play critical roles in the immune response and the pathogenesis of diseases because of the biologic potential of exosomal proteins and contents, including miRNA (Fig. 1a). This potential is exemplified by a recent study on primary biliary cholangitis (PBC).¹ Exosomes can transfer their cargoes (e.g., miRNAs) to target cells. For example, Ying et al.² demonstrated that adipose tissue macrophages secrete exosomes containing miRNAs. These miRNAs can be transferred to insulin target cells and modulate insulin sensitivity and glucose homeostasis. Similarly, Guay et al.³ reported that exosomal miRNAs released by rodents and human T lymphocytes could be transferred to β cells in an active form, resulting in β-cell apoptosis. Specifically, blocking the activities of these miRNAs in recipient β cells reduced both the inflammation within islets and the development of diabetes in nonobese diabetic mice. These findings indicate that exosomal miRNAs are key mediators of cell-to-cell communication and represent potential diagnostic and prognostic biomarkers and therapeutic targets. In addition, exosomes can possess an antigen presentation function and can transfer major histocompatibility complex (MHC) class I, MHC class II, and antigens. In this sense, exosomes are pluripotent and major players in the regulation of immune processes.

Fig. 1.

The release, uptake, and application of exosomal miRNAs. a Exosomes are secreted by many cells and widely distributed in various body fluids. Exosomes derived from antigen-presenting cells (e.g., B cells and dendritic cells) present antigens by transferring major histocompatibility complexes and antigens. Exosomes carry a complex cargo of miRNAs, mRNAs, and proteins and transmit these important signal molecules into target cells, facilitating cell-to-cell transmission. b The key steps for the application of exosomal miRNAs involve isolation of exosomes and miRNAs, detection of miRNAs, and selection of target miRNAs. Exosomes can be isolated from a variety of biological fluids (e.g., plasma, urine, and saliva). There are different methods to detect miRNA expression based on sample vigor and cost

Exosomal miRNA detection involves exosome isolation, miRNA isolation, and miRNA detection (Fig. 1b). However, the results of miRNA expression are impacted by factors including sample vigor, storage, isolation, and detection methods. The lack of standardized detection methods remains a challenge. Due to the characteristics of having a short sequence, low content, and high sequence similarity, it is difficult to accurately detect miRNAs. Currently, conventional and widely used miRNA detection methods include northern blotting, quantitative reverse transcription polymerase chain reaction, microarray, and next generation sequencing. However, all these methods have relative strengths and weaknesses (Supplementary Table 1). Based upon past deficiencies, new strategies, including nanomaterial-based, electrochemical-based, optical-based, molecular biology-based and imaging analysis, have been proposed to improve the sensitivity and specificity of miRNA detection. These methods attempt to detect miRNA both in broad concentration ranges and in low amounts. Recently, detection methods have reached the attomolar level, representing a high degree of sensitivity for detecting miRNAs. Tavallaie et al. detected miRNA concentrations in the range of 10 aM–1 nM using electric-field-induced reconfiguration of a network of gold-coated magnetic nanoparticles modified by probe DNA.⁴ Xue et al. developed an antimonene-based surface plasmon resonance biosensor that also enabled the detection of miRNA at 10 aM⁵. A surface-enhanced Raman scattering-based sensing platform proposed by Lee et al. demonstrated a low detection limit and wide dynamic range (1 aM–100 nM) without an amplification process.⁶ However, a great need remains for more efficient and accurate methods to detect and quantify exosomal miRNAs.

The majority of miRNAs are detectable in human serum, saliva, and urine and are highly enriched in exosomes.⁷ Previous studies demonstrated a relationship between miRNAs and tissue pathology. These characteristics make miRNAs promising biomarkers for diagnosing diseases and predicting responses to drug therapy, including monitoring the progression of a disease.

Autoimmune disease-associated miRNA expression profiles have been studied in many biological materials, including blood plasma, serum, urine, and synovial fluid. As diagnostic tools, hundreds of differential miRNAs have been identified in patients with an autoimmune disease compared with healthy controls. Our group identified 18 differentially expressed miRNAs in serum exosomes between PBC patients and healthy controls.¹ More than 200 differentially expressed miRNAs have been identified in the sera and PBMC of PBC patients. Some of these miRNAs (the panel of hsa-miR-122-5p, hsa-miR-141-3p, and hsa-miR-26b-5p) reflect high diagnostic accuracy. In addition, our previous study found 35 independent miRNAs differentially expressed in liver tissues of PBC patients. Among these miRNAs, miRNA-506 may be an important and potentially useful miRNA in patients with PBC. The upregulation of miRNA-506 can lead to cholestasis by directly targeting the Cl⁻/HCO₃⁻ anion exchanger 2 and type III inositol 1,4,5-trisphosphate receptor. Recently, Erice et al.⁸ demonstrated that the overexpression of miR-506 in biliary epithelial cells leads to PBC-like features and promotes immune activation.

In addition, miRNAs, particularly exosomal miRNAs, have been widely studied in systemic lupus erythematosus (SLE). Perez-Hernandez et al.⁷ found that urinary miRNAs are primarily enriched in exosomes; the level of miR-146a increased in patients with active lupus nephritis compared with patients with inactive lupus nephritis. Sole et al.⁹ reported that miR-29c could serve as a biomarker for determining the degree of chronicity with impressive sensitivity and specificity in patients with lupus nephritis. These findings suggest that miRNAs in urine are a potential diagnostic marker for lupus nephritis.

MiRNAs can also act as potential biomarkers for predicting drug efficacy. As we noted above, miRNAs detected in human serum are concentrated in exosomes. The baseline expression of miR-125b in PBMCs may predict early therapeutic responses in patients with rheumatoid arthritis (RA). MiR-125b was elevated in responder patients compared with nonresponder patients.¹⁰ In addition, increased expression of miR-125b has been demonstrated to be a potential biomarker as a favorable response to rituximab not only in patients with RA but also in patients with B-cell lymphoma and chronic lymphocytic leukemia. IFN-β is a standard treatment for multiple sclerosis (MS) but is associated with side effects. De Felice et al. reported that miR-26a-5p was significantly higher in responder patients than nonresponder patients at baseline, 3 months, and 6 months of IFN-β treatment.¹¹ Similarly, Manna et al.¹² demonstrated that the levels of 14 exosome-associated miRNAs, including miR-26a-5p, were significantly higher in IFN-β-treated patients with a good clinical response than those without a response. These results indicate that exosomal miRNAs may be useful markers for predicting IFN-β responders.

Exosomal miRNAs have been indicated as biomarkers for disease activity and prognosis in autoimmune diseases, including MS and Graves’ Disease. Ebrahimkhani et al.¹³ identified differentially expressed exosomal miRNAs in patients with progressive MS and relapsing-remitting MS (RRMS) compared with healthy controls. Furthermore, they identified a group of miRNAs that distinguished relapsing-remitting patients from those with progressive disease and validated the data in an independent study. Recently, Selmaj et al.¹⁴ also confirmed distinct exosomal miRNA expression in RRMS patients and healthy controls. Hsa-miR-122-5p, hsa-miR-196b-5p, hsa-miR-301a-3p, and hsa-miR-532-5p were significantly decreased during relapse in RRMS patients. Hiratsuka et al.¹⁵ reported differential expression of circulating miRNAs in patients with active versus inactive Graves’ disease. Further, exosomes from intractable Graves’ Disease patients stimulated miRNA expression for IL-1β and TNF-α compared with patients in remission or healthy controls.

MiRNAs are also potential targets for therapeutic strategies in the treatment of cancer, cardiac disease, and autoimmune diseases. Typically, miRNA-based therapy would achieve therapeutic goals by replenishing or inhibiting the sequences of target miRNAs. For each specific disease, the key steps of miRNA-based therapy involve identifying the best miRNA targets, determining miRNA chemistry, optimizing delivery, preclinical testing, and conducting clinical trials. Designing optimal delivery vehicles with higher stable, minimal toxicity, and less off target effects is a major challenge. Delivery methods include viral vectors and nonviral vectors, especially nanoparticles, which enable more miRNA-based therapies to be tested in preclinical and clinic trials. Some miRNA mimics and anti-miRNAs have successfully entered Phase 1 and 2 clinical trials. Miravirsen (an anti-miRNA-122 oligonucleotide) is in a Phase 2 trial for the treatment of hepatitis C. Another similar drug, RG-101, has entered Phase 2 clinical trials.

Several studies have reported suppressive miRNAs in autoimmune disease. For example, nostalgic compensating miR-29, miR-146a, and miR-130b demonstrated potential effects in murine models of RA and SLE.^16–19 MiR-29 is significantly downregulated in SSc fibroblasts and skin sections of SSc patients, resulting in fibrosis via upregulating the expression of type I and type III collagen; these results make miR-29 a promising therapeutic target for SSc.¹⁶ MRG-201 (miR-29 mimic) is being developed for the treatment or prevention of fibrosis. A Phase 2 clinical trial of MRG-201 (NCT03601052) is currently underway. MiR-146a is associated with the severity of arthritis. Ammari et al.¹⁷ delivered miR-146a mimics to Ly6Chigh monocytes in an RA mouse model and demonstrated a reduced accumulation of osteoclasts in the synovial pannus and an alleviation of local bone erosion. Restoring the loss of miR-146a also had benefits in lupus-prone mice by decreasing the expression of autoantibodies, the levels of immunoglobulin G (IgG), and the levels of proinflammatory cytokines.¹⁸ Han et al. found that the injection of miR-130b agomir in mice reduced proteinuria and attenuate glomerulonephritis.¹⁹ Antagonizing miRNAs, such as miR-663, miR-21, and miR-155, were also efficacious in several autoimmunity murine models. For example, miR-663 inhibition reduces the size of spleens and lymph nodes, improves renal function, and decreases IgG in MRL/lpr mice. Treatment with anti-miR-21 can significantly ameliorate splenomegaly in lupus mice. A therapeutic success was reported in the miR-155 antagomir-treated model of lupus alveolar hemorrhage.

A number of promising results have provided evidence for using exosomal miRNAs as diagnostic biomarkers and therapeutic options. However, the application of exosomal miRNAs for patients with an autoimmune disease remains in very early stages with much work to be done.

Competing interests

The authors declares no competing interests.

Supplementary information

The online version of this article (10.1038/s41423-019-0319-9) contains supplementary material.