Lymphoid organs are classified into primary lymphoid tissues (LT), secondary LT, and tertiary LTs or structures (TLSs) based on localization, function, or manner of development.1 TLSs are observed in various organs and often develop at sites of infection or chronic immune stimulation in autoimmune diseases, atherosclerosis, allograft rejection, cancer, or aging.2,3 Although most inflammatory cell infiltrations in chronic diseases are not spatially organized, TLSs are mainly composed of T cells, B cells, and unique stromal cells, and are structurally and functionally organized. TLSs range from small discrete clusters of lymphocytes to large, complex structures. Large, well-developed TLSs contain germinal centers of proliferating B cells surrounded by T cells, and fibroblasts (Figure 1).3 In peripheral tissue TLSs, homeostatic chemokines such as CXCL 13 not only recruit lymphoid cells but also orchestrate their arrangement into TLSs.4 TLSs can initiate adaptive immune response and are local sites of antigen presentation, clonal expansion, proinflammatory cytokine production, and lymphocyte activation. These functional aspects distinguish TLSs from common inflammatory cell infiltration. The impacts of TLSs on host organs can be beneficial or detrimental depending on the clinical context. In infectious diseases and most types of cancers, TLSs play beneficial roles for the host by generating immune responses to pathogens and cancers. Conversely, in autoimmune diseases, TLSs in affected organs are associated with worse clinical outcomes because of autoantibody production and aggravation of organ inflammation.4
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Figure 1.: The localization and cellular components of TLSs in renal parenchyma and UTALS in renal pelvis. The images are periglomerular and perivascular TLS in aged C57BL6J mice subjected ischemic reperfusion injury, subcapsular TLS in aged human, and pelvis UTALS in NZB/NZW F1 mice, which is a model of lupus nephritis.
In the field of nephrology, the existence of TLSs has been reported in autoimmune nephritis, transplantation, and chronic kidney injury.3,56–7 In lupus nephritis patients, TLSs were strongly associated with the immune complex in the tubular basement membrane, implicating them in the pathogenesis of lupus tubulointerstitial inflammation.5 We previously demonstrated that aged kidneys develop multiple TLSs after AKI.3 We also found that TLSs develop and mature through at least three developmental stages that were associated with the severity of kidney injury in mice and humans.8 In addition, we recently demonstrated that stage 2 TLSs in protocol biopsies of kidney transplants 1 year after transplantation predict worse renal outcomes.6 Furthermore, we showed that the administration of CD4 monoclonal antibodies abolishes TLS formation with attenuated inflammation and fibrosis.3 Although this intervention is not specific to TLSs, these results suggest that TLSs possibly play a detrimental role in injured kidneys. The recent research on TLSs in renal parenchyma indicate that autoimmunity and aging are crucial factors to accelerate TLS formation. Understanding the features of TLSs may lead to new therapeutic strategies for kidney disease.
The renal pelvis (RP) is directly connected to parenchyma via connective tissues composed of stromal cells. Several recent studies reported TLSs in the RP of NZB/NZW mice and humans with pyelonephritis.8,9 However, the roles of TLSs in the RP in noninfectious nephritis are unclear.
In this issue of the JASN, Ichii et al.10 reported urinary tract associated lymphoid structures (UTALSs) in humans and mice with noninfectious chronic nephritis and demonstrated the close association between the altered urothelium barrier and UTALS formation (Figure 1). In histologic analysis, UTALSs were localized in the deep portion of the RP and were composed of T cells, B cells, macrophages, antigen-presenting cells, LYVE1+ lymphatic vessels, and high endothelial venules between developed connective tissues. UTALSs in aging mice and MRL/lpr mice, a model of systemic lupus erythematosus, were larger than those in control young MRL/MpJ mice. These results indicate the involvement of aging and systemic immune factors in UTALS development. Furthermore, Ichii et al. demonstrated that the urothelium barrier of the RP was altered in mice and humans with chronic nephritis by scanning electron microscopy and immunostaining analysis, and bovine serum albumin was significantly leaked into developing UTALS in the RP of MRL/lpr mice. Accordingly, they assessed TNF-α and IFN-γ in the urine as candidate molecules causing urothelium barrier defects and confirmed the effect of these cytokines on urothelium barrier integrity using cultured human urothelial cells. In addition, microarray and quantitative PCR data indicated that Ccl1, Ccl8, Ccl20, Cxcl9 and Cxcl13 were highly expressed with their receptor genes in the RP of MRL/lpr mice, and Cxcl9 and Cxcl13 mRNA was detected in interstitial stromal cells of these mice. Accordingly, they stimulated NIH3T3 fibroblasts with MRL/lpr RP-derived urine, which specifically induced CXCL13 expression in these fibroblasts and increased their size. These results strongly suggest that urine-stimulated fibroblasts play an important role in UTALS formation via chemokine production. They also found upregulation of cytokines/chemokines in a coculture of lymphocytes and stromal cells upon stimulation with the urine. In addition, they found a significant correlation between pathologic parameters regarding glomerular lesions, tubulointerstitial lesions, and TLSs, and Cxcl9 and Cxcl13 expression in the RPs.
Next, they focused on collagen families. Microarray analysis of mouse kidneys revealed that Col17a1 was highly expressed in the RPs of MRL/MPJ and MRL/lpr mice. Characteristically, COL17A1+ staining was ectopically localized in the urothelium covering well-developed UTALS. Moreover, predicted promoter region analysis of mouse Col17a1 using JASPAR revealed the highest scores in the binding sites of FOS and JUN, both of which localized in the nuclei in the urothelium covering UTALS. They also showed nuclear localization of FOS in human urothelial cells after stimulation with urine from nephritis patients. Thus, they proposed that urine-urothelium barrier alterations under nephritic conditions result in subsequent leakage of urine into the RP, contributing to UTALS development.
Ichii et al. revealed the unique mechanisms of UTALS formation, and their findings raise intriguing questions. The mice used in their study were MRL/lpr, which are a model of autoimmune disease and accelerated immune senescence and are known to develop TLSs in renal parenchyma in addition to RPs. Thus, chronic inflammation, autoimmune disease, and aging, known to stimulate TLS formation, might also contribute to UTALS formation in these mice. Similarly, TNF-α and IFN-γ in the blood and kidney parenchyma could also contribute to UTALS formation. In addition, the mechanism whereby increased expression of COL17Aa in the urothelium causes the disruption of barrier function remains unclear.
Although these questions remain, this important paper sheds new light on TLSs in the RP and identifies a possible mechanism for urine-triggered disruption of the urothelium barrier and UTALS formation. Elucidation of this mechanism is helpful for a better understanding of non-infectious chronic nephritis.
Disclosures
M. Yanagita reports receiving research grants from Kyowa Kirin, Mitsubishi Tanabe Pharma, and Boehringer Ingelheim; honoraria from Astellas, Kyowa Kirin, Chugai, and others for lecture honoraria; and has other interests/relationships with the Japanese Society of Nephrology and International Society of Nephrology. The remaining author has nothing to disclose.
Funding
This research was supported by Japan Agency for Medical Research and Development grants AMED-CREST21gm1210009, 21gm5010002, and 21zf0127003h001.
Acknowledgments
The authors thank Dr. Yuki Sato, Dr. Makiko Kondo, Dr. Takahisa Yoshikawa, and Dr. Naoya Toriu for valuable discussion and images and Ms. Azusa Okuwa for figure illustration.
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