The Roles of T cells in Bladder Pathologies.

The bladder is frequently afflicted by various forms of chronic inflammatory syndromes. Most of these conditions, such as chronic urethral syndrome, overactive bladder, vulvodynia , endometriosis, etc., have poor and overlapping clinical definitions, probably because their underlying causes are diverse and unclear but might potentially involve common inflammatory and non-inflammatory processes IV ( 54 ). Perhaps, the best studied is interstitial cystitis (IC) or bladder pain syndrome ( 9 , 55 , 67 ). IC is a chronic bladder inflammatory disorder that mainly afflicts subjects aged 50 years and above. It is estimated that 10.6 cases per 100,000 subjects are afflicted by this condition ( 9 , 55 , 56 ). Typical symptoms include urinary frequency, urgency, pelvic pain, nocturia , among others ( 9 , 55 , 56 ). Initially, a diagnostic criterion for IC is the presence of a distinct inflammatory lesion, Hunner’s ulcer, detected by cystoscopy. Later findings suggested that these “classical” cases might only account for less than 10% of total IC patients ( 9 , 55 , 56 ). Moreover, clinical studies have revealed that in almost all IC cases, significant infiltration of T cells into the bladder is evident ( 57 , 58 ). However, how these T cells contribute to the progress of IC has remained elusive. Studies of IC have been severely impaired by the lack of adequate animal models. In recent years, various models of IC have been described ( 59 – 63 ), and they are generally classified into three types based on how they are created. Since IC patients are typically associated with disruption of the bladder epithelial barrier involving thinning of the epithelium or development of lesions such as Hunner’s ulcer, etc., the first model type involves employing small molecules (e.g. protamine sulfate, potassium chloride, etc.) or peptides that directly damage the bladder epithelium in mice ( 61 ). For example, a recent study used a small sialoglycopeptide antiproliferative factor (APF), to successfully damage the bladder epithelium in mice by disrupting expression of uroplakin III and ZO-1, which are important structural proteins forming the uroepithelium structure ( 61 ). The second type involves the utilization of WT or genetically deficient animals which will naturally develop IC symptoms ( 60 , 64 ). For example, a proportion of WT domestic cats naturally develop IC, in which the bladder epithelium becomes damaged and chronic inflammation arises ( 64 ). Another group recently reported that Fcgr2b −/− Pdcd1 −/− BALB/c mice, which are deficient in type IIb Fc receptor for IgG and PD-1, spontaneously develop IC-like symptoms before being 10-weeks-old, in contrast to WT BALB/c mice ( 60 ). Moreover, these mutant mice exhibited bladder associated pathologies consistent with IC, including disruption of bladder epithelium, generation of anti-urothelial autoantibodies, and massive infiltration of immune cells, especially CD4 + T cells ( 60 ). The third type of IC model involves administration of autoimmune peptides such as peptides derived from uroplakin ( 65 ) and the immunogenic peptide T2 ( 62 ). Alternatively, it could involve genetic expression of ovalbumin (OVA) in BECs followed by adoptive transfer of ovalbumin (OVA)- specific OT-II CD4 + T cells in mice ( 59 , 66 ). All of these interventions have resulted in strong T cell infiltration into the bladder epithelium accompanied by chronic bladder inflammation as evidenced by histology. These models have successfully recapitulated several aspects of autoinflammatory responses as seen in IC patients, namely, T cell infiltration and bladder epithelial barrier disruption. The development of these various models has revealed that T cells are essential for the development of bladder tissue damage and IC-like symptoms ( Figure 3 ). Specifically, in nearly all of these models, several T cell subsets, including CD4 + , CD8 + , and γδ T cells, appear to infiltrate the bladder and contribute to tissue damage, as shown by histology or flow cytometry analysis, similarly to what has been observed in IC patients ( 58 , 67 ) ( Key Figure, Figure 4 ). Studies using these models have provided clues as to how T cells may be recruited into the bladder and how they may contribute to IC progress. For example, damage to BECs appears to be a key triggering event, resulting in the release of chemokines such as CXCL10 by the damaged cells – a finding also seen in urine samples from IC patients ( 68 , 69 ). T cells typically expressing the cognate chemokine receptor CXCR3 are then recruited into bladder ( 70 ). T cells which recognize self-antigens such as Uroplakin II, Uroplakin IIIA generated from damaged BECs can selectively aggregate in the epithelium as shown in histologic and gene expression analysis of Uroplakin II- or Uroplakin IIIA-treated vs control protein-treated mice ( 65 , 85 ). Recognition of self-antigens caused CD8 + T cells and Th1 cells to secrete cytotoxic molecules, such as IFNγ and TNFα, which can further damage the bladder tissue by amplifying the chronic inflammation ( 65 , 66 , 71 ). A phase III interventional clinical study of 14 IC patients treated with suplatast tosilate (IPD-1151T, a new immunoregulator that selectively suppresses CD4 + T cells) (JapicCTI-060257) III , in which the primary outcome measure was improvement of the IC symptom score and incidence of adverse drug reactions, also suggested that Th2 derived IL-4 might significantly contribute to IC, although the mechanisms remain unclear ( 72 ). T cell secreted IFNγ can stimulate epithelial cells to secrete CXCL10, which in turn recruits more Th1 cells ( 70 ); T cell secreted TNFα may stimulate mast cells via the receptor for TNFR1 but not TNFR2, which can in turn induce mast cells to produce massive amounts of TNFα, further enhancing T cell activity in murine IC models, such as the model utilizing OVA expression in BEC ( 71 , 73 ). With these positive feedback loops, more T cells and other immune cells including mast cells can accumulate in the bladder and cause chronic inflammatory symptoms resulting in IC in murine models ( 65 , 66 , 71 ). Many of these observations such as robust T cell infiltration, mast cell activation, and TNFα secretion, are also supported by a wealth of data from IC patients ( 58 , 67 , 68 , 69 , 72 , 74 ). Recognizing the pivotal roles of T cells, researchers are actively exploring new therapeutic strategies for IC by applying T cell inhibitory molecules. One group tested RDP58, a novel D-amino acid decapeptide effective in treating human ulcerative colitis and chemotherapy-induced diarrhea, in the BEC expressing OVA murine IC model. Researchers found that RDP58 treatment could significantly reduce T cell numbers in the bladder, decrease the production of TNFα, and limit bladder tissue damage in vivo as evidenced from histology analysis ( 75 ). In vitro RDP58 treatment of cultured OVA antigen-stimulated T cells has also shown that this agent can inhibit IFNγ and TNFα secretion from T cells ( 75 ). Another group treated IC patients with IPD-1151T (JapicCTI-060257) III , found that IC symptoms such as urinary urgency, frequency, and lower abdominal pain were significantly mitigated after treatment using 2 sample Wilcoxon and student t statistical tests ( 72 ). The above research suggests that T cells play various roles in IC, but many questions remain unanswered. First, if disease-causing T cells only infiltrate the bladder when self-antigens and chemokines are released by damaged BECs, what is the initial cause of BEC damage? Several possibilities have been proposed ( 9 , 55 ), including routine renewal of BEC, or the presence of environmental pollutants, as well as mediators released from nerves such as substance P. These relatively minor stimuli might result in the release of small amounts of self-antigen and damage signals, which can be captured and greatly amplified by T cells ( 9 , 55 ). We speculate that another possibility to help explain the seemingly pathologic roles of T cells in IC may be related to the interactions of T cells with different immune cells. But if so, what are these immune cells? As mentioned above, T cells can communicate with mast cells through TNFα signaling ( 71 , 73 ). But whether this results in pathogenic outcomes remains to be more robustly assessed. Finally, because IC describes a heterogeneous collection of symptoms, more in-depth studies in patients and animal models are required to clarify whether distinct subtypes of IC exist, and the roles that T cells play in these conditions.

The prevalence of bladder diseases such as bacterial infection, bladder cancer and chronic inflammation such as IC continues to grow as the population ages. Recent studies clearly show the importance of T cells in the progression of each of these diseases. During bacterial bladder infections, CD8 + T cells, γδ T cells, many Th2 cells and a smaller number of Th1 cells are recruited into the bladder to mediate immune responses ( Key Figure, Figure 4 ). Overall, this response appears to be overwhelmingly biased towards tissue repair, and consequently, bacterial clearance (e.g. of UPEC) is suboptimal, which can promote recurrence of infection. Modulation of these T cell responses towards a more balanced one in UTI prone patients by vaccination or immunotherapy might potentially be an approach to prevent infection recurrence. In bladder cancer, both tumor-fighting CD8 + T cells, cytotoxic CD4 + T cells and Th1 cells, as well as immunosuppressive Tregs infiltrate tumors simultaneously in significant numbers ( Key Figure, Figure 4 ) ( 35 ). For the most part, the immunosuppressive activities of Treg and immunosuppressive signaling emanating from the PD-L1/PD-1 pathway appear to dominate within bladder tumors so that even popular immunotherapies such as anti-PD-L1 antibody treatment may have limited efficacy in most patients ( 35 , 43 , 44 , 49 ). Bladder cancer patients display a wide range of responsiveness to immunotherapy, therefore, understanding the underlying basis for this heterogeneity in responsiveness could uncover new avenues for study. In IC, as a result of damage incurred to the bladder epithelial barrier, large numbers of self-antigen recognizing T cells including CD8 + , Th1, Th2, and γδ T cells ( Key Figure, Figure 4 ) can be recruited into the bladder epithelium by, as yet, unknown mechanisms ( 60 – 74 ). These T cells can further amplify inflammation and increase tissue damage ( 60 – 74 ). Several preliminary studies suggest that modulating T cell activities might be effective in treating IC but remain to be rigorously tested. A major limitation with studies relating to IC is that its symptoms overlap with several other chronic bladder inflammatory syndromes, which makes elucidation of its etiology a challenge (see Outstanding Questions ). Recently, the existence of microbiota in the bladder of healthy humans was reported ( Box 2 ) ( 76 – 80 ). Considering that T cell responses are highly regulated by microbiota in both humans and in mouse models, it would be interesting to explore the interactions between the bladder microbiota and local bladder T cells with the goal of exploring their therapeutic potential ( Box 2 ). In summary, various T cell subsets can contribute significantly, albeit in different ways, to bladder diseases. However, further in-depth studies on the nature of these activities are needed and are eagerly awaited to envision the development of candidate efficacious immunotherapies that might target specific bladder pathologies