Interstitial cystitis/bladder pain syndrome (IC/BPS) is a disabling disease, with a reported prevalence of 52—500/100,000 in the female and 8—41/100,000 in the male population. The etiopathology of IC/BPS is still unclear, impeding causal treatment. Biomarkers could help to better understand the etiopathology, to improve diagnosis, and to develop patient-tailored treatment.
The full-blown disease has a significant social impact since participation in social activities is severely hampered. Up to date, no specific biomarkers have been found, allowing unequivocal diagnosis of the disease. Therefore, diagnosis largely relies on the exclusion of confusable diseases 
. IC/BPS comprises the classic IC/BPS, presenting cystoscopically identifiable Hunner lesions of the urothelium (HIC), contrasting that the non-Hunner-type interstitial cystitis (NHIC) 
and other molecular subtypes may emerge in the future. Furthermore, the etiopathology of IC/BPS is unknown. Since therapeutic options are limited, early detection of patients susceptible to IC/BPS is one of the most urgent clinical challenges.
2. Genetic Factors—Potential Drivers of IC/BPS
Numerous studies suggest an indirect relationship of genetics and the special occurrence of IC/BPS. Frequent comorbidities such as allergies or autoimmune diseases imply a certain share of genetics on the development of IC/BPS (Table 1
), and significant associations have been shown 
. There is also significant evidence for cross-sensitization mechanisms which might drive one or the other organ disease 
. However, up to date there is direct evidence of genetic alteration leading to IC/BPS.
Table 1. Diseases overrepresented and frequently associated with IC/BPS.
|Allergies and autoimmune disorders
|Asthma (especially the non-allergic type)
|Irritable bowel syndrome, Colitis ulcerosa
|Hashimoto’s thyroiditis and Hyperthyroidism
Some studies suggest at least a genetic predisposition of IC/BPS. Monozygotic siblings developed significantly more frequently IC/BPS symptoms (five out of eight) than dizygotic siblings (none of 26 siblings). The risk of developing IC/BPS was 17 times higher in first-grade relatives 
. However, a large study based on the Swedish Twin Registry including >25,000 twins revealed in women that the development of IC/BPS is substantially influenced by environmental factors, whereas genetic factors accounted for less than one-third of the observed variance 
On the other hand, Allen-Brady and coworkers reported a significant excess risk of near relatives for the IC/BPS associated conditions myalgia, fibromyalgia, and constipation. Most interestingly, there was also an excess risk for IC/BPS in patients with myalgia, fibromyalgia, and constipation, supporting the notion of a common underlying genetic factor of those diseases shared with IC/BPS 
. In a recent genetic linkage analysis, the authors found IC/BPS-associated alterations in chromosome 3 and possibly several others on chromosomes 1, 4, 9, and 14, indicating a genetic predisposition for IC/BPS 
. Larger studies are necessary for validation of these interesting data.
Single nucleotide polymorphisms (SNPs) analysis revealed a significant higher prevalence of the homozygote rs11127292 allele (genotype CC) in IC/BPS. Furthermore, the polymorphic allele rs6311 was detected in 90.5% of the patients with severe pain 
While validation of IC/BPS-associated genetic alterations is still pending, the results are promising for the development of novel diagnostic tests. Genetic predisposition might concern the immune system and relate to other autoimmune or allergic diseases.
3. Gene Expression, Networks and Signaling Pathways
Besides alterations in single genes, specific alterations in protein networks and signaling pathways have also been identified in IC/BPS patients by bioinformatic analysis of public expression data sets in the GEO database 
. Despite the number of available expression data sets in IC/BPS was still very small (23 IC lesions vs. 9 normal tissues), the authors found 42 differentially expressed genes (DEGs). A total of 41 of those DEGs formed a protein-protein interaction network (PPI) of 41 knots, of which 11 genes were altered more than 10-times and could be addressed as central knot genes. Those were mainly cell-surface proteins and related to inflammation and immune system activation. In subgroup analyses, 12 DEGs were exclusively associated with HIC, while 27 DEGs were clearly associated with NHIC. Amongst others, the chloride voltage-gated channel 3 (CLCN3) was overexpressed 
. This chloride channel is expressed in the smooth muscle and the urothelium and could trigger pain by spontaneous depolarization. Furthermore, overexpression of several genes of the protein S100 family were identified, playing a key role in NF-κB mediated inflammation. In addition, E2F1 and CCNA2, both associated with cell cycle, were upregulated in IC-lesions. However, their roles in IC/BPS remains unclear.
Gene expression is closely regulated by microRNAs (miRNAs) and disease associated alterations of miRNA expression have been detected, including urothelial carcinoma 
. Recent studies have provided evidence for a significant role of miRNAs in inflammation and tumor. In IC/BPS, Arai et al. found upregulation of 163 and downregulation of 203 miRNAs in IC/BPS-patients compared to healthy controls. Especially members of the miR-320 family were affected, which regulate the expression of the transcription factors E2F1, E2F2 and TUB. Immunohistochemistry supported the overexpression of those transcription factors in IC/BPS 
Many miRNAs were upregulated in IC/BPS-patients, including miRNAs inhibiting the transcription of neurokinin receptor genes (TACR1 and TACR2), correlating with the downregulation of those neurokinin receptors at protein level 
. Upregulation of miR-199a-5p in IC/BPS patients could impair the urothelial barrier via inhibition of the gene expression of several tight junction-associated proteins 
. I In summary, miRNAs are promising molecules for the development of novel molecular diagnostic tests due to their broad involvement in cell proliferation, cell differentiation, inflammatory response, and fibrosis. Such biomarkers might significantly improve the currently tissue-based diagnosis of IC/BPS, suffering from very limited validity of pure histopathological evaluation (e.g., of the mast cell infiltration and fibrosis in bladder biopsies). Furthermore, the pleiotropic effects of miRNAs could also explain the numerous associations of IC/BPS with allergic diseases. Therefore, miRNAs may also prove a valuable new therapeutical approach.
As recently shown, long-noncoding RNA (i.e., maternally expressed gene 3 (MEG3)) is upregulated in HIC patients compared to a healthy control group and upregulates the endosomal toll-like receptor 7 (TLR7), a pattern recognition receptor detecting single strand RNAs (ssRNAs) from bacteria 
, viruses 
and self-antigens 
, finally leading to the release of inflammatory cytokines in HIC bladders 
. TLR7 was found to be upregulated in the urinary bladders of HIC patients 
. Interestingly, the action of MEG3 is enhanced by the downregulation of miR-19a-3p, competing on the RNA binding site in MEG3 
. These findings suggest that HIC etiopathology involves bacterial or viral infection, requiring participation of B-cells, macrophages, and dendritic cells 
4. Occult Urinary Tract Infections and the Microbiome
Several recent studies support the view that IC/BPS might be triggered by occult uropathogens, bacteria or viruses. As shown by Aydogan et al., a special microbiologic culture method and real-time polymerase chain reaction (RT-PCR) can detect different uropathogenic bacteria in the urine of symptomatic IC/BPS patients, previously diagnosed with sterile urine sample 
. Especially the cell wall-deficient, so-called L-form bacteria were detected, which are susceptible to different osmolarities in culture media. This study supports previous findings that only 33% of the bacterial population were detectable by standard culture methods 
An altered microbiome could promote the establishment of pathogenic microbes. However, a comprehensive analysis of the microbiome in urine samples and vaginal smear of 41 pre-menopausal women diagnosed with IC/BPS did not reveal any differences compared to unaffected controls 
. Of interest, however, was a higher correlation between urine and vaginal samples, indicating a reduced diversity of the microbiome. This could be interpreted due to the impaired barrier function in the epithelium. In addition, age-related alterations of the urinary microbiome, and especially the changes in Lactobacillus may contribute to recurrent urinary tract infections in post-menopausal women. The role of the microbiome in IC/BPS is still unclear 
. However, since those studies relied on standard cultivation methods, novel extended cultivation could lead to new insights.
5. Virus Infections in IC/BPS?
New evidence has accumulated indicating a role of viral infections in IC/BPS. As shown by Jhang et al., the Epstein-Barr virus (EBV) was present in 50% of the patients with Hunner-lesion (HIC), but in only 8.6% of the NHIC patients. In controls, the authors could not detect EBV-RNA 
. Also, BK polyoma viruses (BKPyV), which can induce hemorrhagic cystitis 
, were found in the urine of IC/BPS patients and could play a role in the etiopathology of IC/BPS 
. Occult bacterial infection, persisting in macrophage was already demonstrated in endocarditis 
. If this could be a blueprint for the occult persisting viral infection in IC/BPS still has to be investigated.
Recently, evidence was found for a virus-induced cystitis in COVID-19 patients. The infection of a subpopulation of urothelial cells expressing the angiotensin-converting enzyme 2 (ACE2) receptor could indeed be responsible for the occurrence of a de novo urgency in patients showing IC/BPS typic sterile urine cultures. However, it is unclear whether the infection happens luminal via the urine or basal viremic. A local endotheliitis could also play a role and would also provoke local hypoxia in the tissue 
. In addition, the urine of COVID-19 patients with sterile de novo cystitis showed elevated cytokine levels, which could be produced by the urothelial cells or come from renal excretion 
. Elevated levels of cytokines and chemokines were already described in patients with idiopathic urgency and inflammatory bladder alterations 
6. Mast Cells and Lymphocytes as Biomarkers
Mast cell infiltration correlates with the infiltration of other immune cells and cannot serve as a good differential diagnostic criterion 
. Neither in the lamina propria nor in the detrusor significant differences were evident in a recent systematic immunohistochemical study 
. However, the focal clonal expansion of B-lymphocytes and the occurrence of plasma cells seem to be characteristic for HIC 
, raising the question what may be the relevant immunologic processes evoked by the clonal B-cell response in IC/BPS. Clonal B-cell expansion has been observed in several autoimmune diseases, such as Sjögren’s syndrome 
and rheumatoid arthritis 
, but also may be triggered by bacterial infection with Helicobacter pylori 
or by the Epstein-Barr virus 
. In a recent study, almost 60% of the IC/BPS patients turned out to have an occult Epstein-Barr virus infection 
. This might open a new causality chain of the development of IC/BPS, especially of HIC. Interestingly, the virus was mainly located in T-cells, suggesting that not only B-cells but also T-cells are involved 
7. Growth factors and pro-inflammatory mediators modify the bladder histology
Platelet activating factor (PAF), present in the urine of IC/BPS patients , is associated with several allergic diesases, triggering immunologic reactions, including iNOS, COX-2, IL-6 and TNF. Furthermore, PAF can increase the production of MMPs , which can lead to disruption of the urothelial barrier . Growth factors, for sure, play a role in IC/BPS, modulating basic bladder histology. Vascular Endothelial Growth Factor (VEGF), regulating the angiogenesis and lymphangiogenesis, is one interesting candiate. Of special interest is its release by mast cells upon stress via increased corticotropin releasing hormone (CRH) levels. Furthermore, increased CRH-receptor 1 expression in the urothelium and the lamina propria of IC/BPS patients positively correlated with nerve growth factor (NGF) levels . In consequence, acute stress can increase the vascular permeability and may alter nerve fibers in the urinary bladder .
Author growing understanding of the complex gene regulation mechanisms, the role of growth factors and modulators, and the cellular crosstalk with the immune system has greatly improved our view of IC/BPS. A single decisive cause becomes more and more improbable, and several subtypes of IC/BPS crystallizes, furthermost HIC and NHIC. The latter will probably split into several subgroups defined by their specific pathological causality. New, especially molecular biomarkers are at the horizon shedding light into the confusing situation of IC/BPS pathology and etiopathology. PAF enhances MMP-2 production in rat aortic VSMCs via a β-arrestin2-dependent ERK signaling pathway