Intraocular Immune Response in Human Uveitis: Time to Look Beyond Animal Models

Abstract

Purpose

To review the current and future approaches to investigating the intraocular immune response in human uveitis.

Design

Perspective.

Methods

Review of currently available methods for investigating the immune response in ocular tissues and fluids in patients with intraocular inflammation/ uveitis. The advantages and disadvantages of human studies have been compared to those of animal models of uveitis.

Results

Animal models, while being excellent tools for mechanistic studies, do not replicate the clinical and immunologic heterogeneity of human uveitis. Opportunities for immunological studies in human uveitis are mostly limited to histological studies, or sampling of intraocular fluids and peripheral blood. Histopathological studies can be enhanced by revisiting published historical data, tissue repositories, or autopsy specimens. Intraocular fluids can be investigated by a variety of techniques. Among these, flow cytometry and single-cell RNA sequencing (scRNAseq) provide single-cell resolution. While the current technology is costly and labor-intensive, scRNAseq is less limited by the low cellular yield from intraocular fluids and allows unbiased immune profiling enabling discovery of new cellular subsets. Immunological phenotypes uncovered from human data can be further investigated in animal studies.

Conclusion

The diversity of the intraocular immune response in uveitis patients remains challenging but can be studied by multiple techniques including histopathology, flow cytometry, and scRNAseq. Human data can be combined with animal studies for translating uveitis research into novel therapies.

Our understanding of the pathogenesis of human uveitis is largely derived from studies in animal models of uveitis.¹ While providing significant mechanistic insights, these models do not recapitulate many of the fundamental characteristics of human uveitis.² Immunological data from human uveitis has largely been constrained by the limited access to ocular tissue samples from uveitis patients and the relative lack of tools to meaningfully analyze the available samples. In this perspective, we discuss the value of investigating the intraocular immune response in human uveitis and how it can complement data from animal models of uveitis.

DIFFERENCES BETWEEN HUMAN UVEITIS AND ANIMAL MODELS

Human uveitis is a highly heterogeneous entity comprising of more than 30 clinically distinct phenotypes with intraocular inflammation.³ These uveitis conditions may share common inflammatory signs such as anterior chamber or vitreous inflammation or may have unique morphological patterns of retinitis and choroiditis. Various combinations of these inflammatory signs result in a wide array of conditions that are collectively labeled as uveitis. Further, individual patient phenotypes can vary considerably, even within the same disease. It is reasonable to hypothesize that an equally diverse range of immunological responses would be underlying the clinical manifestations of these uveitis conditions. These would be shaped not only by the drivers of the inflammatory response such as infection or autoimmunity, but also sculpted by genetics (particularly the human leukocyte antigen [HLA] alleles), microbiome, and environmental influences.⁴ However, scant data is available on the nature of the intraocular immune responses underlying these uveitis conditions.

Much of our knowledge of the molecular mechanisms of uveitis are based on experimental models of animal uveitis (Table 1) and have been excellently summarized in several review papers on this subject.^1,2 Most experimental uveitis models in rodents involve applying an immunologic trigger either directly into the eye or as a systemic immunization against an ocular-expressed protein. The resulting immune response is generally monophasic, where the inflammation resolves over time. In contrast, the immunologic trigger in human disease remains unclear, and most common human uveitis entities are either chronic or recurrent. While animal models are generally not expected to exactly replicate human disease, the gap between human and experimental models of uveitis may be wider than in other organ-specific diseases, given the differences in ocular anatomy, barriers, and absence of a macula in rodents. This gap may be further increased by the limited number of major histocompatibility complex (MHC) alleles in mice/rats,⁵ the controlled laboratory environment, and inbred genetic status used in experimental uveitis. Thus, animal models of uveitis have a limited range of inflammatory manifestations that cannot match the diversity of human uveitis.

Table 1.

Major animal models of uveitis

Type of model	Method of induction	Primary signs of intraocular inflammation	Major cell types
EAU – ‘classic’ (mice/rats)	Immunization with retinal antigen + CFA, or adoptive transfer of antigen-pulsed T-cells	Vitritis, retinal vasculitis, chorioretinitis, hemorrhages, granuloma, sub-retinal neovascular membrane	CD4+ and CD8+ T-cells, (Th17 lineage for classic EAU, Th1 for adoptive transfer and IRBP TCR spontaneous models)
EAU – spontaneous (mice)	IRBP TCR transgenic; HLA-A29 transgenic; AIRE KO; Transgenic HEL or β-gal under the control of an eye-specific promoter	Vitritis, retinal vasculitis, chorioretinitis
Endotoxin-induced uveitis (mice/ rabbits)	Systemic or intraocular injection of LPS in mice/ local in rabbits	Acute inflammation of the iris and ciliary body	Neutrophils
Primed mycobacterial uveitis (rats/ rabbits)	Immunization with a killed mycobacterial extract followed by intravitreal injection of same extract.	Anterior chamber inflammation	CD68+ macrophages

EAU: Experimental autoimmune uveitis; CFA: Complete Freund's Adjuvant; IRBP: interphotoreceptor retinoid binding protein; TCR: T-cell receptor; AIRE: autoimmune regulator gene; HEL: Hen Egg Lysozyme; LPS: Lipopolysaccharide

Despite these limitations, animal models offer several advantages that are technically not feasible in human studies. Animal models allow controlled genetic manipulations, uveal (and other ocular) tissue evaluation at the peak of inflammation, longitudinal observations of immunological data, and therapeutic interventional studies. Limited heterogeneity can also be modeled in mouse models by using outbred or wild-derived mice, though these are not routinely used. Thus, animal models are invaluable in conducting mechanistic studies required for a complete understanding of human uveitis and treatment response. Indeed, several immunotherapeutic interventions were successfully trialed for efficacy in animal models of uveitis.^6,7 Nonetheless, animal models can only partially represent human disease, including the recurrent and chronic course of human uveitis. For example, while the endotoxin-induced uveitis (EIU) in rats manifests as anterior segment limited inflammation, similar to HLA-B27 uveitis, it does not replicate the chronic, recurrent course of disease found in the latter.⁸ Thus, there is an imminent need to explore investigational tools that allow effective utilization of biological samples available for studying human uveitis. In the subsequent sections, we discuss some of these approaches, including histopathological studies and 2 emerging techniques for investigating intraocular fluids—flow cytometry and single-cell RNA sequencing (RNA-Seq).

HISTOPATHOLOGICAL STUDIES AND REVISITING HISTORICAL DATA

Histopathological studies provide the most direct evidence of the immunopathogenesis of intraocular inflammation in human uveitis. However, uveal, or retinal tissue biopsies are generally not available from patients as they can undermine the structural and/or functional integrity of the eye. Historically, histopathological evaluation provided unique insights into the cellular basis of intraocular inflammation. However, much of this data came from studies that were published prior to the introduction of anti-inflammatory (corticosteroid and immunomodulatory treatments) and antimicrobial therapy. Lack of appropriate and timely therapy at that time often led to progressive inflammation that ultimately required enucleation of the painful eye. Thus, much of the historical histopathological analysis was based on end-stage disease, and not on pathophysiology occurring at a stage amenable to therapeutic intervention. Histopathological studies have also been performed in postmortem samples, of patients documented to have had uveitis. Here again, most specimens are obtained from elderly patients, in whom uveitis that occurred as a young adult would have gone into remission or progressed to complete tissue destruction and no longer represents the active disease state. Yet it is a positive testament to the research progress in this field, especially with therapeutics, that only a minority of eyes now reach the stage of enucleation.

Modern molecular techniques have been applied to existing histopathologic specimens. Tissue sections or paraffin blocks in ocular pathology laboratories can be re-examined using more advanced microscopy, and immunohistochemical studies. For example, recent studies in ocular tuberculosis (TB) localized mycobacteria to the retinal pigment epithelium,⁹ and demonstrated histological and molecular evidence of TB even in eyes from patients with negative TB tests.¹⁰ Additionally, in a newly emerging field termed spatial biology, vast numbers of proteins and RNA molecules can be simultaneously measured in the tissue sections, adding novel information about the subcellular location of these molecules. These techniques are yielding unprecedented amounts of data from tissue, even in historical formalin-fixed paraffin-embedded specimens.¹¹ Given the rapid pace of technical improvements and commercialization, it may soon be viable to systematically re-examine precious historical specimens to extract new insights.

Tissue repositories of different biological samples provide the opportunity for increasing global collaboration for the sharing of rare uveitis specimens among uveitis specialists and pathologists. A recent study on the histopathology and cellular localization of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in ocular tissues from autopsy specimens demonstrates the potential advantage of such repositories.¹² Finally, modern imaging tools such as adaptive optics scanning laser ophthalmoscopy, promise to allow investigation of the disease in living eyes with unprecedented resolution, and can be correlated to histological studies, while providing opportunities for studying different phases of the disease, including early presentations and resolution of inflammation with treatment.¹³

OCULAR FLUID SAMPLING AND INVESTIGATION

Aqueous and vitreous fluids are commonly used to investigate the causes of intraocular inflammation. These fluids are in constant contact with inflamed uveal tissue and the retina and are therefore able to provide some representation of the ongoing inflammation in the tissues. In that sense the aqueous and vitreous can be compared to other body fluids such as the cerebrospinal fluid or the synovial fluids, where correlations between the fluids and their respective tissues have been obtained.¹⁴ In clinical settings, ocular fluids are typically used for the diagnosis of infectious uveitis or masquerade conditions.¹⁵ The tests for infectious uveitis include polymerase chain reaction (PCR) or ELISA-based studies for intraocular antibody production. Additionally, cytology, immunohistochemistry, and flow cytometry are required for the diagnosis of masquerades such as vitreoretinal lymphoma. In contrast, a wider range of tools are available for investigating ocular fluids for research. These include bulk (transcriptomics, proteomics) and single-cell (flow cytometry, transcriptomics) techniques. Bulk approaches measure average values across large populations of cells and are sensitive to detect low abundance signals. However, they are insensitive to heterogeneity across individual cells within the sample, which is better detected using single-cell technologies.

Flow cytometry: Flow cytometry is a technology for targeted multiparametric analysis of single cells in a solution. It allows measurement of targeted protein levels to quantify specific cell populations.¹⁶ The majority of such studies in human uveitis have been restricted to peripheral blood samples from patients (discussed below). Flow cytometric studies in ocular fluid samples from patients have been limited by the relatively low cellular yield from these samples. Table 2 lists the studies performed on aqueous and vitreous samples in the past 10 years.17, 18, 19, 20, 21, 22, 23, 24, 25, 26 Most studies are limited to characterization of the T-cell subsets including the CD4/CD8 ratio, largely due to the low number of cells available for analysis. A high CD4/CD8 ratio was noted in ocular fluids in sarcoidosis (as also in bronchioalveolar lavage fluid),¹⁷ while a low CD4/CD8 ratio (with high CD8⁺ T-cells) was found in viral retinitis.²² Studies in vitreous samples from tubercular (TB) uveitis have noted proinflammatory responses to retinal antigens, besides TB antigens.²¹ The autoreactive T-cells were also resistant to activation-induced cell death. A subsequent study found retinal antigen-specific intraocular T-cell responses in other infectious and noninfectious uveitis as well.²⁴ The main challenge for future flow cytometry studies in intraocular fluids will be to increase the number of parameters for such low input samples. One approach is the application of spectral flow cytometry where, rather than pre-defined bands of wavelengths, the entire spectrum of the fluorochrome is measured, allowing the assessment of up to 40 proteins per experiment.²⁶ This can allow the identification of multiple immune cell subsets in addition to their functional characteristics.

Table 2.

Flow cytometry studies (2014-2023) on aqueous and vitreous infiltrating cells in different uveitis conditions

Study	Disease etiology	Sample size (eyes)	Key findings
Maruyama et al., 201617	Sarcoidosis (tissue-proven)	22	Vitreous fluid had high diagnostic value for sarcoidosis (equal to bronchoalveolar lavage fluid).
Maruyama et al., 201718	Infectious and non-infectious uveitis	234	Classification of uveitis based on CD4/CD8 ratio, T-lymphocyte and B-lymphocyte subset, and pathogen DNA in vitreous samples.
Tagirasa et al., 201719	Tubercular (TB) and non-TB uveitis	35	Vitreous infiltrating cells in TB uveitis comprised of both central and effector memory cells. They respond to antigenic stimulation with both TB and retinal antigens. Retinal antigen-specific cells are more resistant to activation induced cell death.
Sharma et al., 201820	Tubercular uveitis	8	Vitreous infiltrating cells were more proinflammatory and recently activated (CD69+) than matching blood samples. They were also hyporesponsive to TLR9 stimulation.
Tagirasa et al., 202021	Non-infectious uveitis	4	Vitreous CD4+ T-cells expressed significantly higher P-glycoprotein and pro-inflammatory (IL-17, IFNγ, IL17) cytokines than matching blood samples.
Kang et al., 202122	Acute retinal necrosis (ARN) and acute anterior uveitis (AAU)	7	ARN eyes had higher CD8+ T-cells and lower CD4/CD8 ratios than AAU (all aqueous samples).
Carreño et al., 202123	Infectious and non-infectious uveitis	31	T-cells were the main cellular component of both aqueous (n=17) and vitreous (n=14) samples. CD4/CD8 ratios were highly variable.
Alam et al., 202324	TB-immunoreactive uveitis and non-TB uveitis	71	Polyfunctional cytokine response in undifferentiated TB-immunoreactive patients was significantly higher than in true ocular TB patients, likely representing an exaggerated anti-TB immune response rather than active infection.
Deguchi et al., 2023 (abstract)25	Infectious and non-infectious uveitis	58	Higher proportion of regulatory T-cells in vitreous than blood, in viral uveitis; not in sarcoidosis
Kalnitsky ey al., 2023 (abstract)26	Not provided	Not provided	36-parameter spectral flow cytometry could demonstrate wide heterogeneity in immune cell populations even in low input (<100 cells) ocular fluid samples.

Single-cell RNA Sequencing (scRNA-Seq): Single-cell transcriptomics enables unbiased immune cell profiling, including profiling the immune repertoire, or composition of T-cell receptor (TCR) and B-cell receptor (BCR) expression across a population of cells. These techniques have facilitated discovery of cellular states that previously escaped detection by targeted low-parameter approaches such as flow cytometry.²⁷ Given the challenges associated with small-volume ocular samples, scRNA-Seq is ideally suited for analysis of intraocular fluids. Application of these approaches to uveitis has led to discovery of new cellular subtypes in the aqueous and peripheral blood, as well as the demonstration of clonal lymphocyte expansion, implicating an antigen-specific ocular immune response in some cases of uveitis (Table 3).28, 29, 30, 31 A comparison of 3 studies that performed scRNA-Seq on small cohorts of patients suggests that the composition of aqueous inflammatory cells may vary between disease subtypes or etiologies (Figure 1). First, Hassman et al found that clonal expansion of lymphocytes was a shared feature in 4 patients with granulomatous uveitis, suggesting that antigen-driven immune responses may underlie this disease type.²⁸ In contrast, Kasper et al found higher proportion of granulocytes and myeloid cells in acute anterior uveitis (AAU) and endophthalmitis.²⁹ More recently, Kang et al found a significant clonal expansion of CD4⁺ T-cells in Vogt-Koyanagi-Harada disease (VKH), but higher numbers of myeloid cells in Behcet's disease (BD).³¹ Taken together these 3 studies shed light on the spectrum of ocular immune responses present in patient with uveitis and suggest that idiopathic granulomatous uveitis and VKH share ocular infiltrates dominated by adaptive immune mechanisms, while endophthalmitis, AAU and BD feature prominent innate immune responses. scRNA-Seq can also shed light on potential antigen triggers in uveitis. In a recent study, CD8⁺ T-cells from synovial fluid and aqueous from patients with HLA-B27-associated arthritis and uveitis, respectively, were found to express a common complementary-determining region 3β motif in their TCRs that also bound self and microbial peptides.³⁰ Identification of that shared binding motif suggests that microbial antigens may contribute to the pathogenesis of HLA-B27-associated disease. These studies highlight the immense potential of scRNA-Seq technology in uncovering the immunopathogenesis of infectious and noninfectious uveitis. Despite these singular advances, more comprehensive application and use in a clinical setting is still distant given the high costs and specialist bioinformatic expertise required for scRNA-Seq studies.

Table 3.

Single-cell RNA sequencing studies on aqueous infiltrating cells in different uveitis conditions

Study	Disease etiology	Sample size (eyes)	Key findings
Hassman et al., 202128	Active granulomatous uveitis	4	T- and B-cells from aqueous showed evidence of clonal expansion and distinct transcriptional profile compared to peripheral blood samples, suggesting antigen-driven response.
Kasper et al., 202129	Anterior uveitis (HLA B27 positive and negative) and endophthalmitis	7	Plasmacytoid and conventional dendritic cells were more abundant in aqueous humor of HLA-B27-positive vs HLA-B27-negative uveitis patients
Yang et al., 202230	HLA B27 positive ankylosing spondylosis and acute anterior uveitis	4	Enrichment of specific T-cells in joints and eye vs peripheral blood with TCRs cross-reactive for both self-antigens and microbial antigens, suggesting common antigen trigger in uveitis and arthritis. Conserved CD8+ TCR sequences in peripheral blood, synovial fluid and aqueous humor, suggesting a common antigen driven response in eyes and joints.
Kang et al., 202331	Comparison of VKH and Behcet's disease (BD)	6	Relatively more pro-inflammatory CD4+ Th1 cells in VKH, but relatively more myeloid cells and cytotoxic CD8+ T cells in BD, suggested distinct pathophysiology for these two diseases.

FIGURE 1.

Spectrum of inflammatory infiltrates in human uveitis, demonstrated by 3 single cell RNA-Sequencing studies. Kasper et al (REF) found more granulocytes and myeloid cells in acute anterior uveitis and endophthalmitis relative to the other studies (left top panel); Kang et al (REF) found that myeloid cells were more predominant in Behcet's disease while T-cells predominated in Vogt-Koyanagi-Harada disease (center top panel); and Hassman et al (REF) found that clonally expanded T- and B-cells were prevalent in idiopathic granulomatous uveitis (right top panel). Both Kang et al and Hassman et al also found high levels of T- and/or B-cell clonal expansion in VKH and idiopathic granulomatous uveitis, respectively (not shown). Individual disease types analyzed in these studies are arranged in the lower panel from left to right based on the greatest fraction of innate cell types to predominantly adaptive cell types with greater extent of T-/B-cell clonal expansion. (Figure created using biorender)

Proteomics: Many studies have identified chemokines and cytokines present in ocular fluids from patients with noninfectious and infectious uveitis, several of which have had direct therapeutic implication.³² Traditional methods to analyze intraocular proteins rely on antibodies specific for proteins of interest and have evolved significantly over recent decades from single molecule tests to multiplex assays that measure 20 or more proteins in a single sample. Information from these high throughput protein assays can be integrated with high-parameter single-cell transcriptional data to identify roles for specific cell types in ocular inflammation. For example, we found that intraocular macrophages generated a chemokine gradient comprised of CCL2 and CXCL10 to recruit other inflammatory cells in human uveitis.³³ Recently, aptamer-based techniques, which make use of protein-specific nucleotide sequences rather than antibodies, have vastly increased the analysis capacity of small-volume samples. As vast amounts of data are generated from these multiomic techniques, artificial intelligence is used to identify cell signaling networks relevant to ocular disease.³⁴ In diagnostic studies, mass spectrometry has been used for exploring host biomarkers in the vitreous, for the diagnosis of infectious uveitis such as in TB.³⁵

BLOOD VERSUS INTRAOCULAR FLUID SAMPLING IN HUMAN UVEITIS

Since the 1980s, numerous flow cytometry studies of peripheral blood from uveitis patients have identified altered numbers of activated T-cells, particularly Th1/Th17 CD4 T-cells, as well as evidence that alterations in potentially pathogenic T-cell populations and serum cytokine profiles that correlate with disease activity.³⁶ Similarly, specific populations of peripheral blood dendritic cells have also been found to vary with uveitis disease activity and reactivation and to correlate with intraocular dendritic cell populations.³⁷ Peripheral blood gene expression signatures associated with uveitis have also been identified. For example, peripheral blood gene expression differs between patients with uveitis, and associated systemic diseases as well as healthy controls.³⁸

In contrast, studies in other body organs have revealed that blood does not mirror the immunological changes in tissues. More than 95% of the T-cell population resides in the tissues, especially in the lymphoid organs and barrier surfaces (skin and mucosa).³⁹ This was also aptly demonstrated when respiratory tract and circulating immune cells were compared during the COVID-19 pandemic.⁴⁰ Our own research has revealed the T-cell subsets in the vitreous are phenotypically and functionally different from those in paired blood samples in patients with posterior segment uveitis. Specifically, resident CD4+ memory T-cells were the predominant population in the vitreous displayed distinctly higher cytokine responses to ionophore, retinal autoantigens and microbial antigen stimulation as compared to blood CD4 T-cells (SB, manuscript under review).

Whether peripheral blood biomarkers directly reflect active ocular inflammation,³⁷ or dynamic trafficking of cells between lymph nodes and the eye, remains unclear. Alternatively, peripheral blood may better reflect systemic inflammation in multiorgan diseases such as sarcoidosis or Behcet's disease, or simply the immunogenetic environment that predisposes patients to develop uveitis. Notwithstanding, blood samples may still have a role in tracking the intraocular immune response or predicting personalized response to systemic immunosuppressive therapies. Blood offers the advantage of serial sampling to follow the immune response during the course of the disease and patients are more likely to consent to peripheral blood draws than invasive intraocular sampling techniques. However, more studies are required to define the relationships between intraocular inflammation and peripheral blood samples.

CHALLENGES IN INTRAOCULAR FLUID SAMPLING IN HUMAN UVEITIS

Intraocular fluids offer insights into the immune microenvironment of the eye in both infectious and noninfectious uveitis. Nonetheless, sampling of intraocular fluids is an invasive procedure, and therefore carries a risk of complications, albeit low for most patients and may have diagnostic benefit. Thus, careful patient selection and meticulous surgical technique are paramount for sampling in these “hot” eyes. Fortunately, extensive studies have demonstrated that aqueous or vitreous sampling in uveitis patients is safe.^15,24,25

There are several other challenges in using intraocular fluids for immunological investigations in human uveitis. Posterior uveitis conditions may have minimal anterior chamber reaction, or even minimal vitritis (eg, birdshot chorioretinopathy). Such eyes may provide very low cellular yield for flow cytometry or scRNA-Seq studies. Conversely, we may run the risk of oversampling patients with high cellularity in the vitreous or aqueous, which may skew the data towards certain phenotypes. It is not known if aqueous cells can be representative of posterior segment inflammation, or vitreous cells of retinal or choroidal tissue inflammation. Finally, vitreous samples collected from patients undergoing therapeutic vitrectomy for vitreous floaters following resolved inflammation, may contain primarily dead cells and will not yield useful information. Accumulation of data obtained from studies performed in different clinical scenarios and using different experimental protocols should be able to resolve the above issues. In the long-term, novel vitreoretinal approaches or devices for minimally destructive chorioretinal biopsies could be hugely beneficial, alongside immune cell labelling and longitudinal noninvasive tracking approaches. Additionally, advances in in vivo cellular imaging may facilitate noninvasive identification of cell types and alleviate many of the current challenges.

COMBINING HUMAN AND ANIMAL DATA TO GENERATE NEW PERSPECTIVES

Human and animal research play complimentary roles in generating new perspectives on uveitis pathogenesis and treatment. Traditionally, critical research questions are generated through clinical experience. Molecular analysis of patient samples identifies clinical and immunological phenotypes that can be investigated for mechanistic insights using animal models. Finally, preclinical animal research is critical to characterize and validate potential targets before they are tested in humans. This approach has resulted in the successful translation of animal research into the treatment of human disease. In the context of uveitis, the best examples would be sustained release intravitreal dexamethasone injections,⁶ cyclosporine (calcineurin inhibitor),⁷ and the TNF-( inhibitor adalimumab.⁴¹ In contrast, the IL-17 inhibitor secukinumab, could not replicate the success of preclinical research in patients with uveitis although it remains useful in treatment of axial spondyloarthropathy and psoriasis.⁴² Subsequent animal research showed that IL-17A also activates a negative feedback loop in Th17 cells through NF-κB, and inhibition of IL-17A increased the expression of other Th17 cytokines GM-CSF and IL-17F, exacerbating neuroinflammation.⁴³ The story of secukinumab exemplifies the complexity of uveitis research while moving from bench to the bedside and back to the bench while attempting to explain unexpected clinical outcomes.

FUTURE DIRECTIONS

Significant challenges remain in investigating the human intraocular immune response due to analyses based on (1) ocular fluids bathing the tissues, rather than the tissues themselves, and (2) “snapshot” or fixed data, that cannot be correlated to progression or resolution of disease. An approach to overcoming these challenges could be based on prediction strategies used in other fields such as multidimensional cancer pathology data.⁴⁴ Here, phenotypic and functional data obtained from the current investigational tools can be integrated over different time points, either in real time (by repeat liquid biopsies) or by bioinformatic approaches (eg, pseudotime analysis). The outcomes from such studies can then be validated in phenotypically matched patient samples and/or animal experiments. Such a comprehensive approach will not only lead to novel diagnostic and therapeutic solutions, but also feed into the clinical classification criteria for different uveitis entities.

CONCLUSION

New investigational tools have tremendous potential to uncover the intraocular immune response in human uveitis. Combining human data with animal studies will facilitate meaningful translational research into the diagnosis and treatment of this condition.

CRediT authorship contribution statement

SOUMYAVA BASU: Writing – review & editing, Writing – original draft, Funding acquisition, Formal analysis, Conceptualization. LYNN HASSMAN: Writing – review & editing, Writing – original draft, Conceptualization. SHILPA KODATI: Writing – review & editing, Writing – original draft, Conceptualization. COLIN J CHU: Writing – review & editing, Writing – original draft, Conceptualization.