We queried publicly available datasets for CXCR6/CXCL16 expression using Geo2R software. These included human data from Scholtissek et al. GSE95474 (13), Yildiz-Altay et al. (1), and Seremet, Domizio et al. GSE193068 (14, 15); mouse data from Mande et al. (6); and canine data from Garelli & Wong and Amudzi et al. (16, 17). Normalized count values were plotted in GraphPad Prism software v 10. Each dataset was assessed for normality before selection of statistical tests, with normally distributed data employing t tests and non-normally distributed data employing Mann–Whitney U tests. Directional hypothesis testing for one-tailed results was to test the hypothesis that CXCR6/CXCL16 were higher in lupus than controls, which was based on our observations from the Balb/c mouse model and published studies.

Wu et al. (18) examined urinary biomarkers in lupus nephritis using both murine models and human patient samples. Researchers analyzed the urine of lupus-prone mice and SLE patients for VCAM-1, P-selectin, TNFR-1, and CXCL16, finding significantly elevated levels in those with active nephritis. These markers correlated with disease severity and were primarily derived from the kidney rather than the bloodstream. Immunohistochemical analysis confirmed their presence in inflamed renal tissues, particularly in endothelial and tubular cells. In human SLE patients, urinary levels of these biomarkers distinguished those with nephritis from non-nephritic individuals with high sensitivity and specificity. VCAM-1 and P-selectin, both involved in inflammation, were particularly enriched in lupus kidneys and urine, while TNFR-1 and CXCL16 were newly identified as urinary biomarkers. The findings suggest these molecules play roles in disease pathogenesis while serving as potential noninvasive diagnostic tools. Their detection in urine provides a more accessible way to monitor disease progression without the need for invasive kidney biopsies.

Qin et al. (19) investigated serum soluble CXCL16 (sCXCL16) levels in patients with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), and compared these levels to healthy controls. A total of 47 serum samples were collected from 35 SLE patients, with 12 patients providing samples before and after treatment with glucocorticoids and immunosuppressants. The results showed that serum sCXCL16 levels were significantly higher in SLE patients compared to RA patients and healthy controls. The study also found that sCXCL16 levels correlated with disease activity in SLE, as indicated by SLE Disease Activity Index (SLEDAI) scores, and were higher in patients with active disease flare-ups. Furthermore, the levels of sCXCL16 were linked to organ damage, specifically in patients with lupus nephritis and cutaneous involvement, suggesting that elevated sCXCL16 could serve as a marker for these conditions. Following treatment, sCXCL16 levels decreased, reflecting improvements in disease activity, although there was no significant change in chronic organ damage. The findings also suggested that sCXCL16 might help monitor disease progression and guide treatment decisions in SLE patients. The study concluded that sCXCL16 could be a useful biomarker for assessing disease activity and organ involvement, particularly in the skin and kidneys, in SLE.

Ayyappan et al. (20) analyzed changes in various immune markers in patients with systemic lupus erythematosus (SLE). The study included 30 SLE patients and 30 age- and gender-matched healthy controls, with comprehensive exclusion criteria ensuring the exclusion of those with recent infections or steroid use. It found that soluble CD14 (sCD14) levels were significantly elevated in SLE patients, with a positive correlation between sCD14 levels and disease activity, indicating a potential role for endotoxemia in SLE progression. EndoCAb IgM levels were reduced in SLE patients, suggesting that lower endotoxin neutralization might contribute to elevated sCD14. Additionally, increased levels of lysozyme were found in SLE patients, and lysozyme correlated positively with sCD14, LBP, and FABP2, suggesting a shared stimulus for these immune factors. CXCL16 levels were also elevated in SLE patients and correlated with sCD14, supporting the idea of bacterial exposure influencing these immune responses. The study noted no significant changes in galectin-3 levels in SLE patients, though a positive correlation with sCD14 and lysozyme was observed. Immunosuppressive treatments were found to reduce EndoCAb IgM levels but did not affect the ratios of EndoCAbs to total immunoglobulins.

Singh et al. (21) investigated the potential of urinary biomarkers, specifically VCAM-1, MCP-1, and CXCL16, in monitoring lupus nephritis (LN) activity and chronicity. The study involved 74 LN patients, 13 healthy volunteers, and 22 disease controls, with a focus on urinary biomarker levels before initiating any new immunosuppressive therapy. It was found that urinary VCAM-1 and MCP-1 levels were significantly elevated in patients with active renal disease, correlating strongly with disease activity scores and 24-h proteinuria. VCAM-1, in particular, was a strong discriminator of active renal disease and was most elevated in patients with class IV LN, which has a poor prognosis. The study also showed that VCAM-1 levels correlated positively with the acute inflammation index (AI), a measure of kidney inflammation, while negatively correlating with the chronicity index (CI), indicating its role in reflecting inflammation and potential reversibility of kidney damage. Interestingly, while VCAM-1 was not specific to LN and was elevated in other types of nephritis, it appeared to be a useful marker for renal injury. The results suggested that urinary VCAM-1 could serve as a valuable marker for monitoring disease activity and potentially predicting kidney damage in LN.

Wen et al. (22) investigated the potential role of IFN-γ, CXCL16, and suPAR as biomarkers in systemic lupus erythematosus (SLE) and lupus nephritis (LN). Fifty SLE patients were divided into LN and non-LN groups, alongside 15 healthy controls, and their serum, urine, and renal tissue samples were analyzed. The results showed that serum and urine levels of IFN-γ, CXCL16, and suPAR were significantly elevated in SLE patients, particularly in those with LN, and correlated with disease activity indicators such as SLEDAI scores and 24-h proteinuria. Immunohistochemical analysis revealed increased expression of these biomarkers in renal tissues of LN patients, further linking them to kidney inflammation and damage. Strong correlations were observed between serum and urine levels of these markers, suggesting their potential as non-invasive indicators of renal involvement in SLE. Additionally, suPAR showed the strongest association with disease severity, particularly proteinuria, highlighting its potential as a key predictor of renal damage. These findings suggest that IFN-γ, CXCL16, and suPAR interact in the pathogenesis of SLE, likely contributing to local inflammation and systemic disease progression.

le Blanc et al. (23) studied the role of chemokines (CK) in T cell migration into the central nervous system (CNS) during inflammatory conditions. The researchers found that CXCL16 levels were significantly higher in cerebrospinal fluid (CSF) compared to serum, whereas CCL17 and CCL18 were absent in CSF. Elevated CXCL16 was observed in inflammatory conditions such as multiple sclerosis (MS), systemic lupus erythematosus (SLE), and meningitis. Microglia and astrocytes are likely responsible for CXCL16 production, and its increased presence may facilitate immune cell interactions and inflammation. The study also confirmed that CCL18 was elevated in SLE serum but absent in CSF, supporting its role in attracting naïve T cells outside the CNS. The findings suggest that CXCL16 contributes to neuroinflammation by promoting the influx of activated T cells, particularly when the blood-brain barrier (BBB) is compromised. The researchers highlight that CXCL16's role is not limited to neuroinflammatory diseases, as similar mechanisms were observed in rheumatoid arthritis.

El-Gamasy et al. (24) aimed to evaluate urinary neutrophil gelatinase-associated lipocalin (uNGAL) and CXCL16 levels in children and adolescents with systemic lupus erythematosus (SLE) and their role in diagnosing lupus nephritis (LN). A total of 80 SLE patients and 60 age- and sex-matched healthy controls were included, with disease activity assessed using the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) and renal biopsies performed at initial diagnosis. Urinary uNGAL and CXCL16 levels were measured using enzyme-linked immunosorbent assays (ELISA), and statistical analyses were conducted to assess their diagnostic accuracy. The study found significantly higher levels of uNGAL (8.9 ± 3.56 ng/dl) and urinary CXCL16 (1,067 ± 367 µg/L) in SLE patients compared to controls (P < 0.05). uNGAL showed a stronger correlation with 24-h urinary protein levels (r = 0.732) and SLEDAI scores (r = 0.359) than CXCL16, indicating its higher diagnostic value. The sensitivity and specificity of uNGAL in predicting LN were 95% and 90%, respectively, making it a more reliable biomarker than CXCL16. Both biomarkers were found to increase with LN severity, with the highest levels observed in Class IV LN, which is associated with worse outcomes. The findings support the use of uNGAL as a potential early predictor of LN progression, aiding in disease monitoring and treatment decisions. However, the study's limitations include the small sample size and the inability to establish specific cutoff values for different LN classes. Overall, uNGAL and CXCL16 are valuable indicators of LN activity, with uNGAL demonstrating superior diagnostic accuracy.

Klocke et al. (25) investigated the role of chemokines and their receptors in the recruitment of inflammatory cells into the kidneys of lupus nephritis (LN) patients and their potential as biomarkers. The study demonstrated that T cells and macrophages infiltrate the renal interstitium and that specific chemokines such as CCL2, CCL3-5, CXCL10, and CXCL16 play a key role in disease activity. Urine analysis revealed distinct chemokine profiles in active LN patients, with significant differences from serum levels, highlighting their potential as non-invasive biomarkers. Chemokines correlated with disease severity and urinary T-cell and macrophage counts, particularly CXCR3 ligands like CXCL10, which showed strong correlations with CD4+ T-cell levels. The study also identified that CD4+ T cells exhibit diverse chemokine receptor expressions (CCR5, CXCR3, CCR4, CCR6, CXCR6), whereas CD8+ T cells mainly rely on CCR1, CCR5, and CXCR3 for recruitment. Macrophage recruitment appeared to be driven by the CCR2 axis, with elevated CCL2, CCL7, and CCL8 levels in active LN patients. Individual variability in chemokine expression was observed, suggesting that different LN patients may have distinct chemokine recruitment pathways. Urinary chemokine concentrations did not correlate with serum levels, supporting their kidney-specific role. The study provides new insights into leukocyte migration in LN, aligning with findings from rodent models and previous human studies. These findings highlight potential therapeutic targets and reinforce urinary chemokine profiling as a promising diagnostic tool for LN.

Hassan et al. (2) investigated serum sCXCL16 levels in juvenile systemic lupus erythematosus (jSLE) patients and its correlation with disease activity, particularly lupus nephritis. It included 27 Egyptian children with jSLE diagnosed using ACR criteria and 30 healthy, age- and sex-matched controls. Clinical symptoms, laboratory tests, and renal biopsies were performed to assess disease severity, with disease activity classified using the SLEDAI score. Serum sCXCL16 levels were significantly higher in jSLE patients than in controls and correlated positively with lupus nephritis severity, SLEDAI score, anti-dsDNA levels, ESR, and blood pressure. A strong association was observed between sCXCL16 levels and specific cutaneous manifestations, particularly alopecia and malar rash. Patients with more advanced lupus nephritis (class III and IV) had significantly elevated sCXCL16 levels, suggesting its role in disease progression. The study found a negative correlation between sCXCL16 levels and C3, highlighting its potential as a biomarker for disease activity. The findings align with previous studies on the inflammatory role of CXCL16 in lupus nephritis and its association with renal damage. The study suggests that sCXCL16 could serve as a non-invasive marker for monitoring renal involvement in jSLE, reducing the need for repeated renal biopsies. Future therapeutic strategies targeting sCXCL16 could be explored to manage active disease flares and prevent renal complications in jSLE patients.

An et al. (26) studied the role of IL-17-secreting invariant natural killer T (iNKT) cells in lupus nephritis (LN). They found that patients with LN had significantly higher proportions of IL-17-producing NKT cells in peripheral blood mononuclear cells (PBMCs) and increased IL-17 expression in kidney tissues compared to control groups. Using an experimental autoimmune lupus nephritis (eALN) mouse model, researchers observed that NKT cell-deficient mice had less severe disease progression, lower proteinuria, and reduced renal damage than wild-type mice. The absence of NKT cells led to decreased glomerular immune cell infiltration, lower IL-17 and proinflammatory cytokine expression, and increased levels of IL-27, which inhibits Th17 differentiation. Experiments demonstrated that NKT cells contribute to systemic inflammation and IL-17 production, promoting mesangial cell proliferation and disease progression. in vitro studies confirmed that activated NKT cells induced IL-17 secretion, which was diminished upon IL-17 blockade, reducing mesangial cell proliferation. NK1.1+ NKT cells were identified as primary regulators of IL-17 production, with their activation upregulating IL-6, STAT3, and CXCL16 expression, further linking them to LN pathogenesis. A pristane-induced lupus model reinforced these findings, showing that disease severity correlated with IL-17 and NKT cell activation. Blocking IL-17 or depleting NKT cells reduced inflammation and kidney damage, suggesting that IL-17-secreting iNKT cells are central to LN progression. These findings highlight the potential therapeutic targeting of the IL-17/NKT cell axis to mitigate LN severity.

Ko et al. (27) examined gene expression profiles in cutaneous lupus and lupus nephritis to understand their distinct clinical and histological features. Researchers analyzed 16 cutaneous lupus samples (SCLE and CCLE) and 10 normal skin samples, revealing that while lupus skin samples are distinct from normal skin, SCLE and CCLE share highly similar gene expression profiles. Despite clear clinical differences, broad gene expression analysis does not explain these distinctions, suggesting that subtle genetic variations or tissue-specific inflammation may contribute. The study identified a strong type I interferon (IFN) signature in cutaneous lupus, with upregulated genes such as IFI6, IFI44l, and CXCL10, which play key roles in inflammation and immune response. In contrast, lupus nephritis samples showed distinct gene expression patterns, including increased levels of CCL19 and TNFRSF11B, which are associated with kidney inflammation and damage. The data suggest that cutaneous lupus is characterized by strong immune activation, particularly involving T-cell chemokines and type I IFN pathways, whereas lupus nephritis exhibits a more wound-healing and fibrotic response. Using CIBERSORT analysis, the study found that cutaneous lupus had a predominance of activated CD4+ T cells, whereas lupus nephritis showed more resting memory T cells and M2 macrophages, supporting different immune mechanisms. The study proposes that lupus nephritis may result from immune complex deposition originating from inflammation in distant tissues, such as the skin. Despite similarities in early histological presentation, cutaneous lupus and lupus nephritis follow different immune response pathways, emphasizing the complexity of lupus pathogenesis. These findings provide new insights into lupus biology and may help refine diagnostic and therapeutic approaches for different lupus subtypes. A summary of the findings of these studies is presented in the forest plot in Figure 2.

We queried our mouse (1, 6), canine (16, 17) and human (1) CLE transcriptomics datasets to assess CXCR6/CXCL16 expression in skin, and compared these to other publicly available human datasets GSE95474 from Scholtissek et al. (13) and GSE193068 from Seremet, Domizio et al. (14, 15) (Figure 3). Our human dataset is presented in Yildiz-Altay et al. (1), and is comprised of n = 6 healthy and 12 biopsies from 10 DLE patients who were Black. CXCR6 was upregulated in CLE skin vs. controls across all 3 species (Figure 3A–E). CXCL16 was significantly elevated in Balb/c mouse, dog and Scholtissek et al. datasets, but did not reach significance for B6 mouse or skin of color patient samples (Figures 3F–J). To evaluate the possible utility of CXCR6 as a biomarker of cutaneous lupus, we performed Receiver Operator Characteristic curve (ROC) analysis of CXCR6 expression on our dataset, which revealed 100% sensitivity with 83.3% specificity at a normalized count value of 15.33 with an AUC of 0.8333 and P value 0.0246 (Figure 3K). Combining our dataset with Scholtissek et al, which we harmonized by using absolute RNA counts, generated an ROC with 94.4% sensitivity and 81.82% specificity with an AUC of 0.8889 and P value 0.0005 at a normalized count value of >8.071 (Figure 3L).