Incontinentia pigmenti (IP) is a rare X-linked hereditary disease caused by NEMO gene mutation. IP-associated retinopathy (IPR) was classified into 5 stages.1 No report focused on the levels of intraocular cytokines in IPR patients. We measured the aqueous humor cytokines in pediatric IPR eye and analyzed their associations with IPR severity, hoping to provide new insights on pathogenesis and lay the theoretical foundations for treatment.
Materials and Methods
Between March 2015 and March 2017, aqueous humor of eyes receiving intraocular treatment (8 eyes of 6 IPR patients, 18 eyes of 12 congenital cataract patients as control) was enrolled. IPR stages, study design, and procedures followed our previous study1,2 (Supplemental Digital Content 1, Materials and Methods, http://links.lww.com/APJO/A128).
Results
Demographic information and fundus images were collected (Supplemental Digital Content 2, http://links.lww.com/APJO/ A129 and Supplemental Digital Content 3, http://links.lww.com/APJO/A130). Twenty-two tested cytokines concentrations were summarized (Table 1). Three [vascular endothelial growth factor (VEGF), IP-10, intercellular cell adhesion molecule-1 (ICAM-1 )] were significantly higher and 2 [interleukin (IL)-4, granulocyte colony-stimulating factor (G-CSF)] were lower in IPR patients than the control group (P < 0.05). VEGF, IL-8, IP-10 (Interferon-inducible protein), ICAM-1 were significantly increasing and IL-4, G-CSF were significantly decreasing in parallel with increasing IPR stages (P< 0.05).
Table 1 -
Aqueous Humor Cytokine Concentrations (pg/mL) in the IPR Group and the Control Group
|
IPR |
|
|
|
|
Stages of IPR |
| Cytokines |
Stage 2 (n = 2), Mean |
Stage 3 (n = 4), Mean |
Stage 4 (n = 2), Mean |
P value Between IPR Subgroups‡
|
Total (n = 8) |
Control Group (n = 18) |
LOD |
P value Between IPR Group and Control Group |
r
|
P
value§
|
| IL-10 |
0 |
1.39 |
0.065 |
0.374 |
0.00 (0.00,1.44) |
0.00 (0.00, 2.19) |
0.1 |
0.311*
|
−0.197 |
0.334 |
| IL-6 |
5.25 |
7.6075 |
7.7 |
0.327 |
7.04 ±4.27 |
0.93 (0.40, 53.32) |
1.6 |
0.08*
|
0.367 |
0.065 |
| bFGF |
3.875 |
13.54 |
11.75 |
0.513 |
6.83 ±10.97 |
13.01 ±6.73 |
3.4 |
0.51â€
|
−0.135 |
0.509 |
| IL-lβ |
0 |
2.0425 |
0.31 |
0.194 |
0.00 (0.00, 2.37) |
1.05 ± 1.12 |
2.3 |
0.16*
|
−0.263 |
0.194 |
| TNF-α |
0 |
2.82 |
1.62 |
0.147 |
0.00 (0.00, 3.22) |
3.24 ±1.77 |
3.4 |
0.062*
|
−0.329 |
0.101 |
| VEGF |
225.005 |
243.6625 |
191.705 |
0.024
|
226.01 ±86.60 |
23.93 (16.83, 68.24) |
4.5 |
0.001
*
|
0.592 |
0.001
|
| [FN-γ |
0 |
2.735 |
1.44 |
0.161 |
0.00 (0.00, 4.07) |
3.00 ±1.48 |
0.8 |
0.115*
|
−0.284 |
0.159 |
| GM-CSF |
0 |
1.3825 |
0.225 |
0.174 |
0.00 (0.00, 1.73) |
1.18 ± 1.05 |
1.6 |
0.129*
|
−0.288 |
0.154 |
| MlP-lα |
0 |
0.9575 |
0.525 |
0.276 |
0.00 (0.00, 1.32) |
0.28 (0.05, 0.82) |
0.2 |
0.461*
|
−0.09 |
0.661 |
| IL-2 |
0 |
5.165 |
2.975 |
0.11 |
3.33 ±4.64 |
4.81 ±2.63 |
11.2 |
0.31*
|
−0.271 |
0.181 |
| IL-5 |
0 |
0.615 |
0.555 |
0.271 |
0.00 (0.00, 1.06) |
0.62 ±0.36 |
1.1 |
0.238*
|
−0.187 |
0.361 |
| IL-lα |
0 |
0.765 |
0 |
0.307 |
0.00 (0.00, 0.34) |
0.00 (0.00, 0.00) |
1.0 |
0.724*
|
0.111 |
0.590 |
| IL-4 |
0.03 |
0.825 |
0.585 |
0.015
|
0.57 ±0.42 |
1.12 ±O.29 |
1.4 |
0.01
â€
|
−0.546 |
0.004
|
| IL-8 |
13.375 |
6.43 |
24.715 |
0.115 |
12.74±11.37 |
12.46 ±27.20 |
1.2 |
0.98â€
|
0.438 |
0.025
|
| IP-10 |
106.805 |
168.2025 |
176.63 |
0.182 |
154.96±lll.20 |
54.58 (8.29, 127.46) |
0.5 |
0.041
*
|
0.434 |
0.027
|
| G-CSF |
0.1 |
0.825 |
0.615 |
0.05 |
0.59 ± 0.56 |
1.21 (0.87, 1.37) |
1.6 |
0.019
*
|
−0.429 |
0.029
|
| MCP-1 |
381.925 |
213.455 |
738.31 |
0.303 |
280.63 (192.25, 445.88) |
408.36 (219.01, 780.87) |
1.3 |
0.531*
|
−0.118 |
0.565 |
| RANTES |
1.905 |
0.145 |
0.17 |
0.284 |
0.14 (0.00, 0.33) |
0.47 (0.13, 1.56) |
0.0 |
0.09*
|
−0.365 |
0.066 |
| lL-12 |
0.245 |
0.0075 |
0 |
0.599 |
0.00 (0.00, 0.18) |
0.00 (0.00, 0.17) |
0.1 |
0.605*
|
−0.163 |
0.427 |
| Fractalkine |
23.945 |
11.5925 |
4.145 |
0.136 |
8.70 ± 9.82 |
4.32 (2.26, 11.25) |
22.3 |
0.144*
|
0.227 |
0.266 |
| VCAM-1 |
16321.72 |
3232.4225 |
13423.49 |
0.072 |
5515.540 ±7873.21 |
2097.26 (650.16, 7734.86) |
12.2 |
0.41*
|
0.388 |
0.050 |
| lCAM-1 |
1670.56 |
669.665 |
1669.84 |
0.043
|
858.080 ±759.11 |
173.66 (18.53, 528.14) |
25.7 |
0.008
*
|
0.512 |
0.007
|
Values in bold font are statistically significant at P < 0.05.
bFGF indicates basic fibroblast growth factor; G-CSF, granulocyte colony-stimulating factor; GM-CSF, granulocyte-macrophage colony-stimulating factor; ICAM, intercellular cell adhesion molecule; IFN-γ, interferon-γ; IL, interleukin; IP-10, interferon-inducible protein-10; IPR, incontinentia pigmenti-associated retinopathy; LOD, limit of detection; MCP-1, monocyte chemoattractant protein-1; MlP-lα, macrophage inflammatory protein-lα; RANTES, regulated upon the activation of normal T cell expressed and secreted; TNF-α, tumor necrosis factor-α; VCAM, vascular cell adhesion molecule; VEGF, vascular endothelial growth factor.
*Mann-Whitney U test.
†Student t test.
‡Kruskal-Wallis igtest.
§Spearman correlation test.
Discussion
NEMO protein, product of NEMO gene, is a regulatory subunit to activate nuclear factor kappa B (NF-kB), which plays a central role in the regulation of immune functions.3 NF-kB activations resulted in the resistance to transforming growth factor (TGF)-β-mediated suppression of IL-4 signaling.4 In IPR patients, malfunctional NF-kB results in low expression of IL-4.
ICAM-1 and VCAM-1 are major adhesion molecules, expressing during intraocular inflammation and destructing blood-retinal barrier. VCAM-1 (P = 0.05) and ICAM-1 (P = 0.007) significantly increased in parallel with increasing IPR stages, which may be responsive to the pathological change of vascular permeability and leakage as common in IPR.
G-CSF has neuroprotective and antiinflammation effects during retinal ischemia.5 G-CSF decreased in IPR groups, associating with disease severity, which led to dysfunction of retinal pigment epithelium cells and photoreceptor and retinal ganglion cell death. The decline of G-CSF may be an indicator of IPR deterioration.
Inflammatory cytokines, including IL-6, IL-8, monocyte chemoattractant protein-1, macrophage inflammatory protein-1a, and IP-10, were reported to be involved in ocular angiogenic activities and inflam-mation.6,7 Retinal pigment epithelium cells produce IP-10 and IL-8 in response to stimulation of proinflammatory cytokines and T-lymphocyte secretions. The IL-6 (P = 0.08) is elevated in IPR group, and IL-8 and IP-10 are increased and linearly associated with increased IPR severity (P < 0.05). This implies T-lymphocyte infiltration and stimulation, and its mediated inflammation may be involved in the pathogenesis of IPR.
Elevation of VEGF led to pathologic changes of vascular permeability, development of neovascularization and retinal detachment.2 VEGF is linearly associated with increased IPR severity (P = 0.001), and an indicator for bad prognosis. This laid the theoretical foundation for anti-VEGF therapy. Considering elevated VEGF concentration in the stage 2 IPR without NV, the peripheral avascular zone could upregulate VEGF levels indicating need for ablations. In severe stage 2 case with dilated or tortuous vessels, anti-VEGF therapy with laser treatment may offer a better choice.
These altered cytokines are involved with activation of inflammation, angiogenesis, damage on the retina, and blood-retina breakdown in pathogenesis of IPR. The retinal vasculopathy was primary retinal changes of IPR.1 We hypothesize inflammation of the retinal vessels, possibly caused by T cell-mediated inflammation, could be the first pathogenesis followed by vascular occlusion or malformation, VEGF upregulation, vitreous hemorrhage, and tractional retinal detachment. Our results also implied inflammatory cytokines and VEGF elevations may be involved in the pathological changes of IPR. Considering this, anti-inflammation agents, anti-VEGF agents, laser ablations, or a combination may offer a potential treatment for IPR.
Conclusions
Elevated levels of VEGF, IP-10, ICAM-1 and decreased levels of IL-4 and G-CSF may be associated with the pathogenesis of IPR. T-lymphocyte infiltration and stimulation may play a role in IPR. Proinflammatory and proangiogenic cytokines may be potential therapeutic targets for IPR.
References
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Incontinentia pigmenti-associated ocular anomalies of paediatric
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