Sjögren-Like Lacrimal Keratoconjunctivitis in Germ-Free Mice
Abstract
:1. Introduction
2. Results
2.1. Spontaneous Sjögren-Like Lacrimal Keratoconjunctivitis
2.2. Sex Differences in Ocular Surface Manifestations
2.3. Changes in the Ocular Environment in Germ-Free Mice Predispose to Th-1 Cells
2.4. CD4+ T Cells from Germ-Free Mice Induce Sjögren-Like Lacrimal Keratoconjunctivitis
2.5. Fecal Microbiota Transplant Reversed Lacrimal Keratoconjunctivitis in Germ-Free Mice
3. Discussion
4. Methods
4.1. Animals
4.2. Corneal Permeability
4.3. Histology and PAS Staining
4.4. Measurement of Lacrimal Gland Infiltration
4.5. Flow Cytometry Analysis of Infiltrating Cells
4.6. Tear Washings and EGF Enzyme-Linked Immunosorbent Assay
4.7. Caspase-3 Activation Fluorometric Assays
4.8. RNA Isolation and Real-Time PCR
4.9. Isolation of Murine CD4+ T Cells and Adoptive Transfer
4.10. Standard Desiccating Stress Model
4.11. Antibiotic Treatment and Desiccating Stress
4.12. Fecal Microbiota Transplant
4.13. DNA Extraction from Mouse Fecal Samples, 16S rRNA Gene Amplication, and Sequencing
4.14. Microbial Community Analysis
4.15. Statistical Analysis
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Parameters | Sex | CON | GF | p Value | ||||
---|---|---|---|---|---|---|---|---|
F | M | F | M | CON vs. GF | F vs. M (GF) | |||
Ocular surface | OGD intensity (gray levels) | 241 ± 75.6 | 228.2 ± 71.5 | 344 ± 118 | 224 ± 147 | ↑, p < 0.001 (F) | F > M (p < 0.01) | |
Goblet cell (cells/mm) | 56.5 ± 8.8 | 53.2 ± 09.5 | 46.2 ± 7.4 | 43.0 ± 7.0 | ↓, p < 0.001 (F,M) | |||
EGF (pg/mL) | 4282 ± 1288 | 6667 ± 1787 | 2250 ± 750 | 2257 ± 167 | ↓, p < 0.05 (F,M) | |||
LG | Total LG infiltration (%) * | 3.5 ± 3.0 | 1.38 ± 0.4 | 6.36 ± 1.7 | 4.07 ± 2.2 | ↑, p < 0.05 (F,M) | F > M (p < 0.05) | |
CD4+ T cells (%) (flow cytometry) | 6.0 ± 2.8 | 7.5 ± 3.7 | 10.9 ± 4.2 | 11.8 ± 4.8 | ↑, p < 0.05 (F,M) | |||
CD8+ T cells (%) (flow cytometry) | 4.63 ± 1.2 | 3.1 ± 0.9 | 10.1 ± 0.6 | 7.8 ± 2.3 | ↑, p < 0.001 (F,M) | |||
PCR (fold) | IFN-γ | 1.00 ± 0.25 | 0.96 ± 0.11 | 2.84 ± 0.57 | 1.31 ± 0.23 | ↑, p < 0.001 (F) | F > M (p < 0.05) | |
MHC II | 1.00 ± 0.09 | 0.92 ± 0.16 | 1.57 ± 0.10 | 0.89 ± 0.30 | ↑, p < 0.001 (F) | F > M (p < 0.05) | ||
Caspase 3 | 1.00 ± 0.04 | 0.57 ± 0.07 | 1.51 ± 0.10 | 1.00 ± 0.08 | ↑, p < 0.001 (F) | F > M (p < 0.001) | ||
↑, p < 0.01 (M) | ||||||||
IL-1β | 1.00 ± 0.21 | 0.93 ± 0.03 | 0.76 ± 0.18 | 0.61 ± 0.14 | ↓, p < 0.05 (F,M) | |||
IL-12 | 1.00 ± 0.08 | 1.00 ± 0.07 | 2.51 ± 0.7 | 2.28 ± 0.04 | ↑, p < 0.05 (F,M) | F > M (p < 0.01) | ||
TNF-α | 1.00 ± 0.04 | 0.29 ± 0.21 | 0.80 ± 0.10 | 0.55 ± 0.12 | Ø | Ø | ||
Caspase 3 activity (OD) | 2355 ± 871 | 2754 ± 687 | 7499 ± 693 | 4110 ± 1201 | ↑, p < 0.001 (F) | F > M (p < 0.001) | ||
↑, p < 0.01 (M) |
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Wang, C.; Zaheer, M.; Bian, F.; Quach, D.; Swennes, A.G.; Britton, R.A.; Pflugfelder, S.C.; De Paiva, C.S. Sjögren-Like Lacrimal Keratoconjunctivitis in Germ-Free Mice. Int. J. Mol. Sci. 2018, 19, 565. https://doi.org/10.3390/ijms19020565
Wang C, Zaheer M, Bian F, Quach D, Swennes AG, Britton RA, Pflugfelder SC, De Paiva CS. Sjögren-Like Lacrimal Keratoconjunctivitis in Germ-Free Mice. International Journal of Molecular Sciences. 2018; 19(2):565. https://doi.org/10.3390/ijms19020565
Chicago/Turabian StyleWang, Changjun, Mahira Zaheer, Fang Bian, Darin Quach, Alton G. Swennes, Robert A. Britton, Stephen C. Pflugfelder, and Cintia S. De Paiva. 2018. "Sjögren-Like Lacrimal Keratoconjunctivitis in Germ-Free Mice" International Journal of Molecular Sciences 19, no. 2: 565. https://doi.org/10.3390/ijms19020565