Recombinant Human Proteoglycan 4 (rhPRG4) Downregulates TNFα-Stimulated NFκB Activity and FAT10 Expression in Human Corneal Epithelial Cells
Dry Eye Disease (DED) is a complex pathology affecting millions of people with significant impact on quality of life. Corneal inflammation, including via the nuclear factor kappa B (NFκB) pathway, plays a key etiological role in DED. Recombinant human proteoglycan 4 (rhPRG4) has been shown to be a c...
| Main Authors: | , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2022-10-01
|
| Series: | International Journal of Molecular Sciences |
| Subjects: | |
| Online Access: | https://www.mdpi.com/1422-0067/23/21/12711 |
| _version_ | 1797468115904233472 |
|---|---|
| author | Nikhil G. Menon Yasir Suhail Ruchi Goyal Wenqiang Du Adam P. Tanguay Gregory D. Jay Mallika Ghosh Kshitiz Tannin A. Schmidt |
| author_facet | Nikhil G. Menon Yasir Suhail Ruchi Goyal Wenqiang Du Adam P. Tanguay Gregory D. Jay Mallika Ghosh Kshitiz Tannin A. Schmidt |
| author_sort | Nikhil G. Menon |
| collection | DOAJ |
| description | Dry Eye Disease (DED) is a complex pathology affecting millions of people with significant impact on quality of life. Corneal inflammation, including via the nuclear factor kappa B (NFκB) pathway, plays a key etiological role in DED. Recombinant human proteoglycan 4 (rhPRG4) has been shown to be a clinically effective treatment for DED that has anti-inflammatory effects in corneal epithelial cells, but the underlying mechanism is still not understood. Our goal was to understand if rhPRG4 affects tumor necrosis factor α (TNFα)-stimulated inflammatory activity in corneal epithelial cells. We treated hTERT-immortalized corneal epithelial (hTCEpi) cells ± TNFα ± rhPRG4 and performed Western blotting on cell lysate and RNA sequencing. Bioinformatics analysis revealed that rhPRG4 had a significant effect on TNFα-mediated inflammation with potential effects on matricellular homeostasis. rhPRG4 reduced activation of key inflammatory pathways and decreased expression of transcripts for key inflammatory cytokines, interferons, interleukins, and transcription factors. TNFα treatment significantly increased phosphorylation and nuclear translocation of p65, and rhPRG4 significantly reduced both these effects. RNA sequencing identified human leukocyte antigen (HLA)-F adjacent transcript 10 (FAT10), a ubiquitin-like modifier protein which has not been studied in the context of DED, as a key pro-inflammatory transcript increased by TNFα and decreased by rhPRG4. These results were confirmed at the protein level. In summary, rhPRG4 is able to downregulate NFκB activity in hTCEpi cells, suggesting a potential biological mechanism by which it may act as a therapeutic for DED. |
| first_indexed | 2024-03-09T19:02:58Z |
| format | Article |
| id | doaj.art-d1ba189c137b4437a1458cbe983843c0 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-09T19:02:58Z |
| publishDate | 2022-10-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-d1ba189c137b4437a1458cbe983843c02023-11-24T04:55:56ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672022-10-0123211271110.3390/ijms232112711Recombinant Human Proteoglycan 4 (rhPRG4) Downregulates TNFα-Stimulated NFκB Activity and FAT10 Expression in Human Corneal Epithelial CellsNikhil G. Menon0Yasir Suhail1Ruchi Goyal2Wenqiang Du3Adam P. Tanguay4Gregory D. Jay5Mallika Ghosh6Kshitiz7Tannin A. Schmidt8Biomedical Engineering Department, School of Dental Medicine, UConn Health, Farmington, CT 06030, USABiomedical Engineering Department, School of Dental Medicine, UConn Health, Farmington, CT 06030, USABiomedical Engineering Department, School of Dental Medicine, UConn Health, Farmington, CT 06030, USABiomedical Engineering Department, School of Dental Medicine, UConn Health, Farmington, CT 06030, USABiomedical Engineering Department, School of Dental Medicine, UConn Health, Farmington, CT 06030, USADepartment of Emergency Medicine, Warren Alpert Medical School, Brown University, Providence, RI 02903, USADepartment of Cell Biology, School of Medicine, UConn Health, Farmington, CT 06030, USABiomedical Engineering Department, School of Dental Medicine, UConn Health, Farmington, CT 06030, USABiomedical Engineering Department, School of Dental Medicine, UConn Health, Farmington, CT 06030, USADry Eye Disease (DED) is a complex pathology affecting millions of people with significant impact on quality of life. Corneal inflammation, including via the nuclear factor kappa B (NFκB) pathway, plays a key etiological role in DED. Recombinant human proteoglycan 4 (rhPRG4) has been shown to be a clinically effective treatment for DED that has anti-inflammatory effects in corneal epithelial cells, but the underlying mechanism is still not understood. Our goal was to understand if rhPRG4 affects tumor necrosis factor α (TNFα)-stimulated inflammatory activity in corneal epithelial cells. We treated hTERT-immortalized corneal epithelial (hTCEpi) cells ± TNFα ± rhPRG4 and performed Western blotting on cell lysate and RNA sequencing. Bioinformatics analysis revealed that rhPRG4 had a significant effect on TNFα-mediated inflammation with potential effects on matricellular homeostasis. rhPRG4 reduced activation of key inflammatory pathways and decreased expression of transcripts for key inflammatory cytokines, interferons, interleukins, and transcription factors. TNFα treatment significantly increased phosphorylation and nuclear translocation of p65, and rhPRG4 significantly reduced both these effects. RNA sequencing identified human leukocyte antigen (HLA)-F adjacent transcript 10 (FAT10), a ubiquitin-like modifier protein which has not been studied in the context of DED, as a key pro-inflammatory transcript increased by TNFα and decreased by rhPRG4. These results were confirmed at the protein level. In summary, rhPRG4 is able to downregulate NFκB activity in hTCEpi cells, suggesting a potential biological mechanism by which it may act as a therapeutic for DED.https://www.mdpi.com/1422-0067/23/21/12711PRG4proteoglycan 4lubricindry eyecorneal epithelial cells |
| spellingShingle | Nikhil G. Menon Yasir Suhail Ruchi Goyal Wenqiang Du Adam P. Tanguay Gregory D. Jay Mallika Ghosh Kshitiz Tannin A. Schmidt Recombinant Human Proteoglycan 4 (rhPRG4) Downregulates TNFα-Stimulated NFκB Activity and FAT10 Expression in Human Corneal Epithelial Cells International Journal of Molecular Sciences PRG4 proteoglycan 4 lubricin dry eye corneal epithelial cells |
| title | Recombinant Human Proteoglycan 4 (rhPRG4) Downregulates TNFα-Stimulated NFκB Activity and FAT10 Expression in Human Corneal Epithelial Cells |
| title_full | Recombinant Human Proteoglycan 4 (rhPRG4) Downregulates TNFα-Stimulated NFκB Activity and FAT10 Expression in Human Corneal Epithelial Cells |
| title_fullStr | Recombinant Human Proteoglycan 4 (rhPRG4) Downregulates TNFα-Stimulated NFκB Activity and FAT10 Expression in Human Corneal Epithelial Cells |
| title_full_unstemmed | Recombinant Human Proteoglycan 4 (rhPRG4) Downregulates TNFα-Stimulated NFκB Activity and FAT10 Expression in Human Corneal Epithelial Cells |
| title_short | Recombinant Human Proteoglycan 4 (rhPRG4) Downregulates TNFα-Stimulated NFκB Activity and FAT10 Expression in Human Corneal Epithelial Cells |
| title_sort | recombinant human proteoglycan 4 rhprg4 downregulates tnfα stimulated nfκb activity and fat10 expression in human corneal epithelial cells |
| topic | PRG4 proteoglycan 4 lubricin dry eye corneal epithelial cells |
| url | https://www.mdpi.com/1422-0067/23/21/12711 |
| work_keys_str_mv | AT nikhilgmenon recombinanthumanproteoglycan4rhprg4downregulatestnfastimulatednfkbactivityandfat10expressioninhumancornealepithelialcells AT yasirsuhail recombinanthumanproteoglycan4rhprg4downregulatestnfastimulatednfkbactivityandfat10expressioninhumancornealepithelialcells AT ruchigoyal recombinanthumanproteoglycan4rhprg4downregulatestnfastimulatednfkbactivityandfat10expressioninhumancornealepithelialcells AT wenqiangdu recombinanthumanproteoglycan4rhprg4downregulatestnfastimulatednfkbactivityandfat10expressioninhumancornealepithelialcells AT adamptanguay recombinanthumanproteoglycan4rhprg4downregulatestnfastimulatednfkbactivityandfat10expressioninhumancornealepithelialcells AT gregorydjay recombinanthumanproteoglycan4rhprg4downregulatestnfastimulatednfkbactivityandfat10expressioninhumancornealepithelialcells AT mallikaghosh recombinanthumanproteoglycan4rhprg4downregulatestnfastimulatednfkbactivityandfat10expressioninhumancornealepithelialcells AT kshitiz recombinanthumanproteoglycan4rhprg4downregulatestnfastimulatednfkbactivityandfat10expressioninhumancornealepithelialcells AT tanninaschmidt recombinanthumanproteoglycan4rhprg4downregulatestnfastimulatednfkbactivityandfat10expressioninhumancornealepithelialcells |