Journal: Investigative Ophthalmology & Visual Science

Article Title: Fatty Acid Desaturase 1 Knockdown Promotes Wound Healing and Functional Recovery of the Corneal Epithelium in Diabetes

doi: 10.1167/iovs.66.6.6

Figure Lengend Snippet: FADS1 is highly expressed after diabetic corneal epithelial injury, and FADS1 is localized to the ER in corneal epithelial cells. ( A ) Venn diagram illustrating the comparison of metabolism-related DEGs between normal and diabetic rats at 42 h after corneal injury vs. uninjured states, showing the number of unique and overlapping genes. A fold change of >1.5 and P < 0.05. ( B ) Representative immunohistochemical images of FADS1 in uninjured and injured corneal tissues from DM mice. ( C ) Western blotting–based analysis of the subcellular localization of FADS1. TOMM40 and VDAC were used as markers for the mitochondrial outer membrane, and GRP75 and HSP60 were used as markers for the mitochondrial matrix, SDHA was used as a marker for the mitochondrial inner membrane, and Lamin A/C was used as a marker for the nucleus. ( D ) Immunofluorescence images showing the colocalization of FADS1 ( green ) and the ER marker RTN4 ( red ) in HCECs.

Article Snippet: This study used an immortalized human corneal epithelial cell line (SV40-HCEC, Procell, CL-0743) and primary mouse corneal epithelial cells (MCECs, Procell, CP-M120).

Techniques: Comparison, Immunohistochemical staining, Western Blot, Membrane, Marker, Immunofluorescence

Journal: Investigative Ophthalmology & Visual Science

Article Title: Fatty Acid Desaturase 1 Knockdown Promotes Wound Healing and Functional Recovery of the Corneal Epithelium in Diabetes

doi: 10.1167/iovs.66.6.6

Figure Lengend Snippet: FADS1 KD promotes corneal epithelial cell migration under HG conditions. ( A–C ) qRT-PCR and Western blotting analyses of FADS1 in HCECs transfected with NC siRNA or FADS1 siRNA ( n = 3). ( D , E ) Representative scratch assay images and statistical analysis of wound healing rates for HCECs following transfection with NC siRNA or FADS1 siRNA ( n = 9). ( F , G ) Representative transwell migration assay images for HCECs transfected with NC siRNA or FADS1 siRNA, comparing the relative number of migrated cells ( n = 4). ( H , I ) Western blotting analysis of FADS1 in HCECs treated with D5D-IN-326 (2 µM, 48 h), with DMSO as the control ( n = 3). ( J , K ) Representative scratch assay images and statistical analysis of wound healing rates for HCECs treated with D5D-IN-326 (2 µM, 48 h), with DMSO as the control ( n = 9). All values are mean ± SEM. ** P < 0.01; *** P < 0.001; **** P < 0.0001.

Article Snippet: This study used an immortalized human corneal epithelial cell line (SV40-HCEC, Procell, CL-0743) and primary mouse corneal epithelial cells (MCECs, Procell, CP-M120).

Techniques: Migration, Quantitative RT-PCR, Western Blot, Transfection, Wound Healing Assay, Transwell Migration Assay, Control

Journal: Investigative Ophthalmology & Visual Science

Article Title: Fatty Acid Desaturase 1 Knockdown Promotes Wound Healing and Functional Recovery of the Corneal Epithelium in Diabetes

doi: 10.1167/iovs.66.6.6

Figure Lengend Snippet: FADS1 KD promotes HG-induced corneal epithelial cell migration through autophagosome accumulation. ( A – D ) Western blotting analysis of P62 and LC3B in HCECs transfected with NC siRNA or FADS1 siRNA ( n = 3). ( E ) The number of autophagosomes in HECEs was detected by TEM after transfection with NC siRNA or FADS1 siRNA ( n = 3). ( F , G ) Western blotting analysis of LC3B in HCECs treated with 3-MA (5 mM, 6 h) after transfection with NC siRNA or FADS1 siRNA ( n = 3). ( H , I ) Representative scratch assay images and statistical analysis of wound healing rates for HCECs transfected with NC siRNA or FADS1 siRNA, followed by treatment with 3-MA (5 mM, 6 h) ( n = 9). All values are mean ± SEM. * P < 0.05; ** P < 0.01; ns, nonsignificant.

Article Snippet: This study used an immortalized human corneal epithelial cell line (SV40-HCEC, Procell, CL-0743) and primary mouse corneal epithelial cells (MCECs, Procell, CP-M120).

Techniques: Migration, Western Blot, Transfection, Wound Healing Assay

Journal: Investigative Ophthalmology & Visual Science

Article Title: Fatty Acid Desaturase 1 Knockdown Promotes Wound Healing and Functional Recovery of the Corneal Epithelium in Diabetes

doi: 10.1167/iovs.66.6.6

Figure Lengend Snippet: Hyperglycemia impairs corneal epithelial wound healing in diabetic mice. ( A , B ) Blood glucose and body weight of control (Ctrl) and DM mice ( n = 6). ( C ) After scraping of the corneal epithelium in Ctrl and DM mice, sodium fluorescein staining was performed at D0, D1, and D2. ( D ) Histogram illustrating the percentage of the defect area relative to the original wound ( n = 6). All values are mean ± SEM. ** P < 0.01; *** P < 0.001; **** P < 0.0001.

Article Snippet: This study used an immortalized human corneal epithelial cell line (SV40-HCEC, Procell, CL-0743) and primary mouse corneal epithelial cells (MCECs, Procell, CP-M120).

Techniques: Control, Staining

Journal: Investigative Ophthalmology & Visual Science

Article Title: Fatty Acid Desaturase 1 Knockdown Promotes Wound Healing and Functional Recovery of the Corneal Epithelium in Diabetes

doi: 10.1167/iovs.66.6.6

Figure Lengend Snippet: FADS1-specific siRNA treatment promotes corneal epithelial wound healing in diabetic mice. ( A ) Schematic illustrating the establishment and treatment of a diabetic corneal injury model: subconjunctival injection of NC siRNA or FADS1 siRNA was performed at 0 h and 24 h after corneal epithelial debridement, respectively. ( B ) Representative immunohistochemical images of FADS1 in corneal tissue after the injection of NC siRNA or FADS1 siRNA in DM mice with corneal injury ( n = 3). ( C ) After corneal injury in DM mice, NC siRNA or FADS1 siRNA was injected, and sodium fluorescein staining was performed at days (D)0, D1, and D2. ( D ) Histogram illustrating the percentage of the defect area relative to the original wound ( n = 6). ( E ) Representative optical coherence tomography images of diabetic corneal injury following the injection of NC siRNA or FADS1 siRNA compared with normal corneas. ( F ) Central corneal thickness was analyzed in three groups ( n = 3). ( G ) Representative hematoxylin and eosin (HE)–stained images of diabetic corneal injury after the injection of NC siRNA or FADS1 siRNA compared with normal corneas ( n = 3). All values are mean ± SEM. * P < 0.05; ** P < 0.01.

Article Snippet: This study used an immortalized human corneal epithelial cell line (SV40-HCEC, Procell, CL-0743) and primary mouse corneal epithelial cells (MCECs, Procell, CP-M120).

Techniques: Injection, Immunohistochemical staining, Staining, Tomography

Journal: Investigative Ophthalmology & Visual Science

Article Title: Fatty Acid Desaturase 1 Knockdown Promotes Wound Healing and Functional Recovery of the Corneal Epithelium in Diabetes

doi: 10.1167/iovs.66.6.6

Figure Lengend Snippet: FADS1-specific siRNA treatment promotes diabetic corneal epithelial homeostasis and functional recovery. ( A , B ) Representative images of whole cornea immunofluorescence staining revealing the expression and distribution of PAX6 and E-cadherin in each group. ( C , D ) The relative fluorescence intensity of PAX6 and E-cadherin in each group was analyzed based on the immunofluorescence staining results ( n = 3). All values are mean ± SEM. ** P < 0.01.

Article Snippet: This study used an immortalized human corneal epithelial cell line (SV40-HCEC, Procell, CL-0743) and primary mouse corneal epithelial cells (MCECs, Procell, CP-M120).

Techniques: Functional Assay, Immunofluorescence, Staining, Expressing, Fluorescence