Circular RNA expression profile identifies potential circulating biomarkers for keratoconus.

Early diagnosis is important for improving the outcomes of keratoconus (KC). Stable expression and a closed-loop structure of circular RNAs (circRNAs) make them ideal for the diagnosis and treatment of diseases. However, the expression pattern and potential function of circRNAs in KC is not studied yet. Hence, this study explored the circRNA expression profile of KC corneas through transcriptome sequencing and circRNA expression profile analysis. The diagnostic potential of blood circRNAs for KC was explored by analysing the circRNAs' expression levels of fifty paired blood samples from patients with KC and normal controls. The results showed that 107 significantly upregulated and 145 significantly downregulated circRNAs (|fold change| ≥ 2.0, p-value <0.05) were identified in KC tissues. Eight top differently expressed circRNAs were further validated in more cornea samples. Among them, five circRNAs expressed in peripheral blood, and four circRNAs (circ_0006156, circ_0006117, circ_0000284 and circ_0001801) showed significant downregulation in KC patients' peripheral blood too. The blood circ_0000284 expression levels of early, moderate, and advanced KC patients both were significantly lower than the controls. The blood circ_0006117 expression levels present a positive correlation with corrected distance visual acuity values, and a negative correlation with back elevation values of KC eyes. Notably, the expression levels of these circRNAs distinguished KC patients from their healthy counterparts, with the area under the curve (AUC) of circ_0000284, circ_0001801, and circ_0006117 being 0.7306, 0.6871 and 0.6701, respectively. Further, the AUC value for five circRNAs under the logistic regression model was 0.8203, indicating that they can function as effective biomarkers for the KC diagnostics. In conclusion, the expression of circRNAs showed a relationship with KC, with four significantly differentially expressed circRNAs demonstrating potential as biomarkers for the disease.

Bacterial Keratitis Following Small Incision Lenticule Extraction.

Purpose: To describe the development of bacterial keratitis after small incision lenticule extraction in 5 patients and to explore its appropriate therapies. Methods: We retrospectively summarized the clinical treatments of five patients with postoperative bacterial infection after small incision lenticule extraction, who were referred to our hospital from 2019 to 2021. Results: Five male patients had undergone bilateral SMILE in the local hospital due to myopia aged from 18 to 26 years. The onset of keratitis during 1-3 days postoperatively and four of them were severe infection (2 bilateral, 2 unilateral). In five cases, 1 patient (1 eye) who was infected mild keratitis after SMILE was treated with only topical antibiotics; the others who respond poorly to topical antibiotics require surgical treatment, which 1 patient (1 eye) infected necrotic mass of the corneal cap was scraped and irrigated with antibiotic, and 3 patients (5 eyes) were treated by converting the cap to flap, curetting the necrotic tissue and irrigating with the antibiotic solution. In all patients, the duration from onset to resolution was 1-5 weeks. The final uncorrected visual acuity was above 20/32. Conclusion: Owing to the upward popularity of refractive surgery, the incidence of keratitis after SMILE should not be ignored. Early diagnosis and timely treatment of post-SMILE keratitis are essential. For severe keratitis that fails to respond to topical antibiotics, the corneal cap should be opened as a flap.

The involvement of proline-rich protein Mus musculus predicted gene 4736 in ocular surface functions.

AIM: To research the two homologous predicted proline-rich protein genes, Mus musculus predicted gene 4736 (MP4) and proline-rich protein BstNI subfamily 1 (Prb1) which were significantly upregulated in cultured corneal organs when encountering fungal pathogen preparations. This study was to confirm the expression and potential functions of these two genes in ocular surface. METHODS: A Pseudomonas aeruginosa keratitis model was established in Balb/c mice. One day post infection, mRNA level of MP4 was measured using real-time polymerase chain reaction (PCR), and MP4 protein detected by immunohistochemistry (IHC) or Western blot using a customized polyclonal anti-MP4 antibody preparation. Lacrimal glands from normal mice were also subjected to IHC staining for MP4. An online bioinformatics program, BioGPS, was utilized to screen public data to determine other potential locations of MP4. RESULTS: One day after keratitis induction, MP4 was upregulated in the corneas at both mRNA level as measured using real-time PCR and protein levels as measured using Western blot and IHC. BioGPS analysis of public data suggested that the MP4 gene was most abundantly expressed in the lacrimal glands, and IHC revealed that normal murine lacrimal glands were positive for MP4 staining. CONCLUSION: MP4 and Prb1 are closely related with the physiology and pathological processes of the ocular surface. Considering the significance of ocular surface abnormalities like dry eye, we propose that MP4 and Prb1 contribute to homeostasis of ocular surface, and deserve more extensive functional and disease correlation studies.

Serum amyloid A and pairing formyl peptide receptor 2 are expressed in corneas and involved in inflammation-mediated neovascularization.

AIM: To solidify the involvement of Saa-related pathway in corneal neovascularization (CorNV). The pathogenesis of inflammatory CorNV is not fully understood yet, and our previous study implicated that serum amyloid A (Saa) 1 (Saa1) and Saa3 were among the genes up-regulated upon CorNV induction in mice. METHODS: Microarray data obtained during our profiling project on CorNV were analyzed for the genes encoding the four SAA family members (Saa1-4), six reported SAA receptors (formyl peptide receptor 2, Tlr2, Tlr4, Cd36, Scarb1, P2rx7) and seven matrix metallopeptidases (Mmp) 1a, 1b, 2, 3, 9, 10, 13 reportedly to be expressed upon SAA pathway activation. The baseline expression or changes of interested genes were further confirmed in animals with CorNV using molecular or histological methods. CorNV was induced in Balb/c and C57BL/6 mice by placing either three interrupted 10-0 sutures or a 2 mm filter paper soaked with sodium hydroxide in the central area of the cornea. At desired time points, the corneas were harvested for histology examination or for extraction of mRNA and protein. The mRNA levels of Saa1, Saa3, Fpr2, Mmp2 and Mmp3 in corneas were detected using quantitative reverse transcription-PCR, and SAA3 protein in tissues detected using immunohistochemistry or western blotting. RESULTS: Microarray data analysis revealed that Saa1, Saa3, Fpr2, Mmp2, Mmp3 messengers were readily detected in normal corneas and significantly up-regulated upon CorNV induction. The changes of these five genes were confirmed with real-time PCR assay. On the contrary, other SAA members (Saa2, Saa4), other SAA receptors (Tlr2, Tlr4, Cd36, P2rx7, etc), or other Mmps (Mmp1a, Mmp1b, Mmp9, Mmp10, Mmp13) did not show consistent changes. Immunohistochemistry study and western blotting further confirmed the expression of SAA3 products in normal corneas as well as their up-regulation in corneas with CorNV. CONCLUSION: SAA-FPR2 pathway composing genes were expressed in normal murine corneas and, upon inflammatory stimuli challenge to the corneas, their expressions were up-regulated, suggesting their roles in pathogenesis of CorNV. The potential usefulness of SAA-FPR2 targets in future management of CorNV-related diseases deserves investigation.