Differential Expression of Antimicrobial Peptides in Streptococcus pneumoniae Keratitis and STAT3-Dependent Expression of LL-37 by Streptococcus pneumoniae in Human Corneal Epithelial Cells

Abstract

Streptococcus pneumoniae is the leading cause of bacterial keratitis in the developing world with a growing trend of acquiring resistance against various antibiotics. In the current study, we determined the expression of different antimicrobial peptides (AMPs) in response to S. pneumoniae in patients, as well as in primary and immortalized human corneal epithelial cells. We further focused on LL-37 and determined its expression in human cornea infected with S. pneumoniae and studied the killing ability of LL-37 against S. pneumoniae. The expression of AMPs was determined by quantitative PCR and the phosphorylation of signaling proteins was evaluated by immunoblot analysis. LL-37 expression was also determined by immunofluorescence and Western blot method and the killing ability of LL-37 against S. pneumoniae was determined by colony-forming units. Differential expression of antimicrobial peptides was observed in patients with S. pneumoniae keratitis. Although S. pneumoniae induced expression of the AMPs in human corneal epithelial cells (HCEC), it did not induce AMP expression in U937, a human monocyte cell line. S. pneumoniae also caused activation of nuclear factor kappa-light-chain enhancer of activated B cells (NF-κB)and mitogen activated protein kinase (MAPK) pathways in corneal epithelial cells. LL-37 was found to be effective against both laboratory and clinical strains of S. pneumoniae. LL-37 induction by S. pneumoniae in human corneal epithelial cells was mediated by signal transducer and activator of transcription 3 (STAT3) activation, and inhibition of STAT3 activation significantly reduced LL-37 expression. Our study determines an extensive profile of AMPs expressed in the human cornea during S. pneumoniae infection, and suggests the potential of LL-37 to be developed as an alternative therapeutic intervention to fight increasing antibiotic resistance among bacteria.

Recombinant IL ‐22 promotes protection in a murine model of Aspergillus flavus keratitis and mediates host immune responses in human corneal epithelial cells

“…PASH staining of the sections was done to identify fungi and infiltrating cells. The tissue sections were deparaffinised and stained with anti‐IL‐22 (1:50; Novus Biologicals, CO) as described earlier (Sharma et al, 2019) and counterstained with DAPI (Abcam, Cambridge). Sections were also stained with rabbit IgG isotype antibody (1:50; Abcam, Cambridge, MA) as negative controls.…”

Attenuation of Pseudomonas aeruginosa infection by INP0341, a salicylidene acylhydrazide, in a murine model of keratitis

“…PAO1[ 20 ], the mutant strain PAO1 ΔpscC [ 21 ] and two clinical isolates of P. aeruginosa were used in this study. For identification of the clinical isolates, corneal ulcer materials were collected aseptically and investigated following the Institute protocol as described earlier[ 22 ]. Briefly, ulcer materials were placed on glass slides for Gram staining and were inoculated in different specific media for bacterial cultures.…”

Alternative Therapeutic Interventions: Antimicrobial Peptides and Small Molecules to Treat Microbial Keratitis

“…Corneal epithelium is multilayered in nature and acts as the first line of defense during pathogen attack. Cornea expresses several AMPs ( Figure 2 ) that have microbicidal properties or aids in wound healing, either constitutively or in response to microbial antigens ( Mc et al, 1999 ; Mcdermott, 2009 ; Sharma et al, 2018 ; Sharma et al, 2019 ). β-defensins are produced by the ocular surface and α defensins are supplied into the ocular surface fluids by resident or passing neutrophils and lacrimal ductular epithelia secretion ( Kumar et al, 2006 ).…”