Purpose-Human ocular surface epithelia express four antimicrobial peptides (APs): β-defensin (hBD) 1-3 and LL-37. Here the expression of additional APs (hBD 4-6, HE2β1; histatin-1, -3; liver expressed antimicrobial peptide-1, -2; macrophage inflammatory protein (MIP)-3α, and thymosin (T)β-4) was sought and activity against common ocular pathogens studied.Methods-AP expression was determined in human corneal and conjunctival epithelial cells (HCEC, HCjEC) by RT-PCR and in corneal sections by immunostaining. Antimicrobial assays were performed to assess peptide (hBD 1-3, LL-37, MIP-3α, and Tβ4) activity against Pseudomonas aeruginosa (PA), Staphylococcus aureus (SA), and Staphylococcus epidermidis (SE) in the presence of NaCl or tears.Results-HCEC and HCjEC expressed MIP-3α and Tβ4. hBD 1-3, MIP-3α, and Tβ4 showed activity against PA. hBD-3 had potent activity against SA and SE, whereas hBD-2, MIP-3α and Tβ4 had moderate activity and hBD-1 had none. NaCl markedly attenuated, and tears almost completely inhibited the activity of hBD 1-2 and Tβ4, but not that of hBD-3. Conclusions-The ocular surface epithelia additionally express MIP-3α and Tβ4 which have moderate antimicrobial activity. The current data support a role for hBD-3 as an antimicrobial peptide in vivo, but call in to question the effectiveness of some other APs. However, further study is required to conclusively elucidate the physiological role of each AP.
Purpose. Human β-defensin-2 (hBD-2) and cathelicidin LL-37 are salt-sensitive cationic antimicrobial peptides expressed by ocular surface epithelia. The goal of this study was to investigate the effect of preservative-free artificial tears on hBD-2 and LL-37 antimicrobial activity against Pseudomonas aeruginosa.Methods. P. aeruginosa was incubated with hBD-2 or LL-37 in the absence or presence (70% vol/ vol) of different preservative-free artificial tears-Visine Tears (300 mOsm/kg), Tears Naturale Free (261 mOsm/kg), TheraTears (185 mOsm/kg), and Refresh Plus (325 mOsm/kg)-for 2 hours at 37°C. In some experiments, P. aeruginosa was incubated with hBD-2 or LL-37 and Visine Tears or Tears Naturale Free with or without carboxymethylcellulose (0.5% vol/vol final concentration). Plates were inoculated with samples of each reaction mixture and then incubated for 24 hours at 37°C .Results. Visine Tears and Tears Naturale Free had little or no effect on the antimicrobial activity of 100 μg/mL hBD-2 or LL-37. In the presence of Refresh Plus and TheraTears, the activity of 100 μg/mL hBD-2 or LL-37 was reduced by 90% to 100%. Carboxymethylcellulose, at a concentration comparable to that present in Refresh Plus, reduced the effectiveness of hBD-2 or LL-37 by 40% to 90% in the presence of Tears Naturale Free and Visine Tears. Conclusion.Human β-defensin-2 and cathelicidin LL-37 inhibit the growth of P. aeruginosa in vitro, but this activity is markedly reduced in the presence of Refresh Plus and TheraTears. These results suggest that carboxymethylcellulose-containing artificial tears may reduce the activity of the endogenously produced antimicrobial peptides. KeywordsCarboxymethylcellulose; Cationic antimicrobial peptides; Human β-defensin-2; Human cathelicidin LL-37; Preservative-free artificial tears; Pseudomonas aeruginosa Secretion of small cationic antimicrobial peptides by epithelia is an important component of the innate immune response. 1 These peptides, which include β-defensins and cathelicidins, insert themselves into microbial cell membranes and, by pore formation or electrostatic disruption, cause release of intracellular contents leading to death of the organism. 2 By regulating mammalian cell functions, such as chemotaxis and proliferation, defensins and
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