Skip to main content

Advertisement

Log in

Development of a novel ex vivo model of corneal fungal adherence

  • Cornea
  • Published:
Graefe's Archive for Clinical and Experimental Ophthalmology Aims and scope Submit manuscript

Abstract

Purpose

To construct a suitable ex vivo model for the research of molecular mechanisms and the pharmacological screening of fungal adherence on the corneal surface.

Materials and methods

Mouse eyes were divided into three groups as follows: a control group with normal corneal epithelium, a group with corneal epithelium that was needle-scarified, and a group with corneal epithelium that was completely debrided. All 96 corneas were placed in organ culture and inoculated with 5 μl spore suspensions of Candida albicans at 109, 108, or 107 colony-forming units (CFU)/ml and incubated for 0, 30, 60, or 120 min. The corneas were homogenated and diluted for quantification by counting the CFU. The effects of amphotericin B or chondroitin sulfate on the adherence of the fungal spores were evaluated with the ex vivo organ culture model and were also compared with the human corneal epithelium monolayer model in vitro.

Results

Compared with the normal corneas with intact epithelium, the corneas with scarified and debrided epithelium adhered more spores for above two and four folds. The spore adhesion on the corneal surface was in an inoculation concentration- and incubation time-dependent manner. Moreover, both amphotericin B and chondroitin sulfate inhibited the adhesion of C. albicans spores on the corneal surface, but the inhibitory rates were different between the ex vivo corneal organ culture model and the in vitro corneal epithelium monolayer model.

Conclusions

The corneal organ culture was a suitable ex vivo model for the research of fungal adhesion mechanisms and drug screening.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  1. Gopinathan U, Garg P, Fernandes M, Sharma S, Athmanathan S, Rao GN (2002) The epidemiological features and laboratory results of fungal keratitis: a 10-year review at a referral eye care center in South India. Cornea 21:555–559

    Article  PubMed  Google Scholar 

  2. Xie L, Zhai H, Zhao J, Sun S, Shi W, Dong X (2008) Antifungal susceptibility for common pathogens of fungal keratitis in Shandong Province, China. Am J Ophthalmol 146:260–265

    Article  PubMed  CAS  Google Scholar 

  3. Rosa RH Jr, Miller D, Alfonso EC (1994) The changing spectrum of fungal keratitis in south Florida. Ophthalmology 101:1005–1013

    PubMed  Google Scholar 

  4. Leck AK, Thomas PA, Hagan M, Kaliamurthy J, Ackuaku E, John M, Newman MJ, Codjoe FS, Opintan JA, Kalavathy CM, Essuman V, Jesudasan CA, Johnson GJ (2002) Aetiology of suppurative corneal ulcers in Ghana and south India, and epidemiology of fungal keratitis. Br J Ophthalmol 86:1211–1215

    Article  PubMed  CAS  Google Scholar 

  5. Bharathi MJ, Ramakrishnan R, Vasu S, Meenakshi R, Palaniappan R (2003) Epidemiological characteristics and laboratory diagnosis of fungal keratitis. A three-year study. Indian J Ophthalmol 51:315–321

    PubMed  Google Scholar 

  6. Xie L, Zhong W, Shi W, Sun S (2006) Spectrum of fungal keratitis in north China. Ophthalmology 113:1943–1948

    Article  PubMed  Google Scholar 

  7. Mendes-Giannini MJ, Soares CP, da Silva JL, Andreotti PF (2005) Interaction of pathogenic fungi with host cells: molecular and cellular approaches. FEMS Immunol Med Microbiol 45:383–394

    Article  PubMed  CAS  Google Scholar 

  8. Tronchin G, Pihet M, Lopes-Bezerra LM, Bouchara JP (2008) Adherence mechanisms in human pathogenic fungi. Med Mycol 46:749–772

    Article  PubMed  CAS  Google Scholar 

  9. Srinoulprasert Y, Kongtawelert P, Chaiyaroj SC (2006) Chondroitin sulfate B and heparin mediate adhesion of Penicillium marneffei conidia to host extracellular matrices. Microb Pathog 40:126–132

    Article  PubMed  CAS  Google Scholar 

  10. Behling-Kelly E, Vonderheid H, Kim KS, Corbeil LB, Czuprynski CJ (2006) Roles of cellular activation and sulfated glycans in Haemophilus somnus adherence to bovine brain microvascular endothelial cells. Infect Immun 74:5311–5318

    Article  PubMed  CAS  Google Scholar 

  11. Bromley IM, Donaldson K (1996) Binding of Aspergillus fumigatus spores to lung epithelial cells and basement membrane proteins: relevance to the asthmatic lung. Thorax 51:1203–1209

    Article  PubMed  CAS  Google Scholar 

  12. Dabo SM, Confer AW, Hartson SD (2005) Adherence of Pasteurella multocida to fibronectin. Vet Microbiol 110:265–275

    Article  PubMed  CAS  Google Scholar 

  13. Gil ML, Penalver MC, Lopez-Ribot JL, O'Connor JE, Martinez JP (1996) Binding of extracellular matrix proteins to Aspergillus fumigatus conidia. Infect Immun 64:5239–5247

    PubMed  CAS  Google Scholar 

  14. Gonzalez A, Caro E, Munoz C, Restrepo A, Hamilton AJ, Cano LE (2008) Paracoccidioides brasiliensis conidia recognize fibronectin and fibrinogen which subsequently participate in adherence to human type II alveolar cells: involvement of a specific adhesin. Microb Pathog 44:389–401

    Article  PubMed  CAS  Google Scholar 

  15. Hollmer C, Essmann M, Ault K, Larsen B (2006) Adherence and blocking of Candida albicans to cultured vaginal epithelial cells: treatments to decrease adherence. Infect Dis Obstet Gynecol 2006:98218

    PubMed  Google Scholar 

  16. Hostetter MK (1994) Adhesins and ligands involved in the interaction of Candida spp. with epithelial and endothelial surfaces. Clin Microbiol Rev 7:29–42

    PubMed  CAS  Google Scholar 

  17. Rao NA, Riggio DW, Delmage JM, Calandra AJ, Evans S, Lewis W (1985) Adherence of Candida to corneal surface. Curr Eye Res 4:851–856

    Article  PubMed  CAS  Google Scholar 

  18. Dong X, Shi W, Zeng Q, Xie L (2005) Roles of adherence and matrix metalloproteinases in growth patterns of fungal pathogens in cornea. Curr Eye Res 30:613–620

    Article  PubMed  CAS  Google Scholar 

  19. Zhang H, Chen H, Niu J, Wang Y, Xie L (2009) The role of adaptive immunity in the pathogenesis of Candida Albicans keratitis. Invest Ophth Vis Sci 50:2653–2659

    Article  Google Scholar 

  20. Wang S, Wu Z, Sorenson CM, Lawler J, Sheibani N (2003) Thrombospondin-1-deficient mice exhibit increased vascular density during retinal vascular development and are less sensitive to hyperoxia-mediated vessel obliteration. Dev Dyn 228:630–642

    Article  PubMed  CAS  Google Scholar 

  21. Castro-Combs J, Noguera G, Cano M, Yew M, Gehlbach PL, Palmer J, Behrens A (2008) Corneal wound healing is modulated by topical application of amniotic fluid in an ex vivo organ culture model. Exp Eye Res 87:56–63

    Article  PubMed  CAS  Google Scholar 

  22. Singh A, Hazlett LD, Berk RS (1990) Characterization of Pseudomonas aeruginosa adherence to mouse corneas in organ culture. Infect Immun 58:1301–1307

    PubMed  CAS  Google Scholar 

  23. Collin HB, Anderson JA, Richard NR, Binder PS (1995) In vitro model for corneal wound healing; organ-cultured human corneas. Curr Eye Res 14:331–339

    Article  PubMed  CAS  Google Scholar 

  24. Giles SS, Dagenais TR, Botts MR, Keller NP, Hull CM (2009) Elucidating the pathogenesis of spores from the human fungal pathogen Cryptococcus neoformans. Infect Immun 77:3491–3500

    Article  PubMed  CAS  Google Scholar 

  25. Kim SY, Lim JA, Choi JS, Choi EC, Joo CK (2007) Comparison of antibiotic effect and corneal epithelial toxicity of levofloxacin and moxifloxacin in vitro. Cornea 26:720–725

    Article  PubMed  Google Scholar 

  26. Muthusamy A, Achur RN, Valiyaveettil M, Botti JJ, Taylor DW, Leke RF, Gowda DC (2007) Chondroitin sulfate proteoglycan but not hyaluronic acid is the receptor for the adherence of Plasmodium falciparum-infected erythrocytes in human placenta, and infected red blood cell adherence up-regulates the receptor expression. Am J Pathol 170:1989–2000

    Article  PubMed  CAS  Google Scholar 

  27. Tonnaer EL, Hafmans TG, Van Kuppevelt TH, Sanders EA, Verweij PE, Curfs JH (2006) Involvement of glycosaminoglycans in the attachment of pneumococci to nasopharyngeal epithelial cells. Microbes Infect 8:316–322

    Article  PubMed  CAS  Google Scholar 

  28. Ricciuto J, Heimer SR, Gilmore MS, Argueso P (2008) Cell surface O-glycans limit Staphylococcus aureus adherence to corneal epithelial cells. Infect Immun 76:5215–5220

    Article  PubMed  CAS  Google Scholar 

  29. Henriques M, Azeredo J, Oliveira R (2006) Candida species adhesion to oral epithelium: factors involved and experimental methodology used. Crit Rev Microbiol 32:217–226

    Article  PubMed  CAS  Google Scholar 

  30. Chen L, Hobden JA, Masinick SA, Hazlett LD (1998) Environmental factors influence P. aeruginosa binding to the wounded mouse cornea. Curr Eye Res 17:231–237

    Article  PubMed  CAS  Google Scholar 

  31. He XY, Meurman JH, Kari K, Rautemaa R, Samaranayake LP (2006) In vitro adhesion of Candida species to denture base materials. Mycoses 49:80–84

    Article  PubMed  CAS  Google Scholar 

  32. Wasylnka JA, Moore MM (2000) Adhesion of Aspergillus species to extracellular matrix proteins: evidence for involvement of negatively charged carbohydrates on the conidial surface. Infect Immun 68:3377–3384

    Article  PubMed  CAS  Google Scholar 

  33. Ollert MW, Sohnchen R, Korting HC, Ollert U, Brautigam S, Brautigam W (1993) Mechanisms of adherence of Candida albicans to cultured human epidermal keratinocytes. Infect Immun 61:4560–4568

    PubMed  CAS  Google Scholar 

  34. Flanagan JL, Willcox MD (2009) Role of lactoferrin in the tear film. Biochimie 91:35–43

    Article  PubMed  CAS  Google Scholar 

  35. Thakur A, Chauhan A, Willcox MD (1999) Effect of lysozyme on adhesion and toxin release by Staphylococcus aureus. Aust N Z J Ophthalmol 27:224–227

    Article  PubMed  CAS  Google Scholar 

  36. Lan J, Willcox MD, Jackson GD (1999) Effect of tear-specific immunoglobulin A on the adhesion of Pseudomonas aeruginosa I to contact lenses. Aust NZ J Ophthalmol 27:218–220

    Article  CAS  Google Scholar 

  37. Leher H, Zaragoza F, Taherzadeh S, Alizadeh H, Niederkorn JY (1999) Monoclonal IgA antibodies protect against Acanthamoeba keratitis. Exp Eye Res 69:75–84

    Article  PubMed  CAS  Google Scholar 

  38. Halford WP, Schaffer PA (2000) Optimized viral dose and transient immunosuppression enable herpes simplex virus ICP0-null mutants to establish wild-type levels of latency in vivo. J Virol 74:5957–5967

    Article  PubMed  CAS  Google Scholar 

  39. Rediske AM, Koenig AL, Barekzi N, Ameen LC, Slunt JB, Grainger DW (2002) Polyclonal human antibodies reduce bacterial attachment to soft contact lens and corneal cell surfaces. Biomaterials 23:4565–4572

    Article  PubMed  CAS  Google Scholar 

  40. Masinick SA, Montgomery CP, Montgomery PC, Hazlett LD (1997) Secretory IgA inhibits Pseudomonas aeruginosa binding to cornea and protects against keratitis. Invest Ophthalmol Vis Sci 38:910–918

    PubMed  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by the National Natural Science Foundation of China (30630063). Qingjun Zhou was partially supported by the Taishan Scholar Program QDUEYE, Qingdao University, China. The authors thank Dr. Yan Gao and Meili Gao for their technical help with the animal experiments.

Declaration of interest

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Lixin Xie.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Zhou, Q., Chen, H., Qu, M. et al. Development of a novel ex vivo model of corneal fungal adherence. Graefes Arch Clin Exp Ophthalmol 249, 693–700 (2011). https://doi.org/10.1007/s00417-010-1601-9

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00417-010-1601-9

Keywords

Navigation