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Research article

Molecular and Cellular Microbiology / Biomedical Sciences

In Vitro and in Vivo Wound Healing Properties of Plasma and Serum from Crocodylus siamensis Blood

Nisachon Jangpromma 1, 2, Sutthidech Preecharram 3, Thanawan Srilert 2, 4, Surachai Maijaroen 2, 4, Pramote Mahakunakorn 5, Natsajee Nualkaew 5, Sakda Daduang 2, 5 and Sompong Klaynongsruang 2, 4*

1Office of the Dean, Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand, 2Protein and Proteomics Research Center for Commercial and Industrial Purposes (ProCCI), Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand, 3Department of General Science, Faculty of Science and Engineering, Kasetsart University, Chalermphrakiat, Sakon Nakhon Province 47000, Thailand, 4Department of Biochemistry, Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand, 5Department of Pharmacognosy and Toxicology, Faculty of Pharmaceutical Sciences, Khon Kaen University, Khon Kaen 40002, Thailand

Received: January 21, 2016; Accepted: March 8, 2016

J. Microbiol. Biotechnol. 2016; 26(6): 1140-1147

Published June 28, 2016 https://doi.org/10.4014/jmb.1601.01054

Copyright © The Korean Society for Microbiology and Biotechnology.

Abstract

The plasma and serum of Crocodylus siamensis have previously been reported to exhibit potent antimicrobial, antioxidant, and anti-inflammatory activities. During wound healing, these biological properties play a crucial role for supporting the formation of new tissue around the injured skin in the recovery process. Thus, this study aimed to evaluate the wound healing properties of C. siamensis plasma and serum. The collected data demonstrate that crocodile plasma and serum were able to activate in vitro proliferation and migration of HaCaT, a human keratinocyte cell line, which represents an essential phase in the wound healing process. With respect to investigating cell migration, a scratch wound experiment was performed which revealed the ability of plasma and serum to decrease the gap of wounds in a dose-dependent manner. Consistent with the in vitro results, remarkably enhanced wound repair was also observed in a mouse excisional skin wound model after treatment with plasma or serum. The effects of C. siamensis plasma and serum on wound healing were further elucidated by treating wound infections by Staphylococcus aureus ATCC 25923 on mice skin coupled with a histological method. The results indicate that crocodile plasma and serum promote the prevention of wound infection and boost the re-epithelialization necessary for the formation of new skin. Therefore, this work represents the first study to demonstrate the efficiency of C. siamensis plasma and serum with respect to their wound healing properties and strongly supports the utilization of C. siamensis plasma and serum as therapeutic products for injured skin treatment.

Keywords

Wound healing, Wound infection, Scratch assay, Plasma, Serum, Crocodile blood

References

  1. Arunachalam K, Parimelazhagan T. 2013. Anti-inflammatory, wound healing and in-vivo antioxidant properties of the leaves of Ficus amplissima Smith. J. Ethnopharmacol. 145: 139-145.
    Pubmed CrossRef
  2. Buthelezi S, Southway C, Govinden U, Bodenstein J, du Toit K. 2012. An investigation of the antimicrobial and antiinflammatory activities of crocodile oil. J. Ethnopharmacol. 143: 325-230.
    Pubmed CrossRef
  3. Carretero M, Escámez MJ, García M, Duarte B, Holguín A, Retamosa L, et al. 2008. In vitro and in vivo wound healingpromoting activities of human cathelicidin LL-37. J. Invest. Dermatol. 128: 223-236.
    Pubmed CrossRef
  4. Carter CA, Jolly DG, Worden CES, Hendren DG, Kane CJ. 2003. Platelet-rich plasma gel promotes differentiation and regeneration during equine wound healing. Exp. Mol. Pathol. 74: 244-255.
    CrossRef
  5. Demirci S, Do an A, Demirci Y, ahin F. 2014. In vitro wound healing activity of methanol extract of Verbascum speciosum. Int. J. Appl. Res. Nat. Prod. 7: 37-44.
  6. Hamblin MR, Zahra T, Contag CH, McManus AT, Hasan T. 2003. Optical monitoring and treatment of potentially lethal wound infections in vivo. J. Infect. Dis. 187: 1717-1725.
    Pubmed PMC CrossRef
  7. Jorge MP, Madjarof C, Gois Ruiz AL, Fernandes AT, Ferreira Rodrigues RA, de Oliveira Sousa IM, et al. 2008. Evaluation of wound healing properties of Arrabidaea chica Verlot extract. J. Ethnopharmacol. 13: 361-366.
    Pubmed CrossRef
  8. Kommanee J, Phosri S, Daduang S, Temsiripong Y, Dhiravisit A, Thammasirirak S. 2014. Comparisons of antiinflammatory activity of crocodile (Crocodylus siamensis) blood extract. Chiang Mai J. Sci. 41: 627-634.
  9. Kommanee J, Preecharram S, Daduang S, Temsiripong Y, Dhiravisit A, Yamada Y, Thammasirirak S. 2012. Antibacterial activity of plasma from crocodile (Crocodylus siamensis) against pathogenic bacteria. Ann. Clin. Microbiol. Antimicrob. 11: 22.
    Pubmed PMC CrossRef
  10. Kurahashi T, Fujii J. 2015. Roles of antioxidative enzymes in wound healing. J. Dev. Biol. 3: 57-70.
    CrossRef
  11. Leu JG, Chen SA, Chen HM, Wu WM, Hung CF, Yao YD, et al. 2012. The effects of gold nanoparticles in wound healing with antioxidant epigallocatechin gallate and α-lipoic acid. Nanomedicine 8: 767-775.
    CrossRef
  12. Merchant ME, Roche C, Elsey RM, Prudhomme J. 2003. Antibacterial properties of serum from the American alligator (Alligator mississippiensis). Comp. Biochem. Physiol. B Biochem. Mol. Biol. 136: 505-513.
    CrossRef
  13. Merchant ME, Roche CM, Thibodeaux D, Elsey RM. 2005. Identification of alternative pathway serum complement activity in the blood of the American alligator (Alligator mississippiensis). Comp. Biochem. Physiol. B. Biochem. Mol. Biol. 141: 281-288.
    Pubmed CrossRef
  14. Merchant ME, Thibodeaux D, Loubser K, Elsey RM. 2004. Amoebacidal effects of serum from the American alligator (Alligator mississippiensis). J. Parasitol. 90: 1480-1483.
    Pubmed CrossRef
  15. Phosri S, Mahakunakorn P, Lueangsakulthai J, Jangpromma N, Swatsitang P, Daduang S, et al. 2014. An investigation of antioxidant and anti-inflammatory activities from blood components of crocodile (Crocodylus siamensis). Protein J. 33:484-492.
    Pubmed CrossRef
  16. Preecharram S, Daduang S, Bunyatratchata W, Araki T, Thammasirirak S. 2008. Antibacterial activity from Siamese crocodile (Crocodylus siamensis) serum. Afr. J. Biotechnol. 7:3121-3128.
  17. Preecharram S, Jearranaiprepame P, Daduang S, Temsiripong Y, Somdee T, Fukamizo T, et al. 2 01 0. I solation and characterisation of crocosin, an antibacterial compound from crocodile (Crocodylus siamensis) plasma. Anim. Sci. J. 81: 393-401.
    Pubmed CrossRef
  18. Ranzato E, Patrone M, Mazzucco L, Burlando B. 2008. Platelet lysate stimulates wound repair of HaCaT keratinocytes. Br. J. Dermatol. 159: 537-545.
    CrossRef
  19. Sun DP, Yeh CH, So E, Wang LY, Wei TS, Chang MS, Hsing CH. 2013. Interleukin (IL)-19 promoted skin wound healing by increasing fibroblast keratinocyte growth factor expression. Cytokine 62: 360-368.
    Pubmed CrossRef
  20. Tang J, Liu H, Gao C, Mu L, Yang S, Rong M, et al. 2014. A small peptide with potential ability to promote wound healing. PLoS One 9: e92082.
    Pubmed PMC CrossRef
  21. van Hoek ML. 2014. Antimicrobial peptides in reptiles. Pharmaceuticals 7: 723-753.
    Pubmed PMC CrossRef
  22. Yang X, Wang J, Guo SL, Fan KJ, Li J, Wang YL, et al. 2011. miR-21 promotes keratinocyte migration and re-epithelialization during wound healing Int. J. Biol. Sci. 7: 685-690.
    Pubmed PMC CrossRef
  23. Zhang Y, Bai X, Wang Y, Li N, Li X, Han F, et al. 2014. Role for heat shock protein 90a in the proliferation and migration of HaCaT cells and in the deep second-degree burn wound healing in mice. PLoS One 9: e103723.
    Pubmed PMC CrossRef

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Article

Research article

J. Microbiol. Biotechnol. 2016; 26(6): 1140-1147

Published online June 28, 2016 https://doi.org/10.4014/jmb.1601.01054

Copyright © The Korean Society for Microbiology and Biotechnology.

In Vitro and in Vivo Wound Healing Properties of Plasma and Serum from Crocodylus siamensis Blood

Nisachon Jangpromma 1, 2, Sutthidech Preecharram 3, Thanawan Srilert 2, 4, Surachai Maijaroen 2, 4, Pramote Mahakunakorn 5, Natsajee Nualkaew 5, Sakda Daduang 2, 5 and Sompong Klaynongsruang 2, 4*

1Office of the Dean, Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand, 2Protein and Proteomics Research Center for Commercial and Industrial Purposes (ProCCI), Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand, 3Department of General Science, Faculty of Science and Engineering, Kasetsart University, Chalermphrakiat, Sakon Nakhon Province 47000, Thailand, 4Department of Biochemistry, Faculty of Science, Khon Kaen University, Khon Kaen 40002, Thailand, 5Department of Pharmacognosy and Toxicology, Faculty of Pharmaceutical Sciences, Khon Kaen University, Khon Kaen 40002, Thailand

Received: January 21, 2016; Accepted: March 8, 2016

Abstract

The plasma and serum of Crocodylus siamensis have previously been reported to exhibit potent
antimicrobial, antioxidant, and anti-inflammatory activities. During wound healing, these
biological properties play a crucial role for supporting the formation of new tissue around the
injured skin in the recovery process. Thus, this study aimed to evaluate the wound healing
properties of C. siamensis plasma and serum. The collected data demonstrate that crocodile
plasma and serum were able to activate in vitro proliferation and migration of HaCaT, a
human keratinocyte cell line, which represents an essential phase in the wound healing
process. With respect to investigating cell migration, a scratch wound experiment was
performed which revealed the ability of plasma and serum to decrease the gap of wounds in a
dose-dependent manner. Consistent with the in vitro results, remarkably enhanced wound
repair was also observed in a mouse excisional skin wound model after treatment with plasma
or serum. The effects of C. siamensis plasma and serum on wound healing were further
elucidated by treating wound infections by Staphylococcus aureus ATCC 25923 on mice skin
coupled with a histological method. The results indicate that crocodile plasma and serum
promote the prevention of wound infection and boost the re-epithelialization necessary for the
formation of new skin. Therefore, this work represents the first study to demonstrate the
efficiency of C. siamensis plasma and serum with respect to their wound healing properties
and strongly supports the utilization of C. siamensis plasma and serum as therapeutic products
for injured skin treatment.

Keywords: Wound healing, Wound infection, Scratch assay, Plasma, Serum, Crocodile blood

References

  1. Arunachalam K, Parimelazhagan T. 2013. Anti-inflammatory, wound healing and in-vivo antioxidant properties of the leaves of Ficus amplissima Smith. J. Ethnopharmacol. 145: 139-145.
    Pubmed CrossRef
  2. Buthelezi S, Southway C, Govinden U, Bodenstein J, du Toit K. 2012. An investigation of the antimicrobial and antiinflammatory activities of crocodile oil. J. Ethnopharmacol. 143: 325-230.
    Pubmed CrossRef
  3. Carretero M, Escámez MJ, García M, Duarte B, Holguín A, Retamosa L, et al. 2008. In vitro and in vivo wound healingpromoting activities of human cathelicidin LL-37. J. Invest. Dermatol. 128: 223-236.
    Pubmed CrossRef
  4. Carter CA, Jolly DG, Worden CES, Hendren DG, Kane CJ. 2003. Platelet-rich plasma gel promotes differentiation and regeneration during equine wound healing. Exp. Mol. Pathol. 74: 244-255.
    CrossRef
  5. Demirci S, Do an A, Demirci Y, ahin F. 2014. In vitro wound healing activity of methanol extract of Verbascum speciosum. Int. J. Appl. Res. Nat. Prod. 7: 37-44.
  6. Hamblin MR, Zahra T, Contag CH, McManus AT, Hasan T. 2003. Optical monitoring and treatment of potentially lethal wound infections in vivo. J. Infect. Dis. 187: 1717-1725.
    Pubmed KoreaMed CrossRef
  7. Jorge MP, Madjarof C, Gois Ruiz AL, Fernandes AT, Ferreira Rodrigues RA, de Oliveira Sousa IM, et al. 2008. Evaluation of wound healing properties of Arrabidaea chica Verlot extract. J. Ethnopharmacol. 13: 361-366.
    Pubmed CrossRef
  8. Kommanee J, Phosri S, Daduang S, Temsiripong Y, Dhiravisit A, Thammasirirak S. 2014. Comparisons of antiinflammatory activity of crocodile (Crocodylus siamensis) blood extract. Chiang Mai J. Sci. 41: 627-634.
  9. Kommanee J, Preecharram S, Daduang S, Temsiripong Y, Dhiravisit A, Yamada Y, Thammasirirak S. 2012. Antibacterial activity of plasma from crocodile (Crocodylus siamensis) against pathogenic bacteria. Ann. Clin. Microbiol. Antimicrob. 11: 22.
    Pubmed KoreaMed CrossRef
  10. Kurahashi T, Fujii J. 2015. Roles of antioxidative enzymes in wound healing. J. Dev. Biol. 3: 57-70.
    CrossRef
  11. Leu JG, Chen SA, Chen HM, Wu WM, Hung CF, Yao YD, et al. 2012. The effects of gold nanoparticles in wound healing with antioxidant epigallocatechin gallate and α-lipoic acid. Nanomedicine 8: 767-775.
    CrossRef
  12. Merchant ME, Roche C, Elsey RM, Prudhomme J. 2003. Antibacterial properties of serum from the American alligator (Alligator mississippiensis). Comp. Biochem. Physiol. B Biochem. Mol. Biol. 136: 505-513.
    CrossRef
  13. Merchant ME, Roche CM, Thibodeaux D, Elsey RM. 2005. Identification of alternative pathway serum complement activity in the blood of the American alligator (Alligator mississippiensis). Comp. Biochem. Physiol. B. Biochem. Mol. Biol. 141: 281-288.
    Pubmed CrossRef
  14. Merchant ME, Thibodeaux D, Loubser K, Elsey RM. 2004. Amoebacidal effects of serum from the American alligator (Alligator mississippiensis). J. Parasitol. 90: 1480-1483.
    Pubmed CrossRef
  15. Phosri S, Mahakunakorn P, Lueangsakulthai J, Jangpromma N, Swatsitang P, Daduang S, et al. 2014. An investigation of antioxidant and anti-inflammatory activities from blood components of crocodile (Crocodylus siamensis). Protein J. 33:484-492.
    Pubmed CrossRef
  16. Preecharram S, Daduang S, Bunyatratchata W, Araki T, Thammasirirak S. 2008. Antibacterial activity from Siamese crocodile (Crocodylus siamensis) serum. Afr. J. Biotechnol. 7:3121-3128.
  17. Preecharram S, Jearranaiprepame P, Daduang S, Temsiripong Y, Somdee T, Fukamizo T, et al. 2 01 0. I solation and characterisation of crocosin, an antibacterial compound from crocodile (Crocodylus siamensis) plasma. Anim. Sci. J. 81: 393-401.
    Pubmed CrossRef
  18. Ranzato E, Patrone M, Mazzucco L, Burlando B. 2008. Platelet lysate stimulates wound repair of HaCaT keratinocytes. Br. J. Dermatol. 159: 537-545.
    CrossRef
  19. Sun DP, Yeh CH, So E, Wang LY, Wei TS, Chang MS, Hsing CH. 2013. Interleukin (IL)-19 promoted skin wound healing by increasing fibroblast keratinocyte growth factor expression. Cytokine 62: 360-368.
    Pubmed CrossRef
  20. Tang J, Liu H, Gao C, Mu L, Yang S, Rong M, et al. 2014. A small peptide with potential ability to promote wound healing. PLoS One 9: e92082.
    Pubmed KoreaMed CrossRef
  21. van Hoek ML. 2014. Antimicrobial peptides in reptiles. Pharmaceuticals 7: 723-753.
    Pubmed KoreaMed CrossRef
  22. Yang X, Wang J, Guo SL, Fan KJ, Li J, Wang YL, et al. 2011. miR-21 promotes keratinocyte migration and re-epithelialization during wound healing Int. J. Biol. Sci. 7: 685-690.
    Pubmed KoreaMed CrossRef
  23. Zhang Y, Bai X, Wang Y, Li N, Li X, Han F, et al. 2014. Role for heat shock protein 90a in the proliferation and migration of HaCaT cells and in the deep second-degree burn wound healing in mice. PLoS One 9: e103723.
    Pubmed KoreaMed CrossRef