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Hyaluronidase modulates inflammatory response and accelerates the cutaneous wound healing.

Fronza M, Caetano GF, Leite MN, Bitencourt CS, Paula-Silva FW, Andrade TA, Frade MA, Merfort I, Faccioli LH - PLoS ONE (2014)

Bottom Line: Hyaluronidases are enzymes that degrade hyaluronan an important constituent of the extracellular matrix.They have been used as a spreading agent, improving the absorption of drugs and facilitating the subcutaneous infusion of fluids.Moreover, HYAL increased gene expression of peroxisome proliferator-activated receptors (PPAR) γ and PPAR β/δ, the collagen content in the early stages of healing processes as well as angiogenesis.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Análises Clínicas, Toxicológicas e Bromatológicas, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, São Paulo, Brazil; Departamento de Farmácia, Universidade de Vila Velha, Vila Velha, Espirito Santo, Brazil.

ABSTRACT
Hyaluronidases are enzymes that degrade hyaluronan an important constituent of the extracellular matrix. They have been used as a spreading agent, improving the absorption of drugs and facilitating the subcutaneous infusion of fluids. Here, we investigated the influence of bovine testes hyaluronidase (HYAL) during cutaneous wound healing in in vitro and in vivo assays. We demonstrated in the wound scratch assay that HYAL increased the migration and proliferation of fibroblasts in vitro at low concentration, e.g. 0.1 U HYAL enhanced the cell number by 20%. HYAL presented faster and higher reepithelialization in in vivo full-thickness excisional wounds generated on adult Wistar rats back skin already in the early phase at 2nd day post operatory compared to vehicle-control group. Wound closured area observed in the 16 U and 32 U HYAL treated rats reached 38% and 46% compared to 19% in the controls, respectively. Histological and biochemical analyses supported the clinical observations and showed that HYAL treated wounds exhibited increased granulation tissue, diminished edema formation and regulated the inflammatory response by modulating the release of pro and anti-inflammatory cytokines, growth factor and eicosanoids mediators. Moreover, HYAL increased gene expression of peroxisome proliferator-activated receptors (PPAR) γ and PPAR β/δ, the collagen content in the early stages of healing processes as well as angiogenesis. Altogether these data revealed that HYAL accelerates wound healing processes and might be beneficial for treating wound disorders.

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Collagen accumulation in wound areas of HYAL and vehicle-control treated rats at day 2, 7, 14 and 21 after wounding.(A) Representative photomicrograph of wounds tissue sections stained with picrosirius red staining (200x), note the collagen intensity and disposition of fibers (red). (B) Collagen content measured by digital densitometry is shown as a result of collagen content in each specimen in percentage. (C) Determination of wound hydroxyproline content as an indicator of collagen levels. µg of hydroxyproline/mg of dry wound specimen content was measured at day zero, 2, 7, 14 and 21 after injury. Data represent means ± SEM (n = 8 wounds/group), *P<0.05, **P<0.01 compared to control group by one-way-ANOVA.
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pone-0112297-g004: Collagen accumulation in wound areas of HYAL and vehicle-control treated rats at day 2, 7, 14 and 21 after wounding.(A) Representative photomicrograph of wounds tissue sections stained with picrosirius red staining (200x), note the collagen intensity and disposition of fibers (red). (B) Collagen content measured by digital densitometry is shown as a result of collagen content in each specimen in percentage. (C) Determination of wound hydroxyproline content as an indicator of collagen levels. µg of hydroxyproline/mg of dry wound specimen content was measured at day zero, 2, 7, 14 and 21 after injury. Data represent means ± SEM (n = 8 wounds/group), *P<0.05, **P<0.01 compared to control group by one-way-ANOVA.

Mentions: Collagen deposition is an important event in the development of granulation tissue. We could observe that HYAL treated wounds revealed a marked and robust increase in the organization of collagen fibers, detected after staining with Picrosirius red in the wound biopsy, bridging the gaps in the skin compared to the vehicle-treated animals (Figure 4A). At days 2 and 7, the collagen content in the 16 U HYAL treated group was significantly higher than in the vehicle-control group, whereas after 14 and 21 days the collagen amount was similar between the groups with a slight decrease in the HYAL treated group at day 21 post wounding (Figure 4B). To confirm these histological findings, the collagen content was also measured by calculating the amount of hydroxyproline in the homogenate tissue of the wounds after 2, 7, 14 and 21 days post-wounding. The results of the hydroxyproline content in the skin after 2 days and 7 days by daily 16 U HYAL treatment showed a significant higher concentration compared to the controls (Figure 4C). Interestingly, after 21 days of 16 U HYAL wound treatment, the collagen content was significantly reduced compared to the control group.


Hyaluronidase modulates inflammatory response and accelerates the cutaneous wound healing.

Fronza M, Caetano GF, Leite MN, Bitencourt CS, Paula-Silva FW, Andrade TA, Frade MA, Merfort I, Faccioli LH - PLoS ONE (2014)

Collagen accumulation in wound areas of HYAL and vehicle-control treated rats at day 2, 7, 14 and 21 after wounding.(A) Representative photomicrograph of wounds tissue sections stained with picrosirius red staining (200x), note the collagen intensity and disposition of fibers (red). (B) Collagen content measured by digital densitometry is shown as a result of collagen content in each specimen in percentage. (C) Determination of wound hydroxyproline content as an indicator of collagen levels. µg of hydroxyproline/mg of dry wound specimen content was measured at day zero, 2, 7, 14 and 21 after injury. Data represent means ± SEM (n = 8 wounds/group), *P<0.05, **P<0.01 compared to control group by one-way-ANOVA.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4230982&req=5

pone-0112297-g004: Collagen accumulation in wound areas of HYAL and vehicle-control treated rats at day 2, 7, 14 and 21 after wounding.(A) Representative photomicrograph of wounds tissue sections stained with picrosirius red staining (200x), note the collagen intensity and disposition of fibers (red). (B) Collagen content measured by digital densitometry is shown as a result of collagen content in each specimen in percentage. (C) Determination of wound hydroxyproline content as an indicator of collagen levels. µg of hydroxyproline/mg of dry wound specimen content was measured at day zero, 2, 7, 14 and 21 after injury. Data represent means ± SEM (n = 8 wounds/group), *P<0.05, **P<0.01 compared to control group by one-way-ANOVA.
Mentions: Collagen deposition is an important event in the development of granulation tissue. We could observe that HYAL treated wounds revealed a marked and robust increase in the organization of collagen fibers, detected after staining with Picrosirius red in the wound biopsy, bridging the gaps in the skin compared to the vehicle-treated animals (Figure 4A). At days 2 and 7, the collagen content in the 16 U HYAL treated group was significantly higher than in the vehicle-control group, whereas after 14 and 21 days the collagen amount was similar between the groups with a slight decrease in the HYAL treated group at day 21 post wounding (Figure 4B). To confirm these histological findings, the collagen content was also measured by calculating the amount of hydroxyproline in the homogenate tissue of the wounds after 2, 7, 14 and 21 days post-wounding. The results of the hydroxyproline content in the skin after 2 days and 7 days by daily 16 U HYAL treatment showed a significant higher concentration compared to the controls (Figure 4C). Interestingly, after 21 days of 16 U HYAL wound treatment, the collagen content was significantly reduced compared to the control group.

Bottom Line: Hyaluronidases are enzymes that degrade hyaluronan an important constituent of the extracellular matrix.They have been used as a spreading agent, improving the absorption of drugs and facilitating the subcutaneous infusion of fluids.Moreover, HYAL increased gene expression of peroxisome proliferator-activated receptors (PPAR) γ and PPAR β/δ, the collagen content in the early stages of healing processes as well as angiogenesis.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Análises Clínicas, Toxicológicas e Bromatológicas, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, São Paulo, Brazil; Departamento de Farmácia, Universidade de Vila Velha, Vila Velha, Espirito Santo, Brazil.

ABSTRACT
Hyaluronidases are enzymes that degrade hyaluronan an important constituent of the extracellular matrix. They have been used as a spreading agent, improving the absorption of drugs and facilitating the subcutaneous infusion of fluids. Here, we investigated the influence of bovine testes hyaluronidase (HYAL) during cutaneous wound healing in in vitro and in vivo assays. We demonstrated in the wound scratch assay that HYAL increased the migration and proliferation of fibroblasts in vitro at low concentration, e.g. 0.1 U HYAL enhanced the cell number by 20%. HYAL presented faster and higher reepithelialization in in vivo full-thickness excisional wounds generated on adult Wistar rats back skin already in the early phase at 2nd day post operatory compared to vehicle-control group. Wound closured area observed in the 16 U and 32 U HYAL treated rats reached 38% and 46% compared to 19% in the controls, respectively. Histological and biochemical analyses supported the clinical observations and showed that HYAL treated wounds exhibited increased granulation tissue, diminished edema formation and regulated the inflammatory response by modulating the release of pro and anti-inflammatory cytokines, growth factor and eicosanoids mediators. Moreover, HYAL increased gene expression of peroxisome proliferator-activated receptors (PPAR) γ and PPAR β/δ, the collagen content in the early stages of healing processes as well as angiogenesis. Altogether these data revealed that HYAL accelerates wound healing processes and might be beneficial for treating wound disorders.

Show MeSH
Related in: MedlinePlus