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Factor XIII Transglutaminase Supports the Resolution of Mucosal Damage in Experimental Colitis.

Andersson C, Kvist PH, McElhinney K, Baylis R, Gram LK, Pelzer H, Lauritzen B, Holm TL, Hogan S, Wu D, Turpin B, Miller W, Palumbo JS - PLoS ONE (2015)

Bottom Line: The thrombin-activated transglutaminase factor XIII (FXIII) that covalently crosslinks and stablizes provisional fibrin matrices is also thought to support endothelial and epithelial barrier function and to control inflammatory processes.The most striking differences were seen in the resolution of mucosal ulceration, the most severe histopathological manifestation of DSS-induced colitis.These findings directly demonstrate that FXIII is a significant determinant of mucosal healing and clinical outcome following inflammatory colitis induced mucosal injury and provide a proof-of-principle that clinical interventions supporting FXIII activity may be a means to limit colitis pathology and improve resolution of mucosal damage.

View Article: PubMed Central - PubMed

Affiliation: Novo Nordisk A/S, Biopharmaceutical Research Unit, Copenhagen, Denmark.

ABSTRACT
The thrombin-activated transglutaminase factor XIII (FXIII) that covalently crosslinks and stablizes provisional fibrin matrices is also thought to support endothelial and epithelial barrier function and to control inflammatory processes. Here, gene-targeted mice lacking the FXIII catalytic A subunit were employed to directly test the hypothesis that FXIII limits colonic pathologies associated with experimental colitis. Wildtype (WT) and FXIII-/- mice were found to be comparable in their initial development of mucosal damage following exposure to dextran sulfate sodium (DSS) challenge. However, unlike FXIII-sufficient mice, FXIII-deficient cohorts failed to efficiently resolve colonic inflammatory pathologies and mucosal damage following withdrawal of DSS. Consistent with prior evidence of ongoing coagulation factor activation and consumption in individuals with active colitis, plasma FXIII levels were markedly decreased in colitis-challenged WT mice. Treatment of colitis-challenged mice with recombinant human FXIII-A zymogen significantly mitigated weight loss, intestinal bleeding, and diarrhea, regardless of whether cohorts were FXIII-sufficient or were genetically devoid of FXIII. Similarly, both qualitative and quantitative microscopic analyses of colonic tissues revealed that exogenous FXIII improved the resolution of multiple colitis disease parameters in both FXIII-/- and WT mice. The most striking differences were seen in the resolution of mucosal ulceration, the most severe histopathological manifestation of DSS-induced colitis. These findings directly demonstrate that FXIII is a significant determinant of mucosal healing and clinical outcome following inflammatory colitis induced mucosal injury and provide a proof-of-principle that clinical interventions supporting FXIII activity may be a means to limit colitis pathology and improve resolution of mucosal damage.

No MeSH data available.


Related in: MedlinePlus

Treatment with rFXIII ameliorates colitis severity in both WT and FXIII−/− mice.(A) Pharmacological treatment of DSS-challenged WT mice with exogenous rFXIII (◯, n = 12) significantly reduced clinical disease parameters (disease activity index, DAI) relative to vehicle treated WT mice (●, n = 11). (B) A more modest, but significant therapeutic benefit of exogenous rFXIII (◯, n = 14) was also observed in FXIII−/− mice relative to vehicle treatment (●, n = 14). (C) Plasma levels of rFXIII measured at the end of the 14 day experiment were comparable between FXIII−/− and WT mice. As expected, no human FXIII protein was detectable (ND; not detected) in vehicle treated mice. (D) Total plasma FXIII catalytic activity measured at the end of the 14 day study period was predictably greatest in rFXIII treated WT mice. Note that this represents activity derived from both endogenous FXIII and exogenous rFXIII). (E) Representative microscopic view of H&E-stained section of colonic tissue harvested at the end of the 14 day study from a vehicle-treated FXIII−/− mouse. Note the large area of inflammatory edema (*) and mucosal ulceration (**). Treatment of FXIII−/− mice with rFXIII significantly limited colonic histopathology (F), but areas of mucosal damage, including ulceration (not shown), crypt loss (triangles) and edema (*) were still common. Consistent with prior results, colons harvested from vehicle-treated WT mice (G) exhibited significantly less evidence of residual pathology relative to vehicle-treated FXIII−/− mice (E) when collected 7 days after withdrawal of the DSS challenge. Nevertheless, small areas of ulceration (not shown) and more frequent areas of crypt spacing associated with inflammatory infiltrates (arrowhead) were still evident. More significantly, DSS-challenged WT mice treated with rFXIII (H) exhibited generally intact, near-normal appearing mucosa. (I) A detailed microscopic evaluation of tissue sections from all FXIII−/− mice employing the multiparameter histopathological scoring system showed that exogenous rFXIII treatment (white bars) resulted in a significant decrease in ulceration and crypt loss relative to vehicle treatment (black bars). (J) Parallel analyses in WT mice revealed a statistically significant diminution in every disease parameter analysed in rFXIII treated WT mice (white bars) relative to vehicle-treated WT mice (black bars). (K) Both FXIII genotype and treatment with rFXIII were important determinants of mucosal ulceration, shown as a percentage of colon length evaluated. Note that cohorts with the least amount of mucosal ulceration also had the highest level of plasma FXIII activity (Compare panels D and K). Bars in E-H represent 100 μm. *P < 0.05, P values were generated using a 2 way Anova (A & B) or a Mann-Whitney U test.
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pone.0128113.g005: Treatment with rFXIII ameliorates colitis severity in both WT and FXIII−/− mice.(A) Pharmacological treatment of DSS-challenged WT mice with exogenous rFXIII (◯, n = 12) significantly reduced clinical disease parameters (disease activity index, DAI) relative to vehicle treated WT mice (●, n = 11). (B) A more modest, but significant therapeutic benefit of exogenous rFXIII (◯, n = 14) was also observed in FXIII−/− mice relative to vehicle treatment (●, n = 14). (C) Plasma levels of rFXIII measured at the end of the 14 day experiment were comparable between FXIII−/− and WT mice. As expected, no human FXIII protein was detectable (ND; not detected) in vehicle treated mice. (D) Total plasma FXIII catalytic activity measured at the end of the 14 day study period was predictably greatest in rFXIII treated WT mice. Note that this represents activity derived from both endogenous FXIII and exogenous rFXIII). (E) Representative microscopic view of H&E-stained section of colonic tissue harvested at the end of the 14 day study from a vehicle-treated FXIII−/− mouse. Note the large area of inflammatory edema (*) and mucosal ulceration (**). Treatment of FXIII−/− mice with rFXIII significantly limited colonic histopathology (F), but areas of mucosal damage, including ulceration (not shown), crypt loss (triangles) and edema (*) were still common. Consistent with prior results, colons harvested from vehicle-treated WT mice (G) exhibited significantly less evidence of residual pathology relative to vehicle-treated FXIII−/− mice (E) when collected 7 days after withdrawal of the DSS challenge. Nevertheless, small areas of ulceration (not shown) and more frequent areas of crypt spacing associated with inflammatory infiltrates (arrowhead) were still evident. More significantly, DSS-challenged WT mice treated with rFXIII (H) exhibited generally intact, near-normal appearing mucosa. (I) A detailed microscopic evaluation of tissue sections from all FXIII−/− mice employing the multiparameter histopathological scoring system showed that exogenous rFXIII treatment (white bars) resulted in a significant decrease in ulceration and crypt loss relative to vehicle treatment (black bars). (J) Parallel analyses in WT mice revealed a statistically significant diminution in every disease parameter analysed in rFXIII treated WT mice (white bars) relative to vehicle-treated WT mice (black bars). (K) Both FXIII genotype and treatment with rFXIII were important determinants of mucosal ulceration, shown as a percentage of colon length evaluated. Note that cohorts with the least amount of mucosal ulceration also had the highest level of plasma FXIII activity (Compare panels D and K). Bars in E-H represent 100 μm. *P < 0.05, P values were generated using a 2 way Anova (A & B) or a Mann-Whitney U test.

Mentions: In order to determine if rFXIII treatment could ameliorate experimental colitis pathology, cohorts of WT and FXIII−/− mice were challenged with 7 days of DSS followed by water for 7 days and treated throughout the study period once daily with either 4 mg/kg rFXIII or an equivalent volume of vehicle (n = 11 to 15 per cohort). This dosing regimen was based on pharmacokinetic data that determined the half-life of rFXIII to be approximately 30 hours whether rFXIII was administered through intravenous or intraperitoneal injections. Furthermore, the half-life of rFXIII was the same in healthy and DSS-challenged mice (see S1 File for details). Consistent with previous reports showing that stress, including daily investigator handling of animals, exacerbates DSS-induced colitis severity [40, 41], DAI scores were appreciably higher in mice of both genotypes undergoing a daily injection regime over mice merely placed on DSS. Despite the more aggravated course of disease associated with the experimental design, exogenous rFXIII supplementation significantly ameliorated overall colitis disease relative to vehicle-treated animals based on both overall clinical DAI scores and multiparameter histological disease metrics (Fig 5). Note that the data shown in Fig 5 represent the compiled data from three experiments, each with similar results. The most impressive improvement in colitis outcome in mice treated with exogenous rFXIII was actually observed in WT cohorts that experienced a partial loss of FXIII following DSS challenge but still retained the capacity to generate endogenous FXIII (Fig 5A). A statistically significant improvement in disease outcome was also observed in rFXIII-treated FXIII−/− cohorts (Fig 5B). However, with the rFXIII dosing regimen employed the phenotypic benefit of exogenous rFXIII was more modest in FXIII−/− mice than that appreciated in WT cohorts, possibly due to an inability to more effectively restore FXIII levels in mice devoid of any endogenous FXIII.


Factor XIII Transglutaminase Supports the Resolution of Mucosal Damage in Experimental Colitis.

Andersson C, Kvist PH, McElhinney K, Baylis R, Gram LK, Pelzer H, Lauritzen B, Holm TL, Hogan S, Wu D, Turpin B, Miller W, Palumbo JS - PLoS ONE (2015)

Treatment with rFXIII ameliorates colitis severity in both WT and FXIII−/− mice.(A) Pharmacological treatment of DSS-challenged WT mice with exogenous rFXIII (◯, n = 12) significantly reduced clinical disease parameters (disease activity index, DAI) relative to vehicle treated WT mice (●, n = 11). (B) A more modest, but significant therapeutic benefit of exogenous rFXIII (◯, n = 14) was also observed in FXIII−/− mice relative to vehicle treatment (●, n = 14). (C) Plasma levels of rFXIII measured at the end of the 14 day experiment were comparable between FXIII−/− and WT mice. As expected, no human FXIII protein was detectable (ND; not detected) in vehicle treated mice. (D) Total plasma FXIII catalytic activity measured at the end of the 14 day study period was predictably greatest in rFXIII treated WT mice. Note that this represents activity derived from both endogenous FXIII and exogenous rFXIII). (E) Representative microscopic view of H&E-stained section of colonic tissue harvested at the end of the 14 day study from a vehicle-treated FXIII−/− mouse. Note the large area of inflammatory edema (*) and mucosal ulceration (**). Treatment of FXIII−/− mice with rFXIII significantly limited colonic histopathology (F), but areas of mucosal damage, including ulceration (not shown), crypt loss (triangles) and edema (*) were still common. Consistent with prior results, colons harvested from vehicle-treated WT mice (G) exhibited significantly less evidence of residual pathology relative to vehicle-treated FXIII−/− mice (E) when collected 7 days after withdrawal of the DSS challenge. Nevertheless, small areas of ulceration (not shown) and more frequent areas of crypt spacing associated with inflammatory infiltrates (arrowhead) were still evident. More significantly, DSS-challenged WT mice treated with rFXIII (H) exhibited generally intact, near-normal appearing mucosa. (I) A detailed microscopic evaluation of tissue sections from all FXIII−/− mice employing the multiparameter histopathological scoring system showed that exogenous rFXIII treatment (white bars) resulted in a significant decrease in ulceration and crypt loss relative to vehicle treatment (black bars). (J) Parallel analyses in WT mice revealed a statistically significant diminution in every disease parameter analysed in rFXIII treated WT mice (white bars) relative to vehicle-treated WT mice (black bars). (K) Both FXIII genotype and treatment with rFXIII were important determinants of mucosal ulceration, shown as a percentage of colon length evaluated. Note that cohorts with the least amount of mucosal ulceration also had the highest level of plasma FXIII activity (Compare panels D and K). Bars in E-H represent 100 μm. *P < 0.05, P values were generated using a 2 way Anova (A & B) or a Mann-Whitney U test.
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pone.0128113.g005: Treatment with rFXIII ameliorates colitis severity in both WT and FXIII−/− mice.(A) Pharmacological treatment of DSS-challenged WT mice with exogenous rFXIII (◯, n = 12) significantly reduced clinical disease parameters (disease activity index, DAI) relative to vehicle treated WT mice (●, n = 11). (B) A more modest, but significant therapeutic benefit of exogenous rFXIII (◯, n = 14) was also observed in FXIII−/− mice relative to vehicle treatment (●, n = 14). (C) Plasma levels of rFXIII measured at the end of the 14 day experiment were comparable between FXIII−/− and WT mice. As expected, no human FXIII protein was detectable (ND; not detected) in vehicle treated mice. (D) Total plasma FXIII catalytic activity measured at the end of the 14 day study period was predictably greatest in rFXIII treated WT mice. Note that this represents activity derived from both endogenous FXIII and exogenous rFXIII). (E) Representative microscopic view of H&E-stained section of colonic tissue harvested at the end of the 14 day study from a vehicle-treated FXIII−/− mouse. Note the large area of inflammatory edema (*) and mucosal ulceration (**). Treatment of FXIII−/− mice with rFXIII significantly limited colonic histopathology (F), but areas of mucosal damage, including ulceration (not shown), crypt loss (triangles) and edema (*) were still common. Consistent with prior results, colons harvested from vehicle-treated WT mice (G) exhibited significantly less evidence of residual pathology relative to vehicle-treated FXIII−/− mice (E) when collected 7 days after withdrawal of the DSS challenge. Nevertheless, small areas of ulceration (not shown) and more frequent areas of crypt spacing associated with inflammatory infiltrates (arrowhead) were still evident. More significantly, DSS-challenged WT mice treated with rFXIII (H) exhibited generally intact, near-normal appearing mucosa. (I) A detailed microscopic evaluation of tissue sections from all FXIII−/− mice employing the multiparameter histopathological scoring system showed that exogenous rFXIII treatment (white bars) resulted in a significant decrease in ulceration and crypt loss relative to vehicle treatment (black bars). (J) Parallel analyses in WT mice revealed a statistically significant diminution in every disease parameter analysed in rFXIII treated WT mice (white bars) relative to vehicle-treated WT mice (black bars). (K) Both FXIII genotype and treatment with rFXIII were important determinants of mucosal ulceration, shown as a percentage of colon length evaluated. Note that cohorts with the least amount of mucosal ulceration also had the highest level of plasma FXIII activity (Compare panels D and K). Bars in E-H represent 100 μm. *P < 0.05, P values were generated using a 2 way Anova (A & B) or a Mann-Whitney U test.
Mentions: In order to determine if rFXIII treatment could ameliorate experimental colitis pathology, cohorts of WT and FXIII−/− mice were challenged with 7 days of DSS followed by water for 7 days and treated throughout the study period once daily with either 4 mg/kg rFXIII or an equivalent volume of vehicle (n = 11 to 15 per cohort). This dosing regimen was based on pharmacokinetic data that determined the half-life of rFXIII to be approximately 30 hours whether rFXIII was administered through intravenous or intraperitoneal injections. Furthermore, the half-life of rFXIII was the same in healthy and DSS-challenged mice (see S1 File for details). Consistent with previous reports showing that stress, including daily investigator handling of animals, exacerbates DSS-induced colitis severity [40, 41], DAI scores were appreciably higher in mice of both genotypes undergoing a daily injection regime over mice merely placed on DSS. Despite the more aggravated course of disease associated with the experimental design, exogenous rFXIII supplementation significantly ameliorated overall colitis disease relative to vehicle-treated animals based on both overall clinical DAI scores and multiparameter histological disease metrics (Fig 5). Note that the data shown in Fig 5 represent the compiled data from three experiments, each with similar results. The most impressive improvement in colitis outcome in mice treated with exogenous rFXIII was actually observed in WT cohorts that experienced a partial loss of FXIII following DSS challenge but still retained the capacity to generate endogenous FXIII (Fig 5A). A statistically significant improvement in disease outcome was also observed in rFXIII-treated FXIII−/− cohorts (Fig 5B). However, with the rFXIII dosing regimen employed the phenotypic benefit of exogenous rFXIII was more modest in FXIII−/− mice than that appreciated in WT cohorts, possibly due to an inability to more effectively restore FXIII levels in mice devoid of any endogenous FXIII.

Bottom Line: The thrombin-activated transglutaminase factor XIII (FXIII) that covalently crosslinks and stablizes provisional fibrin matrices is also thought to support endothelial and epithelial barrier function and to control inflammatory processes.The most striking differences were seen in the resolution of mucosal ulceration, the most severe histopathological manifestation of DSS-induced colitis.These findings directly demonstrate that FXIII is a significant determinant of mucosal healing and clinical outcome following inflammatory colitis induced mucosal injury and provide a proof-of-principle that clinical interventions supporting FXIII activity may be a means to limit colitis pathology and improve resolution of mucosal damage.

View Article: PubMed Central - PubMed

Affiliation: Novo Nordisk A/S, Biopharmaceutical Research Unit, Copenhagen, Denmark.

ABSTRACT
The thrombin-activated transglutaminase factor XIII (FXIII) that covalently crosslinks and stablizes provisional fibrin matrices is also thought to support endothelial and epithelial barrier function and to control inflammatory processes. Here, gene-targeted mice lacking the FXIII catalytic A subunit were employed to directly test the hypothesis that FXIII limits colonic pathologies associated with experimental colitis. Wildtype (WT) and FXIII-/- mice were found to be comparable in their initial development of mucosal damage following exposure to dextran sulfate sodium (DSS) challenge. However, unlike FXIII-sufficient mice, FXIII-deficient cohorts failed to efficiently resolve colonic inflammatory pathologies and mucosal damage following withdrawal of DSS. Consistent with prior evidence of ongoing coagulation factor activation and consumption in individuals with active colitis, plasma FXIII levels were markedly decreased in colitis-challenged WT mice. Treatment of colitis-challenged mice with recombinant human FXIII-A zymogen significantly mitigated weight loss, intestinal bleeding, and diarrhea, regardless of whether cohorts were FXIII-sufficient or were genetically devoid of FXIII. Similarly, both qualitative and quantitative microscopic analyses of colonic tissues revealed that exogenous FXIII improved the resolution of multiple colitis disease parameters in both FXIII-/- and WT mice. The most striking differences were seen in the resolution of mucosal ulceration, the most severe histopathological manifestation of DSS-induced colitis. These findings directly demonstrate that FXIII is a significant determinant of mucosal healing and clinical outcome following inflammatory colitis induced mucosal injury and provide a proof-of-principle that clinical interventions supporting FXIII activity may be a means to limit colitis pathology and improve resolution of mucosal damage.

No MeSH data available.


Related in: MedlinePlus