Increased neutrophil actin production and translocation of myeloperoxidase in patients with acetaminophen-induced acute liver failure may contribute to multiorgan failure

Godhev K. Manakkat Vijay; Gema Vizcay-Barrena; Thomas Tranah; Leanne Glover; Vishal C. Patel; Christine Bernsmeier; Jennifer M. Ryan; Laura J. Blackmore; Xiaohong Huang; Victoria T. Kronsten; William Bernal; Nicholas J. Taylor; Georg Auzinger; Christopher Willars; Julia Wendon; Yun Ma; Barbara Bain; Alice Warley; Debbie Shawcross

Increased neutrophil actin production and translocation of myeloperoxidase in patients with acetaminophen-induced acute liver failure may contribute to multiorgan failure

Godhev K. Manakkat Vijay, Gema Vizcay-Barrena, Thomas Tranah, Leanne Glover, Vishal C. Patel, Christine Bernsmeier, Jennifer M. Ryan, Laura J. Blackmore, Xiaohong Huang, Victoria T. Kronsten, William Bernal, Nicholas J. Taylor, Georg Auzinger, Christopher Willars, Julia Wendon, Yun Ma, Barbara Bain, Alice Warley, Debbie Shawcross

Centre for Ultrastructural Imaging

Research output: Contribution to journal › Meeting abstract › peer-review

Abstract

Background: There is a marked propensity for patients with acetaminophen-induced acute liver failure (AALF) to develop bacterial and fungal infections which may not only hasten the development of brain edema, but also precipitate multiorgan failure (MOF) and death. Neutrophil dysfunction has recently been shown to be an important biomarker of poor prognosis in AALF. Myeloperoxidase (MPO) is abundantly expressed in neutrophil azurophilic granules and generates reactive oxygen species (ROS) which kill invading pathogens but during AALF also induce bystander damage and MOF. Actin, a cytoskeletal globular protein, plays a key role in neutrophil degranulation. Therefore we sought to determine the neutrophil spontaneous oxidative burst (SOB), MPO degranulation and actin produc-tion in AALF (n=11) compared to healthy controls (HC) (n=10). Methods: Neutrophil SOB was measured by the conversion of dihydrorhodamine to rhodamine by peroxidase. Leukocytes were stained with fluorochrome-bound anti-CD16/CD11b/MPO antibodies to determine the neutrophil extracellular (EC) and intracellular (IC) (after permeabilisation) changes following the addition of E. coli, by flow cytometry. Transmission elec-tron microscopy (TEM) was performed on isolated leukocytes from whole blood and stained with primary mouse (anti-actin and anti-MPO) antibodies independently and then with sec-ondary gold-conjugated antibodies. Stained sections were viewed using TEM and images were acquired with a digital camera. The gold particles present in the cells were counted using ImageJ software and relative labelling index (RLI) was calculated to determine the specificity of the stain (<1-random and non-specific labelling). Plasma cytokines were also mea-sured. Results: SOB was increased in AALF neutrophils com-pared to HC (p<0.0001). Baseline EC MPO was increased in AALF patients compared to HC (p<0.0001). Following E. coli stimulation, EC MPO was increased in HC compared to base-line (p<0.05) but not in AALF. There was no difference in the baseline or post stimulation IC MPO between AALF and HC. TEM revealed no difference in the MPO labelling in AALF com-pared to HC. However, actin was increased in the cytoplasm of neutrophils (RLI>1) in AALF compared to HC (p<0.005; degree of freedom 1). Plasma IL-6 and IL-8 were increased in AALF compared to HC (p<0.0001). Conclusion: These data show that there is no deficiency in the production of MPO in AALF. However the increase in neutrophil SOB, degranulation as measured by actin and EC MPO in AALF implies that gran-ules translocate to the cell surface releasing ROS into the tissues thereby contributing to bystander damage and organ failure.

Original language	English
Article number	738
Pages (from-to)	558A-559A
Number of pages	2
Journal	Hepatology
Volume	60
Publication status	Published - 2014
Event	65th Annual Meeting of the American-Association-for-the-Study-of-Liver-Diseases - Boston, MA, United Kingdom Duration: 7 Nov 2014 → 11 Nov 2014

Cite this

Vijay, G. K. M., Vizcay-Barrena, G., Tranah, T., Glover, L., Patel, V. C., Bernsmeier, C., Ryan, J. M., Blackmore, L. J., Huang, X., Kronsten, V. T., Bernal, W., Taylor, N. J., Auzinger, G., Willars, C., Wendon, J., Ma, Y., Bain, B., Warley, A., & Shawcross, D. (2014). Increased neutrophil actin production and translocation of myeloperoxidase in patients with acetaminophen-induced acute liver failure may contribute to multiorgan failure. Hepatology, 60, 558A-559A. Article 738.

@article{e35d7b7985ca439f92d135687c945742,

title = "Increased neutrophil actin production and translocation of myeloperoxidase in patients with acetaminophen-induced acute liver failure may contribute to multiorgan failure",

abstract = "Background: There is a marked propensity for patients with acetaminophen-induced acute liver failure (AALF) to develop bacterial and fungal infections which may not only hasten the development of brain edema, but also precipitate multiorgan failure (MOF) and death. Neutrophil dysfunction has recently been shown to be an important biomarker of poor prognosis in AALF. Myeloperoxidase (MPO) is abundantly expressed in neutrophil azurophilic granules and generates reactive oxygen species (ROS) which kill invading pathogens but during AALF also induce bystander damage and MOF. Actin, a cytoskeletal globular protein, plays a key role in neutrophil degranulation. Therefore we sought to determine the neutrophil spontaneous oxidative burst (SOB), MPO degranulation and actin produc-tion in AALF (n=11) compared to healthy controls (HC) (n=10). Methods: Neutrophil SOB was measured by the conversion of dihydrorhodamine to rhodamine by peroxidase. Leukocytes were stained with fluorochrome-bound anti-CD16/CD11b/MPO antibodies to determine the neutrophil extracellular (EC) and intracellular (IC) (after permeabilisation) changes following the addition of E. coli, by flow cytometry. Transmission elec-tron microscopy (TEM) was performed on isolated leukocytes from whole blood and stained with primary mouse (anti-actin and anti-MPO) antibodies independently and then with sec-ondary gold-conjugated antibodies. Stained sections were viewed using TEM and images were acquired with a digital camera. The gold particles present in the cells were counted using ImageJ software and relative labelling index (RLI) was calculated to determine the specificity of the stain (<1-random and non-specific labelling). Plasma cytokines were also mea-sured. Results: SOB was increased in AALF neutrophils com-pared to HC (p<0.0001). Baseline EC MPO was increased in AALF patients compared to HC (p<0.0001). Following E. coli stimulation, EC MPO was increased in HC compared to base-line (p<0.05) but not in AALF. There was no difference in the baseline or post stimulation IC MPO between AALF and HC. TEM revealed no difference in the MPO labelling in AALF com-pared to HC. However, actin was increased in the cytoplasm of neutrophils (RLI>1) in AALF compared to HC (p<0.005; degree of freedom 1). Plasma IL-6 and IL-8 were increased in AALF compared to HC (p<0.0001). Conclusion: These data show that there is no deficiency in the production of MPO in AALF. However the increase in neutrophil SOB, degranulation as measured by actin and EC MPO in AALF implies that gran-ules translocate to the cell surface releasing ROS into the tissues thereby contributing to bystander damage and organ failure.",

author = "Vijay, {Godhev K. Manakkat} and Gema Vizcay-Barrena and Thomas Tranah and Leanne Glover and Patel, {Vishal C.} and Christine Bernsmeier and Ryan, {Jennifer M.} and Blackmore, {Laura J.} and Xiaohong Huang and Kronsten, {Victoria T.} and William Bernal and Taylor, {Nicholas J.} and Georg Auzinger and Christopher Willars and Julia Wendon and Yun Ma and Barbara Bain and Alice Warley and Debbie Shawcross",

note = "Publisher: WILEY-BLACKWELL, 111 RIVER ST, HOBOKEN 07030-5774, NJ USA Accession Number: WOS:000344483802234 ISSN: 0270-9139 eISSN: 1527-3350; 65th Annual Meeting of the American-Association-for-the-Study-of-Liver-Diseases ; Conference date: 07-11-2014 Through 11-11-2014",

year = "2014",

language = "English",

volume = "60",

pages = "558A--559A",

journal = "Hepatology",

issn = "0270-9139",

publisher = "Wolters Kluwer Health",

}

Vijay, GKM , Vizcay-Barrena, G, Tranah, T, Glover, L , Patel, VC , Bernsmeier, C , Ryan, JM , Blackmore, LJ , Huang, X, Kronsten, VT, Bernal, W, Taylor, NJ, Auzinger, G, Willars, C, Wendon, J , Ma, Y, Bain, B, Warley, A & Shawcross, D 2014, 'Increased neutrophil actin production and translocation of myeloperoxidase in patients with acetaminophen-induced acute liver failure may contribute to multiorgan failure', Hepatology, vol. 60, 738, pp. 558A-559A.

Increased neutrophil actin production and translocation of myeloperoxidase in patients with acetaminophen-induced acute liver failure may contribute to multiorgan failure. / Vijay, Godhev K. Manakkat ; Vizcay-Barrena, Gema; Tranah, Thomas et al.
In: Hepatology, Vol. 60, 738, 2014, p. 558A-559A.

Research output: Contribution to journal › Meeting abstract › peer-review

TY - JOUR

T1 - Increased neutrophil actin production and translocation of myeloperoxidase in patients with acetaminophen-induced acute liver failure may contribute to multiorgan failure

AU - Vijay, Godhev K. Manakkat

AU - Vizcay-Barrena, Gema

AU - Tranah, Thomas

AU - Glover, Leanne

AU - Patel, Vishal C.

AU - Bernsmeier, Christine

AU - Ryan, Jennifer M.

AU - Blackmore, Laura J.

AU - Huang, Xiaohong

AU - Kronsten, Victoria T.

AU - Bernal, William

AU - Taylor, Nicholas J.

AU - Auzinger, Georg

AU - Willars, Christopher

AU - Wendon, Julia

AU - Ma, Yun

AU - Bain, Barbara

AU - Warley, Alice

AU - Shawcross, Debbie

N1 - Publisher: WILEY-BLACKWELL, 111 RIVER ST, HOBOKEN 07030-5774, NJ USA Accession Number: WOS:000344483802234 ISSN: 0270-9139 eISSN: 1527-3350

PY - 2014

Y1 - 2014

N2 - Background: There is a marked propensity for patients with acetaminophen-induced acute liver failure (AALF) to develop bacterial and fungal infections which may not only hasten the development of brain edema, but also precipitate multiorgan failure (MOF) and death. Neutrophil dysfunction has recently been shown to be an important biomarker of poor prognosis in AALF. Myeloperoxidase (MPO) is abundantly expressed in neutrophil azurophilic granules and generates reactive oxygen species (ROS) which kill invading pathogens but during AALF also induce bystander damage and MOF. Actin, a cytoskeletal globular protein, plays a key role in neutrophil degranulation. Therefore we sought to determine the neutrophil spontaneous oxidative burst (SOB), MPO degranulation and actin produc-tion in AALF (n=11) compared to healthy controls (HC) (n=10). Methods: Neutrophil SOB was measured by the conversion of dihydrorhodamine to rhodamine by peroxidase. Leukocytes were stained with fluorochrome-bound anti-CD16/CD11b/MPO antibodies to determine the neutrophil extracellular (EC) and intracellular (IC) (after permeabilisation) changes following the addition of E. coli, by flow cytometry. Transmission elec-tron microscopy (TEM) was performed on isolated leukocytes from whole blood and stained with primary mouse (anti-actin and anti-MPO) antibodies independently and then with sec-ondary gold-conjugated antibodies. Stained sections were viewed using TEM and images were acquired with a digital camera. The gold particles present in the cells were counted using ImageJ software and relative labelling index (RLI) was calculated to determine the specificity of the stain (<1-random and non-specific labelling). Plasma cytokines were also mea-sured. Results: SOB was increased in AALF neutrophils com-pared to HC (p<0.0001). Baseline EC MPO was increased in AALF patients compared to HC (p<0.0001). Following E. coli stimulation, EC MPO was increased in HC compared to base-line (p<0.05) but not in AALF. There was no difference in the baseline or post stimulation IC MPO between AALF and HC. TEM revealed no difference in the MPO labelling in AALF com-pared to HC. However, actin was increased in the cytoplasm of neutrophils (RLI>1) in AALF compared to HC (p<0.005; degree of freedom 1). Plasma IL-6 and IL-8 were increased in AALF compared to HC (p<0.0001). Conclusion: These data show that there is no deficiency in the production of MPO in AALF. However the increase in neutrophil SOB, degranulation as measured by actin and EC MPO in AALF implies that gran-ules translocate to the cell surface releasing ROS into the tissues thereby contributing to bystander damage and organ failure.

AB - Background: There is a marked propensity for patients with acetaminophen-induced acute liver failure (AALF) to develop bacterial and fungal infections which may not only hasten the development of brain edema, but also precipitate multiorgan failure (MOF) and death. Neutrophil dysfunction has recently been shown to be an important biomarker of poor prognosis in AALF. Myeloperoxidase (MPO) is abundantly expressed in neutrophil azurophilic granules and generates reactive oxygen species (ROS) which kill invading pathogens but during AALF also induce bystander damage and MOF. Actin, a cytoskeletal globular protein, plays a key role in neutrophil degranulation. Therefore we sought to determine the neutrophil spontaneous oxidative burst (SOB), MPO degranulation and actin produc-tion in AALF (n=11) compared to healthy controls (HC) (n=10). Methods: Neutrophil SOB was measured by the conversion of dihydrorhodamine to rhodamine by peroxidase. Leukocytes were stained with fluorochrome-bound anti-CD16/CD11b/MPO antibodies to determine the neutrophil extracellular (EC) and intracellular (IC) (after permeabilisation) changes following the addition of E. coli, by flow cytometry. Transmission elec-tron microscopy (TEM) was performed on isolated leukocytes from whole blood and stained with primary mouse (anti-actin and anti-MPO) antibodies independently and then with sec-ondary gold-conjugated antibodies. Stained sections were viewed using TEM and images were acquired with a digital camera. The gold particles present in the cells were counted using ImageJ software and relative labelling index (RLI) was calculated to determine the specificity of the stain (<1-random and non-specific labelling). Plasma cytokines were also mea-sured. Results: SOB was increased in AALF neutrophils com-pared to HC (p<0.0001). Baseline EC MPO was increased in AALF patients compared to HC (p<0.0001). Following E. coli stimulation, EC MPO was increased in HC compared to base-line (p<0.05) but not in AALF. There was no difference in the baseline or post stimulation IC MPO between AALF and HC. TEM revealed no difference in the MPO labelling in AALF com-pared to HC. However, actin was increased in the cytoplasm of neutrophils (RLI>1) in AALF compared to HC (p<0.005; degree of freedom 1). Plasma IL-6 and IL-8 were increased in AALF compared to HC (p<0.0001). Conclusion: These data show that there is no deficiency in the production of MPO in AALF. However the increase in neutrophil SOB, degranulation as measured by actin and EC MPO in AALF implies that gran-ules translocate to the cell surface releasing ROS into the tissues thereby contributing to bystander damage and organ failure.

M3 - Meeting abstract

SN - 0270-9139

VL - 60

SP - 558A-559A

JO - Hepatology

JF - Hepatology

M1 - 738

T2 - 65th Annual Meeting of the American-Association-for-the-Study-of-Liver-Diseases

Y2 - 7 November 2014 through 11 November 2014

ER -

Increased neutrophil actin production and translocation of myeloperoxidase in patients with acetaminophen-induced acute liver failure may contribute to multiorgan failure

Abstract

Fingerprint

Cite this