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. 2017 Jan 22;482(4):569-574.
doi: 10.1016/j.bbrc.2016.11.075. Epub 2016 Nov 15.

Glycogen storage disease type Ib neutrophils exhibit impaired cell adhesion and migration

Goo-Young Kim  1 , Young Mok Lee  1 , Joon Hyun Kwon  1 , Hyun Sik Jun  2 , Janice Chou  3
Affiliations

Glycogen storage disease type Ib neutrophils exhibit impaired cell adhesion and migration

Goo-Young Kim et al. Biochem Biophys Res Commun. .

Abstract

Glycogen storage disease type Ib (GSD-Ib), characterized by impaired glucose homeostasis, neutropenia, and neutrophil dysfunction, is an inherited autosomal recessive disorder caused by a deficiency in the glucose-6-phosphate transporter (G6PT). Neutrophils play an essential role in the defense against invading pathogens. The recruitment of neutrophils towards the inflammation sites in response to inflammatory stimuli is a tightly regulated process involving rolling, adhesion, and transmigration. In this study, we investigated the role of G6PT in neutrophil adhesion and migration using in vivo and in vitro models. We showed that the GSD-Ib (G6pt-/-) mice manifested severe neutropenia in both blood and bone marrow, and treating G6pt-/- mice with granulocyte colony-stimulating factor (G-CSF) corrected neutropenia. However, upon thioglycolate challenge, neutrophils from both untreated and G-CSF-treated G6pt-/-mice exhibited decreased ability to migrate to the peritoneal cavity. In vitro migration and cell adhesion of G6PT-deficient neutrophils were also significantly impaired. Defects in cell migration were not due to enhanced apoptosis or altered fMLP receptor expression. Remarkably, the expression of the β2 integrins CD11a and CD11b, which are critical for cell adhesion, was greatly decreased in G6PT-deficient neutrophils. This study suggests that deficiencies in G6PT cause impairment in neutrophil adhesion and migration via aberrant expression of β2 integrins, and our finding should facilitate the development of novel therapies for GSD-Ib.

Keywords: CD11a; CD11b; Glucose-6-phosphate transporter; Migration; Neutrophil adhesion.

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Conflict of interest statement

Conflict of interest

None.

Figures

Fig. 1.
Fig. 1.
Neutrophil population in peripheral blood stream and bone marrow (BM) of G6pt−/− and control mice. (A) Representative flow cytometry data (left) and frequency of Gr-1+CD11b+ cells in the blood and bone marrow (BM) of G6pt−/− and control mice (WT) (n = 10 per genotype). (B) Absolute number of white blood cells (WBC) and neutrophils in the blood of G6pt−/− (−/−) and control mice (+/+) (n = 10 per genotype). Each symbol represents an individual mouse, and horizontal lines represent the mean. Data represent the mean ± SEM (n = 10). *P < 0.05, **P < 0.01.
Fig. 2.
Fig. 2.
Neutrophil recruitment to the peritoneal cavity during peritonitis. (A) Representative flow cytometry data (left) and frequency of Gr-1+CD11b+ cells in the blood of G6pt−/− (−/−) and control mice (+/+) (n = 5 per genotype) treated with G-CSF or PBS as a vehicle for 5 days. (B) Quantification of the total number of neutrophils migrated to the peritoneal cavity of G6pt−/− (−/−) and control mice (+/+) (n = 3 per genotype) 4 h after intraperitoneal injection of thioglycolate (3% w/v). Data represent the mean ± SEM (n = 10). *P < 0.05, **P < 0.01.
Fig. 3.
Fig. 3.
In vitro migration and adhesion of neutrophils from G6pt−/− and control mice. (A) Neutrophil migration in response to fMLP. (B) Adhesion of PMA-activated neutrophils to purified mouse ICAM-1 and fibrinogen. Data presented in the chart represent the average of three independent experiments with counting neutrophils in four random fields. (C) Adhesion of PMA-activated neutrophils to epithelial cells. Two epithelial cells, MDCK and Caco2, were treated with TNFα as described in Materials and Methods. Data presented in the charts represent the average of three independent experiments with counting neutrophils in four random fields. Data represent the mean ± SEM. *P < 0.05, **P < 0.01.
Fig. 4.
Fig. 4.
Analysis of the expression of CD11b and CD11a, on neutrophils from G6pt−/− and control mice. (A) Quantification of CD11b mRNA in G6PT-deficient (−/−) and wild-type neutrophils (+/+) by real-time RT-PCR. Expression is normalized to that of β-actin and measured relative to one wild-type neutrophil arbitrarily defined as 1. (B) Western blot analysis of protein extracts of neutrophils. Arrowhead indicates the deglycosylated form of CD11b. Relative CD11b and CD11a protein levels were quantified by densitometry of at least three independent western blots. Data represent the mean ± SEM. *P < 0.05, **P < 0.01.
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