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Role of Fas/FasL in regulation of inflammation in vaginal tissue during HSV-2 infection.

Krzyzowska M, Shestakov A, Eriksson K, Chiodi F - Cell Death Dis (2011)

Bottom Line: Despite upregulation of Fas and FasL, HSV-2-infected keratinocytes and epithelial cells showed a moderate level of apoptosis due to upregulated expression of the anti-apoptotic factors Bcl-2, Akt kinase and NF-κB.Inflammatory lesions within the HSV-2-infected epithelium of C57BL6 mice consisted of infected cells upregulating Fas, FasL and Bcl-2, uninfected cells upregulating Fas and neutrophils expressing both Fas and FasL.We conclude that the Fas pathway participates in regulation of inflammatory response in the vaginal epithelium at the initial stage of HSV-2 infection.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden. krzyzowskam@yahoo.com

ABSTRACT
To assess the role of Fas in lesion development during genital HSV-2 infection, we used a well-established HSV-2 murine model applied to MRL-Fas(lpr)/J (Fas-/-) and C3-Fasl(gld)/J (FasL-/-) C57BL6 mice. In vitro infection of murine keratinocytes and epithelial cells was used to clarify molecular details of HSV-2 infection. Despite upregulation of Fas and FasL, HSV-2-infected keratinocytes and epithelial cells showed a moderate level of apoptosis due to upregulated expression of the anti-apoptotic factors Bcl-2, Akt kinase and NF-κB. Inflammatory lesions within the HSV-2-infected epithelium of C57BL6 mice consisted of infected cells upregulating Fas, FasL and Bcl-2, uninfected cells upregulating Fas and neutrophils expressing both Fas and FasL. Apoptosis was detected in HSV-2-infected cells and to even higher extent in non-infected cells surrounding HSV-2 infection sites. HSV-2 infection of Fas- and FasL-deficient mice led to increased apoptosis and stronger recruitment of neutrophils within the infection sites. We conclude that the Fas pathway participates in regulation of inflammatory response in the vaginal epithelium at the initial stage of HSV-2 infection.

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Percentage of annexin V-positive cells (apoptotic cells) in epithelial Hepa 1–6 (a) and keratinocyte 03C (b) cell cultures subjected to cytotoxic Fas antibody (10 μg/ml) and FasL blocking antibody (10 μg/ml), infected or not, with HSV-2 for 24 h at MOI=1. (c) Percentage of HSV-2-infected keratinocytes at 24 h p.i. with MOI=1, in presence of cytotoxic anti-Fas antibody (10 μg/ml) or FasL blocking antibody (10 μg/ml). (d) Percentage of M30-positive cells (apoptotic cells) at 24 h after infection with MOI=0.1, in presence of cytotoxic anti-Fas antibody (10 μg/ml) or FasL blocking antibody (10 μg/ml) – cells were divided into infected and uninfected keratinocytes. Each bar represents the mean from five experiments (N=3)±S.E.M. *Represents significant differences with P≤0.05, whereas **means P≤0.001
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fig4: Percentage of annexin V-positive cells (apoptotic cells) in epithelial Hepa 1–6 (a) and keratinocyte 03C (b) cell cultures subjected to cytotoxic Fas antibody (10 μg/ml) and FasL blocking antibody (10 μg/ml), infected or not, with HSV-2 for 24 h at MOI=1. (c) Percentage of HSV-2-infected keratinocytes at 24 h p.i. with MOI=1, in presence of cytotoxic anti-Fas antibody (10 μg/ml) or FasL blocking antibody (10 μg/ml). (d) Percentage of M30-positive cells (apoptotic cells) at 24 h after infection with MOI=0.1, in presence of cytotoxic anti-Fas antibody (10 μg/ml) or FasL blocking antibody (10 μg/ml) – cells were divided into infected and uninfected keratinocytes. Each bar represents the mean from five experiments (N=3)±S.E.M. *Represents significant differences with P≤0.05, whereas **means P≤0.001

Mentions: To assess the sensitivity of HSV-2-infected epithelial cells and keratinocytes to Fas-induced apoptosis we used an anti-mouse Fas cytotoxic antibody (Jo-1 clone). In both the Hepa 1–6 epithelial cell cultures and in keratinocytes, addition of the anti-Fas antibody to uninfected cultures resulted in an increased percentage of annexin V-positive cells after 24 h (P<0.001) (Figures 4a and b). Addition of anti-Fas antibody to HSV-2-infected cultures also significantly increased Fas-mediated apoptosis (P<0.05) (Figures 4a and b). FasL blocking antibody (clone MFL-4) did not decrease the numbers of apoptotic cells in HSV-2-infected epithelial cells and keratinocytes (Figures 4a and b).


Role of Fas/FasL in regulation of inflammation in vaginal tissue during HSV-2 infection.

Krzyzowska M, Shestakov A, Eriksson K, Chiodi F - Cell Death Dis (2011)

Percentage of annexin V-positive cells (apoptotic cells) in epithelial Hepa 1–6 (a) and keratinocyte 03C (b) cell cultures subjected to cytotoxic Fas antibody (10 μg/ml) and FasL blocking antibody (10 μg/ml), infected or not, with HSV-2 for 24 h at MOI=1. (c) Percentage of HSV-2-infected keratinocytes at 24 h p.i. with MOI=1, in presence of cytotoxic anti-Fas antibody (10 μg/ml) or FasL blocking antibody (10 μg/ml). (d) Percentage of M30-positive cells (apoptotic cells) at 24 h after infection with MOI=0.1, in presence of cytotoxic anti-Fas antibody (10 μg/ml) or FasL blocking antibody (10 μg/ml) – cells were divided into infected and uninfected keratinocytes. Each bar represents the mean from five experiments (N=3)±S.E.M. *Represents significant differences with P≤0.05, whereas **means P≤0.001
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3101811&req=5

fig4: Percentage of annexin V-positive cells (apoptotic cells) in epithelial Hepa 1–6 (a) and keratinocyte 03C (b) cell cultures subjected to cytotoxic Fas antibody (10 μg/ml) and FasL blocking antibody (10 μg/ml), infected or not, with HSV-2 for 24 h at MOI=1. (c) Percentage of HSV-2-infected keratinocytes at 24 h p.i. with MOI=1, in presence of cytotoxic anti-Fas antibody (10 μg/ml) or FasL blocking antibody (10 μg/ml). (d) Percentage of M30-positive cells (apoptotic cells) at 24 h after infection with MOI=0.1, in presence of cytotoxic anti-Fas antibody (10 μg/ml) or FasL blocking antibody (10 μg/ml) – cells were divided into infected and uninfected keratinocytes. Each bar represents the mean from five experiments (N=3)±S.E.M. *Represents significant differences with P≤0.05, whereas **means P≤0.001
Mentions: To assess the sensitivity of HSV-2-infected epithelial cells and keratinocytes to Fas-induced apoptosis we used an anti-mouse Fas cytotoxic antibody (Jo-1 clone). In both the Hepa 1–6 epithelial cell cultures and in keratinocytes, addition of the anti-Fas antibody to uninfected cultures resulted in an increased percentage of annexin V-positive cells after 24 h (P<0.001) (Figures 4a and b). Addition of anti-Fas antibody to HSV-2-infected cultures also significantly increased Fas-mediated apoptosis (P<0.05) (Figures 4a and b). FasL blocking antibody (clone MFL-4) did not decrease the numbers of apoptotic cells in HSV-2-infected epithelial cells and keratinocytes (Figures 4a and b).

Bottom Line: Despite upregulation of Fas and FasL, HSV-2-infected keratinocytes and epithelial cells showed a moderate level of apoptosis due to upregulated expression of the anti-apoptotic factors Bcl-2, Akt kinase and NF-κB.Inflammatory lesions within the HSV-2-infected epithelium of C57BL6 mice consisted of infected cells upregulating Fas, FasL and Bcl-2, uninfected cells upregulating Fas and neutrophils expressing both Fas and FasL.We conclude that the Fas pathway participates in regulation of inflammatory response in the vaginal epithelium at the initial stage of HSV-2 infection.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden. krzyzowskam@yahoo.com

ABSTRACT
To assess the role of Fas in lesion development during genital HSV-2 infection, we used a well-established HSV-2 murine model applied to MRL-Fas(lpr)/J (Fas-/-) and C3-Fasl(gld)/J (FasL-/-) C57BL6 mice. In vitro infection of murine keratinocytes and epithelial cells was used to clarify molecular details of HSV-2 infection. Despite upregulation of Fas and FasL, HSV-2-infected keratinocytes and epithelial cells showed a moderate level of apoptosis due to upregulated expression of the anti-apoptotic factors Bcl-2, Akt kinase and NF-κB. Inflammatory lesions within the HSV-2-infected epithelium of C57BL6 mice consisted of infected cells upregulating Fas, FasL and Bcl-2, uninfected cells upregulating Fas and neutrophils expressing both Fas and FasL. Apoptosis was detected in HSV-2-infected cells and to even higher extent in non-infected cells surrounding HSV-2 infection sites. HSV-2 infection of Fas- and FasL-deficient mice led to increased apoptosis and stronger recruitment of neutrophils within the infection sites. We conclude that the Fas pathway participates in regulation of inflammatory response in the vaginal epithelium at the initial stage of HSV-2 infection.

Show MeSH
Related in: MedlinePlus