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Increased Susceptibility of Humanized NSG Mice to Panton-Valentine Leukocidin and Staphylococcus aureus Skin Infection.

Tseng CW, Biancotti JC, Berg BL, Gate D, Kolar SL, Müller S, Rodriguez MD, Rezai-Zadeh K, Fan X, Beenhouwer DO, Town T, Liu GY - PLoS Pathog. (2015)

Bottom Line: Staphylococcus aureus is a leading cause of skin and soft-tissue infections worldwide.However a supra-physiologic S. aureus inoculum is required to establish gross murine skin pathology.PMX53, a human C5aR inhibitor, blocked PVL-induced cellular cytotoxicity in vitro and reduced the size difference of lesions induced by the PVL+ and PVL- S. aureus, but PMX53 also reduced recruitment of neutrophils and exacerbated the infection.

View Article: PubMed Central - PubMed

Affiliation: Division of Pediatric Infectious Diseases and the Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California, United States of America.

ABSTRACT
Staphylococcus aureus is a leading cause of skin and soft-tissue infections worldwide. Mice are the most commonly used animals for modeling human staphylococcal infections. However a supra-physiologic S. aureus inoculum is required to establish gross murine skin pathology. Moreover, many staphylococcal factors, including Panton-Valentine leukocidin (PVL) elaborated by community-associated methicillin-resistant S. aureus (CA-MRSA), exhibit selective human tropism and cannot be adequately studied in mice. To overcome these deficiencies, we investigated S. aureus infection in non-obese diabetic (NOD)/severe combined immune deficiency (SCID)/IL2rγ (NSG) mice engrafted with human CD34+ umbilical cord blood cells. These "humanized" NSG mice require one to two log lower inoculum to induce consistent skin lesions compared with control mice, and exhibit larger cutaneous lesions upon infection with PVL+ versus isogenic PVL- S. aureus. Neutrophils appear important for PVL pathology as adoptive transfer of human neutrophils alone to NSG mice was sufficient to induce dermonecrosis following challenge with PVL+ S. aureus but not PVL- S. aureus. PMX53, a human C5aR inhibitor, blocked PVL-induced cellular cytotoxicity in vitro and reduced the size difference of lesions induced by the PVL+ and PVL- S. aureus, but PMX53 also reduced recruitment of neutrophils and exacerbated the infection. Overall, our findings establish humanized mice as an important translational tool for the study of S. aureus infection and provide strong evidence that PVL is a human virulence factor.

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PMX53 reduces the size differences of lesions induced by WT and PVL-S. aureus.Humanized NSG mice were injected i.p. with PBS or PMX53 (5 mg/kg/d). One hour later, the mice were infected on the left flank with ~2 x 106 WT S. aureus and on the right flank with ~2 x 106 CFU PVL- isogenic S. aureus. PBS or PMX53 was subsequently administered once daily. (A) Skin lesions and (B) CFU were measured on d 3. Note 2 mice treated with PMX53 died of an unknown cause prior to d3 and were not included in the analyses. (C—F) Humanized NSG mice were infected with ~2 x 106 WT S. aureus and treated 3 h later and then daily with PBS or PMX53 (5 mg/kg/d) i.p. (C) Skin lesion sizes and (D) bacterial burden on d 3. (E) IL8 release and (F) MPO at the site of infection after 24 h. Red bar = mean, *: p <0.05, **: p <0.01.
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ppat.1005292.g005: PMX53 reduces the size differences of lesions induced by WT and PVL-S. aureus.Humanized NSG mice were injected i.p. with PBS or PMX53 (5 mg/kg/d). One hour later, the mice were infected on the left flank with ~2 x 106 WT S. aureus and on the right flank with ~2 x 106 CFU PVL- isogenic S. aureus. PBS or PMX53 was subsequently administered once daily. (A) Skin lesions and (B) CFU were measured on d 3. Note 2 mice treated with PMX53 died of an unknown cause prior to d3 and were not included in the analyses. (C—F) Humanized NSG mice were infected with ~2 x 106 WT S. aureus and treated 3 h later and then daily with PBS or PMX53 (5 mg/kg/d) i.p. (C) Skin lesion sizes and (D) bacterial burden on d 3. (E) IL8 release and (F) MPO at the site of infection after 24 h. Red bar = mean, *: p <0.05, **: p <0.01.

Mentions: To address the pathogenic role of human C5aR-PVL interaction in vivo and test the efficacy of C5aR blockade as a treatment for S. aureus skin infection, we next infected humanized NSG mice with WT and isogenic PVL- mutant S. aureus strains, and treated the mice either 1 h prior or 3 h after infection with PMX53. Based on several published studies, daily systemic injection of PMX53 at 1 mg/kg effectively blocked various disease manifestations attributable to C5a, but has well-documented immunosuppressive effects [25,26]. Because a relatively high concentration of PMX53 was required to block PVL cytolytic activity in vitro, we selected a PMX53 dose of 5 mg/kg i.p given once daily. As shown in Fig 5A, administration of PMX53 versus PBS prior to infection did not improve skin lesion severity in mice infected with WT PVL+S. aureus, but reduced the lesion size difference between the PVL+ and PVL- infection groups, suggesting blocking of PVL effect by PMX53. For skin infection induced with the isogenic PVL-S. aureus, PMX53 treatment increased lesion sizes, indicating a potential detrimental effect of neutralizing C5aR-C5a interaction.


Increased Susceptibility of Humanized NSG Mice to Panton-Valentine Leukocidin and Staphylococcus aureus Skin Infection.

Tseng CW, Biancotti JC, Berg BL, Gate D, Kolar SL, Müller S, Rodriguez MD, Rezai-Zadeh K, Fan X, Beenhouwer DO, Town T, Liu GY - PLoS Pathog. (2015)

PMX53 reduces the size differences of lesions induced by WT and PVL-S. aureus.Humanized NSG mice were injected i.p. with PBS or PMX53 (5 mg/kg/d). One hour later, the mice were infected on the left flank with ~2 x 106 WT S. aureus and on the right flank with ~2 x 106 CFU PVL- isogenic S. aureus. PBS or PMX53 was subsequently administered once daily. (A) Skin lesions and (B) CFU were measured on d 3. Note 2 mice treated with PMX53 died of an unknown cause prior to d3 and were not included in the analyses. (C—F) Humanized NSG mice were infected with ~2 x 106 WT S. aureus and treated 3 h later and then daily with PBS or PMX53 (5 mg/kg/d) i.p. (C) Skin lesion sizes and (D) bacterial burden on d 3. (E) IL8 release and (F) MPO at the site of infection after 24 h. Red bar = mean, *: p <0.05, **: p <0.01.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4664407&req=5

ppat.1005292.g005: PMX53 reduces the size differences of lesions induced by WT and PVL-S. aureus.Humanized NSG mice were injected i.p. with PBS or PMX53 (5 mg/kg/d). One hour later, the mice were infected on the left flank with ~2 x 106 WT S. aureus and on the right flank with ~2 x 106 CFU PVL- isogenic S. aureus. PBS or PMX53 was subsequently administered once daily. (A) Skin lesions and (B) CFU were measured on d 3. Note 2 mice treated with PMX53 died of an unknown cause prior to d3 and were not included in the analyses. (C—F) Humanized NSG mice were infected with ~2 x 106 WT S. aureus and treated 3 h later and then daily with PBS or PMX53 (5 mg/kg/d) i.p. (C) Skin lesion sizes and (D) bacterial burden on d 3. (E) IL8 release and (F) MPO at the site of infection after 24 h. Red bar = mean, *: p <0.05, **: p <0.01.
Mentions: To address the pathogenic role of human C5aR-PVL interaction in vivo and test the efficacy of C5aR blockade as a treatment for S. aureus skin infection, we next infected humanized NSG mice with WT and isogenic PVL- mutant S. aureus strains, and treated the mice either 1 h prior or 3 h after infection with PMX53. Based on several published studies, daily systemic injection of PMX53 at 1 mg/kg effectively blocked various disease manifestations attributable to C5a, but has well-documented immunosuppressive effects [25,26]. Because a relatively high concentration of PMX53 was required to block PVL cytolytic activity in vitro, we selected a PMX53 dose of 5 mg/kg i.p given once daily. As shown in Fig 5A, administration of PMX53 versus PBS prior to infection did not improve skin lesion severity in mice infected with WT PVL+S. aureus, but reduced the lesion size difference between the PVL+ and PVL- infection groups, suggesting blocking of PVL effect by PMX53. For skin infection induced with the isogenic PVL-S. aureus, PMX53 treatment increased lesion sizes, indicating a potential detrimental effect of neutralizing C5aR-C5a interaction.

Bottom Line: Staphylococcus aureus is a leading cause of skin and soft-tissue infections worldwide.However a supra-physiologic S. aureus inoculum is required to establish gross murine skin pathology.PMX53, a human C5aR inhibitor, blocked PVL-induced cellular cytotoxicity in vitro and reduced the size difference of lesions induced by the PVL+ and PVL- S. aureus, but PMX53 also reduced recruitment of neutrophils and exacerbated the infection.

View Article: PubMed Central - PubMed

Affiliation: Division of Pediatric Infectious Diseases and the Immunobiology Research Institute, Cedars-Sinai Medical Center, Los Angeles, California, United States of America.

ABSTRACT
Staphylococcus aureus is a leading cause of skin and soft-tissue infections worldwide. Mice are the most commonly used animals for modeling human staphylococcal infections. However a supra-physiologic S. aureus inoculum is required to establish gross murine skin pathology. Moreover, many staphylococcal factors, including Panton-Valentine leukocidin (PVL) elaborated by community-associated methicillin-resistant S. aureus (CA-MRSA), exhibit selective human tropism and cannot be adequately studied in mice. To overcome these deficiencies, we investigated S. aureus infection in non-obese diabetic (NOD)/severe combined immune deficiency (SCID)/IL2rγ (NSG) mice engrafted with human CD34+ umbilical cord blood cells. These "humanized" NSG mice require one to two log lower inoculum to induce consistent skin lesions compared with control mice, and exhibit larger cutaneous lesions upon infection with PVL+ versus isogenic PVL- S. aureus. Neutrophils appear important for PVL pathology as adoptive transfer of human neutrophils alone to NSG mice was sufficient to induce dermonecrosis following challenge with PVL+ S. aureus but not PVL- S. aureus. PMX53, a human C5aR inhibitor, blocked PVL-induced cellular cytotoxicity in vitro and reduced the size difference of lesions induced by the PVL+ and PVL- S. aureus, but PMX53 also reduced recruitment of neutrophils and exacerbated the infection. Overall, our findings establish humanized mice as an important translational tool for the study of S. aureus infection and provide strong evidence that PVL is a human virulence factor.

Show MeSH
Related in: MedlinePlus