Limits...
Proteophosophoglycans regurgitated by Leishmania-infected sand flies target the L-arginine metabolism of host macrophages to promote parasite survival.

Rogers M, Kropf P, Choi BS, Dillon R, Podinovskaia M, Bates P, Müller I - PLoS Pathog. (2009)

Bottom Line: This early exacerbation was attributed to two fundamental properties of PSG: Firstly, PSG powerfully recruited macrophages to the dermal site of infection within 24 hours.Secondly, PSG enhanced alternative activation and arginase activity of host macrophages, thereby increasing L-arginine catabolism and the synthesis of polyamines essential for intracellular parasite growth.The increase in arginase activity promoted the intracellular growth of L. mexicana within classically activated macrophages, and inhibition of macrophage arginase completely ablated the early exacerbatory properties of PSG in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Imperial College of Science, Technology and Medicine, London, UK. matthew.rogers@imperial.ac.uk

ABSTRACT
All natural Leishmania infections start in the skin; however, little is known of the contribution made by the sand fly vector to the earliest events in mammalian infection, especially in inflamed skin that can rapidly kill invading parasites. During transmission sand flies regurgitate a proteophosphoglycan gel synthesized by the parasites inside the fly midgut, termed promastigote secretory gel (PSG). Regurgitated PSG can exacerbate cutaneous leishmaniasis. Here, we show that the amount of Leishmania mexicana PSG regurgitated by Lutzomyia longipalpis sand flies is proportional to the size of its original midgut infection and the number of parasites transmitted. Furthermore, PSG could exacerbate cutaneous L. mexicana infection for a wide range of doses (10-10,000 parasites) and enhance infection by as early as 48 hours in inflamed dermal air pouches. This early exacerbation was attributed to two fundamental properties of PSG: Firstly, PSG powerfully recruited macrophages to the dermal site of infection within 24 hours. Secondly, PSG enhanced alternative activation and arginase activity of host macrophages, thereby increasing L-arginine catabolism and the synthesis of polyamines essential for intracellular parasite growth. The increase in arginase activity promoted the intracellular growth of L. mexicana within classically activated macrophages, and inhibition of macrophage arginase completely ablated the early exacerbatory properties of PSG in vitro and in vivo. Thus, PSG is an essential component of the infectious sand fly bite for the early establishment of Leishmania in skin, which should be considered when designing and screening therapies against leishmaniasis.

Show MeSH

Related in: MedlinePlus

PSG recruits macrophages to the site of transmission and enhances their infection with L. mexicana.(A–C) Cellular content within air pouches inflated on the backs of BALB/c mice. Seventy two hour kinetic of neutrophil (A) and macrophage (B) recruitment in response to 1×103 L. mexicana metacyclic promastigotes co-injected with 1 µg L. mexicana PSG and 1 µg Lu. longipalpis saliva. (C) Neutrophil and macrophage content of air pouches in response to 1 µg PSG, 1 µg sand fly saliva or 1 µg of PSG and saliva after 24 hours. Cell type was determined by morphology of at least 200 Giemsa-stained cells per air pouch (4–6 mice per group). Data representative of duplicate (A&B) or triplicate (C) experiments. (D) Macrophages recovered from 48 hour PSG -loaded inflammatory air pouches were infected ex vivo at a MOI of 1∶1 for a further 48 hours. Inflammatory air pouches were generated by the injection of 200 U INFγ and 500 U TNFα 18 hours prior to the injection of PSG. Parasite burden (D) was determined by microscopy of at least 200 Giemsa-stained macrophages in quadruplicate. Representative experiment of duplicates is shown. Unless they are linked with a bar all test groups are compared to their relevant saline control; *, P<0.05; **, P<0.005; ***, P<0.0005.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2722086&req=5

ppat-1000555-g003: PSG recruits macrophages to the site of transmission and enhances their infection with L. mexicana.(A–C) Cellular content within air pouches inflated on the backs of BALB/c mice. Seventy two hour kinetic of neutrophil (A) and macrophage (B) recruitment in response to 1×103 L. mexicana metacyclic promastigotes co-injected with 1 µg L. mexicana PSG and 1 µg Lu. longipalpis saliva. (C) Neutrophil and macrophage content of air pouches in response to 1 µg PSG, 1 µg sand fly saliva or 1 µg of PSG and saliva after 24 hours. Cell type was determined by morphology of at least 200 Giemsa-stained cells per air pouch (4–6 mice per group). Data representative of duplicate (A&B) or triplicate (C) experiments. (D) Macrophages recovered from 48 hour PSG -loaded inflammatory air pouches were infected ex vivo at a MOI of 1∶1 for a further 48 hours. Inflammatory air pouches were generated by the injection of 200 U INFγ and 500 U TNFα 18 hours prior to the injection of PSG. Parasite burden (D) was determined by microscopy of at least 200 Giemsa-stained macrophages in quadruplicate. Representative experiment of duplicates is shown. Unless they are linked with a bar all test groups are compared to their relevant saline control; *, P<0.05; **, P<0.005; ***, P<0.0005.

Mentions: Sand fly saliva is known to attract macrophages to the site of infection [31]; however, the ability of PSG to recruit cells is unknown. To answer this question we tested the recruitment of cells to the site of PSG deposition in vivo using the air pouch model over a 4 to 72 hour period. Our results revealed that the combined presence of 1,000 L. mexicana metacyclic promastigotes, PSG and Lu. longipalpis saliva induced an early and significant attraction of neutrophils and macrophages to the skin compared to the injection of parasites in saline (Fig. 3A&B). In contrast, very few lymphocytes (<10%), eosinophils and basophils (<1%) were recruited to the air pouches under any of the experimental conditions used. By mimicking natural infection by sand fly bite in this way an enhanced cellular recruitment was observed as early as 4 hours post injection (neutrophils, Fig 3A: P<0.0001; macrophages, Fig. 3B: P = 0.0011) and was sustained for the remaining 72 hours. In the next step the contribution of the two main components delivered by sand fly bite to recruit cells, PSG and saliva, were determined alone or in combination. Injection of saline alone recruited mainly neutrophils to the air pouch, indicative of the needle injury to the skin. Compared to the saline group, injection of Lu. longipalpis saliva appeared to enhance this response by inducing a significant recruitment of both neutrophils and macrophages to the air pouch after 24 hours (Fig. 3C: neutrophils: P<0.0001; macrophages: P = 0.0007). The presence of PSG on the other hand proved to be highly efficient for attracting macrophages (P<0.0001), recruiting some 108-fold and 5-fold more macrophages to the air pouch compared to the saline control or saliva injected groups, respectively. When PSG and sand fly saliva were combined to represent the inoculum from a Leishmania-infected sand fly (without the parasites), this attracted the highest number of macrophages to the air pouch (224-fold more macrophages compared to saline controls, P<0.0001), indicating a synergy for macrophage chemotaxis.


Proteophosophoglycans regurgitated by Leishmania-infected sand flies target the L-arginine metabolism of host macrophages to promote parasite survival.

Rogers M, Kropf P, Choi BS, Dillon R, Podinovskaia M, Bates P, Müller I - PLoS Pathog. (2009)

PSG recruits macrophages to the site of transmission and enhances their infection with L. mexicana.(A–C) Cellular content within air pouches inflated on the backs of BALB/c mice. Seventy two hour kinetic of neutrophil (A) and macrophage (B) recruitment in response to 1×103 L. mexicana metacyclic promastigotes co-injected with 1 µg L. mexicana PSG and 1 µg Lu. longipalpis saliva. (C) Neutrophil and macrophage content of air pouches in response to 1 µg PSG, 1 µg sand fly saliva or 1 µg of PSG and saliva after 24 hours. Cell type was determined by morphology of at least 200 Giemsa-stained cells per air pouch (4–6 mice per group). Data representative of duplicate (A&B) or triplicate (C) experiments. (D) Macrophages recovered from 48 hour PSG -loaded inflammatory air pouches were infected ex vivo at a MOI of 1∶1 for a further 48 hours. Inflammatory air pouches were generated by the injection of 200 U INFγ and 500 U TNFα 18 hours prior to the injection of PSG. Parasite burden (D) was determined by microscopy of at least 200 Giemsa-stained macrophages in quadruplicate. Representative experiment of duplicates is shown. Unless they are linked with a bar all test groups are compared to their relevant saline control; *, P<0.05; **, P<0.005; ***, P<0.0005.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2722086&req=5

ppat-1000555-g003: PSG recruits macrophages to the site of transmission and enhances their infection with L. mexicana.(A–C) Cellular content within air pouches inflated on the backs of BALB/c mice. Seventy two hour kinetic of neutrophil (A) and macrophage (B) recruitment in response to 1×103 L. mexicana metacyclic promastigotes co-injected with 1 µg L. mexicana PSG and 1 µg Lu. longipalpis saliva. (C) Neutrophil and macrophage content of air pouches in response to 1 µg PSG, 1 µg sand fly saliva or 1 µg of PSG and saliva after 24 hours. Cell type was determined by morphology of at least 200 Giemsa-stained cells per air pouch (4–6 mice per group). Data representative of duplicate (A&B) or triplicate (C) experiments. (D) Macrophages recovered from 48 hour PSG -loaded inflammatory air pouches were infected ex vivo at a MOI of 1∶1 for a further 48 hours. Inflammatory air pouches were generated by the injection of 200 U INFγ and 500 U TNFα 18 hours prior to the injection of PSG. Parasite burden (D) was determined by microscopy of at least 200 Giemsa-stained macrophages in quadruplicate. Representative experiment of duplicates is shown. Unless they are linked with a bar all test groups are compared to their relevant saline control; *, P<0.05; **, P<0.005; ***, P<0.0005.
Mentions: Sand fly saliva is known to attract macrophages to the site of infection [31]; however, the ability of PSG to recruit cells is unknown. To answer this question we tested the recruitment of cells to the site of PSG deposition in vivo using the air pouch model over a 4 to 72 hour period. Our results revealed that the combined presence of 1,000 L. mexicana metacyclic promastigotes, PSG and Lu. longipalpis saliva induced an early and significant attraction of neutrophils and macrophages to the skin compared to the injection of parasites in saline (Fig. 3A&B). In contrast, very few lymphocytes (<10%), eosinophils and basophils (<1%) were recruited to the air pouches under any of the experimental conditions used. By mimicking natural infection by sand fly bite in this way an enhanced cellular recruitment was observed as early as 4 hours post injection (neutrophils, Fig 3A: P<0.0001; macrophages, Fig. 3B: P = 0.0011) and was sustained for the remaining 72 hours. In the next step the contribution of the two main components delivered by sand fly bite to recruit cells, PSG and saliva, were determined alone or in combination. Injection of saline alone recruited mainly neutrophils to the air pouch, indicative of the needle injury to the skin. Compared to the saline group, injection of Lu. longipalpis saliva appeared to enhance this response by inducing a significant recruitment of both neutrophils and macrophages to the air pouch after 24 hours (Fig. 3C: neutrophils: P<0.0001; macrophages: P = 0.0007). The presence of PSG on the other hand proved to be highly efficient for attracting macrophages (P<0.0001), recruiting some 108-fold and 5-fold more macrophages to the air pouch compared to the saline control or saliva injected groups, respectively. When PSG and sand fly saliva were combined to represent the inoculum from a Leishmania-infected sand fly (without the parasites), this attracted the highest number of macrophages to the air pouch (224-fold more macrophages compared to saline controls, P<0.0001), indicating a synergy for macrophage chemotaxis.

Bottom Line: This early exacerbation was attributed to two fundamental properties of PSG: Firstly, PSG powerfully recruited macrophages to the dermal site of infection within 24 hours.Secondly, PSG enhanced alternative activation and arginase activity of host macrophages, thereby increasing L-arginine catabolism and the synthesis of polyamines essential for intracellular parasite growth.The increase in arginase activity promoted the intracellular growth of L. mexicana within classically activated macrophages, and inhibition of macrophage arginase completely ablated the early exacerbatory properties of PSG in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology, Imperial College of Science, Technology and Medicine, London, UK. matthew.rogers@imperial.ac.uk

ABSTRACT
All natural Leishmania infections start in the skin; however, little is known of the contribution made by the sand fly vector to the earliest events in mammalian infection, especially in inflamed skin that can rapidly kill invading parasites. During transmission sand flies regurgitate a proteophosphoglycan gel synthesized by the parasites inside the fly midgut, termed promastigote secretory gel (PSG). Regurgitated PSG can exacerbate cutaneous leishmaniasis. Here, we show that the amount of Leishmania mexicana PSG regurgitated by Lutzomyia longipalpis sand flies is proportional to the size of its original midgut infection and the number of parasites transmitted. Furthermore, PSG could exacerbate cutaneous L. mexicana infection for a wide range of doses (10-10,000 parasites) and enhance infection by as early as 48 hours in inflamed dermal air pouches. This early exacerbation was attributed to two fundamental properties of PSG: Firstly, PSG powerfully recruited macrophages to the dermal site of infection within 24 hours. Secondly, PSG enhanced alternative activation and arginase activity of host macrophages, thereby increasing L-arginine catabolism and the synthesis of polyamines essential for intracellular parasite growth. The increase in arginase activity promoted the intracellular growth of L. mexicana within classically activated macrophages, and inhibition of macrophage arginase completely ablated the early exacerbatory properties of PSG in vitro and in vivo. Thus, PSG is an essential component of the infectious sand fly bite for the early establishment of Leishmania in skin, which should be considered when designing and screening therapies against leishmaniasis.

Show MeSH
Related in: MedlinePlus