Limits...
Diethylcarbamazine activity against Brugia malayi microfilariae is dependent on inducible nitric-oxide synthase and the cyclooxygenase pathway.

McGarry HF, Plant LD, Taylor MJ - Filaria J (2005)

Bottom Line: Pre-treatment of animals with dexamethasone or indomethacin reduced DEC's efficacy by almost 90% or 56%, respectively, supporting a role for the arachidonic acid and cyclooxygenase pathways in vivo.Furthermore, experiments showed that treatment with DEC results in a reduction in the amount of COX-1 protein in peritoneal exudate cells.These results confirm the important role of the arachidonic acid metabolic pathway in DEC's mechanism of action in vivo and show that in addition to its effects on the 5-lipoxygenase pathway, it targets the cyclooxygenase pathway and COX-1.

View Article: PubMed Central - HTML - PubMed

Affiliation: Filariasis Research Laboratory, Molecular and Biochemical Parasitology, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK. hfcross@liverpool.ac.uk

ABSTRACT

Background: Diethylcarbamazine (DEC) has been used for many years in the treatment of human lymphatic filariasis. Its mode of action is not well understood, but it is known to interact with the arachidonic acid pathway. Here we have investigated the contribution of the nitric oxide and cyclooxygenase (COX) pathways to the activity of DEC against B. malayi microfilariae in mice.

Methods: B. malayi microfilariae were injected intravenously into mice and parasitaemia was measured 24 hours later. DEC was then administered to BALB/c mice with and without pre-treatment with indomethacin or dexamethasone and the parasitaemia monitored. To investigate a role for inducible nitric oxide in DEC's activity, DEC and ivermectin were administered to microfilaraemic iNOS-/- mice and their background strain (129/SV). Western blot analysis was used to determine any effect of DEC on the production of COX and inducible nitric-oxide synthase (iNOS) proteins.

Results: DEC administered alone to BALB/c mice resulted in a rapid and profound reduction in circulating microfilariae within five minutes of treatment. Microfilarial levels began to recover after 24 hours and returned to near pre-treatment levels two weeks later, suggesting that the sequestration of microfilariae occurs independently of parasite killing. Pre-treatment of animals with dexamethasone or indomethacin reduced DEC's efficacy by almost 90% or 56%, respectively, supporting a role for the arachidonic acid and cyclooxygenase pathways in vivo. Furthermore, experiments showed that treatment with DEC results in a reduction in the amount of COX-1 protein in peritoneal exudate cells. Additionally, in iNOS-/- mice infected with B. malayi microfilariae, DEC showed no activity, whereas the efficacy of another antifilarial drug, ivermectin, was unaffected.

Conclusion: These results confirm the important role of the arachidonic acid metabolic pathway in DEC's mechanism of action in vivo and show that in addition to its effects on the 5-lipoxygenase pathway, it targets the cyclooxygenase pathway and COX-1. Moreover, we show for the first time that inducible nitric oxide is essential for the rapid sequestration of microfilariae by DEC.

No MeSH data available.


Related in: MedlinePlus

Western blot detection of COX-1 protein from peritoneal exudate cells. COX-1 protein was detected in 129/SV and iNOS-/- peritoneal exudate cells thirty minutes after i.p. injection of endotoxin-free water (control) or DEC (10 mg/kg). Proteins (10 μg) were separated on a 7.5% denaturing SDS polyacrylamide gel, transferred to PVDF membrane, incubated with rabbit anti-mouse COX-1, then goat anti-rabbit IgG-horse radish peroxidase conjugate and detected by chemiluminescence.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC1173132&req=5

Figure 4: Western blot detection of COX-1 protein from peritoneal exudate cells. COX-1 protein was detected in 129/SV and iNOS-/- peritoneal exudate cells thirty minutes after i.p. injection of endotoxin-free water (control) or DEC (10 mg/kg). Proteins (10 μg) were separated on a 7.5% denaturing SDS polyacrylamide gel, transferred to PVDF membrane, incubated with rabbit anti-mouse COX-1, then goat anti-rabbit IgG-horse radish peroxidase conjugate and detected by chemiluminescence.

Mentions: Thirty minutes after administration of endotoxin-free water to 129/SV and iNOS-/- mice, peritoneal exudate cells were expressing COX-1 protein, whereas those from DEC-exposed animals contained markedly less COX-1 (Fig. 4). Interestingly, there seemed to be a higher level of COX-1 remaining in the iNOS-/- than the 129/SV macrophages after DEC treatment. Neither COX-2 nor iNOS protein was detected in any of the 129/SV or iNOS-/- groups (not shown).


Diethylcarbamazine activity against Brugia malayi microfilariae is dependent on inducible nitric-oxide synthase and the cyclooxygenase pathway.

McGarry HF, Plant LD, Taylor MJ - Filaria J (2005)

Western blot detection of COX-1 protein from peritoneal exudate cells. COX-1 protein was detected in 129/SV and iNOS-/- peritoneal exudate cells thirty minutes after i.p. injection of endotoxin-free water (control) or DEC (10 mg/kg). Proteins (10 μg) were separated on a 7.5% denaturing SDS polyacrylamide gel, transferred to PVDF membrane, incubated with rabbit anti-mouse COX-1, then goat anti-rabbit IgG-horse radish peroxidase conjugate and detected by chemiluminescence.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC1173132&req=5

Figure 4: Western blot detection of COX-1 protein from peritoneal exudate cells. COX-1 protein was detected in 129/SV and iNOS-/- peritoneal exudate cells thirty minutes after i.p. injection of endotoxin-free water (control) or DEC (10 mg/kg). Proteins (10 μg) were separated on a 7.5% denaturing SDS polyacrylamide gel, transferred to PVDF membrane, incubated with rabbit anti-mouse COX-1, then goat anti-rabbit IgG-horse radish peroxidase conjugate and detected by chemiluminescence.
Mentions: Thirty minutes after administration of endotoxin-free water to 129/SV and iNOS-/- mice, peritoneal exudate cells were expressing COX-1 protein, whereas those from DEC-exposed animals contained markedly less COX-1 (Fig. 4). Interestingly, there seemed to be a higher level of COX-1 remaining in the iNOS-/- than the 129/SV macrophages after DEC treatment. Neither COX-2 nor iNOS protein was detected in any of the 129/SV or iNOS-/- groups (not shown).

Bottom Line: Pre-treatment of animals with dexamethasone or indomethacin reduced DEC's efficacy by almost 90% or 56%, respectively, supporting a role for the arachidonic acid and cyclooxygenase pathways in vivo.Furthermore, experiments showed that treatment with DEC results in a reduction in the amount of COX-1 protein in peritoneal exudate cells.These results confirm the important role of the arachidonic acid metabolic pathway in DEC's mechanism of action in vivo and show that in addition to its effects on the 5-lipoxygenase pathway, it targets the cyclooxygenase pathway and COX-1.

View Article: PubMed Central - HTML - PubMed

Affiliation: Filariasis Research Laboratory, Molecular and Biochemical Parasitology, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool L3 5QA, UK. hfcross@liverpool.ac.uk

ABSTRACT

Background: Diethylcarbamazine (DEC) has been used for many years in the treatment of human lymphatic filariasis. Its mode of action is not well understood, but it is known to interact with the arachidonic acid pathway. Here we have investigated the contribution of the nitric oxide and cyclooxygenase (COX) pathways to the activity of DEC against B. malayi microfilariae in mice.

Methods: B. malayi microfilariae were injected intravenously into mice and parasitaemia was measured 24 hours later. DEC was then administered to BALB/c mice with and without pre-treatment with indomethacin or dexamethasone and the parasitaemia monitored. To investigate a role for inducible nitric oxide in DEC's activity, DEC and ivermectin were administered to microfilaraemic iNOS-/- mice and their background strain (129/SV). Western blot analysis was used to determine any effect of DEC on the production of COX and inducible nitric-oxide synthase (iNOS) proteins.

Results: DEC administered alone to BALB/c mice resulted in a rapid and profound reduction in circulating microfilariae within five minutes of treatment. Microfilarial levels began to recover after 24 hours and returned to near pre-treatment levels two weeks later, suggesting that the sequestration of microfilariae occurs independently of parasite killing. Pre-treatment of animals with dexamethasone or indomethacin reduced DEC's efficacy by almost 90% or 56%, respectively, supporting a role for the arachidonic acid and cyclooxygenase pathways in vivo. Furthermore, experiments showed that treatment with DEC results in a reduction in the amount of COX-1 protein in peritoneal exudate cells. Additionally, in iNOS-/- mice infected with B. malayi microfilariae, DEC showed no activity, whereas the efficacy of another antifilarial drug, ivermectin, was unaffected.

Conclusion: These results confirm the important role of the arachidonic acid metabolic pathway in DEC's mechanism of action in vivo and show that in addition to its effects on the 5-lipoxygenase pathway, it targets the cyclooxygenase pathway and COX-1. Moreover, we show for the first time that inducible nitric oxide is essential for the rapid sequestration of microfilariae by DEC.

No MeSH data available.


Related in: MedlinePlus