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A recombinant vaccine effectively induces c5a-specific neutralizing antibodies and prevents arthritis.

Nandakumar KS, Jansson A, Xu B, Rydell N, Ahooghalandari P, Hellman L, Blom AM, Holmdahl R - PLoS ONE (2010)

Bottom Line: Sera from the vaccinated mice developed C5a-specific neutralizing antibodies, however C5 activation and formation of the membrane attack complex by C5b were not significantly altered.Exploitation of host immune response to generate sustained C5a neutralizing antibodies without significantly compromising C5/C5b activity is a useful strategy for developing an effective vaccine for antibody mediated and C5a dependent inflammatory diseases.Further developing of such a therapeutic vaccine would be more optimal and cost effective to attenuate inflammation without affecting host immunity.

View Article: PubMed Central - PubMed

Affiliation: Medical Inflammation Research, Department of Experimental Medicine, Lund University, Lund, Sweden. Nandakumar.kutty-selva@ki.se

ABSTRACT

Objectives: To develop and validate a recombinant vaccine to attenuate inflammation in arthritis by sustained neutralization of the anaphylatoxin C5a.

Methods: We constructed and expressed fusion protein of C5a and maltose binding protein. Efficacy of specific C5a neutralization was tested using the fusion protein as vaccine in three different arthritis mouse models: collagen induced arthritis (CIA), chronic relapsing CIA and collagen antibody induced arthritis (CAIA). Levels of anti-C5a antibodies and anti-collagen type II were measured by ELISA. C5a neutralization assay was done using a rat basophilic leukemia cell-line transfected with the human C5aR. Complement activity was determined using a hemolytic assay and joint morphology was assessed by histology.

Results: Vaccination of mice with MBP-C5a led to significant reduction of arthritis incidence and severity but not anti-collagen antibody synthesis. Histology of the MBP-C5a and control (MBP or PBS) vaccinated mice paws confirmed the vaccination effect. Sera from the vaccinated mice developed C5a-specific neutralizing antibodies, however C5 activation and formation of the membrane attack complex by C5b were not significantly altered.

Conclusions: Exploitation of host immune response to generate sustained C5a neutralizing antibodies without significantly compromising C5/C5b activity is a useful strategy for developing an effective vaccine for antibody mediated and C5a dependent inflammatory diseases. Further developing of such a therapeutic vaccine would be more optimal and cost effective to attenuate inflammation without affecting host immunity.

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Effect of MBP-C5a vaccination on chronic arthritis.Frequency (A), mean arthritis score (B) and levels of C5a-specific antibodies (C) were compared between MBP-C5a and PBS vaccinated groups. B10.Q (BALB/c x B10.Q)F2 mice of both gender (8 weeks old) were immunized with 100 µg of rat CII emulsified in IFA on day 0 at the base of the tail and boosted on day 35 with 50 µg of rat CII in IFA as indicated by arrows. The mice were scored for a period of 210 days for arthritis development. Mice that developed chronic arthritis and at arthritis remission were selected for the treatment experiment. All selected animals received vaccination subcutaneously of 100 µg of MBP-C5a or PBS emulsified in CFA on day 210 and were re-vaccinated on days 228 and 252 with 50 µg of MBP-C5a or PBS emulsified in IFA s.c. at the back. All the animals were re-immunized for synchronization of arthritis induction on day 231 with 50 µg of rat CII in IFA and scored the next 80 days for clinical signs of arthritis up to day 311 (from first day of CII immunization). Serum samples were collected on day 231 and 292. Statistical analysis for mean arthritis score was done on all days (d259 to d311) but the significance between groups was found only on days 275 and 279. All data represent mean ± SEM. *, p<0.05 and **, p<0.005. n, indicates number of mice in each group. All the mice were included for calculation of arthritis susceptibility and severity.
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pone-0013511-g005: Effect of MBP-C5a vaccination on chronic arthritis.Frequency (A), mean arthritis score (B) and levels of C5a-specific antibodies (C) were compared between MBP-C5a and PBS vaccinated groups. B10.Q (BALB/c x B10.Q)F2 mice of both gender (8 weeks old) were immunized with 100 µg of rat CII emulsified in IFA on day 0 at the base of the tail and boosted on day 35 with 50 µg of rat CII in IFA as indicated by arrows. The mice were scored for a period of 210 days for arthritis development. Mice that developed chronic arthritis and at arthritis remission were selected for the treatment experiment. All selected animals received vaccination subcutaneously of 100 µg of MBP-C5a or PBS emulsified in CFA on day 210 and were re-vaccinated on days 228 and 252 with 50 µg of MBP-C5a or PBS emulsified in IFA s.c. at the back. All the animals were re-immunized for synchronization of arthritis induction on day 231 with 50 µg of rat CII in IFA and scored the next 80 days for clinical signs of arthritis up to day 311 (from first day of CII immunization). Serum samples were collected on day 231 and 292. Statistical analysis for mean arthritis score was done on all days (d259 to d311) but the significance between groups was found only on days 275 and 279. All data represent mean ± SEM. *, p<0.05 and **, p<0.005. n, indicates number of mice in each group. All the mice were included for calculation of arthritis susceptibility and severity.

Mentions: To investigate whether MBP-C5a vaccination could be efficacious in the treatment of chronic arthritis, we immunized B10.Q (BALB/c × B10.Q) F2 mice with rat CII in IFA, which develops chronic relapsing arthritis [19]. These are genetically heterogeneous mice, and only mice with severe and active chronic relapsing arthritis for a minimum period of 120 days but at arthritis remission at day 210 were selected. For synchronization of relapse, we reimmunized them with CII emulsified in IFA on day 231. The mice were scored upto day 311 (Fig. 5A). All the mice developed arthritis in the control group, whereas a significant decrease in the incidence of arthritis (Fig. 5B) was observed in MBP-C5a vaccinated group. However, severity of arthritis was significantly reduced only on two different days (275 and 279), when all the mice were included for calculations. On the other hand, when we analyzed significance in severity of arthritis between the groups using sick mice only, the arthritis severity tended to be lower in the C5a treated group but reached significant difference only on day 265 (p = 0.0330). High levels of C5a-specific neutralizing antibodies could be detected in the C5a but not control vaccinated mice (Fig. 5C).


A recombinant vaccine effectively induces c5a-specific neutralizing antibodies and prevents arthritis.

Nandakumar KS, Jansson A, Xu B, Rydell N, Ahooghalandari P, Hellman L, Blom AM, Holmdahl R - PLoS ONE (2010)

Effect of MBP-C5a vaccination on chronic arthritis.Frequency (A), mean arthritis score (B) and levels of C5a-specific antibodies (C) were compared between MBP-C5a and PBS vaccinated groups. B10.Q (BALB/c x B10.Q)F2 mice of both gender (8 weeks old) were immunized with 100 µg of rat CII emulsified in IFA on day 0 at the base of the tail and boosted on day 35 with 50 µg of rat CII in IFA as indicated by arrows. The mice were scored for a period of 210 days for arthritis development. Mice that developed chronic arthritis and at arthritis remission were selected for the treatment experiment. All selected animals received vaccination subcutaneously of 100 µg of MBP-C5a or PBS emulsified in CFA on day 210 and were re-vaccinated on days 228 and 252 with 50 µg of MBP-C5a or PBS emulsified in IFA s.c. at the back. All the animals were re-immunized for synchronization of arthritis induction on day 231 with 50 µg of rat CII in IFA and scored the next 80 days for clinical signs of arthritis up to day 311 (from first day of CII immunization). Serum samples were collected on day 231 and 292. Statistical analysis for mean arthritis score was done on all days (d259 to d311) but the significance between groups was found only on days 275 and 279. All data represent mean ± SEM. *, p<0.05 and **, p<0.005. n, indicates number of mice in each group. All the mice were included for calculation of arthritis susceptibility and severity.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2958150&req=5

pone-0013511-g005: Effect of MBP-C5a vaccination on chronic arthritis.Frequency (A), mean arthritis score (B) and levels of C5a-specific antibodies (C) were compared between MBP-C5a and PBS vaccinated groups. B10.Q (BALB/c x B10.Q)F2 mice of both gender (8 weeks old) were immunized with 100 µg of rat CII emulsified in IFA on day 0 at the base of the tail and boosted on day 35 with 50 µg of rat CII in IFA as indicated by arrows. The mice were scored for a period of 210 days for arthritis development. Mice that developed chronic arthritis and at arthritis remission were selected for the treatment experiment. All selected animals received vaccination subcutaneously of 100 µg of MBP-C5a or PBS emulsified in CFA on day 210 and were re-vaccinated on days 228 and 252 with 50 µg of MBP-C5a or PBS emulsified in IFA s.c. at the back. All the animals were re-immunized for synchronization of arthritis induction on day 231 with 50 µg of rat CII in IFA and scored the next 80 days for clinical signs of arthritis up to day 311 (from first day of CII immunization). Serum samples were collected on day 231 and 292. Statistical analysis for mean arthritis score was done on all days (d259 to d311) but the significance between groups was found only on days 275 and 279. All data represent mean ± SEM. *, p<0.05 and **, p<0.005. n, indicates number of mice in each group. All the mice were included for calculation of arthritis susceptibility and severity.
Mentions: To investigate whether MBP-C5a vaccination could be efficacious in the treatment of chronic arthritis, we immunized B10.Q (BALB/c × B10.Q) F2 mice with rat CII in IFA, which develops chronic relapsing arthritis [19]. These are genetically heterogeneous mice, and only mice with severe and active chronic relapsing arthritis for a minimum period of 120 days but at arthritis remission at day 210 were selected. For synchronization of relapse, we reimmunized them with CII emulsified in IFA on day 231. The mice were scored upto day 311 (Fig. 5A). All the mice developed arthritis in the control group, whereas a significant decrease in the incidence of arthritis (Fig. 5B) was observed in MBP-C5a vaccinated group. However, severity of arthritis was significantly reduced only on two different days (275 and 279), when all the mice were included for calculations. On the other hand, when we analyzed significance in severity of arthritis between the groups using sick mice only, the arthritis severity tended to be lower in the C5a treated group but reached significant difference only on day 265 (p = 0.0330). High levels of C5a-specific neutralizing antibodies could be detected in the C5a but not control vaccinated mice (Fig. 5C).

Bottom Line: Sera from the vaccinated mice developed C5a-specific neutralizing antibodies, however C5 activation and formation of the membrane attack complex by C5b were not significantly altered.Exploitation of host immune response to generate sustained C5a neutralizing antibodies without significantly compromising C5/C5b activity is a useful strategy for developing an effective vaccine for antibody mediated and C5a dependent inflammatory diseases.Further developing of such a therapeutic vaccine would be more optimal and cost effective to attenuate inflammation without affecting host immunity.

View Article: PubMed Central - PubMed

Affiliation: Medical Inflammation Research, Department of Experimental Medicine, Lund University, Lund, Sweden. Nandakumar.kutty-selva@ki.se

ABSTRACT

Objectives: To develop and validate a recombinant vaccine to attenuate inflammation in arthritis by sustained neutralization of the anaphylatoxin C5a.

Methods: We constructed and expressed fusion protein of C5a and maltose binding protein. Efficacy of specific C5a neutralization was tested using the fusion protein as vaccine in three different arthritis mouse models: collagen induced arthritis (CIA), chronic relapsing CIA and collagen antibody induced arthritis (CAIA). Levels of anti-C5a antibodies and anti-collagen type II were measured by ELISA. C5a neutralization assay was done using a rat basophilic leukemia cell-line transfected with the human C5aR. Complement activity was determined using a hemolytic assay and joint morphology was assessed by histology.

Results: Vaccination of mice with MBP-C5a led to significant reduction of arthritis incidence and severity but not anti-collagen antibody synthesis. Histology of the MBP-C5a and control (MBP or PBS) vaccinated mice paws confirmed the vaccination effect. Sera from the vaccinated mice developed C5a-specific neutralizing antibodies, however C5 activation and formation of the membrane attack complex by C5b were not significantly altered.

Conclusions: Exploitation of host immune response to generate sustained C5a neutralizing antibodies without significantly compromising C5/C5b activity is a useful strategy for developing an effective vaccine for antibody mediated and C5a dependent inflammatory diseases. Further developing of such a therapeutic vaccine would be more optimal and cost effective to attenuate inflammation without affecting host immunity.

Show MeSH
Related in: MedlinePlus