Acute phase protein response in an experimental model of ovine caseous lymphadenitis.

BACKGROUND: Caseous lymphadenitis (CLA) is a disease of small ruminants caused by Corynebacterium pseudotuberculosis. The pathogenesis of CLA is a slow process, and produces a chronic rather than an acute disease state. Acute phase proteins (APP) such as haptoglobin (Hp) serum amyloid A (SAA) and alpha1 acid glycoprotein (AGP) are produced by the liver and released into the circulation in response to pro-inflammatory cytokines. The concentration of Hp in serum increases in experimental CLA but it is not known if SAA and AGP respond in parallel or have differing response profiles. RESULTS: The concentration in serum of Hp, SAA and AGP in 6 sheep challenged with 2 x 105 cells of C. pseudotuberculosis showed significant increases (P < 0.05) compared to 3 unchallenged control sheep. By day 7 post infection. (p.i.) the Hp and SAA concentrations reached mean (+/- SEM) values of 1.65 +/- 0.21 g/L and 18.1 +/- 5.2 mg/L respectively. Thereafter, their concentrations fell with no significant difference to those of the control sheep by day 18 p.i.. In contrast, the serum AGP concentration in infected sheep continued to rise to a peak of 0.38 +/- 0.05 g/L on day 13 p.i., after which a slow decline occurred, although the mean concentration remained significantly higher (P < 0.05) than the control group up to 29 days p.i.. Specific IgG to phospholidase D of C. pseudotuberculosis became detectable at 11 days p.i. and continued to rise throughout the experiment. CONCLUSION: The serum concentrations of Hp, SAA and AGP were raised in sheep in an experimental model of CLA. An extended response was found for AGP which occurred at a point when the infection was likely to have been transforming from an acute to a chronic phase. The results suggest that AGP could have a role as a marker for chronic conditions in sheep.

Effect of immunisation against gonadotrophin releasing hormone isoforms (mammalian GnRH-I, chicken GnRH-II and lamprey GnRH-III) on murine spermatogenesis.

In mammals, the hypothalamic decapeptide, gonadotrophin releasing hormone (GnRH-I), is regarded as the major fertility regulating peptide. However, a range of isoforms also exists, varying only in the core region between amino acids 5-8. The physiological role of two of these, GnRH-II and GnRH-III, remains controversial, particularly with regard to fertility. The basis of the present study was to examine whether there is potential for GnRH-II and GnRH-III to be developed into highly specific vaccines, and to determine what the impact of their neutralisation would be on fertility. Computer modelling was used to predict how many common amino acids could be sequentially removed from the N-terminus, without loss of conformational structure. Sequences predicted to retain structure, were synthesised and conjugated to tetanus toxoid. Male mice were actively immunised, in study weeks 0, 2, 4 and 6 and peptide specific ELISA carried out. Mice immunised with TT-GnRH-I, TT-GnRH-II and TT-GnRH-III conjugates induced high antibody titres to the respective peptide. However, serum from TT-GnRH-I treated mice showed cross-reactivity to GnRH-II and GnRH-III peptides, and serum from TT-GnRH-II immunised mice showed cross-reactivity to GnRH-III. On the other hand, serum from only two of the TT-GnRH-III treated animals showed cross-reactivity to GnRH-II. Histological examination of the testes enabled comparative quantification of the disruption to spermatogenesis. Immunisation against TT-GnRH-I and TT-GnRH-III caused 66% and 68%, respectively, of seminiferous tubules viewed to show evidence of spermatogenesis, compared with 82% and 92% against TT-GnRH-II and untreated controls, respectively. Endocrine analysis revealed that only the TT-GnRH-I immunised animals showed significant reduction (p<0.05) in follicle stimulating hormone, while testosterone levels were reduced in the TT-GnRH-I and TT-GnRH-III treated animals. Taken together, our data suggests that GnRH-I and GnRH-III are implicated in spermatogenesis, unlike GnRH-II.

Evaluation of two GnRH-I based vaccine formulations on the testes function of entire Suffolk cross ram lambs.

A modified GnRH peptide (CHWSYGLRPG-NH(2)) was conjugated to tetanus toxoid (TT) or diphtheria toxoid (DT) and formulated with Quil A saponin or a sustained release injectible PLGA (poly(lactide-co-glycolide)/triacetin). For the Quil A formulations, two administrations of TT conjugate at 3-weekly intervals were followed by two booster injections with the DT conjugate in entire ram lambs. With the PLGA formulations, only two injections were administered; the first containing TT and the second DT at 6-weekly intervals. Evaluation was carried out by comparing the specific antibody levels produced in relationship to hormone profiles and testicular changes. The Quil A formulation was considered the most effective, as it caused significant reduction in testosterone and follicle stimulating hormone levels, resulting in marked suppression of spermatogenesis.

Immune responses to a GnRH-based anti-fertility immunogen, induced by different adjuvants and subsequent effect on vaccine efficacy.

A modified GnRH peptide (CHWSYGLRPG-NH2) was conjugated to tetanus toxoid and formulated with different adjuvants (non-ionic surfactant vesicles, aluminium hydroxide, Quil A, PLGA (poly(lactide-co-glycolide)/triacetin), and Quil A/PLGA). A comparison of the anti-fertility efficacy of the formulations was made by examining specific antibody levels, antibody subclasses, endocrine ablation and gonadal atrophy. The production of IgG2b antibody provided the best correlation for castration. PLGA was considered the most effective adjuvant as it produced a consistent anti-fertility response in all the treated animals.