Complement is required for the induction of endotoxic fever in guinea pigs and mice

(1) Cobra venom factor (CVF)-induced hypocomplementemia dose-dependently attenuates the febrile responses of guinea pigs and mice to intraperitoneally (ip) but not to intravenously (iv) injected endotoxic bacterial lipopolysaccharide (LPS). (2) Iv but not ip LPS causes fever in complement component 3 (C3) gene-ablated mice, but neither iv nor ip LPS evokes a body core temperature (T_c) rise when WT and these mice's C5a receptors type 1 are blocked. C5 knockout mice also do not develop fever following either iv or ip LPS. C5a thus appears to be a critical mediator of LPS fever. (3) C5 knockouts develop fever in response to intracerebroventricularly (icv) injected LPS or prostaglandin (PG)E₂; the site of action of C5a is therefore peripheral rather than central. (4) The initiation of the febrile responses to both iv and ip LPS is temporally correlated with the appearance of LPS in the liver's Kupffer cells (Kc). (5) PGE₂ is released by liver in immediate response to the injection of CVF into the portal vein of anesthetized guinea pigs; its level rises quickly to its maximum. LPS injected similarly also evokes a quick release of PGE₂ from the liver; it, however, is prevented by prior hypocomplementation. (6) Neither LPS nor IL-1β induces PGE₂ release from Kc in vitro within the first hour after treatment, but serum C and C plus LPS or IL-1β very quickly trigger PGE₂ increases of similar magnitudes, catalyzed non-differentially by cyclooxygenase (COX)-1 and COX-2. Kc would thus appear to be the principal site of action of C5a, inducing the release of PGE₂. (7) PGE₂ is detectable in the plasma of conscious guinea pigs in temporal correlation with the onset of the T_c rise following ip LPS; cytokines appear significantly later. (8) Taken together, these results indicate that LPS-activated C, rather than LPS or IL-1β by itself, triggers PGE₂ release by Kc. This PGE₂ could be the factor that stimulates vagal afferents, thereby providing the signal to the brain that mediates the febrile response.     

Comparison of the two different approaches of orchiectomy in guinea pigs]

The results of orchiectomy conducted by the abdominal and scrotum routes were compared in guinea pigs. The operation time, surgical complication and incision healing were evaluated. About operation time, there was no distinct difference between two routes. In the abdominal route, the prognosis of this case was favorable. Also the healing of an incised wound was uneventful one week after the operation. While in the scrotum route, the tunica albuginea testis and the visceral vaginal tunic are thin, so that the risks of injuring the testis and/or scattering the contents of testis seem to be high. Inflammation of the scrotum was observed in 4 out of 5 guinea pigs, that were operated by scrotum route. From these results, the abdominal route method is highly recommended in the orchiectomy of guinea pigs.     

DNA augments antigenicity of mycobacterial DNA-binding protein 1 and confers protection against Mycobacterium tuberculosis infection in mice

Mycobacterium consists up to 7% of mycobacterial DNA-binding protein 1 (MDP1) in total cellular proteins. Host immune responses to MDP1 were studied in mice to explore the antigenic properties of this protein. Anti-MDP1 IgG was produced after infection with either bacillus Calmette-Guérin or Mycobacterium tuberculosis in C3H/HeJ mice. However, the level of Ab was remarkably low when purified MDP1 was injected. MDP1 is considered to be associated with DNA in nucleoid, which contains immunostimulatory CpG motif. Therefore, we examined coadministration of MDP1 and DNA derived from M. tuberculosis. Consequently, this procedure significantly enhanced the production of MDP1-specific IgG. Five nanograms of DNA was enough to enhance MDP1-specific IgG production in the administration of 5 microg of MDP1 into mice. Strong immune stimulation by such a small amount of DNA is noteworthy, because >1,000- to 100,000-fold doses of CpG DNAs are used for immune activation. A synthetic peptide-based study showed that B cell epitopes were different between mice administered MDP1 alone and those given a mixture of MDP1 and DNA, suggesting that DNA alters the three-dimensional structure of MDP1. Coadministration of DNA also enhanced MDP1-specific IFN-gamma production and reduced the bacterial burden of a following challenge of M. tuberculosis, showing that MDP1 is a novel vaccine target. Finally, we found that MDP1 remarkably enhanced TLR9-dependent immune stimulation by unmethylated CpG oligo DNA in vitro. To our knowledge, MDP1 is the first protein discovered that remarkably augments the CpG-mediated immune response and is a potential adjuvant for CpG DNA-based immune therapies.     

Comparison of bcg strains for delayed hypersensitivity induction tested in vivo in guinea pigs and in vitro

Previous studies performed on guinea pigs demonstrated a direct dependence of tuberculin reaction size (in vivo hypersensitivity) on immunogenicity in a number of BCG strains. The present work used an in vitro method, MIF detection, for assessing hypersensitivity and compared the results obtained with tuberculin hypersensitivity tests, correlating the data with the immunogenicity of the individual BCG strains employed. The following strains were used: the Czechoslovak BCG strain No. 725, Japanese BCG strain Tokyo, Danish BCG strain Copenhagen and Soviet BCG strain Moscow. The results obtained by the two hypersensitivity testing methods, in vivo and in vitro were in a direct correlation; a direct relationship was also demonstrated between hypersensitivity tested by the in vitro method and immunogenicity. The in vitro method of MIF detection is reproducible and comparable with the other two methods employed and may be used as an alternative approach to BCG vaccine efficacy testing. It might probably also be applicable to estimation of the status of cell-mediated immunity against intracellularly parasitizing bacteria in general.     

Ribosomal dysentery vaccine. II. Biological trials of active protection for guinea pigs and mice]

Ribosomal vaccine from Sh. sonnei injected subcutaneously once or twice in physiological saline or in Freund's complete adjuvant produces a marked protective effect against experimental keratoconjunctivitis in guinea pigs. Inhibition of the protective effect by high doses (above 100 microgram) of ribosomal vaccine is evident after a single, but not repeated injections. Protective effect in mice is achieved by immunization with very low doses of ribosomal vaccine: ED50 is 1.2 ng after challenge with 5.6 LD50. The nature of immunogenic factor responsible for the biological activity of the ribosome vaccine is still obscure. In contrast to Boivin's antigen, ribosomal preparations, even in high doses (1000--2000 microgram), have no toxic effect on mice and guinea pigs.     

Vaccine induced HSV 1 antibodies fail to correlate with protection against HSV 2 in guinea pigs

Guinea pigs immunised with HSV 1 subunit vaccine were bled, and subsequently challenged intravaginally with HSV 2. The clinical response to virus challenge was quantified, and correlations were sought between clinical score and virus-specific serum antibody titre in functional and binding assays (ELISA, neutralization, complement-mediated cytotoxicity and antibody-dependent cellular cytotoxicity). No significant relationships were found, and it was concluded that reactivity in the serological assays chosen did not correlate with protection against HSV 2 genital infection in vaccinated female guinea pigs.     

Vaccine induced HSV 1 antibodies fail to correlate with protection against HSV 2 in guinea pigs

Abstract Guinea pigs immunised with HSV 1 subunit vaccine were bled, and subsequently challenged intravaginally with HSV 2. The clinical response to virus challenge was quantified, and correlations were sought between clinical score and virus-specific serum antibody titre in functional and binding assays (ELISA, neutralization, complement-mediated cytotoxicity and antibody-dependent cellular cytotoxicity). No significant relationships were found, and it was concluded that reactivity in the serological assays chosen did not correlate with protection against HSV 2 genital infection in vaccinated female guinea pigs.     

DNA degradation in the kidney of folic acid-treated guinea pigs

Previous investigators agree on the increased DNA synthesis and destruction of tissues caused by folic acid (FA) administered parenterally. This study aims to clarify whether DNA degradation due to the destruction of cells and nuclei precedes DNA synthesis following FA administration. Forty guinea pigs were divided into four groups: group 1, contained 10 controls; in group 2, ten animals received intraperitoneally 300 mg/kg of body wt FA; in group 3, ten animals received FA and 12 h later frusemide intramuscularly in a dose of 7 mg/kg body wt; and finally in group 4, ten animals received frusemide as in group 3. FA produced necrosis of the epithelial cells of the convoluted tubules as the detection of the beta-aminoisobutyric acid end product of DNA and thymine catabolism indicated. Frusemide administered in group 3 had a favourable effect on the acute renal failure induced by FA.     

Factors influencing protection against experimental tuberculosis in mice by heat-stable cell wall vaccines

Ribi, E. (Rocky Mountain Laboratory, Hamilton, Mont.), R. L. Anacker, W. Brehmer, G. Goode, C. L. Larson, R. H. List, K. C. Milner, and W. C. Wicht. Factors influencing protection against experimental tuberculosis in mice by heat-stable cell wall vaccines. J. Bacteriol. 92:869-879. 1966.-Studies of nonviable, heat-stable vaccines for active protection against experimental tuberculosis have been continued with a test involving aerosol challenge of intravenously vaccinated mice. The previously reported activating effect of light mineral oil on disrupted cells of the BCG strain was found to be shared by certain other mineral oils and a synthetic, 24-carbon hydrocarbon, but not by kerosene or any of several vegetable oils. Dry cell walls coated with a small amount of oil and dispersed in saline with aid of an emulsifier were suitable for intravenous administration and were effective in promoting resistance to challenge. Oil used in this manner, in contrast to water-in-oil emulsions of the Freund type which could not be administered intravenously, did not potentiate the tuberculin-sensitizing activity of the cell walls. Although the amount of oil required for full effect was small (< 0.5 ml/100 mg of dry antigen), there was a critical level below which optimal enhancement was not achieved. More stable suspensions than could be obtained with the other oils were readily prepared from cell walls treated with the synthetic hydrocarbon, 7-n-hexyloctadecane. Extended experience has shown that in this test system both the viable BCG standard vaccine and heated, oil-treated experimental vaccines gave highly reproducible results showing graded responses to graded doses.