Correlation of antigenic expression with progress in antibiotic therapy of acute human brucellosis.

Human brucellosis is a zoonotic disease which is endemic in Saudi Arabia. The aim of this study was to investigate the humoral immune responses and identify the target antigens that persist at different stages in human brucellosis during antibiotic therapy. To do this, an acute case of accidental nosocomial infection was studied experimentally. Blood was collected from the patient at the time of diagnosis, and at weekly intervals during therapy until remission. IgG and IgM immunoblotting was used to characterize specific antigenic determinants, and ELISA antibody titration was performed to quantify the circulating antibodies. Results indicated that protein bands of 12-13.5 kDa bound IgG in the patient's sera but did not bind IgM on immunoblots and are probably not specific for, or important in, early stage infections. However, an 18 kDa band persisted during infection through remission. The pivotal and most important findings were that the number of protein bands seen on immunoblots, the magnitude of ELISA antibody titres and the concomitant changes in the intensity of the polypeptide bands of 42-43 kDa were positively correlated with the stage of infection. High numbers of anti-IgG and -IgM immunoblot bands coupled with high ELISA antibody titres and a concomitant increase in intensity of the 42-43 kDa bands were positively correlated with acute and severe infection. Conversely, a reduction in the number of polypeptide bands as well as a decrease in the intensity, until the complete disappearance of the 42-43 kDa bands, coupled with low (baseline) ELISA antibody titration values indicated successful treatment and remission. The routine use of the methods described here to ascertain the stage of the disease, assess the progress of antimicrobial therapy and monitor cases of relapse in human brucellosis is suggested.

A pilot study of short- and long-term sequelae to rigid fixation across metacarpal physes in a baboon model.

The use of rigid fixation for fractures of the extremities has become commonplace. The short- and long-term effects of rigid fixation on the growing hand, however, have not been studied thoroughly. In this project, the use of rigid fixation across metacarpal growth plates (physes) in growing primate hands was examined. The hypothesis to be tested was that long-term placement of rigid fixation devices across physes during stabilization of mid-shaft osteotomies will cause the physes to close prematurely. Fixation devices with screws placed in the epiphysis and left in place for 4 months or 1 year resulted in open physes, in support of the null hypothesis. However, in physes plated for 1 year, biochemical changes associated with increased bone differentiation were apparent. Findings suggest that rigid fixation across physes for as long as 1 year can be used appropriately in growing individuals when necessary. However, until additional investigation establishes whether the open physes are still capable of producing bone-lengthening hypertrophic chondrocytes, caution should be used in long-term placement of rigid fixation devices.

Interaction of baboon anti-alpha-galactosyl antibody with pig tissues.

As barriers to xenotransplantation are surmounted, such as suppression of hyperacute rejection allowing improved graft survival, it becomes important to define longer-term host-xenograft interactions. To this end we have prepared in baboons high titer anti-alpha-Galactosyl (alphaGal) and anti-porcine aortic endothelial cell antibodies, similar to human natural xenoantibodies and reactive with epitopes of thyroglobulin, laminin, and heparan sulfate proteoglycans. When injected into pigs with a protocol similar to that used in the rat to show the nephritogenic potential of heterologous anti-laminin and anti-heparan sulfate proteoglycan antibodies, baboon immunoglobulins bound first to renal vascular endothelium, and later to interstitial cells, especially fibroblasts and macrophages, and to antigens in basement membranes and extracellular matrix, where they colocalized with laminin- and heparan sulfate proteoglycan-antibodies, and with bound Griffonia simplicifolia B4. A similar binding was observed in other organs. The pigs did not develop an acute complement-dependent inflammation, but rather chronic lesions of the basement membranes and the extracellular matrix. Incubation of renal fibroblasts with baboon anti-alpha-Galactosyl antibodies resulted in increased synthesis of transforming growth factor-beta and collagen, suggesting a possible basis for the fibrotic response. The results demonstrate that in this experimental model a consequence of alphaGal antibody interaction with porcine tissues, is immunoreactivity with alphaGal on matrix molecules and interstitial cells, priming mechanisms leading to fibrosis resembling that in chronic allograft rejection. The possibility that similar lesions may develop in long-surviving pig xenografts is discussed.

Effect of alcohol on spleen cells and their functions in C57BL/6 mice.

Spleen cells from C57BL/6 mice maintained on alcohol containing liquid diet for two weeks were evaluated for different immune functions. On an average, 22% fewer spleen cells were recovered from alcohol-fed mice when compared to cells from control animals. In alcohol-fed mice, the relative frequency of B cells increased, whereas total T cells including CD4+ cells decreased significantly. Alcoholic mice, when challenged with poly(rI) poly(rC), produced significantly less interferon than control mice. In vitro production of interferon alpha and gamma by the spleen cells of alcoholic mice was reduced by 67-90%. No significant differences were seen in the level of natural killer cell activity in spleen cells of control and alcoholic mice. These results suggest that chronic alcohol intake can result in not only changes in the number of immune cells, but more importantly affect their biological functions such as their ability to produce interferons.

Acute toxicity and mutagenicity of the copper complex of pyruvaldehyde-bis (N-4-methylthiosemicarbazone), Cu-PTSM.

Cu-PTSM is a potential imaging agent for the heart and brain when labeled with either 64Cu or 62Cu. Unlabeled Cu-PTSM was evaluated for its acute toxicity and mutagenicity. Cu-PTSM had an i.v. LD50 of 26 mg kg-1 in the rat and 2 mg kg-1 in the rabbit. At necropsy, rats exhibited severely hemorrhagic lungs, histological findings of acute pulmonary congestion, hemorrhage and edema, and mild congestion in kidney, liver and brain. The rabbit displayed marked polymorphonuclear infiltration in alveoli, peribronchial and periarterial areas with marked macrophage hyperplasia, congestion and mild hemorrhage into alveolar spaces. No effects were found in kidney, liver, testes or brain. Administration of 2.16 micrograms kg-1 day-1 for 5 days per week for 2 weeks resulted in no changes in histopathology, hematology or clinical chemistry parameters. This daily dose is at least 300 times the diagnostic dose intended for use in man. Cu-PTSM was not mutagenic when tested in the absence of S9 supernatant, but elicited a weakly mutagenic response in the presence of S9. Since acute effects in the lung occur at doses approaching 300,000 times the diagnostic dose, it is highly unlikely that the clinical use of Cu-PTSM would result in any acute adverse effects.

Adoptive transfer of immunity to Treponema pallidum Nichols infection in inbred strain 2 and C4D guinea pigs.

T lymphocytes purified from lymph nodes and spleens of chancre-immune, inbred strain 2 guinea pigs, when infused into syngeneic guinea pigs, conferred protection against challenge with Treponema pallidum subsp. pallidum Nichols. No protection was conferred by similar injections of cell suspensions from normal guinea pigs or guinea pigs immunized with T. phagedenis biotype Reiter or T. pallidum-free testis supernatants from infected rabbits. Similar results were obtained with homozygous C4D guinea pigs. After several months of infection, 2 of 11 strain 2 and 1 of 8 strain C4D recipients of T. pallidum-immune cells developed an erythematous reaction of short duration at the injection site; 2 of these recipients were positive for T. pallidum. Throughout the experimental period the humoral response to treponemal antigens was substantially lower in the adoptively immune guinea pigs than in various unprotected control groups. Passive immunity to infection with T. pallidum, however, seems to be dose related, since asymptomatic infection persisted for as long as 3 months after challenge in strain 2 guinea pigs transfused with 10(8) T. pallidum-immune lymphocytes, but not in C4D recipients of twice as many immune cells.

Studies of rabbit testes infected with Treponema pallidum. I Immunopathology.

Rabbit testes were injected with suspensions of Treponema pallidum, washed T pallidum, heat killed T pallidum, or Reiter treponemes. The testes were removed three to 24 days after injection and examined for the number of treponemes, the presence of treponemal antibodies, histopathological changes, and presence of T and B cells. In animals infected with T pallidum a substantial number (10(6)-10(7)/ml) of organisms were still present at day 24 in spite of early local production of antibodies and increasing infiltration with plasma cells, T lymphocytes, and macrophages. In animals infected with washed T pallidum a lower degree of inflammation was observed than in those infected with unmodified T pallidum, and the treponemal antibodies were detected simultaneously in samples of testicular fluid and serum. In the groups injected with heat killed T pallidum and Reiter treponemes no macroscopical or microscopical changes were detected, although in the group injected with heat killed T pallidum treponemal antibodies were detected in the testicular fluid on day 24.

Interaction of macrophages and lymphocytes in rat skin allografts.

Light microscopy of 2 microgram sections of rejecting rat skin allografts, embedded in hydroxyethyl methacrylate, revealed among the cells infiltrating the graft base extravascular macrophages containing a small lymphocyte. Toluidine blue staining indicated DNA degradation in some of these phagocytosed lymphocytes. More frequently small lymphocytes were in intimate contact with the surface of the macrophages, resembling "Periopolesis', which others have previously observed in vitro. These macrophage-lymphocyte interactions were not seen in sections of autografts. Despite a previous report that diphenylhydantoin (phenytoin) impairs macrophage function, these macrophage-lymphocyte interactions were present in grafts placed in rats receiving this drug. This treatment did not hasten or delay the onset of graft rejection. These in vivo findings both accord with recent in vitro studies on the mechanisms of phagocytosis and with reports that phagocytosis is one of the effector mechanisms in allograft rejection. However, macrophage phagocytosis of lymphocytes has also been observed in testicular lymph collected from conscious normal sheep.