Degree of Immunity Induced by Killed Vaccines to Experimental Salmonellosis in Mice

Killed vaccines, deoxycholate-extracted or heated, were shown to induce an effective degree of immunity which protected against death (100%), prevented extensive multiplication, and left the mice with low residual salmonella populations in spleen and liver after intravenous (iv) or intraperitoneal (ip) challenge with virulent Salmonella typhimurium. Protection was most effective against the ip challenge route and less effective against the iv route. A study of the kinetics of the population of bacteria in the spleens and livers of immunized animals showed that after ip challenge there was an initial reduction of 99% at 6 hr after challenge, maintenance of levels of less than 10(3) bacteria per organ, and a final population of 10(2) to 10(3) per organ at 21 days. With iv challenge, after an initial reduction of 90% at 6 hr, growth ensued to levels above 10(6) bacteria per organ until 8 days, followed by a steady decline yielding residual populations of 10(3) to 10(4) in some cases. Organ hypertrophy correlated with bacterial population. Morbidity was prevented (as measured by gain in body weight) by immunization against ip challenge but not against iv challenge. Killed vaccines protected by their ability to induce an immune state which reduced the initial challenge population, prevented extensive multiplication, yet allowed "cellular immunity" to develop due to response to the living challenge infection itself. The consequence was a low-level carrier state similar to that induced by recovery from sublethal virulent infection.     

Chimaeric anti-CD4 monoclonal antibody cross-linked by monocyte Fcγ receptor mediates apoptosis of human CD4 lymphocytes

Previous studies have shown that murine anti-CD4 monoclonal antibody, cross-linked by rabbit anti-mouse immunoglobulin, could mediate apoptosis of murine CD4⁺ lymphocytes when they were stimulated by T cell receptor antibody. In this study, we have shown that the murine anti-CD4 monoclonal antibody, OKT4, can induce apoptosis in human CD4⁺ T cells stimulated by the recall antigen tuberculin purified protein derivative (PPD) only when cross-linked by rabbit anti-mouse immunoglobulin. The chimeric anti-CD4 monoclonal antibody, cM-T412 whose Fc fragment is human, was able to cause apoptosis without cross-linking by a second antibody. Similarly, abolition of PPD-induced proliferation of peripheral blood mononuclear cells by cM-T412 did not require cross-linking with rabbit anti-human immunoglobulin. Inhibition of proliferation by cM-T412 could be reduced by pre-treating monocytes with heat-aggregated human IgG. This suggested that monocyte Fcγ receptors might be cross-linking the human Fc of cM-T412. Propidium iodide staining together with immunofluorescence showed that the apoptotic cells were indeed CD4⁺ lymphocytes. It is proposed that during treatment with cM-T412 in autoimmune disease such as rheumatoid arthritis, cM-T412-coated CD4 T cells, when they are subsequently stimulated by the unknown arthritogenic antigen, may undergo apoptotic cell death through cross-linking of cM-T412 on Fey receptor-positive cells within the joint.     

Mariner is defective in myosin VIIA: a zebrafish model for human hereditary deafness

The zebrafish (Danio rerio) possesses two mechanosensory organs believed to be homologous to each other: the inner ear, which is responsible for the senses of audition and equilibrium, and the lateral line organ, which is involved in the detection of water movements. Eight zebrafish circler or auditory/vestibular mutants appear to have defects specific to sensory hair cell function. The circler genes may therefore encode components of the mechanotransduction apparatus and/or be the orthologous counterparts of the genes underlying human hereditary deafness. In this report, we show that the phenotype of the circler mutant, mariner, is due to mutations in the gene encoding Myosin VIIA, an unconventional myosin which is expressed in sensory hair cells and is responsible for various types of hearing disorder in humans, namely Usher 1B syndrome, DFNB2 and DFNA11. Our analysis of the fine structure of hair bundles in the mariner mutants suggests that a missense mutation within the C-terminal FERM domain of the tail of Myosin VIIA has the potential to dissociate the two different functions of the protein in hair bundle integrity and apical endocytosis. Notably, mariner sensory hair cells display morphological and functional defects that are similar to those present in mouse shaker-1 hair cells which are defective in Myosin VIIA. Thus, this study demonstrates the striking conservation of the function of Myosin VIIA throughout vertebrate evolution and establishes mariner as the first fish model for human hereditary deafness.     

A Randomized Clinical Trial of Topical Cysteamine Disulfide (Cystamine) versus Free Thiol (Cysteamine) in the Treatment of Corneal Cystine Crystals in Cystinosis

In nephropathic cystinosis, corneal cystine crystals cause severe photophobia and corneal erosions. Topical cysteamine dissolves these crystals, but cannot be marketed because it rapidly oxidizes to the disulfide form, cystamine, at room temperature. Since cystamine itself could be used commercially, we compared the efficacy of cystamine and cysteamine with respect to cystine crystal dissolution in a randomized, double-masked clinical trial. One eye each of 14 patients with cystinosis was randomized to either cystamine or cysteamine, 0.5%, with 0.01% benzalkonium chloride; the companion eye was treated with the alternate preparation. Corneal crystals were photographed and a density score was assigned to each slide based on 13 standard slides. After 8–20 months, 6 patients showed significant reduction of the corneal crystal score in only one eye. In each case, the improved eye was the cysteamine-treated eye. Theoretically, cysteamine should dissolve both intracellular and extracellular crystals, whereas cystamine should dissolve only intracellular crystals because it must first be reduced to the free thiol by the cytoplasmic-reducing environment. Hence, the lack of efficacy of the disulfide cystamine suggests that some corneal cystine crystals in cystinosis patients are extracellular, and that another form of stable, topical cysteamine must be developed for cystinosis patients.     

Cytogenetic findings in a parent of a patient with Fanconi''s anemia

We have studied a patient with clinically established Fanconi type anemia whose peripheral blood leukocyte culture and bone marrow showed a normal male karyotype, with no morphological alterations. The skin fibroblasts in cultures derived from his father, however, revealed a mosaic pattern, 47 , XY F+/46, XY. The findings of a trisomic cell line in a parent of a patient with Fanconi's anemia has been reported only once before.     

Creatine kinase activity associated with the contractile proteins of the guinea-pig carotid artery

Summary Activity and role of creatine kinase associated with contractile proteins of vascular smooth muscle have been investigated using skinned guinea-pig carotid artery rings. Membrane solubilization was performed with the detergent Triton X-100. Creatine kinase activity, isoenzyme profile as well as mechanics were performed on the Triton skinned carotid artery rings. Total creatine kinase activity was 47.3±9.3 IU g¹ ww and electrophoresis showed BB, MB, and MM isoforms (BB-CK being the predominant isoenzyme). One hour incubation with Triton X-100, produced predominantly BB-CK remaining with the myofibrils with some MB, representing 23% of the preskinned creatine activity. When relaxed carotid artery rings were exposed to pCa 9 in the presence of 250 M ADP, 0 ATP, and 12 mM phosphocreatine, tension was not significantly different from resting tension, but changing to pCa 4.5 caused the carotid artery rings to generate 49.5±4.5% of maximal tension. When a high-tension rigor state was achieved (250 M ADP, 0 ATP, 0 phosphocreatine, and pCa 9), the addition of 12 mM phosphocreatine effected significant relaxation. These observations implicate an endogenous form of creatine kinase, associated with the myofilaments, which is capable of producing enough ATP for submaximal tension generation and significant relaxation from rigor conditions. These results suggest co-localization of ATPase, MLCK, and creatine kinase on the contractile proteins of the carotid artery. Such an enzymic association may play a role in the energetic supply to the contractile apparatus of vascular smooth muscle.Recipient of INSERM/NIH Fogarty International Fellowship, TW01585-01.     

The effects of anti-hypertensive therapy on the structural,mechanical and metabolic properties of the rat aorta

The vascular system exhibits altered growth, calcium responses and metabolism during hypertension. To relate such changes, we compared histological, tension and metabolic responses in the aorta from 32-week-old spontaneously hypertensive rats (SHRs), normotensive Wistar–Kyoto (WKY) rats, and SHRs treated with Verapamil (V) and ACE-inhibitor, Trandolapril (T) as well as a combination of the two treatments (C). Vascular hypertrophy was apparent in the SHRs. Contractile responses induced by 50 mmol/l KCl and 2.5 mmol/l Ca²⁺ were significantly lower in the SHR (64.4 mN/mm² vs. 49.2 mN/mm²), but an associated increase in Ca²⁺-sensitivity (EC₅₀ of extracellular Ca²⁺ (mol/l): SHR, 456 vs. WKY, 616) normalised tension generating ability. All treatments led to significant decreases in blood pressure, although only T and C treated animals became normotensive with concomitant normalisation of vascular hypertrophy. An increase in oxygen consumption was apparent in the SHR aorta, which was associated with significant differences in the activities of key metabolic enzymes. Anti-hypertensive treatment normalised many of the metabolic parameters, with the C therapy being the most efficacious. We conclude that the treatment of hypertension by combined therapy leads to a better normalisation of structural, contractile, and metabolic parameters in the SHR, than either treatment alone and that metabolic changes with the pathology are resolved with appropriate therapy.     

Cell-mediated immune responses to chlamydial antigens in guinea pigs injected with inactivated chlamydiae

Cell-mediated immunity (CMI) to chlamydial antigens was readily induced in guinea pigs by a single injection of Betaprone-inactivated chlamydiae in complete Freund adjuvant. The CMI was measured in vivo by delayed hypersensitivity skin tests, and in vitro by inhibition of migration of peritoneal exudate cells and by proliferation of lymph node lymphocytes. There was an overall correlation between in vivo and in vitro responses. Of the in vitro assays, migration inhibition reflected the state of sensitization, as judged by skin tests, more uniformly than lymphocyte stimulation. Extensive inter- and intra-species cross-reactivity was noted between LB-1, a strain ofC. trachomatis, and three strains ofC. psittaci, 6BC, GPIC, and 562F. Cross-reactivity between LB-1 and 6BC was one-way only, by all three parameters: LB-1 elicited strong cross-reactions in 6BC-immunized animals but not vice versa. Antichlamydial antibodies could not be demonstrated in any of the animals by microimmunofluorescence.