Scrub typhus vaccine candidate Kp r56 induces humoral and cellular immune responses in cynomolgus monkeys

A truncated recombinant 56-kDa outer membrane protein of the Karp strain of Orientia tsutsugamushi (Kp r56) was evaluated in cynomolgus monkeys (Macaca fascicularis) for immunogenicity and safety as a vaccine candidate for the prevention of scrub typhus. This recombinant antigen induced strong humoral and cellular immune responses in two monkeys and was found to be well tolerated. Antigen-specific immunoglobulin M (IgM) and IgG were produced to almost maximal levels within 1 week of a single immunization. Peripheral blood mononuclear cells from vaccinated animals showed an induction of antigen-specific proliferation and gamma interferon production. The Kp r56 was not as efficient as infection with live organisms in preventing reinfection but was able to reduce the inflammation produced at the site of challenge. This report describes the results of the first systematic study of the immunogenicity of a recombinant scrub typhus vaccine candidate in a nonhuman primate model.     

Functional analysis of the cag pathogenicity island in Helicobacter pylori isolates from patients with gastritis, peptic ulcer, and gastric cancer

Helicobacter pylori is the causative agent of a variety of gastric diseases, but the clinical relevance of bacterial virulence factors is still controversial. Virulent strains carrying the cag pathogenicity island (cagPAI) are thought to be key players in disease development. Here, we have compared cagPAI-dependent in vitro responses in H. pylori isolates obtained from 75 patients with gastritis, peptic ulcer, and gastric cancer (n = 25 in each group). AGS gastric epithelial cells were infected with each strain and assayed for (i) CagA expression, (ii) translocation and tyrosine phosphorylation of CagA, (iii) c-Src inactivation, (iv) cortactin dephosphorylation, (v) induction of actin cytoskeletal rearrangements associated with cell elongation, (vi) induction of cellular motility, and (vii) secretion of interleukin-8. Interestingly, we found high but similar prevalences of all of these cagPAI-dependent host cell responses (ranging from 56 to 80%) among the various groups of patients. This study revealed CagA proteins with unique features, CagA subspecies of various sizes, and new functional properties for the phenotypic outcomes. We further showed that induction of AGS cell motility and elongation are two independent processes. Our data corroborate epidemiological studies, which indicate a significant association of cagPAI presence and functionality with histopathological findings in gastritis, peptic ulcer, and gastric cancer patients, thus emphasizing the importance of the cagPAI for the pathogenicity of H. pylori. Nevertheless, we found no significant association of the specific H. pylori-induced responses with any particular patient group. This may indicate that the determination of disease development is highly complex and involves multiple bacterial and/or host factors.     

Effects of lysine on bicarbonate and fluid absorption in the rat proximal tubule

The effects of lysine on bicarbonate and fluid reabsorption in the rat proximal tubule were studied by luminal and capillary perfusion in situ. The proximal tubule and peritubular capillaries were perfused with bicarbonate Ringer solution containing [14C]inulin. The rate of bicarbonate reabsorption (JHCO3) was estimated to be 124 +/- 9.5 peq.min-1.mm-1 using a pH membrane glass electrode. The rate of net fluid reabsorption (Jv) was 2.6 +/- 0.21 nl.min-1.mm-1. When 10 mM L-lysine was added to the luminal perfusate, a 35% reduction in JHCO3 and no change in Jv were observed. Increase of L-lysine concentration in the luminal perfusate to 20 mM did not reduce JHCO3 further nor did it influence Jv.l When 10 mM L-lysine was added to the capillary perfusate, a 13% reduction in JHCO3 was observed (NS). Increase of lysine concentration in the capillary perfusate to 20 mM significantly reduced JHCO3 by 26% (P less than 0.01). There was no significant change in Jv under both conditions. The effect of L-lysine in the lumen was related to its reabsorption kinetics, D-Lysine, which was not reabsorbed significantly, did not affect bicarbonate reabsorption in the proximal tubule. These results indicate that the inhibitory effect of L-lysine is related to the entry of lysine into the cell from the lumen.     

Primary non-Hodgkin's lymphoma of the heart in two patients with the acquired immunodeficiency syndrome

We report two cases of primary cardiac lymphoma that developed in patients suffering from the acquired immunodeficiency syndrome. Both cases of lymphoma were histologically aggressive as generally observed in patients with the acquired immunodeficiency syndrome. The lymphoma cells in the center of a tumor nodule obtained from one patient were monoclonal B-cells, whereas those at the periphery showed a polyclonal pattern of staining. It is postulated that this represents a monoclonal lymphoma evolving from a polyclonal B-cell lymphoproliferation analogous to those reported in some cases of lymphoma in immunosuppressed patients infected with Epstein-Barr virus. The lymphoma cells in the other case failed to stain for cytoplasmic immunoglobulins. The possible underlying basis for the increase in incidence of lymphoma in immunodeficiency and the reasons for prevalence of extranodal sites are discussed.     

Regulation of the immune response: II. Further studies on differences in ability of F(ab′)2 and 7S antibodies to inhibit an antibody response

The ability of F(ab′)₂ antibody preparations to suppress an immune response is much less than that of intact 7S antibody. The activity possessed by F(ab′)₂ preparations withstood repurification procedures, hence contamination with intact 7S antibody is unlikely. Daily or thrice daily injections of antibody did not make equal the suppressive activities of F(ab′)₂ and intact antibody, indicating that rapid excretion of F(ab′)₂ antibody is not the sole factor involved in the difference in immunosuppressive potency between intact 7S and F(ab′)₂ antibody. Some possibilities for distinct differences in the mechanism of the immuno-suppressive action of F(ab′)₂ and 7S antibodies are raised and discussed.     

Real-time PCR for quantitation of porcine reproductive and respiratory syndrome virus and porcine circovirus type 2 in naturally-infected and challenged pigs

Real-time PCR assays were developed for quantitative detection of porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus type 2 (PCV2). The established real-time PCR for the quantitation of PRRSV cDNA and PCV2 DNA were found to be in the 9-log(10) linear dynamic range with excellent linearity and reliable reproducibility. Using these techniques, the distribution and quantitation of PRRSV and PCV2 in naturally infected and challenged pigs were investigated. The viral concentrations were expressed as the mean log(10) viral DNA or cDNA copy numbers per mg or ml of tested samples. For pigs infected naturally with both viruses, the lung, spleen, tonsil and lymphoid organs had the highest viral burdens with ranges from 5.73 to 8.38 and 5.65 to 6.91 for PRRSV and PCV2, respectively. The injection of formalin-inactivated Salmonella choleraesuis emulsified in complete Freund's adjuvant 1 week before and after the inoculation of both viruses resulted in PRRSV replication enhancement 2 weeks post-challenge. However, this facilitated the clearance of PRRSV 4 weeks post-challenge. Results from this study show that the established quantitative PCR could be a useful tool when applied to vaccine development and pathogenesis studies in the future.     

Thrombus of the ascending aorta: a rare cause of myocardial infarction.

We present two cases of a thrombus in the ascending aorta causing an acute myocardial infarction (AMI) and review the 10 other cases previously reported in the literature. This life-threatening condition appears to be more common in female smokers in their fifth decade. Suspicion should be raised in individuals at low risk for atherosclerotic disease with coronary angiographic findings not in keeping with the clinical presentation. The diagnosis may be obtained by transesophageal echocardiography, and we generally recommend surgical thrombectomy.     

Mechanical Analysis of Atherosclerotic Plaques Based on Optical Coherence Tomography

Finite element analysis is a powerful tool for investigating the biomechanics of atherosclerosis and has thereby provided an improved understanding of acute myocardial infarction. Structural analysis of arterial walls is traditionally performed using geometry contours derived from histology. In this paper we demonstrate the first use of a new imaging technique, optical coherence tomography (OCT), as a basis for finite element analysis. There are two primary benefits of OCT relative to histology: 1) imaging is performed without excessive tissue handling, providing a more realistic geometry than histology and avoiding structural artifacts common to histologic processing, and 2) OCT imaging can be performed in vivo, making it possible to study disease progression and the effect of therapeutic treatments in animal models and living patients. Patterns of mechanical stress and strain distributions computed from finite element analysis based on OCT were compared with those from modeling based on "gold standard" histology. Our results indicate that vascular structure and composition determined by OCT provides an adequate basis for investigating the biomechanical factors relevant to atherosclerosis and acute myocardial infarction.     

Mice with neonatally induced inactivation of the vascular cell adhesion molecule-1 fail to control the parasite in Toxoplasma encephalitis

Under various inflammatory conditions, cell adhesion molecules are up-regulated in the central nervous system (CNS) and may contribute to the recruitment of leukocytes to the brain. In the present study, the functional role of vascular cell adhesion molecule (VCAM)-1 in Toxoplasma encephalitis (TE) was addressed using VCAM(flox/flox MxCre) mice. Neonatal inactivation of the VCAM-1 gene resulted in a lack of induction of VCAM-1 on cerebral blood vessel endothelial cells, whereas the constitutive expression of VCAM-1 on choroid plexus epithelial cells and the ependyma was unaffected; in these animals, resistance to T. gondii was abolished, and VCAM(flox/flox MxCre) mice died of chronic TE caused by a failure to control parasites in the CNS. Although leukocyte recruitment to the CNS was unimpaired, the B cell response was significantly reduced as evidenced by reduced serum levels of anti-T. gondii-specific IgM and IgG antibodies. Furthermore, the frequency and activation state of intracerebral T. gondii-specific T cells were decreased, and microglial activation was markedly reduced. Taken together, these data demonstrate the crucial requirement of VCAM-1-mediated immune reactions for the control of an intracerebral infectious pathogen, whereas other cell adhesion molecules can efficiently compensate for VCAM-1-mediated homing across cerebral blood vessels.     

Neuromuscular defects in a Drosophila survival motor neuron gene mutant

Autosomal recessive spinal muscular atrophy (SMA) is linked to mutations in the survival motor neuron (SMN) gene. The SMN protein has been implicated at several levels of mRNA biogenesis and is expressed ubiquitously. Studies in various model organisms have shown that the loss of function of the SMN gene leads to embryonic lethality. The human contains two genes encoding for SMN protein and in patients one of these is disrupted. It is thought the remaining low levels of protein produced by the second SMN gene do not suffice and result in the observed specific loss of lower motor neurons and muscle wasting. The early lethality in the animal mutants has made it difficult to understand why primarily these tissues are affected. We have isolated a Drosophila smn mutant. The fly alleles contain point mutations in smn similar to those found in SMA patients. We find that zygotic smn mutant animals show abnormal motor behavior and that smn gene activity is required in both neurons and muscle to alleviate this phenotype. Physiological experiments on the fly smn mutants show that excitatory post-synaptic currents are reduced while synaptic motor neuron boutons are disorganized, indicating defects at the neuromuscular junction. Clustering of a neurotransmitter receptor subunit in the muscle at the neuromuscular junction is severely reduced. This new Drosophila model for SMA thus proposes a functional role for SMN at the neuromuscular junction in the generation of neuromuscular defects.