Analysis of cell type-specific responses mediated by the type IV secretion system of Helicobacter pylori

Helicobacter pylori persistently infects the human stomach and can cause gastritis, gastric ulceration, and gastric cancer. The type IV secretion system (TFSS) of virulent H. pylori strains translocates the CagA protein, inducing the dephosphorylation of host cell proteins and leading to changes in the morphology or shape of AGS gastric epithelial cells. Furthermore, the TFSS is involved in the induction of proinflammatory cytokines. While the H. pylori genes required for TFSS function have been investigated systematically, little is known about possible host cell factors involved. We infected 19 different mammalian cell lines individually with H. pylori and analyzed CagA translocation, dephosphorylation of host cell proteins, chemokine secretion (interleukin-8 and macrophage inflammatory protein 2), and changes in cellular phenotypes. Our results demonstrate that not only bacterial but also host cell factors determine the cellular response to infection. The identification of such unknown host cell factors will add to our understanding of host-pathogen interactions and might help in the development of new therapeutic strategies.     

Descending projections from the caudal medulla oblongata to the superficial or deep dorsal horn of the rat spinal cord

The location of neurons in the caudal medulla oblongata that project to the superficial or deep dorsal horn was studied in the rat, by means of retrograde labelling from confined spinal injection sites. The tracer cholera toxin subunit B was injected into laminae I–III (fuve rats) or I–V (three rats) at C_4–7 spinal segments. Neurons projecting to the superficial dorsal horn were located in the dorsomedial part of the dorsal reticular nucleus ipsilaterally, the subnucleus commissuralis of the nucleus tractus solitarius bilaterally, and a region occupying the lateralmost part of the ventrolateral reticular formation between the lateral reticular nucleus and the caudal pole of the spinal trigeminal nucleus, pars caudalis, bilaterally. Neurons projecting to the deep dorsal horn, which were only labelled when laminae I–V were filled by the tracer, occurred in the dorsomedial and ventrolateral parts of the dorsal reticular nucleus and in the ventral reticular nucleus bilaterally. A few cells were located in the above described lateralmost portion of the ventrolateral reticular formation bilaterally and in the ventral portion of the ipsilateral cuneate nucleus. In the light of previous data demonstrating that dorsal horn neurons project to the dorsal reticular nucleus, the ventrolateral reticular formation, and the nucleus tractus solitarius, and that neurons in these three medullary regions are involved in pain inhibition at the spinal level, the descending projections demonstrated here suggest the occurrence of spino-medullary-spinal loops mediating the analgesic actions elicited in each nucleus upon the arrival of nociceptive input from the dorsal horn.     

Sporogony and experimental transmission of Babesia equi by Boophilus microplus

The development of Babesia equi in salivary glands of adult female Boophilus microplus was observed under a light microscope using semithin sections stained with toluidine blue. Engorged nymphs were obtained from splenectomized foals experimentally infected with B. equi. As adults, they were then fed on rabbits for 5 days and the salivary glands of manually collected individuals were removed at intervals of 24 h. Sporozoites were found in type III granular acini cells between the 2nd and 5th days following feeding on the rabbits. Sporoblasts and sporozoites were observed in the same or adjacent acini cells in all the glands examined. The formation of the sporozoites occurred following the multiple division of the sporoblasts through a process of radial budding from the periphery of bodies resulting from multiple fission. Sporozoites were detected in smears of adult males stained with Giemsa, between the 2nd and 5th days following feeding by the ticks. Adults of B. microplus, fed during the nymphal phase on foals with patent parasitemia, transmitted sporozoites of B. equi to a splenectomized foal. The role of B. microplus in the transmission and epidemiology of B. equi is discussed. Received: 16 June 1997 / Accepted: 2 September 1997     

Mechanical evaluation of a respiratory device

This article aims to characterize the mechanical behaviour of the Flutter VRP1, a respiratory physiotherapy device designed to aid sputum clearance of the airways of patients. The device resembles a smoking pipe with a conical cavity containing a stainless steel sphere which floats up and down while the patient comes with a forced expiration through it. The sphere's oscillatory movement is function of the air flow rate and angular orientation of the device. When the sphere's oscillatory frequency matches the natural frequency of the patient's chestwall+abdomen system, it will produce resonance which, in turn, will enhance sputum clearance. A dynamical model of the Flutter was formulated and an experimental setup was assembled in order to study the oscillatory frequency of the sphere under different conditions of air flow rate, fluid pressure, angular orientation and sphere's material and weight. Interesting results presented by this article point to eventual mechanical optimization of the device and show information that could be beneficial to the professional of the respiratory physiotherapy.     

The Arabidopsis PILZ group genes encode tubulin-folding cofactor orthologs required for cell division but not cell growth

Plant microtubules are organized into specific cell cycle-dependent arrays that have been implicated in diverse cellular processes, including cell division and organized cell expansion. Mutations in four Arabidopsis genes collectively called the PILZ group result in lethal embryos that consist of one or a few grossly enlarged cells. The mutant embryos lack microtubules but not actin filaments. Whereas the cytokinesis-specific syntaxin KNOLLE is not localized properly, trafficking of the putative auxin efflux carrier PIN1 to the plasma membrane is normal. The four PILZ group genes were isolated by map-based cloning and are shown to encode orthologs of mammalian tubulin-folding cofactors (TFCs) C, D, and E, and associated small G-protein Arl2 that mediate the formation of alpha/beta-tubulin heterodimers in vitro. The TFC C ortholog, PORCINO, was detected in cytosolic protein complexes and did not colocalize with microtubules. Another gene with a related, although weaker, embryo-lethal phenotype, KIESEL, was shown to encode a TFC A ortholog. Our genetic ablation of microtubules shows their requirement in cell division and vesicle trafficking during cytokinesis, whereas cell growth is mediated by microtubule-independent vesicle trafficking to the plasma membrane during interphase.     

DNA damage in cytologically normal urothelial cells of patients with a history of urothelial cell carcinoma

In order to determine if patients with a history of previous urothelial cell carcinoma (UCC) but with current normal urinary cytology have DNA damage in urothelial cells, the single-cell gel electrophoresis (comet) assay was conducted with cells obtained by urinary bladder washings from 44 patients (28 with a history of previous UCC). Increased DNA damage was observed in cytologically "normal" urothelial cells of patients with a history of UCC when compared with referents with no similar history and after correcting the data for smoking status and age (P < 0.018). Increased DNA damage also correlated with the highest tumor grade, irrespective of time or course of the disease after clinical intervention (Kendall tau correlation, 0.37, P = 0.016). Moreover, aneuploidy, as assessed by DNA content ratio (DCR; 75th/25th percentile of total DNA fluorescence of 50 comets/patient) was unaltered by smoking status, but increased with UCC grade: 1.39 +/- 0.12 (median +/- 95% confidence interval; referents); 1.43 +/- 0.11 (Grade I UCC; P = 0.264, against referents); 1.49 +/- 0.16 (Grade II UCC; P = 0.057); 1.57 +/- 0.16 (Grade III UCC; P = 0.003). Micronucleated urothelial cells (MNC) were also scored on Giemsa-stained routine cytological smears and were found not to correlate with DNA damage or DCR. MNC frequencies were higher for patients with a history of UCC and/or smoking than referents with neither history, but there was no statistical difference between groups. Taken together, these results suggest that the normal-appearing urothelium of patients resected for UCC still harbor genetically unstable cells.     

Role of endogenous IFN-gamma in macrophage programming induced by IL-12 and IL-18.

Besides the established role of interleukin-12 (IL-12) and IL-18 on interferon-gamma (IFN-gamma) production by natural killer (NK), T, and B cells, the effects of these cytokines on macrophages are largely unknown. Here, we investigated the role of IL-12/IL-18 on nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) production by CD11b(+) adherent peritoneal cells, focusing on the involvement of endogenously produced IFN-gamma. C57BL/6 cells released substantial amounts of NO when stimulated with IFN-gamma or lipopolysaccharide (LPS), but failed to respond to IL-12 or IL-18 or both. However, IL-12/IL-18 pretreatment was able to program these cells to release 6-8-fold more NO and TNF-alpha in response to LPS or Trypanosoma cruzi stimulation, with NO levels directly correlating with macrophage resistance to intracellular parasite growth. Analysis of IL-12/IL-18-primed cells from mice deficient in IFN-gamma, IFNGR, and IFN regulatory factor-1 (IRF-1) revealed that these molecules were essential for LPS-induced NO release, but TNF-alpha production was IFN-gamma independent. Conversely, the myeloid differentiation factor 88 (MyD88)-dependent pathway was indispensable for IL-12/IL-18-programmed LPS-induced TNF-alpha production, but not for NO release. Contaminant T and NK cells largely modulated the IL-12/IL-18 programming of LPS-induced NO response through IFN-gamma secretion. Nevertheless, a small population of IFN-gamma(+) cells with a macrophage phenotype was also identified, particularly in the peritoneum of chronically T. cruzi-infected mice, reinforcing the notion that macrophages can be an alternative source of IFN-gamma. Taken together, our data contribute to elucidate the molecular basis of the IL-12/IL-18 autocrine pathway of macrophage activation, showing that endogenous IFN-gamma plays an important role in programming the NO response, whereas the TNF-alpha response occurs through an IFN-gamma-independent pathway.     

Peripheral amplification of sweating – a role for calcitonin gene-related peptide

Neuropeptides are the mediators of neurogenic inflammation. Some pain disorders, e.g. complex regional pain syndromes, are characterized by increased neurogenic inflammation and by exaggerated sudomotor function. The aim of this study was to explore whether neuropeptides have a peripheral effect on human sweating. We investigated the effects of different concentrations of calcitonin gene-related peptide (CGRP), vasoactive intestinal peptide (VIP) and substance P (SP) on acetylcholine-induced axon reflex sweating in healthy subjects (total n = 18). All substances were applied via dermal microdialysis. The experiments were done in a parallel setting: ACh alone and ACh combined with CGRP, VIP or SP in various concentrations were applied. Acetylcholine (10⁻² m ) always elicited a sweating response, neuropeptides alone did not. However, CGRP significantly enhanced ACh-induced sweating ( P < 0.01). Post hoc tests revealed that CGRP in physiological concentrations of 10⁻⁷–10⁻⁹ m was most effective. VIP at any concentration had no significant effect on axon reflex sweating. The duration of the sweating response ( P < 0.01), but not the amount of sweat, was reduced by SP. ACh-induced skin blood flow was significantly increased by CGRP ( P < 0.01), but unaltered by VIP and SP. The results indicate that CGRP amplifies axon reflex sweating in human skin.     

Aboriginal status is a prognostic factor for mortality among antiretroviral naïve HIV-positive individuals first initiating HAART

Background  

Although the impact of Aboriginal status on HIV incidence, HIV disease progression, and access to treatment has been investigated previously, little is known about the relationship between Aboriginal ethnicity and outcomes associated with highly active antiretroviral therapy (HAART). We undertook the present analysis to determine if Aboriginal and non-Aboriginal persons respond differently to HAART by measuring HIV plasma viral load response, CD4 cell response and time to all-cause mortality.