Characterization of Multiple Helicobacter bizzozeronii Isolates From a Finnish Patient With Severe Dyspeptic Symptoms and Chronic Active Gastritis

Background:  Helicobacter pylori is the primary cause of gastritis and peptic ulceration in humans. In a minority of patients with upper gastrointestinal symptoms, long tightly coiled spiral bacteria, provisionally named " Helicobacter heilmannii, " are observed in gastric biopsies. These bacteria are extremely fastidious and only one previous study has succeeded in obtaining an isolate in vitro.
Materials and Methods:  We used two different selective media to isolate " H. heilmannii " from the gastric mucosa of a Finnish patient presenting with severe dyspeptic symptoms. The isolates were characterized by testing for urease and catalase activity, by using light and electron microscopy, and by sequencing of the partial 16S rRNA and ureAB genes. Single-enzyme amplified fragment length polymorphism (sAFLP) was used to analyze the genetic diversity among the isolates.
Results:  We obtained 15 isolates from different gastric biopsies prior and three after unsuccessful treatment of the patient. The isolates were identified as Helicobacter bizzozeronii . Eradication therapy was unsuccessful most probably due to high level of resistance to metronidazole. Persistent colonization by the same H. bizzozeronii clone was confirmed by sAFLP, however, small differences between the profiles suggested long-term colonization of the patient.
Conclusions:  Helicobacter bizzozeronii remains the only " H. heilmannii " species isolated from human gastric mucosa although it has been an infrequent observation among " H. heilmannii "-infected patients in PCR-based screening studies. The relevance of H. bizzozeronii and other potentially zoonotic gastric Helicobacter spp. in human disease remains to be determined.     

Evidence for progressive reduction and loss of telocytes in the dermal cellular network of systemic sclerosis

Telocytes, a peculiar type of stromal cells, have been recently identified in a variety of tissues and organs, including human skin. Systemic sclerosis (SSc, scleroderma) is a complex connective tissue disease characterized by fibrosis of the skin and internal organs. We presently investigated telocyte distribution and features in the skin of SSc patients compared with normal skin. By an integrated immunohistochemical and transmission electron microscopy approach, we confirmed that telocytes were present in human dermis, where they were mainly recognizable by their typical ultrastructural features and were immunophenotypically characterized by CD34 expression. Our findings also showed that dermal telocytes were immunophenotypically negative for CD31/PECAM‐1 (endothelial cells), α‐SMA (myofibroblasts, pericytes, vascular smooth muscle cells), CD11c (dendritic cells, macrophages), CD90/Thy‐1 (fibroblasts) and c‐kit/CD117 (mast cells). In normal skin, telocytes were organized to form three‐dimensional networks distributed among collagen bundles and elastic fibres, and surrounded microvessels, nerves and skin adnexa (hair follicles, sebaceous and sweat glands). Telocytes displayed severe ultrastructural damages (swollen mitochondria, cytoplasmic vacuolization, lipofuscinic bodies) suggestive of ischaemia‐induced cell degeneration and were progressively lost from the clinically affected skin of SSc patients. Telocyte damage and loss evolved differently according to SSc subsets and stages, being more rapid and severe in diffuse SSc. Briefly, in human skin telocytes are a distinct stromal cell population. In SSc skin, the progressive loss of telocytes might (i) contribute to the altered three‐dimensional organization of the extracellular matrix, (ii) reduce the control of fibroblast, myofibroblast and mast cell activity, and (iii) impair skin regeneration and/or repair.     

Improved method of canine decerebration

We describe animproved decerebration method for dogs that is suitable for studies ofbrain stem neurons in the absence of anesthesia. Previously reportedtechniques of canine decerebration often lead to respiratory andhemodynamic instability and lack of typical decerebrate rigidity. Wehave developed a precise, visually controlled, midcollicular brain stemtransection technique that overcomes these problems. Our method resultsin only moderate blood loss while preserving carotid and basilar arterycirculations. Consistent levels of brain stem transection routinelylead to stable postdecerebration hemodynamic parameters, allowingprolonged brain stem neuronal recordings. The same model should also be useful for a variety of studies involving other physiological systemsin dogs in the absence of anesthesia and for studies of anestheticeffects.

     

De Novo synthesis of DNA-like molecules by polynucleotide phosphorylase in vitro

In the presence of Mg²⁺ ions, polynucleotide phosphorylase (PNPase, EC 2.7.7.8) is known to synthesize RNA-like polymers using ribonucleoside-5′-diphosphate (NDP) substrates but to be unable to utilize deoxyribonucleoside substrates. Our experiments show that when MgCl₂ is replaced by FeCl₃, PNPase becomes able to synthesize deoxyheteropolymers using deoxyribonucleoside-5′-diphosphates (dNDPs). The deoxyheteropolymer formed from the four dNDPs is degraded by pancreatic DNase, but not by RNase, and is readily used as a template by DNA-dependent DNA polymerase. Synthesis of this DNA-like polymer is accomplished de novo without the help of any primer or preexisting template. What is more, dA/dG and dC/dT ratios of polymers synthesized by different bacterial PNPases closely match ratios found in DNA of the bacterial species the enzyme came from.     

Effect of erythropoietin on primed leucocyte expression profile

Resistance to erythropoietin (EPO) affects a significant number of anaemic patients with end-stage renal disease. Previous reports suggest that inflammation is one of the major independent predictors of EPO resistance, and the effects of EPO treatment on inflammatory mediators are not well established. The aim of this study was to investigate EPO-induced modification to gene expression in primary cultured leucocytes. Microarray experiments were performed on primed ex vivo peripheral blood mononuclear cells (PBMCs) and treated with human EPO-α. Data suggested that EPO-α modulated genes involved in cell movement and interaction in primed PBMCs. Of note, EPO-α exerts anti-inflammatory effects inhibiting the expression of pro-inflammatory cytokine IL-8 and its receptor CXCR2; by contrast, EPO-α increases expression of genes relating to promotion of inflammation encoding for IL-1β and CCL8, and induces de novo synthesis of IL-1α, CXCL1 and CXCL5 in primed cells. The reduction in MAPK p38-α activity is involved in modulating both IL-1β and IL-8 expression. Unlike the induction of MAPK, Erk1/2 activity leads to upregulation of IL-1β, but does not affect IL-8 expression and release. Furthermore, EPO-α treatment of primed cells induces the activation of caspase-1 upstream higher secretion of IL-1β, and this process is not dependent on caspase-8 activation. In conclusion, our findings highlight new potential molecules involved in EPO resistance and confirm the anti-inflammatory role for EPO, but also suggest a plausible in vivo scenario in which the positive correlation found between EPO resistance and elevated levels of some pro-inflammatory mediators is due to treatment with EPO itself.     

Optimized conditions for the fermentation of Helicobacter pylori and production of vacuolating cytotoxin

Abstract We report here improvements to the growth media and fermentation conditions which result in a substantial increase of Helicobacter pylori growth and in the enhanced production of vacuolating cytotoxin. Addition of glucose to the medium resulted in the increase of cell yield, cell viability and a significant improvement in the production of vacuolating cytotoxin.