Endotoxin Binding and Elimination by Monocytes: Secretion of Soluble CD14 Represents an Inducible Mechanism Counteracting Reduced Expression of Membrane CD14 in Patients with Sepsis and in a Patient with Paroxysmal Nocturnal Hemoglobinuria

Little is known about the role of peripheral blood mononuclear cells (PBMCs) in lipopolysaccharide (LPS) elimination. We studied the endotoxin elimination capacities (EEC) of PBMCs of 15 healthy volunteers, 13 patients with sepsis, and 1 patient suffering from paroxysmal nocturnal hemoglobinuria (PNH). Although expression of CD14, the best-characterized receptor for LPS to date, was reduced from 93.6% ± 0.8% in healthy subjects to 50.5% ± 6.5% in patients with sepsis and was 0.3% in a patient with septic PNH, EEC were found to be unchanged. There was no difference in the amount of tumor necrosis factor alpha (TNF-α) released by PBMCs of healthy donors and patients with sepsis. Anti-CD14 antibodies (MEM-18) completely suppressed EEC, binding of fluorescein isothiocyanate-labeled LPS to monocytes as determined by FACScan analysis, and TNF-α release in all three groups studied. The concentrations of soluble CD14 (sCD14) secreted by endotoxin-stimulated PBMCs from healthy donors and patients with sepsis amounted to 4.5 ± 0.4 and 20.1 ± 1.8 ng/ml, respectively. Based on our results, we suggest that PBMCs eliminate LPS by at least two different mechanisms; in healthy subjects, the membrane CD14 (mCD14) receptor is the most important factor for LPS elimination, while in patients with sepsis (including the septic state of PNH), increased sCD14 participates in LPS elimination. Secretion of sCD14 is strongly enhanced under conditions of low expression of mCD14 in order to counteract the reduction of mCD14 and maintain the function of monocytes. This sCD14 may substitute the role of mCD14 in LPS elimination during sepsis. The elimination of LPS by PBMCs correlates with the binding reaction and the secretion of TNF-α.     

PCR m typing: a new method for rapid typing of group a streptococci

A new approach for the M-typing of Streptococcus pyogenes is reported. Oligonucleotide primers were used in a PCR to amplify the N-terminal region of the emm gene. The presence of the PCR amplification product is associated with the corresponding M serotype. This technique offers potential advantages over other molecular typing methods.     

IFNgamma and TNFalpha account for a pro-clonogenic activity secreted by activated murine peritoneal macrophages

In the present study, we found that murine peritoneal macrophages elicited by BCG or Listeria monocytogenes release into the media an activity capable of stimulating the lung colonization as well as the expression of MHC class I antigens in B16 melanoma cells. A similar activity has previously been found in media conditioned by Corynebacterium parvum-elicited macrophages. Analysis by gel filtration chromatography of media conditioned by Corynebacterium parvum-, BCG- or Listeria monocytogenes-elicited macrophages revealed that the material responsible for the pro-clonogenic activity concentrated in chromatographic fractions corresponding to molecular weights (25 to 52 kDa) which are characteristic of certain cytokines. Thus, we challenged the various macrophage-conditioned media with polyclonal antibodies against IFNγand TNFα, and found that the macrophage pro-clonogenic activity was completely abolished in the presence of anti-IFNγantibodies, but only partially inhibited by anti-TNFαantibodies. This finding suggests a cooperative participation of the two cytokines to the pro-clonogenic activity of the media conditioned by Corynebacterium parvum-, BCG- or Listeria monocytogenes-elicited macrophages. This revised version was published online in July 2006 with corrections to the Cover Date.     

Stem cell therapy for neurodegenerative diseases: the issue of transdifferentiation

In the past few years research on stem cells has exploded as a tool to develop potential therapies to treat incurable neurodegenerative diseases. Stem cell transplantation has been effective in several animal models, but the underlying restorative mechanisms are still unknown. Several events such as cell fusion, neurotrophic factor release, endogenous stem cell proliferation, and transdifferentiation (adult cell acquisition of new unexpected identities) may explain therapeutic success, in addition to replacement of lost cells. This issue needs to be clarified further to maximize the potential for effective therapies. Preliminary stem transplantation trials have already been performed for some neurodegenerative diseases. There is no effective pharmacological treatment for amyotrophic lateral sclerosis, but recent preliminary data both in experimental and clinical settings have targeted it as an ideal candidate disease for the development of stem cell therapy in humans. This review summarizes recent advances gained in stem cell research applied to neurodegenerative diseases with a special emphasis to the criticisms put forward.     

Ultrastructural analysis of pancreatic acinar cells from mice fed on genetically modified soybean

No direct evidence that genetically modified (GM) food may represent a possible danger for health has been reported so far; however, the scientific literature in this field is quite poor. Therefore, we investigated the possible effects of a diet containing GM soybean on mouse exocrine pancreas by means of ultrastructural, morphometrical and immunocytochemical analyses. Our observations demonstrate that, although no structural modification occurs in pancreatic acinar cells of mice fed on GM soybean, quantitative changes of some cellular constituents take place in comparison to control animals. In particular, a diet containing significant amount of GM food seems to influence the zymogen synthesis and processing.     

Pharyngotonsillitis caused by Streptococcus pyogenes: clinical and epidemiological aspects and resistance phenotypes towards macrolides

During 2001 we analyzed 1730 pharyngeal swabs for S. pyogenes (SGA): 1142 children (0-10 years old), 132 adolescent subjects (11-17 years old), and 456 adults (18 or more years old). 994 subjects (664 children, 85 adolescent ones, 245 adults) had acute pharyngotonsillitis. In this last group we observed 321 positivities (32.3 %) for SGA: 40.4 % among children, 24.7 % among adolescent people, 13.1 % among adults. The pharyngotonsillitis prevailed during winter and spring. The resistances (R) towards erythromycin were 27.7 % (89 cases), and among children 30.6 % (82 cases), towards clyndamicin 15.3 % (49 cases, and 17.2 %, 46 cases, among children), towards rokytamicin 11.8 % (38 cases, and 13.1 %, 35 cases, among children). These were the phenotypes of R to erythromycin: 25.8 % M-phenotype, 19.1 % inducible (iMLS), 55.1 % constitutive (cMLS); among children respectively 25.6 %, 18.3 % and 53.7 %. Increased resistances towards 16-C macrolides, increased resistances of cMLS to erythromycin, and the persistence of R to 14-C macrolides around 30 % are discussed.     

Cutaneous mucormycosis in a young, immunocompetent girl.

We report a case of cutaneous mucormycosis in a healthy, immunocompetent young girl (age 14 years). The patient had a 5-year history of a slowly enlarging, erythematous plaque with slight elevated, scaling, circinate borders on the right thigh. Histopathology showed a granulomatous infiltrate with broad, pale, non-septate hyphae. Mycological study identified Mucor hiemalis (Wehmer).     

Pattern and distribution of immunoglobulin VH gene usage in a cohort of B-CLL patients from a Northeastern region of Italy.

We analyzed individual VH gene rearrangements in 55 consecutive B-chronic lymphocytic leukemia (B-CLL) patients collected from a northeastern region of Italy, stressing the possible differences related to geographic characteristics of the cohorts studied. Considering the percentage of somatic mutations present in the VH gene sequences and using the 98% cut-off value, 38 of the 55 B-CLL (69%) patients displayed somatic hypermutations and 17 (31%) had a germline configuration. Our results confirm and extend the observations of a bias in the use of certain VH, DH, and JH genes among B-CLL cells. The most frequently used VH genes were VH1-69 (12.7%) with VH3-23 (12.7%) and VH4-34 (10.9%). Collectively these genes accounted for 36.3% of the cases. In the mutated cases, the range of mutations varied from 2% to 15%, with a median of 6.5%. VH1-69 (7 cases, all unmutated) carried few mutations as opposed to VH3-23 (7 cases, 5 of which mutated), VH4-34 (6 cases, all mutated), and VH3-30 (5 cases, all mutated), which show a high load of mutations. D3 family genes were found frequently (38.1%) followed by D2 (27.2%) and D6 (18.1%). The individual D segment most frequently used was D3-3, which was present in 16.3% of cases. There was predominance of the JH4 gene (49%) followed by JH6 (40%). Analysis of the distribution of replacement and silent mutations in the mutated sequences using the method of Lossos showed in 39.4% of cases evidence of antigen selection in the framework region and/or complementary determining regions. In comparison with a recent study on B-CLL patients from the Mediterranean area, the VH4-34 gene was significantly overused in the mutated group at a percentage double that of the Italian cohort reported in this study (10.9% vs. 5%), but at a frequency similar to the entire Mediterranean region (10.7%). We also found an over-representation of VH1-69 usage in the germline group, at a frequency (12.7%) higher than previously described by the same authors (Italian 8%, Mediterranean 10.7%). On the contrary, VH3-07 and VH3-49 were not much used in our study (5.4% and 1.8%, respectively) compared with the Italian group (8% and 5.1%). In our study, VH3-23 gene segment was frequently expressed, at frequency as high as that of VH1-69, a finding in keeping with reported B-CLL Italian data, but higher than the entire series of the Mediterranean area (12.7% vs. 9.2%); VH3-21 gene, frequently expressed in northern European CLL but rarely in the Mediterranean area, was completely absent. This biased usage of VH family genes may reflect a geographic leukemic repertoire, perhaps owing to a peculiar genetic background, depending on variations in germline composition of the IgVH locus or to the effect of a potential environmental element less frequently encountered in different regions.