Brown adipocytes, cardiac protection and a common adipo- and myogenic stem precursor in aged human hearts.

New data on adult stem cells (ASCs) are continuously added by research for use in regenerative medicine. However organ-specific ASC markers are incompletely explored. It was demonstrated that in non-cardiac brown adipose tissue (BAT) CD133+ cells differentiate in cardiomyocytes, and such BAT-derived cells induce bone marrow-derived cells into cardiomyocytes, thus being a promising source for cardiac stem cell therapy. During embryogenesis the subepicardial fat derives from BAT. Although it was not specifically investigated in human adult or aged hearts, it is actually known that metabolically active BAT can be found in many adult humans, is related to antiobesity effects, and it may derive from stem/progenitor cells. Stro-1 can safely identify in situ cardiac stem cells (CSCs) with myogenic and adipogenic potential. It was therefore raised the hypothesis of subepicardial differentiation of CSCs in BAT in adult/aged hearts, which could be viewed, such as in infants, as a mechanism of protection. This could be determined by the reactivation of an embryologic differentiation pattern in which brown adipocytes and muscle cells derive from a common stem ancestor. Such quiescent common stem ancestors could be suggested in adult, or aged, human hearts, when subepicardial BAT is found, and if a Stro-1+/CD133+/Isl-1+ phenotype of CSCs is determined.

Staphylococcus aureus phagocytosis. A new cytofluorometric method using FITC and paraformaldehyde.

A new method for measuring the uptake of Staphylococcus aureus by human polymorphonuclear neutrophils (PMN) using flow cytometry (FCM) is described. Bacteria were labeled with fluorescein isothiocyanate (FITC) and incubated with PMN in a suspension assay. At the end of the assay, phagocytosis was arrested by the addition of cold paraformaldehyde. Thereafter, phagocytosis was quantitated by FCM, using crystal violet to distinguish adherent versus ingested bacteria. The method was validated in multiple samples by reference to our microscopic technique; correlations of phagocytic indices were in good agreement. The FCM method was also found to be reproducible and provided a mean to detect low phagocyte values. Another feature of the approach is that FCM readings can be delayed without any appreciable alterations in the results.

Impaired neutrophil defense against Yersinia enterocolitica in patients with iron overload who are undergoing dialysis.

Increased risk that patients with iron overload who are undergoing dialysis will have bacteremia caused by Yersinia enterocolitica has previously been shown. Iron overload is known to increase the virulence of Y. enterocolitica. Whether alterations of the phagocyte defense against this organism are also involved has not yet been determined. We compared neutrophil defense against a serum-resistant strain of Y. enterocolitica in three groups of individuals: nine patients receiving hemodialysis who had iron overload (group 1), nine patients receiving hemodialysis who did not have iron overload (group 2), and 10 healthy controls (group 3). Y. enterocolitica phagocytosis and killing were studied in the presence of autologous or pooled normal human serum. Phagocytosis was significantly decreased in group 1 compared with that in the other two groups. The use of normal serum for opsonization did not improve the phagocytosis function. Killing was moderately decreased in the group 1, but only in the presence of autologous serum. We conclude that in patients with iron overload who are undergoing dialysis, the high frequency of Yersinia bacteremia is attributable not only to increased virulence of this microorganism but also to disturbances of the mechanisms specifically involved in the neutrophil defense against Yersinia invasion.

Neutrophil dysfunctions in thalassaemia major: the role of cell iron overload.

The susceptibility to infections was recorded in 13 patients with beta thalassaemia major (T.P.). The following parameters were also investigated in their polymorphonuclear neutrophils (PMN): nitro blue tetrazolium (NBT) reduction, heated yeast and Escherichia coli phagocytosis, Escherichia coli killing and myeloperoxydase activity. These results were compared to those obtained in healthy controls (H.C.). The Perls's reaction was performed on PMN and graded according to a scoring system with the aim of quantifying the iron intoxication of PMN. Phagocytosis and Perls's reaction of PMN from H.C. were also studied after 20 h of incubation with thalassaemic serum. 6 T.P. out of 13 developed septicaemia during their lifetime and in all 9 septicaemic episodes were noted. Phagocytosis was greatly impaired, disclosing both cellular and serum abnormalities. The mean percentage of Perls's positive PMN was 13% in T.P., contrasting with the constant negative reaction in H.C. The incubation of PMN from H.C. with serum from T.P. induced the simultaneous appearance of a phagocytosis defect and of a positive Perl's reaction. It was concluded that in beta thalassaemia major the phagocytosis of PMN was altered due to a combination of serum and cellular abnormalities and that both may be related to the iron overload.

Desferrioxamine improves neutrophil phagocytosis in thalassemia major.

The aims of the present study are: first, to assess the toxic role of serum from thalassemic patients in phagocytosis of PMN from healthy controls, and second, to seek to determine whether serum and cellular disturbances of polymorphonuclear neutrophils (PMN) phagocytosis, observed in thalassemic patients, can be prevented and/or corrected by use of desferrioxamine (DFX). Two kinds of in vitro incubations--without or with DFX--were performed. PMN or serum from thalassemic patients or from healthy controls was used. First, a phagocytosis defect of 3 different bacteria species was induced in PMN from healthy controls by incubation in thalassemic serum. Second, DFX could prevent, already at 1 microM, the phagocytic defect induced in normal PMN by the incubation with thalassemic serum, with disappearance of the toxic role of thalassemic serum at higher concentrations. Third, improvement of the phagocytosis defect of PMN from thalassemic patients was also observed at 1 microM of DFX for the 3 bacteria species. Normalization was obtained at higher concentrations for gram-negative bacteria. In vivo studies revealed, after a 3 hr subcutaneous infusion of DFX into 3 thalassemic patients, an improvement of the phagocytosis results and a decrease of the Prussian Blue reactivity of the PMN. It is concluded first that an iron-mediated defect in phagocytosis can be induced in normal neutrophils by incubation in serum from thalassemic patients, and second that a precautious and intensive chelation therapy seems to be advantageous for increasing PMN defense against infectious agents. Special care must nevertheless be taken in order to detect rapidly opportunistic (such as Yersinia) infections.

Recombinant erythropoietin reverses polymorphonuclear granulocyte dysfunction in iron-overloaded dialysis patients.

Iron overload increases the risk of bacterial infection in dialysis patients, partly by impairing functions of the polymorphonuclear granulocytes (PMNs). PMN defence was studied sequentially in haemodialysis patients with transfusional haemosiderosis, treated for 6 +/- 1.5 months (n = 8) to 13 +/- 1.7 months (n = 4) with recombinant human erythropoietin (rHuEpo). Over this period, signs of iron overload (increased serum ferritin and serum iron) improved, and stainable iron disappeared in PMNs. Simultaneously, phagocytosis of Yersinia enterocolitica by PMNs improved. The decrease in serum ferritin was significantly related to the improved phagocytosis. Killing of Y. enterocolitica by PMNs also improved. It is anticipated that rHuEpo therapy in iron-overloaded dialysis patients could decrease the incidence of bacterial infection by improving PMN functions in these patients.