Maria de Sousa

Iron overload in β2-microglobulin-deficient mice

Abstract The present paper describes the results of a comparative histological and quantitative analysis of iron distribution in tissues of β2m−/− and β2m+/− mice of different ages. Progressive hepatic iron overload, indistinguishable from that observed in human hemochromatosis, was found only in mice homozygous for the mutated β2m gene. Total iron measurements done by flame atomic absorption showed statistically significant differences between liver samples from 5 β2m+/− heterozygotes (468 ± 174 μg/g of dry weight) and 9 mice homozygous for the mutated β2m gene with average total hepatic iron levels of 1583 ± 424 μg/g of dry weight.

Haemochromatosis as a window into the study of the immunological system : a novel correlation between CD8+ lymphocytes and iron overload

Abstract: In the present study we report a serial investigation of the numbers of the peripheral blood cells — erythrocytes, polymorphonuclear neutrophils, total lymphocytes, T‐lymphocyte subpopulations (CD2, CD4, CD8), B lymphocytes and monocytes — in a group of 21 patients with haemochromatosis during the time of intensive phlebotomy treatment, i.e., from iron overload until the onset of iron deficiency. A remarkable individual stability of all blood cell populations studied was found in all patients. Patients differed in their relative proportions of CD4+ and CD8+. Each individuals CD4/CD8 ratio, as well as the absolute numbers, remained unaffected with time, confirming the existence of a strict homeostatic regulation of the relative numbers of the two major peripheral T lymphocytes. A significant positive correlation between CD4/CD8 ratios and the amount of iron mobilised by phlebotomy was found during this study. A novel correlation between the relative proportions of CD4+ and CD8+ cells and iron overload is confirmed by the follow‐up of iron re‐entry in the serum transferrin pool in the treated patients.

Major histocompatibility complex class I associations in iron overload: evidence for a new link between the HFE H63D mutation, HLA‐A29, and non‐classical forms of hemochromatosis.

Abstract The present study is an analysis of the frequencies of HFE mutations in patients with different forms of iron overload compared with the frequencies found in healthy subjects from the same region. The frequencies of HLA-A and -B antigens and HLA haplotypes were also analyzed in the same subjects. The study population included: 71 healthy individuals; 39 genetically and clinically well-characterized patients with genetic hemochromatosis (HH); and 25 patients with non-classical forms of iron overload (NCH), excluding secondary hemochromatosis. All subjects were HLA-typed and HFE-genotyped by the oligonucleotide ligation assay (OLA). The gene frequencies found for the C282Y and H63D mutations of HFE were respectively: 0.03 and 0.23 in healthy individuals, 0.86 and 0.04 in HH patients, and 0.08 and 0.48 in NCH patients. An expected significant association between HH and HLA-A3 was observed, which was found to be in linkage disequilibrium with the C282Y mutation. A new association was seen, however, between HLA-A29 and NCH, in linkage disequilibrium with the H63D mutation. Again as expected, the HLA-B antigen B7 was associated with HH in linkage disequilibrium with HLA-A3. In addition, the HLA-B antigen B44 was found to be associated with NCH but not in linkage disequilibrium with either A29 or the H63D mutation. In conclusion, a new association of the HFE H63D mutation with forms of hemochromatosis other than HH and a new association between the HLA phenotype A29 and the HFE H63D mutation were found in the same patients. These findings reinforce evidence for the involvement of the major histocompatibility class I in iron metabolism, supporting the notion of a physiological role for the immunological system in the regulation of iron load.

Recognition of self within self: specific lymphocyte positioning and the extracellular matrix

In this article, Maria de Sousa and colleagues address the importance of interactions between lymphocytes and extracellular matrix components to lymphocyte migration and positioning, emphasizing the role of the basement membrane and the complex stromal matrix of the lymphoid organs. They conclude that such interactions are of great importance, both in the physiological functioning of the immune system and in organ transplantation.

Regulation of expression of a human lymphoid cell surface marker by iron

Abstract Spontaneous rosette formation of human peripheral blood lymphocytes with sheep erythrocytes was inhibited by pretreatment of cells with Fe-citrate, fully saturated TF (but not native TF), fully saturated LF, or native LF. The inhibition was observed within 5 min of incubation; it was relatively temperature dependent, and not mediated by the presence of macrophages, since it occurred after depletion of adherent cells from the suspension. The inhibitory effect of iron was reversed by further treatment of the cells with two highly specific iron-chelating agents, DFX and SHAM, or culture in medium supplemented with 15% FCS. Alternate expression or inhibition of the receptor was observed when the cells were cultured sequentially in low- or high-iron medium for periods of 24 hr. To our knowledge, this is the first demonstration of regulation of expression of a surface marker in mammalian cells by iron. Iron is known to play a similar regulatory role of membrane receptors in bacteria.

Differential inhibition of the MLR by iron: Association with HLA phenotype

The effect of iron on the MLR was examined by pretreating peripheral blood mononuclear cells from 77 unrelated Caucasians with five concentrations of Ferric-citrate (10.0 mM, 1.0 mM, 0.1 mM, 0.01 mM and 0.005 mM). After incubation with the metal, the cells were washed and cultured in a one-way MLR with a pool of stimulator cells. Cell viability remained unchanged (greater than 90 percent) during the 6-day culture period. Citrate per se had no effect on either the responder or the stimulator population. Iron treatment influenced the MLR in the following ways: (1) a variable degree of inhibition was observed which related to the dose of Ferric-citrate used and to HLA phenotype, (2) the responder but not the stimulator cells were affected, (3) no statistically significant differences were seen between female and male donor cells and (4) the mean percent response of cells from HLA-A2 donors were significantly (0.005<P<0.01) less susceptible to iron exposure than those from non-HLA-A2 individuals.The present results indicate that iron can interact with lymphoid cells and influence some immunological functions in vitro. The possibility is discussed that similar interactions take place in vivo which could contribute to the prognosis of certain diseases associated with particular HLA phenotypes.

Inhibition of E-rosette formation by two iron salts

Abstract The effects of four different metal salts on E-rosette formation by human peripheral blood lymphocytes and cells from a human T-cell line were examined. Pretreatment of lymphocytes with FeCl3 and Fe-citrate inhibited rosette formation. The inhibition was related to cell and iron salt concentrations. Zinc chloride and Na-citrate had no significant effect on rosette formation. The results indicate that lymphocytes can bind Fe3+ and the possible implications of this finding are discussed in relation to the known roles played by T lymphocytes in the control of erythropoiesis and cellular immunity.

Control mechanism of lymphocyte traffic: A study of the action of two sulfated polysaccharides on the distribution of 51Cr- and [3H]adenosine-labeled mouse lymph node cells

Abstract The effects of dextrans of varying molecular weights and of pentosan sulfate on the distribution of 51 Cr-labeled mouse lymph node cells were studied in vivo , i.e., in recipients treated with the sulfated polysaccharides, and in vitro , i.e., by following the fate of cells treated in vitro , in intact syngeneic recipients. Both types of experiments demonstrate that dextrans, especially dextran sulfate (DXS) and pentosan sulfate (PS), considerably reduce lymph node entry of lymphocytes, with concomitant increases in the blood and, in the case of DXS, in both the blood and lungs. A parallel quantitative autoradiographic analysis of the distribution of [ 3 H]adenosine-labeled cells confirmed the data with the 51 Cr-labeled cells and, in adidtion, indicated that DXS and PS slow down circulation of lymphocytes through the marginal zone and red pulp of the spleen and, in the case of DXS, in the pulmonary capillary bed. Unusually large numbers of unlabeled lymphocytes were found in the endothelial wall of the post-capillary venules in lymph nodes of PS-treated mice.

The role of cell interactions in the control of lymphocyte traffic

Abstract The effect of pretreatment of 51 Cr-labelled lymph node cells with neuraminidase, trypsin, phospholipase A, Con A and LPS on their fate in intact and splenectomized recipients following intravenous injections was studied. Decreased lymph node entry was observed in all experimental groups in which intact mice were used as recipients. Evidence for this decrease to be the result of a change in the interaction of the circulating cells with the cells in the liver, lungs or spleen and not the result of a specific defect of the interaction between the lympocytes and the post-capillary venule endothelium is presented and discussed.

Lymphocyte interactions and positioning: I. Adhesive interactions

Abstract We report on the effects of factors obtained from short-term cultures of mouse, rat, and human, B, peripheral T, and thymic lymphocytes on the adhesion of these cells and of a range of tissue cells. 1. Heterologous combinations of factor and cell in the mouse system lead to a diminution in lymphocyte to lymphocyte adhesion. Doseresponse curves are reported and a definition of activity is given. Homologous combinations do not affect adhesion. 2. Mouse B and peripheral T cells show no intrinsic specificity of adhesion but addition of one of the factors to a system of cells of both types leads to the appearance of specific aggregates. 3. The sera of normal mice contain both B and T factor activities, while sera from the B mouse contain only B factor activities. 4. The factors are not species specific in the group of organisms mouse, rat, man and also affect the cell to cell adhesion of various other cell types such as embryonic chick liver and neural retina. These cell types also produce factors that affect lymphocyte adhesion. Two CHO cell lines neither produce or are affected by factors. 5. The mouse B and T factors have molecular weights of approximately 3000 and 9000 daltons, respectively. They are heat sensitive molecules. 6. Injection of mouse T factor into a B mouse leads to extensive repositioning of the injected B lymphocytes. 7. The possible roles of such factors in lymphocyte ecotaxis, homing, and positioning are discussed.