Johanna Stefani

Characterization and Histological Localization of Multipotent Mesenchymal Stromal Cells in the Human Postnatal Thymus

The aim of this work was to characterize multipotent mesenchymal stromal cells (MSCs) in the postnatal human thymus and to localize these MSCs in the organ. Adherent cells isolated from thymus samples were characterized by cell-surface antigen expression. This showed that adherent cells have a MSC profile as assessed by the expression of CD73 and CD105 markers and the lack of CD45 expression. These cells are able to differentiate in vitro into adipocytes, osteoblasts, and chondrocytes and to inhibit mixed lymphocyte reaction. This indicates that isolated cells have all of the characteristics of MSC. The fibroblast colony-forming unit (CFU-F) assay was used to determine their frequency in the postnatal thymus. This frequency was 60.9 +/- 14.8 CFU-F per 1 x 10(5) freshly isolated mononuclear cells. Moreover, taking advantage of CD34 and CD105 expression, immunohistological staining allowed us to localize MSC within interlobular trabeculae in close contact with the outer cortex. Polymerase chain reaction experiments indicated that thymic MSC expressed interleukin-7 and stromal cell-derived factor-1 messenger RNA. Overall, these results confirm previous findings of the presence in the adult human thymus of multipotent MSCs with a phenotype similar to adipose-derived adult stem cells. These results also show for the first time a histological localization of MSC in an organ. This suggests a possible role of thymic MSC in intrathymic differentiation.

Effect of nephrotoxic treatment with gentamicin on rats chronically exposed to uranium

Uranium is a radioactive heavy metal with a predominantly chemical toxicity, affecting especially the kidneys and more particularly the proximal tubular structure. Until now, few experimental studies have examined the effect of chronic low-dose exposure to uranium on kidney integrity: these mainly analyse standard markers such as creatinine and urea, and none has studied the effect of additional co-exposure to a nephrotoxic agent on rats chronically exposed to uranium. The aim of the present study is to examine the potential cumulative effect of treating uranium-exposed rats with a nephrotoxic drug. Neither physiological indicators (diuresis and creatinine clearance) nor standard plasma and urine markers (creatinine, urea and total protein) levels were deteriorated when uranium exposure was combined with gentamicin-induced nephrotoxicity. A histological study confirmed the preferential impact of gentamicin on the tubular structure and showed that uranium did not aggravate the histopathological renal lesions. Finally, the use of novel markers of kidney toxicity, such as KIM-1, osteopontin and kallikrein, provides new knowledge about the nephrotoxicity threshold of gentamicin, and allows us to conclude that under our experimental conditions, low dose uranium exposure did not induce signs of nephrotoxicity or enhance renal sensitivity to another nephrotoxicant.

Cholesterol 7α-hydroxylase (CYP7A1) activity is modified after chronic ingestion of depleted uranium in the rat

Depleted uranium (DU) is a radioactive heavy metal derived from the nuclear energy production. Its wide use in civilian and military items increases the risk of its environmental dissemination, and thus the risk of internal contamination of populations living in such contaminated territories. Previous studies have shown that vitamin D and cerebral cholesterol metabolisms were affected following chronic ingestion of DU. Even more than the brain, the liver is a crucial organ in cholesterol homeostasis since it regulates cholesterol distribution and elimination at body level. The aim of this work was to assess the impact of a low-level chronic ingestion of DU on hepatic cholesterol metabolism. Rats were contaminated with DU in their drinking water at a concentration of 40mg/l for 9 months. The major effect induced by DU was a decrease of CYP7A1 specific activity (-60%) correlated with a matching decrease of its product 7alpha-hydroxycholesterol in the plasma. Hepatic gene expression of transporters ABC A1, ABC G5, ABC G8 and of nuclear receptor RXR was increased, whereas that of catabolism enzyme CYP7B1 was decreased. Thus, after a chronic ingestion of DU, rats experience a modulation of cholesterol catabolism but overcome it, since their cholesterolemia is preserved and no pathology is declared.

Chronic exposure to low concentrations of strontium 90 affects bone physiology but not the hematopoietic system in mice

The aim of this work was to delineate the effects of chronic ingestion of strontium 90 (90Sr) at low concentrations on the hematopoiesis and the bone physiology. A mouse model was used for that purpose. Parent animals ingested water containing 20 kBq l−1 of 90Sr two weeks before mating. Offspring were then continuously contaminated with 90Sr through placental transfer during fetal life, through lactation after birth and through drinking water after weaning. At various ages between birth and 20 weeks, animals were tested for hematopoietic parameters such as blood cell counts, colony forming cells in spleen and bone marrow and cytokine concentrations in the plasma. However, we did not find any modification in 90Sr ingesting animals as compared with control animals. By contrast, the analysis of bone physiology showed a modification of gene expression towards bone resorption. This was confirmed by an increase in C‐telopeptide of collagen in the plasma of 90Sr ingesting animals as compared with control animals. This modification in bone metabolism was not linked to a modification of the phosphocalcic homeostasis, as measured by calcium, phosphorus, vitamin D and parathyroid hormone in the blood. Overall these results suggest that the chronic ingestion of 90Sr at low concentration in the long term may induce modifications in bone metabolism but not in hematopoiesis. Copyright

Influence on the mouse immune system of chronic ingestion of 137Cs

The aim of this work was to determine the possible occurrence of damage to the immune system during the course of chronic ingestion of (137)Cs. BALB/C mice were used, with (137)Cs intake via drinking water at a concentration of 20 kBq l(-1). Adults received (137)Cs before mating and offspring were sacrificed at various ages between birth and 20 weeks. Phenotypic analysis of circulating blood cells and thymocytes did not show any significant modification of immune cell populations in animals ingesting (137)Cs as compared with control animals, with the exception of a slight increase in Treg percentage at the age of 12 weeks. Functional tests, including proliferative response to mitogens such as phytohaemagglutinin, response to alloantigens in mixed lymphocyte reaction and immunoglobulin response to vaccine antigens such as tetanus toxin and keyhole limpet haemocyanin did not show any significant functional modification of the immune system in (137)Cs-ingesting animals as compared with control animals. Overall, our results suggest that chronic ingestion of a low concentration of (137)Cs in drinking water in the long term does not have any biologically relevant effect on the immune system.

Bone marrow stromal cells spontaneously produce Flt3-ligand: Influence of ionizing radiations and cytokine stimulation

Purpose: To define the ability of human bone marrow (BM) stromal cells to produce fms-like tyrosine kinase 3 (Flt3)-ligand (FL), and the effect of irradiation, tumour necrosis factor-alpha (TNF) or tumour growth factor beta (TGF) on FL production. Material and methods: Primary BM stromal cell cultures were irradiated at 2–10 Gy or were stimulated with TNFα or TGFβ1. The presence of FL was tested in culture supernatants and in cell lysate. The presence of a membrane-bound form of FL and the level of gene expression were also tested. Results: Primary BM stromal cells spontaneously released FL. This production was increased by TNFα but not by TGFβ1 or by irradiation. Chemical induction of osteoblastic differentiation from BM stromal cells also induced an increase in FL release. Conclusions: Our results suggest that the observed increase in FL concentration after in vivo irradiation is an indirect effect. The possible implication of BM stromal cells in these mechanisms is discussed.

Immune System Modifications Induced in a Mouse Model of Chronic Exposure to 90Sr

Strontium 90 (90Sr) remains in the environment long after a major nuclear disaster occurs. As a result, populations living on contaminated land are potentially exposed to daily ingesting of low quantities of 90Sr. The potential long-term health effects of such chronic contamination are unknown. In this study, we used a mouse model to evaluate the effects of 90Sr ingestion on the immune system, the animals were chronically exposed to 90Sr in drinking water at a concentration of 20 kBq/l, for a daily ingestion of 80–100 Bq/day. This resulted in a reduced number of CD19+ B lymphocytes in the bone marrow and spleen in steady-state conditions. In contrast, the results from a vaccine experiment performed as a functional test of the immune system showed that in response to T-dependent antigens, there was a reduction in IgG specific to tetanus toxin (TT), a balanced Th1/Th2 response inducer antigen, but not to keyhole limpet hemocyanin (KLH), a strong Th2 response inducer antigen. This was accompanied by a reduction in Th1 cells in the spleen, consistent with the observed reduction in specific IgG concentration. The precise mechanisms by which 90Sr acts on the immune system remain to be elucidated. However, our results suggest that 90Sr ingestion may be responsible for some of the reported effects of internal contamination on the immune system in civilian populations exposed to the Chernobyl fallout.