Bone marrow-mesenchymal stem cells as a suitable model for assessment of environmental pollution

Stem cells are considered as ideal model for assessment of environmental toxins on proliferation, multipotency and differentiation. The aim of this study was to investigate the effect of lead as harmaful environmental pollutant on proliferation and neural differentiation of murine bone marrow-MSCs. In this experimental study, MSC cells were exposed to different concentrations of lead [0 to 100 micro M] for 24h, and the level of cell proliferation was determined by 3-[4, 5-dimethyl-2-thiazolyl]-2, 5-dipheny-2H-tetrazolium bromide [MTT] reduction assay. In addition, DNA fragmentation was evaluated with comet assay at a single cell level. To induce the neural phenotype, MSCs were cultured for 2 days in the presence of 50 micro Pb[2+] for 48 h. At the end of this period, the medium was replaced by fresh medium supplemented with 1 mM beta-ME for 24 hr and then fresh medium supplemented with 7 mM beta-ME for 4 hours respectively. The expression of neural marker such as nestin, MAP2, and tau was assessed by immunocytochemistry, while the expression of neuronal specific genes such as Neur-1, Nestin, and beta-tubulin III was determined by RT-PCR analysis. Exposure to lead reduced the level of cell proliferation in a dose-dependent manner. The comet assay of cells exposed to lead showed varying degrees of DNA damaged. Change in cell morphology was observed 1 to 4 hr post neural exposure. The percentage of the MAP2 positive cells was reduced significantly at greater than 40 micro M lead concentration. This observation was further verified by assessment of the expression of neural markers. This study clearly indicated lead is highly cytotoxic to MSCs and these cells appear to be an excellent choice for establishment of guidelines for environmental hazards and drugs on cell proliferation and differentiation

[Characterization of 60 kDa heat shock protein [HSP60] gene in pathogenic dermatophyte Microsporom canis]

The stress-inducible Heat Shock Proteins [i.e. HSP60] constitute one of the highly conserved protein and gene families. As one of the molecular chaperone proteins, they play essential roles in protein metabolism and protein translocation under both stress and non-stress conditions. In the present study, we try to characterize the 60 kDa Heat Shock Protein [HSP60] gene in dermatophyte pathogen Microsporom Canis [M. canis]. This dermatophyte is one of the most important causative agents of dermatophytosis in human and animals. Some properties of M. canis have been investigated in molecular level; however, no information is available regarding the HSP60 in this dermatophyte. In the present study, we try to characterize the 60 kDa Heat Shock Protein [HSP60] gene in dermatophyte pathogen Microsporom Canis [M. canis]. M. canis was obtained from patients with dermatophytosis and cultured in appropriate conditions. High molecular weight DNA was isolated from obtained mycelial mass by standard methods. Pairs of 21 nt primers were designed from highly conserved regions of the HSP60 genes in other eukaryotic cells. Mentioned primers were utilized in PCR using isolated genomic DNA template of M. canis. Predicted molecules have been amplified and were submitted for sequencing. By the time 1550 nucleotides of this gene are sequenced and analysed, that encoding a 497 amino acids protein. In the present study, we report the identification and molecular characterization of a M. canis gene encoding a protein belongs to the 60 kDa Heat Shock Protein family which will here be referred to as McHSP60. Analysis of the amino acid sequence of this gene revealed a considerable identity with other eukaryotic HSP60 such as those of C.immitis [97%], Aspergillus fumigatus [92%] and S.cerevisiae [74%]. Investigation of amino acid composition in HSP60 revealed Alanine, Glutamic acid and Glysine as the most common amino acids in this protein. The amino acid composition of McHSP60 indicates the amount of Cysteine and Tryptophan are poor