Senescence State in Mesenchymal Stem Cells at Low Passages: Implications in Clinical Use.

Mesenchymal stem cells (MSCs) are multipotent cells found in various tissues and are easily cultivated. For use in clinical protocols, MSCs must be expanded to obtain an adequate number of cells, but a senescence state may be instituted after some passages, reducing their replicative potential. In this study, we report a case where MSC derived from an elderly donor acquired a senescence state after three passages. The bone marrow was aspirated from a female patient submitted to a cell therapy for the incontinency urinary protocol; MSCs were cultivated with DMEM low glucose, supplemented with 10% autologous serum (AS) plus 1% L-glutamine and 1% antibiotic/antimycotic. Senescence analysis was performed by ß-galactosidase staining after 24 and 48 h. Controls were established using BM-MSC from healthy donors and used for senescence and gene expression assays. Gene expression was performed using RT-PCR for pluripotency genes, such as SOX2, POU5F1, NANOG, and KLF4. MSC telomere length was measured by the Southern blotting technique, and MSCs were also analyzed for their capacity to differentiate into adipocytes, chondrocytes, and osteocytes. The patient's MSC expansion using AS displayed an early senescence state. In order to understand the role of AS in senescence, MSCs were then submitted to two different culture conditions: 1) with AS or 2) with FBS supplementation. Senescence state was assessed after 24 h, and no statistical differences were observed between the two conditions. However, patients' cells cultured with AS displayed a higher number of senescence cells than FBS medium after 48 h (p = 0.0018). Gene expression was performed in both conditions; increased expression of KLF4 was observed in the patient's cells in comparison to healthy controls (p = 0.0016); reduced gene expression was observed for NANOG (p = 0.0016) and SOX2 (p = 0.0014) genes. Telomere length of the patient's cells was shorter than that of a healthy donor and that of a patient of similar age. Osteocyte differentiation seemed to be more diffuse than that of the healthy donor and that of the patient of similar age. MSCs could enter a senescence state during expansion in early passages and can impact MSC quality for clinical applications, reducing their efficacy when administered.

Biomarkers of exposure, effect, and susceptibility in occupational exposure to traffic-related air pollution: A review.

There is a well-recognized association between environmental air pollution exposure and several human diseases. However, the relationship between diseases related to occupational air pollution exposure on roads and high levels of traffic-related air pollutants (TRAPs) is less substantiated. Biomarkers are essential tools in environmental and occupational toxicology, and studies on new biomarkers are increasingly relevant due to the need to determine early biomarkers to be assessed in exposure conditions. This review aimed to investigate the main advances in the biomonitoring of subjects occupationally exposed to air pollution, as well as to summarize the biomarkers of exposure, effect, and susceptibility. Furthermore, we discuss how biomarkers could be used to complement the current application of methods used to assess occupational exposures to xenobiotics present in air pollution. The databases used in the preparation of this review were PubMed, Scopus, and Science Direct. Considering the significant deleterious effects on health associated with chronic occupational exposure to xenobiotics, this topic deserves attention. As it is difficult to avoid occupational exposure to TRAPs, biomonitoring should be applied as a strategy to reduce the toxic effects of workplace exposure.

Cigarette smoking and antioxidant defences in packed red blood cells prior to storage.

BACKGROUND: Red blood cells from smoking donors can have more lesions from oxidative stress, decreasing the benefits of blood transfusion. We aimed to explore the effect of cigarette smoking on the oxidative status of packed red blood cells (PRBCs) prior to storage. MATERIALS AND METHODS: We compared serum vitamin C, plasmatic malondialdehyde (MDA), and non-protein thiol groups (GSH) levels in PRBCs, as well glutathione peroxidase (GPx) and glutathione s-transferase (GST) activity in PRBCs from smoking (n=36) and non-smoking (n=36) donors. We also correlated urinary cotinine levels with these parameters. RESULTS: Cigarette smoking was associated with decreased serum levels of vitamin C and GPx, and increased GST activity in PRBCs. We found negative correlations between cotinine, GPx activity and vitamin C levels, and a positive correlation between cotinine and GST activity. DISCUSSION: Cigarette smoking changed antioxidant defences of PRBCs prior to storage and these parameters are correlated with cotinine levels. Increased RBC antioxidants such as GST may reflect an exposure to oxidants during erythropoiesis. Because of the inability of mature RBCs to resynthesise antioxidants, PRBCs from smokers may have higher risk of storage lesions than those from non-smoker donors.

Evaluation of immunological, inflammatory, and oxidative stress biomarkers in gasoline station attendants.

BACKGROUND: Gasoline is a complex mixture of saturated and unsaturated hydrocarbons, in which aromatic compounds, such as BTX (benzene, toluene, and xylene) feature as the main constituents. Simultaneous exposure to these aromatic hydrocarbons causes a significant impact on benzene toxicity. In order to detect early alterations caused in gasoline station attendants exposed to BTX compounds, immunological, inflammatory, and oxidative stress biomarkers were evaluated. METHODS: A total of 66 male subjects participated in this study. The gasoline station attendants (GSA) group consisted of 38 gasoline station attendants from Rio Grande do Sul, Brazil. The non-exposed group consisted of 28 subjects who were non-smokers and who had no history of occupational exposure. Environmental and biological monitoring of BTX exposure was performed using blood and urine. RESULTS: The GSA group showed increased BTX concentrations in relation to the non-exposed group (p < 0.001). The GSA group showed elevated protein carbonyl (PCO) levels and pro-inflammatory cytokines, decreased expression of CD80 and CD86 in monocytes, and reduced glutathione S-transferase (GST) activity compared to the non-exposed group (p < 0.05). BTX levels and trans,trans-muconic acid levels were positively correlated with pro-inflammatory cytokines and negatively correlated with interleukin-10 contents (p < 0.001). Increased levels of pro-inflammatory cytokines were accompanied by increased PCO contents and decreased GST activity (p < 0.001). Furthermore, according to the multiple linear regression analysis, benzene exposure was the only factor that significantly contributed to the increased pro-inflammatory cytokines (p < 0.05). CONCLUSIONS: Taken together, these findings show the influence of exposure to BTX compounds, especially benzene, on the immunological, inflammatory, and oxidative stress biomarkers evaluated. Furthermore, the data suggest the relationship among the evaluated biomarkers of effect, which could contribute to providing early signs of damage to biomolecules in subjects occupationally exposed to BTX compounds.

Cytokines Levels and Salivary Microbiome Play A Potential Role in Oral Lichen Planus Diagnosis.

Oral lichen planus (OLP) is a chronic Th1-mediated inflammatory mucocutaneous disease of the skin and oral mucosa that can have various clinical presentations. Lesions are usually bilateral and often painful. While cutaneous Lichen Planus (LP) lesions are self-limiting, the oral lesions are chronic and rarely remissive. The diagnosis of oral lichen planus (OLP) is often challenging, and confirmation by histopathological criterion is generally advised. The aim of our study was to identify the cytokines present in OLP-suggestive lesions and in non-specific inflammatory lesions (NSIL) used as controls. Moreover, assess cytokines protein levels and oral microbiota composition in whole saliva samples. Histopathological analysis, immunohistochemistry and gene expression were used as techniques to analyze the oral mucosal tissue samples. ELISA was conducted to analyze salivary cytokine levels and 16S rRNA sequencing was used to determine the salivary microbiome. As a result we observed larger number of infiltrated lymphocytes (p = 0.025), as well, more T CD4 lymphocytes in the epithelial tissue (p = 0.006) in OLP samples compared to NSIL. In addition, the OLP samples displayed more apoptotic cells compared to NSIL (p = 0.047). Regarding the cytokine analysis, IFN-γ and IL-33 were more expressed in OLP lesions than in NSIL samples (p < 0.001; p = 0.026). Furthermore, our results demonstrated higher levels of IFN-γ protein expression in the saliva of OLP group compared to controls (p = 0.0156). We also observed noted differences in the oral microbiota composition between OLP and NSIL saliva samples. In conclusion, OLP lesions presented larger numbers of apoptotic and inflammatory cells, higher levels of IFN-γ and IL-33 compared to NSIL, and these lesions also differ regarding oral microbiota composition. These results are consistent with the Th-1-mediated chronic inflammation nature of oral lichen planus investigated lesions and displayed unique features that could be used as a diagnostic tool.

Cognitive deficits and ALA-D-inhibition in children exposed to multiple metals.

Children are especially vulnerable to adverse effects of multiple metals exposure. The aim of this study was to assess some metals concentrations such as lead (Pb), arsenic (As), chromium (Cr), manganese (Mn) and iron (Fe) in whole blood, serum, hair and drinking water samples using inductively coupled plasma-mass spectrometry (ICP-MS) in rural and urban children. In addition, evaluate the adverse effects of multiple metals exposure on cognitive function and δ-aminolevulinate dehydratase (ALA-D) activity. The cognitive ability assessment was performed by the Raven's Colored Progressive Matrices (RCPM) test. The ALA-D activity and ALA-D reactivation index (ALA-RE) activity with DTT and ZnCl2 also were determined. Forty-six rural children and 23 urban children were enrolled in this study. Rural children showed percentile IQ scores in the RCPM test significantly decreased in relation to urban children. According to multiple linear regression analysis, the Mn and Fe in hair may account for the cognitive deficits of children. Manganese and Fe in hair also were positively correlated with Mn and Fe in drinking water, respectively. These results suggest that drinking water is possibly a source of metals exposure in children. ALA-D activity was decreased and ALA-RE with DTT and ZnCl2 was increased in rural children in comparison to urban children. Moreover, ALA-D inhibition was correlated with Cr blood levels and ALA-RE/DDT and ALA-RE/ZnCl2 were correlated with levels of Cr and Hg in blood. Thus, our results indicated some adverse effects of children's exposure to multiple metals, such as cognitive deficits and ALA-D inhibition, mainly associated to Mn, Fe, Cr and Hg.

Evaluation of toxic metals and essential elements in children with learning disabilities from a rural area of southern Brazil.

Children's exposure to metals can result in adverse effects such as cognitive function impairments. This study aimed to evaluate some toxic metals and levels of essential trace elements in blood, hair, and drinking water in children from a rural area of Southern Brazil. Cognitive ability and δ-aminolevulinate dehydratase (ALA-D) activity were evaluated. Oxidative stress was evaluated as a main mechanism of metal toxicity, through the quantification of malondialdehyde (MDA) levels. This study included 20 children from a rural area and 20 children from an urban area. Our findings demonstrated increase in blood lead (Pb) levels (BLLs). Also, increased levels of nickel (Ni) in blood and increase of aluminum (Al) levels in hair and drinking water in rural children were found. Deficiency in selenium (Se) levels was observed in rural children as well. Rural children with visual-motor immaturity presented Pb levels in hair significantly increased in relation to rural children without visual-motor immaturity (p < 0.05). Negative correlations between BLLs and ALA-D activity and positive correlations between BLLs and ALA-RE activity were observed. MDA was significantly higher in rural compared to urban children (p < 0.05). Our findings suggest that rural children were co-exposed to toxic metals, especially Al, Pb and Ni. Moreover, a slight deficiency of Se was observed. Low performance on cognitive ability tests and ALA-D inhibition can be related to metal exposure in rural children. Oxidative stress was suggested as a main toxicological mechanism involved in metal exposure.