Biochemical profile of non-enzymatic stress markers in the plant species "Urginea maritima" in a Mediterranean natural reserve exposed to oxidative stress.

Protected areas decrease degrading natural ecosystems due to pollution such as air pollution. In 1981, the inhabitants founded Bentael natural reserve in Byblos, Lebanon, to secure their region against urbanization projects, like the recently constructed road that threatens the biodiversity of the reserve. This study was conducted to determine the oxidative stress resulting from this pollution and that menaces 360 floral species among them a rare species "Urginea maritima." In this research, the biomonitoring approach was experienced to assess the oxidative stress. Biomonitoring possesses has the advantage to be low cost and a constructive method to generate valuable data for further examinations. The studied parameters were air pollutants, ascorbic acid, photosynthetic pigments, leave's pH, relative water content, proline, carbohydrates, and hydrogen peroxide, in three chosen spots, near the pollution source (P1), opposite the latter spot (P2), and in an area relatively far from the source of contamination and which was chosen as the control site (Ctrl). The results showed in P1 detection of air pollutants higher of about 80% than in Ctrl, modifications in stress markers: increased concentration of the reactive oxygen species "hydrogen peroxide," rise in the concentration of the osmoregulator amino acid "proline," and depletion in chlorophyll content, in contrast to an increase in pheophytin. All these findings can be exploited as early diagnosis of air pollution and confirmed the ability to use such biomonitor ("Urginea maritima") as a way to assess the environmental pollution levels and consequently affirm the danger of such landscape activities on natural reserves.

Bone marrow-derived and synovium-derived mesenchymal cells promote Th17 cell expansion and activation through caspase 1 activation: contribution to the chronicity of rheumatoid arthritis.

OBJECTIVE: Th17 cells have been implicated in rheumatoid arthritis (RA). We hypothesized that the interaction of T cells with bone marrow-derived mesenchymal stem cells (BM-MSCs) or with fibroblast- like synoviocytes (FLS) might, with the help of T cell-secreted inflammatory cytokines (i.e., interleukin-17A [IL-17A], tumor necrosis factor α [TNFα], and/or interferon-γ [IFNγ]), promote Th17 cell expansion and activation. METHODS: Peripheral blood mononuclear cells (PBMCs) from healthy blood donors were cocultured with BM-MSCs or FLS from RA patients or osteoarthritis (OA) patients. Cocultures were exposed to phytohemagglutinin with or without IL-17A, TNFα, or IFNγ. Quantitative reverse transcription-polymerase chain reaction analysis, enzyme-linked immunosorbent assay, and cytofluorometry were used to measure IL-17A production. RESULTS: Interaction of PBMCs with BM-MSCs inhibited Th1 and Th2 responses, but promoted Th17 cell expansion, as early as 24 hours, as demonstrated by increases in retinoic acid receptor-related orphan nuclear receptor γ or IL-17A messenger RNA (mRNA) levels, IL-17A secretion levels, and IL-17A-secreting cell frequency, as well as by T cell switching to the Th17 pathway after 2 rounds of stimulation with MSCs. IL-17A production was also increased in PBMCs stimulated with anti-CD3 plus anti-CD28 or in isolated CD3+ or CD45RO+ T cells, thus demonstrating the role of T cell activation. Levels of mRNA for IL-6, IL-8, and IL-1ß were further amplified when T cell-secreted inflammatory cytokines were added. Interestingly, OA FLS or RA FLS also enhanced IL-17A and IL-6 production, but only RA FLS enhanced IFNγ and IL-1ß production. We further demonstrated that MSC-mediated Th17 promotion requires caspase 1 activation by using an inhibitory peptide and measuring its activity. CONCLUSION: We found that the interaction of MSCs or FLS with T cells promotes the activation and expansion of Th17 cells through caspase 1 activation. Since proinflammatory and T cell-secreted inflammatory cytokines are also amplified, this mechanism may participate in the chronicity of RA.

Uremic Serum Induces Inflammation in Cultured Human Endothelial Cells and Triggers Vascular Repair Mechanisms.

Inflammation and cardiovascular disease (CVD) are common in end-stage renal disease (ESRD) patients whose vascular endothelium is in direct contact with the uremic toxins found in the blood. These toxins are believed to affect vascular injury and repair process, which is impaired in ESRD patients. The exact mechanisms behind these interactions are not clear. So, we wanted to investigate what happens at the molecular level of endothelial cells when exposed to uremic serum from ESRD patients with diabetes and/or hypertension and its effect on the expression of molecules associated with vascular injury and repair. Cultured human endothelial cells (ECV304) were incubated in the presence of normal or uremic sera from ESRD patients with diabetes and/or hypertension. The expressions of monocyte chemoattractant protein 1 (MCP-1), vascular endothelial growth factor (VEGF), and stromal cell-derived factor 1 (SDF-1) were investigated in endothelial cells (ECV304) by real-time PCR and ELISA. The expression of MCP-1, VEGF, and SDF-1 was elevated in endothelial cells upon exposure to uremic sera from ESRD patients with diabetes and/or hypertension when compared with cells treated with healthy serum. MCP-1 expression in endothelial cells treated with uremic serum from ESRD patients with hypertension only was significantly increased compared with its expression in other cohorts. Exposure of endothelial cells to uremic serum causes endothelial injury and inflammation characterized by an increase in MCP-1 expression. This injury activates the initiation of vascular repair process in these cells by increasing the expression of VEGF and SDF-1. These molecules can be important biomarkers of chronic kidney disease-associated CVD.

Correlations between serum inflammatory markers and comorbidities in patients with end-stage renal disease.

OBJECTIVES: Chronic inflammatory processes are common in patients with renal disease, especially those with end-stage renal disease (ESRD), in whom inflammatory markers have been shown to increase with renal function deterioration. ESRD is usually accompanied by other chronic diseases such as hypertension and diabetes. The relationships between ESRD comorbidities and serum levels of inflammatory markers have not yet been fully understood. The aim of this study was to assess serum levels of inflammatory markers in different ESRD cohorts and to investigate the correlations between these inflammatory markers and disease comorbidities. METHODS: A total of 147 patients were grouped according to their comorbid conditions: diabetic only, hypertensive only, diabetic and hypertensive, and neither diabetic nor hypertensive. Serum levels of C-reactive protein (CRP), tumour necrosis factor-alpha (TNF-α), and interleukin-1-beta (IL-1ß) were investigated in different ESRD cohorts by enzyme-linked immunosorbent assay. RESULTS: Serum CRP and TNF-α levels were high in diabetic patients (p = 0.0001), hypertensive patients (p = 0.0001), and those who had both diseases (p = 0.0001), when compared to ESRD patients without these comorbidities. There was no significant change in serum IL-1ß levels between patients with diabetes mellitus and/or hypertension compared to patients who did not have these diseases. CONCLUSIONS: Our results showed that, in ESRD patients, CRP and TNF-α seem to be largely affected by patients' comorbidities, unlike IL-1ß, which might be affected more by the dialysis process even in the absence of comorbidities.

Increased sensitivity of rheumatoid synoviocytes to Schnurri-3 expression in TNF-α and IL-17A induced osteoblastic differentiation.

OBJECTIVE: To compare the effects of TNF-α and IL-17A on osteogenic differentiation of isolated fibroblast-like synoviocytes (FLS) from healthy donors, osteoarthritis (OA) and rheumatoid arthritis (RA) patients. METHODS: FLS were cultured in osteogenic medium, with and without TNF-α and/or IL-17A. Extracellular matrix mineralization was evaluated by alizarin red staining and alkaline phosphatase activity (ALP) measurement. mRNA expression was analyzed by qRT-PCR for Wnt5a, BMP2 and Runx2, genes associated with osteogenesis, for DKK1 and RANKL, genes associated with osteogenesis inhibition and Schnurri-3, a new critical gene in the cross talk with osteoclasts. IL-6 and IL-8 production was measured by ELISA. RESULTS: In osteogenic medium, matrix mineralization and increased ALP activity indicated that FLS can undergo osteogenic differentiation, which was increased with TNF-α and IL-17A. The expression of osteogenesis activators (BMP2 and Wnt5a) was increased with cytokines and that of the osteogenesis inhibitor DKK1 was decreased. There was no difference between all three cell types. In contrast, RA FLS were particularly sensitive to the synergistic increase of Shn3 with TNF-α and IL-17A. Levels of IL-6 and IL-8 were also higher for RA-FLS, compared to healthy and OA FLS. CONCLUSION: IL-17A and/or TNF-α treatment favor an osteogenesis induction in isolated FLS, independent of their origin. RA-FLS were more sensitive to the synergistic increase of Schnurri-3 expression. Combined with the higher levels of inflammation, this may in turn activate osteoclastogenesis, leading to increased bone destruction seen in destructive arthritis.

Differential Effects of IL-17A and TNF-α on Osteoblastic Differentiation of Isolated Synoviocytes and on Bone Explants from Arthritis Patients.

OBJECTIVE: TNF-α and IL-17A act on fibroblast-like synoviocytes (FLS) and contribute to cytokine production, inflammation, and tissue destruction in rheumatoid arthritis (RA). The aim of this study was to compare their effects on osteogenic differentiation of isolated FLS and on whole bone explants from RA and osteoarthritis (OA) patients. METHODS: Fibroblast-like synoviocytes and bone explants were cultured in the presence or absence of TNF-α and/or IL-17A. Mineralization of extracellular matrix of FLS was measured by alizarin red and alkaline phosphatase activity (ALP). mRNA expression was analyzed by qRT-PCR for Wnt5a, BMP2, and RUNX2, key genes associated with osteogenesis. IL-6 and IL-8 levels were measured by enzyme-linked immunosorbent assays. Bone explant structure was quantified by histomorphometry. RESULTS: In isolated OA and RA FLS, the combination of TNF-α and IL-17A induced matrix mineralization, increased ALP activity and expression of the osteogenesis-associated genes Wnt5a, BMP2, and Runx2, indicating an osteogenic differentiation. Wnt5a levels increased with TNF-α alone and in combination with IL-17A. BMP2 expression decreased with IL-17A and TNF-α after 12 h with OA FLS and 24 h with RA FLS. Runx2 expression decreased only with combination of TNF-α and IL-17A in OA FLS and with cytokines alone and combined in RA FLS. IL-6 and IL-8 production increased with IL-17A and/or TNF-α in both FLS and bone samples, especially from RA. Treatment of bone explants with cytokine combination increased ALP in OA but not RA samples. A decrease in bone volume was seen with cytokine combination, especially with RA explants. CONCLUSION: Differences were observed for the effects of IL-17A and TNF-α on osteogenic differentiation. In isolated FLS, increased osteoblastogenesis was observed, contrasting with the inhibitory effect in whole bone, specifically in RA. The net effect of IL-17A and TNF-α appears to depend on the disease state and the presence of other cells.

Classical and Paradoxical Effects of TNF-α on Bone Homeostasis.

Tumor necrosis factor-α (TNF-α) plays an essential role in the regulation of bone homeostasis in several chronic immune and inflammatory joint diseases, where inhibition of TNF has led to significant clinical improvement. However, TNF-activated pathways and mechanisms involved in bone remodeling remain unclear. So far, TNF-α was known as an inhibitor of osteoblast differentiation and an activator of osteoclastogenesis. Recent contradictory findings indicated that TNF-α can also activate osteoblastogenesis. The paradoxical role of TNF-α in bone homeostasis seems to depend on the concentration and the differentiation state of the cell type used as well as on the exposure time. This review aims to summarize the recent contradictory findings on the regulation of bone homeostasis by TNF-α at the isolated cell, whole bone, and whole body levels. In addition, the involvement of TNF-α in the bone remodeling imbalance is observed in inflammatory joint diseases including rheumatoid arthritis and ankylosing spondylitis, which are associated with bone destruction and ectopic calcified matrix formation, respectively. Both diseases are associated with systemic/vertebral osteoporosis.

Effects of Interleukin-17A on Osteogenic Differentiation of Isolated Human Mesenchymal Stem Cells.

OBJECTIVES: Rheumatoid arthritis (RA) is characterized by defective bone repair and excessive destruction and ankylosing spondylitis (AS) by increased ectopic bone formation with syndesmophytes. Since TNF-α and IL-17A are involved in both diseases, this study investigated their effects on the osteogenic differentiation of isolated human bone marrow-derived mesenchymal stem cells (hMSCs). METHODS: Differentiation of hMSCs into osteoblasts was induced in the presence or absence of IL-17A and/or TNF-α. Matrix mineralization (MM) was evaluated by alizarin red staining and alkaline phosphatase (ALP) activity. mRNA expression was measured by qRT-PCR for bone morphogenetic protein (BMP)-2 and Runx2, genes associated with osteogenesis, DKK-1, a negative regulator of osteogenesis, Schnurri-3 and receptor activator of nuclear factor kappa B ligand (RANKL), associated with the cross talk with osteoclasts, and TNF-α receptor type I and TNF-α receptor type II (TNFRII). RESULTS: TNF-α alone increased both MM and ALP activity. IL-17A alone increased ALP but not MM. Their combination was more potent. TNF-α alone increased BMP2 mRNA expression at 6 and 12 h. These levels decreased in combination with IL-17A at 6 h only. DKK-1 mRNA expression was inhibited by TNF-α and IL-17A either alone or combined. Supporting an imbalance toward osteoblastogenesis, RANKL expression was inhibited by TNF-α and IL-17A. However, TNF-α but not IL-17 alone decreased Runx2 mRNA expression at 6 h. In parallel, TNF-α but not IL-17 alone increased Schnurri-3 expression with a synergistic effect with their combination. This may be related to an increase of TNFRII overexpression. CONCLUSION: IL-17 increased the effects of TNF-α on bone matrix formation by hMSCs. However, IL-17 decreased the TNF-α-induced BMP2 inhibition. Synergistic interactions between TNF-α and IL-17 were seen for RANKL inhibition and Schnurri-3 induction. Such increase of Schnurri-3 may in turn activate osteoclasts leading to bone destruction as in RA. Conversely, in the absence of osteoclasts, this could promote ectopic bone formation as in AS.

Analysis of epidermal lipids in normal and atopic dogs, before and after administration of an oral omega-6/omega-3 fatty acid feed supplement. A pilot study.

Alterations of the lipid expression in the skin of human and canine atopic subjects may be one of the key factors in the disease development. We have analyzed the ultrastructure of the clinically uninvolved skin of atopic dogs and compared it with the lipid composition of their tape-stripped stratum corneum (SC). The effect of a 2 month treatment of atopic dogs by food supplementation with a mixture of essential fatty acids was evaluated on skin samples taken before and after the treatment period. Electron microscopy revealed that the non-lesional skin of atopic dogs exhibited an abnormal and largely incomplete structure of the lamellar lipids with little cohesion between the corneocyte strata. The SC of atopic dogs was characterized by a significant decrease in the lipid content when compared to the healthy controls. Following oral supplementation with the mixture of essential fatty acids, the overall lipid content of the SC markedly increased. This feature was observed both with the free and, most importantly, with the protein-bound lipids (cholesterol, fatty acids and ceramides), the latter constituting the corneocyte-bound scaffold for ordinate organisation of the extracellular lipid bi-layers. Indeed, the semi-quantitative electron microscopy study revealed that the treatment resulted in a significantly improved organization of the lamellar lipids in the lower SC, comparable to that of the healthy dogs. Our results indicate the potential interest of long-term alimentary supplementation with omega-6 and omega-3 essential fatty acids in canine atopic dermatitis.