Expression of neural markers by undifferentiated mesenchymal-like stem cells from different sources

The spontaneous expression of neural markers, already demonstrated in bone marrow (BM) mesenchymal stem cells (MSCs), has been considered as evidence of the MSCs' predisposition to differentiate toward neural lineages, supporting their use in stem cell-based therapy for neural repair. In this study we have evaluated, by immunocytochemistry, immunoblotting, and flow cytometry experiments, the expression of neural markers in undifferentiated MSCs from different sources: human adipose stem cells (hASCs), human skin-derived mesenchymal stem cells (hS-MSCs), human periodontal ligament stem cells (hPDLSCs,) and human dental pulp stem cells (hDPSCs). Our results demonstrate that the neuronal markers \u3b2III-tubulin and NeuN, unlike other evaluated markers, are spontaneously expressed by a very high percentage of undifferentiated hASCs, hS-MSCs, hPDLSCs, and hDPSCs. Conversely, the neural progenitor marker nestin is expressed only by a high percentage of undifferentiated hPDLSCs and hDPSCs. Our results suggest that the expression of \u3b2III-tubulin and NeuN could be a common feature of stem cells and not exclusive to neuronal cells. This could result in a reassessment of the use of \u3b2III-tubulin and NeuN as the only evidence proving neuronal differentiation. Further studies will be necessary to elucidate the relevance of the spontaneous expression of these markers in stem cells

Monitoring the genomic stability of in vitro cultured rat bone-marrow-derived mesenchymal stem cells

Bone-marrow-derived mesenchymal stem cells (MSCs) are multipotent cells capable of self-renewal and differentiation into multiple cell types. Accumulating preclinical and clinical evidence indicates that MSCs are good candidates to use as cell therapy in many degenerative diseases. For MSC clinical applications, an adequate number of cells are necessary so an extensive expansion is required. However, spontaneous immortalization and malignant transformation of MSCs after culture expansion have been reported in human and mouse, while very few data are present for rat MSCs (rMSCs). In this study, we monitored the chromosomal status of rMSCs at several passages in vitro, also testing the influence of four different cell culture conditions. We first used the conventional traditional cytogenetic techniques, in order to have the opportunity to observe even minor structural abnormalities and to identify low-degree mosaic conditions. Then, a more detailed genomic analysis was conducted by array comparative genomic hybridization. We demonstrated that, irrespective of culture conditions, rMSCs manifested a markedly aneuploid karyotype and a progressive chromosomal instability in all the passages we analyzed and that they are anything but stable during in vitro culture. Despite the fact that the risk of neoplastic transformation associated with this genomic instability needs to be further addressed and considering the apparent genomic stability reported for in vitro cultured human MSCs (hMSCs), our findings underline the fact that rMSCs may not in fact be a good model for effectively exploring the full clinical therapeutic potential of hMSCs

Phytochemical Investigation, Antiulcer, Cyclooxygenase-2, and 15-Lipoxygenase Inhibitory Activities of Echinops erinaceus Kit Tan

Plants of the genus Echinop (Asteraceae) are traditional medicinal plants used to treat several GIT ailments, owing to their diverse bioactive secondary metabolites, including sesquiterpenoids, triterpenoids, phytosterols, phenolics, flavonoids, alkaloids, and essential oils. Echinops erinaceus Kit Tan is a wild perennial herb of the genus Echinops which is endemic to Oman, Saudi Arabia, and Yemen. Currently, there are no previous reports exploring its anti-ulcer and anti-inflammatory effects. Additionally, few reports have described the chemical profile of E. erinaceus Kit Tan. In the current study, the CHCl3 fraction of the aerial parts of the plant was subjected to chromatographic isolation and spectroscopic identification via 1D and 2D NMR, and MS. The plant afforded two new compounds, designated erinaceolic acid (E3) and erinaceoside (E5), in addition to five known compounds, namely taraxasterol acetate (E1), taraxasterol (E2), apigenin (E4), stigmasterol-3-O-β-D-glucoside (E6), and speranskoside (E7). The evaluation of the gastric ulcer protective activity of the total extract and successive fractions of E. erinaceus, using the in vivo ethanol-induced ulcer in rats model, revealed the significant effect of the tested extracts and fractions on the percentage of gastric ulcer protection and ulcer index (500 mg/kg) compared to antodine (20 mg/kg). The tested extracts and fractions also reduced the stomach contents of TNF-α and reduced IL-6 as compared to the untreated group. Histopathological examination of the gastric mucosal tissues of rats supportedprevious results. In addition, the main subfractions and their isolates were assessed for their in vitro anti-inflammatory activity against COX-2 and 15-LOX enzymes. The new compounds erinaceolic acid (E3) and speranskoside (E7) exhibited strong inhibition against COX-2 (3.41 and 2.62 µg/mL) and 15-LOX (10.05 and 5.51 µg/mL), respectively. A molecular docking study was performed to reveal the binding interaction modes of the most active compounds against the binding sites of COX-2 (PDB ID 3LN1) and 15-LOX (PDB ID 1LOX) proteins. Speranskoside (E7) showed a dual binding affinity better than that of the cocrystallized references, celecoxib and (2E)-3-(2-oct-1-yn-1-ylphenyl)acrylic acid (RS7) against both enzymes. This study shed a light on the potential use of E. erinaceus in the protection and treatment of gastric ulcers

Phytochemical, Antimicrobial, Antioxidant, and In Vitro Cytotoxicity Evaluation of Echinops erinaceus Kit Tan

Wild plants are used by many cultures for the treatment of diverse ailments. However, they are formed from mixtures of many wanted and unwanted phytochemicals. Thus, there is a necessity to separate the bioactive compounds responsible for their biological activity. In this study, the chemical composition as well as antimicrobial and cytotoxic activities of Echinops erinaceus Kit Tan (Asteraceae) were investigated. This led to the isolation and identification of seven compounds, two of which are new (erinaceosin C3 and erinaceol C5), in addition to methyl oleate (C1) and ethyl oleate (C2), loliolide (C4), (E)-p-coumaric acid (C6), and 5,7,3`,5`-tetrahydroxy flavanone (C7). The structures of the isolated compounds were elucidated by 1D, 2D NMR, and HR-ESI-MS. The methanol extract showed the highest antimicrobial activity among the tested extracts and fractions. The n-hexane and EtOAc extracts showed remarkable antimicrobial activity against B. subtilus, P. aeruginosa, E. coli, and C. albicans. A cytotoxicity-guided fractionation of the most bioactive chloroform extract resulted in the isolation of bioactive compounds C1/C2, which showed significant cytotoxicity against HCT-116 and CACO2 cell lines (IC50 24.95 and 19.74 µg/mL, respectively), followed by compounds C3 (IC50 82.82 and 76.70 µg/mL) and C5 (IC50 99.09 and 87.27 µg/mL), respectively. The antioxidant activity of the bioactive chloroform fractions was screened. Molecular docking was used to explain the results of the antimicrobial and anticancer activities against five protein targets, including DNA gyrase topoisomerase II, enoyl-acyl carrier protein reductase of S. aureus (FabI), dihydrofolate reductase (DHFR), β-catenin, and human P-glycoprotein (P-gp)

Evaluation of tubulin polymerization and chronic inhibition of proteasome as citotoxicity mechanisms in bortezomib-induced peripheral neuropathy.

Bortezomib (BTZ) is the first proteasome inhibitor entered in clinical practice. Peripheral neuropathy is likely to be a class side effect of these drugs, although its severity is largely variable, and it deserves to be further investigated, since the mechanisms of BTZ-induced peripheral neurotoxicity (BiPN) are still unknown.   In our study, we investigated in vivo and in vitro possible pathogenic events relevant to BiPN using a well-established rat model, with particular reference to the extent of proteasome inhibition and the effects on α-tubulin polymerization in sciatic nerves and dorsal root ganglia specimens obtained from animals treated with chronic regimens at a dose of 0.2 mg/kg intravenously. The same assessments were also performed after a single injection. Moreover, these studies were replicated in vitro using embryonic DRG neurons exposed to 100 nM BTZ and adult DRG neurons exposed to 10-50 nM BTZ for 24 h and 48 h. A significant increase in the polymerized fraction of α-tubulin and prolonged proteasome inhibition were observed after the chronic BTZ treatment in vivo. Recovery to physiological levels was observed after a 4-week follow-up post-treatment period. Proteasome inhibition and increased α-tubulin polymerization were also observed following BTZ treatment of both embryonic and adult DRG neurons in vitro. Our in vivo results suggest that proteasome inhibition and alteration of tubulin dynamics contribute to BiPN. The in vitro systems here described reliably replicate the in vivo results, and might therefore be used for further mechanistic studies on the effects of proteasome inhibitors on neurons

Phytochemical, Antimicrobial, Antioxidant, and In Vitro Cytotoxicity Evaluation of <i>Echinops erinaceus</i> Kit Tan

Wild plants are used by many cultures for the treatment of diverse ailments. However, they are formed from mixtures of many wanted and unwanted phytochemicals. Thus, there is a necessity to separate the bioactive compounds responsible for their biological activity. In this study, the chemical composition as well as antimicrobial and cytotoxic activities of Echinops erinaceus Kit Tan (Asteraceae) were investigated. This led to the isolation and identification of seven compounds, two of which are new (erinaceosin C3 and erinaceol C5), in addition to methyl oleate (C1) and ethyl oleate (C2), loliolide (C4), (E)-p-coumaric acid (C6), and 5,7,3`,5`-tetrahydroxy flavanone (C7). The structures of the isolated compounds were elucidated by 1D, 2D NMR, and HR-ESI-MS. The methanol extract showed the highest antimicrobial activity among the tested extracts and fractions. The n-hexane and EtOAc extracts showed remarkable antimicrobial activity against B. subtilus, P. aeruginosa, E. coli, and C. albicans. A cytotoxicity-guided fractionation of the most bioactive chloroform extract resulted in the isolation of bioactive compounds C1/C2, which showed significant cytotoxicity against HCT-116 and CACO2 cell lines (IC50 24.95 and 19.74 µg/mL, respectively), followed by compounds C3 (IC50 82.82 and 76.70 µg/mL) and C5 (IC50 99.09 and 87.27 µg/mL), respectively. The antioxidant activity of the bioactive chloroform fractions was screened. Molecular docking was used to explain the results of the antimicrobial and anticancer activities against five protein targets, including DNA gyrase topoisomerase II, enoyl-acyl carrier protein reductase of S. aureus (FabI), dihydrofolate reductase (DHFR), β-catenin, and human P-glycoprotein (P-gp)