Carvacrol promotes angiogenic paracrine potential and endothelial differentiation of human mesenchymal stem cells at low concentrations.

OBJECTIVES: Phenolic monoterpene compound, named Carvacrol, has been found to exert different biological outcomes. It has been accepted that the angiogenic activity of human mesenchymal stem cells was crucial in the pursuit of appropriate regeneration. In the current experiment, we investigated the contribution of Carvacrol on the angiogenic behavior of primary human mesenchymal stem cells. METHODS: Mesenchymal stem cells were exposed to Carvacrol in a dose ranging from 25 to 200µM for 48h. We measured cell survival rate by MTT assay and migration rate by a scratch test. The oxidative status was monitored by measuring SOD, GPx activity. The endothelial differentiation was studied by evaluating the level of VE-cadherin and vWF by real-time PCR and ELISA analyses. The content of VEGF and tubulogenesis behavior was monitored in vitro. We also conducted Matrigel plug in vivo CAM assay to assess the angiogenic potential of conditioned media from human mesenchymal stem cells after exposure to Carvacrol. RESULTS: Carvacrol was able to increase mesenchymal stem cell survival and migration rate (p<0.05). An increased activity of SOD was obtained while GPx activity unchanged or reduced. We confirmed the endothelial differentiation of stem cells by detecting vWF and VE-cadherin expression (p<0.05). The VEGF expression was increased and mesenchymal stem cells conditioned media improved angiogenesis tube formation in vitro (p<0.05). Moreover, histological analysis revealed an enhanced microvascular density at the site of Matrigel plug in CAM assay. CONCLUSIONS: Our data shed lights on the possibility of a Carvacrol to induce angiogenesis in human mesenchymal stem cells by modulating cell differentiation and paracrine angiogenic response.

Copper sulfate pentahydrate reduced epithelial cytotoxicity induced by lipopolysaccharide from enterogenic bacteria.

The over usage of multiple antibiotics contributes to the emergence of a whole range of antibiotic-resistant strains of bacteria causing enterogenic infections in poultry science. Therefore, finding an appropriate alternative natural substance carrying an antibacterial capacity would be immensely beneficial. It has been previously discovered that the different types of cupric salts, especially copper sulfate pentahydrate (CuSO4·5H2O), to carry a potent bactericidal capacity. We investigated the neutralizing effect of CuSO4·5H2O (6.25µg/ml) on the reactive oxygen species generation, and expression of MyD88, an essential adaptor protein of Toll-like receptor, and NF-κB in three intestinal epithelial cell lines exposed to 50ng/ml lipopolysaccharide. In order to find the optimal cupric sulfate concentration without enteritis-inducing toxicity, broiler chickens were initially fed with water containing 0.4, 0.5, and 1mg/l during a period of 4days. After determination of appropriate dosage, two broiler chickens and turkey flocks with enteritis were fed with cupric compound for 4days. We found that cupric sulfate can lessen the cytotoxic effect of lipopolysaccharide by reducing the reactive oxygen species content (p<0.05). Additionally, the expression of MyD88 and NF-κB was remarkably down-regulated in the presence of lipopolysaccharide and cupric sulfate. The copper sulfate in doses lower than 0.4mg/ml expressed no cytotoxic effect on the liver, kidney, and the intestinal tract while a concentration of 0.5 and 1mg/ml contributed to a moderate to severe tissue injuries. Pearson Chi-Square analysis revealed the copper cation significantly diminished the rate of mortality during 4-day feeding of broiler chicken and turkey with enteritis (p=0.000). Thus, the results briefed above all confirm the potent anti-bactericidal feature of cupric sulfate during the course of enteritis.

NFκBP65 transcription factor modulates resistance to doxorubicin through ABC transporters in breast cancer.

BACKGROUND: Shedding light on chemoresistance biology of breast cancer could contribute to enhance the clinical outcome. Intrinsic or acquired resistance to chemotherapy is a major problem in breast cancer treatment. METHODS AND MATERIALS: The NFκB pathway by siRNAP65 and JSH-23 as a translocational inhibitor of NFκBP65 in the doxorubicin-resistant MCF-7 (MCF-7/Dox) and MCF-7 cells was blocked. Then, the ABC transporter expression and function were assessed by real-time qRT-PCR and flow cytometry, respectively. Induction of apoptosis was evaluated after inhibition of the NFΚB pathway as well. RESULTS: Our study underlined the upregulation of NFκBP65 and anti-apoptotic Bcl-2 and downregulation of pro-apoptotic Bax in the MCF-7/Dox cells compared with control MCF-7 cells. Here, we showed that interplay between nuclear factor kappa B P65 (NFkBP65) as a transcriptional regulator and ABC transporters in the MCF-7/Dox cancer cells. We found that inhibition of the elevated expression of NFκBP65 in the resistant breast cancer, whether translocational inhibition or silencing by siRNA, decreased the expression and function of MDR1 and MRP1 efflux pumps. Furthermore, the blockade of NFκBP65 promoted apoptosis via modulating Bcl-2 and BAX expression. After inhibition of the NFκBP65 signaling pathway, elevated baseline expression of survival Bcl-2 gene in the resistant breast cells significantly decreased. CONCLUSION: Suppression of the NFκB pathway has a profound dual impact on promoting the intrinsic apoptotic pathway and reducing ABC transporter function and expression, which are some of the chemoresistance features. It was speculated that the NFκB pathway directly acts on doxorubicin-induced MDR1 and MRP1 expression in MCF-7/Dox cells.

Contributory Anti-Inflammatory Effects of Mesenchymal Stem Cells, Not Conditioned Media, On Ovalbumin-Induced Asthmatic Changes in Male Rats.

Our aim in selecting an appropriate cell fraction and conditioned media (CM) was to achieve the suitable candidate for ameliorating long-term chronic asthmatic changes of respiratory tract. Thirty-six rats were classified into healthy and sensitized groups, which were further divided into three subgroups; rats received systemically 50 µl volume of PBS, CM, or 2 × 106 rat bone marrow-derived mesenchymal stem cells (rBMMSCs). Tracheal responsiveness (TR), immunologic responses, and recruitment of rBMMSCs into the lungs were evaluated. A high degree of TR and total WBC and percentages of eosinophils and neutrophils was significantly recorded in all sensitized groups rather than of controls (p < 0.001 to p < 0.05). Concurrently, a significant improvement of TR and eosinophil and neutrophil return toward normal levels was evident in sensitized rats receiving cells as compared to parallel asthmatic animals. Flow cytometric monitoring of lymphocyte subpopulation revealed a decrease in the number of CD3+CD4+ and concurrent increase in CD3+CD8+ in all sensitized rats as compared to control (p < 0.001 to p < 0.05). Noticeably, no significant modulatory effects of either cell or CM administration were achieved on the CD3+CD4+ and CD3+CD8+ populations in non-asthmatic rats. Corroborating our results, the number of CD3+CD4+ tended to increase (p < 0.05) which coincided with a decreased manner of CD3+CD8+ populations as compared to other asthmatic groups (p < 0.01 to p < 0.05). Moreover, stem cells could efficiently transmigrate to the lung parenchyma, albeit the dynamic of asthmatic changes stimulated the rate of recruited cells. Our study shed light on superior effects of mesenchymal stem cells, but not CM, in attenuating chronic asthmatic changes in the model of rat.

Taq1B Polymorphism of Cholesteryl Ester Transfer Protein (CETP) and Its Effects on the Serum Lipid Levels in Metabolic Syndrome Patients.

The metabolic syndrome (MetS) is one of the most important risk factors for type 2 diabetes and cardiovascular disease. This syndrome is characterized by abdominal obesity, hypertension, insulin resistance, and dyslipidemia. The plasma origin of Cholesteryl ester transfer protein (CETP) is responsible for transferring cholesterol esters from high-density lipoprotein particles to apolipoprotein B containing lipoproteins compartment. We conducted this study to investigate the association between CETP gene Taq1B (rs708272) polymorphism in the metabolic syndrome among Iranian subjects. A sample size of 200 patients diagnosed with MetS together with 200 healthy donors as control were enrolled in this study. The investigation of polymorphism was performed by the use of polymerase chain reaction and restriction fragment length polymorphism analysis. To determine the relationship between polymorphism and lipid profile, we measured lipids and CETP concentration in metabolic syndrome and control subjects. Genotype distribution and allelic frequencies of polymorphism were determined and compared in both groups. Our findings showed that all clinical and biochemical characteristics in patients differed from the control group. The results showed that genotype and allele frequency of the Taq1B polymorphism was not significantly different between two groups. Instinctively, CETP was significantly higher in metabolic syndrome (1.64 ± 0.32 µg/ml) than in control (1.53 ± 0.34 µg/ml). A low level of CETP was found in blood of B2B2 typified genotype. In spite of Taq1B polymorphism on ester transfer protein concentration, no direct correlation was found between this polymorphism and metabolic syndrome.

Effects of combination of melatonin and laser irradiation on ovarian cancer cells and endothelial lineage viability.

The main goal of anti-cancer therapeutic approaches is to induce apoptosis in tumor masses but not in the normal tissues. Nevertheless, the combination of photodynamic irradiation with complementary oncostatic agents contributes to better therapeutic performance. Here, we applied two different cell lines; SKOV3 ovarian carcinoma cells and HUVECs umbilical cord cells as in vitro models to pinpoint whether pharmacological concentration of melatonin in combination with photodynamic therapy induces cell cytotoxicity. The cells were separately treated with various concentrations of melatonin (0 to 10 mM) and photodynamic irradiation alone or in combination. Cells were preliminary exposed to increasing concentrations of melatonin for 24 h and subsequently underwent laser irradiation for 60 s with an output power of 80 mW in continuous mode at 675 nm wavelength and a total light dose of 13.22 J/cm2. Cell viability, apoptosis/necrosis rates, and reactive oxygen species levels as well as heat shock protein 70 expression were monitored after single and combined treatments. A statistical analysis was performed by applying one-way analysis of variance (ANOVA) and post hoc Tukey's test. Combination treatment of both cell lines caused a marked increase in apoptosis/necrosis rate, reactive oxygen species generation, and heat shock protein 70 expression compared to incubation of the cells with each agent alone (p < 0.05). SKOV3 cancer cells expressed higher level of heat shock protein 70 under experimental procedure as compared to HUVECs (p < 0.05). Our results introduce melatonin as a potent stimulus for enhancing the efficacy of laser on induction of apoptosis in tumor cells.