Journal: Cell biology international
To examine cytokine production in response to RSV infection, we assessed the levels of 29 cytokines released from RSV-infected human foetal lung fibroblasts. We also examined the relationships between the effects of fluticasone propionate and various signalling pathways in the cells. Twenty-four hours after infection (1MOI), RSV-infected cells released cytokines, for example proinflammatory cytokines (IL-1β, IL-6 and TNF-α), anti-inflammatory (IL-1ra), Th1 (IFN-γ, IFN-λ1a, IL-2 and IL-12), Th2 (IL-4, IL-5, IL-10 and IL-13), granulopoiesis-inducing (G-CSF and GM-CSF), eosinophil recruitment-inducing (eotaxin and RANTES) and neutrophil recruitment-inducing cytokines (IL-8, IP-10, MCP-1 and MIP-1α). Aberrant release of most was significantly suppressed by fluticasone propionate. Twelve hours after RSV infection, increased phosphorylation of Akt, p38 MAPK, ERK1/2 and IκB-α was noted. Fluticasone propionate suppressed the phosphorylation of Akt, p38 MAPK, and ERK1/2, but not IκB-α, in virus-infected cells. TLR-4 expression was unchanged in control and RSV-infected cells, and TLR-3 and RIG-I expression was not detected. The results indicate that RSV infection induces aberrant production and release of certain cytokines through these signalling pathways in human lung fibroblasts. Overproduction and imbalance of these cytokines may be associated with the pathophysiology of RSV-induced excessive and allergic inflammation.
MicroRNA-455 (miRNA-455), which is downregulated in human cancer, potently mediates the multiple steps of carcinogenesis. However, the role of miR-455 in non-small cell lung cancer (NSCLC) carcinogenesis remains unclear. In present study, we determined the mature miRNA-455 expression in NSCLC tissues and cells by real-time PCR. Follow-up studies examined the effects of a miR-455 mimic (gain of function) on cell proliferation, migration and invasion. Our data indicate that miR-455 was significantly down-regulated in NSCLC cell lines and tissues. In functional assays, overexpression of miR-455 suppressed the proliferation, migration and invasion of NSCLC cell lines. Data from reporter assays showed that miR-455 directly binds to 3'UTR of ZEB1 and suppresses the endogenous level of ZEB1 protein expression. Furthermore, overexpression of ZEB1 reverses miR-455-suppressed malignant phenotype of NSCLC cells. Moreover, we found that upregulation of ZEB1 expression is inversely associated with miR-455 expression in NSCLC tissues. Taken together, miR-455 as an anti-oncogene in non-small cell lung cancer through up-regulation of ZEB1 and serve as a potential therapeutic target in NSCLC.
Parkinson’s disease (PD), the second-most prevalent neurodegenerative disease, is primarily characterized by neurodegeneration in the substantia nigra pars compacta, resulting in motor impairment. Loss-of-function mutations in parkin are the major cause of the early-onset familial form of the disease. Although rodents deficient in parkin (parkin(-/-)) have some dopaminergic system dysfunction associated with central oxidative stress and energy metabolism deficiencies, these animals only display nigrostriatal pathway degeneration under inflammatory conditions. This study investigated the impact of the inflammatory stimulus induced by lypopolisaccharide (LPS) on tetrahydrobiopterin (BH4) synthesizing enzymes (de novo and salvage pathways), since this cofactor is essential for dopamine synthesis. The mitochondrial content and architecture was investigated in the striatum of LPS-exposed parkin(-/-) mice. As expected, the LPS (0.33 mg / kg; i.p.) challenge compromised spontaneous locomotion and social interaction with juvenile parkin(-/-) and WT mice. Moreover, the genotype impacted the kinetics of the investigation of the juvenile. The inflammatory scenario did not induce apparent changes in mitochondrial ultrastructure; however, it increased the quantity of mitochondria, which were of smaller size, and provoked the perinuclear distribution of the organelle. Furthermore, the BH4 de novo biosynthetic pathway failed to be up-regulated in the LPS challenge, a well-known stimulus for its activation. The LPS treatment increased sepiapterin reductase (SPR) expression, suggesting compensation by the salvage pathway. This might indicate that dopamine synthesis is compromised in parkin(-/-) mice under inflammatory conditions. Finally, this scenario impaired the striatal expression of the transcription factor BDNF, possibly favoring cell death.
MicroRNAs have been known to function as important regulators in the vascular system, with various physiopathological effects such as vascular remodeling and hypertension modulation. We aimed to explore whether microRNA-150 (miR-150) regulates endothelial cell function and vascular remodeling in acute coronary syndrome (ACS), and the involvement of PTX3 and NF-κB signaling pathway. Ten normal mice and sixty ApoE-/- mice were chosen, and their coronary artery tissues and endothelial cells were extracted. ApoE-/- mice were injected with a series of inhibitors or mimics for miR-150, or siRNA against PTX3. The miR-150 expression, NF-κB1, RELA, and PTX3 mRNA expressions were assessed by reverse transcription quantitative polymerase chain reaction, and pentraxin-3, p-P50, and p-P65 protein expression by western blot analysis. Cell viability and migration were assessed by MTT assay and scratch test. Matrigel tube formation assay was employed to determine vascular remodeling of endothelial cells. The dual-luciferase reporter assay verified that PTX3 was a target of miR-150. Mice with ACS presented with decreased miR-150 but increased PTX3. It was observed that the miR-150 mimics and siRNA against PTX3 reduced levels of PTX3, NF-κB1 and RELA in mice, and the miR-150 inhibitor reversed the tendency. The in vitro cell experimentation proved that miR-150 might facilitate endothelial cell proliferation, migration, and restrain vascular remodeling via inhibiting PTX3 expression. On the basis of the results of this study, it was hypothesized that miR-150 could possibly maintain endothelial cell function and suppress vascular remodeling by inhibiting PTX3 through the NF-κB signaling pathway in mice with ACS.
Colorectal cancer is one of the global causes of cancer deaths. Cancer stem cells (CSCs) inside the tumour niche responsible for metastasis and relapses, and hence need to be targeted for cancer therapeutics. Although dietary fibre and lifestyle changes have been recommended as measures for colorectal cancer prevention, no such recommendations are available for using water soluble vitamins as prophylaxis measure for colorectal cancers. High dose of Vitamin C has been proven to selectively kill colon cancer cells having BRAF and KRAS mutations by inducing oxidative stress. In this study, we show for the first time the opposing effects of the low and high dose of Vitamin C and vitamin B3 on colon CSCs isolated from HT- 29 and HCT-15 colorectal carcinoma cell lines. At small doses, both of these vitamins exerted a cell proliferative effect only on CSCs, while there was no change in the proliferation status of non-stem cancer cells and wild-type (WT) populations. On the other hand, the death effects induced by high doses of Vitamin C and B3 were of the order of 50-60% and ∼30% on CSCs from HT-29 and HCT15 respectively. Interestingly, the control fibroblast cell line (NIH3T3) was highly refractory all the tested concentrations of Vitamin C and B3, except for the highest dose- 10,000 µg of Vitamin C that induced only 15% of cell death. Hence, these results indicate the future scope of use of therapeutic doses of Vitamin C and B3 especially in patients with advanced colorectal cancer.
Sutherlandia frutescens is a medicinal plant, traditionally used to treat various types of human diseases, including cancer. Previous studies of several botanicals link suppression of prostate cancer growth with inhibition of the Gli/hedgehog (Gli/Hh) signaling pathway. Here we hypothesized the anti-cancer effect of S. frutescens was linked to its inhibition of the Gli/Hh signaling in prostate cancer. We found a dose- and time-dependent growth inhibition in human prostate cancer cells, PC3 and LNCaP, and mouse prostate cancer cell, TRAMP-C2, treated with S. frutescens methanol extract (SLE). We also observed a dose-dependent inhibition of the Gli-reporter activity in Shh Light II and TRAMP-C2QGli cells treated with SLE. In addition, SLE can inhibit Gli/Hh signaling by blocking Gli1 and Ptched1 gene expression in the presence of a Gli/Hh signaling agonist (SAG). A diet supplemented with S. frutescens suppressed the formation of poorly differentiated carcinoma in prostates of TRAMP mice. Finally, we found Sutherlandioside D was the most potent compound in the crude extract that could suppress Gli-reporter in Shh Light II cells. Together, this suggests that the S. frutescens extract may exert anti-cancer effect by targeting Gli/Hh signaling, and Sutherlandioside D is one of the active compounds.
Although interleukin-24 (IL-24) has been extensively explored in the immunopathologies of autoimmune diseases, neoplasms, and infections, its role in HIV-1 infection has not been thoroughly elucidated to date. Therefore, the objective of this study was to evaluate the gene and protein expressions of IL-24 at the initial moments of HIV infection in PBMCs. Due to the pro-apoptotic role of IL-24, we evaluated the protein expression of caspase-3, as well as Annexin V/ Propidium Iodide flow cytometry and phosphorylation of ERK, which may induce an apoptotic signal block when phosphorylated. The results of this study demonstrated that HIV-1 infection had an impact on the gene and protein expressions of IL-24 and ERK. Annexin V/ Propidium Iodide assay demonstrated decrease in the mechanisms of apoptosis in infected cells after incubation of IL-24 neutralizing antibody Studies on how HIV-1 regulates IL-24 expression may play a role in characterizing viral persistence mechanisms and designing antiretroviral strategies.
It is a well-known fact, that there is a close interconnection between vascular and neural structures in both embryonic development and postnatal life. Different models have been employed to dissect the mechanisms of these interactions, ranging from in vitro systems (e.g., co-culture of neural and endothelial cells) to in vivo imaging of central neural system recovery in laboratory animals after artificially induced trauma. Nevertheless, most of these models have serious limitations. Here, we describe an ex vivo model, representing an organotypic co-culture of aortic fragments (AF) with longitudinal slices of mouse neonatal spinal cord (SC) or dorsal root ganglia (DRG). The samples were co-cultured in a medium adapted for SC tissue and lacking any pro-angiogenic or neurotrophic growth factors. It was found, that cultivation of AFs in the SC injury zone (transversal dissection of a SC slice) resulted in initiation of active aortic sprouting. Remarkably, the endothelial cells exiting the AFs never invaded the SC tissue, concentrating in a nearby area (negative taxis). In contrast, the DRGs, while also promoting sprouting, were a target of active endothelial CD31+ cell invasion (positive taxis). Thus, the tissues of both central and peripheral nervous systems have a prominent positive effect on aortic sprouting, while the vector of endothelial cell expansion is strictly nervous-tissue-type dependent. The ex vivo AF co-culture with SC or DRG appeared to be a useful and promising model for further endeavor into the mechanisms driving the complex interactions between neural and endothelial tissues.
Reactive oxygen species (ROS) are produced by NADPH oxidase (NOX), an enzyme that reduces oxygen by using NADPH as a substrate. Apocynin (APO) is a catechol that is used as a NOX inhibitor, and N-acetyl-cysteine (NAC) can reduce intracellular ROS levels. In this work, the effect of APO and NAC on osteoclast formation were evaluated. APO and NAC significantly decreased the number of tartrate-resistant acid phosphatase (TRAP)-positive cells and the osteoclast area. We analyzed bone-marrow derived monocyte-macrophages (BMMs) that differentiated into osteoclasts after RANKL stimulation. Stimulation was associated with either APO or NAC treatment and osteoclastogenesis marker expression, including NFATc1, MMP-9, and DC-STAMP, was evaluated. APO decreased the intracellular calcium concentration by calcium channels other than ITPR1 and TPC2. On the other hand, APO reduced Tnfrsf11a (RANK) expression and did not alter Fam102a (EEIG1) expression. Therefore, our results demonstrate that APO inhibits osteoclastogenesis by the RANK-RANKL-related signaling pathways, decreases osteoclast markers, and reduces intracellular calcium concentration.
There are numerous studies which provide support for the use of human adipose tissue-derived stem cells (hASCs) to generate hepatocyte-like cells. However, the produced cells exhibit only a certain level of differentiation, mainly due to inefficient induction conditions. Therefore, based on the important role of insulin-like growth factor (IGF-I) in hepatic function and development, in the current study we evaluated the differentiation efficacy of the mentioned factor to induce hASCs into functional hepatocyte-like cells. To investigate this, using a two-step protocol, hASCs were treated with a combination of HGF, Dex, and OSM in the presence or absence of IGF-I up to 21 days. Hepatic differentiation was evaluated by analyzing specific hepatocyte markers at different time points of differentiation induction. Increased expression of hepatocyte-specific genes including ALB, AFP, CK18, and HNF4a, downregulation of bile duct cells marker (CK19), the higher number of ALB positive cells, increased urea production together with higher glycogen deposit was observed upon the treatment of hASCs with the induction medium containing IGF-I compared to the other treatment. In conclusion, our findings suggest IGF-I as a potent inducer of hepatic differentiation of hASCs and its potential to generate more functional hepatocyte-like cells.