Current Molecular Medicine - Volume 19, Issue 6, 2019

Exosomes are 30-120nm long endocytic membrane-derived vesicles, which are secreted by various types of cells and stably present in body fluids, such as plasma, urine, saliva and breast milk. Exosomes participate in intercellular communication. Recently accumulative studies have suggested that exosomes may serve as novel biomarkers for disease diagnosis and prognosis. Herein, we reviewed the biological features of exosomes, technologies for exosome isolation and identification, as well as progress in exosomal biomarker identification, highlighting the relevance of exosome to human diseases and significance and great potential in translational medicine.

Background: Bcl-2/adenovirus E1B-19kDa-interacting protein (BNIP3), an important target of hypoxia-inducible factors-1 alpha (HIF-1α), was reported to be overexpressed under hypoxic condition. Our previous study demonstrated the protective effect on detached retina by BNIP3-mediated autophagy. The study investigated the role of BNIP3-mediated autophagy in retinal pigment epithelial (RPE) cells under hypoxia, and observed the relationship between BNIP3, vascular endothelial growth factor (VEGF) and inflammatory response in hypoxic RPE cells. Methods: BNIP3 knock down in retinal pigment epithelial cells was performed by small interfering RNA (siRNA) technology in ARPE-19 cells, a human RPE cell line. Both control and BNIP3-knockdown ARPE-19 cells were then subjected to a hypoxic challenge using cobalt (II) chloride (CoCl2). The expression of autophagy-related genes, VEGF and inflammatory factors (IL-18, IL-8, MMP-2, MMP-9, NLRP3, TNF-α) in RPE cells was examined using quantitative Polymerase Chain Reaction (qPCR). The protein levels of HIF-1α, BNIP3, the maker proteins (ATG5, LC3,p62, Beclin-1) of autophagy and the component proteins (p-p70S6K, p70S6K, mTOR, p-mTOR) of the mTORC1 pathway were analyzed by Western blot. BNIP3 subcellualr localization was detected by immunofluorescence. Cell viability was measured with Cell Counting kit-8. Cell apoptosis was examined by TUNEL staining and caspase-3 activity assay. Results: The expression levels of BNIP3, HIF-1α and marker genes of autophagy were upregulated in ARPE-19 cells in response to hypoxia. Importantly, hypoxia-induced autophagy was mediated by the mTORC1 pathway, and was blocked upon BNIP3 knockdown. Additionally, hypoxia reduced cell viability, which was relieved by an mTORC1 inhibitor. Also, autophagy protected ARPE-19 cells from CoCl2-induced cell apoptosis. Moreover, inhibition of autophagy upregulated the expression of VEGF and IL-18, and downregulated the expression of other inflammatory factors in the hypoxic ARPE-19 cells. Conclusion: BNIP3-mediated autophagy under hypoxia is involved in regulating inflammatory response and VEGF expression, which consequently affects the cell viability of RPE cells.

Background: Obesity is a chronic condition associated with poorer cognitive functioning. Wisconsin Card Sorting Test (WCST) is a useful tool for evaluating executive functions. In this study, we assessed the association between dopaminergic gene polymorphisms: DAT1 (SLC6A3), COMTVal158Met, DRD4 (48-bp variable number of tandem repeats - VNTR) and WCST parameters to investigate the functions of the frontal lobes in obese individuals. Objective: To find the significant correlations between polymorphisms of DAT1, COMTVal158Met, DRD4 and executive functions in obese subjects. Methods: The analysis of the frequency of individual alleles was performed in 248 obese patients (179 women, 69 men). Evaluation of the prefrontal cortex function (operating memory and executive functions) was measured with the Wisconsin Card Sorting Test (WCST). Separate analyzes were performed in age subgroups to determine different activities and regulation of genes in younger and older participants. Results: Scores of WCST parameters were different in the subgroups of women and men and in the age subgroups. Regarding the COMT gene, patients with A/A and G/A polymorphisms showed significantly better WCST results in WCST_P, WCST_CC and WCST_1st. Regarding DAT1 men with L/L and L/S made less non-perseverative errors, which was statistically significant. In DRD4, significantly better WCST_1st results were found only in older women with S allele. Conclusion: Obtained results indicate the involvement of dopaminergic transmission in the regulation of prefrontal cortex function. Data analysis indicates that prefrontal cortex function may ensue, from different elements such as genetic factors, metabolic aspects of obesity, and hormonal activity (estrogen).

Background: In this study, the antioxidant property of new synthesized azomethins has been investigated as theoretical and experimental. Methods and Results: Density functional theory (DFT) was employed to investigate the Bond Dissociation Enthalpy (BDE), Mulliken Charges, NBO analysis, Ionization Potential (IP), Electron Affinities (EA), HOMO and LUMO energies, Hardness (η), Softness (S), Electronegativity (μ), Electrophilic Index (ω), Electron Donating Power (ω-), Electron Accepting Power (ω+) and Energy Gap (Eg) in order to deduce scavenging action of the two new synthesized azomethines (FD-1 and FD-2). Spin density calculations and NBO analysis were also carried out to understand the antioxidant activity mechanism. Comparison of BDE of FD-1 and FD-2 indicate the weal antioxidant potential of these structures. Conclusion: FD-1 and FD-2 have very high antioxidant potential due to the planarity and formation of intramolecular hydrogen bonds.

Background: Age-related macular degeneration (AMD) is a progressive and irreversible eye disease. The anti-vascular endothelial growth factor (VEGF) therapy has revolutionized the treatment of neovascular AMD. However, the expense for such treatment is quite high. Methods: We used a traditional Chinese medicine ZQMT as an alternative therapeutic regimen for AMD. We employed two in vivo animal models mimicking dry and wet AMD respectively to assess the therapeutic efficacy of ZQMT on treating AMD-related retinopathy. AMD-related retinopathy in Crb1rd8 mice was evaluated from week 1 to 8 by fundus photography. Laser-induced choroidal neovascularization (CNV) was evaluated by fluorescein angiography and histopathology. Results: ZQMT increased CX3CR1 expression in murine CD4+ T cells either cultured in vitro or directly isolated from animals treated with ZQMT. We also performed both in vitro and in vivo studies to confirm that ZQMT has no apparent toxic effects. ZQMT alleviated AMD-related retinopathy in both Crb1rd8 and CNV models. Depletion of CCL2 and CX3CR1 in Crb1rd8 mice abolished the efficacy of ZQMT, suggesting that CCL2 and/or CX3CR1 may underlie the mechanisms of ZQMT in treating AMD-related retinopathy in mice. Conclusion: In summary, our study supports the protective roles of a traditional Chinese medicine ZQMT in AMD.

Background: Papillary thyroid cancer (PTC) is the cardinal histologic type of thyroid cancer, which is the most prevalent kind of endocrine malignancy. The expression of IL-6 is found higher in thyroid carcinoma (THCA) samples than paired normal tissues based on The Cancer Genome Atlas (TCGA) and Genotype-Tissue expression (GTEx) database. In this study, we aimed to investigate the association between interleukin-6 (IL-6) polymorphisms and the PTC risk. Methods: A case-control study was designed using the following data: 241 PTC patients and 463 healthy controls. Five single nucleotide polymorphisms (SNPs) in IL-6 were selected and genotyped using Agena MassARRAY technology. Results: Our results revealed that SNP rs1800796 was associated with an increased PTC risk in co-dominant model (p = 0.042) and dominant model (p = 0.027). Rs1524107 was also a risk factor for PTC susceptibility in co-dominant model (p = 0.003), dominant model (p = 0.002) and log-additive model (p = 0.044). Moreover, rs2066992 significantly increased the PTC risk in co-dominant model and dominant model (p = 0.011, p = 0.009, respectively). Additionally, rs2069837 variant elevated the PTC risk based on dominant model (p = 0.041). In silico analysis, GTEx results for rs1800796, rs1524107 and rs2066992 variants are known to be associated with IL-6 gene expression. Using HaploReg, we found rs1800796, rs1524107 and rs2066992 in LD with functional importance. Conclusion: Our study indicates that IL-6 variants may be a risk factor involved in the pathogenesis and development of PTC.

Objective: To investigate the protective effects of Gentianella turkestanerum extraction by butanol (designated as GBA) on hepatic cell line L02 injury induced by carbon tetrachloride (CCl4) and hydrogen peroxide (H2O2). Methods: L02 cells were incubated with 5 μg/mL, 10 μg/mL, 20 μg/mL, 40 μg/mL, 60 μg/mL, 80 μg/mL and 100 μg/mL GBA for 24 hours, and then MTT assay was used to screen the cytotoxicity for GBA. Cells were divided into blank control group, CCl4/H2O2 model group, treated by CCl4 (20 mmol/L) or H2O2 (100 μmol/L); silymarin+CCl4/H2O2 group, treated by CCl4 (20 mmol/L) or H2O2 (100 μmol/L) and 5 μg/mL silymarin; GBA+CCl4/H2O2 group, treated by CCl4 (20 mmol/L) or H2O2 (100 μmol/L) and GBA (5 μg/mL, 10 μg/mL and 20 μg/mL). MTT assay was performed to determine the cellular activity. Malondialdehyde (MDA) content was determined using a commercial kit. The alanine transaminase (ALT), aspartate transaminase (AST) in the supernatant was determined. PE-Annexin V/7-ADD method was utilized to determine the apoptosis of cells. RT-PCR was used to evaluate the expression of endoplasmic reticulum stressrelated genes (CHOP, PERK, IRE1 and ATF6) mRNA. Western blot analysis was performed to determine the expression of CHOP, Caspase 12 and NF-ΚB protein. Results: Cellular survival after GBA (5 μg/mL, 10 μg/mL and 20 μg/mL) incubation was ≥ 75%. After GBA incubation, levels of ALT and AST showed a significant decrease (P < 0.05), while that of the MDA showed a significant decrease (P < 0.05). The apoptosis in the CCl4 or H2O2 group showed a significant increase compared to the control group (P < 0.05). In contrast, GBA-preincubation could attenuate the cellular apoptosis compared to the CCl4 or H2O2 group, which displayed a dose-dependent manner (P < 0.05). The expression of CHOP, PERK, IRE1 and ATF6 mRNA was significantly up-regulated in the presence of CCl4 or H2O2 (P < 0.05). Whereas, GBA induced a significant decrease in these mRNA thereafter (P < 0.05), together with a decrease in CHOP and Caspase 12 proteins (P < 0.05). Besides, it could attenuate the expression of NF-ΚB p65 in nuclear protein. Conclusion: G. turkestanerum could inhibit the lipid peroxidation and increase the antioxidant activity. Also, it could inhibit the cellular apoptosis through down-regulating the transcriptional level of ERS related genes and proteins. This process was associated with the nuclear translocation of NF-ΚB p65 protein.