Sargassum thunbergii is a brown alga from which various bioactive compounds can be extracted. Among these, the activities of indole derivatives, particularly as potential inhibitors of matrix metalloproteinases (MMPs), and their underlying mechanisms have been rarely investigated. Therefore, we evaluated the inhibitory effects of indole-6-car-boxaldehyde (I6CA) on MMP-9 by gelatin zymography and western blot anlaysis. We used phorbol 12-myristate 13-acetate (PMA), which is known to induce MMP-9 expression and secretion, to stimulate HT1080 cells. Our results revealed that I6CA significantly inhibited MMP-9 expression and secretion, without significantly affecting the viability of PMA-stimulated HT1080 cells. Our mechanistic studies indicated that I6CA suppressed the phosphorylation and activation of two mitogen-activated protein kinases (MAPKs), c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase 1/2 (ERK). Furthermore, I6CA inhibited the phosphorylation of inhibitor of kappa B alpha (I kappa B alpha) in response to PMA stimulation, which suppressed nuclear factor-kappa B (NF-kappa B) p65 subunit nuclear translocation. Collectively, I6CA was determined to suppress MMP-9 expression and secretion, and effects were proposed to be mediated via the inhibition of the MAPK and NF-kappa B p65 pathways. Therefore, we suggested I6CA to be a potential therapeutic agent for MMP-9-related processes, including tumor invasion and metastasis; however, further investigation is required.
Spindle and kinetochore-associated protein 1 (SKA1), a microtubule-binding subcomplex of the outer kinetochore, is essential for complete chromosomal separation. SKA1 has been suggested as a potential biomarker for various types of cancer. However, the exact role of SKA1 in esophageal squamous cell carcinoma (ESCC) remains unclear. The present study investigated whether SKA1 affects the biological behavior of ESCC. The expression of SKA1 in ESCC tissues was measured using immunohistochemistry and reverse transcription-quantitative polymerase chain reaction. In addition, a SKA1-silencing lentivirus was constructed, which was transfected into TE-1 cells to establish stable SKA1-knockdown TE-1 cells. Proliferation was analyzed using a Celigo image cytometer and a MTS assay. Cell cycle progression and apoptosis were analyzed by flow cytometry, while cell migration was assessed using a Transwell assay. SKA1 was significantly overexpressed in ESCC tissues, and SKA1 overexpression was significantly associated with differentiation, pathological N stage and pathological tumor-node-metastasis stage. SKA1 was determined to be an independent prognostic factor for ESCC. Furthermore, SKA1 was significantly overexpressed in ESCC cells, and SKA1-silencing inhibited cell proliferation and migration, arrested the cell cycle and promoted cell apoptosis. In summary, SKA1 may serve as a potential therapeutic target and prognostic biomarker for ESCC.
Rheumatoid arthritis (RA) is a debilitating joint disease characterized by chronic inflammation, pathologic alteration of fibroblast-like synoviocytes (FLS), destruction of cartilage and bone, and the formation of an invasive pannus. RA-FLS exhibit increased proliferation and resistance to apoptosis. The retinoid X receptor (RXR) has a role in regulating cell cycle, differentiation and apoptosis, and agonism of RXR has been investigated as a treatment strategy in several types of cancer. However, there is little research on the effects of RXR agonism in other diseases. Bexarotene is a novel selective RXR ligand used in the treatment of T-cell lymphoma. In the present study, bexarotene was used to investigate the involvement of RXR in tumor necrosis factor-alpha (TNF-alpha)-induced RA conditions in human FLS. To the best of our knowledge, this is the first time that RXR has been demonstrated to be expressed in FLS and to be downregulated in response to TNF-alpha stimulation. The present study also demonstrated that bexarotene exerted an anti-inflammatory effect by downregulating expression of interleukin (IL)-6, IL-8, monocyte chemoattractant protein-1, and high mobility group box-1. Notably, bexarotene also rescued the TNF-alpha -induced downregulation of the anti-inflammatory cytokines IL-4 and transforming growth factor-beta 1. Bexarotene treatment exhibited a potential protective effect against cartilage degradation by downregulating the expression of matrix metalloproteinase (MMP)-1, MMP-3 and MMP-13. In addition, the present results demonstrated that the effects of bexarotene were mediated through the p38 mitogen-activated protein kinase/nuclear factor-kappa B pathway, via inhibition of p38 protein and the inhibitor alpha of kappa B phosphorylation. Taken together, the present findings demonstrated the potential of RXR agonism using bexarotene as a treatment against the development and progression of RA.
Bladder cancer (BCa) is a common urinary tract malignancy with frequent recurrences after initial resection. Submucosal injection of gemcitabine prior to transurethral resection of bladder tumor (TURBT) may prevent recurrence of urothelial cancer. However, the underlying mechanism remains unknown. In the present study, ultra-performance liquid chromatography Q-Exactive mass spectrometry was used to profile tissue metabolites from 12 BCa patients. The 48 samples included pre- and post-gemcitabine treatment BCa tissues, as well as adjacent normal tissues. Principal component analysis (PCA) revealed that the metabolic profiles of pre-gemcitabine BCa tissues differed significantly from those of pre-gemcitabine normal tissues. A total of 34 significantly altered metabolites were further analyzed. Pathway analysis using MetaboAnalyst identified three metabolic pathways closely associated with BCa, including glutathione, purine and thiamine metabolism, while gluta-thione metabolism was also identified by the enrichment analysis using MetaboAnalyst. In search of the possible targets of gemcitabine, metabolite profiles were compared between the pre-gemcitabine normal and post-gemcitabine BCa tissues. Among the 34 metabolites associated with BCa, the levels of bilirubin and retinal recovered in BCa tissues treated with gemcitabine. When comparing normal bladder tissues with and without gemcitabine treatment, among the 34 metabolites associated with BCa, it was observed that histamine change may be associated with the prevention of relapse, whereas thiamine change may be involved in possible side effects. Therefore, by employing a hypothesis-free tissue-based metabolomics study, the present study investigated the metabolic signatures of BCa and found that bilirubin and retinal may be involved in the mechanism underlying the biomolecular action of submucosal injection of gemcitabine in urothelial BCa.
S-allyl-cysteine sulfoxide (alliin) is the main organosulfur component of garlic and its preparations. The present study aimed to examine the protective effect of alliin on cardiac function and the underlying mechanism in a mouse model of myocardial infarction (MI). Notably, alliin treatment preserved heart function, attenuated the area of infarction in the myocardium of mice and reduced lesions in the myocardium, including cardiomyocyte fibrosis and death. Further mechanistic experiments revealed that alliin inhibited necroptosis but promoted autophagy in vitro and in vivo. Cell viability assays showed that alliin dose-dependently reduced the necroptotic index and inhibited the expression of necroptosis-related receptor-interacting protein 1, receptor-interacting protein 3 and tumor necrosis factor receptor-associated factor 2, whereas the levels of Beclin 1 and microtubule-associated protein 1 light chain 3, which are associated with autophagy, exhibited an opposite trend upon treatment with alliin. In addition, the level of peroxisome proliferator-activated receptor gamma was increased by alliin. Collectively, these findings demonstrate that alliin has the potential to protect cardiomyocytes from necroptosis following MI and that this protective effect occurs via the enhancement of autophagy.
Isosteviol sodium (STVNa), which is a derivate of the natural sweet-tasting glycoside stevioside, has recently been developed and it has been determined that this compound exhibits neuro- and cardio-protective properties. In the current study, whether STVNa interferes with the development of cardiac hypertrophy, which is induced by isoprenaline (Iso), was investigated in an experimental rat model. Rats were treated with a vehicle (0.9% NaCl; control), isoprenaline (Iso; 5 mg/kg) or Iso (5 mg/kg) with STVNa (4 mg/kg; Iso + STVNa). Cardiomyocytes were isolated using enzymatic dissociation and were treated with 5 mu M Iso for 24 h and co-treated with 5 mu M STVNa. Brain natriuretic peptide (BNP) mRNA expression was determined using PCR analysis. Cell surface area, intracellular reactive oxygen species (ROS), mitochondrial transmembrane potential (Delta Psi m), cytoplasmic Ca2+ and Ca(2+ )and contractile function were examined using a laser scanning confocal microscope. The current study demonstrated that STVNa inhibited Iso-induced cardiac hypertrophy by inhibiting cardiomyocyte size. STVNa significantly reduced cell surface area and decreased BNP mRNA expression in ventricular cardiomyocyte Iso-induced hypertrophy. STVNa was also revealed to restore Delta Psi m and reduce ROS generation and intracellular Ca(2+ )concentration when compared with the Iso-treated group. Additionally, STVNa preserved Ca(2+ )transients in hypertrophic cardiomyocytes. In conclusion, the present study demonstrated that STVNa protects against Iso-induced myocardial hypertrophy by reducing oxidative stress, restoring Delta Psi m and maintaining Ca2+ homeostasis.
The senescence of vascular endothelial cells (ECs) plays a critical role in aging-related cardiovascular diseases. We previously reported the causal relation of Jagged1 in ECs and the thickening of the arterial wall in aging mice. The aim of the present study was to further investigate the correlation between insulin-like growth factor-binding protein 1 (IGFBP1), one of the secretory proteins regulated by Jagged1, and the severity of coronary atherosclerosis and patient age, as well as its effect on EC senescence. First, microarray analysis was performed to screen the differentially expressed genes regulated by Jagged1 in human coronary arterial ECs (HCAECs). Inhibition of the Jagged1 expression using a small interfering RNA knockdown method in HCAECs led to the upregulation of 17 and the downregulation of 78 genes by >3-fold, and IGFBP1 was confirmed to be a secretory protein expressed by HCAECs and regulated by Jagged1. Subsequently, in 112 consecutively enrolled patients with acute chest pain who underwent coronary angiography, the circulating level of IGFBP1 was found to be positively correlated with age (r=0.512, P<0.001) and Synergy between PCI with TAXUS and Cardiac Surgery (SYNTAX) score (r=0.409, P<0.001). Among age-comparable patients, the circulating IGFBP1 level was found to be increased in patients with higher SYNTAX scores. In cultured HCAECs, IGFBP1 was shown to protect ECs against passage- or H2O2-induced senescence, and these protective effects of IGFBP1 may be partially reversed by LY294002, a known Akt signaling inhibitor. Therefore, the results of the present study suggested that, as a downstream protein of Jagged1, IGFBP1 was correlated with the severity of coronary atherosclerosis in aging patients, and the increase of circulating IGFBP1 levels with aging may be an adaptive response to counter HCAEC senescence through Akt signaling.
Diallyl disulfide (DADS) is one of the primary components of garlic and it exhibits a broad range of biological activities. In the present study, the effects of DADS on lipid metabolism and its potential role in the modulation of the gut microbiome were determined. Hematoxylin and eosin and oil-red O staining were used to assess the liver and intestinal tissues of mice treated with DADS. The expression of lipid metabolism-associated genes was measured using reverse transcription-quantitative PCR (RT-qPCR). The effects of DADS on the gut microbiome were measured using 16S recombinant (r)DNA gene analysis. The results revealed that the serum non-esterified free fatty acids, high density lipopro-tein-cholesterol, low density lipoprotein-cholesterol, serum total cholesterol, liver triglyceride and total cholesterol levels of the mice fed with a low-dose of DADS was significantly higher when compared with the control. Hematoxylin and eosin and oil-red O staining demonstrated that DADS induced fatty liver in mice. The results of the RT-qPCR revealed that the expression levels of a number of lipid metabolism-associated genes were altered in the livers of mice treated with DADS. The 16S rDNA gene analysis demonstrated that the mice fed on a normal diet treated with a low-dose of DADS had decreased levels of bacteria from the Bacteroidetes phyla and increased levels of bacteria from the Firmicutes phyla. The Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed the top 20 pathways enriched in the low-dose DADS group of mice fed with a normal diet. In the present study, low-dose DADS induced fatty liver and altered the gut micro-biota, similar to the phenotype induced by a high fat diet, by regulating the expression of lipid metabolism associated genes.
Patients with ischemic hearts who have refused coronary vascular reconstruction may exhibit dynamic myocardial remodeling and cardiac dysfunction. In the present study, the role of miRNA-1 and its association with the ubiquitin-proteasome system (UPS) in regulating myocardial remodeling was investigated. A myocardial infarction (MI) model was constructed and the hearts were treated with miRNA-1 antagomir, miRNA-1 lentiviral vectors and the UPS proteasome blocker bortezomib. The expression levels of miRNA-1 were evaluated using reverse transcription PCR and the abundance of the ubiquitin-proteasome protein and caspase-3 were evaluated via western blot analysis. Furthermore, the collagen volume fraction was calculated using Masson's trichrome staining, and the apoptosis index was detected via terminal deoxynucleotidyl transferase dUTP-biotin nick end labeling staining. Transforming growth factor (TGF)-beta expression was assessed via immunohistochemical staining. Echocardiographic characteristics and myocardial infarct size were analyzed. miRNA-1 expression levels were found to be increased following MI. miRNA-1 antagomir administration clearly inhibited miRNA-1 expression, whereas the miRNA-1 lentiviral vector exerted the opposite effect. The levels of 19s proteasome, 20S proteasome and ubiquitin ligase E3 were decreased in the miRNA-1 antagomir group, but were significantly increased in the miRNA-1 lentiviral group; however, only 20S proteasome expression was decreased in the bortezomib group. Collagen hyperplasia and TGF-beta expression were decreased in both the miRNA-1 antagomir and bortezomib groups, although the effects of the miRNA-1 antagomir were more noticeable. The miRNA-1 antagomir and the UPS proteasome blocker both alleviated the ultrastructural impairments, demonstrated by a decreased left ventricular (LV) end-diastolic diameter and LV mass, but the miRNA-1 antagomir was also able to increase LV ejection fraction and LV fractional shortening. miRNA-1 regulated UPS-associated mRNA expression and affected the majority of the UPS components in the myocardium, thereby leading to increased myocardial cell apoptosis, myocardial fibrosis and remodeling. The miRNA-1 antagomir exerted a more prominent cardioprotective effect compared with the UPS proteasome blocker bortezomib.
Previously, a ubiquinol-cytochrome c reductase binding protein (UQCRB) homolog was identified in the house dust mite (HDM) species Dermatophagoides farinae (Der f) as a major allergen. In the present study, the immunodominant immunoglobulin E (IgE) epitope of the protein Der f 24 was investigated. Analysis of the homologous amino acid (aa) sequences in Der f and human UQCRB was performed. Four different recombinant Der f 24 and hybrid proteins formed by integrating Der f and human UQCRB sequences were expressed in Escherichia coli, purified using Ni-NTA resins, and IgE-binding activity was determined using IgE-western blotting and enzyme-linked immunosorbent assay (ELISA) experiments. IgE epitopes were further identified by IgE-dot blotting and IgE-ELISA with synthetic polypeptides and HDM-allergic sera. Three-dimensional (3D) structural modeling was used to analyze the position of the immuno-dominant IgE epitope. The amino acid sequence homology between Der f 24 and the human UQCRB protein was determined to be 39.34%. IgE-ELISA and western blot analysis showed that all of the Der f-human UQCRB hybrid proteins generated, except for the one lacking 59 residues of the N-terminal region of Der f 24, were bound by allergic serum IgE. A synthetic polypeptide consisting of 32 residues of the N-terminal reacted with IgEs from HDM-allergic sera and could be used to generate high titer specific IgG or specific IgE antibodies in immunized mice. The 32-aa N-terminal region of Der f 24 was localized to a structural protrusion, which may facilitate specific IgE-binding. These results indicate that the immunodominant IgE epitope of Der f 24 is located mainly in a 32-residue region of the N-terminus. These findings may inform the mechanisms of HDM allergy sensitization and allergy immunotherapy development.
There is now substantial evidence that myocardial ischemia-reperfusion (IR) injury affects the spinal cord and brain, and that interactions may exist between these two systems. In the present study, the spinal cord proteomes were systematically analyzed after myocardial IR injury, in an attempt to identify the proteins involved in the processes. The myocardial IR injury rat model was first established by cross clamping the left anterior descending coronary artery for 30-min ischemia, followed by reperfusion for 2 h, which resulted in a significant histopathological and functional myocardial injury. Then using the stable isotope dimethyl labeling quantitative proteomics strategy, a total of 2,362 shared proteins with a good distribution and correlation were successfully quantified. Among these proteins, 33 were identified which were upregulated and 57 were downregulated in the spinal cord after myocardial IR injury, which were involved in various biological processes, molecular function and cellular components. Based on these proteins, the spinal cord protein interaction network regulated by IR injury, including apop-tosis, microtubule dynamics, stress-activated signaling and cellular metabolism was established. These heart-spinal cord interactions help explain the apparent randomness of cardiac events and provide new insights into future novel therapies to prevent myocardial I/R injury.
Postoperative cognitive dysfunction (POCD) is a common and well-known complication following surgery, particularly cardiopulmonary bypass (CPB) surgery. There are currently no suitable treatments for POCD, which is associated with increased illness and mortality rates. The present study aimed to identify a novel treatment for POCD. The protective effect of kappa opioid receptor (KOR) agonists on POCD in rats following CPB was determined and the regulatory mechanism of the Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway was examined. The rats were randomly divided into five groups: Sham operation (Sham group), CPB operation (CPB group), KOR agonist + CPB (K group), KOR agonist + norbinaltorphimine (nor-BNI) + CPB (NK group), and KOR agonist + JAK2-STAT3 specific pathway inhibitor + CPB (AG group). A water maze test and neurological function scores were used to evaluate POCD. Hematoxylin and eosin staining was used to observe hippocampal neurons. ELISA was used to detect the levels of inflammatory factors, oxidative stress factors and brain injury markers. Immunofluorescence was used to visualize the neurons. TUNEL staining and western blotting were used to detect neuronal apoptosis, and western blotting was also used to detect JAK2/STAT3 pathway-related proteins. The KOR agonists significantly improved POCD. S-100 beta and NSE detection revealed that KOR agonists alleviated brain damage in CPB rats, and this result was reversed by KOR antagonists. The KOR agonists led to a significantly reduced inflammatory response and oxidative stress, as determined by ELISA detection, and attenuated hippocampal neuronal apoptosis, as revealed by TUNEL staining and western blotting, compared with the results in the CPB group. Finally, the KOR agonists inhibited the expression levels of phosphorylated (p-)JAK2 and p-STAT3, rather than total JAK2 and STAT3, compared with levels in the CPB group. Taken together, KOR agonists improved POCD in rats with CPB by inhibiting the JAK2/STAT3 signaling pathway.
Asthma is a common chronic airway inflammatory disease, characterized by airway inflammation and remodeling. Piperlongumine (PL) has a number of physiological and pharmacological properties. However, the anti-asthmatic effect of PL has not been reported to date. In the present study, ovalbumin (OVA) was used to sensitize and challenge mice to induce asthma. The results revealed that PL pretreatment reduced OVA-induced airway inflammatory cell infiltration, reduced Th2 cytokine expression, both in the bronchoalveolar lavage fluid and in lung tissues, reduced the serum IgE level, pro-inflammatory cytokine [tumor necrosis factor (TNF)-alpha and interleukin (IL)-6] and intercellular adhesion molecule expression, as well as nuclear factor (NF)-kappa B activation. In addition, PL also mitigated OVA-induced goblet cell metaplasia, inhibited mucus protein secretion, mitigated airway fibrosis and downregulated fibrosis marker expression. It was also demonstrated that PL inhibited TNF-alpha induced inflammatory cytokine expression and NF-kappa B activation in vitro. Taken together, the findings of the present study indicated that PL can reduce OVA-induced airway inflammation and remodeling in asthmatic mice, and that these effects may be mediated by inhibiting NF-kappa B signaling.
Long non-coding RNAs (lncRNAs) have been shown to contribute to progression and prognosis of hepatocellular carcinoma (HCC). However, expression profiling and interaction of lncRNAs with messenger RNAs (mRNAs) and microRNAs (miRNAs) remain largely unknown in HCC. The expression profiling of lncRNAs, mRNA and miRNAs was obtained using microarray. The Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes pathway analysis were used to characterize potential functions of differentially expressed mRNAs. Cytoscape was applied to construct an lncRNA-miRNA-mRNA co-expression network and candidate lncRNAs were validated via quantitative PCR in 30 pairs of HCC and adjacent tumor-free tissues. In this study, 1,056 upregulated and 1,288 downregulated lncRNAs were identified, while 2,687 mRNAs and 6 miRNAs were aberrantly expressed in HCC compared with adjacent tumor-free tissues. Potential functions of differentially expressed mRNAs were demonstrated to significantly participate in modulating critical genes in the cell cycle, such as cyclin E1 and cyclin B2. After screening, 95 lncRNAs, 5 miRNAs and 36 mRNAs were recruited for construction of lncRNA-mRNA-miRNA co-expression network in the cell cycle pathway. Subsequently, the top 5 lncRNAs that potentially modulate critical genes in the cell cycle were selected as the candidates for further verification. Kaplan-Meier curves using the Cancer Genome Atlas database showed that 13 targeted mRNAs were associated with overall survival of HCC patients. Finally, three lncRNAs, including ENST00000522221, lnc-HACE1-6:1 and lnc-ICOSLG-11:1, are significantly upregulated in HCC tissues compared with adjacent tumor-free tissues. These findings suggest that lncRNAs play essential roles in the pathogenesis of HCC via regulating coding genes and miRNAs, and may be important targets for diagnosis and treatment of this disease.