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1.
Article in English | WPRIM | ID: wpr-786676

ABSTRACT

BACKGROUND: Low temperature plasma (LTP) was recently shown to be potentially useful for biomedical applications such as bleeding cessation, cancer treatment, and wound healing, among others. Keratinocytes are a major cell type that migrates directionally into the wound bed, and their proliferation leads to complete wound closure during the cutaneous repair/regeneration process. However, the beneficial effects of LTP on human keratinocytes have not been well studied. Therefore, we investigated migration, growth factor production, and cytokine secretion in primary human keratinocytes after LTP treatment.METHODS: Primary cultured keratinocytes were obtained from human skin biopsies. Cell viability was measured with the EZ-Cytox cell viability assay, cell migration was evaluated by an in vitro wound healing assay, gene expression was analyzed by quantitative real-time polymerase chain reaction, and protein expression was measured by enzyme-linked immunosorbent assays and western blotting after LTP treatment.RESULTS: Cell migration, the secretion of several cytokines, and gene and protein levels of angiogenic growth factors increased in LTP-treated human keratinocytes without associated cell toxicity. LTP treatment also significantly induced the expression of hypoxia inducible factor-1α (HIF-1α), an upstream regulator of angiogenesis. Further, the inhibition of HIF-1α expression blocked the production of angiogenic growth factors induced by LTP in human keratinocytes.CONCLUSION: Our results suggest that LTP treatment is an effective approach to modulate wound healing-related molecules in epidermal keratinocytes and might promote angiogenesis, leading to improved wound healing.


Subject(s)
Hypoxia , Biopsy , Blotting, Western , Cell Migration Assays , Cell Movement , Cell Survival , Cytokines , Enzyme-Linked Immunosorbent Assay , Gene Expression , Hemorrhage , Humans , In Vitro Techniques , Intercellular Signaling Peptides and Proteins , Keratinocytes , Plasma , Real-Time Polymerase Chain Reaction , Skin , Wound Healing , Wounds and Injuries
2.
Braz. j. med. biol. res ; 52(4): e7728, 2019. graf
Article in English | LILACS | ID: biblio-1001506

ABSTRACT

Pituitary adenoma is one of the most common tumors in the neuroendocrine system. This study investigated the effects of long non-coding RNAs (lncRNAs) highly up-regulated in liver cancer (HULC) on rat secreting pituitary adenoma GH3 cell viability, migration, invasion, apoptosis, and hormone secretion, as well as the underlying potential mechanisms. Cell transfection and qRT-PCR were used to change and measure the expression levels of HULC, miR-130b, and FOXM1. Cell viability, migration, invasion, and apoptosis were assessed using trypan blue staining assay, MTT assay, two-chamber transwell assay, Guava Nexin assay, and western blotting. The concentrations of prolactin (PRL) and growth hormone (GH) in culture supernatant of GH3 cells were assessed using ELISA. The targeting relationship between miR-130b and FOXM1 was verified using dual luciferase activity. Finally, the expression levels of key factors involved in PI3K/AKT/mTOR and JAK1/STAT3 pathways were evaluated using western blotting. We found that HULC was highly expressed in GH3 cells. Overexpression of HULC promoted GH3 cell viability, migration, invasion, PRL and GH secretion, as well as activated PI3K/AKT/mTOR and JAK1/STAT3 pathways. Knockdown of HULC had opposite effects and induced cell apoptosis. HULC negatively regulated the expression of miR-130b, and miR-130b participated in the effects of HULC on GH3 cells. FOXM1 was a target gene of miR-130b, which was involved in the regulation of GH3 cell viability, migration, invasion, and apoptosis, as well as PI3K/AKT/mTOR and JAK1/STAT3 pathways. In conclusion, HULC tumor-promoting roles in secreting pituitary adenoma might be via down-regulating miR-130b, up-regulating FOXM1, and activating PI3K/AKT/mTOR and JAK1/STAT3 pathways.


Subject(s)
Humans , Animals , Rats , Pituitary Neoplasms/pathology , Adenoma/pathology , RNA, Long Noncoding/physiology , Enzyme-Linked Immunosorbent Assay , Transfection , Adenoma/genetics , Adenoma/metabolism , Cell Movement/physiology , Cell Survival/physiology , Blotting, Western , Apoptosis/physiology , MicroRNAs/analysis , Cell Line, Tumor , STAT3 Transcription Factor/analysis , Janus Kinase 1/analysis , Janus Kinase 1/metabolism , Cell Migration Assays , Forkhead Box Protein M1/analysis , Forkhead Box Protein M1/metabolism , Luciferases
3.
Braz. oral res. (Online) ; 33: e059, 2019. graf
Article in English | LILACS | ID: biblio-1039303

ABSTRACT

Abstract We recently demonstrated that a co-culture system of human umbilical vein endothelial cells (HUVECs) and human dental pulp stem cells (hDPSCs) could enhance angiogenesis ability in vitro. However, whether tumor necrosis factor α (TNF-α) could promote blood vessel formation during pulp regeneration remained unknown. The aim of this study was to investigate the effects of TNF-α on the formation of endothelial tubules and vascular networks in a co-culture system of hDPSCs and HUVECs. hDPSCs were co-cultured with HUVECs at a ratio of 1:5. The Matrigel assay was performed to detect the total tubule branching lengths and numbers of branches, and the Cell-Counting Kit 8 assay was performed to examine the effect of TNF-α on cell proliferation. Real-time polymerase chain reactions and western blot were used to detect vascular endothelial growth factor (VEGF) mRNA and protein expression. The Matrigel assay showed significantly greater total branching lengths and numbers of branches formed in the experimental groups treated with different concentrations of TNF-α compared with the control group. The decomposition times of the tubule structures were also significantly prolonged (P < 0.05). Treatment with 50 ng/ml TNF-α did not significantly change the proliferation of co-cultured cells, but it significantly increased the VEGF mRNA and protein expression levels (p < 0.05). In addition, the migration abilities of HUVECs and hDPSCs increased after co-culture with TNF-α (p < 0.05). TNF-α enhanced angiogenic ability in vitro in the co-culture system of hDPSCs and HUVECs.


Subject(s)
Humans , Adolescent , Adult , Young Adult , Tumor Necrosis Factor-alpha/pharmacology , Neovascularization, Physiologic/drug effects , Dental Pulp/cytology , Dental Pulp/drug effects , Angiogenesis Inducing Agents/pharmacology , Human Umbilical Vein Endothelial Cells/drug effects , Proteoglycans , Reference Values , Time Factors , Cell Count , Cells, Cultured , Blotting, Western , Reproducibility of Results , Collagen , Laminin , Neovascularization, Physiologic/physiology , Dental Pulp/physiology , Vascular Endothelial Growth Factor A/analysis , Vascular Endothelial Growth Factor A/drug effects , Cell Proliferation/drug effects , Cell Proliferation/physiology , Drug Combinations , Cell Migration Assays , Human Umbilical Vein Endothelial Cells/physiology , Real-Time Polymerase Chain Reaction
4.
Braz. j. med. biol. res ; 51(2): e6611, 2018. graf
Article in English | LILACS | ID: biblio-889029

ABSTRACT

Ginsenoside Rg1, one of the most notable active components of Panax ginseng, has been widely reported to exert anti-inflammatory actions. This study aimed to reveal whether ginsenoside Rg1 also exhibits beneficial roles against lipopolysaccharide (LPS)-induced apoptosis and inflammation in human renal tubular epithelial cells, and to evaluate the potential role of the component on tubulointerstitial nephritis treatment. HK-2 cells were treated with various doses of ginsenoside Rg1 (0, 50, 100, 150, and 200 μM) in the absence or presence of 5 μg/mL LPS. Thereafter, CCK-8 assay, flow cytometry, western blot, migration assay, reactive oxygen species (ROS) assay, and ELISA were carried out to respectively assess cell viability, apoptosis, migration, ROS activity, and the release of inflammatory cytokines. As a result, ginsenoside Rg1 protected HK-2 cells from LPS-induced injury, as cell viability was increased, cell apoptosis was decreased, and the release of MCP-1, IL-1β, IL-6, and TNF-α was reduced. Ginsenoside Rg1 functioned to HK-2 cells in a dose-dependent manner, and the 150 μM dose exhibited the most protective functions. Ginsenoside Rg1 had no significant impact on cell migration and ROS activity, while it alleviated LPS-induced ROS release and migration impairment. Furthermore, the down-regulations of p-PI3K, p-AKT, and up-regulations of PTEN, p-IκBα, p-p65, Bcl-3 induced by LPS were recovered to some extent after ginsenoside Rg1 treatment. In conclusion, ginsenoside Rg1 protects HK-2 cells against LPS-induced inflammation and apoptosis via activation of the PI3K/AKT pathway and suppression of NF-κB pathway.


Subject(s)
Humans , Lipopolysaccharides , Apoptosis/drug effects , Ginsenosides/pharmacology , Epithelial Cells/drug effects , Kidney Tubules/cytology , Anti-Inflammatory Agents/pharmacology , Enzyme-Linked Immunosorbent Assay , Cell Line , Cell Survival/drug effects , Blotting, Western , Reproducibility of Results , Analysis of Variance , Cytokines/analysis , Cytokines/drug effects , Cell Migration Assays
5.
Braz. j. med. biol. res ; 51(2): e6793, 2018. graf
Article in English | LILACS | ID: biblio-889023

ABSTRACT

Colon cancer is one of the most common digestive tumors. The present study aimed to explore the functional role, as well as the underlying mechanism of long non-coding RNA LINC00261 in colon cancer. Expression of LINC00261 was analyzed in colon cancer cell lines and human normal cell lines. Acquired resistance cell lines were then built and the acquired resistance efficiency was detected by evaluating cell viability. Thereafter, the effects of LINC00261 overexpression on cisplatin-resistant colon cancer cells were measured, as well as cell apoptosis, viability, migration, and invasion. Subsequently, we investigated the interaction of LINC00261 and β-catenin. The results showed that the LINC00261 gene was down-regulated in colon cancer cell lines and tissues, and in cisplatin-resistant cells. LINC00261 overexpression might relieve cisplatin resistance of colon cancer cells via promoting cell apoptosis, and inhibiting cell viability, migration, and invasion. Moreover, LINC00261 might down-regulate nuclear β-catenin through restraining β-catenin from cytoplasm into nuclei or it could also promote β-catenin degradation and inhibit activation of Wnt pathway. Finally, LINC00261 reduced cisplatin resistance of colon cancer in vivo and enhanced the anti-colon cancer effect of cisplatin through reducing tumor volume and weight.


Subject(s)
Humans , RNA, Long Noncoding/physiology , Antineoplastic Agents/pharmacology , Tetrazolium Salts , Thiazoles , Down-Regulation , Blotting, Western , Reproducibility of Results , Analysis of Variance , Apoptosis/drug effects , Apoptosis/physiology , Apoptosis Regulatory Proteins/drug effects , beta Catenin/physiology , Cell Migration Assays
6.
Braz. oral res. (Online) ; 32: e48, 2018. tab, graf
Article in English | LILACS | ID: biblio-952159

ABSTRACT

Abstract The aim was to investigate the angiogenic effects of concentrated growth factors on human dental pulp cells and human umbilical vein endothelial cells. Cells were treated with concentrated growth factor extracts. The CCK-8 assay and cell cycle assay were conducted to evaluate cell growth. Cell migration was evaluated by the Transwell migration assay. Angiogenesis-associated mRNA and protein expression levels were determined using quantitative real-time PCR and Western blotting, respectively. A tube formation assay was conducted to evaluate the angiogenic capacity in vitro. The data showed that compared with the control, concentrated growth factor extracts significantly promoted dental pulp cell proliferation and differentiation and endothelial cell proliferation and migration in a dose-dependent manner (p < 0.05). Concentrated growth factor extracts also promoted the tube-like structure formation of endothelial cells in vitro. The RT-PCR and Western blot results showed that concentrated growth factor extracts upregulated the expression of angiogenesis-related genes - chemokine receptor-4, platelet-derived growth factor, and vascular endothelial growth factor - in dental pulp cells. In conclusion, concentrated growth factors showed proangiogenic effects on dental pulp cells and endothelial cells and have good application potential for dental pulp revascularization.


Subject(s)
Humans , Male , Adult , Neovascularization, Physiologic/physiology , Intercellular Signaling Peptides and Proteins/physiology , Dental Pulp/cytology , Human Umbilical Vein Endothelial Cells/physiology , Reference Values , Time Factors , Platelet-Derived Growth Factor/analysis , Platelet-Derived Growth Factor/physiology , Cell Cycle/physiology , Cells, Cultured , Blotting, Western , Reproducibility of Results , Analysis of Variance , Receptors, CXCR4/analysis , Receptors, CXCR4/physiology , Intercellular Signaling Peptides and Proteins/analysis , Vascular Endothelial Growth Factor A/analysis , Vascular Endothelial Growth Factor A/physiology , Cell Proliferation/physiology , Cell Migration Assays , Real-Time Polymerase Chain Reaction
7.
Article in Chinese | WPRIM | ID: wpr-303905

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effects of polyunsaturated fatty acids (PUFA) ω-3 and ω-6, and their middle metabolites PGE2 and PGE3 on angiogenesis formation of gastric cancer, and to explore associated mechanism.</p><p><b>METHODS</b>The effects of ω-3, ω-6, PGE2, PGE3 on the proliferation and migration of human umbilical vein endothelial cell (HUVEC) were measured by proliferation and migration assay respectively. The angiogenesis assay in vivo was used to measure the effects of ω-3, ω-6, PGE2 and PGE3 on neovascularization. In all the assays, groups without ω-3, ω-6, PGE2 and PGE3 were designed as the control.</p><p><b>RESULTS</b>With the increased concentration of ω-6 from 1 μmol/L to 10 μmol/L, the proliferation ability of HUVECs enhanced, and the number of migration cells also increased from 28.2±3.0 to 32.8±2.1, which was higher than control group (21.2±3.2) respectively (both P<0.05). With the increased concentration of ω-3 from 1 μmol/L to 10 μmol/L, the proliferation ability of HUVECs was inhibited, and the number of migration cells decreased from 15.8±2.0 to 11.0±2.1, which was lower than control group (22.1±3.0) respectively (both P<0.05). In the angiogenesis assay, compared with control group (standard number: 43 721±4 654), the angiogenesis ability of HUVECs was significantly enhanced by ω-6 in concentration-dependent manner (1 μmol/L group: 63 238±4 795, 10 μmol/L group: 78 166±6 123, all P<0.01). Meanwhile, with the increased concentration of ω-3 from 1 μmol/L to 10 μmol/L, the angiogenesis ability was significantly decreased from 30 129±3 102 to 20 012±1 541(all P<0.01). The proliferation and migration ability of HUVECs were significantly promoted by ω-6 metabolites PGE2 (P<0.05) in a concentration-dependent manner. In contrast, ω-3 metabolites PGE3 significantly inhibited the proliferation and migration ability of HUVECs in a concentration-dependent manner (all P<0.05). After rofecoxib (a COX-2 specific inhibitor) inhibited the expression of COX-2, the expression level of PGE2 was significantly decreased in a dose-dependent manner. In co-culture system, whose gastric cancer cells expressed positive COX-2, ω-6 could increase angiogenesis of gastric cancer cells(P<0.01), but ω-3 could inhibit such angiogenesis(P<0.01). In co-culture system, whose gastric cancer cells did not express COX-2, ω-3 could inhibit the angiogenesis of gastric cancer cells (P<0.05), but ω-6 had no effect on angiogenesis.</p><p><b>CONCLUSIONS</b>The PUFA ω-6 can enhance the angiogenesis via the promotion of proliferation and migration of HUVECs, and COX-2 and PGE2 may play an important role in this process, whereas, the ω-3 can inhibit the angiogenesis through its middle metabolites PGE3 to inhibit the proliferation and migration of HUVECs. Results of this experiment may provide a new approach to inhibit and prevent the spread of gastric cancer.</p>


Subject(s)
Alprostadil , Pharmacology , Angiogenesis Inducing Agents , Metabolism , Pharmacology , Angiogenesis Inhibitors , Pharmacology , Cell Count , Methods , Cell Line, Tumor , Physiology , Cell Migration Assays , Cell Movement , Cell Proliferation , Coculture Techniques , Cyclooxygenase 2 , Pharmacology , Dinoprostone , Metabolism , Pharmacology , Fatty Acids, Omega-3 , Pharmacology , Fatty Acids, Omega-6 , Metabolism , Pharmacology , Fatty Acids, Unsaturated , Pharmacology , Human Umbilical Vein Endothelial Cells , Physiology , Humans , Lactones , Pharmacology , Neovascularization, Pathologic , Stomach Neoplasms , Sulfones , Pharmacology
8.
Yonsei Medical Journal ; : 27-34, 2017.
Article in English | WPRIM | ID: wpr-65066

ABSTRACT

PURPOSE: Ovarian cancer (OC) is the most fatal of gynecological malignancies with a high rate of recurrence. We aimed to evaluate the expression of solute carrier family 6, member 12 (SLC6A12) and methylation of its promoter CpG sites in a xenograft mouse model of metastatic OC, and to investigate the regulatory mechanisms that promote aggressive properties during OC progression. MATERIALS AND METHODS: Expression of SLC6A12 mRNA was determined by reverse-transcription quantitative polymerase chain reaction (RT-qPCR), and DNA methylation status of its promoter CpGs was detected by quantitative methylation-specific PCR. The metastatic potential of SLC6A12 was evaluated by in vitro migration/invasion transwell assays. Gene expression and DNA methylation of SLC6A12 and clinical outcomes were further investigated from publicly available databases from curatedOvarianData and The Cancer Genome Atlas. RESULTS: SLC6A12 expression was 8.1–14.0-fold upregulated and its DNA methylation of promoter CpG sites was 41–62% decreased in tumor metastases. After treatment with DNA methyltransferase inhibitor and/or histone deacetylase inhibitor, the expression of SLC6A12 was profoundly enhanced (~8.0-fold), strongly supporting DNA methylation-dependent epigenetic regulation of SLC6A12. Overexpression of SLC6A12 led to increased migration and invasion of ovarian carcinoma cells in vitro, approximately 2.0-fold and 3.3-fold, respectively. The meta-analysis showed that high expression of SLC6A12 was significantly associated with poor overall survival [hazard ratio (HR)=1.07, p value=0.016] and that low DNA methylation levels of SLC6A12 at specific promoter CpG site negatively affected patient survival. CONCLUSION: Our findings provide novel evidence for the biological and clinical significance of SLC6A12 as a metastasis-promoting gene.


Subject(s)
Animals , Carrier Proteins/genetics , Cell Line, Tumor , Cell Migration Assays , CpG Islands , DNA Methylation , Disease Progression , Epigenesis, Genetic , Female , Gene Expression Regulation, Neoplastic , Humans , Mice , Neoplasm Invasiveness , Neoplasm Transplantation , Ovarian Neoplasms/genetics , Polymerase Chain Reaction , Prognosis , Promoter Regions, Genetic , RNA, Messenger/metabolism , Up-Regulation
9.
Biol. Res ; 50: 1, 2017. tab, graf
Article in English | LILACS | ID: biblio-838973

ABSTRACT

BACKGROUND: Pathologic vascular smooth muscle cell (VSMC) proliferation and migration after vascular injury promotes the development of occlusive vascular disease. Therefore, an effective chemical agent to suppress aberrant proliferation and migration of VSMCs can be a potential therapeutic modality for occlusive vascular disease such as atherosclerosis and restenosis. To find an anti-proliferative chemical agent for VSMCs, we screened an in-house small molecule library, and the selected small molecule was further validated for its anti-proliferative effect on VSMCs using multiple approaches, such as cell proliferation assays, wound healing assays, transwell migration assays, and ex vivo aortic ring assay. RESULTS: Among 43 initially screened small molecule inhibitors of kinases that have no known anti-proliferative effect on VSMCs, a spleen tyrosine kinase (Syk) inhibitor (BAY61-3606) showed significant anti-proliferative effect on VSMCs. Further experiments indicated that BAY61 attenuated the VSMC proliferation in both concentration- and time-dependent manner, and it also significantly suppressed the migration of VSMCs as assessed by both wound healing assays and transwell assays. Additionally, BAY61 suppressed the sprouting of VSMCs from endothelium-removed aortic rings. CONCLUSION: The present study identified a Syk kinase inhibitor as a potent VSMC proliferation and migration inhibitor and warrants further studies to elucidate its underlying molecular mechanisms, such as its primary target, and to validate its in vivo efficacy as a therapeutic agent for restenosis and atherosclerosis.


Subject(s)
Animals , Rats , Pyrimidines/pharmacology , Cell Movement/drug effects , Niacinamide/analogs & derivatives , Myocytes, Smooth Muscle/drug effects , Cell Proliferation/drug effects , Syk Kinase/antagonists & inhibitors , Muscle, Smooth, Vascular/drug effects , Aorta, Thoracic/drug effects , Time Factors , Wound Healing/drug effects , Cells, Cultured , Blotting, Western , Reproducibility of Results , Rats, Sprague-Dawley , Niacinamide/pharmacology , Dose-Response Relationship, Drug , Drug Evaluation, Preclinical , Cell Migration Assays , Muscle, Smooth, Vascular/cytology
10.
Medisan ; 20(1)ene.-ene. 2016.
Article in Spanish | LILACS, CUMED | ID: lil-774461

ABSTRACT

La angiogénesis, que conduce a la formación de redes capilares, tiene un rol importante en un elevado número de eventos fisiológicos y patológicos, entre los cuales figuran el desarrollo embrionario, la cicatrización de heridas, la artritis, el crecimiento tumoral, la metástasis y los procesos isquémicos, como uno de los mecanismos de la protección endógena y exógena. A tales efectos se analizan los métodos más utilizados actualmente tanto in vivo como in vitro para los estudios de angiogénesis; también se destacan las ventajas y desventajas de cada uno y se enfatiza en la combinación de ambos.


The angiogenesis that leads to the formation of capillary nets, has an important role in a high number of physiological and pathological events, among which we can mention the embryonic development, wounds healing, arthritis, tumoral growth, metastasis and in the ischemic processes as one of the mechanisms of the endogenous and exogenous protection. To such effects the most used methods at the moment are analyzed either in vivo or in vitro for the angiogenesis studies; advantages and disadvantages of each one are highlighted too and the combination of both is emphasized.


Subject(s)
In Vitro Techniques , Neovascularization, Physiologic , Essay , Neovascularization, Pathologic , Review , Cell Migration Assays
11.
Braz. j. med. biol. res ; 49(12): e5826, 2016. graf
Article in English | LILACS | ID: biblio-828173

ABSTRACT

Glucagon-like peptide 1 (GLP-1), a kind of gut hormone, is used in the treatment of type 2 diabetes (T2D). Emerging evidence indicates that GLP-1 has anti-inflammatory activity. Chronic inflammation in the adipose tissue of obese individuals is a cause of insulin resistance and T2D. We hypothesized that GLP-1 analogue therapy in patients with T2D could suppress the inflammatory response of macrophages, and therefore inhibit insulin resistance. Our results showed that GLP-1 agonist (exendin-4) not only attenuated macrophage infiltration, but also inhibited the macrophage secretion of inflammatory cytokines including TNF-β, IL-6, and IL-1β. Furthermore, we observed that lipopolysaccharide (LPS)-induced macrophage conditioned media could impair insulin-stimulated glucose uptake. This effect was compensated by treatment with the conditioned media from macrophages treated with the combination of LPS and exendin-4. It was also observed that exendin-4 directly inhibited the activation of NF-κB in macrophages. In conclusion, our results indicated that GLP-1 improved inflammatory macrophage-derived insulin resistance by inhibiting NF-κB pathway and secretion of inflammatory cytokines in macrophages. Furthermore, our observations suggested that the anti-inflammatory effect of GLP-1 on macrophages can contribute to GLP-1 analogue therapy of T2D.


Subject(s)
Humans , Animals , Mice , Glucagon-Like Peptide 1/pharmacology , Inflammation Mediators/pharmacology , Inflammation/drug therapy , Insulin Resistance , Macrophages/drug effects , Peptides/pharmacology , Venoms/pharmacology , Adipose Tissue/metabolism , Cell Migration Assays , Inflammation/metabolism , Macrophages/metabolism
12.
Acta cir. bras ; 30(4): 242-246, 04/2015. graf
Article in English | LILACS | ID: lil-744278

ABSTRACT

PURPOSE: To evaluate the anti-inflammatory and analgesic effects of green tea (Camellia sinensis) in mice. METHODS: The anti-inflammatory effect of alcoholic extracts of green tea (AE) was evaluated in a cell migration assay with four groups of six Swiss mice receiving 0.07g/Kg or 0.14g/Kg EA (treatment groups), saline (negative control) or 10mg/Kg indomethacin (positive control) by gavage. One hour later 300 µg carrageen an was administered intraperitoneally or subcutaneously. The analgesic effect was evaluated using four groups of six animals receiving 0.07g/Kg or 0.14g/Kg EA, saline or 10mg/Kg indomethacin subcutaneously, followed 30 minutes later by 1% acetic acid. RESULTS: When administered subcutaneously at either dose (0.07g/Kg and 0.14g/Kg), AE inhibited carrageenan-induced cell migration (p<0.05). However, when administered by gavage, only the latter (0.14 g/Kg) was efficient (p<0.05). AE at both doses (0.07g/Kg and 0.14g/Kg) inhibited abdominal contortions (p<0.05), but the effect was not dose-dependent. CONCLUSION: Green tea was shown to have analgesic and anti-inflammatory properties and may constitute a natural treatment option in chronic inflammatory disorders. .


Subject(s)
Animals , Male , Mice , Analgesics/therapeutic use , Anti-Inflammatory Agents/therapeutic use , Camellia sinensis/chemistry , Phytotherapy/methods , Plant Extracts/therapeutic use , Tea/chemistry , Cell Migration Assays , Catechin/therapeutic use , Infusions, Subcutaneous , Indomethacin/therapeutic use , Inflammation/drug therapy , Reproducibility of Results , Time Factors , Treatment Outcome
13.
Article in English | WPRIM | ID: wpr-812175

ABSTRACT

Gambogic acid (GA) is an anticancer agent in phase ‖b clinical trial in China but its mechanism of action has not been fully clarified. The present study was designed to search the possible target-related proteins of GA in cancer cells using proteomic method and establish possible network using bioinformatic analysis. Cytotoxicity and anti-migration effects of GA in MDA-MB-231 cells were checked using MTT assay, flow cytometry, wound migration assay, and chamber migration assay. Possible target-related proteins of GA at early (3 h) and late stage (24 h) of treatment were searched using a proteomic technology, two-dimensional electrophoresis (2-DE). The possible network of GA was established using bioinformatic analysis. The intracellular expression levels of vimentin, keratin 18, and calumenin were determined using Western blotting. GA inhibited cell proliferation and induced cell cycle arrest at G2/M phase and apoptosis in MDA-MB-231 cells. Additionally, GA exhibited anti-migration effects at non-toxic doses. In 2-DE analysis, totally 23 possible GA targeted proteins were found, including those with functions in cytoskeleton and transport, regulation of redox state, metabolism, ubiquitin-proteasome system, transcription and translation, protein transport and modification, and cytokine. Network analysis of these proteins suggested that cytoskeleton-related proteins might play important roles in the effects of GA. Results of Western blotting confirmed the cleavage of vimentin, increase in keratin 18, and decrease in calumenin levels in GA-treated cells. In summary, GA is a multi-target compound and its anti-cancer effects may be based on several target-related proteins such as cytoskeleton-related proteins.


Subject(s)
Antineoplastic Agents , Pharmacokinetics , Apoptosis , Breast Neoplasms , Drug Therapy , Metabolism , Calcium-Binding Proteins , Genetics , Cell Line, Tumor , Cell Migration Assays , Cell Migration Inhibition , Cell Proliferation , Computational Biology , Methods , Cytoskeleton , Metabolism , Electrophoresis, Gel, Two-Dimensional , Flow Cytometry , Gene Expression , Humans , Keratin-18 , Genetics , Oxidation-Reduction , Protein Biosynthesis , Protein Transport , Proteomics , Methods , Transcription, Genetic , Ubiquitin-Specific Proteases , Pharmacokinetics , Vimentin , Genetics , Xanthones , Pharmacokinetics
14.
São Paulo; s.n; s.n; 2015. 158 p. graf, tab, ilus.
Thesis in Portuguese | LILACS | ID: biblio-881862

ABSTRACT

Concentrações séricas basais da proteína amiloide sérica A (SAA) estão significativamente aumentadas em pacientes com câncer e alguns autores sugerem uma relação causal. Trabalho anterior do grupo mostrou que a SAA induz a proliferação de duas linhagens de glioblastoma humano e afeta os processos de invasividade in vitro, sustentando um papel pró-tumoral para esta proteína. Com base nesse trabalho, investigamos a abrangência dos efeitos de SAA para outro tipo de célula tumoral e para isso escolhemos um painel de linhagens de melanoma humano e uma linhagem primária obtida a partir de aspirado de linfonodo de paciente com melanoma, por nós isolada. Observamos que apesar da célula precursora de melanomas, isto é, melanócito, não produzir SAA, todas as linhagens de melanoma produziram a proteína e expressaram alguns dos seus receptores. Além disso, quando estas células foram estimuladas com SAA houve uma inibição da proliferação em tempos curtos de exposição (48 horas) e efeitos citotóxicos após um tempo maior (7 dias). A SAA também afetou processos de invasividade e a produção das citocinas IL-6, IL-8 e TNF-α. Aos avaliarmos o efeito da SAA na interação das células de melanoma com células do sistema imune, vimos que a SAA ativou uma resposta imune anti-tumoral aumentando a expressão de moléculas co-estumolatórias, como CD69 e HLA-DR, e sua função citotóxica. Ainda, vimos que a produção de TNF-α, IFN-γ, IL-10, IL-1ß e IL-8 estimuladas por SAA podem contribuir com os efeitos desta. De forma geral estes resultados nos levam a crer que a SAA tem atividade anti-tumoral em melanomas. Finalizando, com base na importância do desenvolvimento da resistência às terapias atuais para o melanoma, observamos que em células resistentes ao PLX4032, um inibidor de BRAF, os efeitos imunomodulatórios induzidos pela SAA estão abolidos, possivelmente identificando um novo componente da resistência


Basal serum concentrations of the protein serum amyloid A are significantly increased in cancer patients and some authors suggest a causal relationship. Previous work of our research group showed that SAA induces proliferation of two cell lines of human glioblastoma and affects invasiveness processes in vitro, supporting a pro-tumor role for this protein. Based on this work, we investigated the extent of SAA effects to another type of tumor cell and we chose a panel of human melanoma cell lines and primary line obtained from a patient with melanoma by lymph node aspirate. Melanoma cells were isolated by us. We observed that while the precursor cells of melanoma, melanocytes, do not produce SAA, all melanoma cell lines expressed the protein and produced some of their receptors. Moreover, when these cells were stimulated with SAA there was an inhibition of proliferation in short exposure times (48 hours) and cytotoxic effects after a longer period (7 days). SAA also affected invasive procedures and the production of the cytokines IL-6, IL-8 and TNF-α. To evaluate the SAA effect in the interaction of melanoma cells with immune system cells, we found that SAA activated an anti-tumor immune response by increasing the expression of co-estimulatory molecules such as CD69 and HLA-DR, and their cytotoxic function. Furthermore, we found that the production of TNF-α, IFN-γ, IL-10, IL-1ß and IL-8 stimulated by SAA can contribute to this effect. In general these results lead us to believe that the SAA has anti-tumor activity in melanomas. Finally, based on the importance of the resistance development to current therapies for melanoma we observed that in cells resistant to PLX4032, a BRAF inhibitor, the immunomodulatory effects induced by SAA are abolished, possibly identifying a new resistance component


Subject(s)
Serum Amyloid A Protein/analysis , Serum Amyloid A Protein/adverse effects , Melanoma/physiopathology , Gene Expression , Proto-Oncogene Proteins B-raf/adverse effects , Toll-Like Receptor 2/genetics , Toll-Like Receptor 4/genetics , Cell Migration Assays/instrumentation , Receptor for Advanced Glycation End Products/genetics
15.
Journal of Breast Cancer ; : 200-206, 2014.
Article in English | WPRIM | ID: wpr-20462

ABSTRACT

PURPOSE: Synuclein-gamma (SNCG), which was initially identified as breast cancer specific gene 1, is highly expressed in advanced breast cancers, but not in normal or benign breast tissue. This study aimed to evaluate the effects of SNCG siRNA-treatment on breast cancer cells and elucidate the associated mechanisms. METHODS: Vectors containing SNCG and negative control (NC) siRNAs were transfected into MDA-MB-231 cells; mRNA levels were determined by real-time polymerase chain reaction. Cell proliferation was evaluated using the MTT assay, cell migration was assessed by the Transwell assay, apoptosis and cell cycle analyses were conducted with the flow cytometer, and Western blot analysis was performed to determine the relative levels of AKT, ERK, p-AKT, and p-ERK expression. RESULTS: SNCG mRNA levels were significantly reduced in MDA-MB-231 cells transfected with SNCG siRNA. Our results indicate that in SNCG siRNA-treated cells, cell migration and proliferation decreased significantly, apoptosis was induced, and the cell cycle was arrested. Western blot analysis indicated that the protein levels of p-AKT and p-ERK were much lower in the SNCG siRNA-treated groups, than in the control and NC groups. CONCLUSION: SNCG siRNA could decrease the migration and proliferation of breast cancer cells by downregulating the phosphorylation of AKT and ERK.


Subject(s)
Apoptosis , Blotting, Western , Breast , Breast Neoplasms , Cell Cycle , Cell Migration Assays , Cell Movement , Cell Proliferation , Extracellular Signal-Regulated MAP Kinases , Phosphorylation , Proto-Oncogene Proteins c-akt , Real-Time Polymerase Chain Reaction , RNA, Messenger , RNA, Small Interfering , Synucleins
16.
Braz. j. med. biol. res ; 46(8): 650-658, ago. 2013. graf
Article in English | LILACS | ID: lil-684524

ABSTRACT

Cisplatin resistance remains one of the major obstacles when treating epithelial ovarian cancer. Because oxaliplatin and nedaplatin are effective against cisplatin-resistant ovarian cancer in clinical trials and signal transducer and activator of transcription 3 (STAT3) is associated with cisplatin resistance, we investigated whether overcoming cisplatin resistance by oxaliplatin and nedaplatin was associated with the STAT3 pathway in ovarian cancer. Alamar blue, clonogenic, and wound healing assays, and Western blot analysis were used to compare the effects of platinum drugs in SKOV-3 cells. At an equitoxic dose, oxaliplatin and nedaplatin exhibited similar inhibitory effects on colony-forming ability and greater inhibition on cell motility than cisplatin in ovarian cancer. Early in the time course of drug administration, cisplatin increased the expression of pSTAT3 (Tyr705), STAT3α, VEGF, survivin, and Bcl-XL, while oxaliplatin and nedaplatin exhibited the opposite effects, and upregulated pSTAT3 (Ser727) and STAT3β. The STAT3 pathway responded early to platinum drugs associated with cisplatin resistance in epithelial ovarian cancer and provided a rationale for new therapeutic strategies to reverse cisplatin resistance.


Subject(s)
Animals , Humans , Rats , Antineoplastic Agents/administration & dosage , Drug Resistance, Neoplasm/physiology , Neoplasms, Glandular and Epithelial/drug therapy , Ovarian Neoplasms/drug therapy , /metabolism , Signal Transduction/drug effects , Apoptosis Regulatory Proteins/genetics , Cell Line, Tumor , Cell Migration Assays/methods , Cell Proliferation/drug effects , Cisplatin/administration & dosage , Drug Resistance, Neoplasm/drug effects , Gene Expression/drug effects , Inhibitor of Apoptosis Proteins/genetics , Organoplatinum Compounds/administration & dosage , Oxazines/pharmacology , Vascular Endothelial Growth Factor A/genetics , Xanthenes/pharmacology , bcl-X Protein/genetics
17.
Clinics ; 67(8): 901-906, Aug. 2012. ilus, graf
Article in English | LILACS | ID: lil-647793

ABSTRACT

OBJECTIVES: Tumor-associated macrophages that generally exhibit an alternatively activated (M2) phenotype have been linked to tumor progression and metastasis. However, the role of M2-polarized macrophages in the growth and metastasis of lung adenocarcinoma remains enigmatic. The aim of this study was to explore the effect of M2 macrophages on the proliferation and migration of mouse Lewis lung carcinoma cells and tumor-induced lymphangiogenesis. METHODS: Trypan blue staining and the Transwell migration assay were performed to evaluate the effects of activated (M1 or M2) macrophages on the proliferation and migration of Lewis cells. Furthermore, vascular endothelial growth factor-C expression in Lewis cells and nitric oxide secretion from activated macrophages were detected during the co-culture assay. Following treatment with activated macrophages, lymphatic endothelial cells differentiated into capillary-like structures, and the induction of Lewis cell migration was assessed using a twodimensional Matrigel-based assay. RESULTS: In the co-culture Transwell system, the proliferation and migration of Lewis cells were promoted by M2 macrophages. Moreover, the co-culture significantly increased the expression of vascular endothelial growth factor-C by Lewis cells and reduced the secretion of nitric oxide from M2 macrophages, which subsequently led to the capillary morphogenesis of lymphatic endothelial cells. Interestingly, following co-culture with Lewis cells, the function of RAW264.7 cells was polarized toward that of the M2 macrophage phenotype. CONCLUSION: M2-polarized macrophages promoted the metastatic behavior of Lewis cells by inducing vascular endothelial growth factor-C expression. Thus, the interruption of signaling between M2 macrophages and Lewis cells may be considered to be a new therapeutic strategy.


Subject(s)
Animals , Mice , Carcinoma, Lewis Lung/secondary , Lung Neoplasms/pathology , Macrophages/physiology , Vascular Endothelial Growth Factor C/metabolism , Cell Line, Tumor , Cell Migration Assays , Cell Movement , Cell Proliferation , Carcinoma, Lewis Lung/metabolism , Endothelial Cells/pathology , Lung Neoplasms/metabolism , Lymphangiogenesis/physiology , Macrophages/cytology , Time Factors , Vascular Endothelial Growth Factor C/physiology
18.
Article in Korean | WPRIM | ID: wpr-30039

ABSTRACT

PURPOSE: Burn injury cause pruritis, pain, psychological and functional sequelae. The one of burn injury sequelae is the hypertrophic scar. It is difficult to control devastating fibrotic condition for hypertrophic scar. The objective of this study was to investigated the therapeutic effect on burn hypertrophic scar and wound healing for sequelae of burn injury by Porcine placenta extract (PPE). METHODS: To investigate the effect of PPE, we performed in vitro cell cytotoxity test (MTT assay), antioxidant activity assay (SOD like activity), melanin content assay, cell migration asssay and RT-PCR. RESULTS: As a result of cell cytotoxity test (MTT assay), PPE showed above 80% cell viability. From Antioxidant activity assay (SOD like activity), this effect was similar to vitamin C. In the melanin content assay, melanin synthesis was inhibited 23% on PPE treatment than control. PPE enhanced cell migration on human fibroblast and decreased the expression of hypertropic scar related gene (a-SMA and P311). CONCLUSION: Our data showed anti-oxidant effect, diminution of melanin and decrease of the expression of hypertropic scar related gene on the treatment of PPE. These results may provide the insight into the potential use of porcine placenta extract as support to control skin fibrosis related to burn hypertrophic scar and alternative medicine for burn sequelae.


Subject(s)
Antioxidants , Ascorbic Acid , Burns , Cell Migration Assays , Cell Movement , Cell Survival , Cicatrix , Cicatrix, Hypertrophic , Complementary Therapies , Fibroblasts , Fibrosis , Humans , Melanins , Placenta , Pruritus , Skin , Wound Healing
19.
Braz. oral res ; 25(1): 13-18, Jan.-Feb. 2011. ilus, graf
Article in English | LILACS | ID: lil-595833

ABSTRACT

Pulp repair is a complex process whose mechanisms are not yet fully understood. The first immune cells to reach the damaged pulp are neutrophils that play an important role in releasing cytokines and in phagocytosis. The objective of this study was to analyze the effect of different pulp-capping materials on the secretion of interleukin-1 beta (IL-1β) and interleukin-8 (IL-8) by migrating human neutrophils. Neutrophils were obtained from the blood of three healthy donors. The experimental groups were calcium hydroxide [Ca(OH)2], an adhesive system (Single Bond), and mineral trioxide aggregate (MTA). Untreated cells were used as control. Transwell chambers were used in performing the assays to mimic an in vivo situation of neutrophil chemotaxis. The pulp-capping materials were placed in the lower chamber and the human neutrophils, in the upper chamber. The cells were counted and the culture medium was assayed using ELISA kits for detecting and quantifying IL-1β and IL8. The data were compared by ANOVA followed by Tukey's test (p < 0.05). The secretion of IL-8 was significantly higher in all groups in comparison to the control group (p < 0.05). The adhesive system group showed higher IL-8 than the MTA group (p < 0.05). The secretion of IL-1β was significantly greater only in the MTA group (p < 0.001). It was concluded that only MTA is able to improve the secretion of IL-1β, and all materials tested increased IL-8 secretion. These results combined with all the other biological advantages of MTA indicate that it could be considered the material of choice for dental pulp capping.


Subject(s)
Humans , Dental Pulp Capping , Dental Pulp/drug effects , Interleukin-1beta , Neutrophils/drug effects , Analysis of Variance , Aluminum Compounds/pharmacology , Bisphenol A-Glycidyl Methacrylate/pharmacology , Bone Cements/pharmacology , Cell Migration Assays , Calcium Compounds/pharmacology , Calcium Hydroxide/pharmacology , Drug Combinations , Dental Pulp/immunology , Materials Testing , Neutrophils , Oxides/pharmacology , Silicates/pharmacology
20.
Article in English | WPRIM | ID: wpr-206103

ABSTRACT

PURPOSE: A composite of aluminum and vanadium (Ti-6Al-4V) is one of the most common compositions of titanium-based alloys. Unfortunately, vanadium has been found to cause adverse reactions. We evaluated the effects of vanadium containing titanium alloy (Ti-6Al-4V) on an osteoblast-like cell line (SaOS-2). MATERIALS AND METHODS: We studied the biologic and morphologic responses of SaOS-2 cell to Ti alloy with grit blasting and Ti coated Ti alloy with grit blasting. We performed energy-dispersive x-ray spectroscopy (EDS) and scanning electron microscopy (SEM) investigations and performed a cell proliferation assay, ALP activity, and cell migration assay of SaOS-2 cells. RESULTS: The morphologic assessment of cells through SEM showed that the two surfaces were covered with similar amounts of small slender osteoblast like cells. The amount of proliferation, ALP activity and the migration extent of SaOS-2 cells on the surfaces of each group were not statistically different. CONCLUSION: We used a grit-blasted Ti-coated Ti alloy, coated using electron beam deposition, and a grit-blasted Ti alloy to evaluate the toxicity of Ti-6Al-4V on SaOS-2 cell. Compared with pure titanium, the vanadium-containing Ti-alloy did not show an adverse effect on SaOS-2 cells.


Subject(s)
Alloys , Aluminum , Cell Line , Cell Migration Assays , Cell Proliferation , Electrons , Microscopy, Electron, Scanning , Osteoblasts , Spectrum Analysis , Titanium , Vanadium
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