摘要
SUMMARY: The objective of this study was to investigate the therapeutic wound healing potential and molecular mechanisms of shikonin as small molecules in vitro. A mouse burn model was used to explore the potential therapeutic effect of shikonin; we traced proliferating cells in vivo to locate the active area of skin cell proliferation. Through the results of conventional pathological staining, we found that shikonin has a good effect on the treatment of burned skin and promoted the normal distribution of skin keratin at the damaged site. At the same time, shikonin also promoted the proliferation of skin cells at the damaged site; importantly, we found a significant increase in the number of fibroblasts at the damaged site treated with shikonin. Most importantly, shikonin promotes fibroblasts to repair skin wounds by regulating the PI3K/AKT signaling pathway. This study shows that shikonin can effectively promote the proliferation of skin cell, and local injection of fibroblasts in burned skin can play a certain therapeutic role.
El objetivo de este trabajo fue investigar el potencial terapéutico de cicatrización de heridas y los mecanismos moleculares de la shikonina como moléculas pequeñas in vitro. Se utilizó un modelo de quemaduras en ratones para explorar el posible efecto terapéutico de la shikonina; Rastreamos las células en proliferación in vivo para localizar el área activa de proliferación de células de la piel. A través de los resultados de la tinción para patología convencional, encontramos que la shikonina tiene un buen efecto en el tratamiento de la piel quemada y promueve la distribución normal de la queratina de la piel en el sitio dañado. Al mismo tiempo, la shikonina también promovió la proliferación de células de la piel en el sitio dañado. Es importante destacar que encontramos un aumento significativo en la cantidad de fibroblastos en el sitio dañado tratado con shikonina. Lo más importante es que la shikonina promueve la función reparadora de fibroblastos en las heridas de la piel regulando la vía de señalización PI3K/ AKT. Este estudio muestra que la shikonina puede promover eficazmente la proliferación de células de la piel y que la inyección local de fibroblastos en la piel quemada puede desempeñar un cierto papel terapéutico.
Subject(s)
Animals , Mice , Wound Healing/drug effects , Burns/drug therapy , Naphthoquinones/administration & dosage , Skin , In Vitro Techniques , Naphthoquinones/pharmacology , Phosphatidylinositol 3-Kinases , Cell Proliferation/drug effects , Disease Models, Animal , Proto-Oncogene Proteins c-akt , Fibroblasts , Mice, Inbred C57BL摘要
SUMMARY: Esophageal cancer is one of the most aggressive gastrointestinal cancers. Invasion and metastasis are the main causes of poor prognosis of esophageal cancer. SPRY2 has been reported to exert promoting effects in human cancers, which controls signal pathways including PI3K/AKT and MAPKs. However, the expression of SPRY2 in esophageal squamous cell carcinoma (ESCC) and its underlying mechanism remain unclear. In the present study, we aimed to investigate the detailed role of SPRY2 in the regulation of cell proliferation, invasion and ERK/AKT signaling pathway in ESCC. It was identified that the expression level of SPRY2 in ESCC was remarkably decreased compared with normal tissues, and it was related to clinicopathologic features and prognosis ESCC patients. The upregulation of SPRY2 expression notably inhibited the proliferation, migration and invasion of Eca-109 cells. In addition, the activity of ERK /AKT signaling was also suppressed by the SPRY2 upregulation in Eca-109 cells. Our study suggests that overexpression of SPRY2 suppress cancer cell proliferation and invasion of by through suppression of the ERK/AKT signaling pathways in ESCC. Therefore, SPRY2 may be a promising prognostic marker and therapeutic target for ESCC.
El cáncer de esófago es uno de los cánceres gastrointestinales más agresivos. La invasión y la metástasis son las principales causas de mal pronóstico del cáncer de esófago. Se ha informado que SPRY2 ejerce efectos promotores en los cánceres humanos, que controla las vías de señales, incluidas PI3K/AKT y MAPK. Sin embargo, la expresión de SPRY2 en el carcinoma de células escamosas de esófago (ESCC) y su mecanismo subyacente aún no están claros. En el presente estudio, nuestro objetivo fue investigar el papel detallado de SPRY2 en la regulación de la proliferación celular, la invasión y la vía de señalización ERK/AKT en ESCC. Se identificó que el nivel de expresión de SPRY2 en ESCC estaba notablemente disminuido en comparación con los tejidos normales, y estaba relacionado con las características clínico-patológicas y el pronóstico de los pacientes con ESCC. La regulación positiva de la expresión de SPRY2 inhibió notablemente la proliferación, migración e invasión de células Eca-109. Además, la actividad de la señalización de ERK/AKT también fue suprimida por la regulación positiva de SPRY2 en las células Eca-109. Nuestro estudio sugiere que la sobreexpresión de SPRY2 suprime la proliferación y la invasión de células cancerosas mediante la supresión de las vías de señalización ERK/AKT en ESCC. Por lo tanto, SPRY2 puede ser un marcador de pronóstico prometedor y un objetivo terapéutico para la ESCC.
Subject(s)
Humans , Esophageal Neoplasms/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Esophageal Squamous Cell Carcinoma/metabolism , Membrane Proteins/metabolism , Immunohistochemistry , Biomarkers, Tumor , Blotting, Western , Extracellular Signal-Regulated MAP Kinases , Cell Proliferation , Proto-Oncogene Proteins c-akt摘要
Objective: To investigate the clinical and pathologic features and diagnosis of follicular lymphoma (FL) with a predominantly diffuse growth pattern (DFL) with 1p36 deletion. Methods: Eight cases of DFL with 1p36 deletion diagnosed at Department of Pathology, Beijing Friendship Hospital, Capital Medical University (n=5) and the Affiliated Cancer Hospital of Zhengzhou University, Henan Cancer Hospital (n=3) from January 2017 to January 2023 were included. Their clinicopathologic features and follow-up data were analyzed. Immunohistochemistry and fluorescence in situ hybridization (FISH) were performed. Results: There were five males and three females, with a median age of 67 years, and inguinal lymphadenopathy was found as the main symptom. Histologically, similar morphologic features were sheared among all cases, with effaced nodal structure and characterized by proliferation of centrocytes in a diffuse pattern, with or without follicular components. The germinal center-related markers such as CD10 and/or bcl-6 were expressed in the tumor cells, and 1p36 deletion but not bcl-2 translocation was appreciable in these cases. Conclusions: DFL with 1p36 deletion is a rare subtype of FL, with some overlaps with other types of FL or indolent B-cell lymphomas in their pathologic features. An accurate diagnosis requires comprehensive considerations based on their clinical, pathologic, immunohistochemical, and molecular features.
Subject(s)
Male , Female , Humans , Aged , Lymphoma, Follicular/pathology , In Situ Hybridization, Fluorescence , Lymphoma, B-Cell/pathology , Chromosome Deletion , Cell Proliferation摘要
Pathological vascular remodeling is a hallmark of various vascular diseases. Previous research has established the significance of andrographolide in maintaining gastric vascular homeostasis and its pivotal role in modulating endothelial barrier dysfunction, which leads to pathological vascular remodeling. Potassium dehydroandrographolide succinate (PDA), a derivative of andrographolide, has been clinically utilized in the treatment of inflammatory diseases precipitated by viral infections. This study investigates the potential of PDA in regulating pathological vascular remodeling. The effect of PDA on vascular remodeling was assessed through the complete ligation of the carotid artery in C57BL/6 mice. Experimental approaches, including rat aortic primary smooth muscle cell culture, flow cytometry, bromodeoxyuridine (BrdU) incorporation assay, Boyden chamber cell migration assay, spheroid sprouting assay, and Matrigel-based tube formation assay, were employed to evaluate the influence of PDA on the proliferation and motility of smooth muscle cells (SMCs). Molecular docking simulations and co-immunoprecipitation assays were conducted to examine protein interactions. The results revealed that PDA exacerbates vascular injury-induced pathological remodeling, as evidenced by enhanced neointima formation. PDA treatment significantly increased the proliferation and migration of SMCs. Further mechanistic studies disclosed that PDA upregulated myeloid differentiation factor 88 (MyD88) expression in SMCs and interacted with T-cadherin (CDH13). This interaction augmented proliferation, migration, and extracellular matrix deposition, culminating in pathological vascular remodeling. Our findings underscore the critical role of PDA in the regulation of pathological vascular remodeling, mediated through the MyD88/CDH13 signaling pathway.
Subject(s)
Mice , Rats , Animals , Myeloid Differentiation Factor 88/metabolism , Vascular Remodeling , Cell Proliferation , Vascular System Injuries/pathology , Carotid Artery Injuries/pathology , Molecular Docking Simulation , Muscle, Smooth, Vascular , Cell Movement , Mice, Inbred C57BL , Signal Transduction , Succinates/pharmacology , Potassium/pharmacology , Cells, Cultured , Diterpenes , Cadherins摘要
Abstract Mammals have a limited capacity to regenerate their tissues and organs. One of the mechanisms associated with natural regeneration is dedifferentiation. Several small molecules such as vitamin C and growth factors could improve reprogramming efficiency. In this study, the NTERA2-D1 (NT2) cells were induced towards differentiation (NT2-RA) with 10-5 M retinoic acid (RA) for three days and then subjected to various amounts of vitreous humor (VH). Results show that the growth rate of these cells was reduced, while this rate was partly restored upon treatment with VH (NT2-RA-VH). Cell cycle analysis with PI method also showed that the numbers of cells at the S phase of the cell cycle in these cells were increased. The levels of SSEA3 and TRA-1-81 antigens in NT2-RA were dropped but they increased in NT2- RA-VH to a level similar to the NT2 cells. The level of SSEA1 had an opposite pattern. Expression of OCT4 gene dropped after RA treatment, but it was recovered in NT2-RA-VH cells. In conclusion, we suggest VH as a potent mixture for improving the cellular reprogramming leading to dedifferentiation.
Resumo Os mamíferos têm uma capacidade limitada de regenerar seus tecidos e órgãos. Um dos mecanismos associados à regeneração natural é a desdiferenciação. Várias moléculas pequenas, como vitamina C e fatores de crescimento, podem melhorar a eficiência da reprogramação. Neste estudo, as células NTERA2-D1 (NT2) foram induzidas à diferenciação (NT2-RA) com ácido retinóico (RA) 10-5 M por três dias e depois submetidas a várias quantidades de humor vítreo (VH). Os resultados mostram que a taxa de crescimento dessas células foi reduzida, enquanto essa taxa foi parcialmente restaurada após o tratamento com VH (NT2-RA-VH). A análise do ciclo celular com o método PI também mostrou que o número de células na fase S do ciclo celular nessas células estava aumentado. Os níveis de antígenos SSEA3 e TRA-1-81 em NT2-RA diminuíram, mas aumentaram em NT2-RA-VH a um nível semelhante ao das células NT2. O nível de SSEA1 teve um padrão oposto. A expressão do gene OCT4 diminuiu após o tratamento com AR, mas foi recuperado em células NT2-RA-VH. Em conclusão, sugerimos o VH como uma mistura potente para melhorar a reprogramação celular levando à desdiferenciação.
Subject(s)
Humans , Vitreous Body , Cell Proliferation , Cell Dedifferentiation , Tretinoin , Tumor Cells, Cultured , Cell Differentiation , Cell Division , Cell Line摘要
OBJECTIVE@#To investigate the role and molecular mechanism of exosomal miR-224-5p in colorectal cancer (CRC).@*METHODS@#The miR-224-5p expression in CRC patient tissues and cell-derived exosomes was measured by laser capture microdissection and qRT-PCR, respectively. Dual-luciferase reporter gene assay was used to determine the target gene of miR-224-5p. The protein expressions of p53 and unc-51 like kinase 2 (ULK2) in CRC cells were detected by western blot. Flow cytometry was used to detect cell cycle and apoptosis. Cell proliferation was measured by CCK8 and EdU assay.@*RESULTS@#The miR-224-5p expression was upregulated in CRC tissues and increased progressively with the rise of CRC stage. CRC cells secreted extracellular miR-224-5p mainly in an exosome-dependent manner, and then miR-224-5p could be transferred to surrounding tumor cells to regulate cell proliferation in the form of autocrine or paracrine. Moreover, ULK2 was characterized as a direct target of miR-224-5p and was downregulated in CRC tissues. Interestingly, ULK2 inhibited CRC cell proliferation in a p53-dependent manner. Furthermore, exosome-derived miR-224-5p partially reversed the proliferation regulation of ULK2 on CRC cells.@*CONCLUSION@#Our findings demonstrate that exosome-transmitted miR-224-5p promotes p53-dependent cell proliferation by targeting ULK2 in CRC, which may offer promising targets for CRC prevention and therapy.
Subject(s)
Humans , MicroRNAs/metabolism , Tumor Suppressor Protein p53/metabolism , Exosomes/metabolism , Cell Proliferation/genetics , Colorectal Neoplasms/genetics , Cell Line, Tumor , Gene Expression Regulation, Neoplastic摘要
BACKGROUND@#Triple-negative breast cancer (TNBC) is a type of highly invasive breast cancer with a poor prognosis. According to new research, long noncoding RNAs (lncRNAs) play a significant role in the progression of cancer. Although the role of lncRNAs in breast cancer has been well reported, few studies have focused on TNBC. This study aimed to explore the biological function and clinical significance of forkhead box C1 promoter upstream transcript (FOXCUT) in triple-negative breast cancer.@*METHODS@#Based on a bioinformatic analysis of the cancer genome atlas (TCGA) database, we detected that the lncRNA FOXCUT was overexpressed in TNBC tissues, which was further validated in an external cohort of tissues from the General Surgery Department of the First Affiliated Hospital of Nanjing Medical University. The functions of FOXCUT in proliferation, migration, and invasion were detected in vitro or in vivo. Luciferase assays and RNA immunoprecipitation (RIP) were performed to reveal that FOXCUT acted as a competitive endogenous RNA (ceRNA) for the microRNA miR-24-3p and consequently inhibited the degradation of p38.@*RESULTS@#lncRNA FOXCUT was markedly highly expressed in breast cancer, which was associated with poor prognosis in some cases. Knockdown of FOXCUT significantly inhibited cancer growth and metastasis in vitro or in vivo. Mechanistically, FOXCUT competitively bounded to miR-24-3p to prevent the degradation of p38, which might act as an oncogene in breast cancer.@*CONCLUSION@#Collectively, this research revealed a novel FOXCUT/miR-24-3p/p38 axis that affected breast cancer progression and suggested that the lncRNA FOXCUT could be a diagnostic marker and therapeutic target for breast cancer.
Subject(s)
Humans , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Gene Expression Regulation, Neoplastic/genetics , MAP Kinase Signaling System , MicroRNAs/metabolism , p38 Mitogen-Activated Protein Kinases/metabolism , RNA, Long Noncoding/metabolism , Triple Negative Breast Neoplasms/pathology摘要
BACKGROUND@#Bladder cancer, characterized by a high potential of tumor recurrence, has high lifelong monitoring and treatment costs. To date, tumor cells with intrinsic softness have been identified to function as cancer stem cells in several cancer types. Nonetheless, the existence of soft tumor cells in bladder tumors remains elusive. Thus, our study aimed to develop a micro-barrier microfluidic chip to efficiently isolate deformable tumor cells from distinct types of bladder cancer cells.@*METHODS@#The stiffness of bladder cancer cells was determined by atomic force microscopy (AFM). The modified microfluidic chip was utilized to separate soft cells, and the 3D Matrigel culture system was to maintain the softness of tumor cells. Expression patterns of integrin β8 (ITGB8), protein kinase B (AKT), and mammalian target of rapamycin (mTOR) were determined by Western blotting. Double immunostaining was conducted to examine the interaction between F-actin and tripartite motif containing 59 (TRIM59). The stem-cell-like characteristics of soft cells were explored by colony formation assay and in vivo studies upon xenografted tumor models.@*RESULTS@#Using our newly designed microfluidic approach, we identified a small fraction of soft tumor cells in bladder cancer cells. More importantly, the existence of soft tumor cells was confirmed in clinical human bladder cancer specimens, in which the number of soft tumor cells was associated with tumor relapse. Furthermore, we demonstrated that the biomechanical stimuli arising from 3D Matrigel activated the F-actin/ITGB8/TRIM59/AKT/mTOR/glycolysis pathways to enhance the softness and tumorigenic capacity of tumor cells. Simultaneously, we detected a remarkable up-regulation in ITGB8, TRIM59, and phospho-AKT in clinical bladder recurrent tumors compared with their non-recurrent counterparts.@*CONCLUSIONS@#The ITGB8/TRIM59/AKT/mTOR/glycolysis axis plays a crucial role in modulating tumor softness and stemness. Meanwhile, the soft tumor cells become more sensitive to chemotherapy after stiffening, that offers new insights for hampering tumor progression and recurrence.
Subject(s)
Animals , Mice , Humans , Proto-Oncogene Proteins c-akt/metabolism , Actins/metabolism , Neoplasm Recurrence, Local , TOR Serine-Threonine Kinases/metabolism , Urinary Bladder Neoplasms , Glycolysis , Cell Line, Tumor , Cell Proliferation , Mammals/metabolism , Tripartite Motif Proteins/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , Integrin beta Chains摘要
PURPOSE@#Mannitol is one of the first-line drugs for reducing cerebral edema through increasing the extracellular osmotic pressure. However, long-term administration of mannitol in the treatment of cerebral edema triggers damage to neurons and astrocytes. Given that neural stem cell (NSC) is a subpopulation of main regenerative cells in the central nervous system after injury, the effect of mannitol on NSC is still elusive. The present study aims to elucidate the role of mannitol in NSC proliferation.@*METHODS@#C57 mice were derived from the animal house of Zunyi Medical University. A total of 15 pregnant mice were employed for the purpose of isolating NSCs in this investigation. Initially, mouse primary NSCs were isolated from the embryonic cortex of mice and subsequently identified through immunofluorescence staining. In order to investigate the impact of mannitol on NSC proliferation, both cell counting kit-8 assays and neurospheres formation assays were conducted. The in vitro effects of mannitol were examined at various doses and time points. In order to elucidate the role of Aquaporin 4 (AQP4) in the suppressive effect of mannitol on NSC proliferation, various assays including reverse transcription polymerase chain reaction, western blotting, and immunocytochemistry were conducted on control and mannitol-treated groups. Additionally, the phosphorylated p38 (p-p38) was examined to explore the potential mechanism underlying the inhibitory effect of mannitol on NSC proliferation. Finally, to further confirm the involvement of the p38 mitogen-activated protein kinase-dependent (MAPK) signaling pathway in the observed inhibition of NSC proliferation by mannitol, SB203580 was employed. All data were analyzed using SPSS 20.0 software (SPSS, Inc., Chicago, IL). The statistical analysis among multiple comparisons was performed using one-way analysis of variance (ANOVA), followed by Turkey's post hoc test in case of the data following a normal distribution using a Shapiro-Wilk normality test. Comparisons between 2 groups were determined using Student's t-test, if the data exhibited a normal distribution using a Shapiro-Wilk normality test. Meanwhile, data were shown as median and interquartile range and analyzed using the Mann-Whitney U test, if the data failed the normality test. A p < 0.05 was considered as significant difference.@*RESULTS@#Primary NSC were isolated from the mice, and the characteristics were identified using immunostaining analysis. Thereafter, the results indicated that mannitol held the capability of inhibiting NSC proliferation in a dose-dependent and time-dependent manner using cell counting kit-8, neurospheres formation, and immunostaining of Nestin and Ki67 assays. During the process of mannitol suppressing NSC proliferation, the expression of AQP4 mRNA and protein was downregulated, while the gene expression of p-p38 was elevated by reverse transcription polymerase chain reaction, immunostaining, and western blotting assays. Subsequently, the administration of SB203580, one of the p38 MAPK signaling pathway inhibitors, partially abrogated this inhibitory effect resulting from mannitol, supporting the fact that the p38 MAPK signaling pathway participated in curbing NSC proliferation induced by mannitol.@*CONCLUSIONS@#Mannitol inhibits NSC proliferation through downregulating AQP4, while upregulating the expression of p-p38 MAPK.
Subject(s)
Humans , Animals , Mannitol/pharmacology , Brain Edema , Neural Stem Cells/metabolism , MAP Kinase Signaling System , p38 Mitogen-Activated Protein Kinases/pharmacology , Cell Proliferation摘要
Objective To establish U251 cells with inhibited expression of interferon-γ inducible protein 30 (IFI30), and to investigate the effect of IFI30 on cell biological function as well as its underlying mechanism. Methods Three knockdown sequences which target IFI30 were designed online and 3 small interfering RNAs (siRNA) were synthesized. After transfection, the inhibition efficiency was detected by real-time quantitative PCR. The siRNA sequence with the highest inhibition efficiency was selected to create short hairpin RNA (shRNA) plasmids. The recombinant plasmids and packaging plasmids were co-transfected into HEK293T cells to prepare lentivirus. The glioma U251 cells were transfected with lentivirus, and the positive cells were screened by puromycin. CCK-8 assay, 5-ethyl-2'-deoxyuridine (EdU) and colony formation assays were used to analyze cell proliferation; the flow cytometry was used to analyze cell cycle and apoptosis; the TranswellTM assay was used to detect cell invasion; the wound-healing assay was employed to detect cell migration, and western blot analysis to detect the protein expresison of cyclin D1, B-cell lymphoma factor 2 (Bcl2), epithelial cadherin (E-cadherin), neural cadherin (N-cadherin), signal transducer and activator of transcription 1 (STAT1). Results The sequence which effectively target IFI30 was screened and U251 cell line capable of inhibiting the IFI30 expression was successfully established. When IFI30 expression was knocked down, the proliferation of U251 cells was inhibited, along with increased ratio of cells in the phase G0/G1, the decreased phase S, the increased rate of cell apoptosis. The cell invasion and migration capabilities was also reduced. The decreased expression of cyclin D1, Bcl2 and N-cadherin were observed in U251 cells, and the expression of E-cadherin and the phosphorylation of STAT1 were found increased. Conclusion Knockdown of IFI30 inhibits the proliferation, invasion and migration of human glioma cell U251 and promotes its apoptosis by activating STAT1.
Subject(s)
Humans , Cyclin D1/genetics , HEK293 Cells , Interferon-gamma , RNA, Small Interfering , Apoptosis/genetics , Cadherins , Cell Proliferation/genetics , Glioma/genetics , Proto-Oncogene Proteins c-bcl-2 , Oxidoreductases Acting on Sulfur Group Donors , STAT1 Transcription Factor/genetics摘要
OBJECTIVE@#To determine whether monotropein has an anticancer effect and explore its potential mechanisms against colorectal cancer (CRC) through network pharmacology and molecular docking combined with experimental verification.@*METHODS@#Network pharmacology and molecular docking were used to predict potential targets of monotropein against CRC. Cell counting kit assay, plate monoclonal assay and microscopic observation were used to investigate the antiproliferative effects of monotropein on CRC cells HCT116, HT29 and LoVo. Flow cytometry and scratch assay were used to analyze apoptosis and cell cycle, as well as cell migration, respectively in HCT116, HT29, and LoVo cells. Western blotting was used to detect the expression of proteins related to apoptosis, cell cycle, and cell migration, and the expression of proteins key to the Akt pathway.@*RESULTS@#The Gene Ontology and Reactome enrichment analyses indicated that the anticancer potential of monotropein against CRC might be involved in multiple cancer-related signaling pathways. Among these pathways, RAC-beta serine/threonine-protein kinase (Akt1, Akt2), cyclin-dependent kinase 6 (CDK6), matrix metalloproteinase-9 (MMP9), epidermal growth factor receptor (EGFR), cell division control protein 42 homolog (CDC42) were shown as the potential anticancer targets of monotropein against CRC. Molecular docking suggested that monotropein may interact with the 6 targets (Akt1, Akt2, CDK6, MMP9, EGFR, CDC42). Subsequently, cell activity of HCT116, HT29 and LoVo cell lines were significantly suppressed by monotropein (P<0.05). Furthermore, our research revealed that monotropein induced cell apoptosis by inhibiting Bcl-2 and increasing Bax, induced G1-S cycle arrest in colorectal cancer by decreasing the expressions of CyclinD1, CDK4 and CDK6, inhibited cell migration by suppressing the expressions of CDC42 and MMP9 (P<0.05), and might play an anticancer role through Akt signaling pathway.@*CONCLUSION@#Monotropein exerts its antitumor effects primarily by arresting the cell cycle, causing cell apoptosis, and inhibiting cell migration. This indicates a high potential for developing novel medication for treating CRC.
Subject(s)
Humans , Proto-Oncogene Proteins c-akt/metabolism , Cell Proliferation , Matrix Metalloproteinase 9 , Molecular Docking Simulation , Cell Cycle , ErbB Receptors , Apoptosis , Colorectal Neoplasms/pathology , Cell Line, Tumor摘要
OBJECTIVE@#To investigate the effect and possible mechanism of hydroxysafflor yellow A (HSYA) on human immortalized keratinocyte cell proliferation and migration.@*METHODS@#HaCaT cells were treated with HSYA. Cell proliferation was detected by the cell counting kit-8 assay, and cell migration was measured using wound healing assay and Transwell migration assay. The mRNA and protein expression levels of heparin-binding epidermal growth factor (EGF)-like growth factor (HBEGF), EGF receptor (EGFR), phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT), mammalian target of rapamycin (mTOR), and hypoxia-inducible factor-1α (HIF-1α) were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot, respectively. Circ_0084443-overexpressing HaCaT cells and empty plasmid HaCaT cells were constructed using the lentiviral stable transfection and treated with HSYA. The expression of circ_0084443 was detected by qRT-PCR.@*RESULTS@#HSYA (800 µmol/L) significantly promoted HaCaT cell proliferation and migration (P<0.05 or P<0.01). It also increased the mRNA and protein expression levels of HBEGF, EGFR, PI3K, AKT, mTOR and HIF-1α, and increased the phosphorylation levels of PI3K and AKT (P<0.05 or P<0.01). Furthermore, HSYA promoted HaCaT cell proliferation and migration via the HBEGF/EGFR and PI3K/AKT/mTOR signaling pathways (P<0.01). Circ_0084443 attenuated the mRNA expression levels of HBEGF, EGFR, PI3K, AKT, mTOR and HIF-1α (P<0.05). HSYA inhibited the circ_0084443 expression, further antagonized the inhibition of circ_0084443 on HBEGF, EGFR, PI3K, AKT, mTOR and HIF-1α, and promoted the proliferation of circ_0084443-overexpressing HaCaT cells (P<0.05 or P<0.01). However, HSYA could not influence the inhibitory effect of circ_0084443 on HaCaT cell migration (P>0.05).@*CONCLUSION@#HSYA played an accelerative role in HaCaT cell proliferation and migration, which may be attributable to activating HBEGF/EGFR and PI3K/AKT signaling pathways, and had a particular inhibitory effect on the keratinocyte negative regulator circ_0084443.
Subject(s)
Humans , Proto-Oncogene Proteins c-akt/metabolism , Phosphatidylinositol 3-Kinase , Phosphatidylinositol 3-Kinases/metabolism , ErbB Receptors/genetics , TOR Serine-Threonine Kinases/metabolism , Cell Proliferation , RNA, Messenger/genetics , Cell Movement , Cell Line, Tumor , Chalcone/analogs & derivatives , Quinones摘要
BACKGROUND@#Lung cancer is a major threat to human health. The molecular mechanisms related to the occurrence and development of lung cancer are complex and poorly known. Exploring molecular markers related to the development of lung cancer is helpful to improve the effect of early diagnosis and treatment. Long non-coding RNA (lncRNA) THAP7-AS1 is known to be highly expressed in gastric cancer, but has been less studied in other cancers. The aim of the study is to explore the role and mechanism of methyltransferase-like 3 (METTL3) mediated up-regulation of N6-methyladenosine (m6A) modified lncRNA THAP7-AS1 expression in promoting the development of lung cancer.@*METHODS@#Samples of 120 lung cancer and corresponding paracancerous tissues were collected. LncRNA microarrays were used to analyze differentially expressed lncRNAs. THAP7-AS1 levels were detected in lung cancer, adjacent normal tissues and lung cancer cell lines by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The diagnostic value of THAP7-AS1 in lung cancer and the relationship between THAP7-AS1 expression and survival rate and clinicopathological parameters were analyzed. Bioinformatics analysis, methylated RNA immunoprecipitation (meRIP), RNA pull-down and RNA-immunoprecipitation (RIP) assay were used to investigate the molecular regulation mechanism of THAP7-AS1. Cell proliferation, migration, invasion and tumorigenesis of SPC-A-1 and NCI-H1299 cells were determined by MTS, colony-formation, scratch, Transwell and xenotransplantation in vivo, respectively. Expression levels of phosphoinositide 3-kinase/protein kenase B (PI3K/AKT) signal pathway related protein were detected by Western blot.@*RESULTS@#Expression levels of THAP7-AS1 were higher in lung cancer tissues and cell lines (P<0.05). THAP7-AS1 has certain diagnostic value in lung cancer [area under the curve (AUC)=0.737], and its expression associated with overall survival rate, tumor size, tumor-node-metastasis (TNM) stage and lymph node metastasis (P<0.05). METTL3-mediated m6A modification enhanced THAP7-AS1 expression. The cell proliferation, migration, invasion and the volume and mass of transplanted tumor were all higher in the THAP7-AS1 group compared with the NC group and sh-NC group of SPC-A-1 and NCI-H1299 cells, while the cell proliferation, migration and invasion were lower in the sh-THAP7-AS1 group (P<0.05). THAP7-AS1 binds specifically to Cullin 4B (CUL4B). The cell proliferation, migration, invasion, and expression levels of phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), phosphoinositide-3 kinase, catalytic subunit delta (PIK3CD), phospho-phosphatidylinositol 3-kinase (p-PI3K), phospho-protein kinase B (p-AKT) and phospho-mammalian target of rapamycin (p-mTOR) were higher in the THAP7-AS1 group compared with the Vector group of SPC-A-1 and NCI-H1299 cells (P<0.05).@*CONCLUSIONS@#LncRNA THAP7-AS1 is stably expressed through m6A modification mediated by METTL3, and combines with CUL4B to activate PI3K/AKT signal pathway, which promotes the occurrence and development of lung cancer.
Subject(s)
Humans , Lung Neoplasms/pathology , RNA, Long Noncoding/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Up-Regulation , Proto-Oncogene Proteins c-akt/metabolism , Cell Line, Tumor , Cell Proliferation/genetics , Gene Expression Regulation, Neoplastic , Methyltransferases/metabolism , Cullin Proteins/genetics摘要
BACKGROUND@#Drug resistance is the main cause of high mortality of lung cancer. This study was conducted to investigate the effect of folic acid (FA) on the resistance of non-small cell lung cancer (NSCLC) cells to Osimertinib (OSM) by regulating the methylation of dual specificity phosphatase 1 (DUSP1).@*METHODS@#The OSM resistant NSCLC cell line PC9R was establishd by gradually escalation of OSM concentration in PC9 cells. PC9R cells were randomly grouped into Control group, OSM group (5 μmol/L OSM), FA group (600 nmol/L FA), methylation inhibitor decitabine (DAC) group (10 μmol/L DAC), FA+OSM group (600 nmol/L FA+5 μmol/L OSM), and FA+OSM+DAC group (600 nmol/L FA+5 μmol/L OSM+10 μmol/L DAC). CCK-8 method was applied to detect cell proliferation ability. Scratch test was applied to test the ability of cell migration. Transwell assay was applied to detect cell invasion ability. Flow cytometry was applied to measure and analyze the apoptosis rate of cells in each group. Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR) method was applied to detect the expression level of DUSP1 mRNA in cells. Methylation specific PCR (MSP) was applied to detect the methylation status of the DUSP1 promoter region in each group. Western blot was applied to analyze the expression levels of DUSP1 protein and key proteins in the DUSP1 downstream mitogen-activated protein kinase (MAPK) signaling pathway in each group.@*RESULTS@#Compared with the Control group, the cell OD450 values (48 h, 72 h), scratch healing rate, number of cell invasions, and expression of DUSP1 in the OSM group were obviously decreased (P<0.05); the apoptosis rate, the methylation level of DUSP1, the expression of p38 MAPK protein, and the phosphorylation level of extracellular regulated protein kinases (ERK) were obviously increased (P<0.05); the cell OD450 values (48, 72 h), scratch healing rate, number of cell invasions, and expression of DUSP1 in the DAC group were obviously increased (P<0.05); the apoptosis rate, the expression of p38 MAPK protein, the phosphorylation level of ERK, and the methylation level of DUSP1 were obviously reduced (P<0.05). Compared with the OSM group, the cell OD450 values (48, 72 h), scratch healing rate, number of cell invasions, and expression of DUSP1 in the FA+OSM group were obviously decreased (P<0.05); the apoptosis rate, the methylation level of DUSP1, the expression of p38 MAPK protein, and the phosphorylation level of ERK were obviously increased (P<0.05). Compared with the FA+OSM group, the cell OD450 values (48, 72 h), scratch healing rate, number of cell invasions, and expression of DUSP1 in the FA+OSM+DAC group were obviously increased; the apoptosis rate, the methylation level of DUSP1, the expression of p38 MAPK protein, and the phosphorylation level of ERK were obviously reduced (P<0.05).@*CONCLUSIONS@#FA may inhibit DUSP1 expression by enhancing DUSP1 methylation, regulate downstream MAPK signal pathway, then promote apoptosis, inhibit cell invasion and metastasis, and ultimately reduce OSM resistance in NSCLC cells.
Subject(s)
Humans , Carcinoma, Non-Small-Cell Lung/genetics , Lung Neoplasms/genetics , Dual Specificity Phosphatase 1/pharmacology , Cell Proliferation , p38 Mitogen-Activated Protein Kinases/pharmacology , Methylation , Apoptosis , Cell Line, Tumor摘要
SUMMARY: Keloid scar is a unique benign fibroproliferative tumor of the human skin. Previously, it was reported that early growth response 1 (EGR1), a transcription factor, promotes keloid fibrosis; however, the mechanism by which EGR1 modulates keloid formation was not elaborated. In this research, the specific function and the microRNA (miRNA) regulatory network of EGR1 in keloids was examined. Keloid fibroblasts (KFs) were transfected with EGR1-small interfering RNA (siEGR1), EGR1-overexpression plasmid (pcDNA3.1-EGR1), and microRNA (miR-183-5p)-mimics to regulate the expression of EGR1 and miR-183-5p. The study employed dual-luciferase reporter assays to explore the targeting regulation of miR-183-5p on EGR1. Additionally, Western blotting, flow cytometry, qRT-PCR, cell count kit-8 (CCK-8), transwell, and wound healing assays, and RNA sequencing were conducted. EGR1 was upregulated in KFs, and EGR1 silencing diminished proliferation, fibrosis, migration, invasion, and apoptosis of cells. In KFs, the expression of miR- 183-5p was reduced, leading to the inhibition of cell proliferation, migration, and invasion. Conversely, it enhanced apoptosis. By targeting EGR1, miR-183-5p partially counteracted the impact of EGR1 on migration, invasion, and fibrosis in KFs. The findings imply that miR-183-5p suppresses keloid formation by targeting EGR1. As a result, EGR1 holds promise as a potential therapeutic target for preventing and treating keloids.
La cicatriz queloide es un tumor fibroproliferativo benigno único de la piel humana. Anteriormente, se informó que la respuesta de crecimiento temprano 1 (EGR1), un factor de transcripción, promueve la fibrosis queloide; sin embargo, no se explicó el mecanismo por el cual EGR1 modula la formación de queloides. En esta investigación, se examinó la función específica y la red reguladora de microARN (miARN) de EGR1 en queloides. Se transfectaron fibroblastos queloides (KF) con ARN de interferencia pequeño de EGR1 (siEGR1), plásmido de sobreexpresión de EGR1 (pcDNA3.1-EGR1) y miméticos de microARN (miR-183-5p) para regular la expresión de EGR1 y miR-183. -5p. El estudio empleó ensayos de indicador de luciferasa dual para explorar la regulación dirigida de miR-183-5p en EGR1. Además, se realizaron pruebas de transferencia Western, citometría de flujo, qRT-PCR, kit de recuento celular-8 (CCK-8), transwell y curación de heridas, y secuenciación de ARN. EGR1 estaba regulado positivamente en KF, y el silenciamiento de EGR1 disminuyó la proliferación, fibrosis, migración, invasión y apoptosis de las células. En KF, la expresión de miR- 183-5p se redujo, lo que llevó a la inhibición de la proliferación, migración e invasión celular. Por el contrario, mejoró la apoptosis. Al apuntar a EGR1, miR-183-5p contrarrestó parcialmente el impacto de EGR1 en la migración, invasión y fibrosis en KF. Los hallazgos implican que miR-183-5p suprime la formación de queloides al apuntar a EGR1. Como resultado, EGR1 es prometedor como objetivo terapéutico potencial para prevenir y tratar los queloides.
Subject(s)
Humans , Male , Female , Adult , Middle Aged , Young Adult , Early Growth Response Protein 1 , Fibroblasts , Keloid/genetics , Keloid/pathology , Wound Healing , Transfection , Down-Regulation , Cell Movement , Blotting, Western , Sequence Analysis, RNA , Apoptosis , MicroRNAs/physiology , Cell Proliferation , Real-Time Polymerase Chain Reaction摘要
SUMMARY: Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that is highly expressed in various types of cancers including breast cancer. However, the role of AhR with its endogenous ligand 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) on the progression of breast cancer remains poorly understood. We aimed to investigate cell proliferation and migration states in breast cancer after activating AhR with the endogenous ligand ITE. Breast cancer tissue was evaluated by cell lines, immunohistochemistry, reverse transcription-polymerase chain reaction, cell proliferation, flow cytometry, migration assays and western blot techniques. We found that AhR was widely expressed in breast cancer tissues and metastasis lymph node tissues, but not in normal tissues. The expression AhR was independent between the age, grades and TNM classifications for breast cancer tissues. ITE treatment significantly induced the activation of AhR in a time-dependent manner in both MCF-7 and T47D breast cancer cell lines. Meanwhile, ITE did not affect the cell migration but significantly suppressed the cell proliferation in estrogen receptor positive (ER+) MCF-7 andT47D cells, which probably attribute to the induction of cell cycle arrest in G1 phase and shortened S phase. Further mechanism study showed that ERK1/2 and AKT signaling were required for the activation of AhR in MCF-7 cells. These data suggest that AhR is a potential new target for treating patients with breast cancer. ITE may be more potentially used for therapeutic intervention for breast cancer with the kind of ER(+).
El receptor de hidrocarburo de arilo (AhR) es un factor de transcripción activado por ligando que se expresa en gran medida en varios tipos de cáncer, incluido el cáncer de mama. Sin embargo, el papel de AhR con su ligando endógeno 2- (1'H-indol-3'-carbonil)-tiazol-4-ácido carboxílico metil éster (ITE) en la progresión del cáncer de mama sigue siendo poco conocido. Nuestro objetivo fue investigar la proliferación celular y los estados de migración en el cáncer de mama después de activar AhR con el ligando endógeno ITE. El tejido de cáncer de mama se evaluó mediante líneas celulares, inmunohistoquímica, reacción en cadena de la polimerasa con transcriptasa inversa, proliferación celular, citometría de flujo, ensayos de migración y técnicas de transferencia Western. Descubrimos que AhR se expresó ampliamente en tejidos de cáncer de mama y en linfonodos con metástasis, pero no en tejidos normales. La expresión AhR fue independiente entre la edad, grados y clasificaciones TNM para tejidos de cáncer de mama. El tratamiento con ITE indujo significativamente la activación de AhR de manera dependiente del tiempo en las líneas celulares de cancer de mama MCF-7 y T47D. Mientras tanto, ITE no afectó la migración celular, pero suprimió significativamente la proliferación celular en células MCF-7 y T47D con receptor de estrógeno positivo (ER+), lo que probablemente se atribuye a la inducción de la detención del ciclo celular en la fase G1 y la fase S acortada. Un estudio adicional del mecanismo mostró que las señales de ERK1/2 y AKT eran necesarias para la activación de AhR en las células MCF-7. Estos datos sugieren que AhR es un nuevo objetivo potencial para el tratamiento de pacientes con cáncer de mama. ITE puede ser utilizado más potencialmente en la intervención terapéutica para el cáncer de mama con el tipo de ER (+).
Subject(s)
Humans , Female , Thiazoles/administration & dosage , Breast Neoplasms/pathology , Receptors, Aryl Hydrocarbon/drug effects , Indoles/administration & dosage , Thiazoles/pharmacology , Immunohistochemistry , Receptors, Estrogen , Blotting, Western , Cytochrome P-450 CYP1A1/genetics , Cell Line, Tumor/drug effects , Cell Proliferation/drug effects , Cell Migration Assays , Cytochrome P-450 CYP1B1/genetics , Flow Cytometry , Indoles/pharmacology摘要
Os hemangiomas são neoplasias benignas dos tecidos moles. São lesões caracterizadas pela proliferação de células endoteliais. Objetivos: O objetivo deste trabalho é demonstrar por meio de uma revisão da literatura os principais hemangiomas de interesse odontológico e suas características clínicas e abordagens terapêuticas. Revisão de literatura: O estudo mostrou prevalência de até 6% na população geral, sendo o tumor mais comum na infância. Essas lesões apresentam rápido crescimento pós-natal, que pode levar meses a dois anos em média, mas geralmente após esse período ocorre sua involução. O diagnóstico é comumente baseado nas características clínicas e na história do paciente. É importante entender que o estudo histopatológico pode ser necessário nos casos em que o diagnóstico é incerto, para diferenciá-lo de outras neoplasias graves. Destacam-se as principais complicações relacionadas a lesões, ulcerações e hemorragias, além de infecções secundárias que podem causar alto índice de morbidade. Assim, é fundamental que o dentista reconheça essas patologias e tenha capacidade para tratá-las. Considerações finais: Foi possível observar que os hemangiomas são manifestações vasculares incomuns para o cirurgião-dentista, porém o profissional deve saber diagnosticá-los e tratá-los. Dentre as áreas acometidas, essas lesões são frequentes na cavidade oral e o tratamento consiste em acompanhamento com intervenções conservadoras(AU)
Hemangiomas are benign soft tissue neoplasms. These are lesions characterized by the proliferation of endothelial cells. Objectives: The objective of this work is to demonstrate through a literature review about the main hemangiomas of dental interest and clinical characteristics and therapeutic approaches. Literature review: The study showed a prevalence of up to 6% in the general population, being the most common tumor in childhood. These lesions presents a rapid postnatal growth, which may take months until two years on average, but usually after this period their involution occurs. The diagnosis is commonly based on clinical characteristics and patient's history. It is important to understand that the histopathological study may be necessary in cases that the diagnostic is uncertainty, in order to differentiate it from other serious neoplasms. Main complications related to injuries, ulceration and hemorrhage can be highlighted, in addition to secondary infections that can cause a high rate of morbidity. Thus, it is essential that dentists recognize such pathologies and be able to treat them. Final considerations: Was possible do observe that hemangiomas are uncommon vascular manifestations for the dental surgeon, however, the professional must know to diagnose and treat them. Among the affected areas, these lesions are frequently seen in the oral cavity and the treatment consists of following up with conservative interventions(AU)
Subject(s)
Soft Tissue Neoplasms , Mouth Neoplasms , Hemangioma , Hemangioma/diagnosis , Hemangioma/therapy , Cell Proliferation , Mouth/injuries , Neoplasms摘要
Objective: The objective of this study was to investigate the morphology, proliferation, and differentiation of gingival mesenchymal stem cells (GMSCs) irradiated with a 970 nm Diode Laser (LLLT). It is essential to validate the efficacy of treatment, optimize irradiation conditions and guarantee the safety and quality of stem cells for future use in dental applications. Materials and Methods: GMSCs were cultured in standard conditions and irradiated with a Diode laser (970 nm, 0.5W) with an energy density of 9J/cm2. Cell proliferation was assessed with the WST-1 proliferation kit. GMSCs were differentiated into chondrogenic and osteogenic lineages. Cell morphology was performed with Hematoxylin/eosin staining, and quantitative nuclear analysis was done. Cell viability was monitored with trypan blue testing. Results: GMSCs subjected to irradiation demonstrated a significant increase in proliferation at 72 hours compared to the non-irradiated controls (p=0.027). This indicates that the 970 nm diode laser has a stimulatory effect on the proliferation of GMSCs. LLLT-stimulated GMSCs exhibited the ability to differentiate into chondrogenic and osteogenic lineages. A substantial decrease in cell viability was observed 24 hours after irradiation (p=0.024). However, after 48 hours, the cell viability recovered without any significant differences. This indicates that there might be a temporary negative impact on cell viability immediately following irradiation, but the cells were able to recover and regain their viability over time. Conclusions: This study support that irradiation with a 970 nm diode laser could stimulate the proliferation of GMSCs, maintain their ability to differentiate into chondrogenic and osteogenic lineages, and has minimal impact on the mor- phological characteristics of the cells. These results support the potential use of NIR Lasers in combination with GMSCs as a promising strategy for dental treatments.
Objetivo: El objetivo de este estudio fue investigar la morfología, proliferación y diferenciación de las células madre mesenquimatosas (GMSC) irradiadas con un láser de diodo de 970 nm (LLLT). Es fundamental validar la eficacia del tratamiento, optimizar las condiciones de irradiación y garantizar la seguridad y calidad de las células madre para su uso futuro en aplicaciones dentales.Materiales y Métodos: Las GMSC se cultivaron en condiciones estándar y se irradiaron con un láser de diodo (970 nm, 0,5 W) con una densidad de energía de 9 J/cm2. La proliferación celular se evaluó con el kit de proliferación WST-1. Las GMSC se diferenciaron en linajes condrogénicos y osteogénicos. La morfología celular se realizó con tinción de hematoxilina/eosina y se realizó un análisis nuclear cuantitativo. La viabilidad celular se controló con prueba de azul de tripano. Resultados: Las GMSC sometidas a irradiación demostraron un aumento significativo en la proliferación a las 72 horas en comparación con los controles no irradiados (p=0,027). Esto indica que el láser de diodo de 970 nm tiene un efecto estimulante sobre la proliferación de GMSC. Las GMSC estimuladas con LLLT exhibieron la capacidad de diferenciarse en linajes condrogénicos y osteogénicos. Se observó una disminución sustancial de la viabilidad celular 24 horas después de la irradiación (p=0,024). Sin embargo, después de 48 horas, la viabilidad celular se recuperó sin diferencias significativas. Esto indica que podría haber un impacto negativo temporal en la viabilidad de las células inmediatamente después de la irradiación, pero las células pudieron recuperarse y recuperar su viabilidad con el tiempo. Conclusión: En conclusión, este estudio respalda que la irradiación con un láser de diodo de 970 nm podría estimular la proliferación de GMSC, mantener su capacidad para diferenciarse en linajes condrogénicos y osteogénicos y tiene un impacto mínimo en las características morfológicas de las células. Estos resultados respaldan el uso potencial de láseres NIR en combinación con GMSC como una estrategia prometedora para tratamientos dentales.
Subject(s)
Humans , Low-Level Light Therapy , Cell Proliferation/radiation effects , Lasers, Semiconductor , Mesenchymal Stem Cells/radiation effects , In Vitro Techniques , Gingiva/radiation effects摘要
Introdução: Os cistos e tumores odontogênicos são lesões que apresentam comportamento biológico heterogêneo e patogênese ainda não totalmente esclarecida. A Yes-associated protein (YAP) atua como um regulador transcricional de genes envolvidos na proliferação celular e na apoptose, participando da ativação de vias associadas ao crescimento cístico e à progressão neoplásica. Objetivo: Analisar a expressão imuno-histoquímica da proteína YAP e correlacioná-la com marcadores envolvidos na proliferação celular e na apoptose em lesões odontogênicas epiteliais benignas. Metodologia: A amostra consistiu de 95 casos de lesões odontogênicas - 25 cistos dentígeros (CDs), 30 CO não sindrômicos (COs), 30 AMB convencionais (AMB-Cs) e 10 AMB unicísticos (AMB-Us) -, além de 10 espécimes de folículo dentários (FD). Foi realizada coleta dos dados clinico-demográficos dos casos, bem como análise morfológica para melhor caracterização da amostra. Os cortes histológicos foram submetidos à técnica imuno-histoquímica através da utilização dos anticorpos YAP, ciclina D1, Ki-67 e Bcl-2, e a análise da expressão destes foi realizada quali-quantitativamente, mediante metodologia adaptada. Os dados coletados seguiram para análise descritiva e estatística (p ≤ 0,05). Resultados: Houve discreta predileção por mulheres (n = 55; 57,6%) e por indivíduos na faixa etária dos 21 aos 40 anos (n = 50; 47,6%), sendo a região posterior de mandíbula mais afetada (64%). A análise da imunoexpressão de YAP revelou maiores níveis de expressão em COs, especialmente nas camadas basal e parabasal, seguido dos AMB-Us e AMB-Cs, que demonstraram moderada imunorreatividade, predominantemente nas células periféricas. Além disso, houve diferenças significativas quanto à imunoexpressão de YAP entre os grupos analisados, com existência de correlações positivas e estatisticamente significativas entre YAP e ciclina D1 em CDs e AMB-Us, e entre YAP e Ki-67 em AMB-Us (p < 0,05). Todavia, entre a imunoexpressão YAP e Bcl-2, foi verificada ausência de correlação estatisticamente significativa. Conclusões: A YAP pode exercer influência sobre a proliferação celular do epitélio de cistos e tumores odontogênicos, auxiliando, assim, na progressão das diferentes lesões odontogênicas (AU).
Background: Odontogenic cysts and tumors present heterogeneous biological behavior, and their etiopathogenesis is not fully understood yet. Yes-associated protein (YAP) acts as a transcriptional regulator of genes involved in cell proliferation and apoptosis, activating pathways associated with cystic growth and neoplastic progression. Objective: To analyze the immunohistochemical expression of YAP protein and correlate it with markers involved in cell proliferation and apoptosis in benign epithelial odontogenic lesions. Methods: The sample consisted of 95 cases of odontogenic lesions - 25 dentigerous cysts (DCs), 30 non-syndromic odontogenic keratocyst (OKCs), 30 conventional AMB (C-AMBs), and 10 unicystic AMB (UAMBs) -, in addition to 10 specimens of dental follicles (DF). Clinicodemographic data collection was carried out, as well as morphological analysis for better characterization of the sample. The histological sections were submitted to the immunohistochemical technique using YAP, cyclin D1, Ki-67, and Bcl-2 antibodies, and their immunoexpression analysis was performed qualitatively and quantitatively, through an adapted methodology. The collected data were submitted for descriptive and statistical analysis (p ≤ 0.05). Results: There was a slight predilection for women (n = 55; 57.6%) and individuals aged between 21 and 40 years (n = 50; 47.6%), with the posterior region of the mandible as the most affected site (64%). Analysis of YAP immunoexpression revealed higher expression levels in OKCs, especially in the basal and parabasal layers, followed by U-AMBs and C-AMBs, which showed moderate immunoreactivity, predominantly in peripheral cells. In addition, there were significant differences in YAP immunoexpression between the analyzed groups, with positive and statistically significant correlations between YAP and cyclin D1 in DCs and U-AMBs, and between YAP and Ki-67 in U-AMBs (p < 0.05). However, between YAP and Bcl-2 immunoexpression, there was no statistically significant correlation. Conclusions: YAP may influence on the cell proliferation of odontogenic cysts and tumors epithelium, thus helping with the progression of the different odontogenic lesions (AU) .
Subject(s)
Cell Proliferation , YAP-Signaling Proteins/metabolism , Transcriptional Coactivator with PDZ-Binding Motif Proteins/metabolism , Dentigerous Cyst/pathology , Biomarkers, Tumor , Medical Records , Retrospective Studies , Data Interpretation, Statistical , Apoptosis , Odontogenic Cyst, Calcifying/pathology , Statistics, Nonparametric , Inhibitor of Differentiation Proteins , Observational Study , Morphological and Microscopic Findings摘要
SUMMARY: Gastrin plays a vital role in the development and progression of gastric cancer (GC). Its expression is up-regulated in GC tissues and several GC cell lines. Yet, the underlying mechanism remains to be investigated. Here, we aim to investigate the role and mechanism of gastrin in GC proliferation. Gastrin-overexpressing GC cell model was constructed using SGC7901 cells. Then the differentially expressed proteins were identified by iTRAQ analysis. Next, we use flow cytometry and immunofluorescence to study the effect of gastrin on the mitochondrial potential and mitochondria-derived ROS production. Finally, we studied the underlying mechanism of gastrin regulating mitochondrial function using Co-IP, mass spectrometry and immunofluorescence. Overexpression of gastrin promoted GC cell proliferation in vitro and in vivo. A total of 173 proteins were expressed differently between the controls and gastrin- overexpression cells and most of these proteins were involved in tumorigenesis and cell proliferation. Among them, Cox17, Cox5B and ATP5J that were all localized to the mitochondrial respiratory chain were down-regulated in gastrin-overexpression cells. Furthermore, gastrin overexpression led to mitochondrial potential decrease and mitochondria-derived ROS increase. Additionally, gastrin-induced ROS generation resulted in the inhibition of cell apoptosis via activating NF-kB, inhibiting Bax expression and promoting Bcl-2 expression. Finally, we found gastrin interacted with mitochondrial membrane protein Annexin A2 using Co-IP and mass spectrometry. Overexpr ession of gastrin inhibits GC cell apoptosis by inducing mitochondrial dysfunction through interacting with mitochondrial protein Annexin A2, then up-regulating ROS production to activate NF-kB and further leading to Bax/Bcl-2 ratio decrease.
La gastrina juega un papel vital en el desarrollo y progresión del cáncer gástrico (CG). Su expresión está regulada al alza en tejidos de CG y en varias líneas celulares de CG. Sin embargo, el mecanismo subyacente aun no se ha investigado. El objetivo de este estudio fue investigar el papel y el mecanismo de la gastrina en la proliferación de CG. El modelo de células CG que sobre expresan gastrina se construyó usando células SGC7901. Luego, las proteínas expresadas diferencialmente se identificaron mediante análisis iTRAQ. A continuación, utilizamos la citometría de flujo y la inmunofluorescencia para estudiar el efecto de la gastrina en el potencial mitocondrial y la producción de ROS derivada de las mitocondrias. Finalmente, estudiamos el mecanismo subyacente de la gastrina que regula la función mitocondrial utilizando Co-IP, espectrometría de masas e inmunofluorescencia. La sobreexpresión de gastrina promovió la proliferación de células CG in vitro e in vivo. Un total de 173 proteínas se expresaron de manera diferente entre los controles y las células con sobreexpresión de gastrina y la mayoría de estas proteínas estaban implicadas en la tumorigenesis y la proliferación celular. Entre estas, Cox17, Cox5B y ATP5J, todas localizadas en la cadena respiratoria mitocondrial, estaban reguladas a la baja en las células con sobreexpresión de gastrina. Además, la sobreexpresión de gastrina provocó una disminución del potencial mitocondrial y un aumento de las ROS derivadas de las mitocondrias. Por otra parte, la generación de ROS inducida por gastrina resultó en la inhibición de la apoptosis celular mediante la activación de NF-kB, inhibiendo la expresión de Bax y promoviendo la expresión de Bcl-2. Finalmente, encontramos que la gastrina interactuaba con la proteína de membrana mitocondrial Anexina A2 usando Co-IP y espectrometría de masas. La sobreexpresión de gastrina inhibe la apoptosis de las células CG al inducir la disfunción mitocondrial a través de la interacción con la proteína mitocondrial Anexina A2, luego regula el aumento de la producción de ROS para activar NF-kB y conduce aún más a la disminución de la relación Bax/Bcl-2.