ABSTRACT
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 ReactionABSTRACT
Intracellular trafficking is a field that has been intensively studied for years and yet there remains much to be learned. Part ofthe reason that there is so much obscurity remaining in this field is due to all the pathways and the stages that define cellulartrafficking. One of the major steps in cellular trafficking is fusion. Fusion is defined as the terminal step that occurs when acargo-laden vesicle arrives at the proper destination. There are two types of fusion within a cell: homotypic and heterotypicfusion. Homotypic fusion occurs when the two membranes merging together are of the same type such as vacuole tovacuole fusion. Heterotypic fusion occurs when the two membranes at play are of different types such as when anendosomal membrane fuses with a Golgi membrane. In this review, we will focus on all the protein components – Rabs,Golgins, Multisubunit tethers, GTPases, protein phosphatases and SNAREs – that have been known to function in both ofthese types of fusion. We hope to develop a model of how all of these constituents function together to achieve membranefusion. Membrane fusion is a biological process absolutely necessary for proper intracellular trafficking. Due to the degreeof importance multiple proteins are required for it to be properly carried through. Whether we are talking about heterotypicor homotypic fusion, any defects in the fusion machinery can result in disease states such as Parkinson’s and Alzheimer’sdisease. Although much research has significantly expanded our knowledge of fusion, there is still much more to belearned.
ABSTRACT
Objective To study the expression and clinical significance of miR-183-5p, TβRⅠ and TβRⅡ in esophageal squamous cell carcinoma (ESCC). Methods The mRNA and protein expression of miR-183-5p, TβRⅠ and TβRⅡ were examined in ESCC cell lines ECA-109, TE-1, normal esophageal epithelial cells, tumor tissues and tumor-free tissues from 72 ESCC patients. Their clinical significance and the relationship between miR-183-5p and the latter two were analyzed. The effects of miR-183-5p on the expression of TβRⅠand TβRⅡ in ECA-109 cells and the cell functions of ECA-109 were also investigated. Results Compared with the normal esophageal epithelia cells, ESCC cell lines TE-1 and ECA-109 were statistically characterized by a high expression of miR-183-5p (all P<0.05) and low expression of TβRⅠand TβRⅡ(all P<0.05). The expression of miR-183-5p in ESCC tissues was higher than that in adjacent normal tissues, while the expressions of TβRⅠ and TβRⅡ were lower (all P< 0.05). The expression of miR-183-5p was closely related to sex, tumor differentiation, tumor staging, distant metastasis, lymphatic metastasis, and tumor location (all P<0.05). TβRⅠlevel was associated with sex, lymph node metastasis and tumor size (all P<0.05). Experimental data showed the negative correlation between the expression of miR-183-5p and TβRⅠin ESCC tissues (r= -0.521, P< 0.05). Over expression of miR-183-5p significantly inhibited the expression of TβRⅠ in ECA-109 cells (P< 0.05) and promoted the growth, invasion and metastasis of ECA-109 cells (P< 0.05). Low expression of miR-183-5p significantly promoted the expression of TβRⅠ in ECA-109 cells (P< 0.05), and suppressed the growth, invasion and metastasis of ECA-109 cells (P< 0.05). There was no significant change in the expression of TβRⅡ in the transfection experiments. Conclusion MiR-183-5p is closely related to the abnormal expression of TβRⅠ, which may exert an important role in the progression of lymphatic metastasis.