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Resumo: O Acidente Vascular Cerebral (AVC) é caracterizado como um déficit neurológico atribuído a uma lesão aguda do sistema nervoso central por uma causa vascular. Constitui-se uma das principais causas de mortes, acometendo principalmente a população idosa e, diante das incapacidades impostas pelo AVC, surge a necessidade de cuidados, que são frequentemente realizados pelo cuidador familiar. Este estudo teve como objetivo construir uma tecnologia educacional direcionada aos cuidadores familiares de pacientes idosos pós-Acidente Vascular Cerebral, dependentes de cuidados básicos para as atividades de vida diária, isto é, uma tecnologia em formato de vídeo, constituindo o produto desta pesquisa. Trata-se de estudo metodológico, realizado em uma unidade de internamento de pacientes neurológicos em um hospital universitário de grande porte localizado na região Sul do país. A amostra foi composta por 14 juizes especialistas em neurologia e/ou gerontologia do território nacional e 15 cuidadores familiares de idosos. A coleta de dados ocorreu no período de dezembro a fevereiro de 2024. O estudo foi composto por três etapas: 1) Pré-produção: construção da sinopse, argumento, roteiro e storyboard do vídeo educacional, para essa etapa foi realizada uma revisão integrativa, visando fundamentação teórica para o desenvolvimento do roteiro para tecnologia educacional com posterior validação do conteúdo do vídeo por juízes especialistas, por meio do Instrumento de Validação de Conteúdo Educacional em Saúde (IVCES). Para análise do instrumento foi utilizado o Índice de Validade de Conteúdo (IVC) e o valor considerado aceitável quando o resultado da proporção de concordância entre os juízes especialistas foi igual ou superior a 0,78; 2) Produção: construção do vídeo educacional e avaliação do vídeo pelos cuidadores familiares dos idosos internados na Neurologia, por meio do instrumento Suitability Assessment of Materials (SAM), com avaliação do conteúdo, exigência de alfabetização, ilustrações, layout e apresentação e adequação cultural; 3) Pós-produção: edição e versão final do vídeo educacional para posterior divulgação. Como resultado da pesquisa, identificado na literatura quais orientações os cuidadores e/ou familiares de pessoa idosas que sofreram AVC deveriam receber sobre os cuidados pós-AVC no preparo para a alta hospitalar. Realizada produção do roteiro do vídeo, que foi validado pela concordância dos juízes especialistas (IVC= 0,93). O vídeo foi composto por 51 cenas em resolução Full HD, com duração de 8 minutos. Houve 99% de concordância entre os cuidadores familiares de idosos participantes, tornando-o adequado e validado. Após finalização dessa etapa, foi preenchido o requerimento para emissão do certificado de produto brasileiro do vídeo educacional na Agência Nacional do Cinema, divulgado em plataforma digital YouTube. O vídeo demonstrou-se uma ferramenta que auxiliou na aquisição de conhecimentos fundamentais sobre o tema, uma vez que é uma forma de disseminar informações de forma atraente, simples, dinâmica e democrática. O produto desenvolvido tem a potencialidade de ser replicado em outras unidades da instituição que atendam ao mesmo perfil de pacientes, uma vez que a tecnologia proporciona orientações e esclarecimentos aos cuidadores familiares de idosos sobre o tema.
Abstract: Cerebral Vascular Accident (CVA) is characterized as a neurological deficit attributed to an acute injury of the central nervous system due to a vascular cause. It is one of the main causes of death, affecting mainly the elderly population and, given the disabilities imposed by stroke, the need for care arises, which is often carried out by the family caregiver. This study aimed to build an educational technology aimed at family caregivers of elderly post-stroke patients, dependent on basic care for activities of daily living, that is, a technology in video format, constituting the product of this research. This is a methodological study, carried out in an inpatient unit for neurological patients in a large university hospital located in the southern region of the country. The sample was made up of 14 judges specializing in neurology and/or gerontology from the national territory and 15 family caregivers of the elderly. Data collection took place from December to February 2024. The study consisted of three stages: 1) Preproduction: construction of the synopsis, argument, script and storyboard of the educational video, for this stage an integrative review was carried out, aiming at theoretical foundation for the development of the script for educational technology with subsequent validation of the video content by expert judges, through the Health Educational Content Validation Instrument (IVCES). To analyze the instrument, the Content Validity Index (CVI) was used and the value was considered acceptable when the result of the proportion of agreement between the expert judges was equal to or greater than 0.78; 2) Production: construction of the educational video and evaluation of the video by family caregivers of elderly people hospitalized in Neurology, using the Suitability Assessment of Materials (SAM) instrument, with content evaluation, literacy requirements, illustrations, layout and presentation and cultural suitability; 3) Postproduction: editing and final version of the educational video for subsequent dissemination. As a result of the research, the literature identified what guidance caregivers and/or family members of elderly people who suffered a stroke should receive regarding post-stroke care in preparation for hospital discharge. The video script was produced, which was validated by the agreement of the expert judges (IVC= 0.93). The video was composed of 51 scenes in Full HD resolution, lasting 8 minutes. There was 99% agreement among family caregivers of participating elderly people, making it appropriate and validated. After completing this stage, the application was completed to issue the Brazilian product certificate for the educational video at the National Cinema Agency, published on the YouTube digital platform. The video proved to be a tool that helped in the acquisition of fundamental knowledge on the topic, as it is a way of disseminating information in an attractive, simple, dynamic and democratic way. The developed product has the potential to be replicated in other units of the institution that serve the same patient profile, as the technology provides guidance and clarification to family caregivers of elderly people on the topic.
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Humans , Male , Female , Adult , Middle Aged , Patient Discharge , Aged , Caregivers , Stroke , Patient Care , Hospital to Home TransitionABSTRACT
ABSTRACT Background: Triple-negative breast cancer (TNBC) is a subtype of breast cancer (BC) that lacks receptors for targeted therapy. Deeper insight into the molecular mechanisms regulating TNBC metastasis is urgently needed. The epithelial-mesenchymal transition process facilitates the metastasis of neighboring epithelial tumor cells. Protein kinase, membrane-associated tyrosine/threonine 1 (PKMYT1), a member of the Wee family of protein kinases, is upregulated in BC, and its high expression predicts poor prognosis in BC patients. Notch signaling activation is a pathognomonic feature of TNBC. PKMYT1 has been found to induce EMT in non-small cell lung cancer by activating Notch signaling. However, whether PKMYT1 exerts effects on TNBC progression by regulating Notch signaling remains unknown. Objectives: The objective of this study was to investigate whether PKMYT1 exerts effects on TNBC progression by regulating Notch signaling. Methods: Fifty cases of surgically resected BC samples (tumor and adjacent non-tumor tissue samples) were collected from patients diagnosed with BC. We measured the expression of PKMYT1 in clinical samples with real-time quantitative polymerase chain reaction (RT-qPCR). For in vitro analysis, RT-qPCR and Western blotting were conducted to evaluate PKMYT1 expression in TNBC cells. Then, the viability, migration, and invasion of TNBC cells were detected by cell counting kit-8 assays, wound healing assays, and Transwell assays. The EMT event was examined by evaluating the levels of EMT-associated proteins. For in vivo analysis, xenograft models in nude mice were established to explore PKMYT1 roles. E-cadherin and Ki67 expression in xenograft models were estimated by immunohistochemistry staining. Hematoxylin and eosin staining was performed to assess tumor metastasis. The underlying mechanisms by which PKMYT1 affected the malignant phenotypes of TNBC cells were explored by Western blotting measuring the pathway-associated proteins. Results: PKMYT1 was upregulated in BC tissues and cells, and its knockdown prevented cell proliferation, migration, invasion, and EMT event in TNBC. Mechanistically, Notch signaling was inactivated by PKMYT1 depletion, and Notch activation abolished the PKMYT1 silencing-induced inhibition in the malignant phenotypes of TNBC cells. For in vivo analysis, PKMYT1 knockdown inhibited tumorigenesis and metastasis of TNBC. Conclusion: PKMYT1 promotes EMT, proliferation, migration, and invasion of TNBC cells and facilitates tumor growth and metastasis by activating Notch signaling.
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Objective @#To investigate the role and mechanism of circular RNA containing pleck strin homology do main of Pleck strin homology domain family M member 3 ( circRNA_PLEKHM3) in regulating epithelial mesenchy mal transition (EMT) behavior in cervical cancer cells through the miR-320 and KLF4 .@*Methods @#The expression levels of circRNA_PLEKHM3 in cervical cancer cells Hela and CaSki were detected by real time quantitative PCR (qRT PCR) . RNA fluorescence in situ hybridization was used to determine the localization of circRNA_PLEKHM3 in human cervical cancer epithelial cells CaSki . Dual luciferase reporter gene experiments were conducted to inves tigate the targeting relationship between circRNA_PLEKHM3 and miR-320 , as well as the targeting relationship be tween miR-320 and KLF4 . CaSki cells were overexpressed with circRNA_PLEKHM3. Additionally, three groups were set up: overexpression of miR-320 on the basis of circRNA_PLEKHM3 overexpression , silencing of KLF4 on the basis of circRNA_PLEKHM3 overexpression , and silencing of KLF4 on the basis of miR-320 overexpression . qRT PCR was performed to detect the expression levels of miR-320 in CaSki . Western blot experiments were con ducted to determine the expression of KLF4 and EMT markers including E-cadherin , N-cadherin , Vimentin , MMP-2 , and MMP-9 in CaSki cells . Transwell assays were performed to measure cell migration and invasion .@*Results@#The expression of circRNA_PLEKHM3 decreased in Hela and CaSki cells (P < 0 05) , mainly localized in the cytoplasm . The dual luciferase reporter gene experiment demonstrated a targeting relationship between miR-320 and circRNA_PLEKHM3 , as well as between KLF4 and miR-320 . Overexpression of circRNA_PLEKHM3 inhibited the protein expression of miR-320 , Ncadherin , Vimentin , MMP-2 , and MMP-9 , up regulated the protein expression of E-cadherin, and reduced cell migration and invasion ( P < 0.05) . Overexpression of miR 320 or silencing of KLF4 on the basis of circRNA_PLEKHM3 overexpression both promoted the protein expression of miR-320, N-cad herin , Vimentin , MMP-2 , and MMP-9 , down regulated the protein expression of E-cadherin , and increased cell migration and invasion (P < 0.05) . However , silencing of KLF4 on the basis of miR-320 overexpression inhibited the protein expression of KLF4 , N-cadherin , Vimentin , MMP-2 , and MMP-9 , up regulated the protein expression of E-cadherin , and reduced cell migration and invasion ( P < 0.05) . @*Conclusion @#Overexpression of circRNA _PLEKHM3 regulates EMT in cervical cancer cells through the miR -320/KLF4 axis .
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The development of animal early embryos commences with the reprogramming of terminally differentiated gametes into totipotent zygotes following fertilization. During the initial stages of embryonic development, the transcriptional levels of zygotic genome remain silent and maternal gene products dominate the regulation of development. As embryonic development progresses, the maternal gene products undergo phased degradation while the zygotic genome gradually activates transcription, marking the transition from the maternal regulation to the zygotic genome regulation in early embryonic development, which is also referred to as the maternal-zygotic transition (MZT). Zygotic genome activation (ZGA) is a critical turning process in this transition, and its accurate occurrence is crucial for early embryonic development and cell fate decisions. However, the regulatory factors and molecular mechanisms of ZGA remain poorly understood. Studies have shown that ZGA varies greatly among different species and may be affected by a variety of regulatory factors such as DNA methylation, histone modification, non-coding RNA, chromatin remodeling and ZGA related factors. Here, we review the research progress of the above regulatory factors affecting ZGA, which can provide valuable insights for further investigations into the ZGA related mechanisms of early embryos.
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Objective:To explore the application of a role transition shock model based on the teach-back technique in knee arthroplasty nursing teaching.Methods:We assigned 50 nursing student interns practicing in the knee arthroplasty team of Orthopedics Department of Nanjing First Hospital between August 2020 and August 2022 into control group ( n=25, traditional teaching) and observation group ( n=25, teach-back-based role transition shock model teaching) according to the order of admission. At the end of internship, the examination scores, the impact of transition shock on comprehensive abilities, and teaching satisfaction of the students were assessed and analyzed using the t test and Fisher's exact test with the use of SPSS 22.0. Results:Compared with the control group, the observation group scored significantly lower in the physical, psychological, knowledge and skills, and sociocultural and developmental dimensions of the transition shock assessment scale ( P<0.05). The observation group showed significantly higher scores of nurse-patient communication, nursing practice, disease observation, health education, humanistic care, team cooperation, clinical thinking, and emergency response than the control group ( P<0.05). The examination results of the observation group were significantly better than those of the control group ( t=12.31, 11.52, P<0.001). The teaching satisfaction rate of the observation group was significantly higher than that of the control group [100.00% (25/25) vs. 68.00% (17/25), χ2=9.52, P=0.002]. Conclusions:The teach-back-based role transition shock model can help alleviate the transition impact faced by nursing student interns when entering clinical practice, and also improve their comprehensive abilities as well as satisfaction with teaching.
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Tumor metastasis is closely related to high mortality rate of cancer.It is well known that glutamine plays an important role in the malignant progression of cancer.Notably,as an important carbon and nitrogen donor,glutamine has been found to be closely related to tumor metastasis in recent years.Glutamine is not only involved in regulating the proliferation of tumor cells,but is also closely related to the migration and invasion of tumor cells.Furthermore,various enzymes along with transporters in the metabolism of glutamine are involved in the process of tumor metastasis through different signaling pathways.This review provided a summary of the role of glutamine in tumor metastasis in recent years and proposed therapeutic targets to provide new strategies for the clinical treatment of tumor metastases.
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AIM:One of the important characteristics of the occurrence and development of triple-negative breast cancer(TNBC)is dysregulated cell metabolism.The aim of this study is to investigate the mechanism of pyruvate dehydrogenase E1 subunit alpha 1(PDHA1),a key enzyme component in aerobic glycolysis,affecting the proliferation,metastasis and invasion of TNBC.METHODS:(1)The expression levels of PDHA1 in breast cancer tissues and adja-cent tissues were analyzed by UALCAN database,KM-plotter database,Gene MANIA database and TCGA database.The expression of PDHA1 was compared according to tumor pathological stage,subtype classification and breast cancer bio-markers.The function of PDHA1 in TNBC was explored by gene enrichment analysis.(2)Immunohistochemistry assays were used to detect the expression of PDHA1 in human TNBC tissue and adjacent tissue samples.(3)Stable PDHA1 knockout and PDHA1 rescue TNBC MDA-MB-231 cells were constructed.The proliferation of MDA-MB-231 cells was de-tected by colony formation assay and cell counting assay.The regulatory effect of PDHA1 on the invasion and migration of MDA-MB-231 cells was detected by in vitro scratch assay and Transwell migration assay.RESULTS:Database analysis showed that the group with high PDHA1 expression in breast cancer had shorter survival and worse prognosis.In clinical specimens,the expression of PDHA1 in cancer tissues was higher than that in adjacent normal tissues.Knockout of PDHA1 inhibited the proliferation,metastasis,invasion and epithelial-mesenchymal transition of MDA-MB-231 cells.CONCLUSION:PDHA1 is overexpressed in TNBC,and it promotes cell proliferation and facilitates TNBC metastasis through the epithelial-mesenchymal transition pathway.
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Objective:To analyze the prognostic value of MET, Cyclin D1, and MET gene copy number (GCN) for non-small cell lung cancer (NSCLC).Methods:This study included 61 patients with NSCLC who received treatment at the Enze Hospital, Taizhou Enze Medical Center (Group) between January 2018 and June 2019. The expression levels of MET and Cyclin D1 were determined using immunohistochemistry. MET GCN was evaluated using a quantitative polymerase chain reaction. Clinicopathological characteristics were compared among patients with different expression levels of these proteins. The Spearman correlation coefficient was used to assess the relationship between MET, Cyclin D1, and MET GCN. The Kaplan-Meier method was used to analyze survival rates. Univariate and multivariate Cox regression analyses were conducted to investigate the correlation between MET, Cyclin D1, and MET GCN and survival rates.Results:Thirty-six cases (59.02%) tested positive for MET, which was mainly expressed in the cytoplasm and membrane. Similarly, 36 cases (59.02%) were positive for Cyclin D1, which was mainly expressed in the cytoplasm. Patients with MET ( χ2 = 6.89, P = 0.009) and MET/Cyclin D1 ( χ2 = 4.05, P = 0.004) had a high proportion of poorly differentiated histology. Moreover, patients with MET GCN ≥ 3 had a relatively high proportion of lymph node metastasis ( χ2 = 8.11, P = 0.004) and TNM stages III-IV ( χ2 = 3.91, P = 0.048). Furthermore, patients with MET GCN ≥ 3/Cyclin D1 also had a high proportion of lymph node metastasis ( χ2 = 6.73, P = 0.009). MET was significantly associated with MET GCN ( r = 0.39, P = 0.002) and Cyclin D1 ( r = 0.39, P = 0.002), while MET GCN was significantly associated with Cyclin D1 ( r = 0.30, P = 0.017). The median survival time of patients with and without MET was 24.0 and 32.5 months, respectively, while the median survival time of patients with MET GCN ≥ 3 and < 3 was 11.0 and 30.5 months, respectively. Multivariate analysis showed that TNM stages III-IV, positive expression of MET, and MET GCN ≥ 3 were significantly associated with a high risk of death. Conclusion:The positive expression of MET and MET GCN ≥ 3 may be adverse prognostic factors in patients with NSCLC. The activation of the MET/Cyclin D1 signaling pathway could potentially contribute to the development and progression of NSCLC.
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Objective To investigate the effects of circDNMT1 on the proliferation,apoptosis and epithelial-mesenchymal transition(EMT)of triple-negative breast cancer(TNBC)cells by regulating miR-377-3p/PUM1 axis.Methods The TNBC tissues and adjacent normal tissues of 24 patients with TNBC treated in Danzhou People's Hospital from 2018 to 2021 were collected.qRT-PCR and Western blot were used to detect the expression of circDNMT1,miR-377-3p,and PUM1 protein in tissue and mouse normal breast epithelial cell line HC11 and TNBC cell lines 4T1,Eph41424,and JC.4T1 cells were divided into the 4T1 group(untransfected),the si-NC group(transfected with si-NC),the si-DNMT1 group(transfected with si-DNMT1),the si-DNMT1+anti-NC group(simultaneously transfected with si-DNMT1 and anti-NC),and the si-DNMT1+anti-miR-377-3p group(simultaneously transfected with si-DNMT1 and anti-miR-377-3p).qRT-PCR was used to detect the expression of circDNMT1 and miR-377-3p of 4T1 cells in each group;CCK-8 was used to detect the proliferation of 4T1 cells in each group;flow cytometry was used to detect the apoptosis of 4T1 cells in each group;Western blot was used to detect the expression of PUM1,EMT-related proteins and apoptosis-related proteins of 4T1 cells in each group;TargetScan website was used to predict the binding sites of miR-377-3p with circDNMT1 and PUM1;dual luciferase report was used to verify the targeting relationships of miR-377-3p with circDNMT1 and PUM1.After inoculation with 4T1 cells,BALB/c mice were randomly divided into the blank control group(injected with equal amount of normal saline),the negative control group(injected with si-NC via tail vein),the DNMT1 silencing group(injected with si-DNMT1 via tail vein),the combined control group(injected with si-DNMT1 and anti-NC via tail vein),and the combined silencing group(injected with si-DNMT1 and anti-miR-377-3p via tail vein),the tumor massess of mice were recorded and the morphological changes of tumors were observed by hematoxylin-eosin staining.Results circDNMT1 and PUM1 were up-regulated in TNBC tissues and cells,and miR-377-3p was down-regulated.The expression difference was most obvious in 4T1 cells,so 4T1 cells were selected for subsequent experiments.Compared with the 4T1 group and the si-NC group,the expression of miR-377-3p,the apoptosis rate of 4T1 cells,the expression levels of Bax,cleaved caspase-3 and E-cadherin protein of 4T1 cells in the si-DNMT1 group were significantly increased(P<0.05),the circ-DNMT1 level,the expression level of PUM1 protein,OD values at 24 hours and 48 hours,the expression level of Bcl-2,N-cadherin,Vimentin protein were significantly decreased(P<0.05).Compared with the si-DNMT1 group and the si-DNMT1+anti-NC group,the expression of miR-377-3p,the apoptosis rate of 4T1 cells,the expression levels of Bax,cleaved caspase-3 and E-cadherin proteins of 4T1 cells in the si-DNMT1+anti-miR-377-3p group were significantly decreased(P<0.05),the expression level of PUM1 protein,OD values at 24 hours and 48 hours,the expression levels of Bcl-2,N-cadherin,Vimentin proteins were significantly increased(P<0.05).Compared with the miR-NC+WT-circDNMT1 group,the cell luciferase activity in the miR-377-3p mimic+WT-circDNMT1 group was significantly decreased(P<0.05);compared with the miR-NC+WT-PUM1 group,the cell luciferase activity in the miR-377-3p mimic+WT-PUM1 group was significantly decreased(P<0.05).The tumor cells in the blank control group and the negative control group were densely arranged with clear boundary;the tumor cells in the DNMT1 silencing group and the combined control group were loosely arranged,the nuclei were pyknotic,and the cell fragments were increased;the tumor cells in the combined silencing group were densely arranged and the boundaries tended to be clear.Compared with the blank control group and the negative control group,the tumor mass of mice in the DNMT1 silencing group was significantly decreased(P<0.05).Compared with the DNMT1 silencing group and the combined control group,the tumor mass of mice in the combined silencing group was significantly increased(P<0.05).Conclusion Silencing circDNMT1 may inhibit the expression of PUM1 by up-regulating miR-377-3p,thereby inhibiting the proliferation and EMT of TNBC cells,and promoting cell apoptosis.
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Objective To investigate the effects and mechanism of circTRIM33-12 on proliferation,apoptosis and epithelial-mesenchymal transition(EMT)of brain glioma cells by miR-191/DAB2 axis.Methods The expressions of circTRIM33-12,miR-191 and DAB2 in brain glioma cell CHG-5 and human normal brain glial epithelial cells HEB were detected by RT-PCR.The cultured CHG-5 cells were divided into the siRNA NC group,the circTRIM33-12 siRNA group,the DAB2 siRNA group;the mimics NC group,the miR-191 mimics group;the circTRIM33-12 WT+mimics NC group,the circTRIM33-12 WT+miR-191 mimics group,the circTRIM33-12 MUT+ mimics NC group,the circTRIM33-12 MUT+miR-191 mimics group;the inhibitor NC group,the miR-191 inhibitor group;the pcDNA+ mimics NC group,the pcDNA-TRIM33-12+mimics NC group,the pcDNA+miR-191 mimics group,the pcDNA-TRIM33-12+miR-191 mimics group;the DAB2 WT+mimics NC group,the DAB2 WT+miR-191mimis group,the DAB2 MUT+mimics NC group,the DAB2 MUT+ miR-191 mimis group.CCK-8 assay was used to detect the effects of the expressions of circTRIM33-12,miR-191 and DAB2 on the prolifera-tion ability of CHG-5 cells;flow cytometry was used to detect the effects of the expressions of circTRIM33-12,miR-191 and DAB2 on the apoptosis of CHG-5 cells;Western blot was used to detect the effects of the expressions of circTRIM33-12,miR-191 and DAB2 on EMT of CHG-5 cells.TargetScan database was used to analyze the correlations among miR-191,circTRIM33-12 and DAB2,and dual luciferase reporter gene assay was used to verify their relationships;RT-qPCR was used to detect the effect of circTRIM33-12 on DAB2 expression through miR-191.Results Compared with HEB cells,the expression of circTRIM33-12 in CHG-5 cells was down-regulated(P<0.01),the expression of miR-191 was up-regulated(P<0.01),and the expression of DAB2 was down-regulated(P<0.01).Compared with the siRNA NC group,the proliferation activity and N-cadherin expression of CHG-5 cells in the circTRIM33-12 siRNA group and the DAB2 siRNA group were significantly increased(P<0.01),while the apoptosis rate and E-cadherin expression were decreased(P<0.01).circTRIM33-12 targeted miR-191,and miR-191 targeted DAB2.Compared with the inhibitor NC group,the proliferation activity and N-cadherin expression of CHG-5 cells in the miR-191 inhibitor group were significantly decreased(P<0.01),while the apoptosis rate and E-cadherin expression were increased(P<0.01).circTRIM33-12 overexpression inhibited CHG-5 cell proliferation and EMT through miR-191.Conclusion circTRIM33-12 may regulate the proliferation,apoptosis and EMT of brain glioma cells through the miR-191/DAB2 axis.
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Objective:To discuss the inhibitory effect of chelerythrine(CHE)on the migration,invasion,and epithelial-mesenchymal transition(EMT)of the human ovarian cancer SKOV3 cells,and to clarify the associated mechanism.Methods:The SKOV3 cells were cultured in vitro and divided into control group and 2.5,5.0,10.0,20.0,and 40.0 μmol·L-1 CHE groups.Methylthiazolydiphenyl-tetrazolium(MTT)assay was used to detect the inhibitory rates of proliferation of the cells in various groups.The SKOV3 cells were cultured in vitro and divided into control group,transforming growth factor-β1(TGF-β1)group,TGF-β1+5 μmol·L-1 CHE group,and TGF-β1+10 μmol·L-1 CHE group.Cell scratch assay was used to detect the migration rates of the cells in various groups;Transwell chamber assay was used to detect the numbers of migration and invasion cells in various groups;Western blotting method was used to detect the expression levels of E-cadherin,N-cadherin,and Vimentin proteins in the cells in various groups;immunofluorescence staining method was used to detect the fluorescence intensities of E-cadherin and N-cadherin in the cells in various groups.Results:The MTT assay results showed that compared with control group,the inhibitory rates of proliferation of the cells in 5.0,10.0,20.0,and 40.0 μmol·L-1 CHE groups were significantly increased(P<0.05 or P<0.01).The cell scratch assay results showed that compared with control group,the migration rate of the cells in TGF-β1 group was increased(P<0.01);compared with TGF-β1 group,the migration rates of the cells in TGF-β1+5 μmol·L-1 CHE group and TGF-β1+10 μmol·L-1 CHE group were significantly decreased(P<0.01).The Transwell chamber assay results showed that compared with control group,the numbers of migration and invasion cells in TGF-β1 group were significantly increased(P<0.05);compared with TGF-β1 group,the numbers of migration and invasion cells in TGF-β1+5 μmo·l L-1 CHE group and TGF-β1+10 μmo·l L-1 CHE group were significantly decreased(P<0.01).The Western blotting results showed that compared with control group,the expression level of E-cadherin protein in the cells in TGF-β1 group was significantly decreased(P<0.01),while the expression levels of N-cadherin and Vimentin proteins were increased(P<0.05 or P<0.01);compared with TGF-β1 group,the expression levels of E-cadherin protein in the cells in TGF-β1+5 μmol·L-1 CHE group and TGF-β1+10 μmol·L-1 CHE group were significantly increased(P<0.01),and the expression levels of N-cadherin and Vimentin proteins were significantly decreased(P<0.01).The immunofluorescence staining results showed that compared with control group,the fluorescence intensity of E-cadherin in the cells in TGF-β1 group was decreased,and the fluorescence intensity of N-cadherin was increased;compared with TGF-β1 group,the fluorescence intensities of E-cadherin in the cells in TGF-β 1+5 μmol·L-1 CHE group and TGF-β1+10 μmol·L-1 CHE group were significantly increased,and the fluorescence intensities of N-cadherin were decreased.Conclusion:CHE can inhibit the proliferation,migration,invasion,and EMT of the human ovarian cancer SKOV3 cells.
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Objective:To discuss the effect of downregulating of high mobility group box protein 2(HMGB2)expression on the biological behavior of the liver cancer cells and the epithelial-mesenchymal transition(EMT)process,and to clarify its mechanism.Methods:The human liver cancer LM3 cells at logarithmic growth phase were divided into negative control group and HMGB2 RNA interference group(HMGB2 siRNA group);the cells in two groups were transfected with RNA oligonucleotides(RNA oligos)with irrelevant sequences and RNA oligos designed to knock down HMGB2,and the Lipofectamine 2000 was regarded as the vector.The expression levels of HMGB2 mRNA and protein in the cells in two groups were detected by real-time fluorescence quantitative PCR(RT-qPCR)and Western blotting methods;cell scratch assay and Transwell chamber assay were used to detect the migration and invasion abilities of the cells in two groups;the expression levels of E-cadherin,N-cadherin,and Vimentin proteins and protein kinase B(AKT)/mammalian target of rapamycin(mTOR)pathway related proteins in the cells in two groups were detected by Western blotting method.Results:Compared with negative control group,the expression levels of HMGB2 mRNA and protein in the cells in HMGB2 siRNA group were significantly decreased(P<0.05),the cell scratch healing rate was significantly decreased(P<0.01),the number of invasion cells was significantly decreased(P<0.01),and the expression level of E-cadherin protein in the cells was significantly increased(P<0.01),while the expression levels of N-cadherin,Vimentin,mTOR,AKT,and phosphorylated AKT(p-AKT)proteins in the cells were significantly decreased(P<0.05 or P<0.01).Conclusion:Downregulating the expression of HMGB2 can reduce the migration and invasion abilities of the liver cancer LM3 cells and inhibit the EMT,and its mechanism may be related to regulating the expression of the AKT/mTOR pathway related proteins.
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Objective To investigate the effect of microRNA(miR)-4645-5p on the proliferation,invasion and epithelial-mesenchymal transition of esophageal cancer cells by targeting mucin 16(MUC16)and its mo-lecular mechanism.Methods The expression of miR-4645-5p in esophageal cancer tissues was analyzed online by TCGA database.The expression level of miR-4645-5p in esophageal cancer cell lines was analyzed by fluo-rescent real-time fluorescence quantitative polymerase chain reaction(qPCR).KYSE-30 cells were transfected with miR-4645-5p mimic and negative control mimic by lipofection technology,and were divided into miR-4645-5p group and control mimic group.The proliferation ability,migration ability and invasion ability of transfected KYSE-30 cells were analyzed by CCK-8 method,scratch test and Transwell test respectively.The target gene of miR-4645-5p was predicted by the bioinformatics website,and the binding of miR-4645-5p to the target gene was detected by the dual-luciferase reporter gene assay.The expression level of MUC16 mR-NA was detected by qPCR,and the protein expression levels of MUC16,transcription factor-1(ZEB-1),zonal atresia protein(ZO-1),tight junction protein-1(Claudin-1)and α-smooth muscle actin(α-SMA)were detected by Western blotting.Results The expression level of miR-4645-5p in esophageal cancer tissues was signifi-cantly lower than that in adjacent tissues(P＜0.01).Compared with HET-1 A,the expression of miR-4645-5p was lower in esophageal cancer cell lines(P＜0.05).After overexpression of miR-4645-5p,the proliferation a-bility of KYSE-30 cells was significantly reduced(P＜0.05),the migration ability was significantly reduced(P＜0.01)and the invasion ability was significantly reduced(P＜0.01).miR-4645-5p targeted and negatively regulated the expression of MUC16 mRNA(P＜0.01).After overexpression of miR-4645-5p,the protein ex-pression levels of MUC16,ZEB-1 and α-SMA were all down-regulated,and the protein expression levels of ZO-1 and Claudin-1 were up-regulated.Conclusion miR-4645-5p regulates the malignant biological behavior of esophageal cancer KYSE-30 cells by targeting MUC16.
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Objective To investigate the correlation between the expression of long non-coding ribonucleic acid growth arrest specific 5(lncRNA GAS5),phospholysine phosphohistidine inorganic pyrophosphate phos-phatase(LHPP)and epithelial-mesenchymal transition(EMT)in cancer tissues of patients with non-small cell lung cancer(NSCLC)and its clinical significance.Methods Cancer tissues and adjacent tissues of 90 patients with NSCLC who underwent surgical resection in the First Hospital Affiliated to Hebei North College from June 2018 to January 2020 were collected.The expressions of lncRNA GAS5,LHPP and EMT-associated pro-teins[E-calmodulin(E-Cad),N-calmodulin(N-Cad),and vimentin(VIM)]were detected by real-time fluores-cence quantitative polymerase chain reaction.The relationship between lncRNA GAS5 and LHPP mRNA and clinicopathological features in cancer tissues of NSCLC patients was analyzed,and the correlation between ln-cRNA GAS5 and LHPP mRNA and EMT-associated proteins expression in cancer tissues of NSCLC patients was analyzed by Pearson correlation.Kaplan-Meier method was used to plot the survival curves of NSCLC pa-tients with different lncRNA GAS5 and LHPP mRNA expressions,and multivariate Cox regression was used to analyze the prognostic factors of NSCLC patients.Results The expressions of lncRNA GAS5,LHPP mR-NA and E-Cad mRNA in cancer tissues of NSCLC patients were lower than those in adjacent tissues,while the expressions of N-Cad mRNA and VIM mRNA were higher than those in adjacent tissues,with statistical sig-nificance(P＜0.05).Pearson correlation analysis showed that lncRNA GAS5 in cancer tissues of NSCLC pa-tients was positively correlated with E-Cad mRNA expression(r=0.724,P＜0.001),and negatively correla-ted with N-Cad mRNA and VIM mRNA expression(r=-0.699,-0.689).P＜0.001);lncRNA GAS5 was positively correlated with LHPP mRNA expression(r=0.651,P＜0.001).The mRNA expressions of ln-cRNA GAS5 and LHPP in cancer tissues of NSCLC patients with different degrees of differentiation,tumor TNM stage and lymph node metastasis were significantly different(P＜0.05).Kaplan-Meier survival curve a-nalysis showed that the 3-year overall survival rate in the lncRNA GAS5 high expression group[68.18％(30/44)]was higher than that in the lncRNA GAS5 low expression group[36.96％(17/46)].The 3-year overall survival rate in the high LHPP mRNA expression group[67.39％(31/46)]was higher than that in the lowLHPP mRNA expression group[36.36％(16/44)],and the difference was statistically significant(x2=10.274,10.322,P＜0.05).Low differentiation,TNM stage Ⅲ and lymph node metastasis were independent risk factors for death in NSCLC patients,and lncRNA GAS5≥1.32 and LHPP mRNA≥1.12 were independ-ent protective factors(P＜0.05).Conclusion The low expression of lncRNA GAS5 and LHPP mRNA in cancer tissues of patients with NSCLC is related to EMT-associated proteins expression,differentiation de-gree,tumor TNM stage,lymph node metastasis and prognosis,and may become a new target for the diagnosis and treatment of NSCLC.
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Objective To investigate the effects of long non-coding RNA FEZ family zinc finger 1 antisense RNA 1(lncRNA FEZF1-AS1)on enhancer of zeste homolog 2(EZH2)in regulation of proliferation,migration,invasion and epithelial-mesenchymal transition(EMT)of pulmonary interstitial cells and its mechanism.Methods The A549 cells human lung adenocarcinoma cell line were divided into control group and model group[model cells were induced into lung interstitial cells after being treated with transforming growth factor β1(TGF-β1)20 ng/mL for 48 h].The protein expression of E-cadherin,N-cadherin and vimentin in each group was detected by Western blot.The expression of lncRNA FEZF1-AS1 and EZH2 in the two groups was detected by RT-qPCR.Cells in the trans-fection group were divided into si NC group,lncRNA FEZF1-AS1+OE vector group and si lncRNA FEZF1-AS1+OE EZH2 group.Cell proliferation was examined by CCK-8 method,cell migration was detected by cell scratch,and cell invasion was detected by Transwell assays.The protein expression of E-cadherin,N-cadherin,vimentin and EZH2 in each group was detected by Western blot.The direct binding effect of FEZF1-AS1 and EZH2 was deter-mined by RNA immuno-precipitation(RIP).Results Compared with the control group,the protein expression level of E-cadherin in the model group was significantly decreased(P＜0.05),and the protein expression of N-cadherin and vimentin was significantly increased(P＜0.05).Compared with the control group,the expression level of lncRNA FEZF1-AS1 and EZH2 genes was significantly increased in the model group(P＜0.05).Compared with si NC group,the proliferation,migration and invasion ability of si lncRNA FEZF1-AS1+OE vector group were decreased,the ex-pression of E-cadherin protein was increased while the expression of N-cadherin,vimentin and EZH2 was decreased(P＜0.05).Compared with si lncRNA FEZF1-AS1+OE vector group,the proliferation,invasion and migration of si lncRNA FEZF1-AS1+OE EZH2 group were increased(P＜0.05).E-cadherin expression was decreased,while N-cad-herin,vimentin and EZH2 expressions were increased(P＜0.05).RIP experiment further confirmed that lncRNA FEZF1-AS1 had direct binding effect with EZH2.Conclusions LncRNA FEZF1-AS1 can promote the proliferation,invasion,metastasis and EMT process of pulmonary fibrosis cells by regulating EZH2.
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Objective To explore the impact of macrophage-to-myofibroblast transition(MMT)on pulmonary fibro-sis induced by acute lung injury by LPS.Methods Totally 21 male mice were randomly classified into 7 groups:control group,model group(LPS-PF)at different time points and intervention group of clodronate-liposomes(CL-LIP)treatement at different time points(n=3).Pulmonary fibrosis was identified by HE and Masson staining microscopy.The immuno-fluorescence technology was used for the evaluation of numbers of macrophage-to-myofi-broblast transition cells(MMT cell which co-expressed CD68 and α-SMA).Bone marrow-derived macrophages(BMDMs)were randomly classified into two group:control(Ctrl)group and TGF-β1-treated group induced by transforming growthfactor-β1.α-SMA,FN and Col1 were detected by RT-qPCR.The expression of α-SMA,Smad3 and p-Smad3 protein was evaluated by Western blot.Results At day 7,the Ashcroft score of lung tissue in LPS-PF mouse model was significantly increased when compared with the Ctrl group(P＜0.01);While the score signifi-cantly declined when the model was pretreated with CL-LIP(P＜0.05).As detected by immuno-fluorescence stai-ning,in CL-LIP group the number of CD68-positive cells co-labeled with α-SMA was obviously less then that of LPS-PF group of the corresponding time point(P＜0.01).When the BMDMs were stimulated by TGF-β1 at 24 h,48 h and 96 h respectively,a higher expression of α-SMA,FN,Col1,were found in TGF-β1-treated group than that in Ctrl group at the corresponding time point(P＜0.01).The expression of Smad3,p-Smad3 significantly higher in LPS-PF group(at both day 7 and day 10)and TGF-β1-treated group(at both 48 h and 96 h)as compared to cor-responding control group(P＜0.01).Conclusions MMT promotes pulmonary fibrosis induced by ALI via LPS.Smad3 is proved to be involved in the MMT process.
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Objective:To investigate the influences of circular RNA(circRNA)BICD2 on the proliferation,apoptosis and epitheli-al-mesenchymal transition(EMT)of oral squamous cell carcinoma(OSCC)cells by regulating the miR-218-5p/Ras homolog gene family member A(RhoA)axis.Methods:qRT-PCR and Western blot were applied to detect CircBICD2,miR-218-5p gene expres-sion and RhoA protein expression in OSCC tissue,paracancerous tissue,human oral epithelial cell line HOEC and OSCC cell line HSC-4,CAL-27 and SCC-15,respectively.SCC-15 cells were divided into 7 groups of Ct(control,A),si-NC(B),si-CircBICD2(C),miR-NC(D),miR-218-5p(E),si-CircBICD2+anti-NC(F)and si-CircBICD2+anti-miR-218-5p(G).qRT-PCR was ap-plied to detect the expression of CircBICD2 and miR-218-5p in the cells;CCK-8 assay and plate cloning assay were applied to detect the cell proliferation;flow cytometry was applied to detect cell apoptosis;Western blot was applied to detect the protein expressions of RhoA,E-cadherin,Vimentin and N-cadherin;dual luciferase was applied to verify the relationship between CircBICD2 and miR-218-5p,miR-218-5p and RhoA,respectively.Results:In OSCC tissues and cells,CircBICD2 and RhoA proteins were highly ex-pressed,miR-218-5p was lowly expressed.In SCC-15 cells,the expression of CircBICD2 and RhoA protein was the highest,and the expression of miR-218-5p was the lowest(P＜0.05),SCC-15 cells in the 7 groups showed that,compared with group B,the expres-sion of CircBICD2 and RhoA in group C was decreased,and the expression of miR-218-5p was increased(P＜0.05);compared with group D,in group E RhoA protein expression level was decreased,and the miR-218-5p expression level was increased(P＜0.05);compared with group C and group F,the expression of miR-218-5p decreased in group G,and the expression of RhoA protein in-creased(P＜0.05).Down-regulation of CircBICD2 or over-expression of miR-218-5p inhibited SCC-15 cell proliferation and EMT,and induced cell apoptosis;down-regulation of miR-218-5p attenuated the inhibition of SCC-15 cell proliferation and EMT,the cell apoptosis was promoted by silencing CircBICD2.Cir-cBICD2 targeted the miR-218-5p/RhoA axis.Conclusion:Silencing CircBICD2 may inhibit RhoA expression by up-regulating miR-218-5p,inhibits SCC-15 cell proliferation and EMT,and induces cell apoptosis.
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Diabetic nephropathy(DN)is one of the most important complications of diabetes.Its pathogenesis is com-plex and has not been fully elucidated.Epithelial-mesenchymal transition(EMT)plays an important role in the development of DN.Relevant data show that glycogen synthesis kinase-3β(GSK-3β)participates in the process of EMT through multiple sig-naling pathways and affects the occurrence and progression of DN.This article reviews the research progress of GSK-3β in-volved in EMT in DN.
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ObjectiveBy observing the effect of Qianyang Yuyin granules on the phenotype of renal tubule epithelial cells， the intervention of Qianyang Yuyin granule on renal interstitial fibrosis was investigated. MethodThe renal tubular epithelial cells （HK-2） were treated with different concentrations of transforming growth factor （TGF）-β1 （5， 10， 15， 20， 25 μg·L-1） for 24 hours， and cell morphology and growth state were observed with an inverted phase contrast microscope. The 20 μg·L-1 was selected as the most appropriate concentration of TGF-β1 according to Western blot results for subsequent experiments. HK-2 cells were divided into six groups： blank group， TGF-β1 group （concentration of 20 μg·L-1）， low， medium， and high dose Qianyang Yuyin granule groups （concentration of 0.5， 1， 2 g·L-1）， and valsartan group （1 × 10-5 mol·L-1）. The cell activity was measured by cell proliferation and cell counting kit-8 （CCK-8）. The cell migration ability was detected by scratch test. The Transwell method was used to detect the invasiveness of cells. Western blot was used to detect levels of fibronectin （FN）， E-cadherin， α-smooth muscle activator （α-SMA）， Vimentin， collagen type Ⅰ（Col Ⅰ）， collagen type Ⅳ（Col Ⅳ）， and other related proteins. ResultTGF-β1 stimulating epithelial-mesenchymal transition （EMT） in renal tubular epithelial cells was time- and concentration-dependent. Compared with the blank group， higher concentration in the TGF-β1 group indicates longer intervention time and more obvious long spindle change of cells， and the migration and invasion ability of the cells was significantly enhanced. The protein expression level of FN， α-SMA， Vimentin， Col Ⅰ， and Col Ⅳ increased significantly （P<0.05， P<0.01）， while the expression level of E-cadherin protein decreased （P<0.05）. Compared with the TGF-β1 group， Qianyang Yuyin granule groups could maintain normal cell morphology， and the migration and invasion ability of the cells was inhibited. The protein expression level of FN， α-SMA， Vimentin， Col Ⅰ， and Col Ⅳ decreased （P<0.05， P<0.01）， and the expression of E-cadherin protein was significantly restored （P<0.05）. ConclusionQianyang Yuyin granule can reverse TGF-β1-induced interstitial transformation of renal tubular epithelial cells by reducing the phenotypic expression of mesenchymal cells and increasing the phenotypic expression of epithelial cells.
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OBJECTIVE To investigate the effects of anlotinib on the malignant phenotype of glioma cells by regulating the nuclear factor-κB （NF-κB） signaling pathway. METHODS Human glioma T98G cells were cultured in vitro， and 5-fluorouracil was used as positive control to investigate the effects of different concentrations of anlotinib （5， 10， 20 μmol/L） on the ability of proliferation， adhesion， migration and invasion， the expressions of epithelial-mesenchymal transition （EMT） related proteins [E-cadherin， N-cadherin， vimentin and fibronectin （FN）]. NF- κB signaling pathway inhibitor （BAY 11-7082） and activator （prostratin） were additionally used to verify the possible mechanism of the above effects of anlotinib. RESULTS Anlotinib with 5， 10， 20 μmol/L could significantly decrease the activity of cell proliferation （except for 5 μmol/L anlotinib group）， migration rate， and the number of adherent cells and invasive cells， could significantly up-regulate the expression of E-cadherin protein while down-regulate the expressions of N-cadherin， vimentin and FN protein （P＜0.05）； the effect of 20 μmol/L anlotinib was similar to that of positive control （P＞0.05）. Compared with 10 μmol/L anlotinib， pathway inhibitor could significantly decrease the ability of proliferation， adhesion， migration and invasion， and the expressions of N-cadherin， vimentin， FN and phosphorylated NF-κB p65 protein， while could significantly up-regulate the expression of E-cadherin protein （P＜0.05）； above indexes were reversed significantly by pathway activator （P＜0.05）. CONCLUSIONS Anlotinib may inhibit the proliferation， adhesion， migration and invasion of human glioma T98G cells， which may be associated with the inhibition of the NF-κB signaling pathway， thus inhibiting cell EMT-like processes.