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1.
J. pediatr. (Rio J.) ; 99(3): 296-301, May-June 2023. tab
Article in English | LILACS-Express | LILACS | ID: biblio-1440463

ABSTRACT

Abstract Objective To verify the performance of the Net Promoter Score (NPS) as a tool to assess parental satisfaction in pediatric intensive care units (PICUs). Methods The authors conducted an observational cross-sectional multicenter study in the PICUs of 5 hospitals in Brazil. Eligible participants were all parents or legal guardians of PICU-admitted children, aged 18 years or over. The NPS was administered together with the EMpowerment of PArents in THe Intensive Care (EMPATHIC-30), used as the gold standard, and a sociodemographic questionnaire. For analysis, the results were dichotomized into values greater than or equal to the median of the tests. The associations between the 2 tools were evaluated and the distribution of their results was compared. Results The parents or legal guardians of 78 PICU-admitted children were interviewed. Of the respondents, 85% were women and 62% were in a private hospital. The median NPS was 10 (IQR, 10-10), and the median EMPATHIC-30 score was 5.7 (IQR, 5.4-5.9). Compared with the gold standard, the NPS had a sensitivity of 100% at all cutoff points, except at cutoff 10, where the sensitivity was slightly lower (97.5%). As for specificity, NPS performance was poorer, with values ranging from 0% (NPS ≥ 5) to 47.4% (NPS = 10). Conclusions NPS proved to be a sensitive tool to assess parental satisfaction, but with poor ability to identify dissatisfied users in the sample.

2.
Indian J Exp Biol ; 2023 Mar; 61(3): 159-166
Article | IMSEAR | ID: sea-222581

ABSTRACT

MicroRNA-20a-5p (miR-20a-5p) has been shown to function as a tumor promoter factor in several cancers. However, its role in small cell lung cancer (SCLC) remains unclear. In this study, we have made an attempt to measure the tumor tissue levels of miR-20a-5p in patients with SCLC using quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). The biological function of miR-20a-5p in SCLC cells was investigated in vitro and in vivo studies, including cell proliferation, migration assays and tumorigenicity in nude mice. Meanwhile?we conducted the luciferase reporter assay to verify the biological relationship between miR-20a-5p and CCNG2. The expression of miR-20a-5p was significantly upregulated in human SCLC compared to that in normal tissues. Kaplan-Meier analysis indicated that patients with high expression of miR-20a-5p are closely related with the shorter survival of SCLC. Further, multivariate analysis showed that miR-20a-5p was an independent prognostic factor. Increasing miR-20a-5p expression promotes the proliferation, migration and invasion of the NCI-H446 cells in vitro and in vivo. Dual-luciferase reporter gene assay demonstrated that miR-20a-5p directly targets CCNG2. These findings suggest that miR-20a-5p levels might be a novel diagnostic and prognostic marker of SCLC. Inhibiting miR-20a-5p could be a promising therapeutic strategy for SCLC.

3.
Article | IMSEAR | ID: sea-223571

ABSTRACT

Background & objectives: Studies have shown that apart from hereditary breast carcinomas, breast cancer susceptibility gene 1 (BRCA1) mutations conferring to its loss are seen in sporadic breast carcinomas (SBC) as well. The aim of the present study was to assess BRCA1 methylation in females presenting at King George’s Medical University, Lucknow, with SBC by both immunohistochemistry (IHC) and methylation PCR with respect to hormonal profile and various morphological prognostic parameters. The primary objective was to look for the association between BRCA1 protein expression and DNA promoter methylation. Methods: 81 mastectomy specimens from SBC of invasive breast carcinoma (no special type) were included in this study. After a detailed morphological assessment, formalin fixed paraffin embedded tissue from a representative tumour area was selected for BRCA1 IHC by heat-mediated antigen retrieval under high pH and DNA extraction and further bisulphate treatment. BRCA1 was studied for methylation by methylated and unmethylated PCR-specific primers. Results: BRCA1 promoter methylation was present in 42/81 (51.9%) participants, with significant BRCA1 protein loss (72.7%; P=0.002). A significant association between BRCA1 loss and hormonal profile was found (P=0.001); maximum in triple negative breast carcinoma (TNBC) (72%; 18/25). Most of the TNBC also harboured methylation (68%). Although not significant grade II and III tumours, lymph vascular invasion, ductal carcinoma in situ, and nodal metastasis (?3) were seen in a higher percentage in methylated tumours. Mortality in SBC was significantly associated with BRCA1 loss (30.3%; P=0.024). Interpretation & conclusions: Study results highlight the concept of “BRCAness” in SBC as well. Hence, we can confer that identification of BRCA1 loss in SBC can make it a perfect candidate for poly ADP- ribose polymerase inhibitors or cisplatin-based therapy like hereditary ones.

4.
Article in Portuguese | LILACS | ID: biblio-1533066

ABSTRACT

Objetivo: analisar os fatores associados a sintomas ansiosos e depressivos de alunos do internato interprofissional da Santa Casa de Belo Horizonte. Analisou-se também a autopercepção de aprendizagem e satisfação. Método: estudo transversal no qual analisou-se os fatores associados a sintomas ansiosos e depressivos estimados pelo Transtorno de Ansiedade Generalizada (GAD-7) e pelo Questionário de Saúde do Paciente (PHQ-9) em alunos de um internato interprofissional de enfrentamento à COVID-19. Ao final do internato, analisou-se a percepção do alcance dos objetivos de aprendizagem e a experiência do aluno utilizando Net Promoter Score (NPS). Resultado: entre os 92 alunos analisados, 22 (23,9%) apresentaram escores elevados para sintomas ansiosos e 26 (28,3%) para sintomas depressivos. A frequência de sintomas ansiosos foi maior entre alunos de farmácia quando comparados aos de medicina ou enfermagem (42,9%, 28,9%, 9,7%, respectivamente, p=0,035). Sintomas ansiosos foram menos frequentes entre alunos que sempre tiveram acesso a equipamento de proteção individual (EPI) quando comparados aos demais (17,7% vs. 36,7%; p=0,046). Alunos que tiveram sintomas de COVID-19, quando comparados aos demais, apresentaram maior frequência de sintomas ansiosos (44,1% vs. 12,1%; p=0,001) e depressivos (41,2% vs. 20,7%; p=0,035). O atendimento a pacientes com COVID-19 não esteve associado a sintomas depressivos nem ansiosos. Observou-se alto nível de percepção do alcance dos objetivos de aprendizagem, maior entre estudantes de enfermagem. O escore geral do NPS foi de 70, com maior frequência de promotores entre alunos de enfermagem (90%), quando comparados aos de farmácia (67%) e medicina (62%). Conclusão: sintomas ansiosos estiveram associados à categoria profissional, acesso a EPI e história prévia de sintomas de COVID-19.A percepção do alcance de objetivos propostos foi elevada e o escore NPS foi satisfatório, com maior proporção de promotores na enfermagem


Objective: to analyze the factors associated with symptoms of anxiety and depression among interprofessional students at Santa Casa de Belo Horizonte. Self-perception of learning and satisfaction were also analyzed. Methods: cross-sectional study that analyzed the factors associated with symptoms of anxiety and depression estimated by the Generalized Anxiety Disorder 7-item (GAD-7) and Patient Health Questionnaire-9 (PHQ-9). At the end of the internship, the perception of achievement of learning objectives and the student's experience were analyzed using the Net Promoter Score (NPS). Results: among the 92 students analyzed, 22 (23,9%) had high scores for anxiety symptoms and 26 (28,3%) for depressive symptoms. The frequency of anxious symptoms was higher among pharmacy students when compared to medicine or nursing students (42.9%, 28.9%, 9.7%, respectively, p=0.035). Anxiety symptoms were less frequent among students who always had access to personal protective equipment (PPE) when compared to the others (17.7% vs. 36.7%; p=0.046). Students who had symptoms of Covid-19, when compared to the others, had a higher frequency of anxiety (44.1% vs. 12.1%; p=0.001) and depressive symptoms (41.2% vs. 20.7%; p =0.035). Caring for Covid-19 patients was not associated with depression or anxiety. There was a high level of perception of achievement of learning objectives, higher among nursing students. Overall NPS score was 70, with a higher frequency of promoters among nursing students (90%), when compared to those in pharmacy (67%) or medicine (62%). Conclusion: anxiety symptoms were associated with the professional category, access to PPE, and previous history of Covid-19. The perception of achievement of learning objectives was high and the NPS score was satisfactory, with more promoters among nursing students


Subject(s)
Humans , Students
5.
Chinese Journal of Biochemistry and Molecular Biology ; (12): 1314-1321, 2023.
Article in Chinese | WPRIM | ID: wpr-1015646

ABSTRACT

Tripartite motif containing protein 7 (TRIM7), as a member of the E3 ubiquitin ligase TRIM family, plays an important regulatory role in immune regulation, metabolism and other physiological processes. The aberrant expression of TRIM7 is closely related to the development and progression of hepatocellular carcinoma (HCC) and it shows a complex regulatory role. However, the regulatory mechanism for the expression of TRIM7 in HCC remains unknown. In this study, multiple online databases were used to analyze the expression of TRIM7 in HCC and data indicated that TRIM7 expression was upregulated in HCC and correlated to poor prognosis. Subsequently, the transcription factor binding sites in the TRIM7 promoter region were analyzed using UCSC and JASPAR databases, and the results showed that TRIM7 promoter contains four SP1 binding sites. In this work, we demonstrated that SP1 could directly bind to its binding sites in TRIM7 promoter and positively regulate the transcriptional activity driven by the TRIM7 promoter using dual luciferase reporter experiments and the ChIP-PCR method. Moreover, our results also showed SP1 overexpression upregulated the expression of TRIM7 at both mRNA and protein levels (P<0. 01),and SP1 inhibitor, mithramycin A, could reverse the activated effect of SP1 on TRIM7 expression (P<0. 01). In conclusion, this study preliminarily reveals the regulatory mechanism of TRIM7 upregulation in HCC, which provides an important theoretical basis for further study of the gene function, early diagnosis and targeted therapy.

6.
Journal of Experimental Hematology ; (6): 489-494, 2023.
Article in Chinese | WPRIM | ID: wpr-982085

ABSTRACT

OBJECTIVE@#To screen better promoters and provide more powerful tools for basic research and gene therapy of hemophilia.@*METHODS@#Bioinformatics methods were used to analyze the promoters expressing housekeeping genes with high abundance, so as to select potential candidate promoters. The GFP reporter gene vector was constructed, and the packaging efficiency of the novel promoter was investigated with EF1 α promoter as control, and the transcription and activities of the reporter gene were investigated too. The activity of the candidate promoter was investigated by loading F9 gene.@*RESULTS@#The most potential RPS6 promoter was obtained by screening. There was no difference in lentiviral packaging between EF1 α-LV and RPS6-LV, and their virus titer were consistent. In 293T cells, the transduction efficiency and mean fluorescence intensity of RPS6pro-LV and EF1 αpro-LV were proportional to the lentiviral dose. The transfection efficiency of both promoters in different types of cells was in the following order: 293T>HEL>MSC; Compared with EF1 αpro-LV, RPS6pro-LV could obtain a higher fluorescence intensity in MSC cells, and RPS6pro-LV was more stable in long-term cultured HEL cells infected with two lentiviruses respectively. The results of RT-qPCR, Western blot and FIX activity (FIX∶C) detection of K562 cell culture supernatant showed that FIX expression in the EF1 α-F9 and RPS6-F9 groups was higher than that in the unloaded control group, and there was no significant difference in FIX expression between the EF1 α-F9 and RPS6-F9 groups.@*CONCLUSION@#After screening and optimization, a promoter was obtained, which can be widely used for exogenous gene expression. The high stability and viability of the promoter were confirmed by long-term culture and active gene expression, which providing a powerful tool for basic research and clinical gene therapy of hemophilia.


Subject(s)
Humans , Transduction, Genetic , Genetic Vectors , Hemophilia A/genetics , Transfection , Blood Coagulation Factors/genetics , Lentivirus/genetics
7.
Chinese Journal of Biotechnology ; (12): 1096-1106, 2023.
Article in Chinese | WPRIM | ID: wpr-970425

ABSTRACT

L-asparaginase (L-ASN) is widely applied in the treatment of malignant tumor and low-acrylamide food production, however, the low expression level hampers its application. Heterologous expression is an effective strategy to increase the expression level of target enzymes, and Bacillus is generally used as the host for efficient production of enzymes. In this study, the expression level of L-asparaginase in Bacillus was enhanced through optimization of expression element and host. Firstly, five signal peptides (SPSacC, SPAmyL, SPAprE, SPYwbN and SPWapA) were screened, among which SPSacC showed the best performance, reaching an activity of 157.61 U/mL. Subsequently, four strong promoters (P43, PykzA-P43, PUbay and PbacA) from Bacillus were screened, and tandem promoter PykzA-P43 showed the highest yield of L-asparaginase, which was 52.94% higher than that of control strain. Finally, three Bacillus expression hosts (B. licheniformis Δ0F3 and BL10, B. subtilis WB800) were investigated, and the maximum L-asparaginase activity, 438.3 U/mL, was reached by B. licheniformis BL10, which was an 81.83% increase compared with that of the control. This is also the highest level of L-asparaginase in shake flask reported to date. Taken together, this study constructed a B. licheniformis strain BL10/PykzA-P43-SPSacC-ansZ capable of efficiently producing L-asparaginase, which laid the foundation for industrial production of L-asparaginase.


Subject(s)
Bacillus licheniformis/metabolism , Asparaginase/genetics , Bacillus/genetics , Protein Sorting Signals , Promoter Regions, Genetic/genetics , Bacillus subtilis/genetics , Bacterial Proteins
8.
China Journal of Chinese Materia Medica ; (24): 2931-2939, 2023.
Article in Chinese | WPRIM | ID: wpr-981425

ABSTRACT

The U6 promoter is an important element driving sgRNA transcription in the CRISPR/Cas9 system. Seven PqU6 promo-ter sequences were cloned from the gDNA of Panax quinquefolium, and the transcriptional activation ability of the seven promoters was studied. In this study, seven PqU6 promoter sequences with a length of about 1 300 bp were cloned from the adventitious roots of P. quinquefolium cultivated for 5 weeks. Bioinformatics tools were used to analyze the sequence characteristics of PqU6 promoters, and the fusion expression vectors of GUS gene driven by PqU6-P were constructed. Tobacco leaves were transformed by Agrobacterium tumefaciens-mediated method for activity detection. The seven PqU6 promoters were truncated from the 5'-end to reach 283, 287, 279, 289, 295, 289, and 283 bp, respectively. The vectors for detection of promoter activity were constructed with GUS as a reported gene and used to transform P. quinquefolium callus and tobacco leaves. The results showed that seven PqU6 promoter sequences(PqU6-1P to PqU6-7P) were cloned from the gDNA of P. quinquefolium, with the length ranged from 1 246 bp to 1 308 bp. Sequence comparison results showed that the seven PqU6 promoter sequences and the AtU6-P promoter all had USE and TATA boxes, which are essential elements affecting the transcriptional activity of the U6 promoter. The results of GUS staining and enzyme activity test showed that all the seven PqU6 promoters had transcriptional activity. The PqU6-7P with a length of 1 269 bp had the highest transcriptional activity, 1.31 times that of the positive control P-35S. When the seven PqU6 promoters were truncated from the 5'-end(PqU6-1PA to PqU6-7PA), their transcriptional activities were different in tobacco leaves and P. quinquefolium callus. The transcriptional activity of PqU6-7PA promoter(283 bp) was 1.59 times that of AtU6-P promoter(292 bp) when the recipient material was P. quinquefolium callus. The findings provide more ideal endogenous U6 promoters for CRISPR/Cas9 technology in ginseng and other medicinal plants.


Subject(s)
Panax/genetics , Promoter Regions, Genetic , Agrobacterium tumefaciens/genetics , Computational Biology , Cloning, Molecular
9.
Acta Academiae Medicinae Sinicae ; (6): 405-409, 2023.
Article in Chinese | WPRIM | ID: wpr-981283

ABSTRACT

Objective To explore the relationship between scavenger receptor class B member 1 (SCARB1) gene promoter methylation and the pathogenesis of coronary artery disease. Methods A total of 120 patients with coronary heart disease treated in Renji Hospital affiliated to Shanghai Jiao Tong University School of Medicine from December 2018 to May 2020 were selected as the case group,while 140 gender and age matched healthy participants were randomly selected as the control group for a case-control study.The methylation status was detected by high-throughput target sequencing after bisulfite converting,and the methylation of CpG sites in the promoter region of SCARB1 gene was compared between the two groups. Results The case group showed higher methylation level of SCARB1+67 and lower methylation level of SCARB1+134 than the control group (both P<0.001),and the differences remained statistically significant in men (both P<0.001) and women (both P<0.001).The overall methylation level in the case group was lower than that in the control group [(80.27±2.14)% vs.(81.11±1.27)%;P=0.006],while this trend was statistically significant only in men (P=0.002). Conclusion The methylation of SCARB1 gene promotor is associated with the pathogenesis and may participate in the occurrence and development of coronary heart disease.


Subject(s)
Male , Humans , Female , Methylation , Case-Control Studies , China , Coronary Artery Disease/genetics , Promoter Regions, Genetic , DNA Methylation , Scavenger Receptors, Class B/genetics
10.
Chinese Journal of Biotechnology ; (12): 2794-2805, 2023.
Article in Chinese | WPRIM | ID: wpr-981233

ABSTRACT

Hevea brasiliensis is the main source of natural rubber. Restricted by its tropical climate conditions, the planting area in China is limited, resulted in a low self-sufficiency. Periploca sepium which can produce natural rubber is a potential substitute plant. cis-prenyltransferase (CPT), small rubber particle protein (SRPP) and rubber elongation factor (REF) are key enzymes involved in the biosynthesis of cis-1, 4-polyisoprene, the main component of natural rubber. In this study, we cloned the promoter sequences of CPT, SRPP and REF through chromosome walking strategy. The spatial expression patterns of the three promoters were analyzed using GUS (β-glucuronidase) as a reporter gene driven by the promoters through Agrobacterium-mediated genetic transformation. The results showed that GUS driven by CPT, SRPP or REF promoter was expressed in leaves and stems, especially in the leaf vein and vascular bundle. The GUS activity in stems was higher than that in leaf. This study provided a basis for analyzing the biosynthesis mechanism of natural rubber and breeding new varieties of high yield natural rubber.


Subject(s)
Peptide Elongation Factors/genetics , Plant Proteins/metabolism , Periploca/metabolism , Rubber , Plant Breeding , Cloning, Molecular
11.
Chinese Journal of Biotechnology ; (12): 1670-1683, 2023.
Article in Chinese | WPRIM | ID: wpr-981162

ABSTRACT

Overexpression of Krüppel like factor 2 (Klf2) or Klf7 inhibits adipocyte formation. However, it remains unclear whether Klf2 regulates klf7 expression in adipose tissue. In this study, oil red O staining and Western blotting were employed to study the effect of Klf2 overexpression on the differentiation of chicken preadipocytes. The results showed that Klf2 overexpression inhibited the differentiation of chicken preadipocytes induced by oleate and the expression of pparγ, while promoted klf7 expression in chicken preadipocytes. Spearman correlation analysis was used to study the correlation between the expression data of klf2 and klf7 in the adipose tissue of both human and chicken. The results showed that there was a significantly positive correlation between the expression of klf2 and klf7 in adipose tissues (r > 0.1). Luciferase reporter assay showed that overexpression of Klf2 significantly promoted the activity of chicken klf7 promoter (-241/-91, -521/-91, -1 845/-91, -2 286/-91, -1 215/-91; P < 0.05). In addition, the activity of klf7 promoter (-241/-91) reporter in chicken preadipocytes was significantly positively correlated with the amount of klf2 overexpression plasmid transfected (Tau=0.917 66, P=1.074×10-7). Moreover, Klf2 overexpression significantly promoted the mRNA expression of klf7 in chicken preadipocytes (P < 0.05). In conclusion, upregulation of klf7 expression might be one of the pathways that Klf2 inhibits chicken adipocyte differentiation, and the sequence from -241 bp to -91 bp upstream chicken klf7 translation start site might mediate the regulation of Klf2 on klf7 transcription.


Subject(s)
Animals , Humans , Chickens/genetics , Kruppel-Like Transcription Factors/metabolism , Transcription Factors/metabolism , Adipocytes/metabolism , Adipose Tissue/metabolism
12.
International Eye Science ; (12): 2031-2034, 2023.
Article in Chinese | WPRIM | ID: wpr-998485

ABSTRACT

AIM: To analyze the relationship between rs128912 single nucleotide polymorphism(SNP)in the promoter region of Toll-like receptor 3(TLR3)gene and cataract in Chinese Han population.METHODS: A total of 263 patients with cataract admitted to our hospital from June 2019 to June 2021 were selected as study group, and 150 patients with lens dislocation were included in control group. Western blotting was used to detect the expression of TLR3 protein in the anterior capsular tissues of lens in the two groups, and direct sequencing method was applied to analyze the polymorphism of rs128912 locus in the promoter region of TLR3 gene. The expression of peripheral blood TLR3 mRNA of patients with different genotypes was detected by real-time quantitative polymerase chain reaction(RT-qPCR).RESULTS: The expression level of TLR3 protein in the anterior capsular tissues in the study group was higher than that in the control group(P&#x003C;0.05). The frequencies of genotypes(AA, AT, TT)at rs128912 locus in the TLR3 gene promoter region in the study group and the control group were in accordance with Hardy-Weinberg genetic equilibrium, and there were differences in the frequencies of genotypes(AA, AT, TT)and frequencies of alleles(A, T)at rs128912 locus in the TLR3 gene promoter region between both groups(P&#x003C;0.05). The relative expression level of peripheral blood TLR3 mRNA in patients with TT genotype in the study group was higher than that in patients with AA or AT genotypes(P&#x003C;0.05).CONCLUSION: The expression of TLR3 protein in anterior capsular tissues of lens of patients with cataract is significantly up-regulated, and rs128912 locus polymorphism in the TLR3 gene promoter region is related to the susceptibility of cataract in Chinese Han population, and people with TT genotype are more prone to cataract.

13.
Chinese Journal of Microbiology and Immunology ; (12): 258-264, 2023.
Article in Chinese | WPRIM | ID: wpr-995284

ABSTRACT

Objective:To investigate the distribution of integrons and plasmid-mediated quinolone resistance (PMQR) genes in clinical isolates of Klebsiella aerogenes and to analyze the relationship between integrons and bacterial resistance to antimicrobial agents. Methods:Ninety-one Klebsiella aerogenes strains isolated from clinical samples in the Fengxian District Central Hospital from November 2015 to March 2021 were used in this study. Class 1 and class 2 integron-integrase genes ( intI1 and intI2) and PMQR genes were screened by PCR. The types of promoters and gene cassette arrays of variable regions were determined by sequencing. Besides, the relationship between integrons and antimicrobial resistance was analyzed. Results:The resistance rate of the 91 Klebsiella aerogenes isolates to aztreonam was more than 40.00% and the resistance rates to other commonly used antimicrobial agents were less than 35.00%. Among the 91 isolates, 30 carried the intI1 gene, while none of them carried the intI2 gene. Seven class 1 integron gene cassette arrays of variable regions were detected and the gene cassette array of aac(6′)-11 C- ΔereA2- IS1247- aac3- arr- ΔereA2 was detected in Klebsiella aerogenes. PcH1 with weak activity was the predominant variable region promoter of class 1 integrons. The detection rates of intI1-positive and intI1-negative isolates in ICU, neurosurgery and other clinical departments were statistically different ( P<0.05). The resistance rate of intI1-positive isolates to some commonly used antibiotics was significantly higher than that of intI1-negative isolates ( P<0.05). qnrS gene was the prevalent PMQR gene. The detection rates of integrons and PMQR genes in Klebsiella aerogenes isolates was low except for the strains isolated in 2016. Conclusions:Antimicrobial resistance in Klebsiella aerogenes was closely related to integrons. The distribution of integrons in Klebsiella aerogenes strains isolated from different clinical departments was different, and the monitoring of drug-resistant strains should be strengthened in ICU and neurosurgery. The resistance to quinolones in Klebsiella aerogenes strains in this region was mainly related to qnrS gene.

14.
Chinese Journal of Pharmacology and Toxicology ; (6): 511-511, 2023.
Article in Chinese | WPRIM | ID: wpr-992192

ABSTRACT

OBJECTIVE To investigate the improve-ment functions of flavonoid compounds on temozolomide(TMZ)-,aging-or AD model-induced dysregulation of hip-pocampal NSC lineage progression,retardancy of den-dritic spine maturation in new-born neurons,as well as impairment of hippocampal-related learning and memory.METHODS We applied 30-week-old neural stem cell(NSC)specific promoter Nestin-GFP and NestinCreERT2:Rosa26-LSL-tdTomato transgenic mice and 16-week-old AD model 5XFAD transgenic mice,together with hippo-campal microinjection(ih),endogenous fluorescence trac-ing and immunofluorescent staining.RESULTS Both fla-vonoid compound A and its functional derivative flavo-noid compound B dose-dependently improved TMZ-,aging-or AD-induced defects of hippocampal NSC lin-eage progression and the maturation of dendritic spines of newborn neurons,thereby improving hippocampus related learning and memory.CONCLUSION This paper provides a new idea and treatment strategy for the devel-opment of new flavonoids that can promote neurogene-sis for neurodegenerative diseases and aging.

15.
Rev. Soc. Bras. Med. Trop ; 55: e0122, 2022. graf
Article in English | LILACS-Express | LILACS | ID: biblio-1407005

ABSTRACT

ABSTRACT Background: Behavioral changes in Rattus norvegicus infected with two strains of Toxoplasma gondii (ME49 and VEG) were investigated. Methods: Rats were evaluated for motor activity and aversion or attraction to cat urine 60 days after infection. After euthanasia, arginine-vasopressin gene methylation in the central nervous system was evaluated. Results: A significant difference was observed in the methylation of the arginine-vasopressin promoter gene between rats infected with the ME49 and VEG strains. Conclusions: Although differences were not observed in many parameters, significant differences were observed in the methylation of the arginine-vasopressin promoter gene in rats infected with the two studied strains.

16.
China Journal of Chinese Materia Medica ; (24): 412-418, 2022.
Article in Chinese | WPRIM | ID: wpr-927983

ABSTRACT

Farnesyl diphosphate synthase(FPPS) is a key enzyme at the branch point of the sesquiterpene biosynthetic pathway, but there are no reports on the transcriptional regulation of FPPS promoter in Pogostemon cabin. In the early stage of this study, we obtained the binding protein PcFBA-1 of FPPS gene promoter in P. cabin. In order to explore the possible mechanism of PcFBA-1 involved in the regulation of patchouli alcohol biosynthesis, this study performed PCR-based cloning and sequencing analysis of PcFBA-1, analyzed the expression patterns of PcFBA-1 in different tissues by fluorescence quantitative PCR and its subcellular localization using the protoplast transformation system, detected the binding of PcFBA-1 protein to the FPPS promoter in vitro with the yeast one-hybrid system, and verified its transcriptional regulatory function by dual-luciferase reporter gene assay. The findings demonstrated that the cloned PcFBA-1 had an open reading frame(ORF) of 1 131 bp, encoding a protein of 376 amino acids, containing two conserved domains named F-box-like superfamily and FBA-1 superfamily, and belonging to the F-box family. Moreover, neither signal peptide nor transmembrane domain was contained, implying that it was an unstable hydrophilic protein. In addition, as revealed by fluorescence quantitative PCR results, PcFBA-1 had the highest expression in leaves, and there was no significant difference in expression in roots or stems. PcFBA-1 protein was proved mainly located in the cytoplasm. Furthermore, yeast one-hybrid screening and dual-luciferase reporter gene assay showed that PcFBA-1 was able to bind to FPPS promoter both in vitro and in vivo to enhance the activity of FPPS promoter. In summary, this study identifies a new transcription factor PcFBA-1 in P. cabin, which directly binds to the FPPS gene promoter to enhance the promoter activity. This had laid a foundation for the biosynthesis of patchouli alcohol and other active ingre-dients and provided a basis for metabolic engineering and genetic improvement of P. cabin.


Subject(s)
Amino Acid Sequence , Cloning, Molecular , Geranyltranstransferase/genetics , Pogostemon , Transcription Factors/genetics
17.
Chinese Journal of Biotechnology ; (12): 831-842, 2022.
Article in Chinese | WPRIM | ID: wpr-927748

ABSTRACT

Promoter is an important genetic tool for fine-tuning of gene expression and has been widely used for metabolic engineering. Corynebacterium glutamicum is an important chassis for industrial biotechnology. However, promoter libraries that are applicable to C. glutamicum have been rarely reported, except for a few developed based on synthetic sequences containing random mutations. In this study, we constructed a promoter library based on the native promoter of odhA gene by mutating the -10 region and the bystanders. Using a red fluorescent protein (RFP) as the reporter, 57 promoter mutants were screened by fluorescence imaging technology in a high-throughput manner. These mutants spanned a strength range between 2.4-fold and 19.6-fold improvements of the wild-type promoter. The strongest mutant exhibited a 2.3-fold higher strength than the widely used strong inducible promoter Ptrc. Sequencing of all 57 mutants revealed that 55 mutants share a 1-4 bases shift (4 bases shift for 68% mutants) of the conserved -10 motif "TANNNT" to the 3' end of the promoter, compared to the wild-type promoter. Conserved T or G bases at different positions were observed for strong, moderate, and weak promoter mutants. Finally, five promoter mutants with different strength were employed to fine-tune the expression of γ-glutamyl kinase (ProB) for L-proline biosynthesis. Increased promoter strength led to enhanced L-proline production and the highest L-proline titer of 6.4 g/L was obtained when a promoter mutant with a 9.8-fold higher strength compared to the wild-type promoter was used for ProB expression. The use of stronger promoter variants did not further improve L-proline production. In conclusion, a promoter library was constructed based on a native C. glutamicum promoter PodhA. The new promoter library should be useful for systems metabolic engineering of C. glutamicum. The strategy of mutating native promoter may also guide the construction of promoter libraries for other microorganisms.


Subject(s)
Corynebacterium glutamicum/metabolism , Gene Library , Metabolic Engineering , Promoter Regions, Genetic/genetics
18.
Acta Pharmaceutica Sinica ; (12): 1526-1536, 2022.
Article in Chinese | WPRIM | ID: wpr-924755

ABSTRACT

On the basis of the Uncaria transcriptome, specific primers were designed for UrSTR. The full-length cDNA of UrSTR (GeneBank: OL310251) was 1 541 bp, encoding 345 amino acid residues, and the promoter region sequence of UrSTR (GeneBank: OL310252) was 1 179 bp. Phylogenetic tree is revealed that UrSTR had a closest relationship with STR from Ophiorrhiza pumila and Ophiorrhiza japonica. Localization of UrSTR protein is revealed located in the vacuole membrane. Plant-care analysis indicated that the promoter region sequence of UrSTR, covering multiple light, stress and hormone-response cis-regulatory elements, and verified transcriptional activity. The results of SDS-PAGE show that pET-28a-UrSTR recombinant protein was successfully expressed, and the size was anticipated. The UrSTR prokaryotic expression system needs to be optimized in the later stage. The research lays the foundation for further purification to study its structure and functional characterization of the UrSTR protein.

19.
Organ Transplantation ; (6): 363-2022.
Article in Chinese | WPRIM | ID: wpr-923583

ABSTRACT

Objective To investigate the effect of compound Fufangteng mixture-containing serum on the proliferation of bone marrow mesenchymal stem cell (BMSC) and its mechanism. Methods Rat BMSC were isolated, cultured and purified in vitro by direct adherence method. Cell morphology was observed. Surface markers were identified by flow cytometry. The rats were treated with compound Fufangteng mixture at a dose of 3 mL/(kg·d) by gavage for 14 d, and then the drug-containing serum was collected. BMSC were divided into the blank control group, drug-containing serum group, Notch1 small interfering ribonucleic acid (siRNA) group and Notch1 siRNA+drug-containing serum group. The proliferation rate of BMSC was detected and the relative expression levels of Notch1 signaling pathway-associated messenger ribonucleic acid (mRNA) and proteins were measured in each group. Results Microscopic observation showed that the first generation BMSC were seen in the long spindle shape, and grown in the parallel or spiral pattern. The third generation BMSC positively expressed CD90 and CD44, whereas were negative for CD45. Compared with the blank control group, the proliferation rate of BMSC in the drug-containing serum group and Notch1 siRNA+ drug-containing serum group was significantly increased, whereas that of BMSC was significantly decreased in the Notch1 siRNA group (all P < 0.05). Compared with the Notch1 siRNA group, the proliferation rate of BMSC was significantly increased in the Notch1 siRNA+drug-containing serum group (P < 0.05). Compared with the blank control group, the relative expression levels of Hey1 and Delta-like ligand (DLL)1 mRNA and proteins were significantly up-regulated in the drug-containing serum group, whereas those were significantly down-regulated in the Notch1 siRNA group and Notch1 siRNA+drug-containing serum group (all P < 0.05). Compared with the Notch1 siRNA group, the relative expression levels of Hey1 and DLL1 mRNA and proteins were significantly up-regulated in the Notch1 siRNA+drug-containing serum group (all P < 0.05). Conclusions Compound Fufangteng mixture-containing serum may promote the proliferation of rat BMSC, and its mechanism is probably associated with the activation of Notch1 signaling pathway.

20.
Organ Transplantation ; (6): 522-2022.
Article in Chinese | WPRIM | ID: wpr-934775

ABSTRACT

Immune deficiency of the host caused by allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the initial factor of reactivation of latent human cytomegalovirus (HCMV). The risk factors of reactivation of HCMV in allo-HSCT recipients consist of the serological status of HCMV in donors and recipients, the matching degree of human leukocyte antigen (HLA) and pretreatment patterns, etc. The reactivation of HCMV is associated with the expression of a series of viral cleavage and proliferation proteins induced by the overexpression of major immediate early promoter/enhancer (MIEP) in the viral genome. In this article, the risk factors of reactivation of HCMV after allo-HSCT, the molecular changes related to maintaining latent infection of HCMV, the key role of MIEP overexpression in reactivation of HCMV, and the molecular pathways involved in reactivation of HCMV after allo-HSCT were reviewed and the major molecular events of reactivation of HCMV after allo-HSCT were elucidated, aiming to provide reference for the prevention and treatment of cytomegaloviral disease (CMVD) after allo-HSCT.

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