Alleviating experimental pulmonary hypertension via co-delivering FoxO1 stimulus and apoptosis activator to hyperproliferating pulmonary arteries

Pulmonary hypertension (PH) is an insidious pulmonary vasculopathy with high mortality and morbidity and its underlying pathogenesis is still poorly delineated. The hyperproliferation and apoptosis resistance of pulmonary artery smooth muscle cells (PASMCs) contributes to pulmonary vascular remodeling in pulmonary hypertension, which is closely linked to the downregulation of fork-head box transcriptional factor O1 (FoxO1) and apoptotic protein caspase 3 (Cas-3). Here, PA-targeted co-delivery of a FoxO1 stimulus (paclitaxel, PTX) and Cas-3 was exploited to alleviate monocrotaline-induced pulmonary hypertension. The co-delivery system is prepared by loading the active protein on paclitaxel-crystal nanoparticles, followed by a glucuronic acid coating to target the glucose transporter-1 on the PASMCs. The co-loaded system (170 nm) circulates in the blood over time, accumulates in the lung, effectively targets the PAs, and profoundly regresses the remodeling of pulmonary arteries and improves hemodynamics, leading to a decrease in pulmonary arterial pressure and Fulton's index. Our mechanistic studies suggest that the targeted co-delivery system alleviates experimental pulmonary hypertension primarily via the regression of PASMC proliferation by inhibiting cell cycle progression and promoting apoptosis. Taken together, this targeted co-delivery approach offers a promising avenue to target PAs and cure the intractable vasculopathy in pulmonary hypertension.

Pharmacological targets and clinical application of traditional Chinese medicine and active components in the prevention and treatment for depression / 中国临床药理学与治疗学

The pathological mechanism for depression is complicated due to the involvement of types of transcriptional factors and signaling pathways in the occurrence and development of depression. Traditional Chinese medicine (TCM) characterized with multi-components and multi-targets exert unique advantage in the prevention and treatment of depression. This review summarized the pharmacological action of TCM and active components on transcriptional factors (CREB, NF-κB, Nrf2) and signaling pathways (BDNF-TrkB, MAPK, GSK-3β, TLR-4-NLRP3-IL-1β) as well as elucidated the clinical application of TCM formula in the prevention and treatment for depression based on relevant literatures from home and abroad.

Expression analysis of transcription factor ERF gene family of Panax ginseng / 中国中药杂志

Ethylene responsive factor(ERF), one of the largest families of transcriptional factors in plants, plays a key role in se-condary metabolism of herbal plants. To analyze the expression of ERF family genes, the heat map clustering method was used by analyzing the ginseng transcriptomes of different parts and different growth years. The contents of ginsenosides Rg_1, Re and Rb_1 in various concentrations of MeJA-treated ginseng adventitious roots were determined by UPLC-MS/MS method. The expression of key genes of ginsenoside biosynthesis(DDS, CYP716A47, CYP716A53v2) and ERF family genes in MeJA-treated ginseng adventitious roots were determined by using real-time quantitative PCR. Pearson correlation was adopted to analyze the gene expression pattern of DDS, CYP716A47, CYP716A53v2 gene and ERF family. The results showed that the content of ginseng diol ginsenoside Rb_1 in ginseng adventitious roots treated with different concentrations of MeJA increased, and the content of ginseng triol ginsenoside Rg_1 and Re decreased. It is consistent with the increase of DDS and CYP716A47 expression and the decrease of CYP716A53v2 gene expression. The expression of ERF003, ERF118 and ERF012 genes was significantly positively correlated with CYP716A53v2, but negatively correlated with DDS. While the expression of ERF1B was significantly negatively correlated with CYP716A47.It is proved that ERF003, ERF118 and ERF012 were likely to inhibit the expression of DDS and promote the expression of CYP716A53v2, and ERF1B was likely to inhibit CYP716A47. This work could provide theoretical basis of ERF functional verification of regulating the biosynthesis of ginsenosides.

Effect of miR-106a expression negatively regulated by KLF4 on the invasive activity of human gastric cancer cells BGC-823 / 西安交通大学学报(医学版)

Objective: To investigate the effect of Krüppel-like factor 4 (KLF4) on the expression of miR-106a and their interaction on the invasive activity of human gastric cancer cells. Methods: The JASPAR database was used to screen the transcriptional factor combined with miR-106a promoter. KLF4 over-expression vector KLF4-pcDNA and miR-106a reporter gene pmiR-106a-WT/MUT-luc were both constructed, and the dual luciferase reporter assay was used to detect the effect of KLF4 on the activity of miR-106a promoter. Forty specimens of human gastric cancer and their adjacent formalin-fixed paraffin-embedded (FFPE) specimens were collected, and Real-time PCR and immunohistochemistry were both used to detect the expression of KLF4. Human gastric cancer cell line BGC-823 was divided into four groups: miR-106a mimic, mimic NC, KLF4+pcDNA, and pcDNA basic. Transwell assay was employed to detect the invasive ability of the gastric cancer cells after four different treatments. Results: Dual luciferase reporter assay indicated that KLF4-pcDNA could antagonize the activity of miR-106a promoter, which suggested that the activity of pmiR-106a-WT-luc luciferase was inhibited (pmiR-106a-WT+pcDNA-basic compared with pmiR-106a-WT+pcDNA-KLF4, P0.05). FFPE samples showed that the relative expression of KLF4 in gastric cancer was 0.69±0.59, which significantly differed from that in adjacent paracancerous tissues (P<0.001). The expression of KLF4 was correlated with differentiation degree, lymph node metastasis, and infiltration depth of human gastric cancer (P<0.05). The positive expression of KLF4 was mainly located in the nucleus and cytoplasm of gastric mucosal epithelial cells. Transwell essay showed that the number of invasive cells in the four different treatment groups was not entirely the same (P<0.001): 89.00±14.85 for miR-106a mimic, 50.90±17.94 for mimic NC, 37.70±12.60 for KLF4+pcDNA, and 66.20±3.19 for pcDNA basic. Conclusion: Transcription factor KLF4 at least has direct binding with miR-106a promoter in vitro, which may be involved in the invasion and metastasis process of gastric cancer cell by negatively regulating the expression of miR-106a at the upstream transcription level.

MarR family transcription regulator HpaR and XC0449 coordinately regulate the virulence of Xanthomonas campestris pv. campestris / 生物工程学报

MarR family transcription regulators are ubiquitous among bacteria and archaea. They extensively control multiple cellular processes and elaborately regulate the expression of genes involved in virulence, stress response and antibiotics at translational level. In Xanthomonas campestris pv. campestris, insertional inactivation of MarR family transcription regulator HpaR (XC2827) resulted in significantly decrease in virulence and increase in the production of the extracellular proteases. Here, we reported that the genome of Xcc 8004 encodes nine MarR family transcription regulators. The MarR family transcription regulators, HpaR (XC2827) and XC0449, were heterologous expressed and purified. In vitro MST and Pull-down assay confirmed the physical interaction between HpaR and XC0449. Phenotypical assay determined that deletion of XC0449 resulted in substantial virulence attenuation. In vitro EMSA, in vivo qRT-PCR and GUS activity assay identified that HpaR and XC0449 coordinately act as the transcriptional activator to regulate the expression of the virulence-associated gene XC0705, and eventually control the bacterial virulence and the production of extracellular proteases.

The promoting effects of SNAI1 activating matrix metalloproteinase on choroidal neovascularization under hypoxia / 中华实验眼科杂志

Objective To investigate whether vascular endothelial cells in choroidal neovascularization whether hypoxia condition can up-regulate SNAI1 and activate matrix metalloproteinase (MMP)2 and MMP9 therefore to participate in choroidal neovascularization(CNV).Methods Sixteen SPF male C57 mice aged 6-8 weeks were divided into control group and model group.CNV models were induced by retinal laser photocoagulation,and flatmount and frozen sections of retinal pigment epithelium (RPE)-choroid-sclera compound were prepared at 7 days after modeling.The CNV in flat-mount was examined by Isolectin B4 staining,and the location of SNAI1,MMP2 and MMP9 in frozen sections was determined by immunofluorescence technology.The expression of SNAI1,MMP2 and MMP9 at mRNA level in CNV was detected by real-time fluorescence quantitative PCR (real-time PCR).The use and care of experimental animals complied with Statement for the Use of Animals in Ophthalmic and Visual Research.The RF/6A cells were divided into normoxia group and hypoxia group and cultured for 24 hours in 5％ CO2condition and mix condition of 94％ N2,5％ CO2 and 1％ O2,respectively.The expression of SNAI1,MMP2 and MMP9 in the cells at mRNA and protein levels was detected by real-time PCR and Western blot assay,respectively.Small interfering RNA of SNAI1 (siSNAI1) was transfected into the cells,and then the expression of MMP2 in the cells at mRNA and protein levels was detected by real-time PCR and Western blot assay,respectively,and the migrating number of the cells was assayed by Transwell chamber assay.Results CD31 and SNAI1 positive-response cells were seen in RPE-choroid-sclera flat-mounts under the laser scanning confocal microscope.The relative expression levels of SNAI1mRNA and MMP2 mRNA in RPE-choroid-sclera tissues were higher in the model group than those in the control group (SNAI1 mRNA:1.291 ±0.060 vs.0.759±0.074,P =0.001;MMP2 mRNA:1.610±0.424 vs.0.772 ±0.080,P =0.044).The expression of MMP9 mRNA was not significantly elevated between model group and control group (P＞0.05).The relative expression level of MMP2 mRNA was higher in comparison with MMP9 mRNA in the model group (P＜0.01).The relative expressions of hypoxic induced factor 1α (HIF-1α),SNAI1 and MMP2 at mRNA level and protein level in RF/6A cells were significantly higher in the hypoxia group than those in the normoxia group (all at P＜0.05) and no considerable difference was seen in MMP9 mRNA expression between the two groups (P＞0.05).The relative expressions of MMP2 mRNA in the cells were 0.217±0.036 and 0.818±0.105,and those of MMP2 protein in the cells were 0.236±0.009 and 1.043±0.120 in the hypoxia+siSNAI1 group and only hypoxia group,respectively,with significant differences between them (P =0.002,0.003).The migrating number of the cells was (254.60 ±71.31)/field in the hypoxia+siSNAI1 group,which was significantly less than (534.10±96.21) /field in the control group (P =0.029).Conclusions The hypoxic environment at CNV can activate MMP2 by up-regulating the expression of SNAI1,which promotes the migration of vascular endothelial cells and therefore participates in CNV formation,and the intervention of SNAI1 activation under the hypoxic condition can inhibit this process.

The effect of Snail gene silencing on cell invasion and proliferation in human pancreatic cancer cell line PANC1 / 中华胰腺病杂志

Objective To observe the effect of silencing Snail gene on the invasion and proliferation ability of human pancreatic cancer cell line PANC1.Methods Lentiviral vectors that can express small hairpin RNA(shRNA) targeting human Snail gene(shRNA-Snail) or shRNA sequence that did not match any known mRNA(shRNA-NC) were constructed,and transfected into PANC1 cells.Untransfected cells served as control.mRNA and protein expression of Snail,α-smooth muscle actin (α-SMA) and E-cadherin was determined by real time quantitative PCR and Western blotting,respectively.In vitro invasion ability was tested by Transwell model.Proliferation ability was measured by CCK-8 assay.Results Compared with those in shRNA-NC group,Snail mRNA (0.27 ± 0.02 vs 0.92 ± 0.03) and protein level (0.26 ± 0.02 vs 0.80 ± 0.02),and α-SMA mRNA (0.33 ±0.04 vs 0.97 ±0.07) and protein level (0.31 ±0.04 vs 0.74 ±0.06) in shRNA-Snail group were obviously decreased,but E-cadherin mRNA (1.57 ± 0.45 vs 0.95 ± 0.08) and protein level (0.86 ± 0.03 vs 0.20 ± 0.03) were greatly increased.The number of cells permeating the septum of transwell [(6.80 ± 0.73)/400 magnification vs (26.80 ± 2.52)/400 magnification,P ＜0.01] was significantly decreased,and cell proliferation was inhibited (0.74 ± 0.05 vs 1.47 ± 0.04,P ＜ 0.01).All the differences above were statistically significant (all P ＜ 0.01).No significant differences were observed between shRNA-NC and normal control group.Conclusions Silencing Snail gene may restrain the invasion and proliferation ability of PANC1 cells to a certain degree.

Identification and functional analysis of a novel HAND1 mutation associated with congenital ventricular septal defect / 中南大学学报(医学版)

Objective:To identify the novel HAND 1 mutation associated with congenital ventricular septal defect (VSD) and to perform the functional analysis.Methods:A total of 125 patients with congenital VSD and 210 control individuals were recruited,and their clinical data and blood samples were collected.The genomic DNA from each study subject was isolated,and all the coding exons of HAND1 were amplified.The amplicons from HAND 1 were sequenced to identify a sequence variation.The functional characteristics of the mutant HAND 1 were analyzed by a dual-luciferase reporter assay system.Results:A novel heterozygous HAND1 mutation c.355G＞T,equivalent to E119X,was identified in a patient with sporadic VSD.This nonsense mutation was absent in the 210 control subjects.Functional analysis revealed that the mutant HAND1 lost the ability to transactivate a target gene.Conclusion:A novel HAND1 mutation with VSD is identified in this study.

Report of 6 cases of Xp11.2 translocation renal cell carcinoma and literature review / 中华泌尿外科杂志

Objective To explore the clinical and pathological characters of Xp1 1.2 translocation renal cell carcinoma.Method We screened patients of renal cell carcinoma of PUMCH between Jan.2011 and Dec.2015,6 patients with Xp11.2 translocation renal cell carcinoma were found.There were 2 males and 4 females,with average age of 39 (ranging from 16 to 73 years old).Diameter of tumor ranged from 1.9cm to 19.0cm,and 9.6cra in average.Among which,3 cases were detected by routine physical examination,1 by severe anemia (Hb 66g/L),1 by gross hematuria,and 1 by flank discomfort.Before treatment,2 cases had local metastasis (local lymph node,renal pelvis invasion),1 had distant metastasis (pulmonary metastasis).CT examination showed that the tumors had soft tissue density / low density,with significant enhancment or uneven enhancement in enhanced scanning,and were all considered malignancy.6 patients were all treated with surgeries,of which 5 patients received radical nephrectomy,1 patient received nephron sparing surgery.Result Pathologically,most clear cells arranged in a papillary,nest like structure,with psaamoma bodies in them.Immunohistochemical examination showed that all patients were positive for TFE3.AE1/AE3,RCC,Vimentin,CD10,EMA,P504 were positive in different degree.According to pathological result,all 6 patients were proved to be Xp1 1.2 translocation renal cell carcinoma.After surgery,2 patients received immunotherapy,2 received targeted drug therapy,and 1 received local radiotherapy.The follow-up duration ranged from 9 to 56 months (average 37 months).Among which,1 patient died from tumor recurrence and multiple metastasis 22 months after surgery,1 had pulmonary metastasis 12 months after surgery,and the tumor had no significant progress after receiving targeted drug therapy.All the other patients survive without tumor recurrence.Conclusions Xp1 1.2 translocation renal cell carcinoma predominantly occurs in children and adults younger than 40 years.Arterial phase enhancement is slightly lower for Xp1 1.2 translocation renal cell carcinoma in CT scan than that of renal clear cell carcinoma.Histological features and immunohistochemical staining of TFE3 positive expression are important means of diagnosis of this disease.If necessary,gene detection could be done to make better diagnose.Surgery is preferred treatment option.Metastatic leads to poor prognosis,and need to be supplemented by targeted drug therapy.

Effect of 1, 25-dihydroxy vitamin D3 combined with mechanical strain treatment on proliferation, differentiation, and expression of OPG and RANKL in osteoblastic MC3T3-E1 cells / 医用生物力学

Objective To investigate the effect of 1,25-(OH)2-vitamin D3 (VD3) or mechanical strain alone and their combined treatment on proliferation and differentiation of pre-osteoblast MC3T3-E1 cells in vitro, as well as gene and protein expression of osteoprotegerin (OPG) and receptor activator of nuclear factor-кB ligand (RANKL) in those cells. Methods MC3T3-E1 cells were treated with 10 nmol/L VD3, intermitted mechanical strain or with a combination of these two factors. Cell proliferation was assessed with flow cytometry, and alkaline phosphatase (ALP) activity was measured using a fluorometric detection kit. The mRNA expression of ALP, runt-related transcriptional factor 2 (Runx2), OPG, and RANKL genes was determined by real-time PCR. The proteins expression of Runx2, OPG, and RANKL was determined by Western blotting. ResultsVD3 inhibited the proliferation of MC3T3-E1 cells, but the mechanical strain had no effect on cell proliferation. Mechanical strain, VD3, and the combined treatment enhanced the ALP activity of MC3T3-E1 cells as well as the protein expression of Runx2. The effect of combined treatment was less pronounced than the effect of VD3 or mechanical strain alone. Mechanical strain promoted the gene and protein expression of osteoprotegerin (OPG) and increased the ratio of OPG/RANKL. However, the combination of VD3 and mechanical strain led to a decrease in ratio of OPG/RANKL. Conclusions Mechanical strain might be effective in inducing osteogenic differentiation and increasing bone formation. A joint stimulation with VD3 and strain can decrease proliferation and osteogenic differentiation and increase RANKL expression, which might affect bone remodeling. This study supplies some new data, which might be important in theoretical and clinical research of osteoporosis (OP) and other related bone diseases.

Bioinformatics analysis of biomarkers and transcriptional factor motifs in Down syndrome

In this study, biomarkers and transcriptional factor motifs were identified in order to investigate the etiology and phenotypic severity of Down syndrome. GSE 1281, GSE 1611, and GSE 5390 were downloaded from the gene expression ominibus (GEO). A robust multiarray analysis (RMA) algorithm was applied to detect differentially expressed genes (DEGs). In order to screen for biological pathways and to interrogate the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database, the database for annotation, visualization, and integrated discovery (DAVID) was used to carry out a gene ontology (GO) function enrichment for DEGs. Finally, a transcriptional regulatory network was constructed, and a hypergeometric distribution test was applied to select for significantly enriched transcriptional factor motifs. CBR1, DYRK1A, HMGN1, ITSN1, RCAN1, SON, TMEM50B, and TTC3 were each up-regulated two-fold in Down syndrome samples compared to normal samples; of these, SON and TTC3 were newly reported. CBR1, DYRK1A, HMGN1, ITSN1, RCAN1, SON, TMEM50B, and TTC3 were located on human chromosome 21 (mouse chromosome 16). The DEGs were significantly enriched in macromolecular complex subunit organization and focal adhesion pathways. Eleven significantly enriched transcription factor motifs (PAX5, EGR1, XBP1, SREBP1, OLF1, MZF1, NFY, NFKAPPAB, MYCMAX, NFE2, and RP58) were identified. The DEGs and transcription factor motifs identified in our study provide biomarkers for the understanding of Down syndrome pathogenesis and progression.

PITX2C loss-of-function mutations responsible for idiopathic atrial fibrillation

OBJECTIVE: This study aimed to identify novel PITX2c mutations responsible for idiopathic atrial fibrillation. METHODS: A cohort of 210 unrelated patients with idiopathic atrial fibrillation and 200 unrelated, ethnically matched healthy individuals used as controls were recruited. The whole coding exons and splice junctions of the PITX2c gene, which encodes a paired-like homeobox transcription factor required for normal cardiovascular morphogenesis, were sequenced in 210 patients and 200 control subjects. The causative potentials of the identified mutations were automatically predicted by MutationTaster and PolyPhen-2. The functional characteristics of the PITX2c mutations were explored using a dual-luciferase reporter assay system. RESULTS: Two novel heterozygous PITX2c mutations (p.Q105L and p.R122C) were identified in 2 of the 210 unrelated patients with idiopathic atrial fibrillation. These missense mutations were absent in the 400 control chromosomes and were both predicted to be pathogenic. Multiple alignments of PITX2c protein sequences across various species showed that the altered amino acids were highly evolutionarily conserved. A functional analysis demonstrated that the mutant PITX2c proteins were both associated with significantly reduced transcriptional activity compared with their wild-type counterparts. CONCLUSION: The findings of this study associate PITX2c loss-of-function mutations with atrial fibrillation, supporting the hypothesis that dysfunctional PITX2c confers enhanced susceptibility to atrial fibrillation and suggesting potential implications for early prophylaxis and allele-specific therapy for this common arrhythmia. .

Expression of transcriptional factor lslet-1 in retina with experimental retinal neovascularization induced by oxygen / 国际眼科杂志(Guoji Yanke Zazhi)

AlM:To evaluate the expression of transcriptional factor lslet-1 in retina in experimental retinal neovascularization induced by oxygen. METHODS: The murine retinal neovascularization were induced by hyperoxia exposure. The morphological observation of retinal neovascularization was performed using angiography by fluorescein dextran injection under the fluorescence microscope, and the new blood vessels were quantified after 5d in room air (17-day-old) by counting the vascular epithelial cell nuclei protruding into viteous cavity using HE stain. Realtime PCR and Western blot were used to examine retinal lslet-1 level in postnatal 7,12, 14,17 and 26d respectively. RESULTS: A lots of new blood vessels were demonstrated in the mouse retina in hyperoxic group by fluorescein angiography and histological method. Moreover, no significant difference was found in retinal lslet-1 level in postnatal 7d between hyperoxic group and control group, but was significantly higher in postnatal 12, 14 and 17d mice compared with control mice. However, mice at postnatal 26d, expression of lslet-1 in retina decreased to normal level. CONCLUSlON: ln processing mouse model of retinal neovascularization, sustained hypoxia retinal tissue induce retinal neovascularization by increas the expression of transcription factor lslet-1.

Prevalence and spectrum of Nkx2.5 mutations associated with idiopathic atrial fibrillation

OBJECTIVE: The aim of this study was to evaluate the prevalence and spectrum of Nkx2.5 mutations associated with idiopathic atrial fibrillation (AF). METHODS: A cohort of 136 unrelated patients with idiopathic atrial fibrillation and 200 unrelated, ethnically matched healthy controls were enrolled. The coding exons and splice junctions of the Nkx2.5 gene were sequenced in 136 atrial fibrillation patients, and the available relatives of mutation carriers and 200 controls were subsequently genotyped for the identified mutations. The functional characteristics of the mutated Nkx2.5 gene were analyzed using a dual-luciferase reporter assay system. RESULTS: Two novel heterozygous Nkx2.5 mutations (p.N19D and p.F186S) were identified in 2 of the 136 unrelated atrial fibrillation cases, with a mutational prevalence of approximately 1.47%. These missense mutations co-segregated with atrial fibrillation in the families and were absent in the 400 control chromosomes. Notably, 2 mutation carriers also had congenital atrial septal defects and atrioventricular block. Multiple alignments of the Nkx2.5 protein sequences across various species revealed that the altered amino acids were completely conserved evolutionarily. Functional analysis demonstrated that the mutant Nkx2.5 proteins were associated with significantly reduced transcriptional activity compared to their wild-type counterpart. CONCLUSION: These findings associate the Nkx2.5 loss-of-function mutation with atrial fibrillation and atrioventricular block and provide novel insights into the molecular mechanism involved in the pathogenesis of atrial fibrillation. These results also have potential implications for early prophylaxis and allele-specific therapy of this common arrhythmia. .

Novel GATA5 loss-of-function mutations underlie familial atrial fibrillation

OBJECTIVE: This study aimed to identify novel GATA5 mutations that underlie familial atrial fibrillation. METHODS: A total of 110 unrelated patients with familial atrial fibrillation and 200 unrelated, ethnically matched healthy controls were recruited. The entire coding region of the GATA5 gene was sequenced in 110 atrial fibrillation probands. The available relatives of the mutation carriers and 200 controls were subsequently genotyped for the identified mutations. The functional effect of the mutated GATA5 was characterized using a luciferase reporter assay system. RESULTS: Two novel heterozygous GATA5 mutations (p.Y138F and p.C210G) were identified in two of the 110 unrelated atrial fibrillation families. These missense mutations cosegregated with AF in the families and were absent in the 400 control chromosomes. A cross-species alignment of GATA5 protein sequence showed that the altered amino acids were completely conserved evolutionarily. A functional analysis revealed that the mutant GATA5 proteins were associated with significantly decreased transcriptional activation when compared with their wild-type counterpart. CONCLUSION: The findings expand the spectrum of GATA5 mutations linked to AF and provide novel insights into the molecular mechanism involved in the pathogenesis of atrial fibrillation, suggesting potential implications for the early prophylaxis and personalized treatment of this common arrhythmia.

Construction of OTX1 Lentiviral Vector and Overexpression Research / 中国生物工程杂志

OTX1 gene is one of the pivotal transcriptional factors involved in the neurogenesis.In order to overexpress the OTX1 gene in distinct cell types and find out its contribution to the proliferation and differentiation of stem cells in vitro,OTX1 cDNA was subcloned into lentiviral vectors.The resulting constructions pDUETOTX1,pDUETGFPOTX1 and pDUETGFP were packaged in 293 cells producing viral particles to transduce 293T cells,SY5Y cells,mouse embryonic stem cells and E15 neural stem cells.It was proved that the transferred OTX1 gene was located in the nuclei of the transduced cells in stead of plasma.Lentivirus is an ideal vector delivering gene to different cells.The overexpression of OTX1 in transduced 293T cells were validated by Western blot and immunofluorescence.

Hypoxia-inducible factors and oxygen sensors / 第二军医大学学报

The erythropoietin(EPO) system was used to study the molecular mechanisms associated with the induction of hypoxia responsive genes and from these investigations the hypoxia-inducible factors (HIFs) was identified as a key transcriptional hypoxic regulator of EPO. Subsequent research has now found that a large number of other hypoxia-inducible genes are also induced by HIF under hypoxic conditions. The HIF transcriptional complex is a heterodimer consisting of an alpha subunit and a beta subunit. Recently, two oxygen sensor, oxygen-dependent prolyl and asparaginyl hydroxylation were found,and it is the first time that the oxygen sensor had been described for higher organisms. This review focused on the structure and functions of HIF and the regulation of HIF proteins by hypoxia and oxygen sensors.

FOXC1 expression in cervical cancer and endometrial cancer / 重庆医科大学学报

Objective:To investingate potential correlations between FOXC1 expression and clinicopathologic features of endometrial cancer and cervical cancer.Methods:FOXC1 protein expression in 54 cases of cervical cancer and 23 of endometrial cancer were investigated with immunohistochemistry.Results:FOXC1 protein expression was observed in cancer tissues,including both endometrial and cervical cancer.Positive FOXC1 expression was revealed in 20(87.0%)of endometrial cancer and 29(53.7%)of cervical cancer.Significant correlation in retrospective study was observed between FOXC1 protein expression and histological grades(P0.05).Conclusion:FOXC1 may play a role in tumorigenesis of cervical cancer.FOXC1 may have no correlation with tumorigenesis of endometrial cancer.

Reactive oxygen species in regulation of hypoxia-inducible factor / 第二军医大学学报

Oxygen is a mandatory for all aerobic organisms. Oxygen-containing free radicals are produced when oxygen is not completely reduced to water in energy-producing oxidation reaction.The radicals may also transform into other reactive compounds through electron transfer and all the compounds with similar functions are referred as reactive oxygen species(ROS).Increased ROS is known to cause damage to proteins,DNA and lipids.Much evidence showed that changes in partial oxygen pressure,hormone,cytokine and chemical stimulation could increase ROS,and ROS,acting as signaling molecules,mediates cell functions.Hypoxia-inducing factor(HIF),a key transcriptional factor for most hypoxia-inducible genes,is a heterodimer consisting of 2 subunits.Recent study found that ROS plays an important role in HIF activity regulation under hypoxic and non-hypoxic conditions.This paper reviews the production of ROS and its role in the regulation of HIF activity.