Molecular Cloning of a Thiol Proteinase Inhibitor Gene and Its Expression in E.coli

A cDNA library was constructed with 1.5×10~6 pfu from rice immature seeds,fromwhich a cDNA clone for rice thiol proteinase inhibitor,oryzacystatin(OC),was isolated byscreening with synthesized oligodeoxynucleotide probe,which contained a 309bp open read-ing frame,84bp 5′-end noncoding region and a poly(A)signal AATAAA at the 3′-end fol-lowed by 31Nt poly(A).Then the coding region of OC was amplified and inserted into thedownstream of λP_RP_L promoter for thermal-inducible expression in E.coli.Shifting the cul-ture temperature from 30℃ to 42℃ led to a high level expression of OC,which exhibited adistinct band of 12.0 kDa and accounted for at least 10% of the total soluble proteins fromSDS-PAGE.The papain-inhibitory activity of the expressed OC was further confirmed.

Studies on Introduction of Arrowhead Proteinase Inhibitor Gene into N． tobacco Protoplasts

The Arrowhead Proteinase Inhibitor(API)gene was introduced into the protoplasts or mesophyll cells of N.tobacco by PEG-mediated.The transformed protoplasts undergoing dirrerentiation of callus and regeneration of plantlet have been growing into transgenic plants. Restriction endonuclease analysis of products amplificated by PCR indicates the existence of the API gene in the transformed plantlet.The extract of the leaves from the transformed plants shows trypsin inhibitory activity,which indicates the expression of the introduced API gene and the transformed plants can accumulate the inhibitor. However, the variation of the inhlbitory activity of the transformed plants reveals the importance of the integration site of the API gene in the genome.

Cotton Plants Transformed with the Activated Chimeric Cry1Ac and API-B Genes

A chimeric gene, Bt29K, composed of coding sequences of activated Cry1Ac insecticidal protein and an endoplasm reticulum-retarding signal peptide, was synthesized. A plant expression vector containing two expression cassettes for the Bt29K and API-B genes was constructed. These two insect-resistant genes were transferred into two cotton ( Gossypium hirsutum L.) varieties ( or lines) via Agrobacterium-mediated transformation and nine homozygous transgenic cotton lines showing a mortality of 90.0% - 99.7% to cotton ballworm (Heliothis armigera) larvae and good agronomic traits were selected through six generations. Molecular biology analysis revealed that one or two copies of the insecticidal protein genes were integrated into the transgenic cotton genome and activated Cry1Ac and API-B protein expression was at a level of 0.17% and 0.09% of the total soluble protein in the transgenic cotton leaves, respectively. Comparison of the insect-resistance of the homozygous lines expressing the activated chimeric Cry1Ac and API-B with that expressing Cry1Ac only revealed that the insect-resistance of the former is apparently higher than the latter. These results also indicate that the strategy to construct a plant expression vector expressing two different insect-resistant genes reported here is reasonable.

EFFECTS OF p53 GENE THERAPY COMBINED WITH CYCLOOXYGENASE-2 INHIBITOR ON CYCLOOXYGENASE-2 GENE EXPRESSION AND GROWTH INHIBITION OF HUMAN LUNG CANCER CELLS

Background Gene therapy by adenovirus-mediated wild-type p53 gene transfer has been shown to inhibit lung cancer growth in vitro,in animal models,and in human clinical trials.The antitumor effect of selective cyclooxygenase(COX)-2 inhibitors has been demonstrated in preclinical studies.However,no information is available on the effects of p53 gene therapy combined with selective COX-2 inhibitor on COX-2 gene expression and growth inhibition of human lung cancer cells.Methods We evaluated the effects of recombinant adenovirus-p53(Ad-p53) gene therapy combined with selective COX-2 inhibitor on the proliferation,apoptosis,cell cycle arrest of human lung adenocarcinoma A549 cell line,and the effects of tumor suppressor exogenous wild type p53 on COX-2 gene expression.ResultsAd-p53 gene therapy combined with selective COX-2 inhibitor celecoxib shows significant synergistic inhibition effects on the growth of human lung adenocarcinoma A549 cell line. Exogenous p53 gene can suppress COX-2 gene expression.ConclusionsSignificant synergistic inhibition effects of A549 cell line by the combined Ad-p53 and selective COX-2 inhibitor celecoxib may be achieved by enhancement of growth inhibition,apoptosis induction and suppression of COX-2 gene expression.This study provides first evidence that the administration of p53 gene therapy in combination with COX-2 inhibitors might be a new clinical strategy for the treatment or prevention of NSCLC.

Hypersensitive Inhibition of the Proliferation of Cells with Mutated DNA Repair-Related Genes by the Catalytic Topoisomerase II Inhibitor 20-O-IngenolEZ

We previously reported that many ingenol compounds derived from Euphoria kansui exhibit topoisomerase inhibitory activity. 20-O-ingenolEZ in these compounds exerted inhibitory effects on both topoisomerase II (topo II) activity and cell proliferative activity. Topoisomerase II inhibitors can be divided into the poison and catalytic inhibitor types and 20-O-ingenolEZ is a catalytic inhibitor and inhibits topo IIα through inhibition of ATPase activity, but induces topo II-mediated DNA damage and apoptosis in BLM-/- DT40 cells through the induction of the DNA damage checkpoint, similar to the poison type inhibitor adriamycin. The ATPase inhibitor of topo II ICRF-193 also showed poison-like characteristics in the same cell line. However, the inhibitory effects of ICRF-193 on the proliferation of BLM-/- DT40 cells differed from those of 20-O-ingenolEZ, as did the specificity of its inhibition of the proliferation of other cell lines. 20-O-ingenolEZ showed hypersensitive inhibition of the proliferation of MCF-7 cells and BLM-/- DT40 cells with mutated DNA repair-related genes.

Inhibiting effect of antisense oligonucleotides phosphorthioate on gene expression of TIMP-1 in rat liver fibrosis

AIM: To observe the inhibition of antisense oligonucleotides (asON) phosphorthioate to the tissue inhibitors metalloproteinase-1 (TIMP-1) gene and protein expression in the liver tissue of immunologically induced hepatic fibrosis rats. The possibility of reversing hepatic fibrosis through gene therapy was observed. METHODS: Human serum albumin (HSA) was used to attack rats, as hepatic fibrosis model, in which asONs were used to block the gene and protein expressing TIMP-1. According to the analysis of modulator, structure protein, coding series of TIMP-1 genome, we designed four different asONs. These asONs were injected into the hepatic fibrosis models through coccygeal vein. The results was observed by RT-PCR for measuring TIMP-1 mRNA expression, immunohistochemistry and in situ hybridization for collagen I, II, special staining of collagen fiber, and electron microscopic examination. RESULTS: Hepatic fibrosis could last within 363 days in our modified model. The expressing level of TIMP-1 was high during hepatic fibrosis process. It has been proved by the immunohistochemical and the electron microscopic examination that the asON phosphorthioate of TIMP-1 could exactly express in vivo. The effect of colchicine was demonstrated to inhibit the expressing level of mRNA and the content of collagen I, III in the liver of experimental hepatic fibrosis rats. However, the electron microscopy research and the pathologic grading of hepatic fibrosis showed that there was no significant difference between the treatment group and the model group (P】 0.05). CONCLUSION: The experimental rat model of hepatic fibrosis is one of the preferable models to estimate the curative effect of anti-hepatic fibrosis drugs. The asON phosphorthioate of TIMP-1 could block the gene and protein expression of TIMP-1 in the liver of experimental hepatic fibrosis rats at the mRNA level. It is possible to reverse hepatic fibrosis, and it is expected to study a new drug of antihepatic fibrosis on the genetic level. Colchicine has very limited therapeutic effect on hepatic fibrosis, furthermore, its toxicity and side effects are obvious.

Possible association of the 5-HTTLPR serotonin transporter promoter gene polymorphism with premature ejaculation in a Turkish population

We evaluated the genotypes of the serotonin transporter gene （5-HTT） in patients with premature ejaculation （PE） to determine the role of genetic factors in the etiopathogenesis of PE and possibly to identify the patient subgroups. A total of 70 PE patients and 70 controls were included in this study. All men were heterosexual, had no other disorders and were either married or in a stable relationship. PE was defined as ejaculation that occurred within 1 min of vaginal intromission. Genomic DNA from patients and controls was analyzed using polymerase chain reaction, and allelic variations of the promoter region of the serotonin transporter gene （5-HTTLPR） were determined. The 5-HTTLPR （serotonin transporter promoter gene） genotypes in PE patients vs. controls were distributed as follows： L/L 16% vs. 17%, L/S 30% vs. 53% and S/S 54% vs. 28%. We examined the haplotype analysis for three polymorphisms of the 5-HTTLPR gene： LL, LS and SS. The appropriateness of the allele frequencies in the 5-HTTLPR gene was analyzed by the Hardy-Weinberg equilibrium using the Z-test. The short （S） allele of the 5-HTTLPR gene was significantly more frequent in PE patients than in controls （P 〈 0.05）. We suggest that the 5-HTTLPR gene plays a role in the pathophysiology of all primary PE cases. Further studies are needed to evaluate the relationship between 5-HTTLPR gene polymorphism and patient subgroup （such as primary and secondary PE） responses to selective serotonin reuptake inhibitors as well as ethnic differences.

Angiotensin-converting enzyme and bradykinin gene polymorphisms and cough:A meta-analysis

AIM:To evaluate the association between genetic polymorphisms and angiotensin converting enzyme in-hibitor (ACEI)-related cough,and the race-or ethnicity-related difference in the prevalence of cough attributed to ACEI therapy.METHODS:We conducted a search in PubMed,EM-BASE,Cinahl,and the Cochrane Database without language limitation.A database of 11 studies on ACEI-related cough,with detailed information regarding ACE I/D or bradykinin B 2 receptor polymorphisms,was created.Eligible studies were synthesized using meta-analysis methods,including cumulative meta-analysis.A subgroup analysis was also performed using ethnicity.RESULTS:Six studies were included on ACE I/D poly-morphism (398 Caucasians,723 East Asians),and three studies were included on bradykinin B 2 receptor poly-morphism (300 East Asians).The distribution of ACE genotypes showed significant differences in the entire population (P=0.004) and in East Asians (P=0.005)but not in Caucasians (P=0.23).Allelic frequencies of ACE showed significant differences in East Asians [odds ratio (OR)=1.49 (1.11-2.02)].The meta-analysis with a random effects model showed a significant associa-tion between ACE allele I/D and ACEI-related cough [random effects (RE) OR=1.49 (1.11-2.02),P=0.009] in East Asians,but not in Caucasians [RE OR=0.90 (0.60-1.35)].The allelic frequencies of the bradykinin B 2 receptor gene were significantly different [OR=2.25 (1.42-3.57)].The distributions of the T/C genotypes of the bradykinin B 2 receptor gene were significantly dif-ferent (χ 2=8.366,P=0.015).The meta-analyses re-vealed that there was a significant association between the bradykinin B 2 receptor allele and ACEI-related cough in East Asians [RE OR=2.29 (1.42-3.69),P=0.001].CONCLUSION:ACE I/D and Bradykinin B 2 receptor polymorphisms contributed to the risk of ACEI-related cough in East Asians,but a negative association be-tween ACE I/D polymorphism and ACEI-related cough was observed in Caucasians.

Association of plasminogen activator inhibitor-1 4G/5G promoter polymorphism with recurrent cerebral infarction in China’s North Jiangsu Province

BACKGROUND： Many international studies have shown that plasminogen activator inhibitor-1 （PAl-l） 4G/5G promoter polymorphism does not increase the risk for cerebral infarction. OBJECTIVE： Using PCR methodology and agarose electrophoresis to detect PAI-1 4G/5G promoter polymorphism in patients with recurrent cerebral infarction in the North Jiangsu Province of China, and to compare results with healthy subjects and patients with first-occurrence cerebral infarction in the same region. DESIGN, TIME AND SETTING： Non-randomized, concurrent, control trial. A total of 122 cerebral infarction patients were admitted to Xuzhou Medical College Hospital＇s Department of Neurology and Xuzhou Central Hospital＇s Department of Neurology between July 2003 and August 2006. PARTICIPANTS： The patients consisted of 63 males and 59 females, aged （62 ± 10） years. They were divided into first-occurrence （n = 58） and recurrence （n = 64） groups. In addition, 50 healthy subjects that underwent physical examination in the outpatient department, including 26 males and 24 females, aged （60 ±12） years, were selected as controls. METHODS AND MAIN OUTCOME MEASURES： PAl-1 4G/5G promoter polymorphism was detected and analyzed using PCR methodology and agarose electrophoresis. RESULTS： Significant differences were determined in terms of genotypic frequency and allele frequency of PAI-1 4G/5G promoter polymorphism, in patients with first-occurrence or recurrent cerebral infarction, when compared with healthy subjects （P 〈 0.05）. There was, however, no significant difference between the first-occurrence and recurrence groups （P 〉 0.05）. CONCLUSION： PAl- 1 4G/5G promoter polymorphism is genetic risk factor for cerebral infarction in China. However, it may be associated with recurrence of cerebral infarction in patients from the North Jiangsu Province of China.

MATRIX METALLOPROTEINASE AND THEIR INHIBITORS: MOLECULAR ASPECTS OF THEIR ROLES IN THE TUMOR METASTASIS

The matrix metalloproteinases (MMPs) are a family of proteolytic enzymes, whose physiological functions include tissue remo-delling and embryogenesis. The importance of this group of proteins in the processes of tumor invasion and metastasis is now widely acknowledged, and has led to the search for MMP inhibitors for use as anticancer treatments in a clinical setting. The review aims to introduce current research relating to MMPs as well as their native and synthetic inhibitor, with particular emphasis on the molecular aspects of their roles in tumor metastasis.

Expression of X-linked Inhibitor of Apoptosis Protein and Its Effect on Chemotherapeutic Sensitivity of Bladder Carcinoma

The expression of X-linked inhibitor of apoptosis protein (XIAP) gene and its effect on chemotherapeutic sensitivity in bladder carcinoma was explored. By using immunohistochemistry, the expression of XIAP was detected in 47 bladder carcinomas and 5 normal bladder tissues. The XIAP gene was transfected into bladder cancer cell line T24 by liposome and the positive clone was screened by G418. Cellular XIAP mRNA level was detected by RT-PCR. Low-dose mitocycin C was administered to induce the apoptosis of T24 cells. The in vitro growth of bladder carcinoma cells was analyzed by MTT colorimetry, and the apoptosis rate was assayed by TUNEL methods. It was found XIAP was moderately expressed in bladder carcinomas with the the positive rate being 78.73% (37/47), but the positive rate was not correlated with carcinoma stages and grades (P<0.05). XIAP mRNA level in transfected T24 cells was significantly increased by 3.8 times as compared with that in the cells not transfected with XIAP. After treatment with low-dose mitomycin C (0.005 and 0.05 mg/mL), the growth rate in XIAP no-transfected control group was increased by (11.60±0.25)% and (16.51±0.87)% (P<0.05), and the apoptosis rate was decreased by (10.1±0.2)% and (11.9±0.2%) (P<0.05) respectively as compared with XIAP transfected group. It was concluded that XIAP was expressed in most of bladder carcimoma samples. Overexpression of XIAP in T24 could significantly reduce the MMC-induced apoptosis of bladder carcinoma, suggesting its effect on the chemothera- peutic sensitivity of T24 cells.

Breast cancer resistance protein(BCRP/ABCG2):its role in multidrug resistance and regulation of its gene expression

Breast cancer resistance protein(BCRP)/ATP-binding cassette subfamily G member 2(ABCG2) is an ATP-binding cassette(ABC) transporter identified as a molecular cause of multidrug resistance(MDR) in diverse cancer cells.BCRP physiologically functions as a part of a self-defense mechanism for the organism;it enhances elimination of toxic xenobiotic substances and harmful agents in the gut and biliary tract,as well as through the blood-brain,placental,and possibly blood-testis barriers.BCRP recognizes and transports numerous anticancer drugs including conventional chemotherapeutic and targeted small therapeutic molecules relatively new in clinical use.Thus,BCRP expression in cancer cells directly causes MDR by active efflux of anticancer drugs.Because BCRP is also known to be a stem cell marker,its expression in cancer cells could be a manifestation of metabolic and signaling pathways that confer multiple mechanisms of drug resistance,self-renewal(stemness),and invasiveness(aggressiveness),and thereby impart a poor prognosis.Therefore,blocking BCRP-mediated active efflux may provide a therapeutic benefit for cancers.Delineating the precise molecular mechanisms for BCRP gene expression may lead to identification of a novel molecular target to modulate BCRP-mediated MDR.Current evidence suggests that BCRP gene transcription is regulated by a number of trans-acting elements including hypoxia inducible factor 1α,estrogen receptor,and peroxisome proliferator-activated receptor.Furthermore,alternative promoter usage,demethylation of the BCRP promoter,and histone modification are likely associated with drug-induced BCRP overexpression in cancer cells.Finally,PI3K/AKT signaling may play a critical role in modulating BCRP function under a variety of conditions.These biological events seem involved in a complicated manner.Untangling the events would be an essential first step to developing a method to modulate BCRP function to aid patients with cancer.This review will present a synopsis of the impact of BCRP-mediated MDR in cancer cells,and the molecular mechanisms of acquired MDR currently postulated in a variety of human cancers.

Transcriptional regulation of human polo-like kinases and early mitotic inhibitors

Human polo-like kinases （PLK1-PLK4） have been implicated in mitotic regulation and carcinogenesis. PLK1 phosphorylates early mitotic inhibitor 1 （Emil） to ensure mitosis entry, whereas Emi2 plays a key role during the meiotic cell cycle. Transcription factor E2F is primarily considered to regulate the G1/S transition of the cell cycle but its involvement in the regulation of mitosis has also been recently suggested. A gap still exists between the molecular basis of E2F and mitotic regulation. The present study was designed to characterize the transcriptional regulation of human PLK and Emi genes. Adenoviral overexpression of E2F1 increased PLK1 and PLK3 mRNA levels in A549 cells. A reporter gene assay revealed that the putative promoter regions of PLK1, PLK3, and PLK4 genes were responsive to activators E2F, E2F1-E2F3. We further characterized the putative promoter regions of Emil and Emi2 genes, and these could be regulated by activators E2F and E2F1-E2F4, respectively. Finally, PLK1-PLK4, Emil, and Emi2 mRNA expression levels in human adult, fetal tissues, and several cell lines indicated that each gene has a unique expression pattern but is uniquely expressed in common tissues and cells such as the testes and thymus. Collectively, these results indicate that E2F can integrate G1/S and G2/M to oscillate the cell cycle by regulating mitotic genes PLK and Emi, leading to determination of the cell fate.

The histone deacetylase inhibitor trichostatin A induces cell cycle arrest and rapid upregulation of gadd45β in LS174T human colon cancer cells

Histone deacetylase (HDAC) inhibitors are considered as promising therapeutic agents against several malignant diseases because they inhibit cancer cell proliferation. The stress sensor genes of the growth arrest and DNA damage-inducible protein (gadd45) family exhibit disordered expression in several types of malignant diseases and are thus a novel target for cancer therapy. However, there have been only few investigations of whether HDAC inhibitors affect the expression of gadd45 genes. We examined the effects of a HDAC inhibitor, trichostatin A (TSA), on the time-dependent expression of gadd45 genes in the human colon cancer cell line LS174T. Addition of TSA to LS174T cells induced inhibition of cell proliferation by arresting the cell cycle. We found that TSA treatment of LS174T cells induced rapid upregulation of gadd45β mRNA expression within 15 min, reaching a peak level at 3 h. Although the time-dependent expression pattern of gadd45β mRNA was similar to that of gadd45β mRNA, the peak level of gadd45β was lower than that of gadd45β. TSA treatment also upregulated the mRNA level of p21Waf1/Cip1, a prolif- eration inhibitor, after 3 h, but downregulated the mRNA levels of cyclin D1, a proliferation inducer, after 3 h, and of c-Myc after 1 h. TSA treatment induced a certain level of apoptosis, but the mRNA level of p53, a potent apoptosis inducer, was down-regulated after 3 h. These results suggest that the up-regulation of p21Waf1/Cip1 and apoptosis was independent of p53 and that the early upregulation of gadd45β gene, which precedes the upregulation of p21Waf1/Cip1 and the downregulation of cyclin D1, are important in TSA-treated LS174T cells.

Aberrant expression of genes and proteins in pterygium and their implications in the pathogenesis

Pterygium is a common ocular surface disease induced by a variety of factors. The exact pathogenesis of pterygium remains unclear. Numbers of genes and proteins are discovered in pterygium and they function differently in the occurrence and development of this disease. We searched the Web of Science and PubMed throughout history for literatures about the subject. The keywords we used contain pterygium, gene, protein, angiogenesis, fibrosis, proliferation, inflammation, pathogenesis and therapy. In this review, we summarize the aberrant expression of a range of genes and proteins in pterygium compared with normal conjunctiva or cornea, including growth factors, matrix metalloproteinases and tissue inhibitors of mefalloproteinases, interleukins, tumor suppressor genes, proliferation related proteins, apoptosis related proteins, cell adhesion molecules, extracellular matrix proteins, heat shock proteins and tight junction proteins. We illustrate their possible mechanisms in the pathogenesis of pterygium as well as the related intervention based on them for pterygium therapy.

Pro-sigmaK inhibitor-BofA基因敲除对嗜热厌氧杆菌SCUT27葡萄糖和木糖利用的影响

嗜热厌氧杆菌SCUT27(Thermoanaerobacterium aotearoense SCUT27,SCUT27)能同时利用木糖和葡萄糖,是发酵廉价木质纤维素获得乙醇及乳酸的优势菌株。为了提高该菌株对木糖和葡萄糖的利用效率,对其在葡萄糖、木糖和混合糖下不同时间点转录组数据分析的基础上,敲除pro-sigmaK抑制因子BofA(pro-sigmak processing inhibitor BofA,BofA)基因,获得基因工程菌ΔbofA。摇瓶发酵结果显示,与出发菌相比,当以木糖为唯一碳源时,突变株ΔbofA对木糖的代谢速率提高51.43%,乳酸产率提高23.53%;以混合糖(葡萄糖:木糖为=1:1)为碳源时,ΔbofA对木糖的代谢速率提高44.44%,乳酸和乙醇的产率分别提高43.75%和20.00%。由此可见,pro-sigmaK抑制因子BofA是木糖利用的负调控因子,其功能有待进一步研究。

Screening Peptide Inhibitors Using Phage Peptide Library with Isocitrate Lyase in Mycobacterium tuberculosis as Target

When devoured by macrophages,Mycobacterium tuberculosis remains persistent in macrophages and gains energy through the glyoxylate bypass to maintain its long-term existence in host cells.Therefore it is possible to stop persistent infections by interdicting the glyoxylate bypass in which the isocitrate lyase（ICL） is the key rate-limiting enzyme and a persistence factor.ICL is the target of anti-TB（TB：tubercular） drugs,which could screen ICL out and effectively inhibit the activity of ICL in Mycobacterium tuberculosis,and because of this,anti-TB drugs can be used to kill persistent Mycobacterium tuberculosis.In this study,the ICL gene of the Mycobacterium tuberculosis H37Rv was cloned successfully and recombinant protein with bioactivity was obtained through the enzyme characteristic appraisal.The specific activity of the recombined ICL is 24μmol·mg-1·min-1.The recombined ICL protein was used as the target,and phages which can specifically combine to ICL were screened in the phage 7 peptide library.According to the results of the ELISA and DNA sequence detection,eventually three 7-peptide chains were synthesized.Then the peptide chains were reacted with ICL,respectively,to detect their inhibitory effects on ICL.The results show that all the three 7-peptide chains possessed varying inhibitory effects on the activity of ICL.This study provided lead compounds for the research and development of new peptide anti-TB drugs.

26S proteasome inhibitors inhibit dexamethasone-dependent increase of tyrosine aminotransferase and tryptophan 2,3-dioxygenase mRNA levels in primary cultured rat hepatocytes

Dexamethasone (Dex), a ligand for transcriptional enhancement of tyrosine aminotransferase (TAT) and tryptophan 2,3-dioxygenase (TO) genes, (100 nM) maximally increased these mRNA levels at 12 h and 7 h in primary cultured rat hepatocytes and the nuclear fraction, respectively. Lactacystin (5 μM) and epoxomicin (0.5 μM), 26S proteasome inhibitors, significantly suppressed the Dex-dependent maximum increase of TAT and TO mRNA levels in the cells and the nuclear fraction. Electrophoretic mobility shift assay demonstrated that lactacystin did not affect binding of glucocorticoid receptor to glucocorticoid responsive element. Furthermore, lactacystin did not affect the activation of GRE luciferase reporter by Dex transfected to the cells. The results demonstrate that 26S proteasome is positively involved in the Dex-dependent increase of TAT and TO mRNA levels in the cells and suggest that the mechanism of action of 26S proteasome may be degradation of some RNase(s), which breaks down TAT and TO mRNAs.

靶向MYC基因抗肿瘤研究进展

MYC是研究最为广泛的原癌基因之一,属于碱性螺旋-环-螺旋亮氨酸拉链(bHLH-Zip)DNA结合蛋白超家族,通过与MAX形成MYC-MAX异二聚体发挥功能。MYC影响蛋白质合成、细胞黏附、增殖、分化、细胞周期、代谢、细胞凋亡和血管形成等多种生物学过程。因其结构与位置的特殊性,难以直接靶向,针对MYC的治疗策略是研究的热点,本文将系统回顾MYC基因的结构、功能和其抑制剂的研究进展,为靶向MYC治疗肿瘤提供新思路。

The Mechanism of Lipofectamine 2000 Mediated Transmembrane Gene Delivery

In this paper, the relatived mechanism between lipofectamine 2000 mediated transmembrane gene delivery and endocytic pathway were investigated. Clathrin and caveolae-mediated endocytic pathway contributions to transfection efficiency were studied. The inhibitors of endocytosis were used to treat HEp-2 cells before lipofectamine 2000/pGFP-N2 transfection. Transfection efficiency was evaluated with green fluorescence protein (GFP) expression assays. Cell viability and cytotoxicity were evaluated with MTT method. The results indicated that inhibitors of clathrin (chlorpromazine or wortmannin) and caveolin (genistein) could reduce the cell transfection efficiency observably. Both clathrin and caveolae-mediated endocytic pathways play important roles in transmembrane gene delivery.