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1.
Clin Oral Investig ; 25(11): 6211-6217, 2021 Nov.
Article in English | MEDLINE | ID: mdl-33818684

ABSTRACT

OBJECTIVES: To evaluate the effect of different endodontic sealers (epoxy resin-based and bioceramic-based) and the time of post-cementation on the bond strength of a fiber post cemented with resin cement. METHODS: Forty human premolars were instrumented and divided into 4 groups. According to the type of sealer and the time of post-cementation: AH-IM (AH Plus, post-cemented immediately after root canal treatment), SP-IM (iRoot SP, post-cemented immediately after root canal treatment), AH-OW (AH Plus, post-cementation after one week), and SP-OW (iRoot SP, post-cementation after one week). In each group, the samples were submitted to push-out test, and failure mode was assessed. Levene's test, one-way ANOVA, and Kruskal-Wallis analysis were applied for statistical analysis (α = 5%). RESULTS: The highest mean push-out bond strength was obtained from the SP-IM group in the apical part (10.45 ± 5.15MPa), while the lowest was observed in samples from the AH-OW group in the middle part (2.63 ± 1.54 MPa). One-way ANOVA showed that within the same root region, the time of post-cementation had a negative influence on the bonding strength in the SP groups in the middle and apical portion (P<0.05), however, when comparing the effect of type of sealers on bonding strength between the OW groups or IM groups within the same root region, no significant difference was observed regardless of the post cementation time (P>0.05). CONCLUSIONS: The bond strength of the fiber post was higher when the post was cemented immediately after root canal treatment when the bioceramic sealer was applied. CLINICAL RELEVANCE: The correct choice of an endodontic sealer and the proper time of post-cementation may help to obtain the best quality of post-and-core restoration.


Subject(s)
Dental Bonding , Post and Core Technique , Root Canal Filling Materials , Cementation , Dentin , Epoxy Resins , Humans , Materials Testing , Resin Cements
2.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-974646

ABSTRACT

Objective To analyze the medical examinations of radiation worker in medical institutions and provide some basic data for radiation protection management. Methods The occupational health examination of 3568 radiation workers from 681 medical institutions who came to our hospital for occupational health examination from January 1 to December 31 in 2020 were summarized and analyzed. Results There was no case of suspected occupational radiation sickness. The abnormal rate was in the range of 2.17%~2.99%, the rate of occupational contraindicated was about 1.44%~2.17%. The total review rate was about 13.00%, more than 79.48% of the radiation workers were checked out other diseases or abnormal. The abnormal examination items are mainly ophthalmology, B ultrasound of liver, gallbladder, spleen and pancreas, liver function, electrocardiogram, blood routine, urine routine, blood pressure, B ultrasound of both kidneys and kidney function. The abnormal rate of ophthalmology in each level of institutions was decreased with the increase of the length of service, while the abnormal results of B-ultrasound of liver, gallbladder, spleen and pancreas, blood pressure, B-ultrasound of both kidneys and renal function were increased with the increase of service. Conclusion Maybe the radiation protection of radiation workers in medical institutions was well in Shenzhen, but there were different effects of the health status of the staff. Therefore, it is important to further strengthen the occupational health monitoring management.

3.
Acta Pharmaceutica Sinica B ; (6): 475-487, 2020.
Article in English | WPRIM (Western Pacific) | ID: wpr-792995

ABSTRACT

ProBiotic-4 is a probiotic preparation composed of , , , and . This study aims to investigate the effects of ProBiotic-4 on the microbiota-gut-brain axis and cognitive deficits, and to explore the underlying molecular mechanism using senescence-accelerated mouse prone 8 (SAMP8) mice. ProBiotic-4 was orally administered to 9-month-old SAMP8 mice for 12 weeks. We observed that ProBiotic-4 significantly improved the memory deficits, cerebral neuronal and synaptic injuries, glial activation, and microbiota composition in the feces and brains of aged SAMP8 mice. ProBiotic-4 substantially attenuated aging-related disruption of the intestinal barrier and blood-brain barrier, decreased interleukin-6 and tumor necrosis factor- at both mRNA and protein levels, reduced plasma and cerebral lipopolysaccharide (LPS) concentration, toll-like receptor 4 (TLR4) expression, and nuclear factor-B (NF-B) nuclear translocation in the brain. In addition, not only did ProBiotic-4 significantly decreased the levels of -H2AX, 8-hydroxydesoxyguanosine, and retinoic-acid-inducible gene-I (RIG-I), it also abrogated RIG-I multimerization in the brain. These findings suggest that targeting gut microbiota with probiotics may have a therapeutic potential for the deficits of the microbiota-gut-brain axis and cognitive function in aging, and that its mechanism is associated with inhibition of both TLR4-and RIG-I-mediated NF-B signaling pathway and inflammatory responses.

4.
Toxicol Lett ; 288: 129-135, 2018 May 15.
Article in English | MEDLINE | ID: mdl-29477354

ABSTRACT

Early diagnosis and treatment of occupational medicamentosa-like dermatitis due to trichloroethylene (OMLDT) are absence of specific and reliable diagnostic/therapeutic biomarkers. This study was conducted on 30 cases of OMLDT, 58 workers exposed to trichloroethylene (TCE) and 40 unexposed controls in order to identify any cytokine signatures that give an index to CD4+T cell differential and serve as biomarkers of OMLDT. Expression profiles of Th1, Th2, Th17 and Treg cell type-specifying transcription factors and cytokines were analyzed using real time quantitative PCR (RT-qPCR) assay. To explore whether such expression profiles reflected their steady state plasma levels, a Luminex liquid fluorescence analysis was conducted. We found that the expression of transcription factors FoxP3 transcription factors (P = 0.006 and P < 0.0001) and IL-10 cytokine (P = 0.0008 and P < 0.0001) of the Treg subset were significantly higher in patients than TCE exposure workers and unexposed controls, suggesting that Treg cells were active after the occurrence of OMLDT. The transcript levels of IL-6 were significantly lower in the TCE exposure groups including patients and exposure workers as compared to the unexposed controls (P < 0.0001 and P = 0.0008). Circulating levels of assessed cytokines of IL-6 (P = 0.001 and P = 0.011) and TFN-α (P = 0.005 and P < 0.0001) were lower in the exposure groups than in the unexposed controls. Compared to the controls, the levels of IL-10 in patients were higher (P = 0.001 and P = 0.0008). There was a significantly positive correlation between the plasma levels IL-6 and IL-10 in TCE exposed workers. These alterations in the expression of transcription factors and cytokines highlight the underlying dysregulation of T cell subsets in OMLDT that reflect an immune tolerance or immune inhibition. Therefore, the elevation of IL-10 level may be a kind of pathogenesis indicator, and the decline in IL-6 level may be a kind of TCE exposure biomarker. These biomarkers need additional longitudinal follow-up studies to warrant to clinically useful biomarkers of OMLDT.


Subject(s)
CD4-Positive T-Lymphocytes/drug effects , Cytokines/biosynthesis , Dermatitis, Occupational/metabolism , Drug Eruptions/metabolism , Trichloroethylene/toxicity , Adult , Biomarkers , Female , Forkhead Transcription Factors/biosynthesis , Forkhead Transcription Factors/blood , Forkhead Transcription Factors/genetics , Humans , Interleukin-10/biosynthesis , Interleukin-10/blood , Interleukin-10/genetics , Interleukin-6/biosynthesis , Interleukin-6/blood , Interleukin-6/genetics , Male , Solvents/toxicity , T-Lymphocyte Subsets , Young Adult
5.
Article in English | WPRIM (Western Pacific) | ID: wpr-758813

ABSTRACT

The subtype H9N2 avian influenza virus greatly threatens the Chinese poultry industry, even with annual vaccination. Waterfowl can be asymptomatically infected with the H9N2 virus. In this study, three H9N2 virus strains, designated A/Goose/Jiangsu/YZ527/2011 (H9N2, Gs/JS/YZ527/11), A/Goose/Jiangsu/SQ119/2012 (H9N2, Gs/JS/SQ119/12), and A/Goose/Jiangsu/JD564/2012 (H9N2, Gs/JS/JD564/12), were isolated from domestic geese. Molecular characterization of the three isolates showed that the Gs/JS/YZ527/11 virus is a double-reassortant virus, combining genes of A/Quail/Hong Kong/G1/97 (H9N2, G1/97)-like and A/Chicken/Shanghai/F/98 (H9N2, F/98)-like; the Gs/JS/SQ119/12 virus is a triple-reassortant virus combining genes of G1/97-like, F/98-like, and A/Duck/Shantou/163/2004 (H9N2, ST/163/04)-like. The sequences of Gs/JS/JD564/12 share high homology with those of the F/98 virus, except for the neuraminidase gene, whereas the internal genes of Gs/JS/YZ527/11 and Gs/JS/SQ119/12 are closely related to those of the H7N9 viruses. An infectivity analysis of the three isolates showed that Gs/JS/SQ119/12 and Gs/JS/YZ527/11 replicated well, with seroconversion, in geese and chickens, the Gs/JS/JD564/12 did not infect well in geese or chickens, and the F/98 virus only infected chickens, with seroconversion. Emergence of these new reassortant H9N2 avian influenza viruses indicates that these viruses can infect both chicken and goose and can produce different types of lesions in each species.


Subject(s)
Animals , Humans , Asian People , Chickens , Geese , Influenza A Virus, H7N9 Subtype , Influenza A Virus, H9N2 Subtype , Influenza in Birds , Neuraminidase , Population Characteristics , Poultry , Sequence Analysis , Seroconversion , Vaccination
6.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-605411

ABSTRACT

Objective To investigate the clinical application of ALT detection based on microtiter plate kinetic method in the au‐tomatic enzyme immunoassay system .Methods By beference to the IFCC recommended kinetic method ,microtiter plate kinetic method was established in the automatic enzyme immunoassay system of ALT detection .The linear ,intro‐batch and inter‐batch du‐plicability of the method were evaluated .ALT test results of 823 samples with microtiter plate kinetic method and automatic bio‐chemistry analyzer method were compared .Results Microtiter plate kinetic method had good linearity where ALT activity <303 U/L ,the intra batch and inter batch variation coefficients < 1/5TEa .The detection results of the clinical samples were correlated with the biochemical analyzer .Conclusion Microtiter plate kinetic method to detection ALT in the automatic enzyme immunoassay system is an ideal method for simultaneous detection of ALT and ELISA in large batch samples ,which is worth popularizing .

7.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-506440

ABSTRACT

Objective To construct a recombinant Escherichia coli ( E. coli) with surface-dis-played lead specific binding protein PbrR and to further study intestinal colonization by the recombinant bac-teria in mice and gastrointestinal tolerance of the bacterial surface-displayed PbrR. Methods Chimeric pro-tein Lpp-OmpA coding sequence was chemically synthesized and inserted into the expression vector pET-21a to construct the outer membrane display vector pLOA. PbrR coding sequence was also obtained by chemical-ly synthesis and inserted into pLOA to generate the outer membrane display plasmid pLOA-pbrr. E. coli BL21 (DE3)pLysS was transformed with pLOA-pbrr and induced by IPTG. The expressed recombinant proteins were analyzed by 15% SDS-PAGE and Western blot assay. Lead adsorption capacity of the cell surface-dis-played PbrR in the simulated intestinal juice and tolerance of the recombinant E. coli to simulated gastric juice were analyzed, respectively. KM mice were orally given the induced recombinant bacteria by gastric lavage for 7 consecutive days and then were continually fed until day 30. The contents of recombinant bacte-ria in stool samples were detected by dilution plate method on day 7, 15 and 30. The recombinant protein with His tag was detected by immunoblotting on day 7 and 15. Results Based on Lpp-OmpA, the PbrR outer membrane display vector was successfully constructed. The recombinant fusion protein Lpp-OmpA-PbrR-His tag was highly expressed in E. coli. The recombinant E. coli strains displaying PbrR on their outer membrane accumulated a significant level of Pb2+ in simulated intestinal juice. Moreover, those strains showed a tolerance to gastric acid in vitro and could colonize in the intestinal tracts of mice via oral infection. The surface-displayed recombinant fusion protein showed a better tolerance to the environment of digestive tract. Conclusion The recombinant E. coli strain displaying PbrR on its surface showed a stronger capabili-ty of lead accumulation from simulated intestinal environment and could colonize in the intestinal tracts of mice. The surface-displayed recombinant PbrR also showed a good tolerance to digestive juice. This study paved the way for further researches on the selective elimination of lead by biosorption based on animal mod-els.

8.
Health Phys ; 109(4): 323-6, 2015 Oct.
Article in English | MEDLINE | ID: mdl-26313591

ABSTRACT

The scoring of the cytokinesis-block micronucleus and dicentric chromosomes in human peripheral blood lymphocytes is used as a dosimeter of radiation exposure. A detailed methodology is presented for human whole blood microculture for cytogenetic analysis. The technique described yields more than sufficient numbers of mitotic lymphocytes for analyzing micronuclei and chromosome aberrations following exposure to radiation.


Subject(s)
Biological Assay/methods , Chromosome Aberrations/radiation effects , Cytogenetic Analysis/methods , Leukocytes, Mononuclear/cytology , Leukocytes, Mononuclear/radiation effects , Radiation Monitoring/methods , Absorption, Radiation , Cells, Cultured , Female , Humans , Leukocytes, Mononuclear/physiology , Male , Middle Aged , Radiation Dosage , Reproducibility of Results , Sensitivity and Specificity
9.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-350538

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the cytotoxicity of indium chloride (InCl₃) and its effects on micro-nucleus formation in primary human lymphocytes cultured in vitro.</p><p><b>METHODS</b>The CCK-8 assay was used to evaluate the cytotoxicity of 24 h exposure to different concentrations of InCl₃(4, 40, 80, 200, 500, and 1 000 µmol/L) in lymphocytes cultured in vitro. The cytokinesis-block method was used to determine the micronucleus level in lymphocytes exposed to different concentrations of InCl₃and the effects of anti-oxidant vitamin C on micronucleus frequency.</p><p><b>RESULTS</b>Lymphocytes exposed to InCl₃of no less than 500 µmol/L had significantly lower survival rates than those in the control group (P < 0.05). Lymphocytes exposed to 80 µmol/L InCl₃had a significantly higher micronucleus frequency than those in the control group (P < 0.05). However, there was no further increase in micronucleus frequency of lymphocytes exposed to 200 µmol/L InCl₃. Lymphocytes cultured in whole blood and exposed to 500 or 1000 µmol/L InCl₃had a significantly increased micronucleus frequency than those in the control group (P < 0.001). The increase in micronucleus frequency of lymphocytes induced by indium could be partially antagonized by 20 or 100 µmol/L vitamin C.</p><p><b>CONCLUSION</b>InCl₃can induce an increase in micronucleus frequency of primary human lymphocytes cultured in vitro, which might be associated with DNA damage induced by oxidative stress.</p>


Subject(s)
Humans , Cell Nucleus , Metabolism , Cytokinesis , DNA Damage , In Vitro Techniques , Indium , Toxicity , Lymphocytes , Oxidative Stress
10.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-326030

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the significance of monitoring procalcitonin (PCT) when applying antibiotics to trichlorethylene (TCE)-induced dermatitis.</p><p><b>METHODS</b>One hundred and two patients who were hospitalized and recovered from TCE-induced dermatitis in our hospital from 2006 to 2013 were enrolled as subjects. Based on whether the PCT level was monitored or not, we divided patients into regular group and PCT group. For the regular group, we applied antibiotic treatment and determined the course of treatment based on clinical symptoms, laboratory test results, medical imaging results, and bacterial culture. For the PCT group, in addition to the above treatments, antibiotic treatment was applied when the PCT level was not lower than 0.25 ng/ml and stopped when the PCT level was lower than 0.25 ng/ml. The distribution of bacterial infection sites, type of bacteria, type of antibiotics, average period of hospitalization, and course of antibiotic treatment were compared between the two groups.</p><p><b>RESULTS</b>There were no significant differences in the distribution of bacterial infection sites, type of bacteria, type of antibiotics, and average period of hospitalization between the two groups (P > 0.05). The course of antibiotic treatment for the PCT group was significantly shorter than that for the regular group (25.37 ± 11.66 vs 20.58 ± 7.53 d, P < 0.05).</p><p><b>CONCLUSION</b>Under similar conditions of bacterial infection, antibiotic treatment of TCE-induced dermatitis based on the serum PCT level can significantly shorten the course of treatment and avoid the abuse of antibiotics.</p>


Subject(s)
Humans , Anti-Bacterial Agents , Therapeutic Uses , Bacteria , Bacterial Infections , Calcitonin , Calcitonin Gene-Related Peptide , Drug Eruptions , Drug Therapy , Hospitalization , Monitoring, Physiologic , Protein Precursors , Trichloroethylene , Toxicity
11.
Chongqing Medicine ; (36): 4504-4506, 2015.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-479664

ABSTRACT

Objective To explore the risk factors of skeletal related events (SREs) in non small cell lung cancer with bone metastases and its effect on the prognosis .Methods Totally 223 cases of NSCLC patients with bone metastasis were retrospective studied from January 2010 to December 2012 in our hospital .The clinical features ,predictive factors for SREs were analysed by sin‐gle factor and multifactor analysis .Results Among 223 cases of NSCLC patients with bone metastasis ,119 cases occured with SREs(53 .4% ) .Univariate analysis showed that the occurrence of SREs in female ,no smoker ,adenocarcinoma ,solitary bone metas‐tasis lesions were less than the male ,smoker non‐adenocarcinoma ,and multiple bone metastases (P0 .05) .The multivariate analysis revealed only multiple bone metastases was an independent risk factor for SREs .The median survival time of the NSCLC patients with bone metastasis was 15 .3 months .Moreover ,survival analysis showed that SREs had no statistical significance on the prognosis of bone metastasis in NSCLC patients (P>0 .05) .Conclusion The female ,adenocarcinoma ,smoking history ,solitary bone metastasis lesions occurred in patients with lower risk SREs .Multiple bone metastasis is an independent risk factor for SREs ,attention should be paid to monitoring and prevention .

12.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-435275

ABSTRACT

Two kinds of incorrect cultivation modes were existed since the launch of cultivation for professional degree postgraduate of clinical medicine:taking professional degree postgraduates as scientific degree postgraduates to cultivate or taking scientific degree postgraduates as professional degree postgraduates to cultivate.In this paper,taking the cultivation of oncologic professional degree postgraduates as example,we put forward some suggestions:strengthening clinical skill training,promoting thinking mode of evidence-based medicine and enhancing clinical trial and statistical training.

13.
Chinese Journal of Dermatology ; (12): 519-521, 2011.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-416995

ABSTRACT

Objective To investigate the expression of peroxisome proliferator-activated receptors (PPARs) in epidermal keratinocytes of patients with psoriasis vulgaris and its significance.Methods Immunohistochemical method was used to examine the expression of PPARα,β/δand.γ in tissue specimens from the normal skin of 5 human controls,lesional and normal skin adjacent to the lesions of 17 patients with psoriasis vulgaris.A semi-quantitative analysis was carried out by image analyzer.Results Different levels of expression of the three PPAR isotypes were observed in the nuclei of epidermal keratinocytes of normal human controls.The expression intensity of PPAR α,β/δand γ was statistically higher in the epidermis of adjacent normal skin than that in psoriatic lesions(all P<0.01)and normal control skin(all P<0.01).Conclusions The decreased expression of PPARα may be associated with the overproliferation and parakeratosis of epidermal keratinocytes in psoriatic lesions,and PPAR β/δ and γ may display a synergistic effect on the maintenance of homeostatic proliferation and difierentiafion of epidermal cells.

14.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-283259

ABSTRACT

<p><b>OBJECTIVE</b>To try to find the ways to enhance the expression of ADS gene encoding amorpha-4,11-diene synthase, a key enzyme in artemisinin biosynthesis pathway catalyzing the formation of amorpha-4,11-diene from farnesyl diphosphate, and accelerate the artemisinin synthesis, the promoter of ADS was isolated and characterized.</p><p><b>METHOD</b>5' untranslated regions of ADS were isolated from Artemisia annua with PCR. For functional characterization, the isolated fragment was fused with GUS reporter gene and introduced into Nicotiana tabacum by Agrobacterium-mediated transformation. The GUS expression regulated by 5' untranslated regions of ADS in transgenic N. tabacum under the normal or stressed conditions were detected by histochemical staining and quantitative spectrophotometry assay.</p><p><b>RESULT</b>The 2 448 bp DNA fragment upstream of ADS coding sequence was isolated from A. annua and introduced into N. tabacum. Histochemical staining showed that the isolated fragment conferred stable GUS expression in transgenic plants. The quantitative results showed that the GUS activity in transgenic tobacco plants treated by low-temperature (4 degrees C) and ultraviolet irradiation were 1. 6 and 2.2 folds higher than that in the controls.</p><p><b>CONCLUSION</b>It was suggested that the isolated fragment had promoter activity and maybe responsive to adverse environmental stresses.</p>


Subject(s)
5' Untranslated Regions , Genetics , Alkyl and Aryl Transferases , Genetics , Metabolism , Artemisia annua , Genetics , Gene Expression Regulation, Plant , Genetic Vectors , Genetics , Molecular Sequence Data , Promoter Regions, Genetic , Genetics
15.
Chinese Journal of Biotechnology ; (12): 1312-1316, 2008.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-275384

ABSTRACT

A single-stranded oligonucleotides containing a 6 histidine sequence, a hydroxylamine cleavage site, a thrombin cleavage site, and stop codon TAA were inserted into the polylinker's downstream of prokaryotic expression vector pBV220 between BamHI and PstI. The resultant vector is named pBV223. Proteins expressed in this vector will have a 6 histidine tail as affinity handy fused to their C terminus and can be quickly purified by one step immobilized metal affinity chromatography (IMAC). This plasmid is verified by restriction map and DNA sequencing. Subsequently, the metastasis suppressor gene nm23-H1 cDNA (without the stop codon) was cloned into vector PBV223 in frame with the 6-histidine sequence, hydroxylamine and thrombin cleavage sites. The soluble nm23-H1 fusion protein was successfully induced in the bacterial DH5a and easily purified with Ni chromatograph. These results indicated that the strategy to clone the single-stranded oligonucleotides directly into the restriction sties between BamH I and Pst I in the pBV220 vector is the simplest and cost-effective method.


Subject(s)
Base Sequence , Chromatography, Affinity , Methods , Cloning, Molecular , DNA, Complementary , Genetics , Escherichia coli , Genetics , Metabolism , Genetic Vectors , Genetics , Molecular Sequence Data , Mutation , NM23 Nucleoside Diphosphate Kinases , Genetics , Recombinant Fusion Proteins , Genetics
16.
Cancer Research and Clinic ; (6): 303-305,309, 2008.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-597143

ABSTRACT

Objective To analyze the association between C12 tumor markers and colorectal cancer,in order to screen for colorectal cancer related tumor markers so as to provide theoretical base for the establishment of colorectal cancer diagnostic biochips. Methods The sera of 173 colorectal cancer patients were detected for 12 common tumor markers including carcinoembryonic antigen (CEA), alpha-fetoprotein (AFP), carbohydrate antigen 19-9 (CA19-9), carbohydrate antigen 242 (CA242), cancer antigen 15-3 (CA15-3), cancer antigen 125 (CA125), prostate specific antigen (PSA), free-PSA, neuron-specific enolase (NSE),human chorionic gonagotropin-beta (β-HCG), human growth hormone (HGH), and ferritin using the C12tumor markers proteinchip, and colorectal cancer related parameters were analyzed by Kappa test and cost-effectiveness analysis to find the most optimal tumor marker program. Results CEA (36.4 %), CA242(19.7 %), CA19-9 (18.5 %), CA125(9.8 %) were major tumor markers increased among the 173 colorectal cancer patients. Kappa test revealed 7 tumor marker programs having strong consistency with the detection results of C12 tumor markers proteinchip, and CEA singly detected was proved to be the best program by cost-effectiveness analysis. Conclusion C12 tumor markers proteinchip system have limited value in the diagnosis of colorectal cancer, but the design of chip is too complicated and costly for widespread screening among the high risk populations. Searching for new colorectal cancer biomarkers and designing small diagnostic chip could significantly enhance the clinical value of tumor markers in terms of diagnostic rate and practical utility.

17.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-580176

ABSTRACT

Objective:To construct prokaryotic expression vector of mutant nm23-H1(S120G,S120G-P96S, S44A), and then express and purify the proteins. Methods: Mutant nm23-H1 fragments were amplified by PCR. The prokaryotic expression vectors of pET28a-nm23-H1 were constructed by gene recombination technique and verified by restriction enzyme analysis and sequencing. The positive clones were transformed into E. coli BL21(DE3) and soluble analysis of the expression was conducted in these systems. The proteins were purified by nickel column chromatography and identified by Western blot. Results: The sequences and open read frames of all the pET28a-nm23-H1 were completely correct. After transforming, these plasmids could express the target proteins. The protein production was very high. Among these mutant proteins, S44A was soluble expression and S120G was partly soluble. However, S120G-P96S was mostly in the inclusion bodies. The molecular weight of these mutant proteins was 20 kD detected by Western blot, which was as the same as the objective protein. Conclusion: We have succeeded in constructing the prokaryotic expression vectors of pET28a-nm23-H1(S120G, S120G-P96S, S44A), and the S120G and S44A proteins can be used in following studies. However, S120G-P96S protein which expressed in this system may not be suitable for the following studies.

18.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-339325

ABSTRACT

<p><b>BACKGROUND</b>Ras to MAPK pathway plays a critical role in the transmission of many growth and developmental signals. A new component of this pathway which is termed kinase suppressor of Ras (KSR) was found in 1995. KSR is as a scaffolding protein that coordinates the assembly of a multiprotein complex containing mitogen-activated protein kinase (MAPK) and its upstream regulators. It has been proven that KSR has many phosphorylation sites, and phosphorylation state changes response to signaling events. Site-directed mutagenesis can precisely change the base sequence and get mutant proteins. The aim of this study is to construct two mutant proteins of KSR by using site-directed mutagenesis, and to express and purify them, therefore to provide basement for studying the functional and biochemical mechanisms of KSR.</p><p><b>METHODS</b>Site-directed mutagenesis of pCMV-Tag2b-KSR gene was performed by modified QuikChangtm site-directed mutagenesis kit method. Two pairs of mutagenic primers were synthesized in vitro and two mutations desired, the recombinant plasmids were verified by restriction enzyme analysis and DNA sequencing. Then positive clones were transfected into 293T cell line. The purified mutant proteins were analyzed by Western blot.</p><p><b>RESULTS</b>Two mutants were successfully constructed. The results of DNA sequencing confirmed that the base sequences of the mutant genes were completely concordant with experiment design, which could be used to be transfected into 293T cell line. The purified mutants were identified by Western blot.</p><p><b>CONCLUSIONS</b>Two mutant KSR genes are successfully constructed. It provides experimental basement for further functional research of KSR.</p>

19.
Chinese Journal of Lung Cancer ; (12): 296-300, 2007.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-339286

ABSTRACT

<p><b>BACKGROUND</b>According to the report of the 11th World Conference on Lung Cancer, lung cancer is the leading cause of cancer related death. So there is important clinical significance to monitor the patients with lung cancer through different ways. The aim of this study is to investigate the clinical significance of multiple tumor marker protein chip in monitoring the recurrence, progression and metastasis of lung cancer.</p><p><b>METHODS</b>Forty-four patients were selected, who were detected at least 4 times with tumor mar-ker protein chip. Based on efficacy and status, patients were classified as six grades. Correlation of expression level of each tumor marker with grade of efficacy and status was analyzed. And the discriminant functions for recurrence, progression and metastasis of lung cancer were established.</p><p><b>RESULTS</b>Grade of efficacy and status was closely related to CA199, CEA, CA242, AFP and CA125 in adenocarcinoma (AC), to CA125 in squamous cell carcinoma (SqCa), and to CA199 and CA125 in small cell lung cancer (SCLC). Based on the discriminant functions, accuracy rate of efficacy and status judgement was 89.4%, 80.4%, 78.3% and 66.7% for female AC, male AC, SqCa and SCLC respectively.</p><p><b>CONCLUSIONS</b>There is important clinical significance of multiple tumor marker protein chip in monitoring the recurrence, progression and metastasis of lung cancer (especially adenocarcinoma).</p>

20.
Article in Chinese | WPRIM (Western Pacific) | ID: wpr-313296

ABSTRACT

<p><b>BACKGROUND</b>The results of our previous studies have proven that nm23-H1 gene can suppress metastasis of lung cancer, which may be associated with suppression of the Wnt signal pathway through up-regulating the activity of glycogen synthase kinase 3β (GSK-3β), the key kinase of the Wnt signal pathway. The aim of this study is to investigate the effect of point mutation of nm23-H1 gene on GSK-3β activity in cytoplasm and nucleus in human high-metastatic large cell lung cancer cell line L9981.</p><p><b>METHODS</b>Using immunoprecipitation and a radioactive isotope scintillation counter, the activity of GSK-3β was detected in cytoplasm and nucleus of human low-metastatic large cell lung cancer cell line NL9980, human high-metastatic large cell lung cancer cell line L9981, L9981-pEGFP (transfected with vector), L9981-nm23-H1-pEGFP (transfected with wild type nm23-H1), L9981-nm23-H1 S44A -pEGFP mutant (transfected with serine 44 to alanineon of nm23-H1 gene), L9981-nm23-H1 P96S -pEGFP mutant (transfected with proline 96 to serine of nm23-H1 gene), L9981-nm23-H1 H118F -pEGFP mutant (transfected with histidine 118 to phenylalanine of nm23-H1 gene) and L9981-nm23-H1 S120G -pEGFP mutant (transfected with serine 120 to glycine of nm23-H1 gene).</p><p><b>RESULTS</b>The GSK-3β activity in cytoplasm and nucleus was remarkably decreased in the transgene lung cancer cell lines transfected with mutant nm23-H1 cDNA (L9981-nm23-H1 S44A -pEGFP, L9981-nm23-H1 P96S -pEGFP, L9981-nm23-H1 H118F -pEGFP and L9981-nm23-H1 S120G -pEGFP). Significant differences of GSK-3β activity in cytoplasm and nucleus were observed (P < 0.05) when L9981-nm23-H1-pEGFP cell line was compared with L9981-nm23-H1 S44A -pEGFP, L9981-nm23-H1 P96S -pEGFP, L9981-nm23-H1 H118F -pEGFP and L9981-nm23-H1 S120G -pEGFP lung cancer cell lines. There was a highly significant difference in GSK-3β activity in the cytoplasm between L9981-nm23-H1-pEGFP cell line and L9981-nm23-H1 P96S -pEGFP lung cancer cell line (P < 0.01 ). A highly significant difference in GSK-3β activity in the nucleus was observed (P < 0.01) when L9981-nm23-H1-pEGFP lung cancer cell line was compared with L9981-nm23-H1 S44A -pEGFP, L9981-nm23-H1 P96S -pEGFP and L9981-nm23-H1 H118F -pEGFP lung cancer cell lines.</p><p><b>CONCLUSIONS</b>(1) Point mutation of nm23-H1 gene can significantly influence the regulating effects on the GSK-3β activity in the cytoplasm and nucleus in the human high-metastatic large cell lung cancer cell line L9981. (2) The effects of nm23-H1 gene on metastatic phenotype may be related to the upregulation of GSK-3β activity in human high-metastatic large cell lung cancer cell line L9981.</p>

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