RESUMO
Bcr-Abl tyrosine kinase inhibitors (TKIs) have been a remarkable success for the treatment of Ph(+) chronic myeloid leukemia (CML). However, a significant proportion of patients treated with TKIs develop resistance because of leukemia stem cells (LSCs) and T315I mutant Bcr-Abl. Here we describe the unknown activity of the natural product berbamine that efficiently eradicates LSCs and T315I mutant Bcr-Abl clones. Unexpectedly, we identify CaMKII γ as a specific and critical target of berbamine for its antileukemia activity. Berbamine specifically binds to the ATP-binding pocket of CaMKII γ, inhibits its phosphorylation and triggers apoptosis of leukemia cells. More importantly, CaMKII γ is highly activated in LSCs but not in normal hematopoietic stem cells and coactivates LSC-related ß-catenin and Stat3 signaling networks. The identification of CaMKII γ as a specific target of berbamine and as a critical molecular switch regulating multiple LSC-related signaling pathways can explain the unique antileukemia activity of berbamine. These findings also suggest that berbamine may be the first ATP-competitive inhibitor of CaMKII γ, and potentially, can serve as a new type of molecular targeted agent through inhibition of the CaMKII γ activity for treatment of leukemia.
Assuntos
Benzilisoquinolinas/farmacologia , Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Células-Tronco Neoplásicas/efeitos dos fármacos , Trifosfato de Adenosina/química , Trifosfato de Adenosina/metabolismo , Animais , Apoptose/efeitos dos fármacos , Benzamidas , Benzilisoquinolinas/química , Benzilisoquinolinas/metabolismo , Western Blotting , Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina/química , Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina/genética , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Proteínas de Fusão bcr-abl/antagonistas & inibidores , Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Células HEK293 , Humanos , Mesilato de Imatinib , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Modelos Moleculares , Mutação , Células-Tronco Neoplásicas/metabolismo , Piperazinas/farmacologia , Ligação Proteica , Inibidores de Proteínas Quinases/farmacologia , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Pirimidinas/farmacologia , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Tumor protein 53 (TP53) mutation predicts an unfavorable prognosis in diffuse large B-cell lymphoma (DLBCL), but the molecular basis for this association remains unclear. In several malignancies, the cytidine deaminase apolipoprotein B mRNA editing enzyme catalytic subunit 3B (APOBEC3B) has been reported to be associated with the TP53 G/C-to-A/T mutation. Here, we show that the frequency of this mutation was significantly higher in relapsed/refractory (R/R) than in non-R/R DLBCL, which was positively associated with the APOBEC3B expression level. APOBEC3B overexpression induced the TP53 G/C-to-A/T mutation in vitro, resulting in a phenotype similar to that of DLBCL specimens. Additionally, APOBEC3B-induced p53 mutants promoted the growth of DLBCL cells and enhanced drug resistance. These results suggest that APOBEC3B is a critical factor in mutant p53-driven R/R DLBCL and is therefore a potential therapeutic target.
Assuntos
Linfoma Difuso de Grandes Células B , Proteína Supressora de Tumor p53 , Citidina Desaminase/metabolismo , Humanos , Linfoma Difuso de Grandes Células B/patologia , Antígenos de Histocompatibilidade Menor/genética , Mutação , Prognóstico , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
OBJECTIVE: To construct eukaryotic expression plasmids containing green fluorescent protein gene and CYP19 wild-type or its variants (W39R, R264C, W39R-R264C) and to observe its expression in MCF-7 and Bcap-37 cells. METHODS: The aromatase WT cDNA sequence was obtained by RT-PCR amplification and cloned into the eukaryotic expression vector pcDNA3.1(+). pcDNA3.1(+)-CYP19-GFP plasmid was then used as the template for site-directed mutation to create variant constructs (W39R, R264C, W39R-R264C). pcDNA3.1(+)-CYP19-GFP was transfected and expressed in MCF-7 and Bcap-37 cells. RESULT: The construction of pcDNA3.1(+)-CYP19-GFP plasmid was confirmed by enzyme digestion and DNA sequencing. pcDNA3.1(+)-CYP19(W39R)-GFP, pcDNA3.1(+)-CYP19(R264C)-GFP, pcDNA3.1(+)- CYP19(W39R-R264C)-GFP plasmids were confirmed by DNA sequencing. The MCF-7 and Bcap-37 cells transfected with the pcDNA3.1(+)-CYP19-GFP plasmid expressed reporter gene of GFP. CONCLUSION: The eukaryotic expression plasmids have been constructed and expressed in MCF-7 and Bcap-37 cells successfully, which lays the foundation for the research of biological activities of CYP19 variant allozymes.
Assuntos
Aromatase/genética , Vetores Genéticos , Plasmídeos/genética , Linhagem Celular Tumoral , Proteínas de Fluorescência Verde/genética , Humanos , Mutagênese Sítio-Dirigida , Proteínas Recombinantes de Fusão/genética , TransfecçãoRESUMO
Hepatoma-derived growth factor (HDGF) is related to tumorigenesis and the development of cancer; it is an independent factor associated with the prognosis of liver cancer, non-small cell lung cancer and pancreatic cancer. However, the molecular mechanism by which HDGF participates in gastric carcinogenesis and development as well as its functional regulation during the development of gastric precancerous lesions needs to be further analyzed. In the present study, we analyzed the effect of HDGF transfection on the proliferation and on the changes of mitogen-activated protein kinase (MAPK), Akt, and nuclear factor-kB (NF-kB) pathways in gastric cancer AGS cells. HDGF transfection significantly activated Erk1/2 in AGS cells and promoted anchorage-independent growth. Further studies showed that HDGF expression gradually increased in the gastric carcinogenesis process and HDGF showed a high expression in poorly differentiated adenocarcinoma prone to lymphoid metastasis; these findings suggest that HDGF is involved in the gastric carcinogenesis process and promotes proliferation and metastasis via Erk1/2 activation.
Assuntos
Transformação Celular Neoplásica/metabolismo , Mucosa Gástrica/enzimologia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Neoplasias Gástricas/enzimologia , Adenocarcinoma/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Mucosa Gástrica/patologia , Humanos , NF-kappa B/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , TransfecçãoRESUMO
AIM: To investigate the expression of SNC73, a trans-cript of the immunoglobulin alpha-1 gene (IgA1-H chain), in human epithelia-derived tumor cells. METHODS: Total RNAs and cell lysates were prepared from five different human epithelial cell lines derived from lung, stomach, liver, skin, and breast, respectively. RT-PCR and immunoblot analysis of these five cell lines were done. Both RT-PCR and immunochemistry were used to detect the expression of SNC73 in these cell lines. We also examined the expression of SNC73 in normal epithelial cells of colon mucosa by in situ hybridization. RT-PCR and immunoblot analysis were used to determine whether the recombination activating gene1/2 (RAG1 and RAG2) is present. The expression of three immunoglobulin transcription factors, EBF, E2A and Pax5, and the heavy chain of IgA1 and two types of light chains of immunoglobulin (kappa and lambda) in the aforementioned cell lines were analyzed by RT-PCR and immunochemistry, respectively. All the RT-PCR products were analyzed by sequencing. RESULTS: The results of RT-PCR and immunochemistry showed that both mRNA and protein of SNC73 were expressed in five human epithelia-derived cancer cell lines. These data were further confirmed in the normal epithelial cells of colon mucosa by in situ hybridization. Also, the heavy chain of IgA1 and kappa light chain were detected in these cells, but no lambda light chain was obse-rved. Both RAG1 and RAG2 were expressed in these human epithelia-derived cancer cell lines and the sequence was identical to that expressed in pre-B and pre-T cells. In addition to RAG1 and RAG2, the mRNA in one of the immunoglobulin transcription factors, EBF, was also detected in these cell lines, and Pax5 was only expressed in SW480 cells, but no expression of E2A was observed in all the five cell lines. CONCLUSION: Immunoglobulin A1 is originally expressed and V(D)J recombination machine is also present in non-lymphoid cells, suggesting that V(D)J recombination machine mediates the assembly of immunoglobulin A1 in non-lymphoid cells as in pre-lymphocytes.
Assuntos
Neoplasias da Mama/metabolismo , Neoplasias Colorretais/metabolismo , Imunoglobulinas/metabolismo , Neoplasias Hepáticas/metabolismo , Neoplasias do Colo do Útero/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Colo/metabolismo , Colo/patologia , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Células Epiteliais/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Células HeLa , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Imunoglobulinas/genética , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fator de Transcrição PAX5/genética , Fator de Transcrição PAX5/metabolismo , Transativadores/genética , Transativadores/metabolismo , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia , Éxons VDJ/genéticaRESUMO
In our previous study, we demonstrated that azurin could selectively trigger apoptosis in human osteosarcoma cell line U2OS cells. However, the rate of apoptosis (35.8 +/- 3.2%) is not very high, and azurin is too expensive to obtain readily. To solve these problems, we constructed a eukaryotic expression plasmid containing the azurin gene with an influenza virus haemagglutinin 9 peptide HA epitope tag, and transfected the recombinant plasmid pcDNA3.1(+)/azurin into U2OS cells. RT-PCR and Western blot analysis validated the successful transfection and the expression of the azurin-HA protein. Conspicuous apoptosis of the transfected cells was detected by flow cytometry (FCM) and the DNA ladder test. The apoptosis rate reached 64.3 +/- 13.1%. The transcriptional levels of the Bax and p53 genes increased significantly in U2OS cells transfected with pcDNA3.1(+)/azurin, but the Bcl-2 mRNA level decreased. There was no difference in the levels of Bcl-xl mRNA and Survivin mRNA. We propose that the transfection of the recombinant plasmid pcDNA3.1(+)/azurin can significantly induce apoptosis in U2OS cells. This is closely associated with the up-regulation of the transcriptional level of the Bax and p53 genes, and the down-regulation of that of the Bcl-2 gene.
Assuntos
Apoptose , Azurina/metabolismo , Células Eucarióticas/metabolismo , Expressão Gênica , Plasmídeos/metabolismo , Transfecção/métodos , Western Blotting , Ciclo Celular , Linhagem Celular Tumoral , Fragmentação do DNA , Citometria de Fluxo , Regulação Neoplásica da Expressão Gênica , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Humanos , Proteínas Inibidoras de Apoptose/genética , Proteínas Inibidoras de Apoptose/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Survivina , Transcrição Gênica , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo , Proteína bcl-X/genética , Proteína bcl-X/metabolismoRESUMO
OBJECTIVE: To investigate the mechanism of apoptosis of chronic myeloid leukemia (CML) cells induced by the novel p210 bcr/abl inhibitor berbamine. METHODS: Human Ph+ CML leukemia K562 cells, which express endogenous p210 bcr/abl protein, were cultured in RPMI 1640 and treated with berbamine as indicated time and dose. Flow cytometry (FCM) and Annexin-V-Fluos/PI staining kit were used to evaluate the apoptosis of leukemic cells; FCM and cytoperm/cytofix plus Caspase-3-McAb-PE were employed to measure the leukemic cells with activated Caspase-3. Phosphorylation of p210 bcr/abl protein in the leukemic cells were assessed by a combination of immunoprecipitation (IP) with c-abl antibody and Western blotting with p-Tyr (pY99) antibody. The protein levels of p210 bcr/abl, Hsp90 and Hsp70 in the leukemic cells were determined by Western blotting with antibodies to c-abl, Hsp90, and Hsp70 respectively. RESULTS: After treatment with berbamine at 8 microg/ml for 48 h, the percentages of leukemic cells expressing activated caspase-3 and apoptotic cells were 45.69% and 48.43% respectively. IP and WB results showed that berbamine at low concentration markedly inhibited phosphorylation of p210 bcr/abl protein in the leukemia cells, and the amount of phosphorylated p210 bcr/abl in the leukemia cells exposed to berbamine at 8 microg/ml for 6 h were only 8.41% of that of untreated leukemia cells without the protein levels of p210 bcr/abl down-regulated. Significantly, berbamine also down-regulated chaperone Hsp90 protein, and the amount of Hsp90 protein in the leukemia cells treated with berbamine at 8 microg/ml for 48 h accounted for 18.37% of that of the untreated leukemia cells. Berbamine at 8 microg/ml had no obvious effect on chaperone Hsp70 protein expression associated with the resistance of leukemia cells to apoptosis. CONCLUSION: (1) Berbamine induces caspase-3-mediated apoptosis of Ph+ leukemia cells through inhibiting phosphorylation of p210 bcr/abl protein and down-regulating its chaperone Hsp90 protein. (2) Unlike Hsp90 inhibitor GA that upregulates Hsp70, berbamine has no obvious effect on chaperone Hsp70 protein expression in leukemia cells, suggesting that berbamine may be a novel class of Hsp90 inhibitor, and further study is required.
Assuntos
Alcaloides/uso terapêutico , Apoptose/efeitos dos fármacos , Benzilisoquinolinas/uso terapêutico , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Caspase 3/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Proteínas Oncogênicas v-abl/metabolismo , Fitoterapia , Proteína de Morte Celular Associada a bcl/metabolismoRESUMO
Human DNA polymerases kappa, eta, iota, and zeta are responsible for the translesion DNA synthesis. Numerous in vitro studies indicated that these enzymes may contribute to DNA lesion-triggered and spontaneous mutation. We measured the transcripts of these 4 enzymes in 131 self-paired cancerous and non-tumor samples, including 23 lung cancers, 49 stomach cancers, and 59 colorectal cancers. Our results indicated that, except pol eta in colorectal cancers, these enzymes are all significantly down-regulated in human lung, stomach, and colorectal cancers, suggesting that these enzymes are probably not closely associated with the elevated mutations in human cancer.
Assuntos
Neoplasias Colorretais/enzimologia , DNA Polimerase Dirigida por DNA/metabolismo , Neoplasias Pulmonares/enzimologia , Neoplasias Gástricas/enzimologia , Regulação para Baixo , Feminino , Humanos , Masculino , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
OBJECTIVE: To investigate the expression level of Shp-2 tyrosine phosphatase in chronic myeloid leukemia (CML) and its relationship with the unlimited growth and apoptosis resistance of p210 bcr/abl-induced malignant cells. METHODS: In this study, p210 bcr/abl positive leukemia cell specimens were obtained from 25 CML cases, meanwhile, bone marrow and peripheral blood cell samples from 8 non-tumor individuals and 10 normal individuals were used as p210 bcr/abl negative controls. K562 and KU812 leukemia cells were used as p210 bcr/abl positive controls, and KG-1 leukemia cell line was used as Shp-2 positive control. Specimens of peripheral blood and bone marrow of 25 adult patients of chronic myelocytic leukemia, 15 males and 10 females, aged 28-64, were collected. Specimens of bone marrow of 8 basically healthy adult volunteers and specimens of peripheral blood of 10 healthy adult volunteers were used as controls. The total cell protein was collected and the expression of Shp-2 was examined by Western blotting. Human leukemia cells of the line K562 were cultured. Shp-2 specific sense and antisense oligonucleotides were added into the culture fluid respectively. The cell apoptosis was detected by flow cytometry (FCM). STI571, specific inhibitor of p210 bcr/abl was added into the cultured fluid of K562 cells, then Western blotting and FCM were used to detect the protein expression of Shp-2 and p210 bcr/abl, and cell apoptosis. RESULTS: Phosphorylated Shp-2 (pShp)-2 protein was overexpressed in 92% (23/25) of the CML cells, but lowly expressed or not expressed in the normal hematopoietic cells. The mean pShp-2 protein/beta-actin ratio of the primary CML leukemia cells was 0.91 +/- 0.62, significantly higher than those of the normal bone marrow cells and peripheral blood hematopoietic cells (0.16 +/- 0.09 and 0.03 +/- 0.05 respectively, both P < 0.01). The apoptotic rates of the CML cells treated by Shp-2 specific antisense oligonucleotide of the concentrations of 1 micromol/L and 4 micromol/L respectively for 72 h was 7.98% and 20.29% respectively, both significantly higher than that of the control group (4.06%, P < 0.01). The number of clone of CML cells treated by 0.25 micromol/L and 1.0 micromol/L Shp-2 specific antisense oligonucleotide for 7 days were 67% (37/60) and 11.9% (5/42) that of the control group. Twenty-four and 48 hours after the stimulation of STI571 the expression level of Shp-2 protein in the CML cells decreased time-dependently and the CML cell apoptotic rates were 31.15% and 38.69% respectively, both lower than that of the control group (33.6%). The number of clone of CML cells effected by 0.25 micromol/L and 1.0 micromol/L Shp-2 specific antisense oligonucleotide for 7 days was 67% (37/60) and 11.9% (5/42) that of the control group. Twenty-four and 48 hours after the stimulation of STI571 the expression level of Shp-2 protein in the CML cells decreased time-dependently and the CML cell apoptotic rates were 31.15% and 38.69% respectively, both lower than that of the control group (33.6%). CONCLUSION: (1) The pShp-2 protein is overexpressed in CML cells, which is associated with the unlimited growth and apoptosis resistance of malignant cells. (2) Shp-2 is upregulated by p210 bcr/abl oncoprotein in CML.
Assuntos
Apoptose , Proteínas de Fusão bcr-abl/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 11/metabolismo , Adulto , Estudos de Casos e Controles , Proliferação de Células , Feminino , Humanos , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Masculino , Pessoa de Meia-Idade , Células Tumorais CultivadasRESUMO
AIM: To explore the activation of signal transduction pathways related with the carcinogenesis of sporadic colon cancers. METHODS: A gene array monitoring the activation of 8 signal transduction pathways (PathwayFinder GEArray) was used to screen the differentially expressed genes between colorectal cancer and normal colon tissues. The differentially expressed genes were further analyzed by RT-PCR, using RNA derived from colorectal cancer and normal colon tissue of 35 patients. RESULTS: The expression of HSF1, HSF27, HSP90 and iNOS was increased in colon cancer tissues compared to normal colon tissue using PathwayFinder GEArray. The RT-PCR results showed that the expression of HSF1 was increased in 86% (30/35) patients and the expression of iNOS was increased in 63% (22/35) patients. CONCLUSION: The induction of HSF1 gene expression is associated with sporadic colon cancer. HSF1 induces heat shock stress signaling pathway, which might play a role in the carcinogenesis of sporadic colorectal cancer.
Assuntos
Neoplasias Colorretais/genética , Neoplasias Colorretais/fisiopatologia , Proteínas de Ligação a DNA/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas de Ciclo Celular , Regulação Neoplásica da Expressão Gênica , Proteínas de Choque Térmico HSP90/genética , Fatores de Transcrição de Choque Térmico , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/genética , Óxido Nítrico Sintase/genética , Óxido Nítrico Sintase Tipo II , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/fisiologia , Fatores de Transcrição , Proteínas Supressoras de TumorRESUMO
OBJECTIVE: To explore the activation of the signal transduction pathways related with the carcinogenesis of sporadic colon cancers. METHODS: A gene microarray monitoring activation of 8 signal transduction pathways (PathwayFinder GEArray) was used to screen the differentially expressed genes between colorectal cancer and normal colon tissue. The differentially expressed genes were further analyzed by RT-PCR, using RNA extracted from cancer tissue and matched normal colon mucosa of 35 patients with colorectal cancer. RESULTS: The expression of hsf1, hsf27 and inos was increased in colon cancer compared with normal colon mucosa using PathwayFinder GEArray. The RT-PCR results showed that the expression of hsf1 was detected in 86% of patients(30/35)and the expression of inos detected in 63% patients(22/35). CONCLUSION: Hsf1 induces heat shock stress signaling pathway, which might play a role in the carcinogenesis of sporadic colorectal cancer.
Assuntos
Neoplasias Colorretais/metabolismo , Proteínas de Ligação a DNA/biossíntese , Proteínas do Tecido Nervoso/biossíntese , Transdução de Sinais/fisiologia , Proteínas de Ciclo Celular , Neoplasias Colorretais/genética , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Fatores de Transcrição de Choque Térmico , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas do Tecido Nervoso/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição , Proteínas Supressoras de Tumor/biossíntese , Proteínas Supressoras de Tumor/genéticaRESUMO
Linalool, a natural small molecule monoterpene, has been shown to have anti-tumor activity against several human tumor cell lines in vitro; however, the anti-leukemia spectrum and molecular mechanisms inhibiting tumor cell growth are not fully understood. In the present study, we demonstrated that linalool preferentially induced growth arrest and apoptosis of a variety of human leukemia cells, but spared normal hematopoietic cells. Treatment of leukemia cells by linalool for 12h led to strong activation of p53, cyclin-dependent kinase inhibitors (CDKIs), GADD45alpha, c-jun and phosphorylated-JNK, suggesting that linalool-induced apoptosis might be associated with activation of p53 and CDKIs. The findings here warrant further investigation of this class of natural product as lead compound for developing novel therapeutic agents for leukemia.
Assuntos
Apoptose/efeitos dos fármacos , Quinases Ciclina-Dependentes/antagonistas & inibidores , Monoterpenos/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Proteína Supressora de Tumor p53/metabolismo , Regulação para Cima/efeitos dos fármacos , Monoterpenos Acíclicos , Linhagem Celular Tumoral , HumanosRESUMO
Protein microarrays for parallel detection of multiple viral antigens and antibodies have not yet been described in the field of human hepatitis virus infections. Here, we describe a simple, rapid and sensitive integrated protein microarray with three different reaction models. The integrated protein microarray could simultaneously determine in human sera two viral antigens (HBsAg, HBeAg) and seven viral antibodies (HBsAb, HBcAb, HBeAb, HCVAb, HDVAb, HEVAb, HGVAb) of human hepatitis viruses within 20 min. The results of the protein microarray were assessed directly by the naked eye but can also be analyzed by a quantitative detector. The detection limit of this protein microarray was 0.1 ng/ml for HBsAg. Overall, >85% concordance was observed between the integrated protein microarrays and an enzyme-linked immunosorbent assay for above hepatitis viral antigen and antibody detections in human sera. This integrated protein microarray can be easily optimized for clinical use and epidemiological screening for multiple hepatitis virus infections.