Urotensin II induces transactivation of the epidermal growth factor receptor via transient oxidation of SHP-2 in the rat renal tubular cell line NRK-52E

Urotensin-II (UII) is a potent vasoactive peptide that has been implicated in cardiac fibrosis and renal diseases. However, the role played by UII in renal tissues is largely unknown. In this study, we investigated the effects of human UII (hUII) on rat renal proximal tubular cells of the NRK-52E line and the role of Src homology 2-containing phosphotyrosine phosphatase (SHP-2) in the hUII-induced transactivation of the epidermal growth factor receptor (EGFR). Exposure to hUII at low concentrations significantly induced proliferation in NRK-52E cells; this effect was inhibited by treatment with an ERK1/2 inhibitor (PD98059). UII treatment increased the phosphorylation of EGFR and induced the generation of reactive oxygen species (ROS). Treatment of the ROS scavenger N-acetyl-cysteine (NAC) inhibited EGFR transactivation and ERK phosphorylation induced by hUII. SHP-2 was found to interact with EGFR and be transiently oxidized following the hUII treatment. In SHP-2 knockdown cells, UII-induced phosphorylation of EGFR was less influenced by NAC, and significantly suppressed by heparin binding (HB)-EGF neutralizing antibody. Our data suggest that the ROS-mediated oxidation of SHP-2 is essential for the hUII-induced mitogenic pathway in NRK-52E cells.     

Src homology 2 domain-containing protein tyrosine phosphatase substrate 1 regulates the induction of Langerhans cell maturation

Recently, we reported that Src homology 2 domain-containing protein tyrosine phosphatase substrate 1 (SHPS-1) plays an important role in the migration of Langerhans cells (LC). Here, we show that SHPS-1 is involved in the maturation of LC. Immunofluorescence analysis on epidermal sheets for I-A or CD86 revealed that LC maturation induced by 2,4-dinitro-1-fluorobenzene (DNFB) or by TNF-alpha was inhibited by pretreatment with an anti-SHPS-1 monoclonal antibody (mAb) or with CD47-Fc fusion protein, a ligand for SHPS-1. Further, FACS analysis demonstrated that I-A(+) LC that had emigrated from skin explants expressed CD80 or CD86, whereas CD47-Fc protein reduced CD80(high+) or CD86(high+) cells. CD47-Fc protein also reduced the up-regulation of surface CD80 or CD86 by LC remaining in the skin explants. In SHPS-1 mutant mice, we observed that the up-regulation of surface CD86 and CCR7 by LC induced by DNFB as well as that of surface CD80 and CD86 by LC in skin explants was attenuated. Finally, contact hypersensitivity (CHS) response was suppressed in SHPS-1 mutant mice and in wild-type mice treated with an anti-SHPS-1 mAb. These observations indicate that SHPS-1 plays an important role in the maturation of LC ex vivo and in vivo, and that SHPS-1-CD47 interaction may negatively regulate CHS.     

表皮生长因子受体和环氧合酶-2在宫颈癌中的表达及临床诊断意义

目的 探讨宫颈癌组织中表皮生长因子受体(epidermal growth factor receptor,EGFR)和环氧合酶2(cyclooxygenase,COX-2)蛋白的表达及其与临床病理特征的关系.方法 采用免疫组化的方法分别检测正常宫颈、宫颈癌中EGFR和COX-2蛋白的表达.结果 EGFR和COX-2蛋白的在宫颈癌组织中的阳性表达率,均显著高于正常宫颈组织.EGFR和COX-2的阳性表达均与患者的年龄及肿瘤大小无关(P＞0.05).多元生存分析显示,EGFR和COX-2是独立的预后因子,相对危险度分别为2.52(P=0.004)和1.88(P=0.039).结论 EGFR和COX-2在宫颈组织中的表达水平可能与肿瘤的发生、发展、浸润和转移密切相关,可作为宫颈癌恶性程度判断和预后的重要指标.     

Seminal plasma and prostaglandin E2 up-regulate fibroblast growth factor 2 expression in endometrial adenocarcinoma cells via E-series prostanoid-2 receptor-mediated transactivation of the epidermal growth factor receptor and extracellular signal-regulated kinase pathway

BACKGROUND: Prostaglandin E(2) (PGE(2)) has been shown to modulate angiogenesis and tumour progression via the E-series prostanoid-2 (EP2) receptor. Endometrial adenocarcinomas may be exposed to endogenous PGE(2) and exogenous PGE(2), present at high concentration in seminal plasma. METHODS: This study investigated fibroblast growth factor 2 (FGF2) mRNA expression and cell signalling in response to seminal plasma or PGE(2), using an endometrial adenocarcinoma (Ishikawa) cell line stably expressing the EP2 receptor (EP2 sense cells) and endometrial adenocarcinoma explants. RESULTS: Seminal plasma and PGE(2) induced a significant up-regulation of FGF2 expression in EP2 sense but not parental untransfected Ishikawa (wild-type) cells (P < 0.05). These effects were inhibited by co-treatment with EP2 receptor antagonist or inhibitors of protein kinase A, c-Src, epidermal growth factor receptor (EGFR) kinase or extracellular signal-regulated kinase (ERK) signalling. The treatment of EP2 sense cells with seminal plasma induced cAMP accumulation and phosphorylation of c-Src, EGFR kinase and ERK via the EP2 receptor. Finally, seminal plasma and PGE(2) significantly increased FGF2 mRNA expression in endometrial adenocarcinoma tissue explants via the EP2 receptor (P < 0.05). CONCLUSIONS: Seminal plasma and PGE(2) can similarly activate FGF2 expression and EP2 receptor signalling in endometrial adenocarcinoma cells. These data highlight the potential for seminal plasma exposure to facilitate tumorigenesis-angiogenesis in endometrial adenocarcinomas in vivo.     

Detection of epidermal growth factor receptor and human epidermal growth factor receptor 2 activating mutations in lung adenocarcinoma by high-resolution melting amplicon analysis: correlation with gene copy number, protein expression, and hormone receptor expression

Activating mutations in the epidermal growth factor receptor (EGFR) (7p12.3-p12.1) and in the human epidermal growth factor receptor 2 (HER2) (17q21.1) characterize a subset of lung carcinomas. These mutations may relate to the response of the tumor to the tyrosine kinase inhibitors gefitinib and erlotinib. High-resolution melting amplicon analysis is a screening technique that has been shown to be able to detect missense mutations as well as deletions and insertions in tumor DNA isolated from paraffin-embedded tissue sections. In this study, we used high-resolution melting amplicon analysis to screen for EGFR and human HER2 activating mutations in 39 patients with primary lung adenocarcinoma. There were 20 cases that showed bronchioloalveolar histology and 19 cases that did not. The EGFR exons screened were exons 18, 19, 20, and 21, and the HER2 exons screened were exons 19 and 20. Six (15%) of the 39 patients had tumors that contained EGFR activating mutations. Four of the mutations were in adenocarcinomas, which had some bronchioloalveolar features, and 2 mutations were in tumors without bronchioloalveolar features. The EGFR mutations were in exon 19 (2 cases), exon 20 (2 cases), and exon 21 (1 case). One case contained mutations in both exons 18 and 20. One (2.6%) of the 39 patients had a tumor that contained an HER2 activating mutation, and the mutation was located in exon 20. Two of the 6 EGFR mutation-positive cases showed polysomy for chromosome 7, and each one showed overexpression of EGFR as determined by immunohistochemical staining. The other EGFR mutation-positive cases did not show EGFR overexpression and appeared disomic for chromosome 7. The HER2 mutation-positive case was in an adenocarcinoma with bronchioloalveolar features. This tumor did not show overexpression of HER2 and was disomic for chromosome 17. For the non-EGFR mutation-positive cases, 4 (13%) of 32 evaluated cases showed polysomy for chromosome 7 and EGFR. No case showed EGFR gene amplification. Polysomy for chromosome 7 was not related to EGFR overexpression as estimated by immunohistochemistry. Estrogen and progesterone receptor expression was not strong in any of the cases and did not correlate with the presence of EGFR or HER2 mutations.     

荧光原位杂交检测非小细胞肺癌EGFR基因拷贝数状况分析

目的 探讨非小细胞肺癌EGFR基因拷贝数状况的临床病理学意义.方法 采用LSI EGFR/CEP 7探针试剂盒对108例非小细胞肺癌石蜡包埋组织标本进行EGFR基因状况的检测,并分析EGFR基因拷贝数与患者临床病理学之间的关系.结果 在108例非小细胞肺癌病例中,64.8%病例判读为EGFR基因低基因拷贝数即FISH阴性,其中二体性21.3%、低度三体性18.5%、高度三体性13.9%、低度多体性11.1%.35.2%病例判读为EGFR基因高基因拷贝数即FISH阳性,其中高度多体性25.9%、基因扩增9.3%.除2例扩增病例(Ratio>2.4)外,所有三体性及多体性,7号染色体与EGFR基因的拷贝数出现一致性的增加,Ratio范围在1.12～1.54之间.EGFR基因状况或EGFR-FISH(-/+)与临床病理特征均无显著相关性.结论 EGFR基因拷贝数增加在非小细胞肺癌是较常发生的事件,可能参与肺癌的发生发展.     

电针干预对缺血再灌注模型大鼠的神经保护与神经调节蛋白1/表皮生长因子受体4信号通路的关系

背景:电针干预曲池、足三里穴位能够起到神经保护作用,改善患者的运动功能,然而针对其作用机制的深入研究则相对较少。目的:基于神经调节蛋白1/表皮生长因子受体4信号通路观察电针干预对脑缺血再灌注损伤模型大鼠的神经保护作用。方法:取90只SPF级雄性Wistar大鼠,采用改良线栓法制备脑缺血再灌注损伤大鼠模型,并随机分为模型组、非穴位组和穴位组;另取30只大鼠只予以左侧颈部血管的分离作为假手术组。造模后3 h,非穴位组用电针刺激右侧肢体腋横纹下和尾骨尖下3 mm的非穴位处,穴位组用电针刺激曲池和足三里,共治疗7 d;假手术组和模型组不予任何治疗,只进行与治疗组相同条件的抓取与固定。造模完成后,以神经功能缺损评分进行模型评价;治疗结束后评价各组大鼠的神经功能缺损评分;检测缺血侧局部脑血流量和血流速度;TTC染色观察并计算脑梗死体积;电镜观察神经元的超微结构;TUNEL染色观察神经细胞凋亡情况;Western blotting检测神经调节蛋白1/表皮生长因子受体4通路蛋白表达水平。结果与结论:①与假手术组相比,模型组和非穴位组大鼠的神经功能缺损评分、脑梗死体积、神经细胞凋亡率、神经调节蛋白1、表皮生长因子受体4蛋白表达水平明显升高,脑血流量、脑血流速度明显下降(P<0.05),神经元超微结构异常,模型组与非穴位组间差异无显著性意义(P>0.05);②与模型组相比,穴位组大鼠的神经功能缺损评分、脑梗死体积、神经细胞凋亡率明显下降,脑血流量、脑血流速度、神经调节蛋白1、表皮生长因子受体4蛋白表达水平明显上升(P<0.05),神经元超微结构明显改善;③说明电针刺激曲池、足三里穴位能够明显改善脑缺血再灌注损伤大鼠的神经功能缺损状态和脑神经元超微结构,抑制神经细胞凋亡,起到神经保护作用,其机制可能与神经调节蛋白1/表皮生长因子受体4信号通路的调控有关。     

乳腺癌中高迁移率族蛋白B1和表皮生长因子受体1的表达及相关性研究

目的 探讨乳腺癌组织中高迁移率族蛋白B1(HMGB1)和表皮生长因子受体1(EGFR)的表达情况、相关性,及其对患者预后的影响。方法 纳入2007年1月-2018年12月东阳市人民医院病理科浸润性乳腺癌石蜡标本392例。患者均为女性;年龄26～90岁,中位年龄52岁。采用免疫组织化学EnVision法,检测标本中HMGB1蛋白在细胞核、细胞质的表达情况以及EGFR蛋白在细胞膜的表达情况,分析HMGB1蛋白与EGFR蛋白表达的相关性。根据HMGB1蛋白在细胞核及细胞质表达状态分为细胞核低表达且细胞质阴性组、细胞核高表达且细胞质阴性组、细胞核低表达且细胞质阳性组、细胞核高表达且细胞质阳性组,比较4组间EGFR蛋白的表达情况。分别观察细胞核HMGB1蛋白高表达+EGFR蛋白阳性、细胞质HMGB1蛋白阳性+EGFR蛋白阳性时对患者预后的影响。结果 (1)乳腺癌中,HMGB1蛋白在细胞核和细胞质阳性表达率分别为80.1%(314/392)和15.1%(59/392),EGFR蛋白在细胞膜阳性表达率为53.6%(210/392)。(2)细胞核HMGB1蛋白高表达患者的EGFR蛋白阳性率56.1%(176/314),高于细胞核HMGB1蛋白低表达的EGFR蛋白阳性率43.6%(34/78),差异有统计学意义(χ²=3.901, P<0.05);Spearman相关分析显示细胞核HMGB1蛋白高表达和EGFR蛋白阳性呈正相关(r=0.100, P<0.05)。细胞质HMGB1蛋白阳性患者的EGFR蛋白阳性率(66.1%,39/59),高于细胞质HMGB1蛋白阴性的EGFR蛋白阳性率(51.4%,171/333),差异有统计学意义(χ²=4.384, P<0.05);Spearman相关分析显示细胞质HMGB1蛋白阳性和EGFR蛋白阳性表达亦呈正相关(r=0.106, P<0.05)。(3)HMGB1蛋白细胞核低表达且细胞质阴性68例、细胞核高表达且细胞质阴性265例、细胞核低表达且细胞质阳性10例、细胞核高表达且细胞质阳性49例,4组中EGFR蛋白阳性率分别为42.6%(29/68)、53.6%(142/265)、50.0%(5/10)、69.4%(34/49),组间比较差异有统计学意义(χ²=8.242, P<0.05)。(4)392例乳腺癌病例中,235例获得了预后生存分析资料。患者术后随访时间为3～80个月,平均60个月。生存分析显示,在235例患者中,5年累积生存率为90.6%,5年无复发累积生存率为81.3%。细胞核HMGB1蛋白高表达且EGFR阳性的患者5年无复发累积生存率为75.2%,低于其他患者的85.8%,差异有统计学意义(χ²=4.171、P<0.05)。细胞核HMGB1蛋白高表达+EGFR阳性的患者5年累积生存率为89.1%,与其他患者的91.8%比较,差异无统计学意义(χ²=0.557、P＞0.05)。细胞质HMGB1蛋白阳性+EGFR阳性的患者5年无复发累积生存率为72.2%,5年累积生存率为83.3%,分别低于其他患者的82.0%、91.2%,但差异均无统计学意义(χ²=1.070、1.307,P值均＞0.05)。结论 细胞核及细胞质HMGB1蛋白的表达均与EGFR蛋白表达正相关,并且细胞核或细胞质HMGB1蛋白与EGFR蛋白同时高表达的患者有更低的5年无复发累积生存率和5年累积生存率。同时抑制HMGB1和EGFR可能成为抗乳腺癌治疗的新策略。     

Expression of c-erbB-2 protein and epidermal growth factor receptor in normal tissues of the female genital tract and in the placenta

Summary The c-erbB-2 (HER-2/neu) protein is a membrane glycoprotein growth factor receptor that has molecular homology with the epidermal growth factor receptor (EGFR). To investigate the relationship between the expression of c-erbB-2 protein and EGFR in the tissues of the human female genital tract and in the placenta, we examined the immunohistochemical reactivity of monoclonal antibodies against both of these proteins. In the müllerian-derived genital tract, epithelial cells of the fallopian tube, endometrium, and endocervix showed reactivity for c-erbB-2 protein, whereas reactivity for EGFR was distributed mainly in the stromal cells throughout the menstrual cycle and during pregnancy. In addition, the staining intensity for EGFR in the endometrial stroma increased with its decidualization. In the exocervical squamous epithelium, basal cells were cerbB-2 protein-negative and EGFR-positive, but the more differentiated squamous cells of the intermediate layer were c-erbB-2 protein-positive and EGFR-negative. In the placental tissues, cytotrophoblasts and syncytiotrophoblasts of the chorionic villi were c-erbB-2 protein-negative and EGFR-positive. In contrast, intermediate trophoblasts in the extravillous space were c-erbB-2 protein-positive and EGFR-negative. Thus, there is an inverse relationship between the expression of c-erbB-2 protein and EGFR in the tissues of the female genital tract and in the placenta. This suggests that there may be a regulatory mechanism(s) for the expression of both proteins that is associated with the differentiation and/or function of cells in the female genital tract and the placenta.     

Mechano growth factor E peptide promotes rat bone marrow-derived mesenchymal stem cell migration through CXCR4-ERK1/2

Abstract

Mechano growth factor (MGF) is a splicing variant of insulin-like growth factor 1 (IGF-1). The unique C-terminal E domain of MGF (MGF-E) makes it distinct from the other variants of IGF-1. Our previous work demonstrated that MGF-25E induces the migration of rat bone marrow-derived mesenchymal stem cells (rMSCs) by altering their mechanical properties, which is accompanied by the activation of the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway. However, the relationship between ERK1/2 activation and the change in mechanical properties has not been illustrated. In the present study, we determined that MGF-25E induced the migration of rMSCs by modulating CXCR4 to activate the ERK1/2 pathway. The analysis of the Young’s modulus and F-actin remodeling indicated that MGF-25E increased the stiffness and the F-actin polymerization of rMSCs through the activation of the CXCR4-ERK1/2 pathway. For the first time, this study clarified the signaling pathway that regulates the mechanical properties of rMSCs and is responsible for MGF-25E-promoted migration.     

Immunohistochemical localization of c-erbB-2 protein and epidermal growth factor receptor in normal surface epithelium,surface inclusion cysts,and common epithelial tumours of the ovary

Summary The c-erbB-2 (HER-2/neu) protein is a membrane glycoprotein growth factor receptor showing molecular homology with the epidermal growth factor receptor (EGFR). We examined the immunohistochemical reactivity of monoclonal antibodies against both of these proteins in normal surface epithelium, surface inclusion cysts, and common epithelial tumours of the ovary. The ovarian tumours were classified as benign (16), borderline malignant (2), and malignant (19). Normal surface ovarian epithelium was weakly positve for both c-erbB-2 protein and EGFR. In surface inclusion cysts, however, the epithelial cells lining the lumen exhibited stronger staining for c-erbB-2 protein, but no staining for EGFR. All 16 benign ovarian tumours and the 2 borderline malignant ovarian tumours were positive for c-erbB-2 protein and negative for EGFR. Of the ovarian carcinomas, 13 of the 19 (68.4%) were positive for c-erbB-2 protein and negative for EGFR, while 4 showed positivity for both c-erbB-2 protein and EGFR. Two cases were negative for both proteins. Expression of both c-erbB-2 protein and EGFR was found in endometrioid carcinoma with squamous differentiation and in clinically advanced poorly differentiated serous carcinomas. Expression of c-erbB-2 protein appears to be increased and that of EGFR is reduced in the early stage of epithelial ovarian oncogenesis. The expression of EGFR with c-erbB-2 protein in ovarian carcinoma is related both to histological differentiation and/or advanced clinical stage.     

A proposal for diagnostically meaningful criteria to classify increased epidermal growth factor receptor and c-erbB-2 gene copy numbers in gastric carcinoma,based on correlation of fluorescence in situ hybridization and immunohistochemical measurements

Amplification of the epidermal growth factor receptor (EGFR) and/or c-erbB-2 oncogenes and overexpression of their proteins are detected in 30% of gastric carcinomas, but there are few reports regarding the correlation between gene amplification and protein overexpression. We examined the correlation between amplification of the EGFR and c-erbB-2 genes, detected using fluorescence in situ hybridization, and overexpression of their proteins, detected using immunohistochemistry, in formalin-fixed tissue sections of 54 surgically resected gastric carcinomas. A mean EGFR copy number per nucleus of four or more and an EGFR/chromosome 7 centromere (CEP7) ratio of 1.7 or more were each detected in 4 specimens (7%). The sensitivity and specificity of both criteria for EGFR protein overexpression were 75% and 92%, respectively. A mean c-erbB-2 copy number per nucleus of 7.0 or more and a c-erbB-2/chromosome 17 centromere (CEP17) ratio of 2.0 or more were detected in six (11%) and eight (15%) specimens, respectively. The sensitivity and specificity of the former criterion to c-erbB-2 overexpression were 83% and 98%, respectively, while those of the latter were 63% and 98%. A mean EGFR gene copy number of 4.0 or more and/or an EGFR/CEP7 ratio of 1.7 and a mean c-erbB-2 gene copy number of 7.0 or more and/or a c-erbB-2/CEP17 ratio of 2.0 or more would be useful in defining increased EGFR and c-erbB-2 gene copy numbers, respectively, in gastric carcinomas.     

Interleukin-1β, interleukin-6, epidermal growth factor and transforming growth factor-α are elevated in the brain from parkinsonian patients

Interleukin (IL)-1β, IL-6, epidermal growth factor (EGF), and transforming growth factor-α (TGF-α) were measured for the first time in the brain (caudate nucleus, putamen and cerebral cortex) from control and parkinsonian patients by highly sensitive sandwich enzyme immunoassays. The concentrations of IL-1β, IL-6, EGF, and TGF-α in the dopaminergic, striatal regions were significantly higher in parkinsonian patients than those in controls, whereas those in the cerebral cortex did not show significant differences between parkinsonian and control subjects. Since these cytokines and growth factors may play important roles as neurotrophic factors in the brain, the present results suggest that they may be produced as compensatory responses in the nigrostriatal dopaminergic regions in Parkinson's disease, and may be related, at least in part, to the process of neurodegeneration in Parkinson's disease.     

Human epidermal growth factor receptor 2, Na+/H+ exchanger regulatory factor 1, and breast cancer susceptibility gene-1 as new biomarkers for familial breast cancers

The aim of this study is to evaluate the analysis of markers related with progression, to further characterize familial breast cancers. Here, we investigated the expression of breast cancer susceptibility gene-1, hypoxia-inducible factor-1α, vascular endothelial growth factor receptor 1, and Na+/H+ exchanger regulatory factor 1 in 187 microarrayed breast carcinomas from 94 familial and 93 sporadic breast cancer patients by immunohistochemical staining. Furthermore, the expression levels of these biomarkers were compared with triple-negative phenotype. Familiarity was significantly associated with younger age (P < .000), higher tumor grade (P = .038), negative estrogen receptor hormonal status (P = .036), and high proliferative activity (P = .029). The familial cancers were immunonegative for membranous Na+/H+ exchanger regulatory factor 1 expression compared with sporadic cancers (P = .001); notably, vascular endothelial growth factor receptor 1 staining correlated with cytoplasmic Na+/H+ exchanger regulatory factor 1 expression in familial tumors (P = .009). In multivariate analysis, the "new biomarkers," including negative human epidermal growth factor receptor 2 status (odds ratio, 4.538; 95% confidence interval, 1.756-11.728), negative membranous Na+/H+ exchanger regulatory factor 1 expression (odds ratio, 7.686; 95% confidence interval, 1.876-31.483) and positive nuclear breast cancer susceptibility gene-1 (odds ratio, 0.3982; 95% confidence interval, 0.169-0.936), significantly correlated with family history of breast cancer. We hypothesize that the evaluation of human epidermal growth factor receptor 2, Na+/H+ exchanger regulatory factor 1, and breast cancer susceptibility gene-1 could be clinically useful to identify familial breast tumors and to select patients candidate to breast cancer susceptibility genes 1/2 gene sequencing.     

Relationship between epidermal growth factor receptor (EGFR) mutation and serum cyclooxygenase-2 Level,and the synergistic effect of celecoxib and gefitinib on EGFR expression in non-small cell lung cancer cells

Epidermal growth factor receptor (EGFR) mutations occur mostly in patients with lung adenocarcinoma; such patients are also more likely to express cyclooxygenase-2 (COX-2), indicating a possible relationship between EGFR mutation and COX-2. The COX-2 and EGFR pathways mutually enhance their procarcinogenic effects in different tumor types. Therefore, simultaneous EGFR and COX-2 inhibition may be a promising therapeutic approach for patients with lung adenocarcinoma. We obtained tissue and serum samples from patients with non-small cell lung cancer (NSCLC) to detect the relationship between EGFR mutation and serum COX-2 level. Subsequently, gefitinib was combined with celecoxib to investigate the efficacy of inhibition in vitro in two NSCLC cell lines: HCC827 (del E746-A750) and A549 (wild-type EGFR). The cells were treated with gefitinib or celecoxib alone or with gefitinib plus celecoxib. Cell proliferation and apoptosis were assessed and correlated with expression of COX-2 and phosphorylated (p)-EGFR. The EGFR mutation rate of the high-COX-2 patients was significantly higher than that in the low-COX-2 patients. Multivariate analysis showed that high COX-2 levels were independently associated with EGFR mutation. Celecoxib and gefitinib inhibited cell growth in both cell lines. At sufficiently high concentrations, celecoxib plus gefitinib significantly mutually enhanced their anti-proliferative and apoptotic effects in both cell lines. At low concentrations, the combination had no additional effects on A549 cells. There was increased down regulation of COX-2 and p-EGFR when both cell lines were treated with high-concentration celecoxib plus gefitinib compared to either agent alone. This study demonstrates that high serum COX-2 levels may indicate EGFR mutations and that the efficacy of combined celecoxib and gefitinib is significantly greater in NSCLC cells with EGFR mutations; at high concentrations, the combination is efficacious in wild-type NSCLC cells.     

Deficiency of SHP1 leads to sustained and increased ERK activation in mast cells, thereby inhibiting IL-3-dependent proliferation and cell death

SHP-1 plays an important role for the regulation of signaling from various hematopoietic cell receptors. In this study, we examined IL-3-induced cell proliferation and IL-3 depletion-induced apoptosis in bone marrow-derived mast cells (BMMC) established from motheaten (me) that lack SHP-1 expression, viable motheaten (me^v) expressing phosphatase-deficient SHP-1, and wild-type (WT) mice. When BMMC were stimulated with IL-3, increased ERK activation was evident in resting state and sustained in me-BMMC relative to WT-BMMC. ERK is known to be involved in the regulation of cell proliferation and apoptosis in some cells. In accordance with sustained ERK activation, apoptosis was decreased in me- and me^v-BMMC compared with WT-BMMC. In contrast to the predicted role of ERK as a pro-survival molecule, IL-3-induced cell proliferation was much lower in me- and me^v-BMMC than WT-BMMC. Stimulation with lower concentration of IL-3 or addition of PD98059, a MEK inhibitor, to the culture resulted in the suppression of decreased apoptosis and cell proliferation in me- and me^v-BMMC. Collectively, these results suggest that SHP-1 positively regulates IL-3-dependent mast cell proliferation and apoptosis by inhibiting ERK activity through its phosphatase activity. Furthermore, our results indicate that ERK would act as a negative regulator for cell proliferation and induce apoptosis when its activity is highly increased.