The association between diastolic blood pressure and massive transfusion in severe trauma: a retrospective single-center study.

OBJECTIVE: To evaluate the association between diastolic blood pressure and massive transfusion in severe trauma. Method: The retrospective study was conducted at a tertiary emergency medical centre in Gwangju , Republic of Korea, and comprised data of severe trauma patients with injury severity score >15 presenting between January 2016 and December 2017. Multivariate logistic regression analysis was performed to evaluate the association between diastolic blood pressure and massive transfusion. Receiver operating characteristic curve analysis was performed to estimate the prognostic performance of diastolic blood pressure. Data was analysed using SPSS 18. RESULTS: Of the 827 patients, 64(7.7%) underwent massive transfusion. After adjusting the confounders, diastolic blood pressure was found to be an independent factor in predicting massive transfusion (odds ratio: 0.965; 95% confidence interval: 0.956-0.975). CONCLUSIONS: Initially low diastolic blood pressure was found to be an independent predictor for massive transfusion in severe trauma cases.

A HIF-1 target, ATIA, protects cells from apoptosis by modulating the mitochondrial thioredoxin, TRX2.

The regulation of apoptosis is critical for controlling tissue homeostasis and preventing tumor formation and growth. Reactive oxygen species (ROS) generation plays a key role in such regulation. Here, we describe a HIF-1 target, Vasn/ATIA (anti-TNFα-induced apoptosis), which protects cells against TNFα- and hypoxia-induced apoptosis. Through the generation of ATIA knockout mice, we show that ATIA protects cells from apoptosis through regulating the function of the mitochondrial antioxidant, thioredoxin-2, and ROS generation. ATIA is highly expressed in human glioblastoma, and ATIA knockdown in glioblastoma cells renders them sensitive to hypoxia-induced apoptosis. Therefore, ATIA is not only a HIF-1 target that regulates mitochondrial redox pathways but also a potentially diagnostic marker and therapeutic target in human glioblastoma.

Biallelic Deletion of Pxdn in Mice Leads to Anophthalmia and Severe Eye Malformation.

Peroxidasin (PXDN) is a unique peroxidase containing extracellular matrix motifs and stabilizes collagen IV networks by forming sulfilimine crosslinks. PXDN gene knockout in Caenorhabditis elegans (C. elegans) and Drosophila results in the demise at the embryonic and larval stages. PXDN mutations lead to severe eye disorders, including microphthalmia, cataract, glaucoma, and anterior segment dysgenesis in humans and mice. To investigate how PXDN loss of function affects organ development, we generated Pxdn knockout mice by deletion of exon 1 and its 5' upstream sequences of the Pxdn gene using the CRISPR/Cas9 system. Loss of both PXDN expression and collagen IV sulfilimine cross-links was detected only in the homozygous mice, which showed completely or almost closed eyelids with small eyes, having no apparent external morphological defects in other organs. In histological analysis of eye tissues, the homozygous mice had extreme defects in eye development, including no eyeballs or drastically disorganized eye structures, whereas the heterozygous mice showed normal eye structure. Visual function tests also revealed no obvious functional abnormalities in the eyes between heterozygous mice and wild-type mice. Thus, these results suggest that PXDN activity is essential in eye development, and also indicate that a single allele of Pxdn gene is sufficient for eye-structure formation and normal visual function.

Prognostic Performance Evaluation of the International Society on Thrombosis and Hemostasis and the Korean Society on Thrombosis and Hemostasis Scores in the Early Phase of Trauma.

BACKGROUND: Disseminated intravascular coagulation (DIC) contributes to poor outcome in the early phase of trauma. We aimed to analyze and compare the prognostic performances of the International Society on Thrombosis and Hemostasis (ISTH) and the Korean Society on Thrombosis and Hemostasis (KSTH) scores in the early phase of trauma. METHODS: Receiver operating characteristics analysis was used to examine the prognostic performance of both scores, and multivariate analysis was used to estimate the prognostic impact of the ISTH and KSTH scores in the early phase of trauma. The primary outcome was 24-hour mortality and the secondary outcome was massive transfusion. RESULTS: Of 1,229 patients included in the study, the 24-hour mortality rate was 7.6% (n = 93), and 8.1% (n = 99) of patients who received massive transfusions. The area under the curves (AUCs) of the KSTH and ISTH scores for 24-hour mortality were 0.784 (95% confidence interval [CI], 0.760-0.807) and 0.744 (95% CI, 0.718-0.768), respectively. The AUC of KSTH and ISTH scores for massive transfusion were 0.758 (95% CI, 0.734-0.782) and 0.646 (95% CI, 0.619-0.673), respectively. The AUCs of the KSTH score was significantly different from those of the ISTH score. Overt DIC according to KSTH criteria only, was independently associated with 24-hour mortality (odds ratio [OR], 2.630; 95% CI, 1.456-4.752). Only the KSTH score was independently associated with massive transfusion (OR, 1.563; 95% CI, 1.182-2.068). CONCLUSION: The KSTH score demonstrates a better prognostic performance for outcomes than the ISTH score in the early phase of trauma.

Myocilin is involved in NgR1/Lingo-1-mediated oligodendrocyte differentiation and myelination of the optic nerve.

Myocilin is a secreted glycoprotein that belongs to a family of olfactomedin domain-containing proteins. Although myocilin is detected in several ocular and nonocular tissues, the only reported human pathology related to mutations in the MYOCILIN gene is primary open-angle glaucoma. Functions of myocilin are poorly understood. Here we demonstrate that myocilin is a mediator of oligodendrocyte differentiation and is involved in the myelination of the optic nerve in mice. Myocilin is expressed and secreted by optic nerve astrocytes. Differentiation of optic nerve oligodendrocytes is delayed in Myocilin-null mice. Optic nerves of Myocilin-null mice contain reduced levels of several myelin-associated proteins including myelin basic protein, myelin proteolipid protein, and 2'3'-cyclic nucleotide 3'-phosphodiesterase compared with those of wild-type littermates. This leads to reduced myelin sheath thickness of optic nerve axons in Myocilin-null mice compared with wild-type littermates, and this difference is more pronounced at early postnatal stages compared with adult mice. Myocilin also affects differentiation of oligodendrocyte precursors in vitro. Its addition to primary cultures of differentiating oligodendrocyte precursors increases levels of tested markers of oligodendrocyte differentiation and stimulates elongation of oligodendrocyte processes. Myocilin stimulation of oligodendrocyte differentiation occurs through the NgR1/Lingo-1 receptor complex. Myocilin physically interacts with Lingo-1 and may be considered as a Lingo-1 ligand. Myocilin-induced elongation of oligodendrocyte processes may be mediated by activation of FYN and suppression of RhoA GTPase.

Qualitative network modeling of the Myc-p53 control system of cell proliferation and differentiation.

A kinetic model of a molecular control system for the cellular decision to proliferate or differentiate is formulated and analyzed for the purpose of understanding how the system can break down in cancer cells. The proposed core of this control system is composed of the transcription factors Myc and p53. The network of interactions between these factors involves negative and positive feedback loops that are linked to pathways involved in differentiation, cell cycle, and apoptosis. Understanding the dynamics of the Myc-p53 control system is aided by the postulate that there exists a cancer zone defined as a range of oncogenic Myc activities where the probability of initiating cancer is high. We propose that an essential role of p53 is to prevent the system from entering or staying too long in the cancer zone by downregulating Myc or, when Myc activity somehow becomes too high, by inducing apoptosis, cell cycle arrest, or differentiation. Kinetic modeling illustrates how deletions or aberrations in PTEN, MDM2, and ARF (genes implicated in various cancers, including glioma) affect the Myc-p53 control system. In addition, computer simulations demonstrate how this control system generates different cellular phenotypes characterized by rates of cellular differentiation and proliferation.

Hepatic stellate cell-specific knockout of transcriptional intermediary factor 1γ aggravates liver fibrosis.

Transforming growth factor ß (TGFß) is a crucial factor in fibrosis, and transcriptional intermediary factor 1γ (TIF1γ) is a negative regulator of the TGFß pathway; however, its role in liver fibrosis is unknown. In this study, mesenchymal stem cells derived from human embryonic stem cells (hE-MSCs) that secrete hepatocyte growth factor (HGF) were used to observe the repair of thioacetamide (TAA)-induced liver fibrosis. Our results showed that TIF1γ was significantly decreased in LX2 cells when exposed to TGFß1. Such decrease of TIF1γ was significantly prevented by co-culture with hE-MSCs. Interaction of TIF1γ with SMAD2/3 and binding to the promoter of the α-smooth muscle gene (αSMA) suppressed αSMA expression. Phosphorylation of cAMP response element-binding protein (CREB) and binding on the TIF1γ promoter region induced TIF1γ expression. Furthermore, hepatic stellate cell-specific TIF1γ-knockout mice showed aggravation of liver fibrosis. In conclusion, loss of TIF1γ aggravates fibrosis, suggesting that a strategy to maintain TIF1γ during liver injury would be a promising therapeutic approach to prevent or reverse liver fibrosis.

Thymosin ß-4 is a novel regulator for primary cilium formation by nephronophthisis 3 in HeLa human cervical cancer cells.

Thymosinß-4(Tß4) is an actin-sequestering protein involved in tumor malignancy. Primary cilia, microtubule-based organelles, are present in most eukaryotic cells, which might be related to tumor cell transformation. Here, we investigated whether ciliogenesis is affected by Tß4 in HeLa human cervical cancer cells. The inhibition of Tß4 attenuated primary cilia formation. The frequency of cilia was increased by Tß4 overexpression. When yeast two-hybrid assay was performed by using Tß4 as a bait, we rescued nephronophthisis 3(NPHP3), one of the components of primary cilia. Interaction of Tß4 with NPHP3 in mammalian cells was confirmed by GST-pulldown assay. Their intracellular co-localization was observed by immunofluorescence staining at peripheral surface of cells. In addition, the number of ciliated cells was reduced by the inhibition of NPHP3. Moreover, NPHP3 expression was decreased by the inhibition of Tß4 but it was increased by Tß4 overexpression. Taken together, the results demonstrate that primary cilia formation could be regulated by Tß4 through its interaction with NPHP3 and/or the control of NPHP3 expression. It suggests that Tß4 is a novel regulator for primary cilia formation by NPHP3. It also suggests that tumorigenesis could be associated with inappropriate regulation of Tß4 and/or NPHP3 expression to maintain primary cilia formation normally.

BIS-mediated STAT3 stabilization regulates glioblastoma stem cell-like phenotypes.

Glioblastoma stem cells (GSCs) are a subpopulation of highly tumorigenic and stem-like cells that are responsible for resistance to conventional therapy. Bcl-2-intreacting cell death suppressor (BIS; also known as BAG3) is an anti-apoptotic protein that is highly expressed in human cancers with various origins, including glioblastoma. In the present study, to investigate the role of BIS in GSC subpopulation, we examined the expression profile of BIS in A172 and U87-MG glioblastoma cell lines under specific in vitro culture conditions that enrich GSC-like cells in spheres. Both BIS mRNA and protein levels significantly increased under the sphere-forming condition as compared with standard culture conditions. BIS depletion resulted in notable decreases in sphere-forming activity and was accompanied with decreases in SOX-2 expression. The expression of STAT3, a master regulator of stemness, also decreased following BIS depletion concomitant with decreases in the nuclear levels of active phosphorylated STAT3, while ectopic STAT3 overexpression resulted in recovery of sphere-forming activity in BIS-knockdown glioblastoma cells. Additionally, immunoprecipitation and confocal microscopy revealed that BIS physically interacts with STAT3. Furthermore, BIS depletion increased STAT3 ubiquitination, suggesting that BIS is necessary for STAT3 stabilization in GSC-like cells. BIS depletion also affected epithelial-to-mesenchymal transition-related genes as evidenced by decrease in SNAIL and MMP-2 expression and increase in E-cadherin expression in GSC-like cells. Our findings suggest that high levels of BIS expression might confer stem-cell-like properties on cancer cells through STAT3 stabilization, indicating that BIS is a potential target in cancer therapy.

Inactivating mutations of the caspase-10 gene in gastric cancer.

We have analysed the genetic alteration of the entire coding region and all splice sites of caspase-8 and -10 genes in 99 gastric cancers by polymerase chain reaction (PCR)-single strand conformation polymorphism (SSCP) and sequencing. We found LOH of the caspase-8 and -10 in nine (28%) of 32 and in four (15%) of 26 informative cases, respectively. Overall, three of 99 gastric cancers (3%) were found to have the caspase-10 mutations, which were identified in the coding regions of the death effector domain (codon 147) and the p17 large protease domain (codons 257 and 410), whereas no mutation was detected in caspase-8. In vitro expression studies, the M147T and Q257stop mutants severely impaired caspase-10-mediated apoptosis, whereas the V410I which was the same mutation detected in ALPS patient had a significant, albeit less severe, effect on apoptosis. The data presented here suggest that somatic alterations of the caspase-10 gene might contribute to the pathogenesis in a subset of gastric cancers through the loss of their apoptotic function.

BRAF and KRAS mutations in stomach cancer.

Ras proteins control signaling pathways that are key regulators of several aspects of normal cell growth and malignant transformation. BRAF, which encodes a RAF family member in the downstream pathway of RAS, is somatically mutated in a number of human cancers. The activating mutation of BRAF is known to play a role in tumor development. As there have been no data on the BRAF mutation in stomach cancer, we analysed the genomic DNAs from 319 stomach carcinomas for the detection of somatic mutations of BRAF. Overall, we detected BRAF mutations in seven stomach carcinomas (2.2%). Five of the seven BRAF mutations involved Val 599, the previously identified hotspot, but the substituted amino acid (V599 M) was different from the most common BRAF mutation (V599E). The remaining two mutations involved a conserved amino acid (D593G). One tumor had both BRAF and KRAS mutations. This is the first report on BRAF mutation in stomach cancer, and the data indicate that BRAF is occasionally mutated in stomach cancer, and suggest that alterations of RAS pathway both by RAS and BRAF mutations contribute to the pathogenesis of stomach cancer.

Inactivating mutations of CASPASE-7 gene in human cancers.

Caspase-7 is a caspase involved in the execution phase of apoptosis. To explore the possibility that the genetic alterations of CASPASE-7 might be involved in the development of human cancers, we analysed the entire coding region and all splice sites of human CASPASE-7 gene for the detection of somatic mutations in a series of human solid cancers, including carcinomas from stomach, colon, head/neck, esophagus, urinary bladder and lung. Overall, we detected CASPASE-7 mutations in two of 98 colon carcinomas (2.0%), one of 50 esophageal carcinomas (2.0%) and one of 33 head/neck carcinomas (3.0%). We expressed the tumor-derived caspase-7 mutants in 293 T cells and found that the apoptosis was reduced compared to the wild-type caspase-7. This is the first report on the CASPASE-7 gene mutations in human malignancies, and our data suggest that the inactivating mutations of the CASPASE-7 gene might lead to the loss of its apoptotic function and contribute to the pathogenesis of some human solid cancers.

Alterations of Fas-pathway genes associated with nodal metastasis in non-small cell lung cancer.

Many types of cancer cells are resistant to Fas-mediated apoptosis by several mechanisms, including the mutations of the genes involved in Fas-mediated apoptosis. In this study, to explore the possibility that the mutations of the genes involved in the proximal pathway of Fas-mediated apoptosis (Fas, FADD, caspase 8 and caspase 10) are involved in cancer metastasis, we have analysed somatic mutation and deletion of these genes in 80 non-small cell lung cancers (NSCLCs) with (n=43) and without (n=37) metastasis to the regional lymph nodes. We found 12 mutations (four Fas, four FADD, and four caspase 10 mutations) in 11 of 80 NSCLCs (13.8%). Interestingly, of these mutations, most mutations (10 out of 12) were detected in the NSCLCs with metastasis, and the frequency in the metastasis lesions (23%) was higher than that in the primary lesions of the NSCLCs without metastasis (5.4%). Furthermore, transfection study revealed that the tumor-derived mutants have decreased apoptosis inductions compared to the wild types. These data suggest that the inactivating mutations of the genes in the proximal pathway of Fas-mediated apoptosis may lead to a decreased cancer cell death and play a role in the metastasis of NSCLC.

The synergistic effect of combination temozolomide and chloroquine treatment is dependent on autophagy formation and p53 status in glioma cells.

Temozolomide (TMZ) is an alkylating agent used for the treatment of glioblastoma. The late autophagy inhibitor chloroquine (CQ) inhibits glioblastoma tumors in a p53-independent and p53-dependent manner. We addressed a possible beneficial effect of combination treatment with TMZ and CQ by examining the molecular and cellular mechanism of co-treatment. Combination treatment of U87 cell (wild type p53) with TMZ and CQ synergistically reduced cell proliferation and enhanced apoptosis, with increased sub-G1 hypodiploid cells and caspase activation. This effect was abolished by a pan-caspase inhibitor, Z-VAD-FMK. TMZ induced autophagy, and the addition of CQ further increased autophagic vacuoles. Inhibition of early stages of autophagy by Beclin 1 knockdown and 3-methyladenine pretreatment prevented the enhanced effect of the combination treatment. The combination treatment also upregulated p53 and phospho-p53 levels, whereas p53 knockdown or overexpression of mutant p53 abolished the combination effect. In contrast, combination therapy had no enhanced effect on U373 cell (mutant p53) proliferation and apoptosis within 3 d, although TMZ induced autophagy and co-treatment with CQ increased autophagic vacuole accumulation. However, long term combination treatment for 9-10 d effectively decreased clonal and cellular growth with increased G2-M arrest. This effect was also abolished by Beclin 1 knockdown. Our data support the beneficial effect of combination treatment with TMZ and CQ in glioma via differential autophagy-associated mechanisms, depending on p53 status.

Genetic analysis of the liver putative tumor suppressor (LPTS) gene in hepatocellular carcinomas.

Recently, a novel liver-related putative tumor suppressor (LPTS), which has a growth inhibitory function in the hepatocellular carcinoma (HCC) cell line, has been identified at chromosome 8p23. To determine the relationship of the LPTS with the development or progression of HCC, we analyzed the genetic alterations and the expression pattern of the LPTS gene in a series of 80 HCCs, six dysplastic nodules, and eight large regenerating nodules, determining the genomic structures. We identified a total of seven exons, of which two were alternative, and three LPTS isoforms, short (LPTS-S), medium (LPTS-M), and long-sizes (LPTS-L). In the genetic alteration study of the LPTS gene, no mutation was detected in the large regenerating nodules, dysplastic nodules, and HCC, whereas ten (34.5%) of 29 informative cases at one or more intragenic polymorphic sites showed loss of heterozygosity (LOH). Interestingly, LOH was identified only in HCC samples with hepatitis B virus (HBV) infection and the frequency of LOH was not statistically related with histologic grade and clinical stage, suggesting that allelic loss of the LPTS gene may occur as an early event in the development of HCC, especially in the cases with HBV infection.

Mutational analysis of Noxa gene in human cancers.

There has been mounting evidence that dysregulation of apoptosis is involved in the mechanisms of cancer development and somatic mutations of apoptosis-related genes have been reported in human cancers. Noxa, a Bcl-2 homology 3 (BH3)-only member of the Bcl-2 family, is known to interact with anti-apoptotic Bcl-2 family members and induces apoptosis. The aim of this study was to explore the possibility that the Noxa gene is mutated in human cancers. We have analyzed the entire coding region and all splice sites of the Noxa gene for the detection of somatic mutations in a series of human cancers, including carcinomas from stomach, colon, liver, urinary bladder and lung by polymerase chain reaction (PCR), single strand conformation polymorphism (SSCP), and DNA sequencing. We found one somatic mutation of the Noxa gene in a transitional cell carcinoma (TCC) of the urinary bladder. To evaluate the functional alterations of the mutant in apoptosis, we overexpressed the mutant and wild-type Noxa in 293T and HeLa cells, but could not find any significant difference in cell death between the wild-type and mutant Noxa. These data suggest that Noxa is rarely mutated in human carcinomas and that the contribution of Noxa gene mutation in the pathogenesis of human cancer might not be related to cell death mechanisms.

Stomach cancer highly expresses both initiator and effector caspases; an immunohistochemical study.

Caspases play an essential role during apoptotic cell death. While caspases 8 and 10 act as initiator caspases of the extrinsic apoptosis pathway, caspase 9 acts as an initiator caspase of the intrinsic apoptosis pathway. Caspase 3 is considered to be the main effector caspase involved in both intrinsic and extrinsic pathways. Alteration of apoptosis is essential for cancer development. Thus, analysis of the expression status of caspases, the main executioners of apoptosis, in cancer tissues is needed for a sophisticated understanding of cancer biology. In the current study, we analyzed the expression of caspases 3, 8, 9 and 10 in 60 advanced gastric adenocarcinomas by immunohistochemistry using a tissue microarray approach. Immunopositivity was observed for caspase 3 in 57 (95%), caspase 8 in 56 (93%), caspase 9 in 54 (90%), and caspase 10 in 58 (97%) of the 60 cancers. While 46 cancers (77%) expressed all of the caspases examined, 14 cancers (23%) showed loss of expression in one or more caspases examined. Normal gastric mucosal cells showed no or weak expression of caspases 3, 8, 9 and 10. Taken together, these results suggest that stomach cancer cells in vivo may need caspase expression for apoptosis. Also, higher expression of the caspases in stomach cancer cells than in normal gastric mucosal cells suggests that apoptosis in susceptible stomach cancer cells might be easily triggered, thereby producing selective pressure to make more apoptosis-resistant cells during tumor development.

Increased expression of FLIP, an inhibitor of Fas-mediated apoptosis, in stomach cancer.

Despite the cell surface expression of Fas (Apo-1/CD95), many types of tumor cells, including stomach cancer cells, are resistant to Fas-mediated apoptosis, indicating the presence of inactivating mechanisms of Fas signaling. Expression of FLICE-like inhibitory protein (FLIP), one of the inhibitory proteins of Fas-mediated apoptosis, has been reported in several cancer types, but not in stomach cancer. In the present study, we analyzed the expression of Fas and FLIP in 60 advanced gastric adenocarcinomas by immunohistochemistry using a tissue microarray approach. Immunopositivity (defined as >/=30% of the neoplastic cells) was observed for Fas in 58 (97%) and FLIP in 54 (90%) of the 60 cancers. All of the tumors with FLIP immunostaining also showed Fas immunostaining. Loss of cell surface Fas immunostaining, another mechanism of Fas resistance, was observed in 45 tumors (75%). By contrast, normal gastric mucosal cells showed no or weak expression of both Fas and FLIP. Taken together, these results indicate that increased expression of FLIP is a frequent event in stomach carcinomas, and suggest that for evading apoptosis stomach carcinoma cells in vivo may need FLIP expression, which might contribute to tumor development.

Immunohistochemical analysis of Omi/HtrA2 expression in stomach cancer.

The serine protease Omi/HtrA2 is released from mitochondria into the cytosol after apoptosis stimuli, inducing apoptosis in a caspase-independent manner through its protease activity and in a caspase-dependent manner by neutralizing the inhibition of inhibitor of apoptosis proteins (IAPs) on caspases. Alteration of apoptosis is essential for cancer development, and cancer cell death by radiation and chemotherapy is largely dependent upon apoptosis. Thus, analysis of the expression status of Omi/HtrA2, a regulator of apoptosis, in cancer tissues is needed for an understanding of cancer development. In the current study, we analyzed the expression of Omi/HtrA2 in 60 advanced gastric adenocarcinomas by immunohistochemistry using a tissue microarray approach. Immunopositivity (defined as >/=30%) was observed for Omi/HtrA2 in 43 (72%) of the 60 cancers. By contrast, the surface mucous cells and mucous neck cells in the normal gastric mucosa showed no or weak expression of Omi/HtrA2. Taken together, these results suggest that stomach cancer cells in vivo may need Omi/HtrA2 expression for apoptosis, and that Omi/HtrA2 expression might be involved in stomach cancer development.

Non-small cell lung cancers frequently express phosphorylated Akt; an immunohistochemical study.

Mounting evidence suggests that the alterations of Akt/protein kinase B (PKB) play an important role in tumorigenesis. Phosphorylated Akt regulates many of the key effector molecules involved in apoptosis, angiogenesis, and cell cycle progression during tumorigenesis. The expression of phosphorylated Akt has been described in some human malignancies, but not in primary human lung cancer. In this study, to understand the role of Akt in lung tumorigenesis we analyzed the expression of phosphorylated Akt in 43 non-small cell lung cancers (NSCLCs) by immunohistochemistry. Phosphorylated Akt was detected either in the cytoplasm (23 cases) or nucleus (6 cases) in 29 of 43 NSCLCs (67.4%). Squamous cell carcinomas, adenocarcinomas, and bronchioloalveolar carcinomas expressed phosphorylated Akt in 68.2%, 61.5% and 75%, respectively. We also analyzed the phosphorylated Akt expression between primary NSCLCs and their corresponding nodal metastasis; the expression was not, however, different between the primary and metastatic lesions. Taken together, these results indicate that Akt 1 is frequently activated in NSCLCs, irrespective of the histological subtypes, and suggest that phosphorylated Akt may play a role in the development of NSCLC rather than in the progression of NSCLC.