Efficiency of serum copper/zinc ratio for differential diagnosis of patients with and without lung cancer

We examined serum copper (Cu), serum zinc (Zn), and the serum copper/zinc ratio (Cu/Zn) in 162 patients. All of them were seen to have an abnormal shadow in the chest X-ray films, that is, 109 patients with lung cancer (LC) and 53 patients with no lung cancer (NLC). The mean Cu and Cu/Zn in LC patients were significantly higher than those in NLC patients (p<0.05). In LC patients, Cu and Cu/Zn were higher and Zn was lower in advanced tumors than early ones. There was a significantly clear relation between Cu or Cu/Zn and the tumor (T) stages. When the relative risk (RR) of LC was estimated, it was seen that the higher Cu and Cu/Zn became, the higher RR became. Furthermore, we showed the sensitivity of the receiver operator characteristic of the test (ROC) curve for Cu, Cu/Zn, and carcinoembryonic antigen (CEA) to diagnose LC, as explained in a paragraph of methods.The determinations of Cu, Zn, and Cu/Zn are simple and inexpensive. They also appear to have a great diagnostic value in determining the local invasion of LC and as a screening test in the high-risk patients for LC.     

Abortive transformation of temperature-sensitive mutants of rat 3Y1 cells by simian virus 40: effect of cellular arrest states on entry into S phase and cellular proliferation

Four temperature-sensitive (ts) mutants of rat 3Y1 fibroblasts, representing independent complementation groups, cease to proliferate predominantly with a 2n DNA content, at the restrictive temperature (39.8 degrees C) (temperature arrest) or at the permissive temperature (33.8 degrees C) at a confluent cell density (density arrest) (Ohno et al., 1984). We studied the temperature- or the density-arrested cells of these mutants infected with simian virus 40 (SV40) or its mutants affecting large T or small t antigen with respect to kinetics at 39.8 degrees C of entry into S phase and cellular proliferation. Three mutants, 3Y1tsD123, 3Y1tsF121 and 3Y1tsG125, expressed T antigen and entered S phase at 39.8 degrees C from both the arrested states after infection with either wild-type, tsA mutants, or a .54/.59 deletion mutant of SV40, whereas in the density-arrested 3Y1tsH203, expression of T antigen and entry into S phase were inefficient and ts. Following the WT-SV40 induced entry into S phase, the temperature-arrested 3Y1tsD123 detached from the substratum with no detectable increase in cell number, whereas the density-arrested ones completed a round of the cell cycle and then detached. 3Y1tsF121 and 3Y1tsG125 in the both arrested states proliferated through more than one generation. 3Y1tsF121 and 3Y1tsG125 in the density-arrested state infected with tsA mutants once proliferated and then ceased to increase in number as the percentage of T-antigen positive population decreased. These results suggest that wild-type and tsA-mutated large T antigens are able to overcome the cellular ts blocks of entry into S phase in the 3 ts mutants of 3Y1 cells in both the arrested states, and that small t antigen is not required to overcome the blocks. It is also suggested that cellular behaviors subsequent to S phase (viability, mitosis, and proliferation in the following generations) depend on cellular arrest states, on traits of cellular ts defects, and on the duration of large T antigen expression.     

Characterization of the Polyamine Biosynthetic Pathways and Salt Stress Response in Brachypodium distachyon

Journal of Plant Growth Regulation - To characterize polyamine (PA) biosynthetic pathways in Brachypodium distachyon, we analyzed the gene-expression patterns and PA contents in various organs....     

Phenotypic characterization of lymphokine-activated killer cells from human lymph node lymphocytes

We previously reported that lymphokine-activated killer (LAK) activity can be generated in human lymph node lymphocytes (LNL) at the same level as that in peripheral blood lymphocytes (PBL), despite the absence of active natural killer (NK) cells. In the present study, we investigated the surface phenotype of LNL-LAK cells by fractionation of lymphocytes, using a panning method. LNL isolated from lung cancer patients were cultured in the presence of recombinant interleukin 2 for 8 days and separated into T cells and non-T cells according to the expression of CD3 antigen. LAK effectors were enriched in the CD3- non-T cells. However, the CD3+ cells also mediated a low but substantial level of LAK activity, which was attributed to a CD8+ T-cell subset. Further investigation of the CD3- cells revealed that most of the CD3- effector cells expressed neither B-cell (CD20) nor NK-cell (CD16) markers. Precursors of this CD3-CD20-CD16- (null) population appeared to be also CD3-, CD20-, and CD16-. From these results, we would stress the significant contribution of CD3-CD20-CD16- null cells to the LAK phenomenon, which has not been focused on in PBL.     

Effects of sodium n-butyrate on entry into S phase in resting rat 3Y1 cells infected with simian virus 40.

In quiescent rat 3Y1 fibroblasts infected with simian virus 40 (SV40), sodium butyrate elongated the time lag before entry into S phase in a concentration-dependent fashion. In spite of the elongated time lags, SV40-infected cells entered S phase in a very synchronous mode, irrespective of the butyrate concentrations. The elongated time lag seemed to be at least partially due to a delayed synthesis and a delayed accumulation of large T antigen caused by butyrate. The entry into S phase was also delayed even when butyrate was added to the cultures after expression of T antigen to an extent sufficient for untreated cells to enter S phase. This suggests that butyrate may also inhibit a cellular event(s) that is required for entry into S phase after expression of the T antigen. In contrast, serum-stimulated cells were more sensitive to butyrate with respect to entry into S phase than SV40-infected cells, and the distribution of the time lag among cell populations increased (i.e., asynchrony in entry into S phase increased) with an increase in the butyrate concentration.     

Clinical,Pathological, and Molecular Features of Lung Adenocarcinomas with AXL Expression

The receptor tyrosine kinase AXL is a member of the Tyro3-Axl-Mer receptor tyrosine kinase subfamily. AXL affects several cellular functions, including growth and migration. AXL aberration is reportedly a marker for poor prognosis and treatment resistance in various cancers. In this study, we analyzed clinical, pathological, and molecular features of AXL expression in lung adenocarcinomas (LADs). We examined 161 LAD specimens from patients who underwent pulmonary resections. When AXL protein expression was quantified (0, 1+, 2+, 3+) according to immunohistochemical staining intensity, results were 0: 35%; 1+: 20%; 2+: 37%; and 3+: 7% for the 161 samples. AXL expression status did not correlate with clinical features, including smoking status and pathological stage. However, patients whose specimens showed strong AXL expression (3+) had markedly poorer prognoses than other groups (P = 0.0033). Strong AXL expression was also significantly associated with downregulation of E-cadherin (P = 0.025) and CD44 (P = 0.0010). In addition, 9 of 12 specimens with strong AXL expression had driver gene mutations (6 with EGFR, 2 with KRAS, 1 with ALK). In conclusion, we found that strong AXL expression in surgically resected LADs was a predictor of poor prognosis. LADs with strong AXL expression were characterized by mesenchymal status, higher expression of stem-cell-like markers, and frequent driver gene mutations.     

Collateral Chemoresistance to Anti-Microtubule Agents in a Lung Cancer Cell Line with Acquired Resistance to Erlotinib

Various alterations underlying acquired resistance to epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) have been described. Although treatment strategies specific for these mechanisms are under development, cytotoxic agents are currently employed to treat many patients following failure of EGFR-TKIs. However, the effect of TKI resistance on sensitivity to these cytotoxic agents is mostly unclear. This study investigated the sensitivity of erlotinib-resistant tumor cells to five cytotoxic agents using an in vitro EGFR-TKI-resistant model. Four erlotinib-sensitive lung adenocarcinoma cell lines and their resistant derivatives were tested. Of the resistant cell lines, all but one showed a similar sensitivity to the tested drugs as their parental cells. HCC4006ER cells with epithelial mesenchymal transition features acquired resistance to the three microtubule-targeting agents, docetaxel, paclitaxel and vinorelbine, but not to cisplatin and gemcitabine. Gene expression array and immunoblotting demonstrated that ATP-binding cassette subfamily B, member 1 (ABCB1) was up-regulated in HCC4006ER cells. ABCB1 knockdown by siRNA partially restored sensitivity to the anti-microtubule agents but not to erlotinib. Moreover, the histone deacetylase inhibitor entinostat sensitized HCC4006ER cells to anti-microtubule agents through ABCB1 suppression. Our study indicates that sensitivity of tumor cells to cytotoxic agents in general does not change before and after failure of EGFR-TKIs. However, we describe that two different molecular alterations confer acquired resistance to EGFR-TKIs and cytotoxic agents, respectively. This phenomenon should be kept in mind in selection of subsequent therapy after failure of EGFR-TKIs.     

Transformation by v-H-ras does not restore proliferation of a set of temperature-sensitive cell-cycle mutants of rat 3Y1 fibroblasts

Three temperature-sensitive cell-cycle mutants of rat 3Y1 fibroblasts (3Y1tsD123, 3Y1tsG125, and 3Y1tsH203, each belonging to distinct complementation groups) were transformed with plasmid DNA carrying Harvey murine sarcoma virus cDNA. The criteria for transformation were increase in saturation cell density, capability to clone in soft agar, and alteration in the cellular morphology. At 39.8 degrees C (restrictive temperature of the parental cell lines), all the transformed sublines of each mutant ceased to proliferate and were arrested reversibly in the G1 phase of the cell cycle like the parental lines. At both 39.8 degrees C and 33.8 degrees C (permissive temperature for the parental lines), all the untransformed parental lines synthesized p21ras at low rate. At 33.8 degrees C, all the transformed sublines synthesized p21ras at much higher rate and expressed the morphological phenotype characteristic to v-H-ras-induced transformation. At 39.8 degrees C, the rate of p21ras synthesis was not changed in the transformed sublines of 3Y1tsD123 and 3Y1tsG125, and the morphology of transformed phenotype also remained intact. In the transformed subline of 3Y1tsH203, the rate of p21ras synthesis was lowered at 39.8 degrees C to that seen in the untransformed parental line, and the transformed phenotype in morphology disappeared. In all of the transformed sublines, the amount of v-H-ras mRNA markedly expressed at both 33.8 degrees C and 39.8 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)     

Invasive human fibrosarcoma DNA mediated induction of a 92 kDa gelatinase/type IV collagenase leads to an invasive phenotype.

Proteolytic enzymes, such as gelatinase/type IV collagenase, play a pivotal role in cancer invasion and metastasis. Invasive human fibrosarcoma cells (HT1080) secrete two species of gelatinase/type IV collagenase, 68-72 kDa and 92 kDa enzymes. The purpose of this study is to elucidate which species of gelatinase/type IV collagenase plays a more important role in invasion. We have found that HT1080 x human fibroblast hybrids have reduced ability to invade a reconstituted basement membrane (Matrigel) in vitro compared to HT1080 cells, and abundantly secrete only the 68-72 kDa gelatinase/type IV collagenase. These data suggest that the 92 kDa gelatinase/type IV collagenase may be more important in HT1080 cell invasion. We next transfected HT1080 genomic DNA into non-invasive mouse C3H/10T1/2 fibroblast cells, which secrete only 68-72 kDa gelatinase/type IV collagenase. Four invasive transfectants were established. These invasive transfectants secreted the 92 kDa gelatinase/type IV collagenase in addition to the 68-72 kDa gelatinase/type IV collagenase, whereas non-invasive control DNA transfectants did not secrete the 92 kDa gelatinase/type IV collagenase. These results suggest that the induction of the 92 kDa gelatinase/type IV collagenase is important in the invasive phenotype.     

Genetic analysis of the catalytic domain of the GAP gene in human lung cancer cell lines

Cell lines of non-small cell lung cancer (nonSCLC) have been shown to contain activating mutation of the K-ras oncogene in about 30% of cases, whereas no small cell lung cancer (SCLC) cell lines displayed these mutations. Biochemically, these mutations result in the ras gene product (p21) being constitutively activated in its GTP-bound form and insensitive to the hydrolytic action of the ras-specific GTPase-activating protein (ras GAP). We hypothesized that, if tumor development is related to the p21 ras being in the active GTP-bound state, then a similar malignant phenotype may result from an inactivating mutation in the ras GAP gene in the region that interacts with ras p21 (so-called catalytic domain). To test this hypothesis, we screened a panel of SCLC and non-SCLC cell lines for major genetic alterations in the catalytic domain of the GAP gene with the Sothern blot technique, and for minor genetic abnormalities (e.g., point mutations) with denaturing gradient gel electrophoresis and single-strand conformation polymorphism. Mutations in the catalytic domain of the GAP gene could not be demonstrated by any technique in any cell line examined. We conclude that mutational inactivation of the catalytic domain of the GAP gene probably does not contribute to the development of lung cancer.