Pre-treatment CD8+ tumour-infiltrating lymphocyte density predicts distant metastasis after definitive treatment in patients with stage III/IV hypopharyngeal squamous cell carcinoma.

OBJECTIVES: Although inflammatory markers, such as the neutrophil-to-lymphocyte ratio, platelet-to-lymphocyte ratio and local immune markers have been shown to have prognostic utility, limited information is available regarding inflammatory and pre-existing tumour-infiltrating lymphocyte density and their association with prognosis in patients with hypopharyngeal squamous cell carcinoma. We investigated the prognostic ability of inflammatory markers and tumour-infiltrating lymphocyte density in stage III and stage IV hypopharyngeal squamous cell carcinoma patients receiving definitive treatment. DESIGN: Retrospective cohort study. SETTING: Kurume University Hospital. PARTICIPANTS: Ninety-six stage III or stage IV hypopharyngeal squamous cell carcinoma patients treated at the Kurume University Hospital between 2000 and 2014. MAIN OUTCOME MEASURES: Inflammatory markers and pre-treatment tumour-infiltrating lymphocyte density were examined from recorded haematologic data and immunohistochemical analysis. RESULTS: Multivariate analyses showed that the CD8+ tumour-infiltrating lymphocyte density was an independent predictive factor for distant metastasis and overall survival, whereas inflammatory markers, including the neutrophil-to-lymphocyte ratio and platelet-to-lymphocyte ratio, were not correlated with distant metastasis or overall survival. CONCLUSIONS: Higher pre-treatment CD8+ tumour-infiltrating lymphocyte density is a useful predictive biomarker for reduced distant metastasis and better prognosis.

SurePath® LBC improves the diagnostic accuracy of intrahepatic and hilar cholangiocarcinoma.

INTRODUCTION: The current study aimed to compare cytology using SurePath® (SP)-LBC and biliary tissue histology (BTH) for the diagnosis of biliary disease. METHODS: Between January 2014 and December 2016, 57 patients underwent endoscopic retrograde cholangiopancreatography for the diagnosis of biliary disease. Biliary cytological samples were processed using SP-LBC and subsequently BTH was performed. A final diagnosis was confirmed by surgery (23 malignant cases) and clinical follow-up (34 benign and malignant cases): 18 extrahepatic cholangiocarcinoma; 17 intrahepatic/hilar cholangiocarcinoma (intra/H-CC); eight other malignant disease; and 14 benign biliary disease. The diagnoses made using SP-LBC and BTH were classified into four categories: (1) benign; (2) indeterminate; (3) suspicious for malignancy/malignant; and (4) inadequate. In addition, diagnostic accuracy was compared between SP-LBC and BTH. RESULTS: Although 23% (13/57) of BTH samples were classified as inadequate, all SP-LBC cases were classified as adequate. Among 43 malignant cases, 11 normal, four indeterminate and 28 suspicious for malignancy/malignant were found using SP-LBC (26%, 9% and 65%, respectively), in contrast to 10 inadequate, nine normal, 10 indeterminate and 14 suspicious for malignancy/malignant observed using BTH (23%, 21%, 23%, and 33%, respectively). The identification of malignant cells was strikingly different between SP-LBC and BTH. Furthermore, limited to intra/H-CC, accuracy was significantly higher using SP-LBC than using BTH (P < .001). CONCLUSIONS: SP-LBC of the biliary tract is a useful and reliable method for diagnosing biliary malignant disease and has an advantage over BTH for detecting malignant cells and accurately diagnosing intra/H-CC.

A Combined test using both cell sediment and supernatant cell-free DNA in pleural effusion shows increased sensitivity in detecting activating EGFR mutation in lung cancer patients.

INTRODUCTION: The aim of this study was to examine whether a combined test using both cell sediment and supernatant cytology cell-free DNA (ccfDNA) is more useful in detecting EGFR mutation than using cell sediment DNA or supernatant ccfDNA alone in pleural effusion of lung cancer patients. METHODS: A total of 74 lung adenocarcinoma patients with paired samples between primary tumour and corresponding metastatic tumour with both cell sediment and supernatant ccfDNA of pleural effusion cytology were enrolled in this study. Cell sediment and supernatant ccfDNA were analysed separately for EGFR mutations by polymerase chain reaction. RESULTS: Out of 45 patients with mutant EGFR in primary tumours, EGFR mutations were detected in 23 cell sediments of corresponding metastases (sensitivity; 51.1%) and 20 supernatant ccfDNA corresponding metastases (sensitivity; 44.4%). By contrast, the combined test detected EGFR mutations in 27 corresponding metastases (sensitivity; 60.0%), and had a higher sensitivity than the cell sediment or the supernatant ccfDNA alone (P < .05). Out of 45 patients with mutant EGFR, 24, three and 18 were cytologically diagnosed as positive, atypical or negative, respectively. The detection rate in the combined test was highest (95.8%) in the positive group, and mutant EGFR was also detected in four of 18 samples (22.2%) in the negative group. CONCLUSIONS: A combined test using both cell sediment DNA and supernatant ccfDNA samples increases the concordance rate of EGFR mutations between primary tumour and corresponding metastases. Our findings indicate that supernatant ccfDNA is useful even in cases where the cytological diagnosis is negative.

Cytological nuclear atypia classification can predict prognosis in patients with endometrial cancer.

OBJECTIVE: Endometrial cancer is one of the leading causes of malignancy in females. Nuclear findings are important for patients with cancer, and can provide valuable information to treating oncologists. We investigated whether nuclear findings were a useful prognostic factor in patients with endometrial cancer. METHOD: We investigated 71 cases of endometrial carcinoma with paired histology and cytology at Kurume University Hospital. We classified endometrial endometrioid adenocarcinoma (EEC) G1 and G2 as type I carcinomas, and uterine papillary serous carcinoma (UPSC), clear cell carcinoma (CC) and EEC G3 as type II carcinomas. For the establishment of the cytological nuclear atypia classification, we examined the following nuclear factors on the cytological smears: mitotic figures, prominent nucleoli, nuclear area and anisonucleosis. RESULTS: There was a significant difference in mitotic figures (P < 0.001) and anisonucleosis (P = 0.026) in cytological smears between type I and type II carcinomas. Based on these findings, we categorized cytological nuclear atypia into three groups, nuclear atypia-1 (57.7%), nuclear atypia-2 (19.7%) and nuclear atypia-3 (22.5%), and this classification system correlated well with prognosis in patients with endometrial cancer (P < 0.001). Furthermore, this classification system was able to extract patients with a good prognosis from those with high-grade carcinomas, such as UPSC+CC+EEC G3, and patients with a poor prognosis from those with EEC G1. CONCLUSIONS: Our system of cytological nuclear atypia classification based on endometrial cytology can predict patient prognosis. Cytological nuclear atypia classification and histological typing may be useful for the treatment and follow-up of patients with endometrial cancer, and should be routinely incorporated into cytological reports.

Association of PD-L1 overexpression with activating EGFR mutations in surgically resected nonsmall-cell lung cancer.

BACKGROUND: Recent clinical trials have shown that immune-checkpoint blockade yields a clinical response in a subset of individuals with advanced nonsmall-cell lung cancer (NSCLC). We examined whether the expression of programmed death-ligand 1 (PD-L1) is related to clinicopathologic or prognostic factors in patients with surgically resected NSCLC. PATIENTS AND METHODS: The expression of PD-L1 was evaluated by immunohistochemical analysis in 164 specimens of surgically resected NSCLC. Cell surface expression of PD-L1 in NSCLC cell lines was quantified by flow cytometry. RESULTS: Expression of PD-L1 in tumor specimens was significantly higher for women than for men, for never smokers than for smokers, and for patients with adenocarcinoma than for those with squamous cell carcinoma. Multivariate analysis revealed that the presence of epidermal growth factor receptor gene (EGFR) mutations and adenocarcinoma histology were significantly associated with increased PD-L1 expression in a manner independent of other factors. Cell surface expression of PD-L1 was also significantly higher in NSCLC cell lines positive for activating EGFR mutations than in those with wild-type EGFR. The EGFR inhibitor erlotinib downregulated PD-L1 expression in the former cell lines but not in the latter, suggesting that PD-L1 expression is increased by EGFR signaling conferred by activating EGFR mutations. A high level of PD-L1 expression in resected tumor tissue was associated with a significantly shorter overall survival for NSCLC patients. CONCLUSIONS: High expression of PD-L1 was associated with the presence of EGFR mutations in surgically resected NSCLC and was an independent negative prognostic factor for this disease.

Jun dimerization protein 2 is a critical component of the Nrf2/MafK complex regulating the response to ROS homeostasis.

Oxidative stress and reactive oxygen species (ROS) are associated with diseases such as cancer, cardiovascular complications, inflammation and neurodegeneration. Cellular defense systems must work constantly to control ROS levels and to prevent their accumulation. We report here that the Jun dimerization protein 2 (JDP2) has a critical role as a cofactor for transcription factors nuclear factor-erythroid 2-related factor 2 (Nrf2) and small Maf protein family K (MafK) in the regulation of the antioxidant-responsive element (ARE) and production of ROS. Chromatin immunoprecipitation-quantitative PCR (qPCR), electrophoresis mobility shift and ARE-driven reporter assays were carried out to examine the role of JDP2 in ROS production. JDP2 bound directly to the ARE core sequence, associated with Nrf2 and MafK (Nrf2-MafK) via basic leucine zipper domains, and increased DNA-binding activity of the Nrf2-MafK complex to the ARE and the transcription of ARE-dependent genes. In mouse embryonic fibroblasts from Jdp2-knockout (Jdp2 KO) mice, the coordinate transcriptional activation of several ARE-containing genes and the ability of Nrf2 to activate expression of target genes were impaired. Moreover, intracellular accumulation of ROS and increased thickness of the epidermis were detected in Jdp2 KO mice in response to oxidative stress-inducing reagents. These data suggest that JDP2 is required to protect against intracellular oxidation, ROS activation and DNA oxidation. qPCR demonstrated that several Nrf2 target genes such as heme oxygenase-1, glutamate-cysteine ligase catalytic and modifier subunits, the notch receptor ligand jagged 1 and NAD(P)H dehydrogenase quinone 1 are also dependent on JDP2 for full expression. Taken together, these results suggest that JDP2 is an integral component of the Nrf2-MafK complex and that it modulates antioxidant and detoxification programs by acting via the ARE.

Convergent evolution of morphology and habitat use in the explosive Hawaiian fancy case caterpillar radiation.

Species occurring in unconnected, but similar habitats and under similar selection pressures often display strikingly comparable morphology, behaviour and life history. On island archipelagos where colonizations and extinctions are common, it is often difficult to separate whether similar traits are a result of in situ diversification or independent colonization without a phylogeny. Here, we use one of Hawaii's most ecologically diverse and explosive endemic species radiations, the Hawaiian fancy case caterpillar genus Hyposmocoma, to test whether in situ diversification resulted in convergence. Specifically, we examine whether similar species utilizing similar microhabitats independently developed largely congruent larval case phenotypes in lineages that are in comparable, but isolated environments. Larvae of these moths are found on all Hawaiian Islands and are characterized by an extraordinary array of ecomorphs and larval case morphology. We focus on the 'purse cases', a group that is largely specialized for living within rotting wood. Purse cases were considered a monophyletic group, because morphological, behavioural and ecological traits appeared to be shared among all members. We constructed a phylogeny based on nuclear and mitochondrial DNA sequences from 38 Hyposmocoma species, including all 14 purse case species and 24 of non-purse case congeners. Divergence time estimation suggests that purse case lineages evolved independently within dead wood and developed nearly identical case morphology twice: once on the distant Northwest Hawaiian Islands between 15.5 and 9 Ma and once on the younger main Hawaiian Islands around 3.0 Ma. Multiple ecomorphs are usually found on each island, and the ancestral ecomorph of Hyposmocoma appears to have lived on tree bark. Unlike most endemic Hawaiian radiations that follow a clear stepwise progression of colonization, purse case Hyposmocoma do not follow a pattern of colonization from older to younger island. We postulate that the diversity of microhabitats and selection from parasitism/predation from endemic predators may have shaped case architecture in this extraordinary endemic radiation of Hawaiian insects.

Efficacy of autologous fat injection laryngoplasty with an adenoviral vector expressing hepatocyte growth factor in a canine model.

OBJECTIVE: The effectiveness of autologous fat injection laryngoplasty may be reduced by resorption of injected fat tissue. The aim of the present study was to clarify the efficacy of fat injection laryngoplasty using autologous fat plus a replication-defective adenoviral vector expressing hepatocyte growth factor, regarding reduction of injected fat tissue resorption. MATERIAL AND METHODS: Four female beagle dogs were used in this study. After sedation, a direct laryngoscope was introduced to enable visualisation of the larynx. In each dog, harvested autologous fat plus an adenoviral vector expressing hepatocyte growth factor was injected into the right true vocal fold, and harvested fat plus an adenoviral vector expressing no gene was injected into the left true vocal fold. A total laryngectomy was performed one year after the intracordal fat injection. Coronal sections of the resected whole larynges were made and the following parameters assessed using light and electron microscopy: size of fat area; number of vasculoendothelial cells surrounding adipocytes; and shape of injected adipocytes in the vocal fold. RESULTS: The fat area was significantly larger and the number of vasculoendothelial cells surrounding adipocytes significantly greater in the intracordal fat injection containing adenoviral vector expressing hepatocyte growth factor, compared with the control intracordal fat injection containing adenoviral vector expressing no gene. When viewed under electron microscopy, the injected adipocytes were observed to have grafted better in the intracordal fat injection with hepatocyte growth factor adenoviral vector, compared with the control intracordal fat injection with adenoviral vector expressing no gene. CONCLUSIONS: Injection into the vocal fold of autologous fat containing an adenoviral vector expressing hepatocyte growth factor can reduce subsequent resorption of injected fat.

Higher expression of deoxyuridine triphosphatase (dUTPase) may predict the metastasis potential of colorectal cancer.

AIMS: 5-Fluorouracil (5-FU) is one of the most widely used anticancer drugs; however, the activity of 5-FU is determined by the presence of several enzymes that limit its activation or degradation, and these include dihydropyrimidine dehydrogenase (DPD), orotate phosphoribosyl transferase (OPRT), thymidylate synthase (TS), thymidine kinase (TK), thymidine phosphorylase (TP) and deoxyuridine triphosphatase (dUTPase). The aim of this study was to compare the expression levels of these enzymes between the primary colorectal cancer of patients with and without distant metastases. Furthermore, there was a comparison of these expression levels between the primary tumour and the corresponding metastasis. METHODS: Of 55 patients with colorectal cancer, 20 had no metastasis and the other 35 had distant metastasis. A strong expression was classified as positive, while weak to moderate or no expression was negative by immunohistochemistry. RESULTS: Of the six 5-FU-related enzymes, the numbers of patients with expression of dUTPase (54% versus 15%; p = 0.005), TK (26% versus 0%; p = 0.019) and DPD (17% versus 45%; p = 0.033) were significantly different in those with primary tumours with metastasis compared with those with non-metastasis, respectively. The altered expression of OPRT (34.3%), TS (40.0%) and dUTPase (42.9%) was significantly greater from primary to metastasis among the 35 patients with metastasis. By contrast, the expression of OPRT, TS and dUTPase was decreased in 6, 5 and 7 patients, respectively, in metastatic sites. CONCLUSIONS: From this comparative study of the six 5-FU-related enzymes in colorectal cancer, the expression of dUTPase was most significantly different between primary tumours and their corresponding metastatic tumour. It is suggested that dUTPase may be a predictive biomarker for the metastatic potential of colorectal cancer.

Critical role of biklf in erythroid cell differentiation in zebrafish.

Hematopoietic cells arise from ventral mesoderm in different vertebrates, but the mechanisms through which various factors contribute to the hematopoietic processes, including erythrogenesis, remain incompletely understood. The Krüppel-like transcription factor Biklf is preferentially expressed in blood islands throughout zebrafish embryogenesis, marking the region of future erythropoiesis [1]. In this paper, we show that expression of biklf is significantly suppressed in the blood-less mutants vampire and m683 in which primitive hematopoiesis is impaired. Knockdown of biklf using morpholino-based antisense oligonucleotides (biklf-MO) led to a potent reduction in the number of circulating blood cells and deficiency in hemoglobin production. Consistently, we found that the expression of beta(e3)globin is strongly suppressed in biklf-MO-injected embryos, while gata1 expression is partly inhibited at the 10-somite stage. In addition, analysis of reporter constructs driven by the GATA1 and beta-globin promoters showed that Biklf can positively regulate both genes. These results indicate that Biklf is required for erythroid cell differentiation in zebrafish.

Necrotic death pathway in Fas receptor signaling.

A caspase 8-deficient subline (JB6) of human Jurkat cells can be killed by the oligomerization of Fas-associated protein with death domain (FADD). This cell death process is not accompanied by caspase activation, but by necrotic morphological changes. Here, we show that the death effector domain of FADD is responsible for the FADD-mediated necrotic pathway. This process was accompanied by a loss of mitochondrial transmembrane potential (DeltaPsim), but not by the release of cytochrome c from mitochondria. Pyrrolidine dithiocarbamate, a metal chelator and antioxidant, efficiently inhibited the FADD-induced reduction of DeltaPsim and necrotic cell death. When human Jurkat, or its transformants, expressing mouse Fas were treated with Fas ligand or anti-mouse Fas antibodies, the cells died, showing characteristics of apoptosis. A broad caspase inhibitor (z-VAD-fmk) blocked the apoptotic morphological changes and the release of cytochrome c. However, the cells still died, and this cell death process was accompanied by a strong reduction in DeltaPsim, as well as necrotic morphological changes. The presence of z-VAD-fmk and pyrrolidine dithiocarbamate together blocked cell death, suggesting that both apoptotic and necrotic pathways can be activated through the Fas death receptor.

Identification of four highly conserved regions in DNA-PKcs.

The gene for the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is responsible for severe combined immune deficiency (SCID) and its products associate with Ku70/Ku86 autoantigens, c-Abl, and other factors to exert its roles. Investigations to date suggest that DNA-PKcs comprises several regions which specifically interact with these known and other as yet unidentified factors. Nevertheless, the relationship between the structure and function of the DNA-PKcs molecule is poorly understood. Here we report cloning of the entire DNA-PKcs cDNA from chicken and Xenopus laevis. Comparative study of the DNA-PKcs polypeptides from four different vertebrates revealed at least three novel conserved regions in addition to the carboxyl-terminal region containing the phosphatidylinositol-3 kinase domain. We also demonstrated that the four vertebrates share the common genomic organization between a licensing factor, MCM4, and DNA-PKcs, both of which locate in a head-to-head manner within a few kilobase intervals. These data provide useful clues for the further genetic study of this huge multifunctional enzyme.

Cytokine-induced nitric oxide production inhibits mitochondrial energy production and impairs contractile function in rat cardiac myocytes.

OBJECTIVES: The present study examined whether nitric oxide (NO) produced by inducible nitric oxide synthase (iNOS) can directly inhibit aerobic energy metabolism and impair cell function in interleukin (IL)-1beta,-stimulated cardiac myocytes. BACKGROUND: Recent reports have indicated that excessive production of NO induced by cytokines can disrupt cellular energy balance through the inhibition of mitochondrial respiration in a variety of cells. However, it is still largely uncertain whether the NO-induced energy depletion affects myocardial contractility. METHODS: Primary cultures of rat neonatal cardiac myocytes were prepared, and NO2-/NO3- (NOx) in the culture media was measured using Griess reagent. RESULTS: Treatment with IL-1beta (10 ng/ml) increased myocyte production of NOx in a time-dependent manner. The myocytes showed a concomitant significant increase in glucose consumption, a marked increase in lactate production, and a significant decrease in cellular ATP (adenosine 5'-triphosphate). These metabolic changes were blocked by co-incubation with N(G)-monomethyl-L-arginine (L-NMMA), an inhibitor of NO synthesis. Sodium nitroprusside (SNP), a NO donor, induced similar metabolic changes in a dose-dependent manner, but 8-bromo-cyclic guanosine 3',5'-monophosphate (8-bromo-cGMP), a cGMP donor, had no effect on these parameters. The activities of the mitochondrial iron-sulfur enzymes, NADH-CoQreductase and succinate-CoQreductase, but not oligomycin-sensitive ATPase, were significantly inhibited in the IL-1beta, or SNP-treated myocytes. Both IL-1beta and SNP significantly elevated maximum diastolic potential, reduced peak calcium current (I(Ca)), and lowered contractility in the myocytes. KT5823, an inhibitor of cGMP-dependent protein kinase, did not block the electrophysiological and contractility effects. CONCLUSIONS: These data suggest that IL-1beta-induced NO production in cardiac myocytes lowers energy production and myocardial contractility through a direct attack on the mitochondria, rather than through cGMP-mediated pathways.

Inhibition of Fas-induced apoptosis by Bcl-2.

Jurkat cells express Fas, and rapidly undergo apoptosis in response to Fas ligand or an agonistic anti-Fas antibody. This apoptotic pathway is mediated by a cascade of caspases. In this report, we show that Fas activation induced the processing of caspase 8 in Jurkat cells with a time frame similar to the activation of caspase 3 and the proteolysis of nuclear proteins. Jurkat cell transformants that overexpress Bcl-2 were partially but not completely resistant to the Fas-induced apoptosis. Little processing of caspase 8 was observed upon Fas activation in these transformants. Furthermore, although caspase 8 was recruited to Fas upon Fas activation in the parental Jurkat cells, the recruitment of caspase 8 to Fas was inhibited in the transformants overexpressing Bcl-2. These results suggest that Bcl-2 inhibits Fas-induced apoptosis by preventing the formation of the death-inducing signaling complex that is composed of Fas, FADD/MORT1, and caspase 8.

Caspase-independent cell killing by Fas-associated protein with death domain.

The binding of Fas ligand to Fas recruits caspase 8 to Fas via an adaptor, FADD/MORT1, and activates a caspase cascade leading to apoptosis. Here, we describe a human Jurkat-derived cell line (JB-6) that is deficient in caspase 8. This cell line was resistant to the apoptosis triggered by Fas engagement. However, the multimerization of Fas-associated protein with death domain, through the use of a dimerizing system, killed the JB-6 cells. This killing process was not accompanied by the activation of caspases or DNA fragmentation. The dying cells showed neither condensation nor fragmentation of cells and nuclei, but the cells and nuclei swelled in a manner similar to that seen in necrosis. These results suggested that Fas-associated protein with death domain can kill the cells via two pathways, one mediated by caspases and another that does not involve them.

Fas-induced DNA fragmentation and proteolysis of nuclear proteins.

BACKGROUND: Fas is a member of the tumour necrosis factor (TNF) receptor family. Activation of Fas by its ligand or an agonistic anti-Fas antibody causes apoptosis in Fas-bearing cells, by activating various members of the caspase family. RESULTS: Specific fluorogenic substrates (MCA-DEVDAPK[dnp] and MCA-VEVDAPK[dnp]) for caspases 3 and 6 were prepared. Using these substrates, a gradual increase of the caspase 3-and 6-like proteases were detected during the Fas engagement in human Jurkat. This activation of caspases correlated well with the cleavage of poly(ADP-ribose) polymerase and lamin B1, as well as with DNA fragmentation. When the recombinant caspases were added to the extracts from Jurkat cells, caspase 3 produced active caspase 6-like protease, while caspase 6 activated the caspase 3 protease, suggesting that these proteases can activate each other. The caspase-treated cell extracts, as well as the extracts from the Fas-activated cells, caused the proteolysis of nuclear proteins and DNA degradation. The cleavage of nuclear proteins was inhibited by caspase inhibitors, while the same inhibitors had no effect on DNA degradation. CONCLUSIONS: At one stage of the caspase cascade, caspases activate each other, and amplify the apoptotic signal. Caspases downstream of the cascade then cause the proteolysis of nuclear proteins and DNA degradation.

Energy metabolism after ischemic preconditioning in streptozotocin-induced diabetic rat hearts.

OBJECTIVES: The aim of this study was to compare the cardioprotective effects of preconditioning in hearts from streptozotocin-induced diabetic rats with its effects in normal rat hearts. BACKGROUND: The protective effect of ischemic preconditioning against myocardial ischemia may come from improved energy balance. However, it is not known whether preconditioning can also afford protection to diabetic hearts. METHODS: Isolated perfused rat hearts were either subjected (preconditioned group) or not subjected (control group) to preconditioning before 30 min of sustained ischemia and 30 min of reperfusion. Preconditioning was achieved with two cycles of 5 min of ischemia followed by 5 min of reperfusion. RESULTS: In the preconditioned groups of both normal and diabetic rats, left ventricular developed pressure, high energy phosphates, mitochondrial adenosine triphosphatase and adenine nucleotide translocase activities were significantly preserved after ischemia-reperfusion; cumulative creatine kinase release was smaller during reperfusion; and myocardial lactate content was significantly lower after sustained ischemia. However, cumulative creatine kinase release was less in the preconditioned group of diabetic rats than in the preconditioned group of normal rats. Under ischemic conditions, more glycolytic metabolites were produced in the diabetic rats (control group) than in the normal rats, and preconditioning inhibited these metabolic changes to a similar extent in both groups. CONCLUSIONS: The present study demonstrates that in both normal and diabetic rats, preservation of mitochondrial oxidative phosphorylation and inhibition of glycolysis during ischemia can contribute to preconditioning-induced cardioprotection. Furthermore, our data suggest that diabetic myocardium may benefit more from preconditioning than normal myocardium, possibly as a result of the reduced production of glycolytic metabolites during sustained ischemia and the concomitant attenuation of intracellular acidosis.

Glutathione S-transferase M1 (GSTM1) and T1 (GSTT1) genetic polymorphism and susceptibility to gastric and colorectal adenocarcinoma.

Genes coding for the glutathione S-transferase M1 (GSTM1) and Theta 1 (GSTT1) proteins are polymorphic in humans and these genes are absent, or homozygous null, in 10-60% of different ethnic populations. These enzymes catalyze the conjugation of glutathione to numerous carcinogenic chemicals and previous epidemiologic studies have associated the null genotypes of these GST genes with higher risk of cancer. In this study the frequency of GSTM1 and GSTT1 null genotypes was determined in Japanese patients with gastric adenocarcinoma and colorectal adenocarcinoma and compared to frequencies determined in a community-based control group. The frequency of the null GSTM1 genotype in patients with gastric adenocarcinoma (56.8%) showed a statistically significant increase compared to the control group frequency (43.6%) (odds ratio (OR) = 1.70; 95% CI, 1.05-2.76). The frequency of GSTM1 null individuals was also higher among all colorectal adenocarcinoma cases, but this increase did not reach statistical significance. After grouping by tumor site, the GSTM1 null genotype was a risk factor among the subgroup with distal colorectal tumors (61.1%) (OR = 2.03; 95% CI, 1.06-3.90). No consistent difference was observed between smoking patients and corresponding controls for the frequency of the GSTM1 null genotype for either cancer, although a large risk (OR = 5.76; 95% CI 1.18-28.3) was associated with the GSTM1 null genotype in the low smoking group of gastric adenocarcinoma patients. On the other hand, no statistically significant differences were observed in the frequency of null GSTT1 genotypes in gastric (47.5%) or colorectal (48.5%) adenocarcinoma patients when compared with the control population (44.4%). These results suggest that the GSTM1 null genotype may be associated with susceptibility to gastric adenocarcinoma and distal colorectal adenocarcinoma in Japanese; however, the associations observed were relatively weak and additional studies will be needed to confirm these findings.