Oxidative DNA damage in human esophageal cancer: clinicopathological analysis of 8-hydroxydeoxyguanosine and its repair enzyme.

Both internal and external oxidative stresses act on DNA and can induce carcinogenesis. 8-hydroxydeoxyguanosine (8-OHdG) is an indicator of oxidative stress and it leads to transversion mutations and carcinogenesis. 8-OHdG is excision-repaired by 8-OHdG DNA glycosylase (OGG1). The purpose of this study is to clarify the effect of oxidative DNA damage and repair enzymes on esophageal carcinogenesis. The levels of 8-OHdG and OGG1 were immunohistochemically evaluated in resected specimens, including squamous cell carcinoma (SCC) in 97 patients with esophageal cancer. Higher levels of 8-OHdG in normal esophageal epithelium were associated with a higher smoking index (P = 0.0464). The 8-OHdG level was higher in cancerous areas than in normal epithelia (P = 0.0061), whereas OGG1 expression was weaker in cancerous areas than in normal epithelia (P < 0.0001). An increase of OGG1 expression in normal epithelium was observed as 8-OHdG levels increased (P = 0.0011). However, this correlation was not observed in cancerous areas. High OGG1 expression in the cytoplasm was related to deeper tumors (P = 0.0023), node metastasis (P = 0.0065) and stage (P = 0.0019). Oxidative DNA damage, which is attributable to smoking as well as disturbances in DNA repair systems, appears to be closely related to esophageal carcinogenesis and its progression.

Fanconi anemia: genetic analysis of a human disease using chicken system.

Fanconi anemia (FA) is a rare hereditary disorder characterized by skeletal abnormalities, bone marrow failure, and an increased incidence of cancer. The basic cellular abnormality in FA has been postulated to lie in the DNA repair mechanisms because cells from FA patients display chromosomal breakage, which is particularly remarkable following induction of DNA crosslinks. However, experimental evidence for this hypothesis has been lacking. To test whether DNA repair is really defective in FA cells, we disrupted several FA genes in chicken B cell line DT40. By measuring efficiency of gene conversion and hypermutation at the Immunoglobulin locus, we have shown that DT40 FA mutant cell lines exhibited defects in homologous DNA recombination, and possibly, translesion synthesis. However, levels of sister chromatid exchange, another important cellular event mediated by HR, were not reduced, possibly indicating the role of FA genes only in a subpathway of HR. Our results indicate that chicken DT40 cells could be highly useful in molecular dissection of basic biochemical processes, which are deficient in a human genetic disorder.

Genetic changes induced in human cells in Space Shuttle experiment (STS-95).

BACKGROUND: Results of past space experiments suggest that the biological effect of space radiation could be enhanced under microgravity. To assess the radiation risk for humans during long-term spaceflight, it is very important to clarify whether human cells exhibit a synergistic effect of radiation and microgravity. HYPOTHESIS: If significant synergism occurs in human cells, genetic changes induced during spaceflight may be detected by using human tumor HCT-116 cells which are hypermutable due to a defect in the DNA mismatch repair system. METHODS: Cultured HCT-116 cells were loaded on the Space Shuttle Discovery (STS-95) and grown during the 9-d mission. After landing, many single-cell clones were isolated, microsatellite repetitive sequences in each clone were amplified by PCR, and mutations in the microsatellite loci were detected as changes in the length of PCR fragments. Mutation frequencies of ouabain-resistant phenotype were also analyzed. RESULTS: The frequencies of microsatellite mutations as well as ouabain-resistant mutations in the flight sample were similar to those of the ground control samples. Some cells were treated in space with bleomycin which mimics the action of radiation, but the frequencies of microsatellite mutations were not significantly different between the flight and the ground control samples. CONCLUSION: Under the present flight conditions, neither space radiation (about 20 mSv during this mission) nor microgravity caused excess mutations in human cells.

Class switch recombination of the chicken IgH chain genes: implications for the primordial switch region repeats.

In mammals and the amphibian, Xenopus, isotypes of antibodies have been shown to be changed through class switch recombination within the IgH chain gene locus. Here, we identified switch (S) repetitive sequences in the 5' introns of the Ig C(mu) and C(gamma) genes of the chicken. The S(mu) region is composed of two homologous regions, S(mu)1 and S(mu)2. The S(mu)1 region is an upstream 3.7 kb sequence composed of 37 repeats of a consensus sequence containing tandem repeats of the decamer ACCAGTATGG. The S(mu)2 region is a downstream 1.4 kb sequence consisting of simple tandem repeats of a decamer CCCAGTACAG. The S(gamma) region contains repeats of the decamer TATGGGGCAG. Analysis of chicken IgG-producing hybridomas revealed that the C(mu) gene was deleted from the chromosome by the recombination occurring between the S(mu) and S(gamma) regions. Recombination breakpoints at the C(mu) gene of splenocytes from an immunized chicken were scattered around the S(mu) region and two such breakpoints, the precise position of which were determined, were located within possible hairpin loop structures at the palindromic sequence of S(mu)1. A primordial palindromic sequence from which the prevalent switch repeat motifs of mammals, chickens and amphibians may have diverged is presented.

Chicken immunoglobulin mu-chain gene: germline organization and tandem repeats characteristic of class switch recombination.

We have isolated the phage clones covering the region spanning from the heavy (H)-chain joining (J) region to the end of the mu-chain gene of the chicken immunoglobulin (Ig). The distance from JH to the first exon of the mu-chain constant (C) region is approximately 13 kb, and introns between the C region exons measure more than 3 kb. These distances are significantly larger than those of known mu-chain genes. We found a region cross-hybridizing to the switch regions of the mouse C mu and C alpha genes just in front of the first exon of C mu. Partial nucleotide sequencing of this region revealed that this region consists of tandem repeats of pentamers (C/T)(C/A)CAG complementary to the mammalian switch repetitive region. These findings suggest that this region is a good candidate for a class switch region of the mu-chain gene of chicken Ig.

Signal joint of immunoglobulin V lambda 1-J lambda and novel joints of chimeric V pseudogenes on extrachromosomal circular DNA from chicken bursa.

We isolated extrachromosomal circular DNA from the bursa of 18-day chick embryos and cloned their BamHI fragments into a phage vector. We found examples of the signal joint fused by V lambda 1-J lambda rearrangement and the sequences homologous to V lambda 1 segments that showed: (1) clustered V pseudogene (psi V) germ-line segments containing new psi V26, (2) a head-to-tail duplication of psi V13-psi V12 region and (3) chimeric structures composed of 5'-V lambda 1 and 3'-psi V segments. Both intrachromosomal tandem duplications and extrachromosomal circles may be generated by unscheduled DNA synthesis and recombination. The chimeric structure of V lambda 1 joined with upstream psi V segments suggests the involvement of V gene replacement for somatic diversification of the immunoglobulin genes in addition to a mechanism of segmental gene conversion.

[Urinary trypsin inhibitor as an acute phase reactant].

Early in this century, trypsin inhibiting activity has already been recognized in patients with acute infection or renal disease. In addition to these, conditions such as coronary thrombosis, surgical operation, artificial fever by heat-killed bacilli, malignancy, leukemia, later stage of normal pregnancy, etc. have been known to cause the elevated excretion of UTI in urine. Typically, maximal excretion of UTI has been observed within one or two days after the onset. It appears that recent studies have overcome the complexity of UTI molecule. Automated measurement of urinary trypsin inhibitor (UTI) in urine sample was carried out by either enzymic or immunologic method. UTI as well as erythrocyte sedimentation rate and C-reactive protein enables us to monitor acute phase response, being confirmed in cases of abdominal surgery.

Physiologic status at 1-year follow-up of obese women engaged in a supervised conditioning program.

A follow-up study was conducted to re-evaluate a group of obese middle-aged women (n = 13), eight of whom had completed an 18-wk supervised (3 d/wk) plus unsupervised (2 d/wk) conditioning intervention program (at least 90 min per day) as the exercise plus diet group; while five of the remainder served as the control group. Each session had included a 25- to 45-min jog/run at intensities between the heart rate (HR) corresponding to lactate threshold (LT) and that slightly above the HR @ LT. During 1 year following the program, the women participated in self-controlled training such as running, aerobic dancing, or jazz dancing 2.6 +/- 1.1 d/wk. Dietary intake averaged approximately 1736 +/- 152 kcal/d at the pre-treatment, 1404 +/- 124 kcal/d at the post-treatment, and 1645 +/- 147 kcal/d 1 year after the post-treatment. Interestingly however, oxygen uptake corresponding to LT (VO2 @ LT), maximal oxygen uptake (VO2max), weight, systolic blood pressure, and the ratio of high-density lipoprotein cholesterol to total cholesterol (HDLC/TC) observed 1 year after the post-treatment were significantly different from the original pre-treatment and/or mid-treatment values. For instance, the significant 42% increase (14.7 +/- 2.4----21.3 +/- 4.2 ml/kg/min) in VO2 @ LT and 18% increase (0.284 +/- 0.106----0.335 +/- 0.093) in HDLC/TC from the pre-test to post-test were maintained throughout the 1-year follow-up period, suggesting no detrimental effect either on a cardiorespiratory fitness factor or on an anti-atherogenic factor. These findings indicate that physiologic status of obese middle-aged women engaged in a conditioning intervention program may not regress to pre-treatment status for at least one year after completion; provided they continue to participate in a 2.6-d/wk self-controlled training program with dietary intake of 1600-1700 kcal/d. Another interesting finding was that significant relationships existed between individual changes (delta) in training frequency and individual changes (delta) in physiologic variables (i.e., delta VO2max, delta VO2 @ LT, delta WT, delta fat, and delta HDLC/TC) during the follow-up study. It is concluded that, although the improved physiologic status of obese women can be maintained fairly well during 1 year following the conditioning program; continuation of training (3 d/wk or more) should be critical, either supervised or self-controlled, for successful maintenance of lost weight (8.2 +/- 2.9 kg) and improved fitness.

[Discriminant function analysis for evaluating the status of coronary heart disease risk].

We investigated to discriminate those individuals categorized by 1. obesity, 2. hypercholesterolemia, 3. hypertension, 4. low maximal oxygen uptake, 5. an abnormal electrocardiogram reflecting ischemic patterns, and/or 6. real sedentary life, from relatively healthier individuals without coronary heart disease (CHD) risk factors. One hundred and six Japanese women, aged 30 to 72 years, all of whom were in the postabsorptive state, were recruited in a series of tests for anthropometric and physiologic profiles both during the resting state and during the submaximal-maximal cycling exercise. Subjects were categorized into two groups--those who possessed four or more of the above 1, 2, 3, 4, 5, and 6 (high-CHD-risk group, n = 15) and apparently healthy individuals with a minimum number of risk factors (low-CHD-risk group, n = 83). Analyses of the data revealed that a combination of 8 variables extracted from among original 25 variables accurately classified 13/15 (87%) of high-CHD-risk group and 77/83 (93%) of low-CHD-risk group (mean = 90/98 or 92%) into their respective groups. The 8 variables were double product, Katsura index, waist girth, chest girth, TG, TC, and skinfold thicknesses at the subscapular and abdominal sites. Subsequent t-test identified significant differences between groups not only for VO2max, SBP and TC but also for DBP, LDLC, TG, Hb, HR, and HRmax. Most of these differences were of a much greater magnitude compared to the existing difference in chronological age. These findings suggest the usefulness and importance of anthropometric and blood lipid variables in the explanation of differences in the health status between high-CHD-risk women and their counterparts.

Exercise-induced lipid peroxidation and leakage of enzymes before and after vitamin E supplementation.

1. The purpose of this study was to investigate the effects of vitamin E on serum levels of malondialdehyde following the acute exhaustive exercise in human, and to determine whether the magnitude of leakage of enzyme would be affected by vitamin E supplementation. 2. Increase of malondialdehyde after exercise before vitamin E supplementation was slight (but statistically significant), however after supplementation with vitamin E, malondialdehyde level after exercise was significantly decreased. 3. Leakage of enzyme was significantly increased after exercise before vitamin E supplementation, but it was lower following exercise after vitamin E supplementation. 4. Lipid peroxidation following a bout of acute heavy exercise can be inhibited by vitamin E supplementation.

Weight Reduction in Athletes May Adversely Affect the Phagocytic Function of Monocytes.

In brief: Monocyte phagocytic function was studied in nine competitive athletes before and after a two-week weight-reduction program of calorie restriction. The phagocytic activity of monocytes before the program was higher in the athletes than in the sedentary controls, but decreased significantly after the weight-reduction program. Furthermore, plasma fibronectin concentrations and the response of lymphocytes to phytohemagglutinin decreased after the calorie restriction. These findings suggest that weight reduction by calorie restriction may adversely affect the integrity of defense mechanisms, even in healthy athletes. Thus, calorie restriction in athletes must be conducted with caution to protect them from adverse effects on their physiologic defense mechanisms.