Sensory system-predominant distribution of leukotriene A4 hydrolase and its colocalization with calretinin in the mouse nervous system.

Leukotriene B4 is a potent lipid mediator, which has been identified as a potent proinflammatory and immunomodulatory compound. Although there has been robust evidence indicating that leukotriene B4 is synthesized in the normal brain, detailed distribution and its functions in the nervous system have been unclear. To obtain insight into the possible neural function of leukotriene B4, we examined the immunohistochemical distribution of leukotriene A4 hydrolase, an enzyme catalyzing the final and committed step in leukotriene B4 biosynthesis, in the mouse nervous system. Immunoreactivity for leukotriene A4 hydrolase showed widespread distribution with preference to the sensory-associated structures; i.e. neurons in the olfactory epithelium and vomeronasal organ, olfactory glomeruli, possibly amacrine cells, neurons in the ganglion cell layer and three bands in the inner plexiform layer of the retina, axons in the optic nerve and tract up to the superior colliculus, inner and outer hair cells and the spiral ganglion cells in the cochlea, vestibulocochlear nerve bundle, spinal trigeminal tract, and lamina II of the spinal cord. Double immunofluorescence staining demonstrated that most of the leukotriene A4-hydrolase-immunopositive neurons coexpressed calretinin, a calcium-binding protein in neurons. The ubiquitous distribution of leukotriene A4 hydrolase was in sharp contrast with the distribution of leukotriene C4 synthase [Shimada A, Satoh M, Chiba Y, Saitoh Y, Kawamura N, Keino H, Hosokawa M, Shimizu T (2005) Highly selective localization of leukotriene C4 synthase in hypothalamic and extrahypothalamic vasopressin systems of mouse brain. Neuroscience 131:683-689] which was confined to the hypothalamic and extrahypothalamic vasopressinergic neurons. These results suggest that leukotriene B4 may exert some neuromodulatory function mainly in the sensory nervous system, in concert with calretinin.

Induction of apoptosis in human colon carcinoma cells HT29 by sublethal cryo-injury: mediation by cytochrome c release.

Cryosurgery is an emerging treatment for human solid tumors, notably colorectal liver metastasis. Cryosurgical procedures generate a thermal gradient of from at least -50 degrees C at the center of the tumor being treated to about 0 degrees C at the periphery. Cell death occurs by necrosis in the center, while the peripheral zone of frozen tumor harbors a mix of viable and dead tissue. In order to understand the mechanisms of cell death and survival in this peripheral area at risk for tumor recurrence, we have established an in vitro freezing system that mimics in vivo conditions of sublethal injury. HT29 colon cancer cells were subjected to freezing temperatures from -6 degrees C to -36 degrees C, thawed at room temperature for 30 min and rewarmed at 37 degrees C for a period of time. Post-freeze-thaw, cryolytic cells were evaluated by trypan blue exclusive assay. We also identified apoptotic cells after rewarming by cell shrinkage, nucleic condensation, TUNEL assay, DNA fragmentation and PARP degradation. The intensity of cryolysis and apoptosis was increased by lowering the freezing temperature. At -36 degrees C, all cells were dead immediately after freeze-thaw. A kinetic analysis of cryo-induced apoptosis showed that the commitment to enter apoptosis occurred right after the freeze-thaw period and lasted less than 8 hr after rewarming. We further demonstrated that freezing triggers one of the caspase cascade involved in apoptosis: release of cytochrome c from mitochondria to cytosol, followed by activation of caspase-9 and degradation of PARP. These results indicate the death of cancer cells under cryo-treatment at sublethal freezing temperature can be attributed 2 different modes, cryolysis as well as apoptosis. HT29 cells carrying p53 mutant have very quick response for induction of apoptosis by cryo-treatment and contain an intact pathway of caspase cascade. Further studies will address if mechanisms in cells with wild-type p53 will differ.

A new mouse model with cochleo-saccular type inner ear defects.

We found a new inner ear mutant exhibiting abnormal behavior, such as circling and head shaking, in a breeding stock of SJL/J mice. The traits are inherited in a simple autosomal-recessive fashion. Animals homozygous for the responsible gene, designated cosa, show no startle response to sounds and an inability to swim. In the inner ears of cosa/cosa homozygous, but not +/cosa heterozygous adults, histopathological features of severe damage that are typical for 'cochleo-saccular' or 'spotting' mutants have been demonstrated. We suggest here that the abnormal mice carry a mutation of a gene that is developmentally switched on in the early stages of development and is involved in endolymph homeostasis.

Mouse myosin X: molecular architecture and tissue expression as revealed by northern blot and in situ hybridization analyses.

The structure of the coding region of mouse myosin X cDNA was determined. The predicted protein sequence indicated an approximately 240 kDa molecular mass with 2062 amino acids. When aligned with the structure predicted for calf myosin X (GenBank Accession No. U55042), extremely highly conserved pleckstrin homology domains and a myosin tail homology 4 domain were apparent in the tail region, suggesting their importance for myosin X's function. Northern blot analysis revealed the existence of a myosin X mRNA, 8.7 kb in size, in various mouse tissues, while a similar size of human type myosin X mRNA was recognized mainly in the testis. In addition to the adult-type transcripts in mice, a smaller embryo-specific mRNA, 4.8 kb in size, was identified in early to late embryonic stages, suggesting the presence of a shorter myosin X isoform in mouse embryos. In situ hybridization experiments with mouse testis revealed that myosin X mRNA was restricted to Sertoli cells at stages VIII-X of the spermatogenesis cycle, suggesting that myosin X is implicated in the supporting cells during the spermatid morphogenesis.

Reproducibility of diagnosis and its influence on the distribution of lung cancer by histologic type in Osaka, Japan.

The histologic types of lung cancer cases diagnosed in 1979-1980 (n=799) and 1987 (n=587) were independently reviewed by two pathologists in order to investigate the reproducibility of the diagnosis of the histologic type when the WHO classification (1981) was used. The specimens from 354 surgical cases and biopsy or cytology specimens from 1032 non-surgical cases were reviewed. The inter-observer agreement was 87.9% (kappa=0.79) for surgical cases and 81.4% (kappa=0.72) for non-surgical cases. When compared to the original diagnosis, the agreement was 86.8% (kappa=0.78) for surgical and 86.4% (kappa=0.79) for non-surgical cases in 1979-1980 and the agreement was 92.8% (kappa=0.87) for surgical and 89.1% (kappa=0.83) for non-surgical cases in 1987. By histologic type, no difference in the agreement was observed except for large cell carcinoma. The distribution of histologic types after the review differed only slightly (less than 6%) from the original distribution. This suggests that in Osaka, Japan, the diagnosis based on the WHO classification (1981) had only a limited influence on the distribution of histologic types, and is not a major reason for the changing trends in lung cancer incidence by histologic type.

[UFT-E granule compliance in postoperative adjuvant chemotherapy].

UFT-E granules were administered as postoperative adjuvant chemotherapy to patients who had undergone surgery for gastric cancer, colorectal cancer or breast cancer. After treatment for one year, the dose conditions were investigated. The subjects were patients under 75 years of age in whom malignant tumors had been confirmed histologically, who had undergone curative resections, had no marked complications, and from whom personal informed consent had been obtained. As a rule, UFT-E granules were administered orally 450 mg (t.i.d.)/day continuously for one year beginning two weeks after surgery. The dose rate was studied from patient records and the tegafur blood concentrations on the 1st, 2nd, 6th, 8th and 12th month. The appearance of complications or clinical lab test abnormalities was also checked. A total of 19 cases were compliant among the 5 gastric cancer, 10 colorectal cancer, and 7 breast cancer patients. The mean administration period was 459 days (29-879 days), and the mean completion rate was 92.5%. The complications in 4 cases (21.1%) were relatively mild. A comparison of the prescribed dosage and patient records revealed a mean dose rate of 86.3%. From these findings, long-term administration of UFT-E granules with mild side effects is considered feasible. However, to achieve high compliance, it is considered necessary to gain a clear picture of dose conditions from patients records and other sources.

Chromosomal localization of a gene responsible for vestibulocochlear defects of BUS/Idr mice: identification as an allele of waltzer.

Mice of the bustling mutant strain BUS/Idr have vestibulocochlear defects. bus/bus homozygotes, but not heterozygotes, are hyperactive and display an abnormal behavior such as circling, head bobbing and head tilting. To characterize BUS mice further, the auditory brain-stem response of the mutant was examined. In +/bus heterozygotes as well as control animals, the auditory brain-stem response was developmentally first recorded as early as 11 days of age and heterozygous and normal adults showed typical auditory brain-stem responses with five peaks in a threshold of 40-45 dB SPL. In contrast, bus/bus homozygotes showed no auditory brain-stem response at any age in response to stimuli up to 130 dB SPL, indicating that they are deaf throughout life. Linkage analysis revealed that the responsible gene, originally designated as bus, maps on chromosome 10, 1.09+/-0.9 cM distal to D10Mit127 and D10Mit59, and 0.72+/-0.51 cM proximal to three markers, D10Mit48, D10Mit112 and D10Mit258, at a site indistinguishable from that of the Albany waltzer, v(A/b). The results of allelism tests between BUS and Albany waltzer indicated that bus is allelic with v(Alb). From these data, we propose here that the bus mutation could represent another allele of waltzer, now designated v(bus).

Trends of lung cancer incidence by histologic type: a population-based study in Osaka, Japan.

We investigated trends of lung cancer incidence from 1974 to 1993 by histologic type, using data from the population-based cancer registry in Osaka, Japan. Since the proportion of cases with histologic types identified was not sufficiently high, sex- and age-specific incidence rates by histologic types were estimated assuming that the distribution of histologic types was the same across the same sex and age group regardless of reporting status. Cumulative risk from 0 to 74 years old for total lung cancer increased 1.3-fold from the period 1974-77 to 1986-89 and then plateaued in the period 1990-93 for both males and females. When divided into histologic types, cumulative risk for incidence of squamous cell carcinoma was almost constant during the study period for both males and females. During the same period, adenocarcinoma increased up to 1.4-fold for both males and females. This increase seemed to have reached a plateau recently for males, but not for females. Small cell carcinoma increased monotonously up to 1.6- to 1.7-fold for both males and females. Large cell carcinoma showed over 2-fold increase for both males and females; however, the estimates fluctuated due to the small number of cases. This study provides further evidence of a relative increase of adenocarcinoma compared to squamous cell carcinoma. Recent trends of tapering increase of lung cancer incidence should be confirmed by further observation.

Degenerative hairlets on the vestibular sensory cells in mutant bustling (BUS/Idr) mice.

The bustling mouse (BUS/Idr: bus) is a mutant mouse strain which exhibits deafness, bustling/hyperkinetic behaviour and functional disorders seemingly related to the vestibular system. This phenotype develops in homozygous (bus/bus) mice and has been shown from cross experiments to be genetically induced by a single autosomal recessive gene. We previously detected, with light and electron microscopy, post-natal degeneration of the inner ear sensory cells in homozygotes. In the present study, we examined, by electron microscopy, the development of pathological changes in the sensory epithelia of the macula acustica and crista ampullaris of homozygous mice of various ages, paying special attention to the detailed morphology of the sensory hairlets. The homozygous mice exhibited specific pathological changes: a decrease in the number of hairs; disarrangement of the kinocilium-stereocilia pattern; and, fused and/or very large stereocilia. Homozygotes also frequently exhibited apical cytoplasmic herniation, or bleb of hair cells, as well as a degenerated kinocilium in the sensory epithelium. Heterozygotes showed similar changes, but to a lesser degree and frequency. As for the vestibular organs, similar pathological changes had developed at day, 17 of gestation. These pathological findings and onset suggest that the BUS mouse may be a mutant mouse strain distinct from other reported strains which display similar behaviour, and may be a useful animal model for the study of human degenerative vestibular disorders.

Multivariate analysis of the prognostic factors of patients with unresectable synchronous liver metastases from colorectal cancer.

PURPOSE: It frequently is observed that widely varying prognoses are given for patients with the same extent of liver metastases from colorectal cancer, even though the same treatment is performed on these patients. One of the reasons for this variance is that prognostic factors for these patients have not been defined. This study was designed to elucidate which clinicopathologic factors were the most important in the prognosis of 73 patients with unresectable synchronous liver metastasis from colorectal cancer. METHODS: Univariate and multivariate analysis of 11 clinicopathologic factors were performed using the Cox proportional hazard model. Survival curves were generated using the Kaplan-Meier method. RESULTS: Extent of liver metastases was the most significant variable in this survival analysis, although the extent of lymph node metastases of the primary lesion also was significant. However, the method of treatment was not a significant determinant in the survival for patients with unresectable liver metastases. Median survival of patients with H1, H2, and H3 was 13, 12, and 6 months, respectively, and there was a significant difference between survival curves for patients with H1 and patients with H3. Median survival of patients with n0, n1, and n2 was 13, 7, and 7 months respectively, and there was a significant difference between survival curves for patients with n0 and patients with n2. Median survival of 6 patients with H1 and n0 and of 17 patients with H3 and n2 was 28 and 4 months, respectively. There was a significant difference in survival curves between these two groups. CONCLUSION: Longevity of patients with unresectable synchronous liver metastases from colorectal cancer is affected adversely by the presence of nodal metastases and extent of liver metastases. This should be considered in the planning treatment.

[The effect of intraportal chemotherapy in terms of administered dose of 5-FU].

Forty-six patients with colorectal cancer were studied for the effects of intraportal chemotherapy in terms of the administered dose of 5-FU. No liver metastases occurred in patients with a total dose of more than 4 g of 5-FU. The five-year survival rate for patients with a total dose of more than 4 g of 5-FU was 98%, which was better than that for control (72%). In patients administered 500 approximately 600 mg/body/day of 5-FU, the concentration of 5-FU in the peripheral blood was 0.022 approximately 0.027 microgram/ml. Liver dysfunction occurred in 22%, 33%, and 80% of patients administered less than 4 g, 4 approximately 4.9 g, or more than 5 g of 5-FU, respectively, but almost all of them were grade 1. These results suggest that intraportal chemotherapy administered with a total of more than 4 g of 5-FU was effective for prevention of liver metastases after resection of colorectal cancer.

Present status and research activities of cancer registries in Japan.

Population-based cancer registries in Japan were first established in the cities of Hiroshima and Nagasaki in 1957-1958 for assessing radiation effects and in Miyagi in 1959 for promoting epidemiological researches, while most other prefectures in Japan set up cancer registries as a part of their own cancer programs. This resulted in the broader use of registry data in Japan. In 1975, the Research Group for Population-based Cancer Registration in Japan was first organized with a research grant under the National Cancer Research Promotion Program. Since then, continuous efforts have been made by the Group to improve the quantity and quality of registry data and to develop methodologies to utilize it. Studies being conducted using registry data cover (1) descriptive epidemiology, (2) analytical epidemiology, (3) evaluation of screening programs, and (4) evaluation of regional cancer medical care. In 1992, 32 regional cancer registries which were operating in Japan set up the Japanese Association of Cancer Registries. However, there are still many difficulties to overcome in order to achieve completeness of reporting in registries. Further improvement of reporting rate, together with standardization of registry data are left for future efforts.

[Pharmacokinetic study in biochemical modulation therapy of UFT with leucovorin tablet].

Preoperative oral treatment with UFT and leucovorin tablet was performed. Pharmacokinetics, degree of degeneration in the tumor tissue and side effects were studied in 34 patients with colorectal cancer preoperatively given 400 mg/day of UFT with 20 mg/day of leucovorin tablet or 400 mg/day of UFT alone. Results were as follows; 1) there was no significant difference between UFTL group and UFT group regarding concentration of FdUMP in the tumor tissue. In UFTL group, concentration of FdUMP was higher in the tumor tissue of moderately differentiated adenocarcinoma than that of well differentiated adenocarcinoma. No significant differences regarding concentration of FdUMP were obtained between diploid and aneuploid groups. 2) TS inhibition rate in the tumor tissue was 66.8% in UFTL group and 53.2% in UFT group, and there was a significant difference between these two groups. TS inhibition rate in the tumor tissue was higher than that in the normal tissue either in UFTL group or UFT group. However, there were no significant differences of TS inhibition rates regarding differentiation of tumor tissue or DNA ploidy pattern. 3) Degree of degeneration of more than Grade 2 was not obtained in any patients of either UFTL or UFT group. 4) There was no change in blood laboratory data between before and after medication. Only one patient complained Grade 1 pruritus in UFTL group. These results suggest that oral biochemical modulation therapy of UFT with leucovorin tablet is effective because of pharmacokinetically high anti-tumor effect and minimal side effects.

Survival rates of childhood cancer patients in Osaka, Japan, 1975-1984.

Survival rates for childhood cancers were analyzed with a total of 2,209 cases who were registered in a population-based cancer registry in Osaka, Japan in 1975-1984. These cases were reclassified according to Birch's classification and the survival rate of each diagnostic group was calculated by Kaplan-Meier methods. Death certificate-only cases, which amounted to 3.9% of all incidence, were excluded from the calculation. The five-year cumulative survival rate for both sexes was 46% for all cancer children. Among 12 major diagnostic groups, the most favorable survival was seen in retinoblastoma (87.5%), followed by renal tumors, epithelial neoplasms, and gonadal and germ-cell tumors. The outcome was unfavorable in leukemias, sympathetic nervous system tumors, hepatic tumors and malignant bone tumors. Comparing the survival in 1975-1979 with that in 1980-1984, the rate for all childhood cancer rose from 41% to 51%. Improvement in survival was also observed in 4 groups; acute lymphocytic leukemia, acute non-lymphocytic leukemia, non-Hodgkin's lymphoma and osteosarcoma. One attributable factor for the rise of survival was proved to be improvement of medical treatment by Cox's hazard model analysis. Comparison of survival rates in Osaka with those in England and the U.S. revealed that the prognosis for acute lymphocytic leukemia and acute non-lymphocytic leukemia was less favorable in Osaka than in England and the U.S.

[Pharmacokinetic studies on continuous intraportal 5-fluorouracil infusion].

Pharmacokinetics of 5-FU administered by continuous intraportal infusion were studied by infusing 5 mg/kg, 10 mg/kg, 20 mg/kg, and 40 mg/kg of 5-FU, respectively, into the ileocecal vein for an hour. Blood samples were collected from portal vein, femoral vein and liver, and small intestine specimens were obtained at proper intervals. The rabbits were sacrificed at 120 minutes from the start of 5-FU infusion. The results were as follows: Extremely high concentrations of 5-FU in the portal vein, femoral vein and liver tissue were observed in the 5-FU group infused with 40 mg/kg. This phenomenon was suggested to rely on the effect by which the 5-FU catabolic enzyme, dihydrouracil dehydrogenase, was saturated with a large inflow of 5-FU. FdUMP concentration of the liver was lower than that of the small intestine in all groups. These results suggest that a large dose of 5-FU infusion is effective to increase the FdUMP concentration in the liver with continuous intraportal 5-FU infusion.

Interhospital differences in cancer survival: magnitude and trend in 1975-1987 in Osaka, Japan.

This study addresses the disparity in cancer survival rates among hospitals in Osaka, Japan. Using data from the Osaka Cancer Registry, four-year survival rates for stomach cancer patients (n = 8,845) diagnosed in 1976, 1981 and 1986, and lung cancer (n = 9,795) and breast cancer patients (n = 7,377) diagnosed in 1975-77, 1980-82 and 1985-87 were calculated according to four hospital categories (teaching hospitals, large hospitals: 400 + beds excluding teaching hospitals, medium-size hospitals: 150-399 beds, and small hospitals: 20-149 beds). Cox's proportional hazards model was employed with adjustment for sex, age, clinical stage at diagnosis, and treatment status. Stomach and lung cancer patients treated in large, medium-size and small hospitals showed significantly higher risks of death than those treated in teaching hospitals in 1975-87. Interhospital differences in breast cancer survival appeared to increase in 1975-87, whereas those in stomach and lung cancer survivals decreased during the same period.

[Role of population-based cancer registry in cancer epidemiology--epidemiological studies in the Cancer Registration Scheme in Osaka, Japan].

A population-based cancer registration scheme started in three areas in Japan in the 1950s solely for studying cancer incidence in their respective areas. Soon thereafter, several prefectural governments started their own schemes as part of their cancer control programs, effectively expanding the aims of cancer registration: to clarify cancer facts, to elevate the medical care for cancer patients, and to plan and evaluate cancer control programs. The Osaka Cancer Registry (OCR) started in 1962, and has been using epidemiological methods as a tool in constructing its registration scheme, as well as analyzing and utilizing registry data. This report deals with the results obtained in the OCR, classifying these results into four activities of epidemiology. 1. Clarifying cancer facts (descriptive epidemiology): The OCR has been observing incidence, medical care for cancer patients, distribution of cases by clinical stage, and the 5-year relative survival rate, and has estimated the prevalence rate, cured-case rate, and future incidence into the 21st century. Population-based data on histology and multiple cancers collected at the OCR have also contributed to the new approaches in cancer epidemiology. 2. Research on risk factors (analytical epidemiology): The OCR developed a computerized record-linkage system in 1970. This not only made registry work more effective and reliable, but many cohort studies were able to be conducted with relatively little effort and highly reliable results. The cancer case file in the OCR has been linked with the newly prepared data file of the study group, and cancer incidence among the study group has been observed. Finally, cancer risks of possible causal factors in that group have been estimated quantitatively. 3. Evaluation of control programs: Secondary prevention programs (early detection) have been conducted in Japan as major cancer control programs, because effective risk factors were not previously defined. OCR data have been used for estimating sensitivity and specificity of screening tests for various cancers, as well as for evaluating the effect of clinical work on improving survival and on decreasing cancer deaths. 4. Planning future cancer control programs: The OCR has reported on the probable rapid increase of cancer incidence in the 21st century, especially of elderly cancer cases, and cancer cases with poor survival. To control these difficult problems, new cancer programs should be urgently designed and implemented. The authors have recommended that programs be prepared by cancer site, and have already presented a detailed program for lung cancer control.

[Trends of lung cancer incidence and its prognosis in Osaka, Japan].

Lung cancer incidence figures in Japan were estimated as 30,000 for males and 11,000 for females for 1989 and represented the second and the fifth leading site of cancer, respectively, according to the Research Group for Population-based Cancer Registration in Japan. It is also estimated that lung cancer will steadily increase in the future. In relative frequencies of the major histological types, squamous cell carcinoma showed a decreasing trend, while small as well as large cell carcinomas showed an increasing trend in Osaka. In the distribution of clinical staging, the proportion of localized cases accounted for only 20% of the lung cancers in 1990. Five-year relative survival rates for lung cancer were 11.3% for all patients and 43.0% for localized cases in Osaka in 1984-1986. The rates were reported as 13.5% and 36.6% for total patients and localized cases, respectively, among caucasians from the SEER Program, a NCI project including 10 population-based cancer registries in the US. No noticeable difference was observed between Japan and the US. To control lung cancer in Japan it is considered vitally important and urgent to develop (1) new examination methods for diagnosing lung cancer in the earlier stage, (2) an efficient and effective cessation campaign for cigarettes smoking and (3) non-smoking based education.