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The instability of microsatellite sequences dispersed in the genome has been linked to a deficiency in cellular mismatch repair. This phenotype has been frequently observed in various human neoplasms and is regarded as a major factor in tumorigenesis. To demonstrate alterations in microsatellite sequences, polymerase chain reaction (PCR) and electrophoretic analysis are techniques often used. However, the electrophoretic profiles of PCR-amplified microsatellite sequences have not been well characterized. Moreover, the conventional method using autoradiography has critical problems in detection characteristics and migration accuracy. We made use of fluorescence-labeled PCR and laser scanning with linear detection characteristics, so as to detect bands quantitatively. Next, we characterized Taq polymerase-dependent modification of the amplified microsatellite sequences, using artificially synthesized microsatellite alleles and we optimized the electrophoretic profiles by enzymatic modification with T4 DNA polymerase. We developed a dual fluorescence co-electrophoresis system, in which both samples derived from cancer and normal tissues are electrophoresed in the same lane, in order to minimize migration errors. These improvements remarkably facilitate precise and objective assessments of microsatellite instability. Analyzing many positive cases in cell lines and tissue specimens, we classified all the patterns of microsatellite alteration and set up new criteria for assessing microsatellite instability.

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The incidence of gastric cancer is much higher in Japan than in other countries even though diagnostics and treatments of such patients have improved. The objective of this study was to present an overview of the past, present and future of surgical treatment for our patients with gastric cancer. We analysed data on 2152 Japanese men and women with gastric cancer who underwent surgical resection from 1965 to 1995 at Kyushu University in Fukuoka, Japan, based on a univariate and the multivariate analysis. We focused on time trends of surgical treatment and the postoperative outcome. Over the years, there have been favourable changes in the numbers of patients with early gastric cancer. In all cases of gastric cancer, the rate of 18% in the first six year period (group 1) was 57% in the last 5 year period (group 6). Size of the tumour was smaller, well-differentiated tumour tissue was more common, and lymphatic involvement was less frequent. Lymph node metastasis, liver metastasis and peritoneal dissemination all decreased. Extensive lymph node dissection was more frequently done and the rate of curative resection (curability A and B) increased. With increases in identifying the early stage of cancer and better perioperative care, mortality rates 30 days after the surgery greatly decreased. Multivariate analysis revealed that the 10 factors of depth of invasion, lymph node metastasis, lymph node dissection, tumour size, liver metastasis, peritoneal dissemination, lymphatic invasion, vascular invasion, lesion in the whole stomach and lesion in the middle stomach were independent factors for determining the prognosis. Detection of the tumour in an early stage, standardized surgical treatment, including routine lymph node dissection, close follow-up schedules and better perioperative management are expected to increase survival time for patients with this malignancy. © 2000 Cancer Research Campaign

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Jun and Fos, the major components of the AP-1 transcription factor family, are induced by various intra-or extracellular signals that trigger cell proliferation or differentiation (reviewed in references 7, 22, 32, and 48). Among five members of Fos family proteins (c-Fos, FosB, AFosB, Fra-1, and Fra-2), AFosB is unique in several respects (9,40,44,57,59). It is a truncated form of FosB that lacks the C-terminal 101 amino acids of FosB and is formed by an alternatively spliced mRNA of fosB. Like other Fos proteins, AFosB is induced by serum stimulation and forms heterodimers with each of the Jun proteins, thereby stimulating their DNA-binding activities. However, AFosB suppresses Jun transcriptional activities, acting on an AP-1-dependent promoter, while c-Fos and FosB dramatically enhance such activities in mouse embryonic carcinoma F9 cells. Furthermore, AFosB counteracts other Fos proteins that repress the c-Fos promoter activated by serum. It has been reported that the constitutive expression of FosB but not AFosB transforms rat fibroblast 208F cells (40,57,59); hence, the C-terminal region and the transcriptional activity of FosB may be essential for cell transformation. However, recent studies on functional domains of the v-Fos protein revealed that the N-terminal region and the DNA binding domain of the protein, which is conserved throughout Fos family proteins, are sufficient for oncogenic transformation and growth stimulation (35,51,60). The N-terminal region of c-Fos protein to which the DNA binding domain of the yeast GCN4 protein is fused is capable of transforming rat embryo fibroblasts together with Ras, but the transcriptional activity of the fusion protein is much less than that of GCN4 or c-Fos itself (46). Since there are more than 70 identical and 20 similar amino acid residues between the N-terminal amino acid sequence of c-Fos and the N-terminal amino acid sequence of FosB or AFosB (61), we considered that AFosB, which carries the N-terminal region and the DNA binding domain of FosB, may have the * Corresponding author.potential to regulate cell proliferation, as does a c-Fos-GCN4 fusion protein.To address this question, we set up an experimental system to express AFosB protein in Rat-lA cells. Here, we report that artificial expression of AFosB in quiescent Rat-lA cells results in reentry of the quiescent cells into the cell cycle and progression through the entire cell cycle, without activating the AP-1-responsive transin gene. MATERIALS AND METHODSPlasmids. Plasmid pcDEBA is a derivative of pcDEB (21) harboring the SRa promoter and hygromycin B phosphotransferase (hph) gene, used for selection in mammalian cells. pcDEBA lacks oriP and carries multicloning sites. Murine FosB and AFosB cDNAs (44) and estrogen receptor (ER) fusion constructs (see Fig. 2) were placed under control of the SRa promoter in pcDEBA. Plasmid HE14 carrying a cDNA region for the estrogen binding domain of human ER (30) was kindly provided by P. Chambon.Chemicals. Estrogen (3-estradiol) was obtained from Sigma. 4-Hy...

Depressed heart rate variability (HRV) in septic patients is known to be associated with poor outcome. However, neither etiology of depression of HRV nor its clinical significance has been clearly determined. Because hypercytokinemia plays an important role in sepsis, we investigated the relationships between depressed HRV and IL-6 blood level. The subjects of this study were 45 septic patients treated in our intensive care unit. IL-6 blood level upon admission exhibited significant negative correlations with two HRV indices, low-frequency power (LF) (r = -0.76; P < 0.01) and high-frequency power (HF) (r = -0.53; P < 0.01). Multivariate analysis revealed strong correlations between IL-6 blood level and LF (P = 0.01) and HF (P = 0.01), respectively, even when the effects of patient background factors and therapeutic intervention were taken into account. Among the patients who developed septic shock, a high IL-6 blood level and a low LF were observed in both the survivor and nonsurvivor groups on the day of admission. The HF was lower than normal at the same time points in both groups. However, the HF was significantly higher in the nonsurvivor group than in the survivor group. By the time of discharge from the intensive care unit, both IL-6 blood level and HRV indices had become significantly closer to the normal ranges in the survivor group, but not in the nonsurvivor group. A significant negative correlation was observed between LF upon admission and percent decline in blood pressure (r = -0.76, P < 0.01). These findings indicate that reduction in HRV indices is associated with hypercytokinemia, indicating that the autonomic nervous system and the inflammatory response mediated by the cytokine network affect each other. These results also suggest that depression of HRV is closely related to rapid changes in blood pressure. Thus, heart rate variability indices are associated with both the severity and poor outcome of sepsis.