Impacts of skipping breakfast and late dinner on the incidence of being overweight: a 3-year retrospective cohort study of men aged 20-49 years.

BACKGROUND: Most studies on the dietary habits and overweight status of men aged 20-49 years have been cross-sectional, with longitudinal studies being scarce. One-quarter of Japanese men aged 20-49 years skip breakfast or have dinner within 2 h of bedtime (late dinner); therefore, the effects of these eating habits on men's increasing body weight need to be determined. METHODS: We conducted a retrospective cohort study using health check-up data provided from several health insurance societies in Japan. Participants comprised 45 524 men employees aged 20-49 years who were followed up for 3 years. The primary outcome investigated was body mass index (BMI) ≥25 kg m-2 . We conducted a multivariable logistic regression analysis and calculated the odds ratios for skipping breakfast and late dinner, as well as baseline age, body mass index, smoking status, eating speed, snack-eating status, alcohol drinking frequency, physical activity, sleep habits, and the interaction between skipping breakfast and late dinner. RESULTS: Of the participants, 17 706 (38.8%) skipped breakfast and 25 987 (57.1%) had a late dinner. At the 3-year follow-up, 5093 (11.2%) had a BMI ≥25 kg m-2 . The odds ratios of men skipping breakfast and having a late dinner were 1.18 (95% confidence interval = 1.04-1.33) and 0.92 (95% confidence interval = 0.84-1.01), respectively. The interaction between these factors was nonsignificant. CONCLUSIONS: We suggest that skipping breakfast among men aged 20-49 years was one predictor of being overweight; however, having dinner within 2 h of bedtime was not a predictor.

Early-stage malignant peripheral nerve sheath tumour arising from a solitary neurofibroma.

We report a case of early-stage malignant peripheral nerve sheath tumour (MPNST) found in a solitary neurofibroma, and its CT and MRI findings. A 19-year-old male with no known history of a neurofibromatosis presented with a painless swelling in the left forearm. CT and MRI scans showed a well-circumscribed, intermuscular mass, which was 6.0 cm in diameter and contained a strongly enhanced 1.0 cm nodular structure with surrounding oedema. Peripheral nerve continuity with the mass was not seen. Histological evaluation proved the nodular structure was an MPNST component completely surrounded by neurofibroma. Following an excisional biopsy with wide margins, the patient was followed up for a year without treatment and no recurrence was observed.

Accumulation of NEDD8 in neuronal and glial inclusions of neurodegenerative disorders.

NEDD8 (neural precursor cell expressed, developmentally down-regulated 8) is a ubiquitin-like protein that controls vital biological events through its conjugation to members of the cullin family, which are components of certain ubiquitin E3 ligases. Recent studies have shown that NEDD8 is incorporated into Lewy bodies (LBs) in Parkinson's disease, Mallory bodies in alcoholic liver disease and Rosenthal fibres in astrocytoma. In order to examine whether NEDD8 plays a role in the formation of ubiquitinated inclusions, we performed immunohistochemical staining of brain tissue from patients with various neurodegenerative disorders, using an affinity-purified polyclonal antibody raised against NEDD8 that did not cross-react with ubiquitin. In LB disease, NEDD8 immunoreactivity was present in almost all of the LBs and Lewy neurites. Moreover, NEDD8 immunoreactivity was found in a variety of ubiquitinated inclusions, including neuronal and oligodendroglial inclusions in multiple system atrophy, neurofibrillary tangles in Alzheimer's disease, ubiquitinated inclusions in motor neurone disease, and intranuclear inclusions in triplet repeat diseases. These findings suggest that NEDD8 is involved in the formation of various ubiquitinated inclusions via the ubiquitin-proteasome system.

Incidence of p53 and ras gene mutations in DMBA-induced rat leukemias.

Leukemia, a form of haematological malignancy, is a multi-stage disease and a wide range of diverse genes has been speculated to correlate with its initiation and development. Ras has been speculated to be an initiating gene for haematological malignancy, but more investigation will be needed to determine the genes associated with the progression of the disease. 7,12-dimethylbenz(a)anthracene (DMBA)-induced rat leukemia provides a good tool for research into various stages of the disease. The entire coding regions of p53 and ras genes were examined for mutations in the present study. In this experiment, we used fluorescence-labeled polymerase chain reaction single-stranded conformation polymorphism analysis (PCR-SSCP) and direct sequencing to detect mutations of both genes on rat erythroleukemia. Fifteen out of 18 (83.3%) rat leukemias were found to have N-ras codon 61 mutation, consistent with previous results. The result of direct sequencing showed a single base substitution (CAA to CTA), resulting in an amino-acid change from Gln to Leu. No mutations were found in H-ras, K-ras or codon 12 of N-ras. The incidence of p53 gene mutation was 16.6% (3/18) in rat leukemia at late-stage. In the present study, mutation of the p53 gene was detected in three DMBA-induced leukemias as follows: a single-base substitution (CAT to CGT) at codon 177 (exon 5), resulting in an amino-acid change from Arg to Leu, a CGG to CTG/CGG changed at codon 211 (exon 6) resulting in an amino-acid change from His to Arg/His, and a GGG to TGG at codon 242 (exon 6) resulting in an amino-acid change from Gly to Trp, respectively. Thus, mutations of p53 gene do not seem to respond to the carcinogenesis of the DMBA-induced leukemia, in contrast to mutation of the N-ras oncogene, and may possibly be involved in the progress of multi-stage leukemogenesis.

Targeting of NEDD8 and its conjugates for proteasomal degradation by NUB1.

NEDD8 is a ubiquitin-like protein that controls vital biological events through its conjugation to cullin family members. Recently, we identified a negative regulator of the NEDD8 conjugation system, NUB1, which interacts with NEDD8 and down-regulates NEDD8 expression post-transcriptionally (Kito, K., Yeh, E. T. H., and Kamitani, T. (2001) J. Biol. Chem. 276, 20603-20609). Here, we show that NUB1 possesses a ubiquitin-like domain at the N-terminal region and binds to S5a of the 19 S proteasome activator (PA700). A GST pull-down assay revealed that the overexpression of NUB1 leads to a greater precipitation of NEDD8 conjugates with GST-S5a, suggesting that NUB1 might have an adaptor function between S5a and NEDD8. Furthermore, proteasome inhibitors completely block NUB1-mediated down-regulation of NEDD8 expression. These results suggest that NUB1 recruits NEDD8 and its conjugates to the proteasome for degradation, providing a direct functional link between the NEDD8 conjugation system and the proteasomal degradation pathway.

Cloning and characterization of a p53-related protein kinase expressed in interleukin-2-activated cytotoxic T-cells, epithelial tumor cell lines, and the testes.

A human protein kinase, p53-related protein kinase (PRPK), was cloned from an interleukin-2-activated cytotoxic T-cell subtraction library. PRPK appears to be a homologue of a growth-related yeast serine/threonine protein kinase, YGR262c. However, a complementation assay using YGR262c-disrupted yeast indicated that PRPK is not functionally identical to the yeast enzyme. PRPK expression was observed in interleukin-2-activated cytotoxic T-cells, some human epithelial tumor cell lines, and the testes. The intrinsic transcriptional activity of p53 was up-regulated by a transient transfection of PRPK to COS-7 cells. PRPK was shown to bind to p53 and to phosphorylate p53 at Ser-15. These results indicate that PRPK may play an important role in the cell cycle and cell apoptosis through phosphorylation of p53.

MCP-1 receptor binding affinity is up-regulated by pre-stimulation with MCP-1 in an actin polymerization-dependent manner.

Monocyte chemoattractant protein-1 (MCP-1) induces monocyte chemotaxis via interaction with the MCP-1 receptor CCR2. We found that MCP-1 binding to monocytic THP-1 cells was increased by pre-treatment with MCP-1. The amount of CCR2 mRNA and the cell-surface expression of CCR2 were not affected by MCP-1 stimuli. In contrast, the MCP-1-treated THP-1 cells showed a sixfold increase in MCP-1 binding affinity compared with untreated cells. MCP-1 binding to CCR2B-transfected HEK-293 cells was also enhanced by pre-treatment with MCP-1, and MCP-1 binding affinity increased by sixfold. In both cell lines, the enhancement of MCP-1 binding by stimulation with MCP-1 was blocked by cytochalasin D, an inhibitor of actin polymerization. This effect of pre-treatment with MCP-1 is insensitive to pertussis toxin and partially blocked by U73122, an inhibitor of phospholipase C. These results demonstrate that the MCP-1 receptor binding affinity is up-regulated by MCP-1 stimuli in an actin polymerization-dependent manner.

NUB1, a NEDD8-interacting protein, is induced by interferon and down-regulates the NEDD8 expression.

NEDD8, a ubiquitin-like protein, covalently conjugates to cullin family members. It appears to control vital biological events through its conjugation to cullins. To study how this conjugation pathway is regulated, we performed yeast two-hybrid screening by using NEDD8 as a bait and isolated a cDNA fragment encoding a potent down-regulator of the NEDD8 expression. Here, we report this novel regulator, NUB1 (NEDD8 Ultimate Buster-1). NUB1 is composed of 601 residues with a calculated 69.1-kDa molecular mass. It is an interferon-inducible protein and predominantly localized in the nucleus. The NUB1 message is specifically expressed in adult human testis, ovary, heart, and skeletal muscle tissues and is developmentally down-regulated in mouse embryos. In biochemical analysis, we found that NUB1 overexpression leads to severe reduction of NEDD8 monomer and NEDD8 conjugates in cells. This reduction is not due to down-regulation of NEDD8 transcription, but due to post-transcriptional mechanism. As expected from this activity, overexpression of NUB1 had a profound growth-inhibitory effect on U2OS cells. Thus, NUB1 is a strong down-regulator of the NEDD8 expression and appears to play critical roles in regulating biological events, including cell growth.

[Left main coronary trunk compression by dilated main pulmonary artery in a patient with atrial septal defect].

A 12-year-old girl with atrial septal defect combined with pulmonary hypertension and 90% stenosis of the left main coronary artery caused by dilated pulmonary artery was scheduled for atrial septal closure and coronary artery bypass graft under general anesthesia. During the echocardiographic examination to evaluate the anatomical relationship between the pulmonary artery and left main coronary trunk, bradycardia and a depression of ST-segment on electrocardiogram appeared suddenly when the operator compressed the pulmonary artery with a probe of echocardiography from the operative field. The circulatory collapse and ischemic change on electrocardiogram might have been caused by a further reduction of blood flow to the left main coronary trunk narrowed originally by dilated pulmonary artery. Although various etiologies, such as atherosclerosis, syphilis, and congenital abnormalities are widely known to cause stenosis of the left main coronary trunk, external compression by dilated pulmonary artery has not been widely known. Malignant arrhythmias from coronary artery compression with subsequent ischemia could contribute to an incidence of sudden death. Coronary angiography and magnetic resonance imaging are useful for the preoperative evaluation. Careful management is needed to protect such a patient from ischemic event in the perioperative period.

Characterization of complex chromosomal abnormalities in B-cell lymphoma by a combined spectral karyotyping (SKY) analysis and fluorescence in situ hybridization (FISH) using a 14q telomere probe.

We report a case of non-Hodgkin's lymphoma of unknown origin with invasion into bone marrow and brain. This case showed complex chromosomal abnormalities, including five clonal marker chromosomes (mar) and four additional materials of unknown origin (add) that could not be identified by means of conventional G-banding. Spectral karyotyping (SKY) analysis could not only determine the origin and organization of all thus far unidentified structural chromosomal abnormalities but also detect two cryptic unbalanced translocations, which had been erroneously considered to be normal on the basis of G-banding analysis, and correct one abnormality misidentified by G banding. Among these abnormalities, we identified the new partner site of the 14q32 translocation, 22q13, and the jumping translocations involving 2p23 as a new donor chromosome. Furthermore, by using fluorescence in situ hybridization (FISH) with the probes specific for the 14q telomere, we could identify the unbalanced translocation of t(3;14)(q27;q32), which had been erroneously considered to be normal chromosome 3 on the basis of not only G-banding but also of SKY analysis. This translocation is one of the most frequent chromosomal abnormalities in B-cell lymphoma, especially diffuse large cell lymphoma. After SKY and FISH analysis, the original descriptions in the G-band karyotype were modified for a total of 13 chromosomes. The combination of SKY and FISH using the 14q telomere probe was therefore considered very useful for the characterization of complex cytogenetic cases in B-cell lymphoma.

Cloning and expression of a novel MAPKK-like protein kinase, lymphokine-activated killer T-cell-originated protein kinase, specifically expressed in the testis and activated lymphoid cells.

A novel protein kinase, TOPK (T-LAK cell-originated protein kinase), was isolated from a lymphokine-activated killer T (T-LAK) cell subtraction cDNA fragment library. The open reading frame of the TOPK gene encodes a protein of 322 amino acids, possessing a protein kinase domain profile. The cap site analysis of the 5'-end of TOPK mRNA revealed two forms, a major full-length form and a minor spliced form at the 5'-site, both encoding the same protein. A BLAST homology search and phylogenetic analysis indicated that TOPK is related to dual specific mitogen-activated protein kinase kinase (MAPKK). The transfection of the TOPK gene to COS-7 cells up-regulated a phosphorylation of p38 MAPK but not ERK1/2 or SAPK/JNK. Gel precipitation study indicated that TOPK protein can be associated with p38 in vitro. Tissue distribution of TOPK mRNA expression was specific for the testis, T-LAK cells, activated lymphoid cells, and lymphoid tumors. On the other hand, deactivated T-LAK cells did not show TOPK mRNA expression. These data suggest that TOPK is a newly identified member of a novel MEK3/6-related MAPKK that may be enrolled in the activation of lymphoid cells and support testicular functions.

Generalized intraperitoneal seeding of hepatocellular carcinoma after microwave coagulation therapy: a case report.

We first describe a case of generalized intraperitoneal seeding of hepatocellular carcinoma (HCC) after microwave coagulation therapy (MCT). A 61 year-old man underwent operative MCT for an exophytic HCC, 60 mm in diameter, in segment IV of his cirrhotic liver. Despite successful tumor ablation, the serum alpha-fetoprotein levels continuously rose after MCT. Five months later, radiographic examinations delineated several perihepatic masses with hypervascularity, and the patient presented with constipation. At the second laparotomy, there were numerous small peritoneal metastases involving the entire peritoneal cavity and slightly bloody ascites. An omental mass, 50 mm in diameter, involved the transverse colon. Most of these intraabdominal masses were removed together with the involved colon. Histologically, the initial tumor was a moderately differentiated HCC, and the peritoneal masses were poorly differentiated HCCs. The patient died of rapid tumor progression and bleeding 2 months later. In conclusion, we should be aware of the possible occurrence of peritoneal seeding after MCT for HCC. Every effort should be made to prevent this serious complication, particularly in cases of superficial, large, and less differentiated HCCs.

Association of replication error positive phenotype with lymphocyte infiltration in endometrial cancers.

Microsatellite instability (MI) has been detected in certain sporadic cancers as well as in hereditary non-polyposis colorectal cancer (HNPCC). In order to determine the precise clinicopathological characteristics of MI in endometrial cancer, we examined 90 sporadic endometrial cancers (83 endometrioid adenocarcinomas, 3 adenosquamous carcinomas, 3 papillary serous carcinomas, and 1 clear cell carcinoma) and eight lesions of endometrial hyperplasia for replication error (RER) using polymerase chain reaction amplification of CA repeated microsatellite sequences at 15 loci. RER was observed in 23 (28%) of the 83 endometrioid adenocarcinomas at at least one locus and in 19 (23%) at two or more loci (RER+ phenotype) in the seven most commonly observed loci, but not in carcinomas of other histological types or in endometrial hyperplasia. Lymphocyte infiltration around carcinoma cells, which is one of the histological features seen in tumors from HNPCC, was severer in RER+ phenotype tumors (79%, 11/14) than in the RER- tumors (25%, 11/44) (marked/moderate infiltration versus slight, P < 0.001, chi 2 test), when 58 tumors with muscular invasion were examined. The RER+ phenotype was associated with a higher parity and gravidity (P < 0.05, Wilcoxon test). However, RER+ phenotype was not associated with tumor stage, histological grade, muscular invasion, lymph node metastasis or patient survival. In conclusion, MI occurs in a subset of endometrial cancers, which often show marked infiltration of lymphocytes around the tumor.

Duodenum-preserving resection of the pancreatic head for mucinous ductal ectasia without overt carcinoma.

BACKGROUND/AIMS: The clinical characteristics of mucinous ductal ectasia (MDE) of the pancreas without overt carcinoma have not been clarified. To clarify MDE and assess the optimal treatment procedure, including the technique of duodenum-preserving resection of the pancreatic head (DpRPH), we studied four patients. METHODOLOGY: Our patients consisted of three men and one woman, with a mean age of 71 years. The patients underwent DpRPH (n=3) or the pylorus-preserving Whipple procedure (PpW) (n=1). Clinicopathological features, postoperative pancreatic function, and technique to preserve duodenal blood flow were studied. RESULTS: All patients had intraductal mucin-hypersecretion and multilocular cysts lined by hyperplastic epithelium. The lesions were located in the uncinate process (n=3) or head-body (n=1) of the pancreas. DpRPH totally removed the lesions in the uncinate process. Of the three patients receiving DpRPH, dusky duodenum and a postoperative duodenal ulcer developed in two whose gastroduodenal arteries (GDA) were divided, but did not develop in one with undivided GDA. Postoperative glucose tolerance test and peptide para-aminobenzoic acid test after DpRPH showed better values than those after PpW. All patients are alive and well 22 to 40 months after surgery. CONCLUSIONS: DpRPH is a new standard for MDE. During DpRPH, preservation of the GDA and the superior portion of the pancreatic head is recommended to maintain an adequate duodenal blood flow.

Identification of three major sentrinization sites in PML.

Acute promyelocytic leukemia arises following a reciprocal chromosome translocation t(15;17), which generates PML-retinoic acid receptor alpha fusion proteins (PML-RARalpha). We have shown previously that wild type PML, but not PML-RARalpha, is covalently modified by the sentrin family of ubiquitin-like proteins (Kamitani, T., Nguyen, H. P., Kito, K., Fukuda-Kamitani, T., and Yeh, E. T. H. (1998) J. Biol. Chem. 273, 3117-3120). To understand the mechanisms underlying the differential sentrinization of PML versus PML-RARalpha, extensive mutational analysis was carried out to determine which Lys residues are sentrinized. We show that Lys65 in the RING finger domain, Lys160 in the B1 Box, and Lys490 in the nuclear localization signal contributes three major sentrinization sites. The PML mutant with Lys to Arg substitutions in all three sites is expressed normally, but cannot be sentrinized. Furthermore, the triple substitution mutant is localized predominantly to the nucleoplasm, in contrast to wild type PML, which is localized to the nuclear bodies. Thus, sentrinization of PML, in the context of the RING finger and the B1 box, regulates nuclear body formation. Furthermore, we showed that sentrinization of PML-RARalpha could be restored by overexpression of sentrin, but not by retinoic acid treatment. These studies provide novel insight into the pathobiochemistry of acute promyelocytic leukemia and the sentrinization pathway.

Characterization of a second member of the sentrin family of ubiquitin-like proteins.

Sentrin is a novel ubiquitin-like protein that can be conjugated to other proteins in a manner analogous to ubiquitination. Two additional cDNA sequences that encode proteins highly homologous to sentrin have been reported to GenBankTM. It is not known whether these sentrin-like proteins could also function as protein modifiers. In this report, a second member of the sentrin family was characterized in detail. Sentrin-2 is a 95-amino acid polypeptide that is 46% identical and 66% homologous to sentrin-1. Northern blot analysis showed that the sentrin-2 message was expressed in all tissues, but was barely detectable in the liver and placenta. The ability of sentrin-2 to conjugate to other proteins was tested by expressing hemagglutinin epitope-tagged sentrin-2 in COS cells. Western blot analysis showed that sentrin-2 could be transferred to other proteins in a pattern similar to that of sentrin-1 conjugation and had similar C-terminal processing. We further showed that both sentrin-1 and sentrin-2 could covalently modify RanGAP1, a Ran GTPase-activating protein critically involved in nuclear transport. Immunocytochemical analysis showed that sentrin-2 derivatives were highly enriched in the nucleus. Taken together, our results demonstrate that sentrin-2 is another protein modifier for the sentrinization pathway.

Increased expression of sucrase and intestinal-type alkaline phosphatase in human gastric carcinomas with progression.

The activities of sucrase, total alkaline phosphatase (total ALP) and intestinal-type alkaline phosphatase (I-ALP) were assayed in gastric carcinomas and in their surrounding mucosae from 57 patients with advanced cancers, and the localization of sucrase in 203 carcinomas, including 86 early cancers, was examined immunohistochemically using polyclonal anti-sucrase antibody. All three enzymes were active in the 57 carcinomas as well as in their surrounding mucosae, but the levels were fairly low as compared to those in normal jejunum mucosa. A considerable part of the total ALP activity in tumor specimens was assumed to be due to I-ALP itself. Increased sucrase and I-ALP were found with greater depth of invasion by undifferentiated-type carcinomas. The pattern of immunohistochemical localization of sucrase in the 203 carcinomas also clearly indicated increased expression with greater depth of invasion even in differentiated-type carcinomas.

Covalent modification of PML by the sentrin family of ubiquitin-like proteins.

PML, a RING finger protein with tumor suppressor activity, has been implicated in the pathogenesis of acute promyelocytic leukemia that arises following a reciprocal chromosomal translocation that fuses the PML gene with the retinoic acid receptor alpha (RARalpha) gene. Immunocytochemical analysis has demonstrated that PML is co-localized with a novel ubiquitin-like protein in the nuclear bodies, which could be disrupted by the PML-RARalpha fusion protein. The physical nature of this co-localization is unknown. Using a COS cell expression system, we show that PML is covalently modified by all three members of the sentrin family of ubiquitin-like proteins. Covalent modification of PML requires the conserved Gly residue near the C termini of sentrin proteins. Sentrinization of PML is highly specific because neither NEDD8 nor ubiquitin could modify PML. Similar specificity is also observed for the covalent modification of RanGAP1 by the sentrin member of ubiquitin-like proteins. These observations highlight the fine substrate specificity of the sentrinization pathway. In acute promyelocytic leukemia, two forms of PML-RARalpha fusion proteins have been reported. Remarkably, both forms of PML-RARalpha fusion proteins could not be sentrinized. Thus differential sentrinization of PML and PML-RARalpha could play an important role in regulating the biological function of PML and in the pathogenesis of acute promyelocytic leukemia.

Characterization of NEDD8, a developmentally down-regulated ubiquitin-like protein.

NEDD8 is a novel 81 amino acid polypeptide which is 60% identical and 80% homologous to ubiquitin. Northern blot analysis showed that the NEDD8 message was developmentally down-regulated. In adult tissues, NEDD8 expression was mostly restricted to the heart and skeletal muscle. Antiserum specific for NEDD8 detected a 6-kDa monomer in SK-N-SH, BJAB, and HL60 cell lysates. A 14-kDa band was also detected in BJAB, HL60, and SK-MEL28 but not in SK-N-SH and K562 cell lysates. An approximately 90-kDa band was detected in all cell lines tested. Thus, NEDD8 is likely to be conjugated to other proteins in a manner analogous to ubiquitination. However, the conjugation pattern of NEDD8 is entirely different from that of ubiquitin in all cell lines tested. To study NEDD8 conjugation in more detail, hemagglutinin-epitope-tagged NEDD8 was expressed in COS cells. Western blot analysis revealed an NEDD8 monomer and a series of higher molecular weight NEDD8-conjugated proteins or NEDD8 multimers. Immunocytochemical analysis showed that NEDD8 expression was highly enriched in the nucleus and was much weaker in the cytosol. In contrast, ubiquitin expression was detectable equally well in the nucleus and cytosol. Mutational analysis showed that the C terminus of NEDD8 was efficiently cleaved and that Gly-76 was required for conjugation of NEDD8 to other proteins. Taken together, NEDD8 provides another substrate for covalent protein modification and may play a unique role during development.