Tumor promotion by 12-O-tetradecanoylphorbol-13-acetate in an ultra-short-term skin carcinogenesis bioassay using rasH2 mice.

Assessment of the skin tumor-promoting potential of 12-O-tetradecanoylphorbol-13-acetate (TPA) after initiation with 7,12-dimethylbenz[a]anthracene (DMBA) was conducted using rasH2 transgenic (Tg) mice and their nontransgenic (non-Tg) littermates. Mice were treated with DMBA (50 µg/100 µL acetone) on clipped back skin at the commencement of the study, and 1 week thereafter, TPA was applied at 8 µg/200 µL or 4 µg/200 µL acetone, once or twice weekly, for 7 weeks. Skin nodules were observed in the rasH2 Tg mice from week 4, and the incidence reached 100% at weeks 5 and 6. The number of skin nodules (multiplicity) in the 8-µg twice-weekly, 8-µg once-weekly, 4-µg twice-weekly, and 4-µg once-weekly groups was 62.4, 46.2, 62.6, and 36.9, respectively. The non-Tg mice also developed skin nodules, but the sensitivity to induction in the rasH2 Tg mice was higher. No nodules were observed in the acetone groups, but single nodules were apparent in the no-treatment rasH2 Tg and non-Tg groups. In conclusion, skin promotion effects could be detected within only 8 weeks in the rasH2 mice, and the concentration of 4 µg TPA once weekly was sufficient as a positive control. This short-term skin carcinogenesis bioassay using rasH2 mice could represent a useful tool for the assessment of drug and chemical safety with cutaneous treatment.

Prognostic significance of S-phase kinase-associated protein 2 and p27kip1 in patients with diffuse large B-cell lymphoma: effects of rituximab.

BACKGROUND: The F-box protein S-phase kinase-associated protein 2 (Skp2) positively regulates the G1-S transition by promoting degradation of the cyclin-dependent kinase inhibitor p27(kip1) (p27). Recent evidence has indicated an oncogenic role of Skp2 in not only carcinogenesis but also lymphomagenesis. MATERIALS AND METHODS: Clinicopathologic features and immunohistochemical expression of Skp2 and p27 were studied retrospectively in 671 patients treated with cyclophosphamide, vincristine, doxorubicin and prednisolone (CHOP) or cyclophosphamide, vincristine, doxorubicin and prednisolone plus rituximab (R-CHOP). The median follow-up periods were 43.2 months in the CHOP group (n = 425) and 24.0 months in the R-CHOP group (n = 246). RESULTS: High Skp2 or low p27 expression correlated significantly with poor overall survival (OS) and progression-free survival (P < 0.001) in both treatment groups. The prognostic value of Skp2 or p27 expression was independent of the parameters included in the International Prognostic Index by multivariate analysis. Patients with high Skp2 expression in combination with low p27 expression showed the worst survival. CONCLUSIONS: Addition of rituximab to the CHOP regimen did not provide a beneficial outcome to patients with diffuse large B-cell lymphoma with high Skp2 expression and low p27 expression. Skp2 and p27 may be useful prognostic markers in the rituximab era.

Studies of severe acute respiratory syndrome coronavirus pathology in human cases and animal models.

During the severe acute respiratory syndrome (SARS) outbreak of 2003, approximately 10% of SARS patients developed progressive respiratory failure and died. Since then, several animal models have been established to study SARS coronavirus, with the aim of developing new antiviral agents and vaccines. This short review describes the pathologic features of SARS in relation to their clinical presentation in human cases. It also looks at animal susceptibility after experimental infection, animal models of SARS, and the pathogenesis of this disease. It seems that adaptation of the virus within the host animal and the subsequent abnormal immune responses may be key factors in the pathogenesis of this new and fatal respiratory disease. The proteases produced in the lung during inflammation could also play an important role for exacerbation of SARS in animals.

Recognition signal for the C-terminal processing protease of D1 precursor protein in the photosystem II reaction center. An analysis using synthetic oligopeptides.

Synthetic oligopeptides of different chain lengths of 11 to 38 amino acids, corresponding to the carboxyl-terminal sequence of D1 precursor protein of the photosystem II reaction center, were subjected to a proteolytic cleavage by a processing enzyme isolated from spinach, in order to analyze the recognition signal. Practically the same Km and Vmax values were obtained for the oligopeptides consisting of more than 19 amino acids; a decrease in affinity, without affecting the Vmax value, was observed for the peptide consisting of 16 amino acids; no detectable activity was found for the peptide with 11 amino acids. When Asp-342 (12th residue from C-terminus) was replaced with Asn, for the peptide consisting of 16 amino acids, the enzymatic activity was completely abolished. In contrast, replacing Asp-342 with Glu had little effect. The efficiency of these oligopeptides as a substrate mentioned above, together with their effectiveness as an inhibitor, clearly demonstrated that the negative charge on Asp-342 plays a crucial role in the recognition, i.e., binding and cleavage, of the substrate by the processing enzyme, and suggested that the carboxyl-terminal extension consisting of 9 amino acids, by itself is not important in the binding.

Type A behavior is associated with an increased risk of left ventricular hypertrophy in male patients with essential hypertension.

OBJECTIVE: To determine whether type A behavior, which is associated with a risk of coronary heart disease, affects left ventricular hypertrophy in patients with essential hypertension. DESIGN: Cross-sectional study of 88 untreated patients with mild to moderate essential hypertension (33 men, mean +/- SEM age 54 +/- 1 years). METHODS: We measured the type A behavior score using a standardized questionnaire, left ventricular mass index using M-mode echocardiography and 24 h mean ambulatory blood pressure (recorded every 30 min). Beat-to-beat blood pressure was also measured using a Finapres device in patients at rest and during mental stress (counting backward) to determine the blood pressure response to stress. RESULTS: The left ventricular mass index was correlated with the type A behavior score (r = 0.214, P < 0.05), age (r = 0.266, P < 0.05), 24 h mean systolic and diastolic blood pressures (r = 0.391, P < 0.001, and r = 0.382, P < 0.001, respectively), systolic blood pressure both at rest and during stress (r = 0.255, P < 0.05, and r = 0.215, P < 0.05, respectively), and the variability of both systolic and diastolic blood pressures at rest (r = 0.253, P < 0.05, and r = 0.321, P < 0.01, respectively). Stepwise multiple linear regression analysis demonstrated that age was associated with an increase in the left ventricular mass index for both sexes (P = 0.004 for males, P = 0.003 for females). The type A behavior score predicted a greater increase in left ventricular mass index in men (P = 0.018) but not in women. The 24 h mean systolic blood pressure was associated with a greater increase in left ventricular mass index in women (P < 0.001) but not in men. CONCLUSION: Type A behavior is an independent risk factor for left ventricular hypertrophy in male patients with essential hypertension.

Psychobehavioral factors involved in the isolated office hypertension: comparison with stress-induced hypertension.

OBJECTIVE: To investigate the psychobehavioral factors involved in the isolated clinic blood pressure elevation and hypertension induced by mental stress. DESIGN AND METHODS: We studied 73 untreated patients with essential hypertension defined as World Health Organization stage I or II (28 men and 45 women, mean age 55 +/- 11 years). The amount of isolated clinic blood pressure elevation was examined in terms of the difference between clinic and daytime ambulatory blood pressures. Blood pressure (measured using a Finapres device) and R-R interval (measured electrocardiographically) were continuously monitored with subjects at rest and under mental stress (counting backward) to examine the cardiovascular response to the stress. Psychobehavioral characteristics such as anger, anxiety, tension, type A behavior pattern, and nervousness were evaluated and scored using structured interviews and self-reporting questionnaires. RESULTS: The anger score was inversely correlated to the clinic-ambulatory blood pressure difference for the systolic (r = -0.308, P < 0.01) and diastolic (r = -0.233, P < 0.05) blood pressures. The score for type A behavior pattern tended to be inversely correlated to the clinic-ambulatory blood pressure difference for diastolic blood pressure (r = -0.209, P < 0.1). The nervousness score was positively correlated to stress-induced increase in the systolic (r = 0.249, P < 0.05) and diastolic (r = 0.232, P < 0.05) blood pressures. The clinic-ambulatory blood pressure difference was not related to the blood pressure rise induced by mental stress (r = 0.170 for systolic blood pressure; r = 0.112 for diastolic blood pressure). CONCLUSION: The isolated clinic blood pressure elevation and hypertension due to mental stress were related to different psychobehavioral factors.

Neurovirulence of six different murine coronavirus JHMV variants for rats.

Six variant viruses of the JHMV strain of murine coronavirus with large (cl-2, CNSV, DL and DS) or small (sp-4 and JHM-X) S proteins were compared in terms of their relative neurovirulence in weanling Lewis rats. Inoculation of various doses of the variants revealed that the cl-2 and CNSV were highly virulent and DL and DS were low-virulent, while sp-4 and JHM-X were avirulent. Pathological examination of rats infected with variants cl-2, DL and sp-4 showed that the cl-2 and DL induced severe and mild acute encephalomyelitis, respectively, while no lesions were observed in the central nervous system of rats infected with sp-4. Virus growth and distribution of antigen in rat brains correlated strongly with neurovirulence. These results suggest that S protein plays a role in neurovirulence in rats. In addition, these variant viruses were shown to be useful tools for further analysis of JHMV neurovirulence in animals as well as in cultured cells.

Unique N-linked glycosylation of murine coronavirus MHV-2 membrane protein at the conserved O-linked glycosylation site.

The membrane (M) proteins of murine coronavirus (MHV) strains have been reported to contain only O-linked oligosaccharides. The predicted O-glycosylation site consisting of four amino acid residues of Ser-Ser-Thr-Thr is located immediately adjacent to the initiator Met and is well conserved among MHV strains investigated so far. We analyzed the nucleotide sequence of a highly virulent strain MHV-2 M-coding region and demonstrated that MHV-2 had a unique amino acid, Asn, at position 2 at the conserved O-glycosylation site. We also demonstrated that this substitution added N-linked glycans to MHV-2 M protein resulting in increment of molecular mass of MHV-2 M protein compared with JHM strain having only O-linked glycans.

Localization of major neutralizing epitopes on the S1 polypeptide of the murine coronavirus peplomer glycoprotein.

A recombinant baculovirus system has been used to express the amino terminal half of the murine coronavirus (JHMV) peplomer glycoprotein in insect cells. The expressed polypeptide is glycosylated and is recognized by a set of monoclonal antibodies (mAbs) specific for JHMV S protein. Three of these mAbs have a very high neutralizing activity for JHMV but not for other MHV strains. These results indicate that JHMV-specific, major neutralizing epitopes reside in the amino terminal S1 subunit of the peplomer glycoprotein.

Protective effects of murine recombinant interferon-beta administered by intravenous, intramuscular or subcutaneous route on mouse hepatitis virus infection.

The significance of the route for administration of murine recombinant interferon-beta (IFN-beta) for inducing its therapeutic effects has been studied. BALB/c mice were daily injected intravenously, intramuscularly or subcutaneously with 1.5x10(3), 1. 5x10(4), or 1.5x10(5) IU of IFN-beta, from one day before to 8th day after mouse hepatitis virus (MHV-2) challenge. All mice received IFN-beta survived significantly longer than those without IFN. In the liver of those IFN-treated mice, viral growth and the histopathological damages were extremely alleviated. These results suggest that, irrespective of the differences in the route of administration, IFN-beta markedly suppressed viral activity when its administration was started prior to viral infection. For clinical use, however, further studies are needed on the optimal route for administration if IFN-beta is given after viral infection.

Hemorrhagic cystitis after bone marrow transplantation: importance of a thin sectioning technique on urinary sediments for diagnosis.

We used a thin-sectioning technique for the electron microscopic detection of viral particles within the cells of urinary sediments in three recipients who developed hemorrhagic cystitis after allogeneic bone marrow transplantation. Results of viral cultures of urine and electron microscopic (EM) observations on urinary sediments were consistent in only one recipient. In this recipient, EM observations revealed many viral particles within the cells of urinary sediments with diameter of about 80 nm corresponding to adenovirus, of which type 11 was produced in viral cultures. In one of the other two recipients many viral particles with a mean diameter of 41.6 nm corresponding to papovavirus were observed, but viral cultures using conventional cells were negative. Re-cultures using HEK cells produced polyomavirus BK. EM observation was a clue to the correct diagnosis. In the remaining recipient, no viral particles were observed within the cells of urinary sediments, suggesting the hemorrhagic cystitis to be of non-viral origin, despite a positive result of viral culture. These results suggest that a thin-sectioning technique on the cells of urinary sediments is important for the differential diagnosis between a viral-induced and non-viral hemorrhagic cystitis.

Isolation of a subclonal cell line of PC12 transfected with dexamethasone-regulated ras oncogene: morphological differentiation, biochemical properties, and tumorigenicity.

We have isolated and characterized a new subclonal cell line designated as MR31, which was obtained by transfection of PC12 cells with a glucocorticoid-regulated ras oncogene. The mRNA derived from the c-Ha-ras gene was proved to be expressed on exposure of the MR31 cells to dexamethasone, the highest value being attained at 8 h. MR31 cells rapidly extended neurite-like processes within 24 h in response to dexamethasone as well as nerve growth factor (NGF). The time of onset of neurite outgrowth induced by dexamethasone corresponded to the time when the highest ras mRNA level was observed. The catecholamine content of MR31 cells was found to be twice that of PC12 cells. A time course study on the effects of dexamethasone or NGF on cells showed that the former caused an increase in dopamine, a major catecholamine, to twofold the control level at 48 h after the treatment, while the latter caused a decrease in the dopamine level. These effects on catecholamines were almost the same in MR31 and PC12 cells. The acetylcholinesterase activity of MR31 cells was enhanced by both dexamethasone and NGF, whereas that of PC12 cells was enhanced by NGF, but not by dexamethasone. The changes in acetylcholinesterase activity were correlated with neurite outgrowth. Electron-microscopically, MR31 cells were not different from PC12 cells. MR31 cells exhibited extremely decreased tumorigenicity as compared with PC12 cells. The morphological and biochemical properties of MR31 cells remained constant, even after repeated passages.(ABSTRACT TRUNCATED AT 250 WORDS)

Opsonisation of group B streptococci and restriction endonuclease digestion patterns of their chromosomal DNA.

Isolates of group B streptococci (GBS) from neonates with early-onset septicaemia are associated with particular restriction endonuclease digestion patterns (RDP types Ia-3 and III-3) of chromosomal DNA. Opsonophagocytosis of serotype Ia and serotype III GBS isolates was studied by the luminol-enhanced phagocytic chemiluminescence (CL) assay. Pools of serum containing GBS type-specific antibody levels equivalent to or just above levels typically found in sera from mothers of infected infants were used. CL intensities induced by GBS isolates of RDP types Ia-2, Ia-3 and III-3 were lower than those of the other RDP types of the same serotype. Opsonophagocytosis was more efficient with serum containing higher concentrations of type-specific antibodies but for RDP type III-3 strains these differences were much less marked than for other RDP types. CL intensity did not correlate with cell surface charge, hydrophobicity or sialic acid content of GBS. Results demonstrate that certain GBS RDP types are more resistant to opsonophagocytosis and suggest that potentially virulent strains with genetic homogeneity may exist.

Restriction endonuclease digest patterns of chromosomal DNA from group B beta-haemolytic streptococci.

Scanning densitometry and computer-assisted numerical analysis were used to examine restriction endonuclease digest patterns (RDPs) of chromosomal DNA from 26 infecting strains and 44 vaginal isolates of group B beta-haemolytic streptococci (GBS). At the 95% similarity level, HindIII RDPs of serotype Ia and III strains clustered into four and three RDP types, respectively. Nine of 10 strains from neonates with early-onset septicaemia belonged to two particular RDP types (Ia-3 and III-3). In contrast, serotype III GBS strains from meningitis cases were not characterised by particular RDP types. Associations between RDPs and certain phenotypic characteristics were also found.

Hypo-triploid acute myeloid leukemia with isochromosome 11q.

A 78-year-old male developed acute myeloid leukemia (AML, M2) with an isochromosome 11q (i(11q)) in hypo-triploid populations. He died 2 months later from the leukemia without having responded to chemotherapy. We reviewed 12 cases in the literature with i(11q) in hematopoietic neoplasms and found that the i(11q) is associated with complex chromosome abnormalities and with elderly patients. Ten out of the 13 patients had an AML phenotype, and they had poor response to chemotherapy and a short survival. The i(11q) may be a non-random chromosome aberration in hematopoietic neoplasms.

Deletion of chromosome 6q in two cases of acute myeloblastic leukemia and a review of the literature.

Two cases of acute myeloblastic leukemia (AML M2) associated with a deletion of chromosome 6q are described. One was a 38-year-old man with constitutional inversion of chromosome 9, and another was a 57-year-old female atomic-bomb survivor. The karyotype of these patients were 46,XY,del(6)(q12q14),inv(9)(p11q13), and 47,XX,6q-,+min, respectively. In both cases c-myb protooncogene, which is located in chromosome 6q, was neither deleted nor rearranged, and c-myb messenger RNA level was not elevated. These results suggest that c-myb is not involved in the leukemogenesis of AML with 6q- as well as lymphoid malignancies with 6q-. Out of 23 AML cases with 6q- reviewed, 6 cases had erythroleukemia, and 4 developed in Down syndrome patients.

Neutralizing antibody to herpes simplex virus by a novel neutralization method with peroxidase-labelled complement Clq.

A theoretically new neutralization method was developed for rapid and quantitative detection of virus neutralizing antibodies. The method involves the specific measurement of viral antigens produced by unneutralized virus by antiviral immune serum and peroxidase-labelled complement Clq (P*-Clq). After incubation of 37 degrees C for 18 h for amplification of herpes simplex virus (HSV) as a model, to detect unneutralized HSV, the amounts of HSV antigens produced in cells are measured by OD reading of enzymatic activities of P*-Clq bound to the HSV antigen-antibody complex formed after addition of anti-HSV probe serum and P*-Clq. Results showed that the OD reading of bound P*-Clq was proportional to the input m.o.i. of HSV and that neutralization of HSV with immune serum or human sera resulted in significant reduction of HSV-specific OD readings, depending on the antibody titers of serum samples used. The neutralizing antibody activity can be expressed as a percentage (NT%) of the quantity of neutralized virus, and the entire assay procedure can be completed within 24 h. This new method can replace the conventional neutralization test.

Patterns of cardiovascular responses to stress as a function of race and parental hypertension in men.

This study investigated cardiovascular responses to two stressors known to elicit either beta-adrenergic (mental arithmetic) or alpha-adrenergic (forehead cold pressor) reactivity in Black and White men. Participants in each group were selected for presence or absence of parental hypertension. Based on previous research, Blacks were expected to show smaller cardiovascular responses to the beta-adrenergic mental arithmetic task and greater responses to the alpha-adrenergic cold pressor relative to the Whites. Systolic blood pressure (SBP), diastolic blood pressure (DBP), heart rate, forearm blood flow, and forearm vascular resistance were assessed during a resting baseline, a prestress period, and during and after each experimental procedure. Unlike previous findings, no significant racial differences in cardiovascular responses were found during either task. However, Black participants had significantly higher SBP and DBP levels throughout the cold pressor periods. Parental history did not significantly influence cardiovascular responses in either group. The results are discussed in relation to previous research on racial differences in stress reactivity and their implications for future research.

Analysis of bile acids in colon residual liquid or fecal material in patients with colorectal neoplasia and control subjects.

Bile acids are believed to play a role in the etiology of colorectal cancer. To examine the relationship between bile acids and colorectal neoplasia, bile acids in colon residual liquid or fecal material were analyzed in 18 patients with colorectal adenoma, 12 patients with colorectal cancer, and 18 healthy control subjects. High-performance liquid chromatography combined with immobilized 3 alpha-hydroxysteroid dehydrogenase in column form showed a significant elevation in the proportion of deoxycholic acid (P < 0.05), lithocholic acid (P < 0.05), secondary bile acids (deoxycholic acid plus lithocholic acid) (P < 0.02), and the chenodeoxycholic acid-lithocholic acid family (chenodeoxycholic acid plus lithocholic acid) (P < 0.05) in the colon residual liquid or fecal material of the patients with colorectal adenoma compared with proportions in the control subjects. A similar trend was noted in the patients with colorectal cancer compared to the control subjects. These findings suggested that an increase in the proportion of secondary bile acids, in particular, of lithocholic acid, was closely related to the pathogenesis of colorectal neoplasia.

Clinical significance of serum iron and ferritin in patients with colorectal cancer.

To clarify the significance of serum iron and ferritin as indicators of iron loss caused by continuous bleeding, and, thus, to determine their value as markers of colorectal cancer, values for the two were compared in male patients with early and advanced colorectal cancer and age-matched male controls. The mean value of serum iron levels in patients with advanced colorectal cancer was significantly decreased compared with values in patients with early colorectal cancer and controls, 50.5 +/- 38.6 micrograms/dl vs 93.0 +/- 32.1 micrograms/dl and 107.1 +/- 32.9 micrograms/dl, respectively (p < 0.001). The mean value of serum ferritin levels in patients with early and advanced colorectal cancer was also significantly decreased compared with controls, 80.5 +/- 35.0 ng/ml (p < 0.01) and 48.8 +/- 72.8 ng/ml (p < 0.001), respectively, vs 117.1 +/- 46.8 ng/ml. However, there was no significant difference between mean serum iron levels in patients with early colorectal cancer and controls. Eighteen (78.3%) of the 23 patients with advanced colorectal cancer and 3 (16.7%) of the 18 patients with early colorectal cancer had serum iron levels below 85 micrograms/dl and serum ferritin levels below 60 ng/ml. Levels of both serum iron and ferritin, without clinically evident anemia, are useful indicators of advanced colorectal cancer.