Detection of pre-C and core region mutants of hepatitis B virus in chronic hepatitis B virus carriers.

We analyzed the pre-C and core region of hepatitis B virus (HBV) DNA by a polymerase chain reaction in 22 chronic carriers. In 9 hepatitis B e antigen-positive asymptomatic carriers, a single DNA band was detected at the expected size, whereas additional shorter DNA bands were observed in 7 out of 11 patients with chronic hepatitis. The smaller-sized DNAs from one chronic hepatitis patient had various lengths of deletions spanning from 105 to 183 bp in the middle of the core gene, and all deletions included common nucleotide sequences. All of the smaller-sized DNAs from the other patients proved to be variant core genes. They were deleted in similar regions by Southern analysis using oligonucleotide probes. A follow-up study revealed that four out of seven chronic hepatitis patients with a short core gene seroconverted to antibody to hepatitis B e antigen, but those with only a "wild type" did not. In another set of sequence studies, clones isolated from two chronic carriers displayed heterogeneity of the pre-C and core gene which was more often present in sera with normal alanine aminotransferase levels than with abnormal levels. These results suggest that mutant HBV alters the host immune response, and may modulate the clinical course of HBV infection. An alternative possibility is that chronic hepatitis selects for mutant forms.     

Pain and Bradykinin Receptors--sensory transduction mechanism in the nociceptor terminals and expression change of bradykinin receptors in inflamed condition]

Bradykinin (BK), an endogenous algesic and sensitizing substance, excited nociceptors and sensitized their heat responses. These effects were mediated by B2 receptors (B2Rs) in normal condition, and B1 receptors were additionally recruited in inflammation. B2Rs were coupled with Gq/11 and their activation resulted in diacylglycerol and inositol triphosphate release. Diacylglycerol activated protein kinase (PK) Cepsilon in sensory neurons. To clarify what channel was modulated by PKC to depolarize nociceptor terminals, we examined the heat activation threshold (Tt) of heat-sensitive capsaicin receptor (TRPV1). Tt was lowered down to 31 degrees C by BK in concentration dependent manner through activation of PKCepsilon in cells heterologously expressing TRPV1 and B2Rs. Thus both excitation and sensitization to heat could be explained by one mechanism, lowering Tt of TRPV1. The same was observed in capsaicin-sensitive primary sensory neurons. However, TRPV1 knockout mice showed almost no change in BK-induced nociceptive behavior and nociceptor excitation, although BK-induced heat hyperalgesia completely disappeared, suggesting that TRPV1 was not the sole channel that was modulated by BK to depolarize nociceptor terminals. In addition nociceptor sensitivity to BK was augmented in inflamed animals, with B2R mRNA and protein upregulated. The mechanism for prostaglandin-induced augmentation of BK response is left open for future study.     

Identification of CCND3 and BYSL as candidate targets for the 6p21 amplification in diffuse large B-cell lymphoma.

PURPOSE: Increases in gene dosage through DNA amplification represents a common feature of many tumors and can result in the up-regulation of tumor-promoting genes. Our recent genome-wide, array-based comparative genomic hybridization analysis of 66 cases of diffuse large B-cell lymphoma found that genomic gain of 6p21 was observed in as many as 17 cases, including 14 cases with low-level copy number gain and three cases with high-level copy number gains (amplifications). EXPERIMENTAL DESIGN AND RESULTS: To identify the target gene(s) for 6p21 amplification, we constructed a detailed amplicon map at the region of genomic amplification with the aid of high-resolution contig array-based comparative genomic hybridization glass slides, consisting of contiguously ordered bacterial artificial chromosome/P1-derived artificial chromosome clones covering 3 Mb throughout the 6p21 amplification region. Alignment of the amplifications identified a minimally overlapping 800 kb segment containing 15 genes. Quantitative expression analysis of the genes from both patient samples and the SUDHL9 cell line revealed that CCND3 and BYSL (1.9 kb telomeric to the CCND3 gene locus) are the targets of 6p21 genomic gain/amplification. CONCLUSIONS: Although it is known that t(6;14)(p21;q32) induces aberrant overexpression of CCND3 in B-cell malignancies, we were able to show that CCND3, which encodes the cyclin D family member protein that controls the G1-S phase of cell cycle regulation, can also be a target of genomic gain/amplification. Overexpression of CCND3 through genomic amplification is likely to lead to aberrant cell cycle control, although the precise biological role of BYSL with respect to tumorigenesis remains to be determined.     

A dose-finding study of glycosylated G-CSF (Lenograstim) combined with CHOP therapy for stem cell mobilization in patients with non-Hodgkin's lymphoma

BACKGROUND: Peripheral blood stem cell (PBSC) reinfusion has been widely used for hematopoietic reconstitution after high-dose chemotherapy. However, the optimal dose of granulocyte colony-stimulating factor (G-CSF) for PBSC mobilization in combination with chemotherapy for autograft remains unknown. METHODS: To find the optimal dose of glycosylated G-CSF (lenograstim) for PBSC mobilization in combination with chemotherapy for aggressive non-Hodgkin's lymphoma (NHL), we conducted a dose-finding study on 43 newly diagnosed patients who had unfavorable prognostic factors. They received four to six courses of cyclophosphamide, doxorubicin, vincristine and prednisolone combined with lenograstim every 2 weeks (biweekly CHOP therapy). PBSC apheresis was started after the third course of biweekly CHOP therapy. Lenograstim was given daily from day 3 until the day of the last apheresis. The optimum dose of lenograstim was assessed based on mobilization efficacy and safety profiles at a daily single dose of 2, 5 and 10 microg/kg for eight patients in each level. RESULTS: The collected number of CD34+ cells in the first apheresis products was higher in the 5 microg/kg group than in the 2 microg/kg group (median, 4.22 x 10(6) vs 2.49 x 10(6) CD34+ cells/kg, P = 0.051). The highest dose of 10 microg/kg (median, 2.99 x 10(6) CD34+ cells/kg) failed to show a dose dependence in PBSC mobilization. The efficacy and safety of the 5 microg/kg dose were further confirmed in an additional 19 patients. CONCLUSIONS: The present study suggests that the recommended dose of lenograstim for PBSC mobilization with CHOP therapy in untreated NHL is 5 microg/kg.     

Pitavastatin, an HMG-CoA reductase inhibitor, exerts eNOS-independent protective actions against angiotensin II induced cardiovascular remodeling and renal insufficiency

Angiotensin II (Ang II) plays a pivotal role in cardiovascular remodeling leading to hypertension, myocardial infarction, and stroke. Pitavastatin, an HMG-CoA reductase inihibitor, is known to have pleiotropic actions against the development of cardiovascular remodeling. The objectives of this study were to clarify the beneficial effects as well as the mechanism of action of pitavastatin against Ang II-induced organ damage. C57BL6/J mice at 10 weeks of age were infused with Ang II for 2 weeks and were simultaneously administered pitavastatin or a vehicle. Pitavastatin treatment improved Ang II-induced left ventricular hypertrophy and diastolic dysfunction and attenuated enhancement of cardiac fibrosis, cardiomyocyte hypertrophy, coronary perivascular fibrosis, and medial thickening. Ang II-induced oxidative stress, cardiac TGFbeta-1 expression, and Smad 2/3 phosphorylation were all attenuated by pitavastatin treatment. Pitavastatin also reduced Ang II-induced cardiac remodeling and diastolic dysfunction in eNOS-/- mice as in wild-type mice. In eNOS-/- mice, the Ang II-induced cardiac oxidative stress and TGF-beta-Smad 2/3 signaling pathway were enhanced, and pitavastatin treatment attenuated the enhanced oxidative stress and the signaling pathway. Moreover, pitavastatin treatment reduced the high mortality rate and improved renal insufficiency in Ang II-treated eNOS-/- mice, with suppression of glomerular oxidative stress and TGF-beta-Smad 2/3 signaling pathway. In conclusion, pitavastatin exerts eNOS-independent protective actions against Ang II-induced cardiovascular remodeling and renal insufficiency through inhibition of the TGF-beta-Smad 2/3 signaling pathway by suppression of oxidative stress.     

Correlation between myocardial uptake of technetium-99m-sestamibi and pressure-derived myocardial fractional flow reserve

OBJECTIVES: Development of the coronary pressure wire has facilitated the measurement of fractional flow reserve (FFR) to assess the functional severity of coronary artery stenoses. METHODS: This study evaluated the correlations between FFR and myocardial direct counts of technetium-99m(99mTc)-sestamibi in 20 patients (16 men, 4 women, mean age 66 +/- 8 years) who underwent 99mTc-sestamibi single-photon emission computed tomography (SPECT) with the 2-day protocol using 740 MBq of 99mTc-sestamibi each day. Visual assessment of myocardial imaging and quantitative analysis with the measurement of percent uptake and direct count of 99mTc-sestamibi were performed. RESULTS: Visual assessment of myocardial imaging revealed that reversibility of 99mTc-sestamibi perfusion defects was correlated with FFR of < 0.75, which is regarded as functionally important stenosis (17/20 vs 3/20, kappa = 0.71, p < 0.002). Regional reversibility score did not correlate with FFR (r = -0.40, p = NS). Quantitative analysis revealed that the change in 99mTc-sestamibi percent uptake with pharmacologic stress using adenosine triphosphate disodium (ATP) also did not correlate with FFR (r = 0.35, p = NS). In contrast, percent increase in 99mTc direct counts with ATP was lower in patients with FFR of < 0.75 than in those with FFR of > = 0.75 (-4 +/- 16% vs 24 +/- 30%, p < 0.01). In addition, a significant correlation (r = 0.70, p < 0.001) was observed between percent increase in 99mTc direct counts with ATP and FFR. CONCLUSIONS: These results suggest that quantitative analysis of 99mTc-sestamibi scintigraphy enables the assessment of the magnitude of functional significance of coronary stenosis.     

Comparative analysis of clinical outcomes after allogeneic bone marrow transplantation versus peripheral blood stem cell transplantation from a related donor in Japanese patients

A reduced incidence of graft versus host disease (GvHD) has been documented among Japanese allogeneic bone marrow transplantation (BMT) patients, as the Japanese are genetically more homogeneous than western populations. To clarify whether this ethnic difference affects the results of allogeneic peripheral blood stem cell transplantation (PBSCT), we conducted a nationwide survey to compare clinical outcomes of allogeneic PBSCT (n = 214) and BMT (n = 295) from a human leucocyte antigen-identical-related donor in Japanese patients. The cumulative incidence of grades II-IV acute GvHD was 37.4% for PBSCT and 32.0% for BMT. The cumulative incidence of extensive chronic GvHD at 1 year was significantly higher after PBSCT than BMT (42% vs. 27%; P < 0.01). The organ involvement patterns of GvHD were different between the two groups. By multivariate analyses, the incidence of chronic GvHD was significantly increased in PBSCT, whereas the stem cell source did not affect the incidence of acute GvHD, transplant-related mortality, relapse or survival. We concluded that Japanese PBSCT patients have an increased risk of chronic GvHD compared with BMT patients, but the incidence of acute GvHD was still lower than in western populations. Thus, the choice of haematopoietic stem cell source should be considered based on data for individual ethnic populations.