Valuable antiviral therapeutic options for the treatment of chronic hepatitis C patients with thrombocytopenia.

Thrombocytopenia in patients with chronic hepatitis C may represent an obstacle for the initiation of antiviral treatment. The aim of this study was to evaluate factors predictive of successful pegylated interferon (PEG-IFN) α2b and ribavirin (RBV) treatment for patients with thrombocytopenia with no history of splenectomy or partial splenic embolization. One hundred and fifty-one chronic hepatitis C patients (genotype 1: n = 110, genotype 2: n = 41) with TCP (<100 × 10(9) /L) at baseline were enrolled. Pretreatment variables included interleukin 28B (IL28B) genotype (rs8099917) and homoeostasis model assessment of insulin resistance score (HOMA-IR). The kinetics of haemoglobin and platelets according to the inosine triphosphatase (ITPA) genotype (rs1127354) were investigated. Sustained virological response (SVR) was significantly more frequent in hepatitis C virus (HCV) genotype 2 (65.9%) than in genotype 1 (34.5%) patients (P < 0.0001). Multiple logistic regression analysis of HCV genotype 1 extracted IL28B TT genotype [odds ratio (OR) 5.97, P = 0.006] and HOMA-IR <2.5 (OR 7.14, P = 0.0016) as significant independent pretreatment predictors of SVR. The analyses of HCV genotype 2 showed that HOMA-IR was significantly related to SVR, but IL28B genotype was not. Patients with ITPA CC genotype showed a significant haemoglobin reduction and lower degree of platelets decrease than those with ITPA CA/AA genotypes. The most common reason for premature discontinuation of treatment was the development of hepatocellular carcinoma (n = 8, 5.3%). In conclusion, HOMA-IR is a useful predictor of SVR for patients with thrombocytopenia infected with HCV genotype 1 or 2 treated with PEG-IFNα2b and RBV. The inclusion of IL28B, ITPA genotypes and HOMA-IR adds valuable therapeutic information.

Evaluation of the effects of freeze-dried soybean curd intake on cholesterol levels using a novel biomarker.

To evaluate the effects of freeze-dried soybean curd intake on serum cholesterol levels, we examined the subclasses of cholesterol for healthy adult volunteers who continued to eat a piece of freeze-dried soybean curd each day along with their ordinary diet for four weeks. Of 12 subjects, the soybean curd diet proved effective in 2 cases; small dense LDL (sd-LDL) cholesterol levels were significantly reduced in association with the decrease in LDL cholesterol levels. These results suggested that daily intake of freeze-dried soybean curd may lead to an improvement in cholesterol metabolism in subjects with originally higher cholesterol levels and that sd-LDL cholesterol can be a novel biomarker for evaluation of the cholesterol lowering-action of functional food.

Protective effect of sivelestat sodium (Eraspol) on postoperative lung dysfunction in patients with type A acute aortic dissection: a pilot study.

AIM: The authors evaluated the protective effect of sivelestat sodium on postoperative lung dysfunction in patients with type A acute aortic dissection who underwent aortic arch surgery with cardiopulmonary bypass (CPB) under deep hypothermia with circulatory arrest (DHCA). METHODS: Twelve patients with type A acute aortic dissection who underwent aortic arch replacement under CPB with DHCA and were pretreated with or without sivelestat sodium (sivelestat group, N.=7 patients; control group, N.=5 patients) were observed. The ratio of arterial oxygen tension to inspired oxygen fraction (P/F ratio) was measured as a parameter of pulmonary function before and after operation. The number of white blood cells was also counted as an index of inflammatory reaction before and after the operation. RESULTS: The P/F ratio decreased significantly after operation in the control group. However, the P/F ratio was unchanged between before and after operation in the sivelestat group. The number of white blood cells tended to increase after operation in the control group, whereas it decreased significantly after operation in the sivelestat group. CONCLUSION: The present study demonstrated the protective effect of sivelestat sodium on postoperative lung injury in patients with acute type A aortic dissection undergoing aortic arch surgery under CPB with DHCA.

The effects of a herbal medicine (Mao-to) in patients with chronic hepatitis C after injection of IFN-beta.

We found that a herbal medicine (Mao-to) relieves the side effects of interferon (IFN)-beta and the combination therapy improves the biochemical response rate. However, the exact mechanism by which Mao-to is effective remains to be established. We conducted a controlled trial to clarify the effects of Mao-to. The study was carried out in 18 patients with chronic hepatitis C, and we examined subjective symptoms, body temperature and cytokines such as interleukin (IL)-beta, IL-1receptor antagonist (ra), IL-6 and TNF-alpha. Each patient received 6 million units of IFN-beta intravenously. Mao-to was given orally just before, just after, and 1 hour after IFN administration. The control study was carried out 6 months after the combination therapy of Mao-to and IFN-beta. The scores for general malaise, arthralgia and discomfort were significantly lower in the combination group than in control group. Body temperature did not significantly differ between the two groups. Plasma IL-6 level and IL-1ra were significantly elevated in the combination group compared to control (P = 0.0057 and 0.0003, respectively). Mao-to did not affect plasma concentrations of IL-1beta and TNF-alpha. We considered the increment of IL-1ra caused by Mao-to is to be one of the key factors involved in reducing the flu-like symptoms accompanying IFN-beta and improving the biochemical response rate.

The efficacy of a herbal medicine (Mao-to) in combination with intravenous natural interferon-beta for patients with chronic hepatitis C, genotype 1b and high viral load: a pilot study.

Patients with chronic hepatitis C, with a high serum viral load (> or = 1 Meq/ml) and genotype 1b seem to be resistant to interferon (IFN) therapy. To evaluate the efficacy of a herbal medicine (Mao-to) in combination with natural IFN-beta for the treatment of these patients, eighteen Japanese patients were enrolled in this study. Every patient received 6 million units (MU) of IFN-beta intravenously daily for 8 weeks. Mao-to was given orally 3-4 times a day during the IFN-beta administration, Sixteen of the 18 patients (89%) became negative for serum HCV RNA at the end of treatment, but only 2 of them (11%) remained negative for the virus RNA at 6 months of follow-up. Serum ALT levels normalized in 17 patients (94%) at 2 weeks of follow-up after the cessation of therapy, and 11 patients (61%) retained normal ALT levels for more than 6 months of follow-up. This rate of biochemical response was high as compared with that of therapy with IFN-beta alone (19%) in the largest IFN-beta trial in Japan. Serum hyaluronic acid levels were decreased significantly from 147.0 +/- 110.5 ng/ml to 77.4 +/- 67.4 ng/ml in the sustained biochemical response group (P = 0.003). None of the patients needed to interrupt therapy because of side effects of IFN-beta. Thus, Mao-to administration together with IFN-beta treatment could increase the sustained biochemical response rate, and reduce liver fibrosis.

A comparative study between excess-dose users and regular-dose users of rhubarb contained in Kampo medicines.

With prolonged use of rhubarb-containing Kampo medicines, some patients come to ask for high-dose rhubarb because of deteriorated reactivity to rhubarb. We divided patients into two groups in terms of rhubarb-dose, and compared clinical backgrounds between regular-dose group and excess-dose group. Patients who were treated with rhubarb-containing Kampo extracts (manufactured prescriptions) or Kampo formulae (decoctions) for more than 12 months were enrolled. These two groups were compared for age, sex, shape of stool, abdominal symptoms, existence of hemorrhoids, Kampo diagnosis of abdomen, past stimulant laxative use, duration of stimulant laxative use before the first administration of rhubarb, duration of rhubarb use in our hospital, and initial existence of stimulant pain caused by taking stimulant laxatives for the first time. No significant difference was shown between the two groups in terms of age, duration of stimulant laxative-use before the first prescription of rhubarb, shape of stool, abdominal symptoms, existence of hemorrhoids, or duration of rhubarb-use. However, most patients in the regular-dose group had initial stimulant pain of the abdomen upon taking stimulant laxatives for the first time, but most patients in the excess-dose group did not (p < 0.001). All patients except one in the regular dose group had the sign of "umbilical region tenderness on pressure", but half of the excess-dose group did not have it (p = 0.041). Based on these findings, the absence of "initial stimulant pain" and the absence of "umbilical region tenderness on pressure" may predict increasing or excess use of rhubarb, and long-term use of rhubarb should be discouraged more strongly in the patients without these signs.

Depletion and deletion analyses of eucaryotic translation initiation factor 1A in Saccharomyces cerevisiae.

Translation initiation factor eIF1A is a highly conserved, small, acidic protein that is required for cell growth in yeast. Biochemical studies in vitro implicate eIF1A in dissociating ribosomes, promoting methionyl-tRNA(i) binding to 40S ribosomal subunits, scanning of mRNAs and recognizing the AUG initiation codon. To elucidate the pleiotropic functions of eIF1A in vivo, the factor was depleted by placing its gene behind the repressible GAL1 promoter. After Saccharomyces cerevisiae cells were shifted to glucose medium, depletion of eIF1A was seen after 3-4 generations, corresponding with cessation of cell growth. Polysome profiles of the depleted strain showed ribosome run-off from mRNAs, indicating that eIF1A is involved in the initiation phase of translation. A decrease in free 40S ribosomes and an apparent increase in free 60S ribosomes were attributed to the formation of 40S subunit dimers. The result suggests that one of the functions of eIF1A is to prevent formation of 40S dimers. Mutant forms of eIF1A lacking either the positively charged N-terminal region or the negatively charged C-terminal region were constructed and tested for their ability to confer cell growth as the sole source of eIF1A. Either deletion supports cell growth, albeit at a slower rate, and causes a reduction in polysomes, although eIF1A lacking the N-terminal region is more deleterious. Therefore the charged terminal regions contribute to, but are not absolutely essential for, eIF1A function.

Overexpression of HUT1 gene stimulates in vivo galactosylation by enhancing UDP-galactose transport activity in Saccharomyces cerevisiae.

Transfer of activated sugar-nucleotides from the cytoplasm to the lumen of the Golgi is an essential requirement for glycosylation of glycoproteins, proteoglycans and glycosphingolipids. Although mannosylation is the major modification in the yeast Saccharomyces cerevisiae, several reports suggest the presence of galactose residues on yeast proteins and sphingolipids. We have detected alpha-galactosylated O-linked chitinase by lectin blotting from cells that functionally express the gma12(+) gene, encoding alpha 1,2-galactosyltransferase from Schizosaccharomyces pombe. This result implies the presence of a UDP-galactose transporter in S. cerevisiae. A conserved gene, HUT1, which encodes a putative multi-transmembrane protein, was cloned and characterized for its possible involvement in galactosylation. The HUT1 gene is not essential and is expressed at a relatively low level under the physiological conditions we examined. The disruption of this gene did not show any apparent impairments in glycosylation. However, a temperature- and concentration-dependent increase in UDP--galactose transport activity was detected from cells overexpressing HUT1 in the presence of gma12(+). The surface of these cells was confirmed to carry galactose residues by staining with FITC-conjugated alpha-galactose-specific lectin. These results suggest a role for Hut1p in the transport of UDP--galactose from the cytosol into the Golgi lumen in S. cerevisiae.

Coexpression of alpha1,2 galactosyltransferase and UDP-galactose transporter efficiently galactosylates N- and O-glycans in Saccharomyces cerevisiae.

We have studied in vivo neo-galactosylation in Saccharomyces cerevisiae and analyzed the critical factors involved in this system. Two heterologous genes, gma12(+) encoding alpha1, 2-galactosyltransferase (alpha1,2 GalT) from Schizosaccharomyces pombe and UGT2 encoding UDP-galactose (UDP-Gal) transporter from human, were functionally expressed to examine the intracellular conditions required for galactosylation. Detection by fluorescence labeled alpha-galactose specific lectin revealed that 50% of the cells incorporated galactose to cell surface mannoproteins only when the gma12(+) and hUGT2 genes were coexpressed in galactose media. Integration of both genes in the Delta mnn1 background cells increased galactosylation to 80% of the cells. Correlation between cell surface galactosylation and UDP-galactose transport activity indicated that an exogenous supply of UDP-Gal transporter rather than alpha1,2 GalT played a key role for efficient galactosylation in S.cerevisiae. In addition, this heterologous system enabled us to study the in vivo function of S. pombe alpha1,2 GalT to prove that it transfers galactose to both N - and O -linked oligosaccharides. Structural analysis indicated that this enzyme transfers galactose to O -mannosyl residue attached to polypeptides and produces Galalpha1,2-Man1-O-Ser/Thr structure. Thus, we have successfully generated a system for efficient galactose incorporation which is originally absent in S. cerevisiae, suggesting further possibilities for in vivo glycan remodeling toward therapeutically useful galactose containing heterologous proteins in S. cerevisiae.

The 39-kilodalton subunit of eukaryotic translation initiation factor 3 is essential for the complex's integrity and for cell viability in Saccharomyces cerevisiae.

Eukaryotic translation initiation factor 3 (eIF3) in the yeast Saccharomyces cerevisiae comprises about eight polypeptides and plays a central role in the binding of methionyl-tRNAi and mRNA to the 40S ribosomal subunit. The fourth largest subunit, eIF3-p39, was gel purified, and a 12-amino-acid tryptic peptide was sequenced, enabling the cloning of the TIF34 gene. TIF34 encodes a 38,753-Da protein that corresponds to eIF3-p39 in size and antigenicity. Disruption of TIF34 is lethal, and depletion of eIF3-p39 by glucose repression of TIF34 expressed from a GAL promoter results in cessation of cell growth. As eIF3-p39 levels fall, polysomes become smaller, indicating a role for eIF3-p39 in the initiation phase of protein synthesis. Unexpectedly, depletion results in degradation of all of the subunit proteins of eIF3 at a rate much faster than the normal turnover rates of these proteins. eIF3-p39 has 46% sequence identity with the p36 subunit of human eIF3. Both proteins are members of the WD-repeat family of proteins, possessing five to seven repeat elements. Taken together, the results indicate that eIF3-p39 plays an important, although not necessarily direct, role in the initiation phase of protein synthesis and suggest that it may be required for the assembly and maintenance of the eIF3 complex in eukaryotic cells.

Continuous urine oxygen tension monitoring in patients undergoing cardiac surgery.

OBJECTIVES: To evaluate the effect of cardiopulmonary bypass (CPB) on urine oxygen tension (PuO2) and to determine whether perioperative PuO2 can predict postoperative renal dysfunction in patients undergoing cardiac surgery. DESIGN: Prospective clinical study. SETTING: A university research laboratory, a university-affiliated hospital. PARTICIPANTS: Ninety-eight consecutive adult patients undergoing coronary artery bypass surgery or valvular surgery. INTERVENTIONS: PuO2 was continuously measured by inserting a polarographic electrode into the urinary tube connected to a Foley catheter. MEASUREMENTS AND MAIN RESULTS: PuO2 was constant before CPB and then progressively decreased after the start of CPB. It partially recovered at weaning from CPB but did not completely return to its original level until the end of surgery. Postoperative serum creatinine concentrations were significantly higher in patients whose PuO2 decreased after CPB, as compared with those whose PuO2 was constant or increased. The amplitude and the rate of recovery in PuO2 after CPB were significantly associated with peak values of postoperative serum creatinine concentrations. CONCLUSIONS: These results suggest the possibility of PuO2 detecting an early stage of renal dysfunction in cardiac surgery, although further studies will be required to substantiate it.

Characterization of yeast translation initiation factor 1A and cloning of its essential gene.

Translation initiation factor eIF1A is required in vitro for maximal rates of protein synthesis in mammalian systems. It functions primarily by dissociating ribosomes and stabilizing 40 S preinitiation complexes. To better elucidate its precise role in promoting the translation initiation process, the yeast form of eIF1A has been identified in Saccharomyces cerevisiae and purified to homogeneity on the basis of its cross-reaction with antibodies prepared against mammalian eIF1A. The apparent mass of yeast eIF1A (22 kDa) resembles that of the mammalian homolog (20 kDa), and the yeast factor is active in stimulating methionyl-puromycin synthesis in an assay composed of mammalian components. The gene encoding yeast eIF1A, named TIF11, was cloned and shown to be single copy. TIF11 encodes a protein comprising 153 amino acids (17.4 kDa); the deduced amino acid sequence exhibits 65% identity with the sequence of human eIF1A. Both human and yeast eIF1A contain clusters of positive residues at the N terminus and negative residues at the C terminus. Deletion/disruption of TIF11 demonstrates that eIF1A is essential for cell growth. Expression of human eIF1A cDNA rescues the growth defect of TIF11-disrupted cells, indicating that the structure/function of yeast and mammalian eIF1A is highly conserved.