Hepatitis C virus infection: a risk factor for Parkinson's disease.

Recent studies found that hepatitis C virus (HCV) may invade the central nervous system, and both HCV and Parkinson's disease (PD) have in common the overexpression of inflammatory biomarkers. We analysed data from a community-based integrated screening programme based on a total of 62,276 subjects. We used logistic regression models to investigate association between HCV infection and PD. The neurotoxicity of HCV was evaluated in the midbrain neuron-glia coculture system in rats. The cytokine/chemokine array was performed to measure the differences of amounts of cytokines released from midbrain in the presence and absence of HCV. The crude odds ratios (ORs) for having PD were 0.62 [95% confidence interval (CI), 0.48-0.81] and 1.91 (95% CI, 1.48-2.47) for hepatitis B virus (HBV) and HCV. After controlling for potential confounders, the association between HCV and PD remained statistically significant (adjusted OR = 1.39; 95% CI, 1.07-1.80), but not significantly different between HBV and PD. The HCV induced 60% dopaminergic neuron death in the midbrain neuron-glia coculture system in rats, similar to that of 1-methyl-4-phenylpyridinium (MPP(+) ) but not caused by HBV. This link was further supported by the finding that HCV infection may release the inflammatory cytokines, which may play a role in the pathogenesis of PD. In conclusion, our study demonstrated a significantly positive epidemiological association between HCV infection and PD and corroborated the dopaminergic toxicity of HCV similar to that of MPP(+) .

Risk of new-onset type II diabetes after appendicectomy.

BACKGROUND: Altered immune function after appendicectomy has been associated with autoimmune disease, even though the mechanisms are not clearly elucidated. This study aimed to investigate whether the frequency of new-onset type II diabetes was increased after appendicectomy in a case-control study. METHODS: This was a retrospective cohort study from the Taiwan Longitudinal Health Insurance Database 2000. The relative risk was compared with that in the general population using population-based data. Each patient was tracked for a 3-year interval to identify those who developed type II diabetes. Cox proportional hazard regression analysis was used to assess the risk of type II diabetes during follow-up. RESULTS: A total of 31,512 patients were included in the study, of whom 5252 had an appendicectomy (study cohort) and 26,260 were matched for comparison. Some 714 patients (2.3 per cent) developed type II diabetes during the 3-year follow-up, 161 in the study cohort (3.1 per cent) and 553 in the comparison cohort (2.1 per cent). The adjusted hazard ratio (HR) for type II diabetes in the study cohort was 1.45 (95 per cent c.i. 1.22 to 1.74). This increased risk was most pronounced in men (adjusted HR 1.47, 1.16 to 1.88) and in those with a perforated appendix (adjusted HR 2.28, 1.71 to 3.03), and applied only to patients younger than 65 years of age. CONCLUSION: An increased risk of new-onset type II diabetes within 3 years after appendicectomy was found in patients aged less than 65 years. The risk was highest in men and in those with complicated appendicitis.

Development of a cerebral microvascular dysplasia model in rodents.

Normal vasculature development of the central nervous system is extremely important because patients with vascular malformations are at life-threatening risk for intracranial hemorrhage or cerebral ischemia. The etiology and pathogenesis of abnormal vasculature development in the central nervous system are unknown, and progress is hampered by the lack of animal models for human cerebrovascular diseases. Here, we report our current study on cerebral microvascular dysplasia (CMVD) development. Using vascular endothelial growth factor hyper-stimulation, we demonstrated that aberrant microvessels could be developed in the rodent brain under certain conditions (such as genetic deficient background, local cytokine and chemokine release, or exogenous vessel dilating stimulation) that may speed up focal angiogenesis and lead to cerebral vascular dysplasia.

Changes in ploidy distributions in human liver carcinogenesis.

Cellular and nuclear DNA content was measured by flow cytometry and the fraction of binucleated cells by fluorescence microscopy in normal adult human livers, hepatocellular carcinomas, cirrhotic livers surrounding tumors, and in some benign liver conditions. In five normal livers about one-half of the hepatocytes were polyploid; the majority of these were binucleated tetraploids containing two diploid nuclei. Thus, polyploidization in human liver does not progress as far as, for example, in the rat, where 80%-90% of adult hepatocytes are polyploid, mostly with tetraploid or octoploid nuclei. In five human euploid hepatocellular carcinomas and one investigated case of focal nodular hyperplasia, the percentage of polyploid cells was significantly reduced. Four other carcinomas exhibited a prominent aneuploid (hypotetraploid) peak in addition to the diploid peak. An abnormally low fraction of binucleated cells was also indicated in these tumors. Liver tissue surrounding the tumors had a ploidy distribution similar to that of normal liver. The results suggest that, like in several models of experimental hepatocarcinogenesis, human hepatocellular tumor growth is associated with a decreased polyploidization tendency and a corresponding increase in diploid, divisional growth, which may give the tumors a growth advantage relative to the surrounding liver.

Telomerase activity and telomere length in human hepatocellular carcinoma.

Telomerase activity is activated and telomere length altered in various types of cancers, including hepatocellular carcinoma (HCC). A total of 39 HCC tissues and the corresponding non-tumour livers were analysed and correlated with clinical parameters. Telomere length was determined by terminal restriction fragment assay, and telomerase activity was assayed by telomeric repeat amplification protocol. Telomerase activity was positive in 24 of the 39 tumour tissues (1.15-285.13 total product generated (TPG) units) and in six of the 39 non-tumour liver tissues (1.05-1.73 TPG units). In the 28 cases analysed for telomere length, telomere length was shortened in 11 cases, lengthened in six cases, and unaltered in 11 cases compared with non-tumour tissues. Neither telomere length nor telomerase activity was correlated to any clinical parameters.

Interferon-alpha reduces insulin resistance and beta-cell secretion in responders among patients with chronic hepatitis B and C.

This study aimed at elucidating the effects of interferon (IFN)-alpha on glucose metabolism in patients with chronic hepatitis B and C infections. Twenty-eight biopsy-proven patients with chronic hepatitis B (ten cases) and hepatitis C (18 cases) were given IFN-alpha for a total of 24 weeks. The patients received a 75 g oral glucose tolerance test (OGTT), glucagon stimulation test, tests for type 1 diabetes-related autoantibodies and an insulin suppression test before and after IFN-alpha therapy. Ten of the 28 patients responded to IFN-alpha therapy. Steady-state plasma glucose of the insulin suppression test decreased significantly in responders (13.32+/-1.48 (S.E.M.) vs 11.33+/-1.19 mmol/l, P=0.0501) but not in non-responders (12.29+/-1.24 vs 11.11+/-0.99 mmol/l, P=0.2110) immediately after completion of IFN-alpha treatment. In the oral glucose tolerance test, no significant difference was observed in plasma glucose in either responders (10.17+/-0.23 vs 10.03+/-0.22 mmol/l) or non-responders (10.11+/-0.22 vs 9.97+/-0.21 mmol/l) 3 Months after completion of IFN-alpha treatment. However, significant differences were noted in C-peptide in both responders (2.90+/-0.13 vs 2.20+/-0.09 nmol/l, P=0.0040) and non-responders (2.45+/-0.11 vs 2.22+/-0.08 nmol/l, P=0.0287) before vs after treatment. The changes of C-peptide in an OGTT between responders and non-responders were also significantly different (P=0.0028), with responders reporting a greater reduction in C-peptide. No case developed autoantibodies during the treatment. In patients who were successfully treated with IFN-alpha, insulin sensitivity improved and their plasma glucose stayed at the same level without secreting as much insulin from islet beta-cells.

Quantitative detection of hepatitis B virus DNA in human sera by branched-DNA signal amplification.

Serum samples from 116 patients with hepatitis B surface antigen (HBsAg), from 7 patients without detectable HBsAg and from 71 healthy blood donors were tested by a branched DNA signal amplification (bDNA) method. Hepatitis B virus (HBV) DNA was detected in 39 (34%) of the 116 samples with HBsAg, including 19 (70%) of the 27 patients who were also positive for hepatitis B e antigen (HBeAg). In contrast, one of the 7 patients without HBsAg and none of the 71 blood donors were positive for HBV DNA. The titers of serum HBV DNA did not correlate with the serum alanine aminotransferase levels. All the samples positive by the bDNA assay were positive by the polymerase chain reaction (PCR). However, 59% of the PCR-positive samples were bDNA-negative. None of the PCR-negative samples was positive by the bDNA method. Although the sensitivity of bDNA method is not entirely satisfactory, it showed excellent specificity and reproducibility. Thus it may be considered as an alternative for quantitative detection of HBV DNA in serum samples of patients with relatively high titers of HBV viremia.

Oncopolitical issues: obstacles and options for success in a comprehensive breast center.

Over the last 20 years of breast center development, it has been found that for every obstacle there is an option for resolution. The solutions are not uniform in each setting, but rather they are a reflection of the medical staff politics, interests, and strengths, the resources and commitment of the institution, and the needs of the community. The most important message of this article is to validate local medical staff issues as legitimate concerns. The crucial obstacle is the lack of cooperation among the medical specialties and the lack of trust between the medical staff and management. The obstacles need to be appropriately addressed by the creation of an effective organizational structure that is successful in the local environment and by the programmatic development that reflects the medical staff's interests and strengths. This will ensure comprehensive breast center success in the local political environment and ultimately translates into enhanced expertise, pride, and higher quality patient care.

Intratumor injection of absolute ethanol under ultrasound guidance for the treatment of small hepatocellular carcinoma.

For the treatment of small hepatocellular carcinoma, intratumor injection of absolute ethanol under ultrasound guidance was performed in 27 tumors in 23 patients, with a tumor diameter of between 1.0 and 3.3 cm. The initially elevated serum alpha-fetoprotein levels in 15 patients decreased during treatment, with 13 returning to normal after this regimen. In the 6 patients who finally received surgical resection, 4 had complete necrosis of the tumor, while the other 2 had a small peripheral residual cancer nest. In the remaining non-resected 17 cases, follow-up CT, multiple biopsies and angiography revealed evidence of viable tumor in only 3 cases. After additional ethanol injections, these 3 cases were successfully treated. Inhomogeneous distribution of the injected ethanol and difficulty in identifying the tumor after previous injections accounted for the incomplete necrosis of the tumor. To cope with these problems, a steel coil was implanted in the tumor before treatment, and a needle with multiple side holes was used in the last 3 cases, with satisfactory results. Ethanol injection is promising and may even be curative in the treatment of small hepatocellular carcinoma.

Tissue hepatocyte growth factor and proliferating cell nuclear antigen in hepatocellular carcinoma.

To better understand the roles of hepatocyte growth factor (HGF) and proliferating cell nuclear antigen (PCNA) in hepatocellular carcinoma (HCC), 37 surgically resected HCCs and corresponding nontumorous liver tissue specimens were collected and the expression of these two factors was quantified by Western blot analysis. Both HGF and PCNA expression levels were significantly higher in tumor tissue than in nontumorous liver tissue. However, their expression levels in HCC tissue and nontumorous tissue did not show any significant correlation with the recurrence of HCC. In addition, HGF and PCNA did not correlate with Edmondson's grade, invasiveness of tumor, presence of tumor capsule, or tumor size. No correlation was found between the expression levels of HGF and PCNA in HCC tissue. We conclude that, although both HGF and PCNA are present at higher levels in HCC tissue than in nontumorous liver tissue, they play little role in the clinicopathologic manifestations of this tumor. HGF appears to contribute little, if at all, to the proliferative activity of HCC cells.

Development of a uniform system for mortality review.

Four Veterans Administration Medical Centers in Medical District 14 of the Great Lakes region collaborated in the development and implementation of a uniform, education-oriented mortality review system. Because the new review format emphasizes evaluation of care already delivered as well as planning for treatment in similar situations, the process and product of such reviews is expected to be very useful. Data derived from these mortality reviews may also be used for trending both within and between medical centers.

Cancer Program Administrators' survey results.

In summary, cancer programs across the country have been developing since 1975. A more formal organizational structure has been adopted to program activities since 1980. The full-time Cancer Program Administrator position exists in about one-third of the programs reporting. Persons occupying the position are at least bachelors degreed with many also having a masters degree. Strengths, weaknesses, and obstacles to program development are basically the same items across the country. It appears that programs that have one problem resolved are still working towards the solution of another problem and vice versa among programs. All programs are struggling for identity within the hospital and are in developmental stages with a lot of activity focused on clarifying the position of an organized cancer program within the hospital organization.

Hepatitis delta antigen expressed by recombinant baculoviruses: comparison of biochemical properties and post-translational modifications between the large and small forms.

Hepatitis delta virus (HDV) encodes only one protein, the hepatitis delta antigen (HDAg). Two forms of HDAg, a large (27 kDa) and a small (24 kDa) one, participate in the various steps of HDV replication. To further understand the properties of HDAg, we have constructed recombinant baculoviruses and expressed both forms of the HDAg in insect cells. The gene encoding HDAg was placed under the control of the polyhedrin promoter of Autographa Californica nuclear polyhedrosis virus (AcNPV) by homologous recombination. When Spodoptera frugiperda (Sf9) cells were infected with the recombinant viruses, both the small HDAg and the large HDAg were expressed at high levels. The HDAgs produced by the recombinants were similar in size and antigenic properties to those of the proteins produced in mammalian hepatoma cell lines. It was also localized exclusively in the nuclei. In addition, both proteins bound to HDV RNA in an in vitro assay. No difference in the RNA-binding affinity was noted between the two forms of HDAg, suggesting that the trans-dominant inhibitory activity of the large HDAg on HDV replication is not due to its competition with the small HDAg for RNA binding. Two RNA-protein complexes could be detected, suggesting either that there are at least two binding sites on the HDV RNA or that HDAg binds to HDV RNA in two multimeric forms. We have further shown that both the large and the small HDAgs are phosphoproteins, with the former having an approximately sixfold higher level of phosphorylation. Finally, it was demonstrated that the large HDAg was isoprenylated, while the small one was not. These differences in post-translational modifications are the first differences in biochemical properties demonstrated between the two forms and may explain the differential effects of the large and small HDAgs on HDV RNA replication and virus packaging.

Large hepatitis delta antigen in packaging and replication inhibition: role of the carboxyl-terminal 19 amino acids and amino-terminal sequences.

Hepatitis delta virus (HDV) encodes two proteins, the small delta antigen (SHDAg) and large delta antigen (LHDAg). The latter is identical to the former except for the presence of additional 19 amino acids at the C terminus. While SHDAg is required for HDV replication, LHDAg inhibits replication and, together with hepatitis B surface antigen (HBsAg), is required for the assembly of HDV. The last 19 C-terminal amino acids of LHDAg are essential for HDV assembly. Most of LHDAg (amino acids 19 to 146 and 163 to 195) had been shown to be dispensable for packaging with HBsAg. To discern whether the last 19 C-terminal amino acids solely constitute the signal for packaging with HBsAg, we constructed two LHDAg deletion mutants and tested their abilities to be packaged with HBsAg in cotransfection experiments. We found that deletion of amino acids 2 to 21 and 142 to 165 did not affect LHDAg packaging. This result suggested that only the last 19 C-terminal amino acids of LHDAg are required for packaging. We further constructed two plasmids which expressed c-H-ras with or without additional 19 C-terminal amino acids identical to those in LHDAg. Only c-H-ras with additional 19 amino acids could be cosecreted with HBsAg in the cotransfection experiment. This result confirmed that the C-terminal 19 amino acids are the packaging signal for HBsAg. We also tested the trans activation activity and trans-dominant inhibitory activity of the deletion mutants of SHDAg and LHDAg, respectively. In contrast to deletion of amino acids 142 to 165, deletion of amino acids 2 to 21 impaired the trans-dominant inhibitory activity of LHDAg. Deletion of amino acids 2 to 21 and 142 to 165 did not affect the trans activation activity of SHDAg. This result suggested that a functional domain which is important for the trans-dominant inhibitory activity of LHDAg exists in the amino terminus of HDAg.

RNA-binding activity of hepatitis delta antigen involves two arginine-rich motifs and is required for hepatitis delta virus RNA replication.

Hepatitis delta antigen (HDAg) is an RNA-binding protein with binding specificity for hepatitis delta virus (HDV) RNA (J. H. Lin, M. F. Chang, S. C. Baker, S. Govindarajan, and M. M. C. Lai, J. Virol. 64:4051-4058, 1990). By amino acid sequence homology search, we have identified within its RNA-binding domain two stretches of an arginine-rich motif (ARM), which is present in many prokaryotic and eukaryotic RNA-binding proteins. The first one is KERQDHRRRKA and the second is EDEKRERRIAG, and they are separated by 29 amino acids. Deletion of either one of these ARM sequences resulted in the total loss of the in vitro RNA-binding activity of HDAg. Thus, HDAg is different from other RNA-binding proteins in that it requires two ARM-like sequences for its RNA-binding activity. Replacement of the spacer sequence between the two ARMs with a shorter stretch of sequence also reduced RNA binding in vitro. Furthermore, site-specific mutations of the basic amino acid residues in both ARMs resulted in the total loss or reduction of RNA-binding activity. The biological significance of the RNA-binding activity was studied by examining the trans-activating activity of the RNA-binding mutants. The plasmids expressing HDAgs with various mutations in the RNA-binding motifs were cotransfected with a replication-defective HDV dimer cDNA construct into COS cells. It was found that all the HDAg mutants which had lost the in vitro RNA-binding activity also lost the ability to complement the defect of HDV RNA replication. We conclude that the trans-activating function of HDAg requires its binding to HDV RNA.

Isoprenylation of large hepatitis delta antigen is necessary but not sufficient for hepatitis delta virus assembly.

Hepatitis delta virus (HDV) encodes two proteins, the small hepatitis delta antigen (SHDAg) and large hepatitis delta antigen (LHDAg). Both proteins are identical except for the presence of additional 19 amino acids at the C terminus of LHDAg. While SHDAg is required for HDV RNA replication, LHDAg inhibits replication and is required together with hepatitis B surface antigen for the assembly of HDV. The C-terminal last 4 amino acids of LHDAg (Cys-Arg-Pro-Gln) is an isoprenylation motif. It has previously been shown that the mutation of the Cys inhibited the assembly of HDV. In order to discern whether this effect is due to change of amino acid residue or abolition of isoprenylation, we constructed several LHDAg mutants of the terminal three amino acid residues and tested their abilities to be packaged with HBsAg by cotransfection experiments. We also made GST-fusion proteins of these mutants and tested their abilities to be isoprenylated in rabbit reticulocyte lysate system. We found that some, but not all, of the substitutions of the amino acid residues other than the Cys also inhibited isoprenylation and that the status of isoprenylation of these mutant proteins correlated well with their abilities to be packaged with HBsAg into virions. This result indicates that isoprenylation, rather than the primary amino acid sequence, is required for LHDAg packaging. Furthermore, we found that the attachment of an isoprenylation motif to SHDAg did not enable it to be packaged with HBsAg and that the deletions of any 5 amino acids in the last 15 amino acids (amino acids 196 to 210) unique to the LHDAg abolished the packaging ability. In contrast, the deletion of 33 amino acids (amino acids 163 to 195) upstream of the last C-terminal 19 amino acids of LHDAg did not interfere with its packaging ability. Therefore, we conclude that the 15 amino acids upstream of the isoprenylation site of LHDAg are also essential for HDV assembly, and a large portion of the alleged C-terminal Pro/Gly-rich region (amino acids 146 to 195) is not required for the assembly process.

Effect of in utero and postnatal exposure to environmental tobacco smoke on the developmental expression of pulmonary cytochrome P450 monooxygenases.

Pulmonary cytochrome P450 monooxygenases metabolize xenobiotic chemicals, including those found in environmental tobacco smoke (ETS). Exposure to ETS beginning at birth has been shown to induce the P450 CYP1A1 by seven days of life. The effects of perinatal exposure to ETS of the rat lung on the expression of CYP1A1, 1B1, 2B1, and NADPH cytochrome P450 reductase were measured using semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Timed pregnant dams and their pups were exposed to aged and diluted sidestream cigarette smoke (ADSS) as a surrogate for ETS for four hours/ day from gestational day 5 through postnatal day 21. For all genes analyzed, mRNA could be detected in the fetal lung beginning at gestational day 17 but were not altered by ADSS. In contrast, intraperitoneal injection of dams with beta-naphthoflavone significantly elevated both CYP1A1 and 1B1 at gestational day 21, indicating that these genes are inducible. Continued exposure to ADSS significantly induced CYP1A1 but not other P450 genes as early as one day after birth.. We conclude that (1) ADSS induces pulmonary CYP1A1 in the first day of life; (2) fetal cytochrome P450 genes are not induced by maternal exposure to ADSS; and (3) in the fetal lung, CYP1A1 and 1B1 can be induced by beta-naphthoflavone.

Acid-induced gene expression in Helicobacter pylori: study in genomic scale by microarray.

To understand the RNA expression in response to acid stress of Helicobacter pylori in genomic scale, a microarray membrane containing 1,534 open reading frames (ORFs) from strain 26695 was used. Total RNAs of H. pylori under growth conditions of pH 7.2 and 5.5 were extracted, reverse transcribed into cDNA, and labeled with biotin. Each microarray membrane was hybridized with cDNA probe from the same strain under two different pH conditions and developed by a catalyzed reporter deposition method. Gene expression of all ORFs was measured by densitometry. Among the 1,534 ORFs, 53 ORFs were highly expressed (> or = 30% of rRNA control in densitometry ratios). There were 445 ORFs which were stably expressed (<30% of rRNA in densitometry) under both pH conditions without significant variation. A total of 80 ORFs had significantly increased expression levels at low pH, while expressions of 4 ORFs were suppressed under acidic condition. The remaining 952 ORFs were not detectable under either pH condition. These data were highly reproducible and comparable to those obtained by the RNA slot blot method. Our results suggest that microarray can be used in monitoring prokaryotic gene expression in genomic scale.

Effect of hepatitis C antibody screening in blood donors on post-transfusion hepatitis in Taiwan.

A national screening programme for antibody to hepatitis C virus (HCV) in blood donors in Taiwan began in July 1992 using a second-generation immunoassay. To study the impact of this screening on post-transfusion hepatitis in Taiwan, a prospective study on post-transfusion hepatitis, that was started in 1987, was continued. As of June 1994, 245 patients who received a blood transfusion after July 1992 had completed a follow-up period for more than 6 months post-transfusion. Of them, seven (2.8%) recipients developed acute post-transfusion hepatitis. The hepatitis in six cases could not be attributed to infection by hepatitis A, B, C, D, E viruses or cytomegalovirus (CMV) or Epstein-Barr virus (EBV). The remaining patient seroconverted to both IgG and IgM anti-CMV. All seven patients recovered in 6 months without development of chronicity, and the mean peak alanine aminotransferase level was lower compared with that of the cases before anti-HCV screening (i.e. pre-July 1992). These results indicate that the current anti-HCV screening has effectively interrupted HCV transmission through blood transfusion in Taiwan.

Packaging of hepatitis delta virus RNA via the RNA-binding domain of hepatitis delta antigens: different roles for the small and large delta antigens.

Hepatitis delta virus (HDV) is composed of four specific components. The first component is envelope protein which contains hepatitis B surface antigens. The second and third components are nucleocapsid proteins, referred to as small and large hepatitis delta antigens (HDAgs). The final component is a single-stranded circular RNA molecule known as the viral genome. In order to study the mechanism of HDV RNA packaging, a four-plasmid cotransfection system in which each viral component was provided by a separate plasmid was employed. Virus-like particles released from Huh-7 cells receiving such a cotransfection were found to contain HDV RNA along with three proteins. Therefore, the four-plasmid cotransfection system could lead to successful HDV RNA packaging in vitro. The system was then used to show that the large HDAg alone was able to achieve a low level of HDV RNA packaging. Analysis of a variety of large HDAg mutants revealed that the RNA-binding domain was essential for viral RNA packaging. By increasing the incorporation of small HDAg into virus-like particles, we found a three- to fourfold enhancement of HDV RNA packaging. This effect was probably through a direct binding of HDV RNA, independent from that of large HDAg, with the small HDAg. The subsequent RNA-protein complex was packaged into particles. The results provided insight into the roles and functional domains of small and large HDAgs in HDV RNA packaging.