Characterization of integrated hepatitis B virus DNA harboring pre-S mutations in hepatocellular carcinoma patients with ground glass hepatocytes.

Ground glass hepatocytes (GGHs) have been associated with hepatocellular carcinoma (HCC) recurrence and poor prognosis. We previously demonstrated that pre-S expression in some GGHs is resistant to current hepatitis B virus (HBV) antiviral therapies. This study aimed to investigate whether integrated HBV DNA (iDNA) is the primary HBV DNA species responsible for sustained pre-S expression in GGH after effective antiviral therapy. We characterized 10 sets of micro-dissected, formalin-fixed-paraffin-embedded, and frozen GGH, HCC, and adjacent hepatitis B surface antigen-negative stained tissues for iDNA, pre-S deletions, and the quantity of covalently closed circular DNA. Eight patients had detectable pre-S deletions, and nine had detectable iDNA. Interestingly, eight patients had integrations within the TERT and CCNE1 genes, which are known recurrent integration sites associated with HCC. Furthermore, we observed a recurrent integration in the ABCC13 gene. Additionally, we identified variations in the type and quantity of pre-S deletions within individual sets of tissues by junction-specific PacBio long-read sequencing. The data from long-read sequencing indicate that some pre-S deletions were acquired following the integration events. Our findings demonstrate that iDNA exists in GGH and can be responsible for sustained pre-S expression in GGH after effective antiviral therapy.

Detection of Hepatitis B Virus-Host Junction Sequences in Urine of Infected Patients.

Integrated hepatitis B virus (HBV) DNA, found in more than 85% of HBV-associated hepatocellular carcinomas (HBV-HCCs), can play a significant role in HBV-related liver disease progression. HBV-host junction sequences (HBV-JSs), created through integration events, have been used to determine HBV-HCC clonality. Here, we investigate the feasibility of analyzing HBV integration in a noninvasive urine liquid biopsy. Using an HBV-targeted next-generation sequencing (NGS) assay, we first identified HBV-JSs in eight HBV-HCC tissues and designed short-amplicon junction-specific polymerase chain reaction assays to detect HBV-JSs in matched urine. We detected and validated tissue-derived junctions in five of eight matched urine samples. Next, we screened 32 urine samples collected from 25 patients infected with HBV (5 with hepatitis, 10 with cirrhosis, 4 with HCC, and 6 post-HCC). Encouragingly, all 32 urine samples contained HBV-JSs detectable by HBV-targeted NGS. Of the 712 total HBV-JSs detected in urine, 351 were in gene-coding regions, 11 of which, including TERT (telomerase reverse transcriptase), had previously been reported as recurrent integration sites in HCC tissue and were found only in the urine patients with cirrhosis or HCC. The integration breakpoints of HBV DNA detected in urine were found predominantly (~70%) at a previously identified integration hotspot, HBV DR1-2 (down-regulator of transcription 1-2). Conclusion: HBV viral-host junction DNA can be detected in urine of patients infected with HBV. This study demonstrates the potential for a noninvasive urine liquid biopsy of integrated HBV DNA to monitor patients infected with HBV for HBV-associated liver diseases and the efficacy of antiviral therapy.

Characterization of the hepatitis B virus DNA detected in urine of chronic hepatitis B patients.

BACKGROUND: Detection of human hepatitis B virus (HBV) DNA in the urine of patients with chronic hepatitis B infection (CHB) has been reported previously, suggesting urine could provide a potential route of horizontal HBV transmission. However, it is not clear whether the HBV DNA detected in urine is indeed full-length, infectious viral DNA. The aim of this study is to assess the potential infectivity of urine from patients with CHB and to correlate HBV DNA detection in urine with clinical parameters, such as serum viral load and HBeAg status. METHODS: Urine from 60 CHB patients with serum viral loads ranging from undetectable to 108 IU/mL were analyzed for HBV DNA and serum immune markers. HBV DNA was detected from total urine DNA and size-fractionated urine DNA (separated into ≤1 kb and > 1 kb fractions) by PCR analysis of six regions of the HBV genome. RESULTS: Twenty-seven of 59 (45.7%) patients with HBV serum viral load (≥20 IU/mL) contained at least 20 copies per mL of fragmented HBV DNA in urine detected in at least 1 of the 6 PCR assay regions. Only one patient contained HBV DNA detected by all six regions, and was found to have evidence of blood in the urine. Sixteen of 25 urine samples with high viral load (> 105 IU/mL) and 11 of 34 urine samples with low viral load (< 105 IU/mL) contained detectable HBV DNA. Twelve of 27 (44.44%) patients with detectable HBV DNA in urine were HBeAg positive, and only 5 of these HBeAg positive patients were in the group of 33 (15.15%) patients with no detectable HBV DNA in urine. By Fishers' exact test, HBV DNA in urine is significantly associated with high serum viral load (P = 0.0197) and HBeAg (P = 0.0203). CONCLUSIONS: We conclude that urine from CHB patients with healthy kidney function should not contain full-length HBV DNA, and therefore should not be infectious.

Risk factors for acquiring extended-spectrum ß-lactamase-producing Enterobacteriaceae in geriatric patients with multiple comorbidities in respiratory care wards.

AIM: Extended-spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae is associated with a high mortality rate and increased medical care costs. Elderly patients might receive mechanical ventilation with respiratory treatment for the long term in respiratory care wards (RCW). This retrospective case-control study sought to determine the risk factors for geriatric patients who acquire a urinary tract infection with ESBL-producing Escherichia coli or Klebsiella pneumoniae in this type of hospital. METHODS: Two RCW participated in this study from September 2006 to March 2007. Patients suspected of having a UTI were enrolled in this study. Urine samples were collected for culture. The medical records and demographic data of patients, including days of hospitalization, comorbidities and duration of invasive instruments, were recorded. UTI was diagnosed by physicians. ESBL-producing isolates were detected using the phenotypic confirmatory tests according to the Clinical and Laboratory Standard Institute standards. RESULTS: There were 109 patients having 240 sets of clinical data and laboratory specimens. The prevalence of ESBL-producing isolates of E. coli. and K. pneumoniae were 39.5% and 69.7%, respectively. Patients with multiple underlying comorbidities (OR=2.88, P<0.05) or receiving more than two antimicrobial agents (OR=3.71, P<0.05) were at an increased risk for acquiring the ESBL-producing microorganisms after adjustment for sex, age and days of hospitalization. CONCLUSIONS: Geriatric patients with recent exposure to two or more antibiotics and two or more numbers of comorbidities were at risk for ESBL-producing organism infection. Our results suggest that infection control procedures in RCW should be concerned with reducing antimicrobial prescriptions and patient comorbidities.

The misplacement of external ventricular drain by freehand method in emergent neurosurgery.

External ventricular drain (EVD) placement is one of the most basic and common neurosurgical procedure which most was performed by young neurosurgical trainees. This study is conducted to determinate the safe and accuracy of EVD placement by freehand method. About 129 EVD placements were evaluated in this study. Eighty-three catheters (64.3%) were located in the ipsilateral frontal horn or third ventricle. The functional accuracy was 86%. Of eighteen misplaced catheters, only 4 (3.1%) catheters were nonfunctional, requiring a replacement or reposition. The higher misplaced rate was significantly observed in patients whose head CT scans revealed the lower hydrocephalus ratio (28.85%) and the smaller ventricular size (5.6 mm). Twenty-one (16.2%) new hemorrhages associated with EVD placements were observed. Using the freehand method, EVD placement is a safe and effective procedure in management of these emergent neurosurgical diseases.

Blastocystis hominis infection in long-term care facilities in Taiwan: prevalence and associated clinical factors.

Blastocystis hominis is probably the most common protozoan found in the human gut worldwide. In Taiwan, the prevalence of B. hominis infection is yet to be determined but is expected to be relatively higher among foreign workers. No data is available on the prevalence of B. hominis infection in long-term care facilities in Taiwan. This study included 713 subjects (552 residents and 161 care workers) from ten long-term care facilities in Taiwan who completed stool microscopic examinations with Merthiolate-iodine-formalin stain technique. The prevalence rate of blastocystosis was the highest among foreign and domestic care workers followed by residents (12.2%, 4.6%, and 2.7%, respectively). Older age (p = 0.04) and lower educational level (p = 0.008) were significantly associated with blastocystosis among care workers. Among residents, B. hominis infection was negatively associated with prolonged use of antibiotics within 3 months prior to examination (p = 0.05) and positively associated with tracheostomy in-place (p = 0.028). In conclusion, B. hominis infection was the most prevalent intestinal parasitic infection among both care workers and residents of long-term care facilities in Taiwan. Use of antibiotics was negatively associated with B. hominis infection among residents. Additionally, appropriate preventive measures should be implemented to older care workers with lesser educational attainment in order to reduce the risk of blastocystosis infection.

Association between C-reactive protein and biomarkers of bone and mineral metabolism in chronic hemodialysis patients: a cross-sectional study.

OBJECTIVE: Both chronic inflammation and dysregulation of bone and mineral metabolism are closely related with long-term outcomes of dialysis patients. Our objective was to investigate the relationship between these two abnormalities. DESIGN: This was a cross-sectional study. SETTING: This study was performed at a hospital-based hemodialysis center. PATIENTS: We enrolled 448 (male, 198; female, 250) clinically stable hemodialysis patients. Patients with chronic inflammatory disease, malignancy, or viral hepatitis were excluded. Their age (mean +/- SD) was 57.4 +/- 12.5 years. MAIN OUTCOME MEASURES: Biomarkers, including high-sensitivity C-reactive protein (hsCRP), total calcium, phosphate, and intact parathyroid hormone levels, were measured and compared with the recommended range in the K/DOQI guidelines. Correlations between these parameters were analyzed, and factors independently associated with hsCRP and the calcium phosphate product (Ca x P) were identified by regression analysis. RESULTS: Most patients did not achieve the K/DOQI recommended therapeutic range in the four parameters, and only 50 patients (11%) met their treatment goals. The hsCRP level was directly related to calcium, phosphate, and Ca x P. Patients who achieved the guidelines' range had lower hsCRP levels (1.97 mg/L vs. 2.71 mg/L, P < .05). A high hsCRP level (> or = 10 mg/L) was associated with higher calcium, phosphate, and Ca x P levels, and lower albumin levels. Serum albumin, Ca x P, alkaline phosphatase, and diabetes independently predicted hsCRP levels. CONCLUSION: There is a strong association between chronic inflammation and the disturbance of bone mineral metabolism in chronic hemodialysis patients.

MYC pathway is activated in clear cell renal cell carcinoma and essential for proliferation of clear cell renal cell carcinoma cells.

Clear cell renal cell carcinoma (ccRCC) is the major and aggressive subtype of RCC. Previously, we identified 383 differentially expressed genes by analyzing full-length cDNA libraries of ccRCC and normal kidney tissues. In this study, we applied functional network analysis to the differentially expressed genes for identifying deregulated molecular pathways in ccRCC, and the results indicated that MYC showed a prominent role in the highest scoring network. The upregulation of MYC expression was validated in ccRCC tissues and cell lines. Furthermore, Knockdown of MYC expression by MYC-specific siRNA significantly inhibited the abilities of uncontrolled proliferation, anchorage-independent growth and arrested cell cycle in the G0/G1 phase in ccRCC cells. Moreover, we found that 37 differentially expressed genes were shown to be MYC-target genes, and the upregulation of the MYC-target genes BCL2, CCND1, PCNA, PGK1, and VEGFA were demonstrated. The expression of these MYC-target genes was significantly correlated with the expression of MYC in ccRCC tissues, and knockdown of MYC also suppressed the expression of these MYC-target genes in ccRCC cells. The recruitment of MYC to the promoter regions of BCL2, CCND1, PCNA, PGK1, and VEGFA was shown by Chromatin immunoprecipitation assay. These results suggest that MYC pathway is activated and plays an essential role in the proliferation of ccRCC cells.

Complete fracture-dislocation of the thoracolumbar spine without paraplegia.

Complete fracture-dislocation of thoracolumbar spine without significant neurologic deficits is a rare entity. The possible mechanism may result from the spontaneous decompressive fractures of the spinal elements with sparing of the spinal cord. Here, we report a case of a healthy 50-year-old man who presented with complete fracture-dislocation of T12 through L1 vertebrae without paraplegia. Early diagnosis and early surgery with decompression, reduction, and stabilization is the best policy in management of this unstable spine injury.

Inflammatory marker but not adipokine predicts mortality among long-term hemodialysis patients.

AIMS: chronic inflammation contributes significantly to the morbidity and mortality of chronic hemodialysis patients. A recent research has shown that adipokines were associated with inflammation in these patients. We aim to investigate whether biomarkers of inflammation, adipokines, and clinical features can predict the outcome of hemodialysis patients. MATERIALS AND METHODS: we enrolled 181 hemodialysis patients (men: 97, mean age: 56.3+/-13.6) and analyzed predictors of long-term outcomes. RESULTS: during the 3-year followup period, 41 patients died; the main causes of death were infection and cardiovascular disease. Elevated serum levels of hsCRP and albumin and advanced age were highly associated with death (all P<.001). Leptin and adiponectin levels were not significantly different between deceased patients and survivors. Cox-regression analysis indicated that age, diabetes, albumin level, and hsCRP were independent factors predicting mortality. CONCLUSION: the presence of underlying disease, advanced age, and markers of chronic inflammation is strongly related to survival rate in long-term hemodialysis patients.

Multiple intracranial hemorrhages after evacuation of bilateral subdural effusions.

Postoperative intracerebral hemorrhages occurring after evacuation of subdural fluid collections have been infrequently reported and remain a devastating complication. The pathophysiological mechanism is still unclear. Disturbed autoregulation and restoration of normal cerebral flow seems to play an important role in this type of event because of rapid decompression. Herein, we present a case of multiple intracerebral hemorrhages in the putamen and cerebellar hemisphere following evacuation of bilateral subdural effusions, and review the relevant literature.

Gene expression analysis of human hepatocellular carcinoma by using full-length cDNA library.

Hepatocellular carcinoma (HCC) is a leading cause of death worldwide. Hepatitis B virus (HBV) or hepatitis C virus (HCV) infection has been shown to cause hepatic carcinogenesis. A total 58,251 of cDNA clones of full-length cDNA libraries of HBV and HCV-infected HCC and their surrounding non-tumor tissues, respectively, were sequenced and analyzed by blasting against GENEBANK maintained by NCBI. About 180 and 279 of genes were shown an obviously increased and decreased expression patterns between HCC tissue and its adjacent non-tumor tissue. The candidate genes consisted of the genes encoded liver specific metabolism enzymes, secretory functional proteins, proteases and their inhibitors, protein chaperon, cell cycle components, apoptosis-related proteins, transcriptional factors, and DNA binding proteins. Several genes were further investigated by using real-time PCR to confirm the gene expression levels in at least 24 pairs of HCC tissues and adjacent non-tumor tissues. The results showed that genes encoded reticulon 4, RGS-1, antiplasmin, and kallikrein B were down-regulated with the average of 2.8, 8.5, 3.2, and 10.5-fold, respectively. Our results provide crucial candidate genes to develop clinical diagnosis and gene therapy of HCC.

Identification of differentially expressed genes in clear cell renal cell carcinoma by analysis of full-length enriched cDNA library.

Renal cell carcinoma (RCC) is the most common malignancy in adult kidney, and accounts for 3% of malignancies worldwide with increasing incidence. Clear cell RCC (ccRCC) is the major type in RCC. Resection by surgery is the main treatment because the response of ccRCC to traditional therapies is very poor. To identify the tumor-associated genes for better understanding the molecular mechanism of ccRCC, the full-length enriched cDNA libraries of ccRCC and normal kidney tissues were constructed by the oligo-capping method. Nucleotide sequences of the cDNA libraries of ccRCC and normal kidney tissues were sequenced. From the sequence analysis of 19,425 and 12,400 clones of ccRCC and normal kidney tissues, 4356 and 3055 genes were identified, respectively. By comparing the gene-expression patterns of ccRCC and normal tissues, the up- or down-regulated genes were identified. Among these identified genes, the differential expression of annexin A2 and argininosuccinate synthetase genes were further confirmed by quantitative real-time PCR and Western blot analysis.

Additive effect of smoking and cotton dust exposure on respiratory symptoms and pulmonary function of cotton textile workers.

One hundred and sixty-nine and 175 cotton textile workers (CTWs) were enrolled in the first (1991) and second (1996) surveys to investigate the prevalence of byssinosis. The synergistic effect of smoking on cotton dust exposure was also evaluated. Although the difference in prevalence of abnormal pulmonary function between the first (38.5%) and second study (38.9%) was not statistically significant, smokers had significantly higher frequency than nonsmokers in both surveys. A significant trend existed between the cotton dust levels and the frequency of abnormal lung function. The significant trend was also noted in both smokers and nonsmokers. The frequency of respiratory symptoms and the prevalence of severe byssinosis in the second survey (14.9% and 12.6%, respectively) were significantly lower than that in the first survey (39.7% and 21.9%, respectively). The reduction of symptoms was due to remodeling of this old cotton mill. The prevalences of respiratory symptoms and byssinosis in smokers being significantly higher than in nonsmokers only found in the first survey, but not found in the second survey. These results indicate that smoking potentiates the effect of cotton dust exposure on respiratory symptoms and byssinosis. The second study reveals high prevalence of byssinosis still existed in Taiwanese cotton mill, although the prevalence was declining. Smoking was found to show an additive effect on cotton dust exposure. Anti-smoking campaign, occupational health program to reduce the dust exposure, and periodical medical examination are measures to prevent from byssinosis.

Study on the effect of food on the absorption of theophylline.

BACKGROUND: The pharmacokinetics of theophylline under fasting and normal meal have been widely studied, but that of Euphyllin Retard in Chinese subjects has not been reported. Since various food-induced absorption changes occur with sustained-release theophylline, it is of interest to study the food effect (especially Chinese food) on this drug product. METHODS: A total of 10 non-smoking healthy male volunteers were involved in the study with a 2-period crossover comparison. They were randomly divided into 2 groups. In the first phase study, group A took a single dose of 350 mg Euphyllin Retard (theophylline-ethylenediamine formulation, containing 255 anhydrous theophylline) under fasting condition. Group B took the same dose with breakfast. Blood samples were collected before and during the 36 hours following administration of the drug. For the second-phase study after 2 weeks, the group A acted as fasting group and group B as non-fasting group. The difference in the absorption of theophylline with fasting versus non-fasting administration was assessed using pharmacokinetic parameters derived from a serum theophylline concentration (STC) vs time curve. RESULTS: The means of maximum drug concentration (Cmax), unextrapolated area under the concentration vs time curve (AUC) from time 0 to 24 hours (AUCun), extrapolated AUC from time 0 to infinity (AUCex), and terminal elimination rate constant (Kel) were higher in the non-fasting group. The fasting group showed a more delayed time to maximum concentration (Tmax). The mean of half-life (T50%) was slightly higher in fasting group. In comparing each of the variables, no statistically significant differences were demonstrated between the 2 modes of administration except Cmax. CONCLUSION: Food increases the rate but not the extent of the absorption of Euphyllin Retard, and one should be aware of the possibility of unwanted side effects caused by high peak concentration. There were wide variations in serum drug levels among individuals, so serum theophylline level monitoring is necessary for an optimal effect. This study was performed in a limited number of normal healthy subjects and the same result is yet to be in asthmatic patients and a larger population of normal subjects.