Detection of Mycobacterium tuberculosis Complex in Paraffin-Embedded Tissues by the New Automated Abbott RealTime MTB Assay.

OBJECTIVE: The Abbott RealTime MTB assay, launched in June 2014, has been shown to have a competitive performance in the detection of the Mycobacterium tuberculosis (MTB) complex in respiratory specimens. The present study was conducted to investigate the usefulness of the Abbott MTB Realtime assay in the detection of MTB in formalin-fixed paraffin-embedded (FFPE) tissues. METHODS: A total of 96 FFPE specimens obtained from microbiologically proven MTB cases (N=60) and nontuberculous Mycobacterium cases (N=36) were analyzed. The performance of the Abbott MTB Realtime assay was compared with that of the Roche Cobas TaqMan MTB assay. RESULTS: The overall sensitivity and specificity of the Abbott assay were 63.3% and 97.2%, respectively, compared with 11.7% and 100% for the Cobas assay. The detection rate of the Abbott assay was much higher among 37 acid-fast-positive specimens than among 23 acid-fast-negative specimens (89.3% versus 21.7%, respectively). The detection rate of the assay was higher among 29 resection specimens than among 31 small biopsy specimens (86.2% versus 41.9%, respectively). CONCLUSION: Our results suggest that the Abbott RealTime MTB assay can be used to differentiate MTB from nontuberculous mycobacterial infections in acid-fast-positive FFPE tissues.

Deregulation of SLIT2-mediated Cdc42 activity is associated with esophageal cancer metastasis and poor prognosis.

INTRODUCTION: SLIT2, a secreted protein, has been found to inactivate Cdc42 GTPase to modulate neural cell migration. However, alteration of SLIT2-mediated Cdc42 in terms of migration regulation remains undefined in esophageal squamous cell carcinoma (ESCC). METHODS: We report here in ESCC cell, animal, and clinical models that SLIT2 acts as a migration suppressor and serves as a prognostic biomarker. RESULTS: The immunohistochemistry data indicated that 31.8% (49 of 154) of tumors from ESCC patients showed low expression of SLIT2 protein which correlated with poor overall survival and disease-free survival. DNA methylation analysis suggested that promoter hypermethylation is responsible for low expression of SLIT2 in ESCC. Knockdown of SLIT2 increased ESCC cell migration, while SLIT2 stable overexpression reduced cell migration. ESCC cells treated with conditioned media from cells overexpressing SLIT2 also suppressed cell migration. Importantly, silencing of SLIT2 decreased the complex formation, and thus induced Cdc42 activity and promoted membrane localization of focal adhesion kinase and Paxillin. Anti-metastatic effect of SLIT2 was confirmed in an experimental metastasis model of SLIT2 knockdown ESCC cells. CONCLUSION: Our results provide novel evidence that low expression of SLIT2 correlates with poor prognosis and promotes metastasis in ESCC, which may be regulated by the Cdc42-mediated pathways.

Low SOX17 expression is a prognostic factor and drives transcriptional dysregulation and esophageal cancer progression.

The transcriptional network of the SRY (sex determining region Y)-box 17 (SOX17) and the prognostic impact of SOX17 protein expression in human cancers remain largely unclear. In this study, we evaluated the prognostic effect of low SOX17 protein expression and its dysregulation of transcriptional network in esophageal squamous cell carcinoma (ESCC). Low SOX17 protein expression was found in 47.4% (73 of 154) of ESCC patients with predicted poor prognosis. Re-expression of SOX17 in ESCC cells caused reduced foci formation, cell motility, decreased ESCC xenograft growth and metastasis in animals. Knockdown of SOX17 increased foci formation in ESCC and normal esophageal cells. Notably, 489 significantly differential genes involved in cell growth and motility controls were identified by expression array upon SOX17 overexpression and 47 genes contained putative SRY element in their promoters. Using quantitative chromatin immunoprecipitation-PCR and promoter activity assays, we confirmed that MACC1, MALAT1, NBN, NFAT5, CSNK1A1, FN1 and SERBP1 genes were suppressed by SOX17 via the SRY binding-mediated transcriptional regulation. Overexpression of FN1 and MACC1 abolished SOX17-mediated migration and invasion suppression. The inverse correlation between SOX17 and FN1 protein expression in ESCC clinical samples further strengthened our conclusion that FN1 is a transcriptional repression target gene of SOX17. This study provides compelling clinical evidence that low SOX17 protein expression is a prognostic biomarker and novel cell and animal data of SOX17-mediated suppression of ESCC metastasis. We establish the first transcriptional network and identify new suppressive downstream genes of SOX17 which can be potential therapeutic targets for ESCC.

Metformin enhances cisplatin cytotoxicity by suppressing signal transducer and activator of transcription-3 activity independently of the liver kinase B1-AMP-activated protein kinase pathway.

Metformin has been used as first-line treatment in patients with type 2 diabetes, and is reported to reduce cancer risk and progression by activating the liver kinase B1 (LKB1)-AMP-activated protein kinase (AMPK) pathway. Cisplatin remains the main drug for treating advanced non-small-cell lung cancer. However, drug resistance often develops through several mechanisms during the treatment course, including one mechanism mediated by the activation of the IL-6/signal transducer and activator of transcription (STAT)-3 pathway, related to the generation of reactive oxygen species (ROS). This study demonstrated a correlation between STAT3 phosphorylation and cisplatin cytotoxicity, using AS2 (PC14PE6/AS2)-derived cell lines (AS2/S3C) that contained constitutively active STAT3 plasmids as a model. A STAT3 inhibitor (JSI-124) enhanced the cisplatin sensitivity in AS2 cells, whereas metformin inhibited STAT3 phosphorylation and enhanced cisplatin cytotoxicity. By contrast, another AMPK activator (5-aminoimidazole-4-carboxamide-riboside) failed to produce these effects. LKB1-AMPK silencing by small, interfering RNA or mammalian target of rapamycin (mTOR) inhibition by rapamycin or pp242 did not alter the effect of metformin on STAT3 activity suppression, suggesting that metformin can modulate the STAT3 pathway through an LKB1-AMPK-independent and probably mTOR-independent mechanism. Metformin also inhibited cisplatin-induced ROS production and autocrine IL-6 secretion in AS2 cells. Both mechanisms contributed to the ability of metformin to suppress STAT3 activation in cancer cells, which resulted in the decreased secretion of vascular endothelial growth factor by cancer cells. The growth of subcutaneous tumor xenografts was significantly delayed by a combination of cisplatin and metformin. This is the first study to demonstrate that metformin suppresses STAT3 activation via LKB1-AMPK-mTOR-independent but ROS-related and autocrine IL-6 production-related pathways. Thus, metformin helps to overcome tumor drug resistance by targeting STAT3.

Anti-IL-20 monoclonal antibody suppresses breast cancer progression and bone osteolysis in murine models.

IL-20 is a proinflammatory cytokine involved in rheumatoid arthritis, atherosclerosis, and stroke. However, little is known about its role in breast cancer. We explored the function of IL-20 in tumor growth and metastasis, as well as in clinical outcome. Tumor expression of IL-20 was assessed by immunohistochemical staining among 198 patients with invasive ductal carcinoma of the breast, using available clinical and survival data. IL-20 expression was associated with advanced tumor stage, greater tumor metastasis, and worse survival. Reverse transcription quantitative polymerase chain reaction showed that clinical breast tumor tissue expressed higher levels of IL-20 and its receptors than did nontumorous breast tissue. IL-20 was also highly expressed in breast cancer bone-metastasis tissue. In vitro, IL-20 upregulated matrix metalloproteinase-9, matrix metalloproteinase-12, cathepsin K, and cathepsin G, and enhanced proliferation and migration of breast cancer cells, which were inhibited by anti-IL-20 mAb 7E. In vivo, we generated murine models to evaluate the therapeutic potential of 7E, using luminescence intensity, radiological scans, and micro-computed tomography. 7E reduced tumor growth, suppressed bone colonization, diminished tumor-mediated osteolysis, and lessened bone density decrement in mice injected with breast cancer cells. In conclusion, our results suggest that IL-20 plays pivotal roles in the tumor progression of breast cancer. IL-20 expression in breast cancer tissue is associated with a poor clinical outcome. Anti-IL-20 mAb 7E suppressed bone colonization and decreased osteolytic bone lesions. Therefore, IL-20 may be a novel target in treating breast tumor-induced osteolysis.

Heteroresistance to cephalosporins and penicillins in Acinetobacter baumannii.

Heteroresistance to antimicrobial agents may affect susceptibility test results and therapeutic success. In this study, we investigated heteroresistance to cephalosporins and penicillins in Acinetobacter baumannii, a major pathogen causing nosocomial infections. Two A. baumannii isolates exhibited heteroresistance to ampicillin-sulbactam, ticarcillin-clavulanic acid, cefepime, and cefpirome, showing a distinct colony morphology of circular rings within the inhibition halos. Pulsed-field gel electrophoresis (PFGE) and outer membrane protein (OMP) analysis demonstrated that subpopulations around the disks/Etest strips and the original strains all belonged to the same PFGE type and OMP profile. Population analysis profile (PAP) showed the presence of heteroresistant subpopulations with high cefepime resistance levels in two isolates (008 and 328). Interestingly, A. baumannii 008 contained two peaks: one was grown in the presence of up to 1 µg of cefepime/ml, the other apparently occurred when the concentration of cefepime was raised to 256 µg/ml. After serial passages without exposure to cefepime, the PAP curve maintained the same trend observed for the original strain of A. baumannii 008. However, the PAP curve showed a shift to relatively lower cefepime resistance (from 256 to 64 µg/ml) in A. baumannii 328 after 10 passages in antibiotic-free Mueller-Hinton agar plates. Convergence to a monotypic resistance phenotype did not occur. Growth rate analysis revealed that slower growth in resistant subpopulations may provide a strategy against antibiotic challenge. To our knowledge, this is the first report of heteroresistance to cephalosporins and penicillins in A. baumannii.

Apolipoprotein E expression promotes lung adenocarcinoma proliferation and migration and as a potential survival marker in lung cancer.

Many human lung cancer cell lines express apolipoprotein E (ApoE), especially cells derived from malignant pleural effusions (MPE) in patients with lung adenocarcinoma. This study aimed to investigate the influence of ApoE expression on lung cancer. In lung cancer tissues, ApoE expression was more frequently found in malignant pleural effusions (MPE)-associated lung adenocarcinoma than in lung adenocarcinoma or squamous cell carcinoma without MPE (P<0.05), indicating that ApoE is associated with the pathogenesis of MPE in patients with lung adenocarcinoma. Next, we examined the roles of ApoE in an MPE-derived lung adenocarcinoma cell line that endogenously over-expresses ApoE, PC14PE6/AS2 (AS2). In that experiment we inhibited ApoE expression by transfection of a plasmid carrying ApoE siRNAs into AS2 cells to generate AS-S2 and AS-S3 cells. Compared to vector-control cells and parental AS2 cells, AS2-S2 and AS2-S3 cells grew slower (P<0.05), were more sensitive to cisplatin, and had significantly impaired cellular migration (P<0.05). Furthermore, over-expression of ApoE was independently associated with poor survival in lung adenocarcinoma patients who had MPE at the time of diagnosis (P<0.001). Conclusively, ApoE over-expression promotes cancer proliferation and migration and contributes to an aggressive clinical course in patients with lung adenocarcinoma and MPE.

Single cell phospho-specific flow cytometry can detect dynamic changes of phospho-Stat1 level in lung cancer cells.

Single cell phospho-specific flow cytometry (SCPFC) enables the investigation of signaling network interactions and the categorization of disease outcome. While this method has been successfully used to study hematologic disorders, its application on solid tumors has not been examined. This study aimed to demonstrate the ability of SCPFC to detect dynamic changes of Tyrosine phospho-Stat1 (pStat1) in solid tumor models and in human tumor samples. In the human lung cancer cell line PC14PE6/AS2, the fluorescence intensity changes of pStat1 after IFN-γ stimulation were compatible to results obtained by Western blot analysis. In metastatic animal models, cancer cells from subcutaneous tumors, malignant ascites, and peritoneal tumors responded to IFN-γ. The pStat1 was activated in these cells after IFN-γ stimulation, with a 1.5- to 2.5-fold increase in fluorescence intensity compared to the unstimulated control. To examine the potential clinical application of SCPFC, cancer cells were collected from malignant pleural effusions (MPEs) of lung cancer patients to observe the activation of pStat1 after IFN-γ stimulation. Cell apoptosis after cisplatin treatment was evaluated by Annexin V staining, which showed that MPE cancer cells with higher pStat1 changes after IFN-γ stimulation were more resistant to cisplatin. In conclusion, there is a preliminary application of SCPFC to solid tumors and links to drug sensitivity are promising.

Extended-spectrum beta-lactamase-producing phenotype signifies a poor prognosis for patients with cefpodoxime-resistant Escherichia coli or Klebsiella pneumoniae bacteremia.

BACKGROUND AND PURPOSE: Bloodstream infections caused by multidrug-resistant Enterobacteriaceae are a major concern. This study explored the clinical impact of extended-spectrum beta-lactamase (ESBL) production among cefpodoxime-resistant Escherichia coli and Klebsiella pneumoniae bacteremia. METHODS: The medical charts and microbiological results of patients with cefpodoxime-resistant E. coli or K. pneumoniae bacteremia in a tertiary hospital in southern Taiwan between June 2003 and December 2006 were retrospectively reviewed. The clinical characteristics, medical histories, and clinical outcomes were evaluated. ESBL production was indicated by the double-disk synergy test. RESULTS: 278 episodes of bacteremia caused by cefpodoxime-resistant K. pneumoniae or E. coli were identified, of which 115 (41%) were ESBL producing. Compared with non-ESBL-producing bacteremia, bacteremic episodes caused by ESBL producers were less often community acquired (4.3% vs 26.4%; p < 0.001). Underlying diabetes mellitus (48.7% vs 35.0%; p = 0.02), liver cirrhosis (22.6% vs 11.7%; p = 0.02), or uremia (21.7% vs 3.7%; p < 0.001) were more common in ESBL-producing bacteremia. In contrast, solid tumors were more frequent in non-ESBL-producing bacteremia (44.8% vs 27.8%; p = 0.004). Overall, patients with ESBL-producing bacteremia had higher disease severity indicated by a Pittsburgh bacteremia score > or = 4, longer duration of hospital stay (51.1 days vs 31.9 days; p = 0.007), more admission to intensive care units (19.1% vs 8.0%; p = 0.006), and a higher mortality rate at 28 days (34.8% vs 23.9%; p = 0.03). CONCLUSIONS: ESBL production signifies a poor clinical outcome for patients with bacteremia caused by cefpodoxime-resistant E. coli or K. pneumoniae.

Angiogenetic biomarkers in non-small cell lung cancer with malignant pleural effusion: correlations with patient survival and pleural effusion control.

OBJECTIVE: To evaluate how the angiogenetic biomarkers vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and interleukin-8 (IL-8) in the fluid of non-small cell lung cancer (NSCLC) with malignant pleural effusion (MPE) correlate with patient survival and pleural effusion control. DESIGN: Prospective study. PATIENTS AND METHODS: From April 1, 1998 to April 30, 2005, we used thoracoscopic biopsy to collect pleural specimens and pleural effusion from 97 patients with NSCLC and MPE. Paired blood samples were harvested. We used enzyme-linked immunosorbent assays to evaluate levels of angiogenic factors in MPE and blood, and immunohistochemical staining to evaluate them in pleural specimens. Related data, such as patient survival and PE control, were collected for correlation analysis. RESULTS: Smoking and PE VEGF >1350 ng/mL were both significant negative predictors of patient survival. A trapped lung was the only significant factor for poor PE control. The serum level, the amount of PE, and the number of red blood cells in PE correlated well with PE VEGF level. Immunohistochemical staining of pleural samples showed that VEGF was secreted by both mesothelial and tumor cells. The level of PE IL-8 weakly correlated with PE VEGF, and the level of bFGF was not significant. CONCLUSIONS: PE VEGF was a useful angiogenetic factor for the amount of fluid in patients with NSCLC and MPE. In addition to smoking, PE VEGF >1350 ng/mL was a significant negative predictor of patient survival.

IL-20 may contribute to the pathogenesis of human intervertebral disc herniation.

STUDY DESIGN: The gene expression of interleukin (IL)-20 on human herniated intervertebral disc. OBJECTIVE.: To elucidate the role of novel cytokine IL-20 in the pathogenesis of human intervertebral disc (IVD) herniation. SUMMARY OF BACKGROUND DATA: IL-20 is involved in inflammatory diseases such as psoriasis, atherosclerosis, and rheumatoid arthritis, etc. However, IL-20 is never reported to be associated with the pathogenesis of human disc herniation. METHODS: Twenty consecutive patients who were diagnosed with IVD herniation and received open discectomy were included in this study. The retrieved disc material specimens and the isolated primarily cultured disc cells were immunohistochemically stained to detect the expression of IL-20 and its receptor subunits (IL-20R1, IL-20R2, and IL-22R1). Besides, to investigate the in vitro response of IL-20 on human herniated intervertebral disc, we analyzed the effects of IL-20 alone, in combination with IL-1beta, and IL-1beta alone on the gene expression and protein levels of various cytokines, chemokines, matrix metalloproteinases (MMPs), etc. RESULTS: IL-20 and its receptors were detectable in human herniated disc tissues and isolated disc cells. In vitro, IL-1beta induced the expression of IL-20. Furthermore, IL-20 induced transcripts of IL-1beta, IL-6, vascular endothelial growth factor (VEGF), MMP-3, and monocyte chemoattractant protein (MCP-1) on primarily cultured human disc cells. IL-1beta induced transcripts of IL-1beta, IL-6, IL-8, VEGF, MMP3, and MCP-1. IL-20 combined with IL-1beta induced transcripts of tumor necrosis factor-alpha (TNF-alpha), IL-1beta, IL-6, IL-8, MMP-3, and MCP-1 to a level higher than those found in cells treated with IL-20 or IL-1beta alone.Enzyme-linked immunosorbent assay, analysis also showed that IL-20 combined with IL-1beta up-regulated the secretion of TNF-alpha, IL-6, IL-8, and MCP-1. CONCLUSION: IL-20 induces proinflammatory, chemotaxtic, and matrix degradative responses in IVD cells especially in combination with IL-1beta. Our study suggests that IL-20 plays an important role in the pathogenesis of disc herniation.

Langerhans cell histiocytosis as a possible differential diagnosis of painful scoliosis.

We report on a 5-year 8-month-old boy suffering from spinal Langerhans cell histiocytosis (LCH), who had initial symptoms of back and abdominal pain, as well as tilting of the shoulder that mimicked hemivertebra of T10 with scoliosis, as revealed by radiography. The LCH-involved vertebra did not demonstrate the classic radiographic picture of vertebra plana until the vertebral body symmetrically collapsed about 6 months later, when the patient's scoliosis disappeared. The delayed diagnosis of LCH was confirmed by biopsy after another 6 months. Polyostotic lesions affecting C6, T5, T9-12, and L2 were found. This case represented an unusual presentation of LCH as an early disease entity, which resulted in a misdiagnosis of painful scoliosis. We believe we are the first to report LCH as a differential diagnosis of painful scoliosis.

The in vivo biological effects of intradiscal recombinant human bone morphogenetic protein-2 on the injured intervertebral disc: an animal experiment.

STUDY DESIGN: Prospective analysis. OBJECTIVE: To investigate biologic influences of recombinant human bone morphogenetic protein (rhBMP)-2 on intervertebral discs after anular tears. SUMMARY OF BACKGROUND DATA: Treatments for intervertebral disc injury or degeneration are unsatisfactory. rhBMP-2, a high-potency osteoinductive and chondroinductive substance, is approved for use in anterior lumbar interbody fusions. rhBMP-2 stimulates the proliferation of rat disc cells and the secretion of extracellular matrix in vitro. In vivo responses in the intervertebral disc after anular tears are rarely studied. METHODS: Twenty New Zealand white rabbits received full-thickness anular tears and intradiscal injections of saline (control) and rhBMP-2 0.1 mg with and without coral grafts at L2-L3, L3-L4, and L4-L5, respectively. Three died or had infection. Therefore, 17 underwent radiography and sacrifice at 12 weeks. Spinal sections were stained with hematoxylin and eosin to examine responses to rhBMP-2. RESULTS: Radiographs revealed degenerative changes, such as disc space narrowing and irregularity, subchondral sclerosis, osteophyte formation, and hypertrophy of vertebral endplates in all groups. Degeneration was more frequent and severe with rhBMP-2 with (P < 0.01) and without (P < 0.05) coral than with saline. Two rabbits receiving rhBMP-2 and coral achieved solid interbody bony fusion. New bone formation was noted in 2 controls, in 3 animals treated with rhBMP-2, and in 4 treated with rhBMP-2 and coral. Vascularity and fibroblast proliferation increased with rhBMP-2 (n = 14) and rhBMP-2 with coral (n = 9) compared with control (n = 3; P < 0.01 and P = 0.03, respectively). Inflammatory infiltrates increased with rhBMP-2 (n = 8) compared with control (n = 2; P = 0.03). CONCLUSIONS: Degenerative changes were more frequent and severe in the groups treated with rhBMP-2 with or without coral in radiographic findings. In histopathologic findings, rhBMP-2 promoted hypervascularity and fibroblast proliferation of the intervertebral disc after an anular tear.

Clinical significance and distribution of putative virulence markers of 116 consecutive clinical Aeromonas isolates in southern Taiwan.

OBJECTIVES: The objectives of this study were to elucidate the clinical manifestations of Aeromonas infections and the association of putative virulence genes with clinical invasiveness. METHODS: 116 consecutive clinical Aeromonas isolates collected from July 1999 to June 2001 in a medical center in southern Taiwan were included. All isolates were identified by biochemical phenotyping and their genomic sequences encoding eight putative virulence factors, including cytolytic enterotoxin (AHCYTOEN), aerolysin (aerA), hemolysin (hlyA), heat-labile enterotoxin (alt), heat-stable enterotoxin (ast), and components of type III secretion system (ascV, aexT and ascF-ascG) were analyzed using polymerase chain reaction and colony blot hybridization. The association of clinical diseases of the patients with the putative virulence genes in the isolates was analyzed. RESULTS: Sixty-two percent of Aeromonas isolates caused clinically evident infections, of which the major clinical manifestations were primary bacteremia (40%), followed by soft tissue infections (27%), and hepatobiliary tract infections (15%). Liver cirrhosis (36%), malignancy (25%), and hepatobiliary diseases (13%) were the major underlying diseases in patients with Aeromonas bacteremia. The majority (64%) of patients with Aeromonas hepatobiliary infections had underlying hepatobiliary diseases, whereas 71% of those with soft tissue infections had antecedent water- or soil-related injuries. The crude fatality rate for Aeromonas infections was 26%. Aeromonas hydrophila complex was the most common (52%) of the three major complex groups investigated, followed by Aeromonas sobria complex (24%) and Aeromonas caviae complex (23%). None of the eight putative virulence factors was associated with invasiveness or bacteremia. CONCLUSIONS: Primary bacteremia, soft tissue infections, and hepatobiliary tract infections are the three major clinical manifestations of invasive Aeromonas infections in southern Taiwan. This study found no association between the presence of AHCYTOEN, aerA, hlyA, alt, ast, ascV, aexT or ascF-ascG genes in Aeromonas isolates and the development of extra-intestinal infections or bacteremia.

Transesophageal echocardiographic recognition of infiltrative cardiac sarcoma mimicking mitral stenosis.

Primary cardiac sarcomas are very rare. Infiltrative cardiac tumors may be difficult to diagnose by transthoracic echocardiography (TTE) only. Herein, we report a case of primary unclassified cardiac sarcoma with clinical and echocardiographic manifestations of mitral stenosis (MS). The tumor was not identified by TTE preoperatively because of its diffuse infiltration of the left atrial wall and both mitral leaflets without protruding mass, and was only discovered by intraoperative transesophageal echocardiography (TEE). This report alerts clinicians that TEE is a necessary adjunctive tool to facilitate the correct diagnosis in patients with obscure etiologies of mitral valve diseases especially when they will receive surgical intervention.

Histopathologic findings of retrieved specimens of vertebroplasty with polymethylmethacrylate cement: case control study.

STUDY DESIGN: Case control study. OBJECTIVE: To investigate the histopathologic findings of 2 retrieved specimens from failed vertebroplasty with polymethylmethacrylate (PMMA) cement. SUMMARY OF BACKGROUND DATA: Vertebroplasty using PMMA cement has been commonly used to treat debilitating back pain from compression fracture, angiomas, and metastatic cancer. However, there was concern about the unpredictable future results with PMMA cement. The histopathologic changes were rarely reported. METHODS: There were 2 PMMA augmented and 3 nonaugmented fractured vertebral bodies retrieved for histopathologic study. Between the 2 groups, we compared the findings of bone necrosis, foreign body reaction, fibrotic wall formation, and neovascularization. RESULTS: Bone necrosis was noted in the periphery of PMMA cement, which was surrounded by fibrotic tissues. In contrast, no fibrotic wall formation could be found in the nonaugmented control group. Foreign body reaction was only noted in PMMA augmented cases, and neovascularization was only noted in the control cases. CONCLUSION: PMMA cement might not be as bioinert as we considered. Therefore, the long-term safety of vertebroplasty should be further evaluated.

A mouse-adapted enterovirus 71 strain causes neurological disease in mice after oral infection.

A mouse-adapted enterovirus 71 (EV71) strain with increased virulence in mice, MP4, was generated after four serial passages of the parental EV71 strain 4643 in mice. Strain MP4 exhibited a larger plaque size, grew more rapidly, and was more cytotoxic in vitro than strain 4643. Although strains 4643 and MP4 both induced apoptosis of SK-N-SH human neuroblastoma cells, MP4 was more virulent than 4643 in 1-day-old mice (50% lethal doses, 10(2) and 10(4) PFU/mouse, respectively). Strain MP4 (5 x 10(6) PFU/mouse), but not 4643, could orally infect 7-day-old mice, resulting in rear-limb paralysis followed by death 5 to 9 days after inoculation with the virus. Histopathologically, neuronal loss and apoptosis were evident in the spinal cords as well as the brain stems of the infected mice. The limb muscles displayed massive necrosis. There was early and transient virus replication in the intestines, whereas the spinal cord, brain, and muscle became the sites of viral replication during the late phase of the infection. Virus transmission occurred among infected and noninfected cagemates, as demonstrated by the occurrence of seroconversion and the presence of viable viruses in the stool samples of the latter. Protection against EV71 challenge was demonstrated following administration of hyperimmune serum 1 day after inoculation with the virus. Nucleotide sequence analysis of the genome of EV71 strain MP4 revealed four nucleotide changes on the 5' untranslated region, three on the VP2 region, and eight on the 2C region, resulting in one and four amino acid substitutions in the VP2 and 2C proteins, respectively.