Comparison of contrast-enhanced ultrasonography and MRI results obtained by expert and novice radiologists indicating short-term response after transarterial chemoembolisation for hepatocellular carcinoma.

AIM: To evaluate inter-reader agreement between novice and expert radiologists in assessing contrast-enhanced ultrasonography (CEUS) and magnetic resonance imaging (MRI) images for detecting viable tumours with different sizes after conventional transarterial chemoembolisation (cTACE). MATERIALS AND METHODS: This prospective study included patients who had less than five hepatomas and who underwent cTACE. Hepatomas with one or two feeding arteries were selected as target lesions. CEUS and MRI were performed within 1 week after cTACE to evaluate viable tumours. RESULTS: The expert group had higher kappa values in evaluating all tumour sizes via CEUS compared with MRI. The novice group had similar kappa values. In patients with tumours measuring ≤3 cm, the expert group had higher kappa values in reading CEUS compared with MRI images; however, in the novice group, the kappa value was lower in evaluating CEUS compared with MRI images. In patients with tumours measuring >3 cm, the expert and novice groups had good to excellent kappa values. The confidence level of the two groups in reading MRI images was high; however, the novice group had a lower confidence level. CONCLUSION: CEUS is a convenient, cost-effective, and easy to apply imaging tool that can help interventionists perform early detection of viable hepatocellular carcinoma post-TACE. It has a higher inter-rater agreement in interpreting CEUS images compared with MRI images among expert radiologists even when they are extremely familiar with post-cTACE MRI images. In novice radiologists, there may be a learning curve to achieve good consistency in CEUS interpretation.

A case of GNE myopathy mimicking hereditary motor neuropathy.

A 36-year-old woman who presented with upper limb distal weakness since the age of 15 years, with gradual progression to the lower limbs, is reported. Hereditary motor neuropathy was initially suspected based on distal weakness and hyporeflexia; however, whole exome sequencing accidentally revealed a compound heterozygous variant in the GNE gene, and ultrasound revealed increased homogeneous echogenicity in the involved muscles, which is characteristic of myopathic changes. Muscle magnetic resonance imaging revealed fatty infiltration in all limb muscles, sparing the triceps brachii, vastus lateralis and vastus medialis. Muscle biopsy revealed intracytoplasmic rimmed vacuole, supporting the diagnosis of GNE myopathy.

Clinical presentations and skin denervation in amyloid neuropathy due to transthyretin Ala97Ser.

OBJECTIVE: Familial amyloid polyneuropathy (FAP) due to amyloidogenic transthyretin (TTR) is often associated with impairment of thermonociceptive functions. This study investigated skin innervation and its clinical significance in genetically defined FAP due to a hot-spot Ala97Ser TTR mutation (Ala97Ser). METHODS: Skin biopsies were performed on the distal leg of patients with Ala97Ser, and intraepidermal nerve fiber (IENF) densities were quantified. RESULTS: There were 19 unrelated patients with Ala97Ser manifesting a late-onset (59.47 +/- 5.70 years) generalized neuropathy with disabling motor, sensory, and autonomic symptoms. Against a background of a slowly progressive course, 7 patients (36.8%) exhibited additional rapid declines in neurologic deficits, which were associated with elevation of the protein content in the CSF (p < 0.001). The IENF density was markedly reduced in Ala97Ser patients compared to age- and gender-matched controls (0.99 +/- 1.11 vs 8.31 +/- 2.87 fibers/mm, p < 0.001). Skin denervation was present in all patients and was lower in patients with a higher disability grade (0.17 +/- 0.26 vs 1.37 +/- 1.16 fibers/mm, p = 0.003). Albuminocytologic dissociation in the CSF was observed in 14 patients (73.7%), and the IENF density was negatively correlated with the CSF protein concentration (p = 0.015). CONCLUSIONS: Skin denervation was common in Ala97Ser, and degeneration of cutaneous nerve terminals was correlated with the severity of clinical phenotypes and the level of CSF protein.

Involvement of Gas7 along the ERK1/2 MAP kinase and SOX9 pathway in chondrogenesis of human marrow-derived mesenchymal stem cells.

OBJECTIVE: The growth-arrest-specific protein, Gas7, has been shown to be involved in reorganization of the cytoskeleton and for inducing changes in cell shape during cell differentiation. The goals of this study were to investigate the novel role of human Gas7 (hGas7) in chondrogenic differentiation of human mesenchymal stem cells (hMSCs) and to identify the relationship between hGas7, extracellular signal-regulated kinase (ERK1/2) and SOX9 in the chondrogenic pathway. METHODS: Bone marrow-derived hMSCs were induced to undergo chondrogenic differentiation with transforming growth factor-beta1 (TGF-beta1) in an aggregate culture system. The expression of hGas7 and SOX9 and phosphorylation of ERK1/2 at multiple time points were investigated. Chondrogenic capacity was evaluated by the size of aggregates, by glycosaminoglycan content, and by type II collagen and proteoglycan deposition after interfering with expression of hGas7, ERK1/2 or SOX9. To delineate the functional role of these genes in chondrogenesis, inhibition of individual gene's expression in hMSCs, by antisense oligonucleotides or interference RNA (siRNA), and the effect on chondrogenic differentiation were also investigated. RESULTS: Treatment of hMSCs with TGF-beta1 resulted in a transient up-regulation of hGas7b, one of the hGas7 isoforms (day 3-day 5), a transient phosphorylation of ERK1/2 (0.5-4 h) and an up-regulation of SOX9 (2 h to day 14). Transient expression of hGas7b was also detected in hMSCs by reverse transcription-polymerase chain reaction at day 2 and day 3 following TGF-beta1 treatment. Interference with hGas7b production by hGas7b-specific antisense oligonucleotide or inhibition of p-ERK with PD98059, a specific inhibitor of ERK signaling pathway, or interference with SOX9 production by SOX9 siRNA all caused adverse effects of chondrogenic differentiation of hMSCs. Meanwhile, inhibition of p-ERK or SOX9 both blocked the expression of hGas7b. However, the p-ERK and SOX9 pathway was not affected by inhibition of hGas7b. CONCLUSION: These results provide evidence that the transient expression of hGas7b, regulated by activation of ERK1/2 and SOX9 pathway, is essential for chondrogenic differentiation of hMSCs.

Painful neuropathy with skin denervation after prolonged use of linezolid.

The prolonged use of linezolid, a new antibiotic against drug-resistant gram-positive pathogens, might cause painful neuropathy. This finding raises the possibility that small-diameter sensory nerves in the skin, which are responsible for transmitting nociceptive information, might be affected. We report a 53-year-old female who developed pure small-fibre painful neuropathy (visual analogue scale, VAS = 82 on 0-100 scale) with marked skin denervation in the leg (epidermal nerve density, END = 2.32 fibres/mm, norm <5.88 fibres/mm) and significant elevation of the warm threshold in the foot (40.0 degrees C, norm <39.4 degrees C) after the use of linezolid for 6 months. Eight months after the discontinuation of linezolid, the skin became fully reinnervated (END = 9.04 fibres/mm), with disappearance of neuropathic pain (VAS = 0) and normalisation of the warm threshold (36.3 degrees C). Nerve conduction studies for large-diameter motor and sensory nerves were normal. This report documents a pure small-fibre sensory neuropathy after prolonged use of linezolid, and the relationship between skin innervation and corresponding neuropathic pain.

Expression patterns of cell cycle and apoptosis-related genes in a multidrug-resistant human colon carcinoma cell line.

BACKGROUND: An in vitro multidrug resistance (MDR) system from a human colonic cancer cell line (SW620-MDR) has been established. To further study the mechanisms at molecular level and prevention of multidrug resistance in clinical practice, it was demonstrated that the expressions of several apoptosis-related and cell cycle regulator genes were changed in the cells. METHODS: A multidrug-resistant colonic cell line (SW620-MDR) was established, and the Atlas human cDNA expression array was used for studying the pattern of gene expression in this cell line. Furthermore, Northern hybridization or real-time PCR analysis confirmed the pattern of gene expression. RESULTS: In the SW620-MDR cell line the pro-apoptosis genes, CASP4, BIK, PDCD2, and TACE were expressed with decreased levels, and the antiapoptosis genes CD27-L and IGFBP2 were over-expressed. Furthermore, the cell cycle regulator genes such as CDK6, CCND1, CDC27HS, CDC16HS, Wee1Hu, MAPKK1, and IGFBP6 were expressed with decreased levels in the drug-resistant cell line. CONCLUSIONS: It is worthwhile investigating whether the differentially expressed pattern of the aforementioned genes exists in the drug-resistant cancer specimens, and to further understand their functions in the cancer drug-resistance mechanism.

Apaf-1 overexpression partially overcomes apoptotic resistance in a cisplatin-selected HeLa cell line.

Inhibition of caspase-3-mediated apoptosis has been hypothesized to be associated with chemoresistance. Investigations of apoptosis revealed that cytosolic cytochrome c is associated with a complex of apoptotic protease activating factor-1 (Apaf-1), an adapter molecule, and caspase-9 to activate caspase-3. However, whether these apoptotic molecules are involved in acquired cisplatin resistance is not understood. The present work shows reduced activation of caspase-3 and apoptosis in a cisplatin-selected HeLa cell line. Ac-DEVD-CHO, a caspase-3 inhibitor, inhibited cisplatin-induced apoptosis about 60-70% in both cell lines. Ac-LEHD-CHO, a caspase-9 inhibitor or Ac-IETD-CHO, a caspase-8 inhibitor, inhibited cisplatin-induced caspase-3 activation and apoptosis similarly in both cell lines. In addition, cisplatin induced the activation of caspase-9, the upstream activator of caspase-3, in a dose-dependent manner, and the activation of caspase-9 was less induced in resistant cells. The accumulation of cytosolic cytochrome c, an activator of caspase-9, and the induction of the mitochondrial membrane-associated voltage-dependent anion channel were also reduced in cisplatin-resistant cells. However, the concentration of Bcl-2 family proteins in cisplatin-resistant cells was normal. The concentration of Apaf-1 was unaltered in both cell lines. Increasing the cellular concentration of Apaf-1 through the transient expression of the gene increased the induction of apoptosis in resistant cells, associated with enhanced activation of caspase-9, caspase-3 and DNA fragmentation factor. Regression analysis reveals that the modification factor, the ratio of the slope in the linear range of the dose-response curve with Apaf-1 to the slope without Apaf-1, is 1.5 and 4.75 in the HeLa and cisplatin-resistant HeLa cells, respectively. These results indicate that apoptosis and caspases are less induced in cisplatin-selected HeLa cells. They also suggest that ectopic overexpression of Apaf-1 may partially reverse the acquired cisplatin resistance.

Signal transduction pathways and apoptosis in bacteria infected chondrocytes.

The mechanism underlying chronic destructive arthropathy after pyogenic arthritis is not clear. This study evaluated the role of apoptosis in Staphylococcus aureus infected human articular chondrocytes and investigated the signal transduction pathways activated by bacterial infection. Chondrocytes cultured in monolayer were challenged with bacteria for 6 h and were analyzed after incubation for 2, 18, and 24 h. Chondrocytes showed morphologic and biochemical evidences of apoptosis after infection and the following incubation period. Although treatment with extensive washing and vancomycin could ameliorate the amount of apoptosis from 31% to 15% at 2 h, from 48% to 23% at 18 h, and from 58% to 33% at 24 h, the infected samples with treatment still had higher amount of apoptosis than the un-infected controls (ANOVA P < 0.001). Accompanying with the increasing amount of apoptosis, the caspase activity was upregulated in bacteria infected samples and remained high in samples with treatment (ANOVA P < 0.05). Signal transduction pathways activated by bacterial infection were assessed by co-transfection technique. After infection, the c-Jun N-terminal kinase, extracellular signal-regulated kinase, and cyclic AMP-dependent protein kinase activities were elevated by 7.6-, 7.3-, and 3.2-fold, respectively, compared to the uninfected controls. The data support the hypothesis that human chondrocytes will undergo apoptosis after infection by a single organism. Apoptosis and activated intracellular kinase activities may be related to the pathogenesis of post-infectious destructive arthropathy.

A critical role for lymphotoxin-beta receptor in the development of diabetes in nonobese diabetic mice.

To assess the role of lymphotoxin-beta receptor (LTbetaR) in diabetes pathogenesis, we expressed an LTbetaR-Fc fusion protein in nonobese diabetic (NOD) mice. The fusion protein was expressed in the embryo, reached high levels for the first 2 wk after birth, and then declined progressively with age. High expression of LTbetaR-Fc blocked diabetes development but not insulitis. After the decline in chimeric protein concentration, mice became diabetic with kinetics similar to the controls. Early expression of fusion protein resulted in disrupted splenic architecture. However, primary follicles and follicular dendritic cells, but not marginal zones, developed in aged mice. Hence, LTbetaR signaling is required for diabetes development and regulates follicular and marginal zone structures via qualitatively or quantitatively distinct mechanisms.

U50488 inhibits HIV-1 expression in acutely infected monocyte-derived macrophages.

Opioids may play an immunomodulatory role in the pathogenesis of human immunodeficiency virus-1 (HIV-1) infection. Recently, synthetic kappa-opioid receptor (KOR) ligands have been found to have anti-human immunodeficiency virus type 1 activity in acutely infected brain macrophages. In the present study, we investigated whether the selective KOR ligand U50488 would exert such an anti-HIV-1 effect in acutely infected blood monocyte-derived macrophages (MDM). Treatment of acutely infected MDM with U50488 induced a concentration-dependent inhibition of HIV-1 expression. The dose--response relationship of U50488 was U-shaped with a peak effect observed at 10(-13) M, which was evident at both 7 and 14 days post-infection. The KOR antagonist nor-binaltorphimine blocked the anti-HIV-1 effect of U50488 by 73%, indicating involvement of a KOR-mediated mechanism. Also, expression of KOR mRNA and binding activity with a fluorescence-labeled KOR ligand supported the existence of KOR on MDM. Antibodies to the beta-chemokine, RANTES (regulated on activation normal T-cell expressed and secreted), but not to various other cytokines, blocked U50488 inhibition by 56% suggesting that the anti-HIV-1 effect of U50488 involved, in part, the production of RANTES by MDM. Taken together, these in vitro findings support the anti-HIV-1 property of U50488, and suggest that KOR ligands may have therapeutic potential for treating patients with acquired immunodeficiency syndrome.

Cytokine expression in the mouse brain in response to immune activation by Corynebacterium parvum.

Cytokine expression in the brain has been suggested to mediate various sickness behaviors. Here we report that intraperitoneal injection of a Corynebacterium parvum antigen in C57BL/6 mice was followed by prolonged upregulation of cytokines in the cerebral cortex and subcortical structures in a time course that coincided with reduced spontaneous running activity.

Aberrant expression of TSG101 in Taiwan Chinese breast cancer.

Functional inactivation of the tsg101 gene in mouse fibroblasts results in cell transformation and the ability to form metastatic tumors in nude mice. The human tsg101 gene was mapped to chromosome 11q15.1-2 and found to mutate in some cancer patients. To test the expression pattern of the tsg 101 gene in Chinese breast cancer patients, we analyzed the mRNA by RT-PCR in 51 breast cancer patients. The full-length tsg101 and 7 truncated transcripts were detected in both normal and matched tumor tissues. A short transcript with a deletion of nucleotides 154-1054 is frequently presented in late-stage breast cancers. TSG101 protein expression was also detected by Western blot analysis in 30 breast cancer patients. A predicted full-length 46 kDa and three proteins with smaller molecular weight were detected. The full-length 46 kDa protein was less expressed in tumor specimens. Immunohistochemical stains from 10 patients of each stage 0-4 revealed that TSG101 protein was predominantly present in the cytoplasm. Cell nuclei were occasionally immunopositive and the chromosomes were deeply stained during cell division. The intracellular location and the expression of TSG101 protein were both not stage-dependent in primary breast cancers. In addition, normal mammary glands were more homogenously immunopositive than invasive ductal carcinoma. These results support the notion that the aberrant expression of TSG101 in breast cancer is associated with altered cell growth.

The agent of human granulocytic ehrlichiosis induces the production of myelosuppressing chemokines without induction of proinflammatory cytokines.

Infection by human granulocytic ehrlichiosis (HGE) is characterized clinically by cytopenias out of proportion to the number of cells seen to be infected directly. To study the pathogenic role of inflammatory mediators in HGE infection, cytokine production by untreated and dimethyl sulfoxide-treated HL-60 cells, which demonstrate enhanced infection because of granulocytic differentiation, and by normal bone marrow cells was measured using modified sandwich ELISA assays on samples obtained sequentially after inoculation with the HGE agent. All infected cells produced physiological concentrations of CC (monocyte chemotactic protein-1, macrophage inflammatory protein-1alpha and -beta, and RANTES) and CXC (interleukin [IL]-8) chemokines in amounts significantly greater than those produced by uninfected controls. In contrast, infected cells did not secrete the classic proinflammatory cytokines IL-1, IL-6, or tumor necrosis factor-alpha. The striking production of chemokines, powerful leukocyte chemoattractants capable of suppressing hematopoiesis, by susceptible target cells, is likely to be of pathogenic importance both in the observed cytopenias and in mediation of inflammation and host defenses during infection.

Cytokine effects on glutamate uptake by human astrocytes.

Glutamate uptake by astrocytes has been postulated to play a neuroprotective role during brain inflammation. Using primary human fetal astrocyte cultures, we investigated the influence of selected cytokines on glutamate uptake activity. Interleukin (IL)-1beta and tumor necrosis factor-alpha dose-dependently inhibited astrocyte glutamate uptake, whereas interferon (IFN)-gamma alone stimulated this activity. The nitric oxide synthase inhibitor, N(G)-monomethyl-L-arginine, blocked IL-1beta-mediated inhibition of glutamate uptake, suggesting involvement of nitric oxide in the effect of IL-1beta. IL-1 receptor antagonist protein totally reversed the inhibitory effect of cytokines, suggesting a critical role of IL-1beta. The anti-inflammatory cytokine IFN-beta blocked cytokine (IL-1beta plus IFN-gamma)-induced inhibition of glutamate uptake with a corresponding reduction in nitric oxide generation. Taken together, these findings suggest that proinflammatory cytokines inhibit astrocyte glutamate uptake by a mechanism involving nitric oxide, and that IFN-beta may exert a therapeutically beneficial effect by blocking cytokine-induced nitric oxide production in inflammatory diseases of the brain.

Differing functional recovery of donor-derived immune cells after purified haploidentical and fully mismatched hematopoietic stem cell transplantation in mice.

Functional recovery of the immune system is critical for long-term survival in hematopoietic stem cell transplant recipients. In this study, two donor-recipient allogeneic transplant settings (haploidentical and fully mismatched) are used to investigate the functional activity of donor-derived B and T cells in animals grafted with purified c-kit(+), Thy 1.1(lo), Lin(-/lo), and Sca-1(+) hematopoietic stem cells (KTLS HSC).Ovalbumin-specific immunoglobulin G, polyclonal immunoglobulin isotypes, and B- and T-cell proliferation were examined on the recipients who received haploidentical or fully mismatched HSC.A severe deficiency of antigen-specific immunoglobulin response occurs in fully engrafted mice that received KTLS HSC from fully mismatched, but not haploidentical, donors. This lack of B-cell-specific immunity is not due to a deficiency of polyclonal immunoglobulins in serum. B cells from both fully mismatched and haploidentical recipients proliferate normally after stimulation with anti-mu and the percentage of mature B cells is normal. The T-cell response to anti-CD3 in fully mismatched recipients was much weaker than that of their untransplanted controls. However, T cells from haploidentical recipients respond normally to anti-CD3. This study demonstrates that numerical recovery of donor-derived cells in the periphery of recipients does not represent a functional reconstitution, particularly in animals that receive fully mismatched transplants. Defects of specific B-cell immunity and T-cell proliferation are observed in fully mismatched, purified HSC transplant recipients with a quantitative recovery within the normal range of donor-derived lymphocytes.

Role of Iron in the Cellular Effects of Asbestos.

Research to understand how the physical characteristics of asbestos fibers impact the pathological effects has intensified in the past 10 years. The role that iron, intrinsic or acquired, may play has been the subject of many of these investigations. Asbestos catalyzes many of the same reactions that iron does, including lipid peroxidation and DNA damage. At physiological pH, mobilization of iron from the fibers by a low-molecular-weight chelator greatly enhances these reactions. Iron is also mobilized from asbestos fibers in human lung epithelial (A549) cells after the fibers are endocytized. The iron appears to move from a low-molecular-weight pool to nonferritin proteins and ferritin. The amount of iron in the low-molecular-weight pool was directly correlated with the toxicity of the asbestos. Glutathione (CSH) levels in these cells were dramatically reduced after asbestos treatment as a result of CSH efflux. This was not related to the iron associated with the fibers, but appeared to be due in some way to the silicate structure. The presence of the mobilized iron in the cells depleted of CSH creates a very oxidizing environment. Asbestos treatment of cells resulted in DNA oxidation, as assessed by formation of 8-hydroxy-2'-deoxyguanosine (8-oxo-dCJ. The DNA oxidation was dependent upon the iron associated with the fibers and the enzymatic generation of (•) NO by the cells. The induction of the inducible form of nitric oxide synthase (iNOS), responsible for the production of "NO, was dependent upon the presence of iron from the fibers and the decrease in CSH. One without the other did not lead to induction of iNOS. Asbestos was mutagenic in hgprt-gpt(+) V79 Chinese hamster lung cells. The mutagenicity was dependent upon iron, intrinsic or acquired. Addition of (•) NO synergistically increased the mutagenicity of the asbestos, suggesting that in cells that respond to asbestos by making (•) NO, the response will be enhanced. This also suggests that the responses will be enhanced in the presence of activated macrophages producing (•) NO or in the presence of cigarette smoke, because of the presence of (•) NO and iron chelators in the smoke.

Neuroprotective mechanism of glial cell line-derived neurotrophic factor on dopamine neurons: role of antioxidation.

Recombinant human GDNF was infused into the rat striatum either acutely or subchronically. Its effects and its interactions with MPP+ on antioxidant enzyme activities were examined. Results indicated that acute GDNF infusion significantly increased glutathione peroxidase, superoxide dismutase and catalase activities. Subchronic GDNF treatment decreased the DA level and enhanced DA turnover. Pre-treatment with GDNF markedly protected DA neurons against MPP+-induced toxicity. These results suggest that GDNF protects DA neurons through its activation of the antioxidant enzyme systems.

Inhibition of microglial cell RANTES production by IL-10 and TGF-beta.

Using human fetal microglial cell cultures, we found that the gram-negative bacterial cell wall component lipopolysaccharide (LPS) stimulated RANTES (regulated upon activation of normal T cell expressed and secreted) production through the protein kinase C signaling pathway and that activation of transcription nuclear factor (NF)-kappaB was required for this effect. Similarly, the proinflammatory cytokines interleukin (IL)-1beta and tumor necrosis factor-alpha dose-dependently stimulated microglial cell RANTES production via NF-kappaB activation. Anti-inflammatory cytokines, IL-10, and transforming growth factor (TGF)-beta sequentially inhibited LPS- and cytokine-induced microglial cell NF-kappaB activation, RANTES mRNA expression, and protein release. Proinflammatory cytokines but not LPS also stimulated RANTES production by human astrocytes. These findings demonstrate that human microglia synthesize RANTES in response to proinflammatory stimuli, and that the anti-inflammatory cytokines IL-10 and TGF-beta down-regulate the production of this beta-chemokine. These results may have important therapeutic implications for inflammatory diseases of the brain.