Blockade of Apoptosis Signal-Regulating Kinase 1 Attenuates Matrix Metalloproteinase 9 Activity in Brain Endothelial Cells and the Subsequent Apoptosis in Neurons after Ischemic Injury.

Conditions of increased oxidative stress including cerebral ischemia can lead to blood-brain barrier dysfunction via matrix metalloproteinase (MMP). It is known that MMP-9 in particular is released from brain endothelial cells is involved in the neuronal cell death that occurs after cerebral ischemia. In the intracellular signaling network, apoptosis signal-regulating kinase 1 (ASK1) is the main activator of the oxidative stress that is part of the pathogenesis of cerebral ischemia. ASK1 also promotes apoptotic cell death and brain infarction after ischemia and is associated with vascular permeability and the formation of brain edema. However, the relationship between ASK1 and MMP-9 after cerebral ischemia remains unknown. Therefore, the aim of the present study was to determine whether blocking ASK1 would affect MMP-9 activity in the ischemic brain and cultured brain endothelial cells. Our results showed that ASK1 inhibition efficiently reduced MMP-9 activity in vivo and in vitro. In endothelial cell cultures, ASK1 inhibition upregulated phosphatidylinositol 3-kinase/Akt/nuclear factor erythroid 2 [NF-E2]-related factor 2/heme oxygenase-1 signals and downregulated cyclooxygenase-2 signals after hypoxia/reperfusion. Additionally, in neuronal cell cultures, cell death occurred when neurons were incubated with endothelial cell-conditioned medium (EC-CM) obtained from the hypoxia/reperfusion group. However, after incubation with EC-CM and following treatment with the ASK1 inhibitor NQDI-1, neuronal cell death was efficiently decreased. We conclude that suppressing ASK1 decreases MMP-9 activity in brain endothelial cells, and leads to decreased neuronal cell death after ischemic injury.

Changes in site of obstruction in obstructive sleep apnea patients according to sleep position: a DISE study.

OBJECTIVES/HYPOTHESIS: This study aimed to evaluate changes in obstruction site in obstructive sleep apnea (OSA) patients according to sleep position. STUDY DESIGN: Prospective case series. METHODS: Eighty-five patients who had undergone level 1 sleep study and drug-induced sleep endoscopy in the supine and lateral positions were included. Obstruction sites were classified as soft palate (SP), tongue base (TB), lateral wall (LW), and larynx (LX). Subgroup analysis was performed according to lateral apnea-hypopnea index (AHI): those with an AHI of ≥ 10 (lateral obstructors, LO) and those with an AHI of < 10 (lateral nonobstructors, LNO). RESULTS: Prevalence in obstruction site of SP, TB, and LX decreased significantly after change from supine to lateral position (P < 0.05). However, the prevalence of LW obstruction was not affected by position change. LW collapse in moderate OSA decreased (from 66.7% to 35.9%) after change to lateral sleep, whereas it persisted in severe OSA patients (81.6%-89.5%). In the lateral position, persistent obstruction at the LW was observed more frequently in the LO group compared to the LNO group (83.3% vs. 33.3%). CONCLUSION: When sleep posture is changed from supine to lateral, obstruction due to structures such as tongue base and larynx improves dramatically. Obstruction in lateral position is mostly due to obstruction at the oropharyngeal LWs. Therefore, position dependency is mostly determined by LW collapsibility. Evaluating the changes of the upper airway according to sleep position can further characterize the upper airway collapsibility and can be used for tailored treatment planning.

The effectiveness of the oncolytic activity induced by Ad5/F35 adenoviral vector is dependent on the cumulative cellular conditions of survival and autophagy.

To overcome the poor tumor transduction efficiency of adenovirus serotype 5 (Ad5) observed in several types of cancer, the fiber region of Ad5, apart from its tail, was replaced by adenovirus serotype 35 (Ad35). The chimeric Ad5/F35 adenoviral vector did not exhibit any significant enhancement of transduction efficiency. CD46, a receptor for Ad35, was expressed in relatively small amounts in most of the cancer cells examined. Therefore, we investigated the pivotal factor(s) that render cancer cells susceptible to transduction. We discovered that the tumor transduction efficiency of Ad5/F35 was enhanced in the presence of rapamycin, an autophagy inducer, in some cancer cells. Analysis of survival potential and cell proliferation rates revealed that Ad5/F35 exerted a more pronounced oncolytic effect in cancer cells with higher survival potential in the presence of rapamycin.

Practices and predictors of 2009 H1N1 vaccination in cancer patients: a nationwide survey in Korea.

BACKGROUND: Because patients with cancer are considered to be at high-risk for influenza infection and related complications, annual vaccination is recommended. The emergence of the novel H1N1 influenza virus in 2009 complicated the medical care of patients with cancer. The present study examined H1N1 vaccination practices among patients with cancer during the pandemic season and investigated factors related to the vaccination. METHODS: A national multicenter cross-sectional survey of patient-doctor dyads was performed; A total of 97 oncologists (response rates of invited participants, 87.4%) and 495 patients (response rates of recruited participants, 86.5%) were included. Patients with cancer provided information concerning vaccination practices and reasons for/against it. Oncologists answered questions about their recommendations and knowledge of H1N1 vaccination. Mixed logistic regression was used to identify patient-level and physician-level predictors of H1N1 vaccination. RESULTS: Only 34.1% of the patients had received H1N1 vaccination, and 53.5% had not considered the need for vaccination. The H1N1 vaccine was proactively recommended by physicians in only a small fraction of patients (8.3%). Increasing age, higher educational status, longer time since the cancer diagnosis, comorbidities, and greater knowledge of H1N1 vaccination among oncologists were significant predictors of patients being vaccinated. CONCLUSIONS: The present results showed low levels of utilization and poor interaction between patients and physicians with regard to the need for vaccination. In addition, the oncologist's level of knowledge affected the adoption of preventive services. Intervention strategies are needed to maximize the rapid adoption of preventive methods to confront future pandemic threats in the cancer patient population.

HSP27 modulates survival signaling networks in cells treated with curcumin and TRAIL.

The combination of curcumin and TRAIL and their role in enhancing apoptotic cell death has been reported by many studies. However, the exact molecular mechanism of apoptosis mediated by curcumin and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is not yet completely understood. In this study, we observed a close connection between dephosphorylated Akt and an increase in phosphorylated heat shock protein 27 (HSP27) during combined treatment with curcumin and TRAIL. Akt dephosphorylation was cumulatively regulated by protein phosphatase 1 (PP1), phosphoinositide-dependent kinase-1 (PDK1), and src. PP1 and PDK1 directly interacted with HSP27, whereas src indirectly interacted with HSP27 via the tumor necrosis factor receptor-associated factor 6 complex. In conclusion, HSP27 modulated cell survival by its interactions with various binding partners, depending on the level of phosphorylated HSP27.

MEKK1/MEKK4 are responsible for TRAIL-induced JNK/p38 phosphorylation.

The tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been shown to activate mitogen-activated protein kinases (MAPKs) depending on caspase and mammalian sterile 20-like kinase 1 activations. However, the upstream molecule of MAPKs has not yet been identified. The mitogen-activated protein kinase kinase 1 (MEKK1) and the apoptosis signal-regulating kinase 1 (ASK1) are considered to be possible candidates for the action of MAPKKKs induced by TRAIL and the possibility of reactive oxygen species involvement has also been investigated. We found that MEKK1/MEKK4 as opposed to ASK1, are responsible for TRAIL-induced c-Jun NH2-terminal kinase (JNK) or p38 activation, and that their catalytic activity is repressed by the caspase-8 inhibitor, suggesting that the caspase-8 activation induced by TRAIL is indispensible for MEKK activation. The 14-3-3 θ was also shown to interact with and to dissociate from MEKK1 by TRAIL treatment, thus implicating the 14-3-3 protein as a negative regulator of MEKK1 activation. Taken together, we show herein that the upstream molecule of the TRAIL-induced MAPK activation is MEKK, as opposed to ASK1, via the mediation of its signal through JNK/p38 in a caspase-8-dependent manner.

TRAIL-induced caspase/p38 activation is responsible for the increased catalytic and invasive activities of Akt.

We previously observed that TRAIL induces acquired TRAIL resistance coinciding with increased Akt phosphorylation brought about by the Src-PI3K-Akt signaling pathways and mediated by c-Cbl. c-Cbl, a ubiquitously expressed cytoplasmic adaptor protein, is simultaneously involved in the rapid degradation of TRAIL receptors and Akt phosphorylation during TRAIL treatment. Here, we show that Akt phosphorylation is not exclusively responsible for acquired TRAIL resistance. Akt catalytic activation is known to increase during metabolic oxidative stress, but we show that TRAIL also dramatically induces the catalytic activation of Akt in TRAIL-sensitive cells, but not in TRAIL-resistant cells. This suggests that Akt catalytic activation during TRAIL-induced apoptosis is likely to play a compensatory role in the maintenance of cell homeostasis. In addition, activated p38 and phosphorylated HSP27 were found to act as downstream effector molecules of p38 during TRAIL treatment and were shown to be responsible for increased Akt catalytic and invasive activities.

A novel homozygous missense mutation in the factor VII gene of severe factor VII deficiency in a newborn baby.

A term male infant born to nonconsanguineous parents was admitted to the hospital for evaluation of lethargy and a pale appearance on the third day of life. He had anemia from an intracranial hemorrhage, and his coagulation factor assay revealed that his bleeding episode was due to severe congenital factor VII deficiency (5% of normal activity). An A-to-G point mutation in the acceptor splice site of intron 5 was identified at nucleotide position 9418. Sequence analysis of the factor VII gene in the parents revealed that they were both heterozygous for a G-to-A transversion at nucleotide position 9418 (IVS5-1) between intron 5 and exon 6. A genetic study involving a patient with a congenitally inherited disease and the parents can confirm the genetic background of the disease and can be used for prenatal guidance to exclude severe bleeding disorders.

A mixture of trans, trans conjugated linoleic acid induces apoptosis in MCF-7 human breast cancer cells with reciprocal expression of Bax and Bcl-2.

The growth inhibitory effect of a mixture of trans, trans conjugated linoleic acid isomers (t, t CLA) was investigated in a human breast cancer cell line, MCF-7, with references to c9, t11 CLA, t10, c12 CLA, and linoleic acid. The t, t CLA treatment effectively induced a cytotoxic effect in a time-dependent (0-6 days) and concentration-dependent (0-40 microM) manner, as compared to the reference and control treatments. The apoptotic parameters were measured on cells treated with 40 microM t, t CLA for 4 days. The occurrence of the characteristic morphological changes and DNA fragmentation confirmed apoptosis. The t, t CLA treatment led to an increase in the level of p53 tumor suppressor protein and Bax protein, but suppressed the expression of Bcl-2 protein. In addition, cytochrome c was released from the mitochondria into the cytosol, and the activation of caspase-3 led to the cleavage of poly(ADP-ribose) polymerase (PARP). Moreover, the composition of the linoleic and arachidonic acids was decreased in cellular membranes. These findings suggest that incorporation of t, t CLA in the membrane induces a mitochondria-mediated apoptosis that can enhance the antiproliferative effect of t, t CLA in MCF-7 cells.

Transduced human PEP-1-heat shock protein 27 efficiently protects against brain ischemic insult.

Reactive oxygen species contribute to the development of various human diseases. Ischemia is characterized by both significant oxidative stress and characteristic changes in the antioxidant defense mechanism. Heat shock protein 27 (HSP27) has a potent ability to increase cell survival in response to oxidative stress. In the present study, we have investigated the protective effects of PEP-1-HSP27 against cell death and ischemic insults. When PEP-1-HSP27 fusion protein was added to the culture medium of astrocyte and primary neuronal cells, it rapidly entered the cells and protected them against cell death induced by oxidative stress. Immunohistochemical analysis revealed that, when PEP-1-HSP27 fusion protein was intraperitoneally injected into gerbils, it prevented neuronal cell death in the CA1 region of the hippocampus in response to transient forebrain ischemia. Our results demonstrate that transduced PEP-1-HSP27 protects against cell death in vitro and in vivo, and suggest that transduction of PEP-1-HSP27 fusion protein provides a potential strategy for therapeutic delivery in various human diseases in which reactive oxygen species are implicated, including stroke.

Detection of porcine circovirus type 2 in feces of pigs with or without enteric disease by polymerase chain reaction.

To establish the sensitive polymerase chain reaction(PCR) method and detect porcine circovirus type 2 (PCV2) from intestines and feces of commercial swine herds with or without enteric disease, intestinal samples from 68 pigs and 29 fecal samples from commercial swine farms were collected. A primer set, forward primer 5'-GAAGAATGGAAGAAGCGG-3' and reverse primer 5'-CTCACAGCAGTAGACAGGT-3', could detect the virus at a concentration as low as 2 infectious virions per milliliter under controlled conditions using PK-15 cell-adapted PCV2. The genomic nucleotide sequences of open reading frame 1 (ORF1) PCR products from fecal samples were found to have complete homology with other PCV2s deposited in the GenBank database. Transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV) as the other enteric pathogens were also investigated by performing duplex reverse transcription-PCR (RT-PCR). Among 63 pigs with clinical enteric disease, 18 PCV2s (14 from intestines and 4 from feces), 7 TGEVs from intestines, and 18 PEDVs (14 from intestines and 4 from feces) were detected by PCR and the duplex RT-PCR. In 34 pigs (14 from intestines and 20 from feces) without clinical enteric disease, only PCV2 was detected in 19 pigs (3 from intestines and 16 from feces). Both PEDV and PCV2 were found in 6 pigs with clinical enteric disease. Among 15 PCV2 samples that were PCR-positive, 4 were culture-positive at passage level 3 in PK-15 cells. These results reveal that PCV2 is shed through the feces of pigs without clinical enteric disease, which suggests the potentiality of the fecal-oral transmission of PCV2 in feces.