Extramedullary hematopoiesis in pyogenic granuloma.

Extramedullary hematopoiesis (EMH) suggests the presence of hematopoietic stem cells (HSC) outside bone marrow. EMH has been reported, albeit rarely, in pyogenic granuloma (PG), a polypoid lobular capillary hemangioma. However, statistical data have hitherto been lacking on the actual incidence of EMH in PG. Therefore, we here reviewed 157 consecutive cases using routine diagnostic surgical slides and found unequivocal EMH in 17 (10.8%). This indicates that EMH is a rather common finding in PG, which could thus have strong potential to be an important resource for the study of HSC.

Appearance of nuclear-sorted caspase-12 fragments in cerebral cortical and hippocampal neurons in rats damaged by autologous blood clot embolic brain infarctions.

Following endoplasmic reticulum (ER) stress, cerebral infarctions have been reported to involve an apoptotic process, including the activation of the caspase cascade. To confirm whether fragmented caspase-12, which is activated by cleavage and is detectable during ER stress, is also involved in embolic cerebral infarctions in rats, we adopted an autologous blood clot model for the analysis of cerebral infarctions. We performed experiments in rats with brain infarctions, which are closely related to embolic cerebral infarctions. We utilized a homologous blood clot, i.e., natural materials, to form the infarct area. Our findings reveal that caspase-12 is fragmented when infarct areas form in cerebral cortical neurons. Interestingly, we observed that these fragments translocated to the nuclei of not only cerebral cortical neurons but hippocampal neurons. We further found that glucose-regulated protein 78 (GRP78), a marker of ER stress, is up-regulated in both cerebral cortical and hippocampal neurons during cerebral infarction. This result suggests that the fragmentation of caspase-12 and the subsequent nuclear translocation of these fragments are involved in the brain infarction process in rats.

Heterochromatin protein 1gamma epigenetically regulates cell differentiation and exhibits potential as a therapeutic target for various types of cancers.

Heterochromatin protein 1 (HP1) is a chromosomal protein that participates in both chromatin packaging and gene silencing. Three HP1 isoforms (alpha, beta, and gamma) occur in mammals, but their functional differences are still incompletely understood. In this study, we found that HP1gamma levels are decreased during adipocyte differentiation, whereas HP1alpha and beta levels are expressed constitutively during adipogenesis in cultured preadipocyte cells. In addition, ectopic overexpression of HP1gamma inhibited adipogenesis. Furthermore, we did not detect any HP1gamma protein in the differentiated cells of various normal human tissues. These results suggest that the loss of HP1gamma is required for cell differentiation to occur. On the other hand, the methylation levels of lysine 20 (K20) on histone H4 showed a significant correlation with HP1gamma expression in both these preadipocyte cells and normal tissue samples. However, all cancer tissues examined were positive for HP1gamma but were often negative for trimethylated histone H4 K20. Thus, a dissociation of the correlation between HP1gamma expression and histone H4 K20 trimethylation may reflect the malfunction of epigenetic control. Finally, suppression of HP1gamma expression restrained cell growth in various cancer-derived cell lines, suggesting that HP1gamma may be an effective target for gene therapy against various human cancers. Taken together, our results demonstrate the novel function of HP1gamma in the epigenetic regulation of both cell differentiation and cancer development.

USP15 plays an essential role for caspase-3 activation during Paclitaxel-induced apoptosis.

Paclitaxel (also known as Taxol) is a well-known anticancer agent that blocks cell mitosis and kills tumor cells, and is often used in clinic to treat cancers. Despite the success of Paclitaxel, the development of drug resistance prevents its clinical applicability. Here, we screened an siRNA library against the entire human genomes using HeLa cells, and have find that lack of USP15 (ubiquitin-specific protease 15) causes Paclitaxel resistance. We also observed the decreased expression of USP15 in Paclitaxel-resistant human ovarian cancer samples. In addition, we have demonstrated that USP15 plays an essential role for stability and activity of caspase-3 during Paclitaxel-induced apoptosis. Thus, USP15 may be a candidate diagnostic marker and therapeutic target for Paclitaxel-resistant cancers.

Effects of 915 MHz electromagnetic-field radiation in TEM cell on the blood-brain barrier and neurons in the rat brain.

The aim of this study was to determine whether albumin leakage and dark neurons were present in rat brains 14 and 50 days after a single 2-h exposure to a 915 MHz electromagnetic field, as reported by Salford et al. (Environ. Health Perspect. 111, 881-883, 203). Sixty-four male F344 rats (12 weeks old) were exposed to a 915 MHz electromagnetic field at whole-body average specific absorption rates of 0, 0.02, 0.2 and 2.0 W/kg in TEM cells for 2 h, following the protocol reported by Salford et al. The brains were examined histologically and immunohistochemically. No albumin immunoreactivity was observed in the exposed groups. In addition, dark neurons, assessed using hematoxylin and eosin staining, were rarely present, with no statistically significant difference between exposed and sham-exposed animals. This study thus failed to confirm the results of Salford et al.

Glucocorticoid prevents brain-derived neurotrophic factor-mediated maturation of synaptic function in developing hippocampal neurons through reduction in the activity of mitogen-activated protein kinase.

An increased level of glucocorticoid may be related to the pathophysiology of depressive disorder. The involvement of brain-derived neurotrophic factor (BDNF) in the antidepressive effect has also been suggested; however, the possible influence of glucocorticoid on the action of BDNF in the developing central nervous system has not been elucidated. In this study, we investigated the effect of glucocorticoid (dexamethasone, DEX) on synaptic maturation and function enhanced by BDNF in early developing hippocampal neurons. In the immature stage, BDNF increased the outgrowth of dendrites and the expression of synaptic proteins including glutamate receptors and presynaptic proteins. Pretreatment with DEX significantly inhibited the BDNF-dependent up-regulation of both dendritic outgrowth and synaptic proteins. In the more mature stage, the BDNF-reinforced postsynaptic Ca(2+) influx was decreased by DEX. BDNF-enhanced presynaptic glutamate release was also suppressed. RU486, a glucocorticoid receptor antagonist, canceled the DEX-dependent blocking effect on the action of BDNF. After down-regulation of glucocorticoid receptor by small interfering RNA application, no inhibitory effect of DEX on the BDNF-increased synaptic proteins was observed. Interestingly, the BDNF-activated MAPK/ERK pathway, which is an essential intracellular signaling pathway for the BDNF-increased synaptic proteins, was reduced by DEX. These results suggest that BDNF-mediated synaptic maturation is disturbed after neurons are exposed to high-level glucocorticoid in their development stage.

Monofocal acute inflammatory demyelination manifesting as open ring sign. Case report.

A 48-year-old woman with a history of viral influenza infection was admitted with rapidly progressive numbness and weakness of the right extremities. On admission, general physical examination revealed no abnormality. Cerebrospinal fluid analysis showed no abnormal findings. Brain computed tomography and magnetic resonance imaging showed an open ring-like enhanced lesion in the white matter of the right parietal lobe with massive perifocal edema. Cerebral angiography showed no tumor staining and thallium-201 single photon emission computed tomography showed no abnormal uptake. The preoperative diagnosis was malignant glioma and partial resection was performed. Histological examination showed perivascular accumulation of small lymphocytes and a large number of macrophages with reactive astrocytes. Phagocytosis of myelin was observed in the macrophages and nuclear fragmentation in the reactive astrocytes. The histological diagnosis was acute inflammatory demyelinating disease. After therapy with methylprednisolone, her neurological symptoms improved gradually and no relapse occurred during 18 months of follow up. Tumor-like masses of demyelination may occupy an intermediate position between acute multiple sclerosis and postinfectious encephalitis. Open ring sign may be a pathognomonic feature of these lesions.

Nerve growth factor attenuates endoplasmic reticulum stress-mediated apoptosis via suppression of caspase-12 activity.

Following endoplasmic reticulum (ER) stress, which occurs via inhibition of the glycosylation of newly synthesized proteins, caspase family proteins are activated to promote ER stress-mediated apoptosis. Here we report that nerve growth factor (NGF) suppressed the ER stress-mediated apoptosis in tunicamycin-treated PC12 cells through an extensive decrease of the caspase-3/-9/-12 activity. Detailed analysis of the mechanism underlying the NGF-mediated cell survival revealed that the activities of all seriate caspases were reduced through the phosphatidylinositol 3-kinase (PI3-K) signaling pathway induced by NGF. Moreover, we found that the activity of c-Jun N-terminal kinase (JNK) was not essential for the tunicamycin-induced apoptosis of PC12 cells. These results demonstrate that the inactivation of caspase-12 via the NGF-mediated PI3-K signaling pathway leads to inactivation of the caspase cascade including caspase-3 and -9.

Prevention of endoplasmic reticulum stress-induced cell death by brain-derived neurotrophic factor in cultured cerebral cortical neurons.

Brain-derived neurotrophic factor (BDNF), one of the neurotrophic factors acting in the central nervous system (CNS), prevents ordinary types of neuronal cell death induced by various stimulants. On the other hand, an accumulation of unfolded proteins in the endoplasmic reticulum (ER) leads to ER stress and then induces ER stress-mediated cell death. The ER stress-mediated cell death is distinctive because the caspase-12 activity plays a crucial role in the progression of cell death. We previously showed that nerve growth factor (NGF) attenuated ER stress-mediated cell death in non-neuronal PC12 cells. Here, we report that BDNF suppressed the ER stress-mediated cell death in tunicamycin (Tm)-treated cerebral cortical neurons. An analysis using a specific inhibitor of phosphatidylinositol 3-kinase (PI3-K), LY294002, revealed that BDNF prevented this cell death via the PI3-K signaling pathway. We found that the number of NeuN/TUNEL-double positive cells and the activity of caspase-3 suppressed by BDNF were increased by LY294002. We also discovered that LY294002 diminished the effect of BDNF on the activation of caspase-12, indicating that BDNF prevents ER stress-mediated cell death via a PI3-K-dependent mechanism by suppressing the activation of caspase-12 in cultured CNS neurons.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine has a transient proliferative effect on PC12h cells and nerve growth factor additively promotes this effect: possible involvement of distinct mechanisms of activation of MAP kinase family proteins.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a potent inducer of cell death, producing reactive oxygen species (ROS) and causing apoptosis in PC12h cells at 1 mM [Shimoke et al., J. Neurosci. Res. 63 (2001) 402-409]. We showed here that MPTP also had a weak proliferative effect on PC12h at 500 microM when treated for 24 h. The proliferative effect was additive within 24 h cells when nerve growth factor (NGF) was present in the culture medium, but NGF promoted cell differentiation 2 or 3 days after. Use of PD98059, a specific inhibitor of MEK1 located upstream of extracellular signal-regulated kinases (ERKs), revealed that the NGF- and MPTP-induced proliferative effect depends on the MEK1 pathway because PD98059 diminished the proliferation completely, and interestingly, NGF and MPTP promoted sustained activation of ERKs. Moreover, we observed that MPTP increased the activity of p38 MAPK but not c-jun N-terminal kinase (JNK) in 30 min. We also observed that SB203580, a specific inhibitor of p38 MAPK, decreased cell viability. These results suggest that NGF and MPTP cooperate to promote acute cell proliferation via the sustained ERKs and the p38 MAPK pathway within 24 h in PC12h cells.

MPTP-induced reactive oxygen species promote cell death through a gradual activation of caspase-3 without expression of GRP78/Bip as a preventive measure against ER stress in PC12 cells.

Glucose-regulated protein 78 (GRP78)/Immunoglobulin binding protein (Bip) is a chaperone which functions to protect cells from endoplasmic reticulum (ER) stress. GRP78/Bip is expressed following ER stress induced by thapsigargin, tunicamycin or chemical factors. However, the mechanism of progression of ER stress against stress factors is still obscure. We examined whether reactive oxygen species (ROS) were involved in GRP78/Bip expression and caspase-3 activity was induced in PC12 cells using 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to produce ROS. We report that PC12 cells lost viability in the presence of MPTP for 24 hours as a partial effect of ROS. We also show that N-acetyl-L-cysteine diminished the MPTP-induced apoptosis with expunction of ROS. Furthermore, we observed that GRP78/Bip was not up-regulated and the caspase-3 activity was increased in the presence of MPTP. These results suggest that insubstantial ROS do not contribute to the ER stress-mediated cell death while caspase-3 is involved in ROS-promoted cell death in MPTP-treated cells.

Actions of insulin-like growth factor binding protein-5 (IGFBP-5) are potentially regulated by tissue kallikrein in rat brains.

Tissue kallikrein (EC 3.4.21.35) that hydrolyzes kininogen and releases a physiologically active peptide, kinin, is found in rat brains. Insulin-like growth factor binding proteins (IGFBPs) that modulate IGF actions are also expressed in a variety of tissues including rat brains, and one of the major IGFBPs expressed in brain is known to be IGFBP-5, which is reported to be hydrolyzed in vitro by prostate-specific antigen (PSA) and gamma-nerve growth factor (gamma-NGF), both of which belong to the member of the kallikrein gene family. This study was designed to determine whether or not kallikrein has a potential to hydrolyze IGFBP-5 and their topographic proximity was investigated in rat brain using double immunohistochemical staining method. Immunohistochemically, IGFBP-5 positive cells were numerous and widespread in the cerebral cortex and belonged to neurons in the cell configuration. IGFBP-5 positive cells were negative for S-100 protein and were positive for betaIII tubulin, confirming them to be neurons. In addition, kallikrein positive cells were virtually all IGFBP-5 positive cells. IGFBP-5 was clearly hydrolyzed by kallikrein with cleavage sites of Arg188-Met189 and Arg136-Arg137 of IGFBP-5. Therefore, there is a possibility that kallikrein plays an important role in brain physiology, specifically in the neurons by regulating the actions of IGFBP-5 and IGF.

Clinical diagnosis of gliomatosis cerebri: report of three cases.

Distinction of gliomatosis cerebri (GC), a rare entity characterized by a widespread infiltration of the brain by tumor, from diffuse glioma is a difficult clinical problem. Most previously reported cases of GC have been autopsy cases because of the lack of objective and quantitative clinical diagnostic criteria. In order to better define this entity, we report the neuroradiological and pathological findings of three cases of GC. Three patients (one man and two women, aged 46-71 years) presented with symptoms of mild increased intracranial pressure, cognitive impairment, or seizure. Magnetic resonance imaging (MRI) was done with T1-weighted images after gadolinium injection, and with T2-weighted images and fluid attenuated inversion recovery (FLAIR) in all cases. Histological confirmation of glial proliferation was obtained in all cases by craniotomy. The topography of the tumoral infiltration was characteristic, involving mainly the white matter, basal ganglia, thalamus, and commissural fibers. More than two cerebral lobes were affected. Contrast enhancement was absent, and mass effects were minimal compared with the extent of tumoral infiltration, but one patient presented with a small frontal enhanced mass during the clinical course. The pathological analyses demonstrated infiltration of the brains by variably differentiated neoplastic glial cells with destruction of the myelin sheath, but the involved axis cylinder and neuronal cells were preserved. Diagnosis of GC should be faithful to the pathological diagnosis criteria of Scheinker and Evans, and therefore the precise assessment of MRI findings according to these criteria is required for clinical, antemortem diagnosis of GC.

Clinicopathological significance of expression of nestin, a neural stem/progenitor cell marker, in human glioma tissue.

The purpose of the study was to investigate the pathological and clinical significance of the expression of nestin, a type-VI intermediate filament transiently expressed during brain development, in glioma tissue. This study was conducted in 70 patients with newly diagnosed adult supratentorial gliomas who underwent multimodality treatment in our department, including surgery. The pathological diagnosis was grade II in 6 patients, grade III in 21 patients, and grade IV in 43 patients. Two specimen sections, one from the bulk of the removed tumor and one from the border between the tumor and normal brain tissue, were subjected to immunostaining with a mouse anti-human nestin monoclonal antibody. Analyses were performed to investigate possible correlation with pathological features, the relationship between nestin expression and the continuity of tumor with the subventricular zone (SVZ), correlation with the therapeutic prognosis, etc. Nestin was expressed specifically in astrocytoma lineage cells. In oligodendroglial tumors, nestin was expressed only in less-differentiated cells and cells suggestive of the presence of astrocytoma. In astrocytic tumors, the rate and level of nestin expression increased as the degree of malignancy increased. There was no significant correlation between the expression level of nestin and the continuity of tumor with the SVZ in the contrast-enhanced imaging before surgery. In addition, no correlation with the therapeutic prognosis was observed. Nestin, a neural stem cell marker, was specifically expressed in astrocytoma lineage cells in glioma tissue. A positive correlation was observed between the degree of malignancy and the level of nestin expression. However, the level of nestin expression was not related to the tumor localization in the SVZ and was not correlated with the therapeutic prognosis.

Cyclophosphamide promotes cell survival via activation of intracellular signaling in cultured cortical neurons.

Cyclophosphamide (CP) has been used as an antitumour agent or immunosuppressant clinically, though the potential biological role of CP in the central nervous system (CNS) has not been clarified. In the present study, we found that pretreatment with CP prevented neuronal cell death caused by serum deprivation in cultured cortical neurons. Interestingly, CP stimulated activation of PI3K (phosphatidylinositol 3 kinase) and MAPK/ERK (mitogen-activated protein kinase/extracellular signal-regulated kinase) pathways, which are known as survival-promoting intracellular signalings. Furthermore, CP increased the expression of Bcl2, an anti-apoptotic factor. In the presence of inhibitors for PI3K or MAPK/ERK pathways, the CP-dependent neuronal survival and Bcl-2 up-regulation were both abolished. Importantly, significant increase in BDNF (brain-derived neurotrophic factor) expression was induced by CP application, implying that BDNF up-regulation is involved in the CP effect. We propose that CP has a protective effect on CNS neurons via the activation of intracellular signalings, and up-regulation of Bcl2 and BDNF.

SA4503, a sigma-1 receptor agonist, prevents cultured cortical neurons from oxidative stress-induced cell death via suppression of MAPK pathway activation and glutamate receptor expression.

Many studies suggest that antidepressants act as neuroprotective agents in the central nervous system (CNS), though the underlying mechanism has not been fully elucidated. In the present study, we examined the effect of SA4503, which is a sigma-1 receptor agonist and a novel antidepressant candidate, on oxidative stress-induced cell death in cultured cortical neurons. Exposure of the neurons to H(2)O(2) induced cell death, while pretreatment with SA4503 inhibited neuronal cell death. The SA4503-dependent survival effect was reversed by co-application with BD1047 (an antagonist of sigma-1/2 receptors). Previously we found that H(2)O(2) triggers a series of events including over-activation of mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) and intracellular Ca(2+) accumulation via voltage-gated Ca(2+) channels and ionotropic glutamate receptors, resulting in neuronal cell death (Numakawa et al. (2007) [20]). Importantly, we found in this study that SA4503 reduced the activation of the MAPK/ERK pathway and down-regulated the ionotropic glutamate receptor, GluR1. Taking these findings together, it is possible that SA4503 blocks neuronal cell death via repressing activation of the MAPK/ERK pathway and, consequently, expression levels of glutamate receptors.

Radiopathological characteristics of cerebellar malignant glioma in adults.

Our aim was to extract the radiopathological features of cerebellar malignant glioma in adults from the four cases we encountered. All four cases (two men and two women, aged 52-80 years; mean age, 67 years) had a floating sensation or vertigo at the onset of their disease. Initially, these patients were given a diagnosis of cerebellar infarction or cavernous angioma, or had faint abnormalities in the cerebellum that were overlooked. These patients were followed up for 2-14 months (mean, 6 months), and the tumor was detected when their clinical symptoms deteriorated. The tumor was located in the hemisphere in one patient and in the vermis in three patients. MRI revealed ring-like enhancement in two patients and nodular enhancement in two patients. All patients underwent subtotal tumor resection, followed by postoperative radiochemotherapy. However, three patients died at 16 to 44 months (mean, 28 months), and cerebrospinal fluid (CSF) dissemination was observed in three patients. Two cases were classified as WHO grade III and two as WHO grade IV. The pathological features were typical of malignant glioma, partially presenting the features of low-grade glioma, such as pilocytic, astrocytic, or oligodendroglial components. Nuclear pleomorphism and vascular endothelial proliferation were prominent, and micronecrosis was relatively less evident. The MIB-1 index was 12%-40%, and most of the patients were p53 protein positive. At the onset of cerebellar malignant glioma, diagnostic imaging shows few signs of brain tumor. Thereafter, tumors grow in a short period of time, following a rapid clinical course. Because cerebellar malignant glioma is a rare disease, it is difficult to differentiate it from metastatic brain tumors and primary central nervous system lymphoma by preoperative imaging. Some patho logical features suggesting malignant transformation from low-grade glioma were detected.

Down-regulation of miR-92 in human plasma is a novel marker for acute leukemia patients.

BACKGROUND: MicroRNAs are a family of 19- to 25-nucleotides noncoding small RNAs that primarily function as gene regulators. Aberrant microRNA expression has been described for several human malignancies, and this new class of small regulatory RNAs has both oncogenic and tumor suppressor functions. Despite this knowledge, there is little information regarding microRNAs in plasma especially because microRNAs in plasma, if exist, were thought to be digested by RNase. Recent studies, however, have revealed that microRNAs exist and escape digestion in plasma. METHODOLOGY/PRINCIPAL FINDINGS: We performed microRNA microaray to obtain insight into microRNA deregulation in the plasma of a leukemia patient. We have revealed that microRNA-638 (miR-638) is stably present in human plasmas, and microRNA-92a (miR-92a) dramatically decreased in the plasmas of acute leukemia patients. Especially, the ratio of miR-92a/miR-638 in plasma was very useful for distinguishing leukemia patients from healthy body. CONCLUSIONS/SIGNIFICANCE: The ratio of miR-92a/miR-638 in plasma has strong potential for clinical application as a novel biomarker for detection of leukemia.

Interleukin-27 directly induces differentiation in hematopoietic stem cells.

Interleukin (IL)-27, one of the most recently discovered IL-6 family cytokines, activates both the signal transducer and activator of transcription (STAT)1 and STAT3, and plays multiple roles in pro- and anti-inflammatory immune responses. IL-27 acts on various types of cells including T, B, and macrophage through the common signal-transducing receptor gp130 and its specific receptor WSX-1, but the effect of IL-27 on hematopoietic stem cells (HSCs) remains unknown. Here, we show that IL-27 together with stem cell factor (SCF) directly acts on HSCs and supports their early differentiation in vitro and in vivo. CD34(-/low)c-Kit(+)Sca-1(+)lineage marker(-) (CD34(-)KSL) cells, a population highly enriched in mouse HSCs, were found to express both IL-27 receptor subunits. In vitro cultures of CD34(-)KSL cells with IL-27 and SCF resulted in an expansion of progenitors including short-term repopulating cells, while some of their long-term repopulating activity also was maintained. To examine its in vivo effect, transgenic mice expressing IL-27 were generated. These mice exhibited enhanced myelopoiesis and impaired B lymphopoiesis in the bone marrow with extramedullary hematopoiesis in the spleen. Moreover, IL-27 similarly acted on human CD34(+) cells. These results suggest that IL-27 is one of the limited cytokines that play a role in HSC regulation.