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RB1gene is often homozygously deleted or mutated in prostate adenocarcinomas following acquirement of castration resistance and/or metastatic ability. We found that SUCLA2gene is frequently involved in the deletion of the RB1gene region in advanced prostate cancer. SUCLA2 constitutes the β-subunit of succinate CoA ligase heterodimer that reversibly converts succinyl CoA into succinate. We sought the possibility that deletion of SUCLA2gives rise to a metabolic vulnerability that could be targeted therapeutically. We found a significant metabolic shift in SUCLA2-deleted prostate cancer cells, including lower mitochondrial respiratory activity. By screening a number of libraries for compounds that induce cell death selectively in SUCLA2-deficient prostate cancer cells, we identified thymoquinone (2-isopropyl-5-methylbenzo-1,4-quinone) and PMA (phorbol-12-myristate-13-acetate) from a natural compound library. These findings indicate that the metabolic vulnerability in SUCLA2-deficient prostate cancer cells is pharmacologically targetable.

Have you ever imagined reactions of alkenes with hydrogen that result in anything other than hydrogenation or hydrogenative C–C coupling? We have long sought to develop not only hydrogenation catalysts that activate H2as hydride ions but also electron transfer catalysts that activate H2as a direct electron donor. Here, we report the reductive cyclopropanation of alkenes using an iridium electron storage catalyst with H2as the electron source without releasing metal waste from the reductant. We discuss the catalytic mechanism with selectivity to give the trans-isomer. These findings are based on the isolation of three complexes and density functional theory calculations.

Tumor recurrence is attributable to cancer stem-like cells (CSCs), the metabolic mechanisms of which currently remain obscure. Here, we uncovered the critical role of folate-mediated one-carbon (1C) metabolism involving mitochondrial methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) and its downstream purine synthesis pathway. MTHFD2knockdown greatly reduced tumorigenesis and stem-like properties, which were associated with purine nucleotide deficiency, and caused marked accumulation of 5-aminoimidazole carboxamide ribonucleotide (AICAR)—the final intermediate of the purine synthesis pathway. Lung cancer cells with acquired resistance to the targeted drug gefitinib, caused by elevated expression of components of the β-catenin pathway, exhibited increased stem-like properties and enhanced expression of MTHFD2. MTHFD2knockdown or treatment with AICAR reduced the stem-like properties and restored gefitinib sensitivity in these gefitinib-resistant cancer cells. Moreover, overexpression of MTHFD2 in gefitinib-sensitive lung cancer cells conferred resistance to gefitinib. Thus, MTHFD2-mediated mitochondrial 1C metabolism appears critical for cancer stem-like properties and resistance to drugs including gefitinib through consumption of AICAR, leading to depletion of the intracellular pool of AICAR. Because CSCs are dependent on MTHFD2, therapies targeting MTHFD2 may eradicate tumors and prevent recurrence.

Supplemental Digital Content is available in the text.Liver fibrosis is one of the common complications of transient myeloproliferative disorder (TMD) in Down syndrome (DS), but the exact molecular pathogenesis is largely unknown. We herein report a neonate of DS with liver fibrosis associated with TMD, in which we performed the serial profibrogenic cytokines analyses. We found the active monocyte chemoattractant protein-1 expression in the affected liver tissue and also found that both serum and urinary monocyte chemoattractant protein-1 concentrations are noninvasive biomarkers of liver fibrosis. We also showed a prospective of the future anticytokine therapy with herbal medicine for the liver fibrosis associated with TMD in DS.

Retinoblastoma tumor suppressor protein (RB) is inactivated more frequently during tumor progression than during tumor initiation. However, its exact role in controlling the malignant features associated with tumor progression is poorly understood. We established in vivo and in vitro models to investigate the undifferentiated state induced by Rb inactivation. Rbheterozygous mice develop well‐differentiated thyroid medullary carcinoma. We found that additional deletion of Trp53, without change in lineage, converted these Rb‐deficient tumors to a poorly differentiated type associated with higher self‐renewal activity. Freshly prepared mouse embryonic fibroblasts (MEFs) of Rb−/−; Trp53−/−background formed stem cell‐like spheres that expressed significant levels of embryonic genes despite of lacking the ability to form colonies on soft agar or tumors in immune‐deficient mice. This suggested that Rb‐p53 double inactivation resulted in an undifferentiated status but without carcinogenic conversion. We next established Rb−/−; N‐ras−/−MEFs that harbored a spontaneous carcinogenic mutation in Trp53. These cells (RN6), in an Rb‐dependent manner, efficiently generated spheres that expressed very high levels of embryonic genes, and appeared to be carcinogenic. We then screened an FDA‐approved drug library to search for agents that suppressed the spherogenic activity of RN6 cells. Data revealed that RN6 cells were sensitive to specific agents including ones those are effective against cancer stem cells. Taken together, all these findings suggest that the genetic interaction between Rb and p53 is a critical determinant of the undifferentiated state in normal and tumor cells. StemCells2015;33:1657–1669

Reactive oxygen species (ROS) activate uncoupler protein 1 (UCP1) in brown adipose tissue (BAT) under physiological cold exposure and noradrenaline (NA) stimulation to increase thermogenesis. However, the endogenous regulator of ROS in activated BAT and its role in pathological conditions remain unclear. We show that serum levels of selenoprotein P (SeP; encoded by SELENOP) negatively correlate with BAT activity in humans. Physiological cold exposure downregulates Selenopin BAT. Selenopknockout mice show higher rectal temperatures and UCP1 sulfenylation during cold exposure. SeP treatment to brown adipocytes eliminated the NA-induced mitochondrial ROS by upregulating glutathione peroxidase 4 and impaired cellular thermogenesis. A high-fat/high-sucrose diet elevates serum SeP levels and diminishes the elevated NA-induced thermogenesis in BAT-SelenopKO mice. Therefore, SeP is the intrinsic factor inducing reductive stress that impairs thermogenesis in BAT and may be a potential therapeutic target for obesity and diabetes.

Cerebrovascular white matter (WM) lesions contribute to cognitive impairment and motor dysfunction in the elderly. A disruption of the blood–brain barrier (BBB) is believed to be a critical early event leading to these WM lesions. Previous studies have suggested the involvement of matrix metalloproteinase-2 (MMP-2) in BBB disruptions and the upregulation of MMP-2 after chronic cerebral hypoperfusion in a rat model. In the present study, we asked whether MMP-2 is involved in the BBB disruption and the subsequent WM lesions after chronic cerebral hypoperfusion.

RECKwas first isolated as a transformation suppressor gene by cDNA expression cloning in a mouse fibroblast cell line transformed by an activated RASoncogene. Subsequently, reduced expression of RECKin transformed cells and cancer cells were demonstrated. Moreover, in several types of tumors, positive correlation between RECK expression and survival of patients have been noted. RECKencodes a GPI-anchored glycoprotein harboring three protease inhibitor-like domains. The RECK protein regulates at least three members of the matrix metalloproteinase (MMP) family, MMP-2, MMP-9, and MT1-MMP, in vitroor in cultured cells. Restored expression of RECKin cancer cell lines results in strong suppression of invasion, metastasis, and tumor angiogenesis. Mice lacking RECKdie in uterowith reduced integrity of blood vessels, the neural tube, and mesenchymal tissues. In these mice, MMP activity is elevated, and the amount of collagen type I greatly reduced. The RECKnull phenotype is partially rescued (half day delay of death and marked recovery of tissue integrity) by MMP-2null mutation, demonstrating functional interaction between RECK and MMP-2 in vivoand involvement of other target(s) for RECK in the lethal phenotype. These findings indicate that (i) RECK is an important regulator of extracellular matrix remodeling and that (ii) down-regulation of RECK by oncogenic signaling leads to the excessive activation of MMPs thereby promoting malignant behavior of cancer cells such as invasion, metastasis, and angiogenesis.

A human placenta cDNA expression library was screened for genes inducing flat reversion when transfected into a v‐K‐ras‐transformed NIH3T3 cell line, DT. One such gene was found to encode a Kunitz‐type serine protease inhibitor, tissue factor pathway inhibitor‐2 (TFPI‐2). While the TFPI‐2mRNA can be detected in normal human fibroblasts (MRC‐5), it is down‐regulated in MRC‐5 cells expressing an activated H‐rasoncogene and in the human fibrosarcoma cell line, HT1080. Restored expression of the TFPI‐2gene in HT1080 cells resulted in the suppression of matrix invasion activity in vitro with concomitant decrease in the relative amount of active matrix metalloproteinase‐2 secreted from the cells. When DT cells were cultured in the presence of conditioned medium and extracellular matrix prepared from TFPI‐2‐transfected HT1080 cells, increased attachment and flat reversion were observed. These results suggest that TFPI‐2 may be required for the maintenance of the integrity of extracellular matrix in normal tissues and its down‐regulation as a result of oncogene activation may contribute to the malignant phenotypes of tumor cells.

We have isolated and characterized the 5′-flanking region of the mouse RECKgene aiming to understand the mechanism of oncogene-mediated suppression of RECKgene expression. The upstream 52-base region was found to contain a promoter activity which is, to some extent, suppressed by the rasoncogene. This region contains two Sp1-binding motifs, one cEBPb-binding motif, and one CAAT box. Although both of the Sp1 sites were found to associate with Sp1 as well as Sp3 proteins, rasresponsiveness seems to be mediated only by the downstream Sp1 site. Our data indicate that the Sp1 motif in certain contexts can serve as a negative target for the Ras signal.

1Recombinant ATP‐sensitive K+channels (KATPchannels) were heterologously expressed in the NIH3T3 mouse cell line, and the electrophysiological properties were studied using patch‐clamp techniques.2The NIH3T3 cell lines transfected with the inwardly rectifying K+channel Kir6.1 alone or with both Kir6.1 and cystic fibrosis transmembrane conductance regulator (CFTR) exhibited time‐independent K+currents with weak inward rectification. In contrast, no measurable K+conductance was observed in mock‐transfected cells or in cells transfected with CFTR alone. Regardless of co‐transfection with Kir6.1, the transfection with CFTR produced a Cl−conductance that was activated by cell dialysis with cAMP (1 mM). The conductance was reversibly suppressed by glibenclamide (30 μM).3Whole‐cell currents at +60 mV were blocked in a concentration‐dependent manner by Ba2+ions with similar IC50values: 89.3 ± 23.3 μM (Kir6.1 alone) and 67.3 ± 24.9 μM (Kir6.1‐CFTR).4The currents recorded from Kir6.1‐transfected cells were not affected by glibenclamide, whereas glibenclamide did inhibit the conductance expressed in cells co‐transfected with CFTR (IC50= 35.9 ± 6.6 μM).5In the cell‐attached mode with a 150 mM K+pipette solution, both Kir6.1‐ and Kir6.1‐CFTR‐transfected cells displayed a class of K+channels showing weak inward rectification and a slope conductance of 50.7 ± 1.0 and 52.4 ± 4.9 pS, respectively.6In the inside‐out mode, the single‐channel currents recorded from both types of cells were not inhibited by intracellular ATP (1 mM). However, glibenclamide was found to block the single‐channel activities in the co‐transfected cells.

GDE2 controls surface amounts of the metalloprotease inhibitor RECK to promote ADAM10 α-secretase activity and prevent Aβ production and synapse loss.

Infarction located in the midbrain and pons presents various ophthalmic symptoms, because of the damage of the nuclei that control the movement of internal and external ocular and palpebral muscles. We experienced a case which presented with rare ocular symptoms and course. A 61-year-old man presented with left hemiparesis and dysarthria, bilateral ptosis, and bilateral impaired eyeball movement: right eyeball movement was totally impaired and left could only perform slight adduction. MRI showed fresh stroke in the right thalamus, cerebral crus, and posterior lobe and cuneate lesion on bilateral paramedian portion of the midbrain. MRA showed occlusion in the P1 area of the posterior cerebral artery (PCA). Transesophageal echocardiography (TEE) showed findings of a patent foramen ovale (PFO). These findings suggested cardioembolic stroke as a cause of PCA occlusion and we prescribed rivaroxaban. The patient’s eyeball and eyelid movement, only on the left side, was improved imperfectly 2 weeks later. We thought that neurological findings and course of this case may have arisen from dysfunction of the oculomotor nucleus and oculomotor fascicles, and MLF results from the presence of the lesion in paramedian midbrain and pons.

We recently experienced two patients with stroke-like symptoms and ascending aortic dissection (AAD) in our outpatient department. Both patients were transferred to our hospital presenting with neurological deficit such as hemiparesis and conjugate deviation. They did not complain from any chest or abdominal pain. Their MRI did not show fresh infarction or main branch occlusion. A chest CT image showed AAD. The former patient was immediately transferred to a tertiary hospital and the latter received conservative management in the cardiovascular department. Discussion. As neither patient was experiencing any pain, we initially diagnosed them with ischemic stroke and began treatment. Fortunately, bleeding complications did not occur. In such cases, problems are caused when intravenous tissue plasminogen activator (t-PA) injection is administered with the aim of reopening the occluded intracranial arteries. In fact, patients with AAD undergoing t-PA injection have been reported to die from bleeding complications without any recognition of the dissection. These findings suggest that confirmation using carotid ultrasound, carotid MR angiography, and a D-dimer test is crucial and should be adopted in emergency departments.