Kenji Kasai

Primary lymphoma of the heart: Case report and literature review

Primary cardiac lymphoma (PCL) is a rare and usually fatal neoplasma. A case of PCL in a 78‐year‐old man who complained of exertional dyspnea and peripheral edema is presented. Echocardiography revealed a mass in the right atrium and a diagnosis of low‐grade B‐cell lymphoma was obtained with the surgically resected tumor. The lesion appeared to have originated in the right atrium and involved the right ventricle. The patient died of bronchopneumonia 8 months after the initial consultation. The present case and 39 patients with PCL reported between 1995 and 2002 were reviewed. Forty patients showed various and non‐specific symptoms such as dyspnea, edema, arrhythmia and pericardial effusion. Primary cardiac lymphoma occurred slightly more often in male patients (M : F = 23:17) and in the elderly in general (mean age, 67 years), with lesions found in the following locations, listed in order of frequency: right atrium, pericardium, right ventricle, left atrium, left ventricle, and other sites. Antemortem diagnosis was obtained in 37 of the 40 patients. Thirty‐seven cases were of B‐cell lineage and two cases were of T‐cell lineage. Complete remission was obtained in only 15 of the 40 patients. Although PCL antemortem diagnoses have been made in the majority of recent cases, the prognosis still remains poor.

The G12 family of heterotrimeric G proteins and Rho GTPase mediate Sonic hedgehog signalling

Sonic hedgehog (Shh) is a secreted morphogen crucial for cell fate decision, cellular proliferation, and patterning during vertebrate development. The intracellular Shh signalling is transduced by Smoothened (Smo), a seven‐transmembrane spanning protein that belongs to the G‐protein coupled receptor family. Among four families of Gα subunits, Gαi has been thought to be responsible for transducing Shh signalling, while several lines of evidence indicated that other signalling pathways may be involved. We found that the G12 family of heterotrimeric G proteins and the small GTPase RhoA are involved in Shh/Smo‐mediated cellular responses, including stimulation of target gene promoter and inhibition of neurite outgrowth of neuroblastoma cells. We also found that the G12/RhoA pathway is responsible for Smo‐induced nuclear import of GLI3 which is thought to transduce Shh signals to nucleus. Furthermore, misexpression of a G12‐specific GTPase‐activating protein in rat neural tubes leads to pertubation of motor neurone and interneurone development, mimicking the effects of decreased Shh signalling. These results show that Shh signalling is mediated in part by activating G12 family coupled signalling pathways. The participation of RhoA, a pivotal molecular switch in many signal transduction pathways, may help explain how Shh can trigger a variety of cellular responses.

SCL/TAL1 Interrupting Locus Derepresses GLI1 from the Negative Control of Suppressor-of-Fused in Pancreatic Cancer Cell

As a physically binding protein of GLI1 transcription factor, Suppressor-of-Fused (SUFU) has been placed in the center of negative regulation of Hedgehog (Hh) signaling. SUFU tethers GLI1 in cytoplasm, and in some circumstances, it moves into the nucleus in association with GLI1, leading to the suppression of GLI1 target gene expression by recruiting a corepressor complex. The activated transcriptional function of GLI1 is important for cellular proliferation in a variety of human cancers. However, it has not been revealed how GLI1 is derepressed from SUFU-mediated suppression. Here, we show SCL/TAL1 interrupting locus (SIL) product, a cytoplasmic protein overexpressed in pancreatic ductal adenocarcinoma (PDA), is responsible for the derepression of GLI1. We found SIL associated with the carboxyl terminus of SUFU, one of two distinct GLI1-binding domains, and this association was responsible for cytoplasmic tethering of SUFU. Overexpressed SIL attenuated SUFU-mediated cytoplasmic tethering and target gene suppression of GLI1. Knockdown of SIL in PDA cells conversely induced the nuclear accumulation of SUFU in association with GLI1 and the transcriptional suppression of GLI1 target genes. Importantly, we also showed that oncogenic K-RAS, and not Sonic hedgehog, enhanced the SIL association with the amino-terminus of SUFU, the other GLI1-binding domain that led to further increase of nuclear translocation of GLI1. These results uncover the role of SIL in derepressing GLI1 from the negative control of SUFU, which is a crucial step for activating Hh signaling in cancer cells.

Rat Small Intestinal Goblet Cell Kinetics in the Process of Restitution of Surface Epithelium Subjected to Ischemia-Reperfusion Injury

Repair of superficial damage to gastrointestinal mucosa occurs by a process called restitution. Goblet cells reside throughout the length of the intestine and are responsible for the production of mucus. However, a kinetic analysis of goblet cell dynamics of small intestine in restitution has hitherto not been reported. The aim of the present study was to investigate the role of goblet cells in the process of restitution of rat small intestine subjected to ischemia and ischemia–reperfusion injury, and therefore intestinal epithelium from rats subjected to both ischemia and ischemia–reperfusion was studied. Detachment of enterocytes was observed after 5-min of reperfusion. After 20–30 minutes of reperfusion, the denuded villous tips were covered with goblet cells. Within 75 min of reperfusion the epithelium restitution was complete. On the other hand, restitution was not observed in ischemia group. These data suggest that goblet cells may play an important role in restitution after ischemia–reperfusion injury.

GLI1 Interferes with the DNA Mismatch Repair System in Pancreatic Cancer through BHLHE41-Mediated Suppression of MLH1

The mismatch repair (MMR) system is indispensable for the fidelity of DNA replication, the impairment of which predisposes to the development and progression of many types of cancers. To date, GLI1 transcription factor, a key molecule of the Hedgehog signaling pathway, has been shown to regulate the expression of several genes crucial for a variety of cancer cell properties in many types of cancers, including pancreatic ductal adenocarcinoma (PDAC), but whether GLI1 could control the MMR system was not known. Here, we showed that GLI1 and GLI2 indirectly suppressed the expression of MLH1 in PDAC cells. Through GLI1 target gene screening, we found that GLI1 and GLI2 activated the expression of a basic helix-loop-helix type suppressor BHLHE41/DEC2/SHARP1 through a GLI-binding site in the promoter. Consistent with a previous report that BHLHE41 suppresses the MLH1 promoter activity, we found that the activation of GLI1 led to the BHLHE41-dependent suppression of MLH1, and a double knockdown of GLI1 and GLI2 conversely increased the MLH1 protein in PDAC cells. Using TALEN-based modification of the MLH1 gene, we further showed that GLI1 expression was indeed associated with an increased tolerance to a methylating agent, methylnitrosourea cooperatively with a lower copy number status of MLH1. Finally, GLI1 expression was immunohistochemically related positively with BHLHE41 and inversely with MLH1 in PDAC cells and precancerous lesions of the pancreas. On the basis of these results, we propose that GLI1 depresses the MMR activity and might contribute to the development and progression of PDAC.

Paracrine factors of vascular endothelial cells facilitate cardiomyocyte differentiation of mouse embryonic stem cells

For myocardial regeneration therapy, the low differentiation capability of functional cardiomyocytes sufficient to replace the damaged myocardial tissue is one of the major difficulties. Using Nkx2.5-GFP knock-in ES cells, we show a new efficient method to obtain cardiomyocytes from embryonic stem (ES) cells. The proportion of GFP-positive cells was significantly increased when ES cells were cultured with a conditioned medium from aortic endothelial cells (ECs), accompanied by upregulation of cardiac-specific genes as well as other mesodermal genes. The promotion was more prominent when EC-conditioned medium was added at an early stage of ES cell differentiation culture (Day 0-3). Inhibitors of bone morphogenic protein (BMP), cyclooxygenase (COX), and nitric oxide synthetase (NO) prevented the promotion of cardiomyogenesis by EC-conditioned medium. These results suggest that supplementation of EC-conditioned medium enables cardiomyocytes to be obtained efficiently through promotion of mesoderm induction, which is regulated by BMP, COX, and NOS.

Diversity of Restitution After Deoxycholic Acid-Induced Small Intestinal Mucosal Injury in the Rat

Repair of superficial damage to gastrointestinal mucosa occurs by a process called restitution, with epithelial integrity and continuity reestablished before cell proliferation occurs. The aim of the present study was to investigate the diversity of restitution in rat jejunum exposed to different concentrations of deoxycholic acid (DOC; 1.5–100 mmol/liter). Following a 30-min exposure, the intestine was allowed to recover for 15–330 minutes. DOC caused dose-dependent tissue destruction. Exfoliating epithelial cells were already observed after 5 min of exposure (1.5 mmol/liter), with simple sloughing off and resealing of the tips. Moderately affected epithelium (20 mmol/liter) demonstrated denudation of villous tips and then became covered with goblet cells. Severely affected epithelium (100 mmol/liter) also appeared to be replaced with goblet cells. These data suggest that the reversibility of mucosal damage induced by DOC is due to a variety of processes, which depend on the severity of the mucosal insult.

Addiction of pancreatic cancer cells to zinc-finger transcription factor ZIC2

Activity of GLI transcription factors of Hedgehog signaling is key for various cancer cell properties, especially in pancreatic ductal adenocarcinoma (PDAC). Zinc-finger transcriptional regulators ZIC1 to ZIC5 of ZIC gene family were demonstrated to associate with GLI to increase the nuclear accumulation and transcriptional activity of GLI. Notwithstanding this supportive role for GLI-dependent transcription, it was not fully understood whether ZIC plays an independent role in cancer cell biology. Here, we found that ZIC2 is indispensable in the regulation of PDAC cell apoptosis. We found that human PDAC cell lines uniquely express ZIC2. ZIC2 knockdown induced PDAC cell apoptosis; conversely, ZIC2 over-expression enhanced the cellular proliferation. Through a comprehensive screening, we identified fibroblast growth factor receptor 3 (FGFR3) and ANNEXIN A8 (ANXA8) as genes up-regulated by ZIC2 in PDAC cells. The forced expression of these two genes cooperatively rescued the apoptosis of ZIC2-knockdown cells. Immunohistochemical analyses further supported the correlation of ZIC2 expression and these genes in human pancreata harboring PDAC. Intriguingly, the ZIC2-mediated up-regulation of FGFR3 and ANXA8 was indicated to be GLI -independent. This evidence highlights the indispensable role of ZIC2 in regulating cellular proliferation and apoptosis during PDAC development and suggests a potential therapeutic target for PDAC.

GLI1 orchestrates CXCR4/CXCR7 signaling to enhance migration and metastasis of breast cancer cells

The up-regulation of chemokine receptors CXCR4 and CXCR7 impacts on the distant metastasis and prognosis of breast cancer, though knowledge about the regulatory mechanism of their expressions is limited. Meanwhile, the GLI transcription factors of Hedgehog signaling have been reported to play a pivotal role in the development and progression of many types of human cancer. In breast cancer, the increased expression of GLI1 correlated with metastasis and unfavorable overall prognosis, though its molecular mechanism is also not fully understood. Based on our findings that GLI1 enhanced the lung metastasis of breast cancer cells in a mouse model system, we comprehensively screened for genes up-regulated by GLI1 in breast cancer cells, and as such identified CXCR4, CXCR7/ACKR3, and actin-binding protein LCP1/L-PLASTIN, all of which have been reported to be involved in CXCL12-stimulating signaling. In breast cancer cells, we found that GLI1 and GLI2 up-regulated these expressions, while treatment with GLI-specific inhibitor GANT61 reduced the expressions. As for CXCR4, we confirmed it as a direct target of GLI1 through the reporter assay and the chromatin immunoprecipitation assay. We also found that GLI1 enhanced CXCL12-induced ERK phosphorylation and cell migration, both of which were blocked by either CXCR4-specific inhibitor or knockdown of CXCR7 or LCP1. These evidences suggest an indispensable role of GLI1 in the migration and metastasis of breast cancer cells through CXCL12/CXCR4 signaling enhancement.

GLI1 Modulates EMT in Pancreatic Cancer—Letter

Joost and colleagues reported that GLI1 directly transactivates E-cadherin expression and modulates the TGFβ-induced epithelial–mesenchymal transition in pancreatic cancer cells ([1][1]). This conclusion was mainly drawn from the observation that siRNA-mediated knockdown of GLI1 and GLI2