Tadahiko Enoki

DNA damage signaling is activated during cancer progression in human colorectal carcinoma

Purpose: Recent studies have shown that the DNA damage response (DDR) is activated in precancerous lesions, suggesting that neoplastic cells may avoid apoptosis by impairing the DDR which acts as a barrier against tumor progression. To define the role of the DDR pathway in human colorectal carcinoma, we investigated the level of phosphorylated proteins of the DDR pathway. Experimental Design: Colorectal tissue samples were obtained at the time of surgery, from 55 patients at two hospitals. The tissues were classified into four groups according to pathology: normal mucosa, adenoma, early carcinoma and advanced carcinoma. We evaluated phosphorylated ataxia telangiectasia mutated (pATM), phosphorylated H2AX (γH2AX) and Chk2 (pChk2) protein levels by immunohistochemistry and Western blot analysis. We also evaluated apoptosis by the TUNEL assay. Results: Immunostaining for pATM, γH2AX and pChk2 revealed that all were significantly expressed during tumor progression in advanced carcinoma (vs. normal tissue for pATM [p

Operative injury accelerates tumor growth by inducing mobilization and recruitment of bone marrow-derived stem cells.

BACKGROUND Although operative injury is thought generally to worsen the prognosis of cancer patients, the relevant mechanisms are not yet understood fully. We tested the hypothesis that operative injury induces mobilization and recruitment of bone marrow stem cells, thereby enhancing angiogenesis and accelerating tumor growth. METHODS Mice were subjected to an open gastrotomy, and naïve mice were used as controls. The mobilization of bone marrow stem cells was monitored after operation. Using an established tumor model in green fluorescent protein (GFP)(+) bone marrow-transplanted chimera mice, we investigated further whether the mobilized stem cells affected tumor growth. RESULTS Compared with the control, gastrotomy increased the populations of CD34(+) cells (6.9 ± 4.5 % vs 3.3 ± 0.4%, P < .05) and CD34(+)/Flk-1(+) cells (0.08 ± 0.02% vs 0.05 ± 0.01%, P < .05) in peripheral blood 12 h after operation. Twelve days after operation, the tumor volume almost doubled in mice after gastrotomy compared with control (580 ± 106 mm(3) vs 299 ± 162 mm(3), P < .05). A histologic analysis of tumor tissue revealed that the microvessel density and number of proliferating cells were significantly greater, but those of apoptotic cells were significantly less, in mice after gastrotomy as compared with control. Furthermore, the number of GFP(+) cells found in tumor tissue was significantly greater in mice that underwent gastrotomy than in controls. Some of the stained GFP(+) cells were positive for CD34 and had been incorporated into microvessels. Administration of AMD3100, which is an antagonist of stromal-cell-derived factor (SDF)-1/CXCR4 signaling pathway, inhibited the recruitment of GFP(+) cells and negated completely the acceleration in tumor growth after operation (345 ± 172 mm(3), P < .05). CONCLUSION Operative injury may induce the mobilization and recruitment of bone marrow stem cells, thereby enhancing angiogenesis and accelerating tumor growth. Inhibition of the SDF-1/CXCR4 signals may represent a new therapeutic strategy for preventing acceleration of tumor growth after operation.

The c‐MYC‐ABCB5 axis plays a pivotal role in 5‐fluorouracil resistance in human colon cancer cells

c‐MYC overexpression is frequently observed in various cancers including colon cancer and regulates many biological activities such as aberrant cell proliferation, apoptosis, genomic instability, immortalization and drug resistance. However, the mechanism by which c‐MYC confers drug resistance remains to be fully elucidated. In this study, we found that the c‐MYC expression level in primary colorectal cancer tissues correlated with the recurrence rate following 5‐fluorouracil (5‐FU)‐based adjuvant chemotherapy. Supporting this finding, overexpression of exogenous c‐MYC increased the survival rate following 5‐FU treatment in human colon cancer cells, and knockdown of endogenous c‐MYC decreased it. Furthermore, c‐MYC knockdown decreased the expression level of ABCB5, which is involved in 5‐FU resistance. Using a chromatin immunoprecipitation assay, we found that c‐MYC bound to the ABCB5 promoter region. c‐MYC inhibitor (10058‐F4) treatment inhibited c‐MYC binding to the ABCB5 promoter, leading to a decrease in ABCB5 expression level. ABCB5 knockdown decreased the survival rate following 5‐FU treatment as expected, and the ABCB5 expression level was increased in 5‐FU‐resistant human colon cancer cells. Finally, using a human colon cancer xenograft murine model, we found that the combined 5‐FU and 10058‐F4 treatment significantly decreased tumorigenicity in nude mice compared with 5‐FU or 10058‐F4 treatment alone. 10058‐F4 treatment decreased the ABCB5 expression level in the presence or absence of 5‐FU. In contrast, 5‐FU treatment alone increased the ABCB5 expression level. Taken together, these results suggest that c‐MYC confers resistance to 5‐FU through regulating ABCB5 expression in human colon cancer cells.

JAB1 regulates unphosphorylated STAT3 DNA-binding activity through protein–protein interaction in human colon cancer cells

Recent studies have revealed that unphosphorylated STAT3 forms a dimer, translocates to the nucleus, binds to the STAT3 binding site, and activates the transcription of STAT3 target genes, thereby playing an important role in oncogenesis in addition to phosphorylated STAT3. Among signaling steps of unphosphorylated STAT3, nuclear translocation and target DNA-binding are the critical steps for its activation. Therefore, elucidating the regulatory mechanism of these signaling steps of unphosphorylated STAT3 is a potential step in the discovery of a novel cancer drug. However, the mechanism of unphosphorylated STAT3 binding to the promoter of target genes remains unclear. In this study, we focused on Jun activation domain-binding protein 1 (JAB1) as a candidate protein that regulates unphosphorylated STAT3 DNA-binding activity. Initially, we observed that both unphosphorylated STAT3 and JAB1 existed in the nucleus of human colon cancer cell line COLO205 at the basal state (no cytokine stimulation). On the other hand, phosphorylated STAT3 did not exist in the nucleus of COLO205 cells at the basal state. Immunoprecipitation using nuclear extract of COLO205 cells revealed that JAB1 interacted with unphosphorylated STAT3. To investigate the effect of JAB1 on unphosphorylated STAT3 activity, RNAi studies were performed. Although JAB1 knockdown tended to increase nuclear STAT3 expression, it significantly decreased unphosphorylated STAT3 DNA-binding activity. Subsequently, JAB1 knockdown significantly decreased the expression levels of MDR1, NANOG, and VEGF, which are STAT3 target genes. Furthermore, the expression level of nuclear JAB1, but not nuclear STAT3, correlated with unphosphorylated STAT3 DNA-binding activity between COLO205 and LoVo cells. Taken together, these results suggest that nuclear JAB1 positively regulates unphosphorylated STAT3 DNA-binding activity through protein-protein interaction in human colon cancer cell line COLO205.

Combined right hepatic and retrohepatic caval resection with reconstruction using a polytetrafluoroethylene graft for primary leiomyosarcoma of the liver: Report of case

We present herein the case of a 53-year-old woman who underwent successful surgical treatment for a leiomyosarcoma of the liver that originated from the posterior hepatic segment and involved the retrohepatic inferior vena cava (IVC). A computed tomographic scan and magnetic resonance imaging demonstrated a large tumor, with rich vascularity, in the liver. The IVC was found to be occluded on these scans, which was confirmed by venacavography. The patient underwent a combined right hepatic and caval resection with reconstruction using an expanded polytetra-fluoroethylene graft. The tumor consisted of spindle-shaped cells with cigar-shaped nuclei. It also had a moderate degree of cellularity and ten mitotic figures per ten high-power fields. Immunohistologically, desmin and alpha-smooth muscle actin were stained positive in the tumor cells, implying that the tumor was derived from smooth muscle cells. The patient is alive and well 15 months after her operation.

Long-term survival of xenogeneic heart grafts achieved by costimulatory blockade and transient mixed chimerism.

Background. Xenotransplantation holds great promise in clinical medicine, but is limited by the vigorous rejection response elicited against solid organs transplanted across species barriers. In this study, we investigated the role of anti-CD40L monoclonal antibody (mAb) in inducing xenogeneic mixed chimerism and donor-specific heart transplantation tolerance. Methods. One day before heart transplantation, mice were injected intraperitoneally with anti-mouse CD8/NK1.1/Thy1.2 mAbs. On day 0, the mice received 3 Gy total body irradiation (TBI), an intravenous injection of unseparated bone marrow (BM) harvested from F344 rats, and an intraperitoneal injection of hamster antimouse CD40L mAb, MR1. Heart grafts from F344 rats were heterotopically transplanted into the abdomen of B6 mouse recipients. Using flow cytometric analysis of peripheral white blood cells, we assessed donor hematopoiesis at various times after bone marrow transplantation (BMT). Results. Chimerism subsided gradually and disappeared completely 18 weeks after BMT. The cardiac graft survived permanently, even after the mixed chimerism disappeared. To determine if the mice acquired donor-specific tolerance, second rat heart grafts were transplanted 120 days after the first heart transplantation. The second transplanted hearts were also accepted over 60 days. Histological analysis revealed no remarkable vasculopathy in the coronary vessels at any stage. Conclusions. These findings clearly show that costimulatory blockade plays an important role in inducing xenochimerism, and that transient mixed chimerism can induce permanent acceptance of rat to mouse cardiac xenografts. Transplantation of xenogeneic bone marrow cells under costimulatory blockade at the time of heart transplantation may induce transplantation tolerance.

Congenital Segmental Dilatation of the Duodenum : Report of a Case

We herein report what, to our knowledge, is only the fourth known case of segmental dilatation of the duodenum. Antenatal ultrasonography revealed an intraabdominal cyst in the fetus, but the exact location of the segmental dilatation was difficult to find preoperatively. Moreover, even using computed tomography, it was not possible to make a diagnosis prior to surgery. The anatomic characteristics of duodenal dilatation made it difficult to perform the usual resection techniques. In fact, the surgical procedure was different from the previously reported cases. We performed a partial resection of the duodenum followed by a tapering procedure to preserve the ampulla of Vater. The infant had an uneventful postoperative course, and sufficient growth and development has been achieved.

Surgical Stress and the Development of Complications in Relation to Polymorphonuclear Leukocyte Elastase (PMNE) Levels

This study was conducted to examine the effects of surgical stress on changes in polymorphonuclear leukocyte elastase (PMNE) levels, and to evaluate the relationship of these changes to the development of postoperative complication. A total of 69 patients who underwent alimentary surgery were subsequently divided into three groups: a complicated group, comprised of 25 patients; an uncomplicated group with a high blood loss (H) of more than 1 000 ml, comprised of 18 patients; and an uncomplicated group with a low blood loss (L) of less than 1 000 ml, comprised of 26 patients. The changes in the levels of PMNE, fibronectin (FN), and antithrombin III (AT III) were compared among these three groups. In the uncomplicated H and L groups the PMNE levels rose significantly on postoperative day (POD) 1. On POD 3, high levels of PMNE were still evident in the uncomplicated H group, but a decline was observed in the uncomplicated L group. From POD 7 onwards the levels decreased to the preoperative values in both uncomplicated groups; however, the complicated group continued to show high levels even on POD 14. Significantly decreased FN levels were observed for the first 3 PODs in each group. The uncomplicated H and L groups regained their preoperative levels on PODs 7 and 14, respectively, but no recovery was found in the complicated group. The AT III levels showed similar changes to the FN levels in all groups. These findings indicate that monitoring the PMNE levels could be a useful index for the early detection of postoperative complications following alimentary surgery.

Adenoma of the Nipple, Focusing on the Contrast-Enhanced Magnetic Resonance Imaging Findings: Report of a Case

A woman in her 50s was referred to our hospital for an investigation of a right breast tumor. The tumor was palpated below the nipple, but there was no erosion or nipple discharge. Mammography showed a well-defined high-density tumor, measuring 2 cm in diameter, without calcification, and ultrasonography showed a low-echoic mass with a fluid component with posterior echo enhancement and a lateral shadow. Contrast-enhanced magnetic resonance imaging (CE-MRI) demonstrated a 1.3 × 0.8 cm solid component and a gradually increasing time-intensity curve. We performed lumpectomy and the pathological findings were adenoma of the nipple. The pattern of the time-intensity curve might be attributed to moderate fibrosis of the tumor. Contrasten-hanced MRI is therefore considered to be very useful in the diagnosis of breast disease because it can show the nature and extent of the breast lesion; however, we should be aware that various patterns have been observed on CE-MRI for adenoma of the nipple.

The Mobilization and Recruitment of C-Kit+ Cells Contribute to Wound Healing after Surgery

Delayed wound healing is a serious clinical problem in patients after surgery. A recent study has demonstrated that bone marrow-derived c-kit-positive (c-kit+) cells play important roles in repairing and regenerating various tissues and organs. To examine the hypothesis that surgical injury induces the mobilization and recruitment of c-kit+ cells to accelerate wound healing. Mice were subjected to a left pneumonectomy. The mobilization of c-kit+ cells was monitored after surgery. Using green fluorescent protein (GFP+) bone marrow-transplanted chimera mice, we investigated further whether the mobilized c-kit+ cells were recruited to effect wound healing in a skin puncture model. The group with left pneumonectomies increased the c-kit+ and CD34+ stem cells in peripheral blood 24 h after surgery. At 3 days after surgery, the skin wound size was observed to be significantly smaller, and the number of bone marrow-derived GFP+ cells and GFP+/c-kit+ cells in the wound tissue was significantly greater in mice that had received pneumonectomies, as compared with those that had received a sham operation. Furthermore, some of these GFP+ cells were positively expressed specific markers of macrophages (F4/80), endothelial cells (CD31), and myofibroblasts (αSMA). The administration of AMD3100, an antagonist of a stromal-cell derived factor (SDF)-1/CXCR4 signaling pathway, reduced the number of GFP+ cells in wound tissue and completely negated the accelerated wound healing. Surgical injury induces the mobilization and recruitment of c-kit+ cells to contribute to wound healing. Regulating c-kit+ cells may provide a new approach that accelerates wound healing after surgery.