Susumu Tamakawa

Hypoxia-Independent Overexpression of Hypoxia-Inducible Factor 1α as an Early Change in Mouse Hepatocarcinogenesis

Hypoxia-inducible factor 1 (HIF-1) is involved in tumor progression/metastasis and activated in various cancers. Here we show that HIF-1alpha, which plays a major role in HIF-1 activation, is overexpressed in preneoplastic hepatocytic lesions from a very early stage during hepatocarcinogenesis in mice and man. Transcriptional targets of HIF-1, such as vascular endothelial growth factor, glut-1, c-met, and insulin-like growth factor II (IGF-II), were also overexpressed in mouse lesions. Oxygen tension within the lesions was not different from that of the normal hepatic tissues, indicating that HIF-1alpha expression was independent of hypoxia. On the other hand, Akt, the pathway of which can up-regulate HIF-1alpha expression, was activated in the mouse lesions, whereas HIF-1alpha was markedly down-regulated in the mouse hepatocellular carcinoma (HCC) cell lines after treatment with a phosphatidylinositol 3-kinase (PI3K) inhibitor, LY294002, indicating that HIF-1alpha expression is dependent on PI3K/Akt signaling. Conversely, HIF-1alpha knockdown by short interfering RNA in the HCC cell line resulted in decreased expression of activated Akt together with the HIF-1 target genes, indicating that Akt activation is reversely dependent on HIF-1 activation. Treating the HCC cells with IGF-II or epidermal growth factor (EGF) up-regulated both phospho-Akt and HIF-1alpha, whereas inhibition of IGF-II or EGF signaling down-regulated them both, suggesting that IGF-II and EGF can, at least in part, mediate the activation of Akt and HIF-1alpha. However, Akt was not activated by IGF-II or EGF in the HIF-1alpha knockdown cells, indicating that expression of the HIF-1 target genes is necessary for the Akt activation. These findings suggest that the reciprocal activation of PI3K/Akt signaling and HIF-1alpha may be important in the progression of hepatocarcinogenesis.

Activated hepatic stellate cells overexpress p75NTR after partial hepatectomy and undergo apoptosis on nerve growth factor stimulation.

Abstract: Background: Expression of neurotrophins (NTs) and their receptors is increased during hepatic regeneration, but their role is not well understood.

Neoplastic hepatocyte growth associated with cyclin D1 redistribution from the cytoplasm to the nucleus in mouse hepatocarcinogenesis

Cyclin D1 overexpression is a frequent change in hepatocellular carcinomas (HCCs). Our present study demonstrated that cyclin D1 overexpression with abundant cyclin E, cdk4, cdk2, and p27Kip1 (p27) occurred in neoplastic hepatocytes from the early stage of mouse hepatocarcinogenesis. While cyclin D1 expression was mainly found in the cytoplasm of the tumor cells, it shifted to the nucleus in association with cell proliferation after the animals were subjected to a partial hepatectomy (PH), and then returned once more to the cytoplasm when the cells became quiescent. Inhibition of PI3 kinase (PI3K) by Ly294002 in mouse HCC cells in vitro suppressed the nuclear shift of cyclin D1 as well as cell proliferation, while PI3K activation by PTEN suppression failed to induce nuclear shift of cyclin D1, suggesting that PI3K activation is essential but not sufficient for the cyclin D1 nuclear shift. While MEK‐ERK1/2 inhibition by PD98059 and mTOR inhibition by rapamycin affected the cyclin D1 nuclear shift and cell proliferation to a lesser extent, both these inhibitors reduced cyclin D1 levels. Finally, although p27, cdk4 and calmodulin (CaM) were detected in the cyclin D1 immunoprecipitates from both quiescent and proliferating HCC cells, Hsc70 and SSeCKS were detected only in the immunoprecipitate from quiescent cells, and p21Waf1/Cip1 (p21) was detected only in that from proliferating cells, suggesting that the cyclin D1 complex is different in quiescent and proliferating cells. These observations indicate that the nuclear/cytoplasmic localization of cyclin D1 plays an important role in the proliferation/quiescence of neoplastic hepatocytes.

Expression of NGF in hepatocellular carcinoma cells with its receptors in non-tumor cell components

Nerve growth factor (NGF) is suggested to have a role in tumor progression in addition to its role in differentiation and survival of neuronal cells. We investigated expression of NGF and its receptors, TrkA and p75NTR, in hepatocellular carcinomas (HCCs). Although hepatocytes and hepatic stellate cells (HSCs) showed respectively weak and intense NGF immunostaining in the background livers of patients suffering from liver cirrhosis (LC) or chronic hepatitis (CH), intense staining was demonstrated in HCC cells of 33 of 54 (61.1%) tumors. RT‐PCR detected NGF mRNA in 7 freshly‐isolated HCC samples, and in 2 of 4 cases, in which both background livers and tumors could be analyzed, NGF mRNA was more abundant in the tumors than the background livers. TrkA was detected in the smooth muscle cells of hepatic arteries, but it was negative in tumor cells as well as non‐neoplastic hepatocytes. p75NTR and α‐smooth muscle actin (αSMA) was expressed in HSCs in the background liver and fibroblast‐like cells in stromal septa, whereas HSCs within the HCC tissues were mostly negative for p75NTR but positive for αSMA. This suggests that HSCs in HCC have a different property from those in background livers. Furthermore, the stromal septa contained abundant nerve fibers, which may be related to the increased NGF expression in HCC cells. NGF and its receptors are then thought to have a role in cellular interactions involving HCC cells, HSCs, arterial cells and nerve cells in HCC tissues.

Low p38 MAPK and JNK activation in cultured hepatocytes of DRH rats; a strain highly resistant to hepatocarcinogenesis

DRH rats are a hepatocarcinogenesis‐resistant strain isolated from hepatocarcinogenesis‐sensitive Donryu rats, and the liver of DRH shows less histological damage and fewer/smaller neoplastic hepatic lesions by the treatment with hepatocarcinogens. To investigate the mechanism of the resistance, the properties of hepatocytes of DRH and Donryu were compared. In primary culture, DRH hepatocytes exhibited higher proliferation and less apoptosis than Donryu hepatocytes in the presence of EGF and insulin. However, such difference was not correlated to the degree of DNA damage associated with cell culture or cell cycle checkpoint function. Although the mitogen‐activated protein kinases [EGF receptor (EGFR) and extracellular signal regulating kinases (ERK1/2)] were activated to the same degree, the stress‐activated protein kinases [p38 mitogen‐activated protein kinase (p38) and c‐jun N‐terminal kinase (JNK)] were activated to a lesser degree in the DRH hepatocytes. Treatment with 2‐acetylaminofluorene (2‐AAF) in vivo also resulted in less JNK and p38 activation in the DRH livers. Furthermore, apoptosis signal‐regulating kinase 1 (ASK1) was inhibited by the lysate from the DRH but not by the Donryu hepatocytes. The low activation of the stress‐activated protein kinases may be linked to the resistance to cellular stress, which may underlie the hepatocarcinogenesis‐resistance in DRH rats.

A case of herpes zoster affecting the glossopharyngeal nerve

not eat because of a sore throat, which was confined to the right side of pharynx. She also complained of absence of taste on the right side of the tongue and of absence of sweet taste on the left side of the tongue. Small vesicles were seen on the right portion of the soft palate and the right palatine tonsil (Fig. 1). No other cranial nerve seemed to be affected. Complement-fixing (CF) antibody titers for varicellazoster were 1 :32 (the normal range is less than 4 times), and varicella-zoster virus IgG (VZVIgG) titers on enzyme immunoassay were 103.2 (the normal range is less than 2.0). Using the PCR technique, we then found pieces of DNA specific for varicella-zoster virus in specimens from the vesicle and peripheral blood samples (Fig. 2). We diagnosed her as suffering from herpes zoster. She was admitted the day following the definitive diagnosis. After admission, the patient was given acyclovir 800mg five times per day orally for 5 days and loxoprofen sodium 180mg three times per day orally for 14 days. A stellate ganglion block with 5 ml of 1% mepivacaine was also performed for 28 days. The patient could eat liquid food three days after admission. CF antibody titers and VZVIgG titers decreased over several days. The vesicles became crusted 7 days after the commencement of treatment. The pain in the right ear and the sore throat were relieved within 1 month. Taste recovered completely on the left side of the tongue within 1 month, but absence of taste on the right side of the tongue has persisted.

High-dose intrathecal morphine to control cancer pain--a case report.

A 52-year-old woman with recurrent parotid gland cancer was referred to our hospital for uncontrollable severe pain in her right leg. We implanted a port in the chest wall and started a morphine infusion into the intrathecal space. The pain increased daily, and the patient finally needed 140 mg/day of morphine to control the pain. Cerebrospinal fluid was noted at the site of puncture and spinal catheter implantation. We removed the port and intrathecal catheter, then implanted the port again on the other side of the chest wall.

Different growth capacity between infant and adult mouse hepatocytes in vitro correlates to the cyclin D1 level without relation to oxidative DNA damage

Abstract: Background: Proliferating capacity of hepatocytes is rapidly decreased during growth into maturity, but its exact reason(s) are not well known.

Stellate ganglion block therapy for a patient with Tietze's syndrome

A 45-year-old woman came to our clinic because of chest wall pain. It was 6 years earlier when she first felt slight chest pain. The pain was in the infraclavicular region and ran to the left arm when pressure was applied. When the pain increased, she felt not only left upper chest pain but also paresthesia and weakness of the left arm. Physicians noted costal cartilage tenderness, but did not note swelling or flush over the skin. Laboratory studies revealed a white blood cell count to be 7000mm -3 and the concentration of C-reactive protein to be under 0.5mg.d1-1. Computed tomography (CT) showed swelling of the first left costal cartilage. Radionuclide imaging with 67Ga showed inflammation of the left first costochondral junction. A non steroid