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Supplementary Table 1 from Loss of Hepatocyte Growth Factor/c-Met Signaling Pathway Accelerates Early Stages of N-nitrosodiethylamine–Induced Hepatocarcinogenesis

Supplementary Methods and Materials from Loss of Hepatocyte Growth Factor/c-Met Signaling Pathway Accelerates Early Stages of N-nitrosodiethylamine–Induced Hepatocarcinogenesis

Hepatocyte growth factor (HGF) has been reported to have both positive and negative effects on carcinogenesis. Here, we show that the loss of c-Met signaling in hepatocytes enhanced rather than suppressed the early stages of chemical hepatocarcinogenesis. c-Met conditional knockout mice (c-metfl/fl, AlbCre+/−; MetLivKO) treated with N-nitrosodiethylamine developed significantly more and bigger tumors and with a shorter latency compared with control (w/w, AlbCre+/−; Cre-Ctrl) mice. Accelerated tumor development was associated with increased rate of cell proliferation and prolonged activation of epidermal growth factor receptor (EGFR) signaling. MetLivKO livers treated with N-nitrosodiethylamine also displayed elevated lipid peroxidation, decreased ratio of reduced glutathione to oxidized glutathione, and up-regulation of superoxide dismutase 1 and heat shock protein 70, all consistent with increased oxidative stress. Likewise, gene expression profiling done at 3 and 5 months after N-nitrosodiethylamine treatment revealed up-regulation of genes associated with cell proliferation and stress responses in c-Met mutant livers. The negative effects of c-Met deficiency were reversed by chronic p.o. administration of antioxidant N–acetyl–l-cysteine. N–acetyl–l-cysteine blocked the EGFR activation and reduced the N-nitrosodiethylamine–initiated hepatocarcinogenesis to the levels of Cre-Ctrl mice. These results argue that intact HGF/c-Met signaling is essential for maintaining normal redox homeostasis in the liver and has tumor suppressor effect(s) during the early stages of N-nitrosodiethylamine–induced hepatocarcinogenesis. [Cancer Res 2007;67(20):9844–51]

During skull base surgery, reconstruction of the dura is a very important procedure. Here, the use of a temporofrontal pedicled fascia flap tunnelled between temporal muscle fibres is described. This technique is easily performed and has a low postoperative morbidity, including skin and temporal muscle complications.

Liver cirrhosis (LC) is often complicated by hyperinsulinemia due to insulin resistance (IR), which is considered to be closely related to shunt formation and impaired liver function. This study evaluates whether balloon-occluded retrograde transvenous obliteration (B-RTO) can affect glucose and insulin metabolism in patients with LC. Twenty-five cirrhotic patients (mean age = 69.6 years; female/male = 12/13; hepatitis C virus/alcohol/nonalcoholic steatohepatitis = 14/6/5; Child-Pugh’s class A/B = 10/15) with gastric varices and/or hepatic encephalopathy caused by portosystemic shunts (PSS) due to portal hypertension (PH) underwent B-RTO at our hospital. Testing was performed before and at 1 month after the procedure. Shunt occlusion resulted in a decrease in extrahepatic collateral blood flow and an increase in portal venous flow, as well as a dramatic improvement in hepatic function markers. In addition, B-RTO significantly decreased homeostasis model assessment (HOMA) of IR without a statistical decline of HOMA of β-cell function. The 75-g oral glucose tolerance test (75-OGTT) revealed that occlusion of PSS reduced both fasting immunoreactive insulin (IRI) levels and the area under the curve for IRI. However, no significant change in preprandial or postprandial plasma glucose levels was observed. Furthermore, according to the criteria of the American Diabetes Association, B-RTO led to an improved 75-OGTT profile in 58.3 % of patients who had impaired glucose tolerance or diabetes mellitus before the procedure. Shunt occlusion improves IR-related hyperinsulinemia through increased portal venous flow, ameliorated liver function, and consequent augmented hepatic insulin clearance in cirrhotic patients with PH.

Aim Insulin resistance (IR) increases during the early stages of hepatitis C virus (HCV)-related chronic liver disease and is a sign of poor prognosis as well as a risk factor for hepatic fibrosis and hepatocellular carcinoma. We aimed to determine the factors affecting IR in HCV-related chronic liver disease. Methods We retrospectively examined 71 patients with HCV-related chronic liver disease and analyzed various parameters, including amino acids, as possible predictors of IR. IR was assessed using the Homeostasis Model of Assessment – Insulin Resistance (HOMA-IR). Amino acids were assayed by examining branched-chain amino acids (BCAA), tyrosine level, and the ratio of BCAA to tyrosine level (BTR). Results HOMA-IR was significantly correlated with body mass index, platelet count, prothrombin time, hemoglobin, total bilirubin, total protein, albumin, total cholesterol, fasting glucose, BTR (r = −0.46, P = 0.0001) and tyrosine (r = 0.55, P

Aim We recently reported that the iron chelator deferoxamine (DFO) is efficacious in advanced hepatocellular carcinoma (HCC) patients. Iron regulation may thus have an important impact in HCC therapy. Because transferrin is a native chelator that regulates iron homeostasis, it may act as an anticancer agent in a similar manner as DFO. The objective of this study was to evaluate serum transferrin as a prognostic predictor in advanced HCC patients undergoing hepatic arterial infusion chemotherapy (HAIC). Methods We retrospectively studied 44 patients receiving HAIC and analyzed various parameters for their possible use as prognostic predictors. Results The 1-, 2- and 3-year cumulative survival rates were 36.4%, 18.2% and 8.5%, respectively, and the median survival time (MST) was 7.0 months. The survival rates of patients who had serum transferrin of 190 mg/dL or more (MST, 12.0 months) were significantly better than those of patients who had serum transferrin of less than 190 mg/dL (MST, 4.9 months). Multivariate analysis identified serum transferrin of 190 mg/dL or more (hazard ratio [HR], 0.282; 95% confidence interval [CI], 0.132–0.603; P = 0.001) and Child–Pugh score B (HR, 1.956; 95% CI, 1.034–3.700; P = 0.039) as independent prognostic predictors. There was a significant correlation between serum transferrin level and therapeutic effect (P

HGF/c-Met signaling plays a pivotal role in hepatocyte survival and tissue remodeling during liver regeneration. HGF treatment accelerates resolution of fibrosis in experimental animal models. Here, we utilized Metfl/fl;Alb-Cre+/− conditional knockout mice and a carbon tetrachloride(CCl4)-induced liver fibrosis model to formally address the role of c-Met signaling in hepatocytes in the context of chronic tissue injury. Histological changes during injury (4weeks) and healing phase (4weeks) were monitored by immunohistochemistry; expression levels of selected key fibrotic molecules were evaluated by western blotting, and time-dependent global transcriptomic changes were examined using a microarray platform. Loss of hepatocyte c-Met signaling altered hepatic microenvironment and aggravated hepatic fibrogenesis. Greater liver damage was associated with decreased hepatocyte proliferation, excessive stellate cell activation and rapid dystrophic calcification of necrotic areas. Global transcriptome analysis revealed a broad impact of c-Met on critical signaling pathways associated with fibrosis. Loss of hepatocyte c-Met caused a strong deregulation of chemotactic and inflammatory signaling (MCP-1, RANTES, Cxcl10) in addition to modulation of genes involved in reorganization of the cytoskeletal network (Actb, Tuba1a, Tuba8), intercellular communications and adhesion (Adam8, Icam1, Itgb2), control of cell proliferation (Ccng2, Csnk2a, Cdc6, cdk10), DNA damage and stress response (Rad9, Rad52, Ercc4, Gsta1 and 2, Jun). Our study demonstrates that deletion of c-Met receptor in hepatocytes results in pronounced changes in hepatic metabolism and microenvironment, and establishes an essential role for c-Met in maintaining the structural integrity and adaptive plasticity of the liver under adverse conditions.

57歳の男性．B型肝硬変・肝細胞癌に対して外来経過観察中，下血に対する精査加療目的で当科に緊急入院した．上部消化管内視鏡検査（EGD）で十二指腸下行脚に静脈瘤が認められたものの活動性出血および血液貯留の所見はなく，下部消化管内視鏡検査・カプセル内視鏡検査・経肛門的ダブルバルーン内視鏡検査においても回腸～結腸内に凝血塊の貯留が認められたのみで出血源の同定には至らなかった．腹部血管造影検査では十二指腸周辺に著明な門脈系-大循環系短絡路の発達が認められ，血管造影下CTで十二指腸下行脚付近に造影剤の血管外漏出が疑われた．直後の緊急EGDにより十二指腸静脈瘤破裂と診断し，内視鏡的静脈瘤結紮術（EVL）にて一次止血に成功した．しかしその２日後に出血性ショックに陥ったため，EVLで止血した上で引き続きバルーン閉塞下逆行性経静脈的塞栓術（B-RTO）を施行して完全止血を得た．十二指腸静脈瘤破裂は非常に稀な難治性消化管出血であるが，今回我々はEVL・B-RTO併用療法で止血し救命し得た症例を経験したので報告する．

左脈絡膜悪性黒色腫に対する手術歴（2004年4月，左眼球摘出術）を有する79歳の男性患者が2009年10月に右季肋部痛を主訴に当院外来を受診した．血液検査にて軽度の肝機能障害と高度の凝固系異常が認められたため精査加療目的で同日当科に緊急入院となった．HBs抗原・抗HBs抗体・抗HBc-IgG抗体・抗HCV抗体はいずれも陰性であり，腹部超音波検査・CT検査・MRI検査・血管造影検査で肝両葉に多血性腫瘤が多数認められた．悪性黒色腫に特異的な腫瘍マーカーである5-S-cysteinyldopaが異常高値を呈したがPIVKA IIも著明に上昇していたため画像・血液検査所見からは悪性黒色腫肝転移と肝細胞癌の鑑別には至らなかった．超音波下肝腫瘍生検にて悪性黒色腫の確定診断を得た上で同病変に対してシスプラチンを用いた肝動脈化学塞栓療法を施行した．今回我々は「脈絡膜悪性黒色腫肝転移」という非常に稀な一例を経験したので報告する．

We examined the effect of cyclosporin A (CsA) on the pathogenesis of acute experimental liver injury in rats induced by injection of heat-killed Propionibacterium acnes (P. acnes) and subsequent injection of lipopolysaccharide (LPS). Pretreatment with CsA significantly reduced serum alanine aminotransferase (ALT), serum tumor necrosis factor-alpha (TNF-alpha) production, without changing the TNF-alpha mRNA level in the liver, and plasma interferon-gamma (IFN-gamma), following LPS injection in this model. Twenty-four-hour mortality was also markedly improved, from 100% in the P. acnes plus LPS group to 0% in the CsA-pretreated group. Although direct addition of CsA to isolated hepatic macrophages from P. acnes-pretreated rats did not prevent the production of TNF-alpha and active oxygen species, isolated hepatic macrophages from P. acnes plus CsA-pretreated rats significantly reduced their production in response to the addition of LPS. These results suggest that CsA protects against P. acnes plus LPS-induced acute liver injury, not by direct inhibition of hepatic macrophage activation, but by indirect prevention of hepatic macrophage activation, presumably related to the reduction in plasma IFN-gamma levels.

AIM A late evening snack (LES) is recommended for protein-energy malnutrition in patients with liver cirrhosis. However, many cases of liver cirrhosis have accompanying impaired glucose tolerance and there are concerns that LESs might aggravate glucose intolerance. In this study, we concomitantly used an alpha-glucosidase inhibitor with a LES and examined the effects on glucose tolerance. In addition, we examined whether or not there was an improvement in energy metabolism by slowing glucose absorption with the concomitant use of the alpha-glucosidase inhibitor. METHODS The subjects were 11 patients with liver cirrhosis. From before the study, all the patients had been taking a LES supplementation with a branched-chain amino acid (BCAA)-enriched nutrient mixture. The patients were started on the concomitant use of alpha-glucosidase inhibitor (0.2 mg) taken just prior to the LES. The change of glucose tolerance and energy metabolism were examined using a 75-g oral glucose tolerance test and indirect calorimetry. RESULTS One week and three months after the start of the concomitant use of the alpha-glucosidase inhibitor, the area under the concentration curve for plasma glucose was significantly decreased. Three months after the concomitant use, the non-protein respiratory quotient was significantly improved. There were no serious side effects during the follow-ups. CONCLUSION The concomitant use of the alpha-glucosidase inhibitor use with LES showed the possibility of improving glucose tolerance and energy metabolism. In patients with impaired glucose tolerance, the concomitant use of an alpha-glucosidase inhibitor with LES might be a useful measure for nutritional management.

Rennin-angiotensin system is involved in liver fibrogenesis through activating hepatic stellate cells (HSCs). Telmisartan (Tel) is an angiotensin II type 1 receptor antagonist, could function as a selective peroxisome proliferator-activated receptor gamma activator. Here we studied the effect of Tel on liver fibrosis, pre-neoplastic lesions in vivo and primary HSCs in vitro. In vivo study, we used the choline-deficient L-amino acid-defined (CDAA)-diet induced rat NASH model. The rats were fed the CDAA diet for 8 weeks to induce liver fibrosis and pre-neoplastic lesions, and then co-administrated with Tel for another 10 weeks. Tel prevented liver fibrogenesis and pre-neoplastic lesions by down-regulating TGFbeta1 and TIMP-1, 2 and increasing MMP-13 expression. Tel inhibited HSCs activation and proliferation. These results suggested that Tel could be a promising drug for NASH related liver fibrosis.

Aim: We investigated the effect of supplementation with branched-chain amino acids (BCAA) in patients with liver cirrhosis on the change of energy metabolism as well as glucose tolerance. Methods: Thirty liver cirrhosis patients underwent nutrient supervision by a dietician for one week. They were then prescribed oral supplementation with three packs of a BCAA nutrient (Livact 4.15 g/pack; Ajinomoto Pharma, Tokyo, Japan), taken three times a day: after breakfast, dinner and before sleep. The change in energy metabolism and glucose tolerance was examined using an indirect calorimeter and 75 g oral glucose tolerance test (75 g OGTT). Results: Non-protein respiratory quotient (npRQ) as well as branched-chain amino acid/tyrosine ratio (BTR) showed significant improvement, especially in patients with a creatinine height index (CHI) greater than 80. There was also a significant correlation between npRQ after one week of BCAA supplementation and the CHI. The patients with CHI greater than 80 and those with borderline pattern assessed by 75 g OGTT showed significant improvement in impaired glucose tolerance. Conclusion: Liver cirrhosis patients with CHI greater than 80 are the first candidates for BCAA supplementation. These patients showed improvement not only in energy metabolism and BTR, but also glucose tolerance.

Although sorafenib is expected to have a chemopreventive effect on hepatocellular carcinoma (HCC) recurrence, there are limitations to its use because of adverse effects, including effects on liver function. We have reported that the iron chelator, deferoxamine can prevent liver fibrosis and preneoplastic lesions. We investigated the influence of administering a new oral iron chelator, deferasirox (DFX), on the effects of sorafenib. We used the choline-deficient l-amino acid-defined (CDAA) diet-induced rat liver fibrosis and HCC model. We divided rats into four groups: CDAA diet only (control group), CDAA diet with sorafenib (sorafenib group), CDAA diet with DFX (DFX group), and CDAA diet with DFX and sorafenib (DFX + sorafenib group). Liver fibrosis and development of preneoplastic lesions were assessed. In addition, we assessed adverse effects such as changes in body and liver weight, skin damage (eruption, dryness, and hair loss), which is defined as hand-foot skin syndrome, in the sorafenib and DFX + sorafenib groups. The combination of DFX + sorafenib markedly prevented liver fibrosis and preneoplastic lesions better than the other treatments. Furthermore, the combination therapy significantly decreased adverse effects compared with the sorafenib group. In conclusion, the combination therapy with DFX and sorafenib may be a useful adjuvant therapy to prevent recurrence after curative treatment of HCC.

Background & Aims Hepatic arterial infusion chemotherapy (HAIC) is an option for treating advanced hepatocellular carcinoma (HCC). Because of the poor prognosis in HAIC non-responders, it is important to identify patients who may benefit from continuous HAIC treatment; however, there are currently no therapeutic assessment scores for this identification. Therefore, we aimed to establish a new therapeutic assessment score for such patients. Methods We retrospectively analyzed 90 advanced HCC patients with elevated baseline alpha-fetoprotein (AFP) and/or des-gamma-carboxy prothrombin (DCP) levels and analyzed various parameters for their possible use as predictors of response and survival. AFP and DCP responses were assessed after half a course of HAIC (2 weeks); a positive-response was defined as a reduction of ≥ 20% from baseline. Results Multivariate analysis identified DCP response (odds ratio 16.03, p < 0.001) as an independent predictor of treatment response. In multivariate analysis, Child-Pugh class A (hazard ratio [HR] 1.99, p = 0.018), AFP response (HR 2.17, p = 0.007), and DCP response (HR 1.90, p = 0.030) were independent prognostic predictors. We developed an Assessment for Continuous Treatment with HAIC (ACTH) score, including the above 3 factors, which ranged from 0 to 3. Patients stratified into two groups according to this score showed significantly different prognoses (≤1 vs. ≥2 points: median survival time, 15.1 vs. 8.7 months; p = 0.003). Conclusions The ACTH score may be useful in the therapeutic assessment of HCC patients receiving HAIC.

Leptin is now recognized as a proinflammatory cytokine and thought to be a progressive factor for non-alcoholic steatohepatitis (NASH). Here we showed the effects of leptin on the production of TNF-alpha (tumor necrosis factor-alpha) by Kupffer cells (KCs) with signal transduction. Leptin enhanced TNF-alpha production accompanied by a dose-dependent increase of MAPK activity in lipopolysaccharide (LPS)-stimulated KCs. SB203580 and JNK inhibitor I, specific inhibitors of P38 and JNK, inhibited TNF-alpha production in KCs but PD98059, an inhibitor of the ERK pathway, did not affect TNF-alpha production by KCs. Recombinant constitutively active adenovirus (Ad)-MKK6 and-MKK7 increased TNF-alpha production in KCs with activation of P38 and JNK without any change by Ad-MEK1 delivery. On the other hand, KCs isolated from the Zucker rat (fa/fa), a leptin receptor-deficient rat, showed reduced production of TNF-alpha on stimulation with LPS. The delivery of Ad-MKK6 and-MKK7, but not Ad-MEK1, increased TNF-alpha production in KCs of Zucker rats with activation of P38 and JNK. Addition of leptin to normal rats increased LPS-induced hepatic TNF-alpha production in vivo and leptin receptor-deficient Zucker rats showed reduced hepatic TNF-alpha production on addition of LPS in vivo. These findings indicate that P38 and JNK pathways are involved in the signal transduction of leptin enhancement of LPS-induced TNF-alpha production.

Background Temporal bone chondroblastoma is a rare primary bone tumor that affects the floor of the middle cranial fossa. This tumor is known to have high recurrence rate after curettage, and wide resection is therefore recommended. However, the literature provides little information regarding long-term results after wide resection of temporal bone chondroblastoma. Methods Four cases of surgically treated temporal bone chondroblastoma underwent long-term follow-up. Results Four patients, 3 males and 1 female, with mean age of 34, were surgically treated at the neurosurgery department of Keio University Hospital. Two patients were treated for recurrent tumor and the other two for new disease. In all cases the tumor mainly involved the mandibular fossa with variable degree of infiltration into tympanic and petrous parts. The tumor was totally removed via zygomatic approach in all patients. In 3 patients, the mandibular condyle was removed to expose the tumor. These patients had temporary malocclusion and restricted motion postoperatively, which resolved within 3 to 12 months with conservative treatment. All patients have no recurrence to date with a mean follow-up period of 9 years. Conclusion Temporal bone chondroblastoma was removed totally with skull base surgical technique and no recurrence has occurred for 6 to 13 years postoperatively. We found that removal of the mandibular head does not cause permanent problems of mastication in patients with normal dentures.

Jugular foramen meningioma is very rare. Papillary meningioma, the histological aspects and clinical behavior of which are highly malignant, is also very rare. Only 55 cases of jugular foramen meningioma have been reported in the literature. To our knowledge, this is the first report of a jugular foramen papillary meningioma. A 25-year-old woman presented with hoarseness and right hearing disturbance. Neurological examination showed deficits of the right hypoglossal and vagus nerves with palsy. Magnetic resonance imaging (MRI) revealed a mass on the right jugular foramen extending intracranially and extracranially, with weak contrast enhancement. The intracranial tumor was removed via the right lateral suboccipital approach. Histopathological examination confirmed the diagnosis of papillary meningioma. We report here the first case of jugular foramen papillary meningioma. No completely effective treatment for papillary meningiomas exists at present, with the exception of surgery.

BACKGROUND Only 2 cases of abducens nerve schwannoma solely inside the cavernous sinus have been reported. In both cases, abducens nerve palsy remained after operation. We report the first case of abducens nerve schwannoma inside the cavernous sinus proper with postoperative recovery from abducens nerve palsy. CASE DESCRIPTION The patient was a 47-year-old female who developed left abducens and trigeminal nerve palsies. Neuroradiological examination revealed left intra-cavernous sinus tumor. Total removal of the tumor was performed. The location of the tumor was confirmed intraoperatively inside the cavernous sinus itself, with no relation to the trigeminal nerve. Further, the relation of the tumor to one particular nerve fiber within the abducens nerve bundle was confirmed inside the cavernous sinus. After surgery, the patient had transient abducens nerve palsy. It had totally disappeared by 6 months. CONCLUSION When the tumor origin is just within the spacious cavernous sinus rather than more posterior in the narrow dural tunnel of Dorello's canal, successful preservation of the nerve function is possible postoperatively through a thorough knowledge of the membranous anatomy and careful preoperative study of the radiographic findings.

The goal of this study was to determine the membranous protection of the abducens nerve in the petroclival region. The petroclival portion of the abducens nerve was studied in ten dissections from five cadaveric head specimens. One of the heads was used for histological sections. Four heads were injected with colored latex for microsurgical dissections. The histological sections were prepared from petroclival dura mater, embedded in paraffin blocks, stained, sectioned in the axial, coronal, and sagittal planes, and evaluated by light microscopy. The abducens nerve was covered by a dural sleeve and arachnoidal membrane during its course within the petroclival area. Following the petrous apex, the abducens nerve was fixed by a sympathetic plexus and connective tissue extensions to the lateral wall of the cavernous segment of the internal carotid artery and to the medial wall of Meckel's cave. Fibrous trabeculations inside the venous space were attached to the dural sleeve. The lateral clival artery accompanied the dural sleeve of the abducens nerve and supplied the petroclival dura mater. The arterioles accompanying the abducens nerve through the subarachnoid space supplied the nerve within the dural sleeve. The arachnoid membrane covered the abducens nerve within the dural sleeve to the petrous apex, and arachnoid granulations found on the dural sleeve protruded into the venous space. The extension of the arachnoid membrane to the petrous apex and the presence of arachnoid granulations on the dural sleeve suggest that the subarachnoid space continues in the dural sleeve.

The aim of this study was to investigate the role of leptin in the development of liver fibrosis with Kupffer cell function using leptin receptor deficient rats. Male Zucker (fa/fa) and control (fa/-) rats received pig serum for 8 weeks. Animals were sacrificed to estimate the degree of liver fibrosis and stellate cell activation with the expression of alpha smooth muscle actin (alphaSMA). Microarray analysis was performed. Isolated Kuppfer cells of Zucker and control rats were treated with LPS. LPS uptake and TNF-alpha production were examined. Stellate cells were also isolated from Zucker and control rats. The expression of procollagen type I mRNAs was examined. Control rats developed liver fibrosis 8 weeks after injection of pig serum and showed an increased liver hydroxyproline content of 348 +/- 34 microg/g (n = 10) compared with Zucker rats (225 +/- 13, n = 10, P0.01). The procollagen type I mRNA level and alphaSMA expression of Zucker rats were also significantly reduced. Microarray analysis indicated significantly reduced expression of TNF-alpha, LPS-binding protein, urokinase-type plasminogen activator (uPA), IGF, IGF-binding protein (IGFBP)-3,5, and increased expression of apolipoprotein IV. Isolated Kupffer cells of Zucker rats showed significantly reduced LPS uptake as well as TNF-alpha production compared with control rats. However, no significant change was observed in procollagen type I mRNA levels of isolated stellate cells after 4 days of culture on plastic dishes. These results suggest that leptin receptor deficiency retards the development of liver fibrosis due to the dysfunction of Kuppfer cells.

Cerebral infarction results in impairment of motor and cognitive functions. We performed intracranial transplantation of multipotential neuro-epithelial stem cells with mesenchyme into experimentally large ischemic lesions to study their potential to relieve deficits.Wistar albino rats were subjected to transient middle cerebral artery occlusion for 60 minutes, producing an extensive ischemic lesion in the ipsilateral striatum and adjacent cerebral cortex. The rat mesencephalic neural plate at the early somite stage (embryonic day 10.5) together with adjacent ventral mesenchyme was used as donor material. We performed histological and immunohistochemical studies, with antibodies against tyrosine hydroxylase, and dopamine- and adenosine 3': 5'-monophosphate-regulated phosphoprotein 32 (DARPP-32; a striatal marker). Micro-angiograms were made by using Microfil silicone rubber. Morris Water-maze learning and treadmill task were employed to evaluate motor and cognitive functions.A viable non-tumoral mass was recognized in the rat striatum, up to as long as 156 days after transplantation. There were many cells positive for tyrosine hydroxylase or DARPP-32 in the graft. Some of the DARPP-32 positive cells within the graft had extended their dendrites into host tissues. In the micro-angiograms, many fine vessels were observed within the graft and dilated vessels meandered around the graft. Transplanted animals recovered significantly better in motor and cognitive functions than animals injected with only culture medium.Neuro-epithelial stem cells may follow several lines of differentiation; along the naturally genetically programmed line of differentiation, or along other cell lines depending on different environments. Grafting of neuro-epithelial stem cells with mesenchyme may merit a further study as a treatment for cerebral infarction.

The petroclival or prepontine tumors originating medial to the trigeminal nerve include meningiomas, trigeminal neurinomas, laterally extended chordomas and prepontine epidermoids. They frequently show middle fossa or parasellar extension and have been one of the difficult tumors requiring a combined large craniotomy or multiple step surgery. Here, we will present a surgical method of tumor removal through a keyhole, epidurally created in a petrous apex. The technical points presented are based on my experience of 95 cases. This approach has the following advantages: (1) One-stage epidural surgery to the petroclival and dumbbell parasellar–clival lesions through a small craniotomy; (2) Minimal venous complications compared to the presigmoid approach; (3) Minimal facial and lower cranial nerves injury by jumping into the petroclival area medial to the internal auditory meatus; (4) Bloodless tumor removal being possible by detachment of the tentorial artery; (5) Hearing preservation by limited petrous resection.

The purpose of this study was to elucidate the biological significance and the possibility of intracerebral grafting of neuroepithelial stem cells derived from the mesencephalic neural plate. Immunohistological studies of embryonic day 10.5 (E10.5) Wister rats revealed strong nestin expression in the mesencephalic part of the neural plate. Mesencephalic neural plates removed from E10.5 rats were processed to either tissue or cell dissociation culture. They were cultured in vitro under various conditions and were analyzed 7 days after the primary culture. When they were cultured as a tissue, cell proliferation and differentiation into neurons extending long neurites were obvious in a serum-free medium, in a medium containing 3% serum, and in a medium containing 20 ng/ml epidermal growth factor. On the other hand, in a medium containing 10 ng/ml basic fibroblast growth factor (bFGF), both vigorous cell proliferation and sphere formation were recognized. Furthermore, marked neurite growth was rarely seen in this culture. When they were plated in a dissociation culture, cell proliferation and neurosphere generation were also recognized only in a medium containing bFGF, depending on the initial cell concentration. The spheres, generated 7 days after the primary cell culture, were positively stained by nestin. These data suggested that bFGF was able to amplify the stem cell population present in the mesencephalic neural plate derived from early embryos. This might make it possible to obtain a large number of stem cells as donor material for neural transplantation on demand.

Tumor necrosis factor-alpha (TNF-alpha), may be involved in the pathogenesis of inflammatory bowel diseases (IBDs). The aim of this study was to evaluate the effect of TNF-alpha on the inflammatory activity of IBD.TNF-alpha mRNA expression in intestinal mucosal biopsy specimens from IBD patients [ulcerative colitis (UC), n = 54; and Crohn's disease (CD), n = 11] was analyzed using a competitive polymerase chain reaction. The degree of macrophage infiltration was analyzed by immunohistochemistry, using an antihuman CD68 antibody.TNF-alpha mRNA expression was increased in UC patients, corresponding to the inflammatory activity. However, in CD, TNF-alpha mRNA expression was not correlated with the endoscopic findings.We clarified that TNF-alpha mRNA expression was responsible for the inflammatory activity in UC. However, TNF-alpha mRNA expression was not correlated with the mucosal injury in CD.

OBJECTIVE: The goals of this investigation were to perform a detailed analysis of petroclival microanatomic features, to investigate the course of the abducens nerve in the petroclival region, and to identify potential causes of injury to neurovascular structures when anterior transpetrosal or transvenous endovascular approaches are used to treat pathological lesions in the petroclival region. METHODS: Petroclival microanatomic features were studied bilaterally in seven cadaveric head specimens, which were injected with colored silicone before microdissection. Another cadaveric head was used for histological section analyses. RESULTS: A lateral or medial location of the abducens nerve dural entrance porus, relative to the midline, was correlated with the course and angulation of the abducens nerve in the petroclival region. The angulation of the abducens nerve was greater and the nerve was closer to the petrous ridge in the lateral type, compared with the medial type. The abducens nerve exhibited three changes in direction, which represented the angulations in the petroclival region, at the dural entrance porus, the petrous apex, and the lateral wall of the internal carotid artery. The abducens nerve was covered by the dural sleeve and the arachnoid membrane, which became attenuated between the second and third angulation points. The abducens nerve was anastomosed with the sympathetic plexus and fixed by connective tissue extensions to the lateral wall of the internal carotid artery and the medial wall of Meckel's cave at the third angulation point. There were two types of trabeculations inside the sinuses around the petroclival region (tough and delicate). CONCLUSION: The petroclival part of the abducens nerve was protected in a dural sleeve accompanied by the arachnoid membrane. Therefore, the risk of abducens nerve injury during petrous apex resection via the anterior transpetrosal approach, with the use of the transvenous route through the inferior petrosal sinus to the cavernous sinus, should be lower than expected. The presence of two anatomic variations in the course of the abducens nerve, in addition to findings regarding nerve angulation and tethering points, may explain the relationships between adjacent structures and the susceptibility to nerve injury with either surgical or endovascular approaches. Venous anatomic variations may account for previously reported cases of subarachnoid hemorrhage with the endovascular approach.

A 40-year-old female was admitted to the hospital with complaints of headache worsening gradually over a 1-month duration. Her past history included surgery to treat a left cerebellar cystic lesion 3 years before, and an untreated small solid right supracerebellar lesion of 1 cm diameter. On admission, magnetic resonance imaging showed that the right cerebellar lesion had grown to approximately 4 cm diameter abutting the tentorium and causing obstructive hydrocephalus. She also had two more small lesions, a right supratentorial solid lesion with cystic component near the splenium and an intramedullary cystic lesion at the C-2 level. Right suboccipital craniectomy was done. The vascular attachments between the superior aspect of the tumor and the tentorium were coagulated and the tumor was totally removed. C1-2 laminectomy was also performed to drain the intramedullary cyst. The patient deteriorated and lost consciousness with respiratory arrest 6 hours postoperatively and was reoperated for intracerebellar hematoma due to oozing from the tentorial vessels. Histological investigation revealed hemangioblastoma. Dural tentorial vascular attachments in solid hemangioblastomas located subjacent to the tentorium may cause early postoperative complications of hematoma at the site of vascular attachment following the resection. Computed tomography study in the early postoperative period is helpful to identify this problem.

A 17-year-old male presented with a wound on the right temporal region, oozing hemorrhagic necrotic brain tissue and cerebrospinal fluid, following a fall. Computed tomography showed temporoparietal and petrous apex fractures on the right. Neurological examination revealed abducens nerve paresis, ptosis, and myosis on the right side. The patient was treated surgically for the removal of the free bony fragments at the fracture site and to close the dural tear. The abducens nerve paresis, ptosis, and myosis persisted at the 3rd monthly postoperative follow-up examination. The anatomy of the abducens nerve at the petroclival region was studied in four cadaveric heads. Two silicone-injected heads were used for microsurgical dissections and two for histological sections. The abducens nerve has three different angulations in the petroclival region, located at the dural entrance porus, the petrous apex, and the lateral wall of the cavernous segment of the internal carotid artery. The abducens nerve had fine anastomoses with the trigeminal nerve and the periarterial sympathetic plexus. There were fibrous connections extending inside the venous space of the petroclival area. The abducens nerve seems to be vulnerable to damage in the petroclival region, either directly by trauma to its dural porus and petrous apex or indirectly by stretching of the nerve through the nervous and/or fibrous connections. Concurrent functional loss of the abducens nerve and the periarterial sympathetic plexus clinically manifested as incomplete Horner's syndrome in our patient.

A 39-year-old woman was admitted with complaints of headache and nasal discharge on the left for 3 months which was later on proved to be cerebrospinal fluid (CSF). Neurological examination found no abnormalities except bilateral papilledema. Neuroimaging demonstrated enlargement of the lamina cribrosa foramina through which the olfactory nerves pass, as well as empty sella and cerebral cortical atrophy. Bone mineral densitometry showed osteopenia. CSF Ca++ and blood parathyroid hormone levels were elevated. CSF pressure was 280 mmH2O. Bilateral frontal craniotomy was performed to expose the anterior fossa. Foraminal enlargement at the lamina cribrosa was confirmed, and islands of extra-osseous calcifications on the arachnoid membrane were identified. The base of the anterior fossa was repaired intradurally with fascial graft and fibrin glue on both sides. No CSF leakage was noted at 1-year follow up. Spontaneous CSF leakage probably resulted from enlargement of the foramina at the lamina cribrosa due to Ca++ mobilization from bones and pseudotumor cerebri not to the extent of hydrocephalus caused by poor CSF absorption at the arachnoid granulations obliterated by extra-osseous calcareous accumulation.

The aim of this study was to investigate the association of stellate cell activation with serum fibrosis markers in a rat model of hepatic fibrosis prepared using a choline-deficient l-amino acid (CDAA) defined diet. CDAA diet administration resulted in increased liver hydroxyproline contents in a time-dependent manner with activated stellate cells, expressing α-smooth muscle actin (α-SMA) as well as increased serum concentrations of amino-terminal procollagen type III peptide (PIIIP) and the 7S fragment of type IV collagen. Hydroxyproline content of the liver showed a closer correlation with the serum 7S (r = 0.75, P < 0.01) concentration than with the serum PIIIP (r = 0.51, P < 0.01) concentration. The percent area of α-SMA-positive cells showed stronger correlation with the serum PIIIP concentration (r = 0.85, P < 0.01) than with the 7S concentration (r = 0.50, P < 0.01). These results indicate that the serum PIIIP concentration reflects the activity of fibrogenesis, while the serum 7S concentration reflects the accumulation of collagen fibers in the liver.

The aim of the study was to investigate the effect of the immunosuppressant FK 506 (tacrolimus hydrate) on acute liver injury induced by Propionibacterium acnes and lipopolysaccharide (LPS). Acute liver injury was induced in male Wistar rats by injecting the animals with P. acnes (10 mg/rat), and administering LPS (10 μg/rat) seven days later. One group was given FK 506 (1 mg/kg) 24 and 2 hr before administration of LPS, and the other group was given the same dose of saline. The 24-hr survival rate, serum alanine aminotransferase (ALT) concentration, and tumor necrosis factor (TNF) -α mRNA and protein concentrations in the liver and spleen were then compared. Hepatic macrophages were also isolated from rats seven days after P. acnes injection, LPS, and FK 506 or saline were added to the culture supernatant, and TNF-α production was studied. The 24-hr survival rate was 100% in the FK 506-treated group, in contrast with 16.6% in the saline group. Four hours after LPS injection, the serum ALT concentration was 755 ± 401 in the saline group versus 119 ± 42 units/ml (P < 0.01) in the FK 506-treated group. The serum TNF-α concentration was lower in the FK 506-treated group (1419 ± 957 pg/ml) than in the saline group (9205 ± 2215) (P < 0.01). The mRNA and protein concentrations in the liver and spleen in the two groups did not differ significantly 1 hr after LPS injection but were significantly lower in the FK 506-treated group after 4 hr. FK 506 did not directly inhibit TNF-α production by isolated cultured hepatic macrophages. FK 506 is unable to inhibit initial TNF-α production by hepatic macrophages (or probably that by splenic macrophages either) stimulated by injection of LPS in P. acnes + LPS-induced acute liver injury. However, the immunosuppressant does limit hepatic damage by inhibiting subsequent aggravation of inflammation by the cytokine network.

Injection of pig serum into rats twice a week for eight weeks induced transforming growth factor-β1 (TGF-β1) mRNA expression and protein production resulting in liver fibrosis without parenchymal cell injury. Eight-week treatment with pig serum reduced bromodeoxyuridine (BrdU) -positive hepatocytes 24 hr after 70% partial hepatectomy compared to that in the livers of rats treated with saline for eight weeks. Administration of a choline-deficient l-amino acid-defined (CDAA) diet for six weeks with pig serum coadministration, after pretreatment with pig serum for eight weeks, led to the development of preneoplastic lesions that were positive for the placental form of glutathione S-transferase (GSTP). Eight-week pretreatment with pig serum induced more GSTP-positive lesions and TGF-β1 mRNA expression and protein concentration in the livers of rats subsequently fed a CDAA diet for six weeks than in rats fed the CDAA diet with saline treatment. These results indicate that TGF-β1 induced by pig serum treatment inhibited hepatocyte proliferation but failed to prevent the development of preneoplastic lesions in a CDAA diet model.

Japanese upstream companies have consistently underperformed private independents. A recent audit by the Ministry of International Trade and Industry (MITI) underlines the magnitude of financial losses and the necessity for sectoral restructuring. MITI's proposals involve refocusing rather than redefining upstream policy, including recognition that official intervention should be reduced in favour of greater control by the more successful local production companies. Some commentators claim the proposed reforms do not go far enough. The ultimate success of MITI's initiatives, it is claimed, depends upon changes in prevailing corporate governance arrangements and the progressive replacement of existing senior managers with those having superior professional or technical qualification. These changes have not been directly mandated by Japanese authorities. On the other hand, MITI's proposals do imply a redisrtribution of corporate oversight in favour of greater market control, a major step in the direction of increased operational and financial efficiency.

After cerebral infarction, necrosis in neural tissues is not usually repaired or reconstructed by the injured brain. We therefore examined the effects on postinfarction repair of implanting central nervous system (CNS) stem cells together with mesenchyme, because CNS stem cells can be expected to adapt and survive in the adult brain. Cerebral infarction was induced by the Koizumi-Longa method, using the adult male spontaneous hypertensive rat model. Reperfusion was performed an hour after middle cerebral artery occlusion. The rat mesencephalic neural plate at the early somite stage (embryonic day 10.5) together with the adjacent ventral mesenchymal tissues was dissected out under the microscope and immediately implanted into the ischemic rat striatum. One month later, the cognitive function was evaluated by the Morris water maze method. Histologic and immunohistochemical examinations of the graft were made with hematoxylin-eosin (H&E), neurofilament-200, and tyrosine hydroxylase (TH) stains. In the water maze study, mean latency times required to reach an escape platform in the implanted animals with surviving grafts were found to be shorter than in those without grafts, but longer than in normal animals. In the spatial probe trial, the number of animals seen to cross the area in the pool where the platform had been located was greater in the implanted rats with surviving grafts than in other groups. Multiple vascularization in the grafted area was observed histologically in H&E-stained tissues, and neurofilament-200-positive cells were recognized in the graft. TH staining revealed within the graft many immunoreactive neuron-like cell bodies with long dendrites. It was suggested that grafted CNS stem cells with mesenchyme may survive and differentiate into mature CNS tissue within the adult ischemic rat brain, constructing vessels in and around the grafts, and may therefore have the potential to be effective in the recovery of the cognitive function of the rat model.

The proliferation and differentiation of stellate (Ito, or fat-storing) cells into myofibroblast-like cells is responsible for the development of liver fibrosis. Using computer image analysis, we evaluated the changes of alpha smooth muscle actin-positive stellate cells and liver fibrosis after interferon-alpha or -beta (IFN-alpha, beta) therapy in patients with chronic hepatitis C.Patients with chronic hepatitis C were treated with IFN-alpha or -beta and were divided into three groups on the basis of clinical criteria; a complete responder group (CR, 18 of 51), a partial responder group (PR, 17 to 51), and a nonresponder group (NR, 16 of 51). Liver fibrosis was assessed from specimens stained with Sirius red and was quantitated by computer image analysis. We also evaluated alpha-smooth muscle actin expression in the liver before and after IFN therapy by a semiquantitative scoring method (the alpha-smooth muscle actin index).Before IFN therapy, a large number of stellate cells expressing a-smooth muscle actin were present in the liver biopsy specimens. There was a significant correlation (r = 0.699, p0.05) between the change in the percent area of fibrosis and the alpha-smooth muscle actin index before and after IFN therapy in all groups. The complete responder group also showed a significant reduction of a-smooth muscle actin-expressing cells that was correlated with the reduction of serum ALT (r = 0.686, p0.05).These results suggest a-smooth muscle actin-expressing cells are responsible for liver fibrosis, and the elimination of factors stimulating matrix synthesis (e.g., hepatitis virus) may decrease liver fibrosis.

We have recently had many opportunities to operate on patients who presented with no neurologic deficits and were incidentally detected to have unruptured cerebral aneurysms. Therefore, it is very important to know how craniotomy affects the normal brain when making decisions regarding surgery for asymptomatic cerebral diseases.Thirty patients with unruptured cerebral aneurysms were evaluated. Aneurysm sites were as follows: internal carotid artery (ICA) (eight cases), anterior communicating artery (ACoA) (seven), middle cerebral artery (MCA) (five), basilar artery-superior cerebellar artery (BA-SCA) (two), and multiple aneurysms (three) [bilateral MCA: (two), ICA + Basilar top: (one)]. These patients underwent neuropsychological examinations [the Mini-Mental State (MMS) test, the "Kana-hiroi" test, and the "Maze" test] and single photon emission computed tomography (SPECT) before and after surgery.The details of the 17 cases whose Kana-hiroi tests deteriorated 1 month after surgery are as follows: ACoA: seven, MCA: five, ICA: three, BA: two. SPECT showed a decrease in CBF in nine cases (ACoA: six, ICA: two, MCA: one). There was a statistically significant difference (p0.05) between patients with ACoA aneurysms and ICA aneurysms. Three months after operation, all of the patients with neuropsychological deterioration and four of the six low CBF patients recovered to preoperative levels.These results indicated that the operation for unruptured cerebral aneurysms is moderately safe and meaningful; such a study should be required to determine when those patients could return to normal life.

The aim of this study was to investigate wheThe ran iron chelator, deferoxamine (DFO) can prevent lipidperoxidation, resulting in reduced liver injury as wellas reducing preneoplastic lesions induced by a choline-deficient L-amino acid-defined(CDAA) diet. CDAA diet administration resulted in anincreased serum ALT level (367 ± 58) after twoweeks, but simultaneous DFO treatment for two weeksreduced this elevation of ALT as well asmalondialdehyde (MDA) production in the liver. Feedingrats a CDAA diet for 12 weeks led to the development ofsevere liver fibrosis and preneoplastic lesions detectedas enzymealtered lesions. DFO treatment preventedthe expression of activated stellate cells, resulting inThe reduction of liver fibrosis as well as reducing thedevelopment of preneoplastic lesions. These results indicate that iron chelation can reducethe development of preneoplastic lesions in a CDAA dietmodel.

Injection of pig serum into rats twice a week for 8 weeks induced stellate cell activation resulting in liver fibrosis without parenchymal cell injury. Administration of a choline deficient L-amino acid defined (CDAA) diet for 6 weeks with or without pig serum pretreatment led to the development of preneoplastic lesions that were positive for the placental form of glutathione S-transferase (GSTP). Pig serum pretreatment induced more activated stellate cells in the livers of rats subsequently fed a CDAA diet for 6 weeks compared with rats fed the CDAA diet alone. Activated stellate cells were detected as α smooth muscle actin (αSMA)-positive cells and by the expression of αSMA messenger RNA. These cells caused severe fibrosis as assessed by the hepatic hydroxyproline content. Pre-existing fibrosis induced by the activation of stellate cells with pig serum pretreatment increased hepatic malondialdehyde (MDA) level in parallel with GSTP-positive lesions. These results indicate that pre-existing fibrosis with the activated stellate cells accelerates the development of preneoplastic lesions in a CDAA diet model.

It is of extreme importance to prevent liver fibrosis and subsequent progression to liver cirrhosis. The aim of our study was to elucidate in vitro whether Sho-saiko-to exerted inhibitory effects on hepatic stellate cells.Hepatic stellate cells were isolated from male Wistar rats. Water-soluble ingredients of Sho-saiko-to were obtained at concentrations of 10, 100, 250, 500 and 1000 microg/ml. Morphological transformation was observed under a phase-contrast microscope. Flow cytometric analysis was performed on day 4 after culture to evaluate the potential to proliferate of the stellate cells by analyzing cell cycles. Northern blot analysis was carried out on day 3 after culture to determine the expressions of type I and type III procollagen mRNAs.(i) Sho-saiko-to 500 and 1000 microg/ml inhibited morphological transformation of the stellate cells to myofibroblast-like cells. (ii) Sho-saiko-to 500 and 1000 microg/ml significantly (p0.0001) accumulated the cells in the G0/G1 phase (118.8+/-0.7%, 119.2+/-0.5%, respectively as compared with control) and significantly (p0.0001) decreased cell numbers subsequently in G2/M phase (47.5+/-8.1%, 48.9+/-2.0%, respectively). (iii) Sho-saiko-to 500 and 1000 microg/ml also significantly (p0.05 or p0.0001) suppressed procollagen mRNA expression of type I to 51.5+/-6.4%, 34.9+/-3.7%, respectively, and type III to 51.3+/-12.3%, 46.7+/-11.4%, respectively.We have clarified the inhibitory effects of Sho-saiko-to on hepatic stellate cells in vitro. Sho-saiko-to could be a potent inhibitor in the pathogenesis of liver fibrosis.

The aim of this study was to investigate the effect of interferons- γ (IFN- γ ) and - α (IFN- α ) on the synthesis of matrix proteins such as collagens as well as the gene expression of tissue inhibitor of metalloproteinase-1 (TIMP-1) in vivo. We investigated the effects of IFN- γ and - α in a model of liver fibrosis induced by pig serum in male Wistar rats, which develop fibrosis without an increase in serum ALT (i.e. without hepatocyte injury). Rats were injected with 0.5 ml of pig serum twice a week for 8 weeks with or without 50 000 U of IFN- γ or 100 000 U of IFN- α . IFN- γ at the dose of 50 000 U day −1 prevented fibrosis, as indicated by reduced hydroxyproline content in the liver. IFN- γ at 50 000 U day −1 also reduced expression of type I procollagen as well as TIMP-1 in the liver. However, fibrosis was not reduced by IFN- α . Histologically, IFN- γ at 50 000 U day −1 also reduced the number of myofibroblast-like cells (activated stellate cells). These results indicate that IFN- γ , but not IFN- α , can prevent fibrosis by inhibiting the activation and proliferation of stellate cells, resulting in reduced expression of procollagen and TIMP-1 mRNA in pig serum-induced rat liver fibrosis in vivo.

Recent advances in technology have not substantially changed the high mortality rate associated with acute renal failure (ARF). To obtain a simple, valid prognostic index, we retrospectively evaluated the relative importance of demographic data, causes (acute insults) of renal failure, and comorbid clinical conditions for the outcome in 102 ARF patients who received renal replacement therapy with an overall mortality rate of 65% (66 of 102). There were no significant differences between survivors and nonsurvivors in age and gender. Mortality according to acute insults was similar to that of the whole population studied. Of the 10 clinical conditions at the time of the first renal replacement therapy, mechanical ventilation (p = 0.0002), cardiac failure (p = 0.0006), hepatic failure (p = 0.003), central nervous system dysfunction (p = 0.005), and oliguria (p = 0.04) were found to be significantly related to mortality by univariate analysis. Furthermore, multivariate analysis demonstrated that only mechanical ventilation, cardiac failure, and hepatic failure were significant risk factors. Survival was directly related to the number of significant variables in univariate analysis: zero, 89% (8 of 9); one, 62% (21 of 34); two, 19% (5 of 27); three, 10% (2 of 20); four, 0% (0 of 8); five, 0% (0 of 4). This simple and early prognostic index, derived from the assessment of clinical conditions which were easily determined at the patient's bedside, could be useful for outcome prediction in ARF patients requiring renal replacement therapy.