Impact of elobixibat on liver tumors, microbiome, and bile acid levels in a mouse model of nonalcoholic steatohepatitis.

BACKGROUND: Elevated bile acid levels have been associated with liver tumors in fatty liver. Ileal bile acid transporter inhibitors may inhibit bile acid absorption in the distal ileum and increase bile acid levels in the colon, potentially decreasing the serum and hepatic bile acid levels. This study aimed to investigate the impact of these factors on liver tumor. METHODS: C57BL/6J mice received a one-time intraperitoneal injection of 25-mg/kg diethylnitrosamine. They were fed a choline-deficient high-fat diet for 20 weeks starting from 8 weeks of age, with or without elobixibat (EA Pharma, Tokyo, Japan). RESULTS: Both groups showed liver fat accumulation and fibrosis, with no significant differences between the two groups. However, mice with elobixibat showed fewer liver tumors. The total serum bile acid levels, including free, tauro-conjugated, glyco-conjugated, and tauro-α/ß-muricholic acids in the liver, were noticeably reduced following elobixibat treatment. The proportion of gram-positive bacteria in feces was significantly lower in the group treated with elobixibat (5.4%) than in the group without elobixibat (33.7%). CONCLUSION: Elobixibat suppressed tumor growth by inhibiting bile acid reabsorption, and decreasing total bile acid and primary bile acid levels in the serum and liver. Additionally, the presence of bile acids in the colon may have led to a significant reduction in the proportion of gram-positive bacteria, potentially resulting in decreased secondary bile acid synthesis.

An improved method to assess skeletal muscle mass in patients with liver cirrhosis based on computed tomography images.

AIM: Conventionally, the skeletal muscle area with computed tomography (CT) attenuation ranging from -29 to +150 Hounsfield unit (HU) divided by height squared (the conventional skeletal muscle index [SMI]) was used as an index of skeletal muscle mass. However, it includes fat-infiltrated skeletal muscle, which is known to have poor function. This study aims to determine whether the low-fat SMI, which uses skeletal muscle mass with CT attenuation ranging from +30 to +150 HU, or conventional SMI appropriately reflects the function of skeletal muscle. METHODS: We retrospectively analyzed 120 patients with cirrhosis whose handgrip strength was measured. Among them, 48 patients underwent a physical performance assessment such as liver frailty index (LFI) and short physical performance battery (SPPB), and 80 underwent quality of life (QOL) assessment. The relationships between each SMI and handgrip strength, LFI, SPPB, and QOL were evaluated. RESULTS: Low-fat SMI was significantly correlated with handgrip strength (males, R = 0.393, p = 0.002; females, R = 0.423, p < 0.001) and LFI (males, R = -0.535, p = 0.035; females, R = -0.368, p = 0.039), whereas conventional SMI was not. When using low-fat SMI, patients with low skeletal muscle mass had significantly low handgrip strength, LFI, SPPB, and physical and social-related QOL score than those without. By contrast, no significant differences were found for any items when using conventional SMI. CONCLUSIONS: Low-fat SMI is a good index of skeletal muscle mass that appropriately reflects skeletal muscle function.

Obesity and myosteatosis: the two characteristics of dynapenia in patients with cirrhosis.

OBJECTIVE: In patients with liver cirrhosis, the clinical characteristics of dynapenia, a condition in which skeletal muscle mass is maintained but muscle strength is reduced, are not yet known. This study aimed to clarify the characteristics of dynapenia and its impact on quality of life (QOL) in patients with liver cirrhosis. METHODS: We retrospectively analyzed 116 patients with cirrhosis. Based on grip strength and skeletal muscle mass measured by the bioelectrical impedance analysis method, patients were divided into four groups: normal muscle status, dynapenia, pre-sarcopenia (a condition involving only low muscle mass), and sarcopenia. The characteristics of dynapenia and its influence on QOL were examined. RESULTS: Fourteen patients had dynapenia. Liver function did not differ among the four groups. In patients with dynapenia, BMI was highest and computed tomography attenuation of skeletal muscle at the third lumbar spine vertebra was lowest among the four groups. The percentage of patients with both BMI ≥25 kg/m2 and myosteatosis was significantly higher in patients with dynapenia [9/14 (64.3%)] than in those with sarcopenia [2/23 (8.7%), P = 0.004] and pre-sarcopenia [0/18 (0%), P < 0.001] and tended to be higher than those with normal muscle status [16/61 (26.2%), P = 0.065]. The physical QOL in patients with dynapenia was as low as that in those with sarcopenia and significantly lower than that in those with normal muscle status. CONCLUSION: Cirrhotic patients with dynapenia had high BMI and myosteatosis, and impaired physical QOL.

A Rare Case of Epidermoid Cyst in the Pancreatic Tail Invaginated from the Splenic Hilum: The Long-term Changes in the Imaging Findings.

Epidermoid cysts arising from both the pancreas and spleen are rare. We herein report a case of a surgically resected epidermoid cyst in the pancreatic tail invaginated from the spleen. A multi-locular cyst, 2 cm in diameter, without a solid component was discovered incidentally in the pancreatic tail. During the 11-year follow-up, the emergence of satellite cystic lesions with distinct appearances was seen, and surgical resection was selected despite the lack of any associated symptoms or evidence of cytological abnormalities. Histologically, these cysts were lined with benign multi-layered flattened epithelium surrounded by a thin layer consisting of cells positive for CD8 and CD68 and connecting to the spleen.

Primary Poorly Differentiated Squamous Cell Carcinoma of the Extrahepatic Bile Duct.

Squamous cell carcinoma of the extrahepatic bile duct is quite rare. A 77-year-old woman with jaundice and general fatigue was referred to our hospital. Multiphase contrast-enhanced computed tomography visualized a 17-mm solid mass in the junction of the cystic and common bile ducts. The patient underwent pylorus-preserving pancreaticoduodenectomy. The pathological findings demonstrated keratin-positive poorly differentiated squamous cell carcinoma of the extrahepatic bile duct (T3N0M0, stage IIIA). Although adjuvant chemotherapy with gemcitabine was administered, the patient exhibited local recurrence at the site of anastomosis of biliojejunostomy 20 months after resection and died 32 months after resection.

Multiple Cancers of the Biliary Tract and Pancreatic Duct after Cholecystectomy for Gallbladder Cancer in a Patient with Pancreaticobiliary Maljunction.

We herein report the rare case of a 76-year-old woman who underwent cholecystectomy with bile duct resection for advanced gallbladder cancer associated with pancreaticobiliary maljunction (PBM) and subsequently developed multiple cancers of the pancreaticobiliary system (the distal bile duct, intrahepatic duct and pancreatic duct) after the operation. We performed conventional endoscopic retrograde cholangiopancreatography (ERCP) using a side-viewing scope to evaluate the masses in the distal bile duct and the pancreatic duct. We also performed ERCP using double-balloon enteroscopy (DBE) to observe the mass in the intrahepatic duct. It was possible to directly observe the lesion using DBE and to perform a biopsy under visual control. All lesions were correctly diagnosed by the combination of ERCP using different endoscopes. The present case suggests that it is necessary to pay close attention (with regard to carcinogenesis) to the whole pancreaticobiliary system in patients with PBM. In addition, the combination of ERCP using DBE and a side-viewing scope may be useful for making a precise diagnosis in patients with altered biliary anatomy who have multiple cancers of the pancreaticobiliary system.

Ex vivo activation of angiogenic property in human peripheral blood-derived monocytes by thrombopoietin.

Human peripheral blood mononuclear cells (PB-MNCs) have angiogenic properties, which make them promising cells for use in angiogenic therapy approaches in regenerative medicine. To explore an efficient method for expanding pro-angiogenic cells from PB-MNCs, we developed a novel serum-free culture system composed of X-VIVO15 medium supplemented with vascular endothelial growth factor, basic fibroblast growth factor, and thrombopoietin (TPO). Using this ex vivo culture, we obtained floating spheres composed mainly of CD11b(+) monocytes expressing c-Mpl (TPO receptor) and which exhibited acetylated low-density lipoprotein uptake and phagocytosis. Expression of IL-8, CXCR4, and vasohibin-2 mRNA was upregulated in these cells. In the presence of TPO, the number and size of the spheres were increased. In a nude mouse hind-limb ischemia model, the intramuscular injection of spheroid cells treated with TPO rescued blood perfusion more effectively than that without TPO. These results indicate that the ex vivo addition of TPO augments the pro-angiogenic activity of peripheral CD11b(+) monocytes, suggesting that this method shows promise for uses in human cell therapy aimed at the induction of vascular regeneration by activating the angiogenic properties of human peripheral blood-derived monocytes.

Transplanting normal vascular proangiogenic cells to tumor-bearing mice triggers vascular remodeling and reduces hypoxia in tumors.

Blood vessels deliver oxygen and nutrients to tissues, and vascular networks are spatially organized to meet the metabolic needs for maintaining homeostasis. In contrast, the vasculature of tumors is immature and leaky, resulting in insufficient delivery of nutrients and oxygen. Vasculogenic processes occur normally in adult tissues to repair "injured" blood vessels, leading us to hypothesize that bone marrow mononuclear cells (BMMNC) may be able to restore appropriate vessel function in the tumor vasculature. Culturing BMMNCs in endothelial growth medium resulted in the early outgrowth of spindle-shaped attached cells expressing CD11b/Flt1/Tie2/c-Kit/CXCR4 with proangiogenic activity. Intravenous administration of these cultured vascular proangiogenic cells (VPC) into nude mice bearing pancreatic cancer xenografts and Pdx1-Cre;LSL-Kras(G12D);p53(lox/+) genetically engineered mice that develop pancreatic ductal adenocarcinoma significantly reduced areas of hypoxia without enhancing tumor growth. The resulting vasculature structurally mimicked normal vessels with intensive pericyte coverage. Increases in vascularized areas within VPC-injected xenografts were visualized with an ultrasound diagnostic system during injection of a microbubble-based contrast agent (Sonazoid), indicating a functional "normalization" of the tumor vasculature. In addition, gene expression profiles in the VPC-transplanted xenografts revealed a marked reduction in major factors involved in drug resistance and "stemness" of cancer cells. Together, our findings identify a novel alternate approach to regulate abnormal tumor vessels, offering the potential to improve the delivery and efficacy of anticancer drugs to hypoxic tumors.

Incidence of synchronous and metachronous pancreatic carcinoma in 168 patients with branch duct intraductal papillary mucinous neoplasm.

BACKGROUND/AIMS: Although branch duct intraductal papillary mucinous neoplasms of the pancreas (BD-IPMN) are being diagnosed with increasing frequency, the incidence of concomitant pancreatic carcinoma (PC) is not well known. We investigated the incidence and clinical features of synchronous and metachronous PC in patients with BD-IPMN. METHODS: We studied 168 BD-IPMN patients diagnosed by various imaging modalities, including endoscopic retrograde pancreatography, between 1990 and 2008. We reviewed the medical records and clinical features in both patients developing and not developing PC. The diagnosis of PC was histologically verified in all patients. RESULTS: PC was observed in 9 (5.4%) of 168 patients. Five were synchronously detected at the time of BD-IPMN diagnosis, whereas four were metachronously identified during the follow-up period. All PCs occurred in regions separate from the BD-IPMN lesion. All PCs represented histologically invasive ductal adenocarcinomas, whereas the BD-IPMN lesion was diagnosed as adenoma. Patients developing PC were significantly older than patients not developing PC (p = 0.017). The diameters of the BD-IPMN lesions and main pancreatic ducts were significantly smaller in patients developing PC than patients not developing PC (p = 0.013 and p < 0.001, respectively). CONCLUSIONS: It was not infrequent for PC to occur in the pancreas with BD-IPMN. Particular attention should therefore be paid to the development of PC, even in low-risk BD-IPMN, as well as to changes in BD-IPMN.

[A case of pancreatic carcinoma presenting as pancreaticopleural fistula with pancreatic pleural effusion].

A 63-year-old man was admitted with left pleural effusion, and an amylase level of 30994IU/l. A diagnosis of pancreaticopleural fistula was made, based on the findings of magnetic resonance cholangiopancreatography and endoscopic retrograde pancreatography (ERP). After the placement of an endoscopic naso-pancreatic drainage tube, the pleural effusion markedly reduced. When ERP was performed for internal drainage, the main pancreatic duct and stricture were biopsied and showed pancreatic ductal adenocarcinoma histologically. CT revealed a mass in the head of the pancreas. He underwent pylorus-preserving pancreaticoduodenectomy. To the best of our knowledge this is the first case of pancreatic carcinoma presenting as pancreaticopleural fistula with pancreatic pleural effusion. Clinicians should pay attention to the possible presence of cancer and pancreaticopleural fistula in patients with pancreatic pleural effusion.

Hedgehog promotes neovascularization in pancreatic cancers by regulating Ang-1 and IGF-1 expression in bone-marrow derived pro-angiogenic cells.

BACKGROUND: The hedgehog (Hh) pathway has been implicated in the pathogenesis of cancer including pancreatic ductal adenocarcinoma (PDAC). Recent studies have suggested that the oncogenic function of Hh in PDAC involves signaling in the stromal cells rather than cell autonomous effects on the tumor cells. However, the origin and nature of the stromal cell type(s) that are responsive to Hh signaling remained unknown. Since Hh signaling plays a crucial role during embryonic and postnatal vasculogenesis, we speculated that Hh ligand may act on tumor vasculature specifically focusing on bone marrow (BM)-derived cells. METHODOLOGY/PRINCIPAL FINDINGS: Cyclopamine was utilized to inhibit the Hh pathway in human PDAC cell lines and their xenografts. BM transplants, co-culture systems of tumor cells and BM-derived pro-angiogenic cells (BMPCs) were employed to assess the role of tumor-derived Hh in regulating the BM compartment and the contribution of BM-derived cells to angiogenesis in PDAC. Cyclopamine administration attenuated Hh signaling in the stroma rather than in the cancer cells as reflected by decreased expression of full length Gli2 protein and Gli1 mRNA specifically in the compartment. Cyclopamine inhibited the growth of PDAC xenografts in association with regression of the tumor vasculature and reduced homing of BM-derived cells to the tumor. Host-derived Ang-1 and IGF-1 mRNA levels were downregulated by cyclopamine in the tumor xenografts. In vitro co-culture and matrigel plug assays demonstrated that PDAC cell-derived Shh induced Ang-1 and IGF-1 production in BMPCs, resulting in their enhanced migration and capillary morphogenesis activity. CONCLUSIONS/SIGNIFICANCE: We identified the BMPCs as alternative stromal targets of Hh-ligand in PDAC suggesting that the tumor vasculature is an attractive therapeutic target of Hh blockade. Our data is consistent with the emerging concept that BM-derived cells make important contributions to epithelial tumorigenesis.

Pancreatic ductal adenocarcinomas in long-term follow-up patients with branch duct intraductal papillary mucinous neoplasms.

OBJECTIVE: Although branch duct intraductal papillary mucinous neoplasms (BD-IPMNs) are slow-growing tumors with a favorable prognosis, the synchronous occurrence of pancreatic ductal adenocarcinomas (PDAs) in patients with BD-IPMNs has been reported. This study was aimed to elucidate the development of PDAs in long-term follow-up patients with BD-IPMNs. METHODS: We investigated 89 BD-IPMN patients who had no mural nodules and followed them up conservatively at least 2 years (median follow-up, 64 months; range, 25-158 months). All subjects underwent examinations by imaging modalities including endoscopic retrograde pancreatography. We calculated the standardized incidence ratio (SIR) from the vital statistics compiled by the Ministry of Health, Labor, and Welfare of Japan. RESULTS: Among the 89 patients, 4 cases of PDAs distant from BD-IPMN were observed in 552 patient-years of follow-up (7.2 per 1000 patient-years). The expected number was 0.25, and the SIR of PDAs was 15.8 (95% confidence interval [CI], 4.3-40.4; P = 0.00014). Subgroup analyses showed that the incidence of PDAs was significantly increased in patients 70 years or older (SIR 16.7; 95% CI, 3.4-48.7; P = 0.0008) and in women (SIR 22.5; 95% CI, 2.7-81.1; P = 0.0037). CONCLUSIONS: Patients with BD-IPMNs are at a high risk for PDAs. During the follow-up, careful examination is required to detect the development of PDAs in patients with BD-IPMNs.

[Gemcitabine in combination with S-1 or UFT in patients with advanced pancreatic cancer].

The present retrospective study aimed to evaluate the anti-tumor activity and toxicity of combination chemotherapy with gemcitabine (GEM) and oral S-1 or UFT in patients with advanced or metastatic pancreatic cancer. Ninety-four patients received chemotherapy. Among them, sixty-three were treated with GEM alone, twenty-two with UFT and GEM (UFT/GEM), and nine with S-1 and GEM(S-1/GEM). The median survival time was 8.7 months with GEM, 7.3 months with UFT/GEM, and 23.3 months with S-1/GEM. The overall response rate was 11.1%, 10.0%, and 22.2%, respectively. The 1-year survival rate was 29.5%, 36.4%, and 85.7%, respectively. Although the treatment-related adverse effects were not infrequent in patients treated with S-1/GEM, they were moderate in intensity. The combination chemotherapy with S-1/GEM was well tolerated and yielded a high response rate in patients with pancreatic cancer.

Sonic hedgehog derived from human pancreatic cancer cells augments angiogenic function of endothelial progenitor cells.

Hedgehog signaling is important in the pathogenesis of pancreatic cancer. Several recent observations suggest the involvement of sonic hedgehog (SHH) in postnatal neovascularization. We identified a novel role for SHH in tumor-associated angiogenesis in pancreatic cancer. Immunohistochemical analysis revealed that patched homolog 1 (PTCH1), both a receptor for and transcriptional target of hedgehog signaling, was expressed in a small fraction of endothelial cells within pancreatic cancer, but not in normal pancreatic tissue. When endothelial progenitor cells (EPC) isolated from human peripheral blood were cultured with supernatant from SHH-transfected 293 cells or pancreatic cancer cells, mRNA levels of vascular endothelial growth factor (VEGF), stromal cell-derived factor-1 and angiopoietin-1 were significantly increased, whereas no such induction was observed in human umbilical vein endothelial cell (HUVEC) and human dermal microvascular endothelial cell (HMVEC). HUVEC tube formation was stimulated when cocultured with EPC, and preconditioning EPC with supernatant from KP-1 N pancreatic cancer cells highly expressing SHH significantly enhanced the effect. The effect was partially attenuated by specific inhibition of SHH with cyclopamine or a neutralizing antibody. These findings suggest that tumor-derived SHH can induce angiogenesis, and this is mediated by its effects on EPC specifically. Targeting SHH would be a novel therapeutic approach that can inhibit not only proliferation of cancer cells but also EPC-mediated angiogenesis.

Identification of transmembrane protein functions by binary topology patterns.

We propose a novel method for identifying and classifying the functions of transmembrane (TM) proteins based on their TM topology [the number of TM segments (tms), the loop length and the N-terminus location]. In this method, the TM topology is expressed as a string of '0' and '1', and this is designated the binary topology pattern (BTP). We focused on TM proteins with up to 12 tms, with the exception of 1 and 9 tms, and classified them into 37 functional groups by the number of tms and the functional annotation. These grouped TM protein sequences were used to determine BTPs which are specific to the individual functional groups. Since the evaluated accuracies (sensitivity, specificity and self-consistency) of these patterns in functional identification were quite high overall, i.e. 0.940, 0.934 and 0.935, respectively, as averaged over the 37 functional groups, we confirmed that TM protein function can be identified by the number of tms and the characteristics of loop lengths, i.e. BTPs.

Rapid appearance of pathological changes of neurons and glia cells in the cerebellum of microsphere-embolized rats.

Neuropathological changes in the cerebellar cortex of microsphere-embolized rats were studied at 30 min and 3 h after the embolism. Necrotic processes including a sponge-like vacuolation in the molecular layer, a vague outline of some Purkinje cells, and a few pyknotic granule cells having small and dark profiles were identified at sometime between 30 min and 3 h after microsphere-induced embolism in Nissl staining. Glial fibrillary acidic protein staining shows an apparent reduction in the number of Bergmann glial processes in some of the areas where there was necrosis of the molecular layer and poor astroglia processes in the areas subjacent to the pyknotic granule cells. These data demonstrate that within a short time, microsphere-induced cerebral ischemia produces necrosis of cerebellar neurons (i.e. Purkinje and granule cells) and changes in cerebellar glia cells (i.e. Bergmann and astroglia cells), and that these neuropathological changes are secondary phenomenon caused by microsphere blockage of cerebellar blood flow.