Takuma Arai

Vascular Endothelial Adrenomedullin-RAMP2 System Is Essential for Vascular Integrity and Organ Homeostasis

Background— Revealing the mechanisms underlying the functional integrity of the vascular system could make available novel therapeutic approaches. We previously showed that knocking out the widely expressed peptide adrenomedullin (AM) or receptor activity-modifying protein 2 (RAMP2), an AM-receptor accessory protein, causes vascular abnormalities and is embryonically lethal. Our aim was to investigate the function of the vascular AM-RAMP2 system directly. Methods and Results— We generated endothelial cell–specific RAMP2 and AM knockout mice (E-RAMP2−/− and E-AM−/−). Most E-RAMP2−/− mice died perinatally. In surviving adults, vasculitis occurred spontaneously. With aging, E-RAMP2−/− mice showed severe organ fibrosis with marked oxidative stress and accelerated vascular senescence. Later, liver cirrhosis, cardiac fibrosis, and hydronephrosis developed. We next used a line of drug-inducible E-RAMP2−/− mice (DI-E-RAMP2−/−) to induce RAMP2 deletion in adults, which enabled us to analyze the initial causes of the aforementioned vascular and organ damage. Early after the induction, pronounced edema with enhanced vascular leakage occurred. In vitro analysis revealed the vascular leakage to be caused by actin disarrangement and detachment of endothelial cells. We found that the AM-RAMP2 system regulates the Rac1-GTP/RhoA-GTP ratio and cortical actin formation and that a defect in this system causes the disruption of actin formation, leading to vascular and organ damage at the chronic stage after the gene deletion. Conclusions— Our findings show that the AM-RAMP2 system is a key determinant of vascular integrity and homeostasis from prenatal stages through adulthood. Furthermore, our models demonstrate how endothelial cells regulate vascular integrity and how their dysregulation leads to organ damage.

Adrenomedullin in sinusoidal endothelial cells play protective roles against cold injury of liver.

Donor organ damage caused by cold preservation is a major problem affecting liver transplantation. Cold preservation most easily damages liver sinusoidal endothelial cells (LSECs), and information about the molecules modulating LSECs function can provide the basis for new therapeutic strategies. Adrenomedullin (AM) is a peptide known to possess anti-apoptotic and anti-inflammatory properties. AM is abundant in vascular endothelial cells, but levels are comparatively low in liver, and little is known about its function there. In this study, we demonstrated both AM and its receptors are expressed in LSECs. AM treatment reduced LSECs loss and apoptosis under cold treatment. AM also downregulated cold-induced expression of TNFalpha, IL1beta, IL6, ICAM1 and VCAM1. AM reduced apoptosis and expression of ICAM1 and VCAM1 in an in vivo liver model subjected to cold storage. Conversely, apoptosis was exacerbated in livers from AM and RAMP2 (AM receptor activity-modifying protein) knockout mice. These results suggest that AM expressed in LSECs exerts a protective effect against cold-organ damage through modulation of apoptosis and inflammation.

Functional differentiation of RAMP2 and RAMP3 in their regulation of the vascular system

Adrenomedullin (AM) is a vasoactive peptide that possesses various bioactivities. AM receptors are dimers consisting of CLR with one of two accessory proteins, RAMP2 or RAMP3. The functional difference between CLR/RAMP2 and CLR/RAMP3 and the relationship between the two receptors remain unclear. To address these issues, we generated RAMP2 and RAMP3 knockout (-/-) mice and have been studying their physiological activities in the vascular system. AM-/- and RAMP2-/- mice die in utero due to blood vessel abnormalities, which is indicative of their essential roles in vascular development. In contrast, RAMP3-/- mice were born normally without any major abnormalities. In adult RAMP3-/- mice, postnatal angiogenesis was normal, but lymphangiography using indocyanine green (ICG) showed delayed drainage of subcutaneous lymphatic vessels. Moreover, chyle transport by intestinal lymphatics was delayed in RAMP3-/- mice, which also showed more severe interstitial edema than wild-type mice in a tail lymphedema model, with characteristic dilatation of lymphatic capillaries and accumulation of inflammatory cells. In scratch-wound assays, migration of isolated RAMP3-/- lymphatic endothelial cells was delayed as compared to wild-type cells, and AM administration failed to enhance the re-endothelialization. The delay in re-endothelialization was due to a primary migration defect rather than a decrease in proliferation. These results suggest that RAMP3 regulates drainage through lymphatic vessels, and that the AM-RAMP3 system could be a novel therapeutic target for controlling postoperative lymphedema.

Impact of advanced age on the short- and long-term outcomes in patients undergoing hepatectomy for hepatocellular carcinoma: a single-center analysis over a 20-year period

BACKGROUND The purpose of this study was to analyze the influence of age on both the risk of hepatectomy and the prognosis in patients with hepatocellular carcinoma (HCC). METHODS Patients undergoing an initial hepatectomy for HCC were classified into 2 age groups: 75 years or over (n = 113) and less than 75 years (n = 499). RESULTS A zero 90-day mortality was achieved in the elderly. Although the recurrence rate and recurrence sites were almost similar between the 2 groups, the 5-year survival rate in the elderly patients was significantly lower than that in the younger patients (46.0% vs 57.6%; P = .018), possibly because of the higher incidence of deaths from other causes (26.8% vs 10.4%; P = .011) in the elderly. CONCLUSION Selected elderly HCC patients can undergo a hepatectomy safely and can benefit from long-term HCC control comparable with that of their younger counterparts.

Regulation of adrenomedullin and its family peptide by RAMP system--lessons from genetically engineered mice.

Adrenomedullin (ADM), originally identified as a vasodilating peptide, is now recognized to be a pleiotropic molecule involved in both the pathogenesis of cardiovascular diseases and circulatory homeostasis. Homozygotes of ADM knockout mice (ADM-/-) were lethal at mid-gestation with abnormalities of vascular development and this finding clarified the angiogenic potency of ADM. Calcitonin gene-related peptide (CGRP), which has a structure and function similar to that of ADM, has been identified as a family peptide of ADM. Unlike ADM-/-, CGRP-/- were apparently normal. Therefore, the study of knockout mice first clarified the distinctly different physiological roles between ADM and CGRP. In contrast, heterozygotes of ADM knockout mice (ADM+/-) were alive but showed blood pressure elevation, reduced neovascularization, and enhanced neointimal formation by arterial injury. Based on these observations, there was hope ADM would have a therapeutic use. However, ADM has a short half-life in the blood stream and its application in chronic disease has limitations. Therefore, we focused on the ADM receptor system. The calcitonin-receptor-like receptor (CLR), which is the ADM receptor, associates with one of the accessory proteins, called receptor activity-modifying proteins (RAMPs). By interacting with RAMP1, CLR exhibits a high affinity for CGRP, whereas by interacting with either RAMP2 or -3, CLR exhibits a high affinity for ADM. We generated RAMP knockout mice and found that vascular phenotypes similar to ADM-/- were reproduced only in RAMP2-/-. This shows that RAMP2 is the key determinant of the vascular functions of ADM. RAMP2 could be an attractive therapeutic target in cardiovascular diseases.

Hepatocellular carcinoma associated with sarcoidosis

INTRODUCTION The association of hepatic sarcoidosis with hepatocellular carcinoma (HCC) is considerably rare. Here we report a rare case of HCC associated with sarcoidosis. PRESENTATION OF CASE A 75-year-old male with no history of alcohol addiction or viral hepatitis was referred to our hospital because of an abnormal liver mass. Subsegmentectomy of the liver was performed for the diagnosis of HCC. A histopathological examination revealed small non-necrotizing granulomas with a tendency to coalesce that were scattered in and around the carcinoma. No features of cirrhosis, steatohepatitis, and any other liver diseases were observed. Furthermore, swelling of the bilateral lung hilar lymph nodes with uptake of 18F-fluorodeoxyglucose was found on positron emission tomography/computed tomography and the tuberculin reaction test results were negative. On the basis of these findings, the final diagnosis of HCC associated with sarcoidosis was confirmed. DISCUSSION By reviewing previous cases, we found only five cases that described patients diagnosed with HCC associated with sarcoidosis. Of these, four patients died within two years after diagnosis because of ruptures or inoperable huge tumors. In contrast, radical hepatectomy was performed at an earlier stage of disease in two patients, including ours, and both these patients have remained healthy with no recurrences or metastases at the latest follow-up visit. CONCLUSION Periodic checkups of the liver should be conducted for patients with systemic sarcoidosis, regardless of the presence of liver cirrhosis.

Efficacy of self-expanding metallic stent for right-sided colonic obstruction due to carcinoma before 1-stage laparoscopic surgery.

Purpose: The aim of this small study was to assess the clinical outcomes of self-expanding metallic stent (SEMS) placement followed by 1-stage laparoscopic resection for treatment of acute right-sided colonic obstruction due to carcinoma. Method: From January 2012 to December 2012, we performed 1-stage laparoscopic colectomy after placement of SEMS for right-sided obstructive colon carcinoma in 4 patients. Results: SEMS placement was technically successful in all cases and symptoms were immediately relieved. The presence of the endoluminal stent did not prevent a laparoscopic approach; moreover, decompression of the bowel obstruction provided a wide field of view and working space for the laparoscopic procedure. There were no cases of conversion to laparotomy and no intraoperative complications. Conclusion: One-stage laparoscopic surgery, as a minimally invasive therapy, can be safely performed by SEMS placement in patients with acute right-sided colonic obstruction due to carcinoma. A large sample study is required to confirm the efficacy of this procedure.

Induction of LYVE-1/stabilin-2-positive liver sinusoidal endothelial-like cells from embryoid bodies by modulation of adrenomedullin-RAMP2 signaling.

Embryonic stem cells (ESCs) are a useful source for various cell lineages. So far, however, progress toward reconstitution of mature liver morphology and function has been limited. We have shown that knockout mice deficient in adrenomedullin (AM), a multifunctional endogenous peptide, or its receptor-activity modifying protein (RAMP2) die in utero due to poor vascular development and hemorrhage within the liver. In this study, using embryoid bodies (EBs)-culture system, we successfully induced liver sinusoidal endothelial-like cells by modulation of AM-RAMP2. In an EB differentiation system, we found that co-administration of AM and SB431542, an inhibitor of transforming growth factor β (TGFβ) receptor type 1, markedly enhanced differentiation of lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1)/stabilin-2-positive endothelial cells. These cells showed robust endocytosis of acetylated low-density lipoprotein (Ac-LDL) and upregulated expression of liver sinusoidal endothelial cells (LSECs)-specific markers, including factor 8 (F8), Fc-γ receptor 2b (Fcgr2b), and mannose receptor C type 1 (Mrc1), and also possessed fenestrae-like structure, a key morphological feature of LSECs. In RAMP2-null liver, by contrast, LYVE-1 was downregulated in LSECs, and the sinusoidal structure was disrupted. Our findings highlight the importance of AM-RAMP2 signaling for development of LSECs.

Cystic mixed adenoneuroendocrine carcinoma of the pancreas: A case report

Highlights • There are few reports about pancreatic MANEC with cystic features.• Mixed tumor of the pancreas may arise from totipotent stem cells.• The treatment strategy is unclear, but surgery may be the first choice if possible.