Impact of Lifestyle Behaviors on Postprandial Hyperglycemia during Continuous Glucose Monitoring in Adult Males with Overweight/Obesity but without Diabetes.

Data regarding hyperglycemia-related factors were scarce in people without diabetes. Fifty males (age 50-65 years) with overweight/obesity but without diagnosis of diabetes were recruited. After excluding participants with the 2 h plasma glucose value during a 75 g oral glucose tolerance test ≥200 mg/dL, continuous glucose monitoring (CGM) was performed for 6 days. Subjects with ≥1800 CGM readings were included (n = 36). The CGM indices of hyperglycemia were significantly associated with disposition index and snacking frequency. In receiver-operating characteristic analysis for predicting the maximal CGM glucose ≥200 mg/dL, the area under curves of disposition index, snacking frequency, and minimal daily step counts during the study were 0.69, 0.63, and 0.68, whereas the cutoff values were 1.57, once daily, and 2499 steps, respectively. After adjustments, the lower disposition index (≤1.57), higher snacking frequency (≥1 per day), and lower minimal step (≤2499 steps per day) categories conferred 14.5, 14.5, and 6.6-fold increased probabilities for having the maximum level ≥ 200 mg/dL, respectively. In addition, the snacking habits were significantly associated with insulin resistance and compensatory hyperinsulinemia. In conclusion, in middle aged males with overweight/obesity but without diabetes, snacking and physical inactivity serve as the major drivers of postprandial hyperglycemia independently of ß-cell function.

A Case of Recurrent Hemorrhagic Corpus Luteum with Elevated Follicle-Stimulating Hormone, Controlled by Estrogen/Gestagen Therapy.

A high secretion of follicle-stimulating hormone (FSH) in reproductive-aged women is unusual. We report a case of recurrent corpus luteum hemorrhage and subsequent ovarian torsion with markedly elevated FSH levels in a reproductive-aged woman in the absence of functional gonadotroph adenoma (FGA) or premature ovarian failure (POF). A 22-year-old nulligravid woman with a history of bilateral hemorrhagic corpus luteum and subsequent ovarian torsion presented with acute abdominal pain. An emergency salpingo-oophorectomy of the right side was performed, and the right ovarian torsion due to hemorrhagic corpus luteum was diagnosed. Laboratory tests revealed markedly elevated FSH levels (77.6 mIU/mL). FGA was suspected, but no evidence of tumor was identified. The left ovary enlarged again at one-month follow-up. Estrogen/gestagen therapy (EGT) was started, which reduced the enlarged ovary to normal size. Two years later, her pituitary hormonal status was evaluated in detail. Besides markedly elevated FSH level, slightly elevated LH (31.2 mIU/mL), normal total inhibin B (35.3 pg/ml), abnormally low anti-Müllerian hormone (AMH) (<0.03 ng/mL), and poor FSH response to gonadotropin-releasing hormone stimulation test were found. In the absence of FGA, we conclude that certain disorders of inhibitory factors for FSH function, including inhibin and AMH may exist, which could attribute to the patient's symptoms. EGT was very effective in suppressing the ovarian hyperactivity.

Plasma brain natriuretic peptide levels are elevated in patients with cancer.

BACKGROUND: Natriuretic peptides have been proposed as biomarkers of cardiovascular disease, especially heart failure. Brain natriuretic peptide (BNP) has also been shown to be upregulated at the transcriptional and translational levels by pro-inflammatory cytokines in cardiac myocytes. Although we often measure plasma BNP levels in cancer patients, it remains unknown whether cancer-related inflammation affects the plasma BNP levels. We investigated the relationship between the BNP and human cancers. METHODS: We retrospectively studied 2,923 patients in whom the plasma BNP levels and serum C-reactive protein (CRP) were measured and echocardiography was performed. Patients with clinically evident heart failure (NYHA II or higher), heart disease requiring medical treatment or surgery, renal dysfunction, and inflammatory disease were excluded. There were 234 patients in the final analysis. Blood sampling was performed before surgery and chemotherapy. In addition, we evaluated the relationship between the inflammation and plasma BNP levels in mouse models of colon cancer. RESULTS: Of the 234 patients, 80 were diagnosed with cancer. Both the plasma BNP and serum CRP levels were significantly higher in cancer patients than those without. There were no significant differences in the echocardiographic parameters. There was a significant positive correlation between the plasma BNP and serum CRP levels in cancer patients (r = 0.360, P<0.01) but not in those without. In cancer patients, only the CRP correlated with the BNP independent of the age, creatinine level, hypertension, and body mass index. In addition, in nude mice with subcutaneous colon cancer, the plasma BNP level was elevated compared with that in non-cancer mice, and there was a significant relationship between the plasma BNP and serum levels of the inflammatory markers. CONCLUSIONS: In cancer patients, as well as colon cancer model mice, the plasma BNP levels were elevated, possibly due to cancer-related inflammation. The effect of cancer on the BNP levels should be considered when using BNP as an indicator of heart failure in cancer patients.

Ghrelin and Blood Pressure Regulation.

Ghrelin is a growth hormone-releasing polypeptide that was first isolated from the rat stomach in 1999. High expression of growth hormone secretagogue receptor, the ghrelin receptor, in the heart, kidney, and blood vessels provides evidence of ghrelin activity in blood pressure regulation. Circulating ghrelin concentrations are reported to be inversely correlated with blood pressure, and the acute and chronic effects of ghrelin in decreasing blood pressure have been reported in animals with normal blood pressure, healthy individuals, animals and patients with heart failure, and animals with hypertension. The mechanism by which ghrelin regulates blood pressure appears to be related to modulation of the autonomic nervous system, direct vasodilatory activities, and kidney diuresis. Thus, modulation of the signaling pathway through ghrelin may provide a novel concept for treating hypertension. In this review, we discuss the current evidence and potential mechanisms of ghrelin activity in blood pressure regulation.

Importance of Endogenous Atrial and Brain Natriuretic Peptides in Murine Embryonic Vascular and Organ Development.

Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) bind to the receptor guanylyl cyclase (GC)-A, leading to diuresis, natriuresis, and blood vessel dilation. In addition, ANP and BNP have various angiogenic properties in ischemic tissue. When breeding mice devoid of GC-A, we noted significant skewing of the Mendelian ratio in the offspring, suggesting embryonic lethality due to knockout of GC-A. Consequently, we here investigated the roles of endogenous ANP and BNP in embryonic neovascularization and organ morphogenesis. Embryos resulting from GC-A(-/-) × GC-A(+/-) crosses developed hydrops fetalis (HF) beginning at embryonic day (E)14.5. All embryos with HF had the genotype GC-A(-/-). At E17.5, 33.3% (12 of 36) of GC-A(-/-) embryos had HF, and all GC-A(-/-) embryos with HF were dead. Beginning at E16.0, HF-GC-A(-/-) embryos demonstrated poorly developed superficial vascular vessels and sc hemorrhage, the fetal side of the placenta appeared ischemic, and vitelline vessels on the yolk sac were poorly developed. Furthermore, HF-GC-A(-/-) embryos also showed abnormal constriction of umbilical cord vascular vessels, few cardiac trabeculae and a thin compact zone, hepatic hemorrhage, and poor bone development. Electron microscopy of E16.5 HF-GC-A(-/-) embryos revealed severe vacuolar degeneration in endothelial cells, and the expected 3-layer structure of the smooth muscle wall of the umbilical artery was indistinct. These data demonstrate the importance of the endogenous ANP/BNP-GC-A system not only in the neovascularization of ischemic tissues but also in embryonic vascular development and organ morphogenesis.

Characterization of Autosomal Dominant Hypercholesterolemia Caused by PCSK9 Gain of Function Mutations and Its Specific Treatment With Alirocumab, a PCSK9 Monoclonal Antibody.

BACKGROUND: Patients with PCSK9 gene gain of function (GOF) mutations have a rare form of autosomal dominant hypercholesterolemia. However, data examining their clinical characteristics and geographic distribution are lacking. Furthermore, no randomized treatment study in this population has been reported. METHODS AND RESULTS: We compiled clinical characteristics of PCSK9 GOF mutation carriers in a multinational retrospective, cross-sectional, observational study. We then performed a randomized placebo-phase, double-blind study of alirocumab 150 mg administered subcutaneously every 2 weeks to 13 patients representing 4 different PCSK9 GOF mutations with low-density lipoprotein cholesterol (LDL-C) ≥70 mg/dL on their current lipid-lowering therapies at baseline. Observational study: among 164 patients, 16 different PCSK9 GOF mutations distributed throughout the gene were associated with varying severity of untreated LDL-C levels. Coronary artery disease was common (33%; average age of onset, 49.4 years), and untreated LDL-C concentrations were higher compared with matched carriers of mutations in the LDLR (n=2126) or apolipoprotein B (n=470) genes. Intervention study: in PCSK9 GOF mutation patients randomly assigned to receive alirocumab, mean percent reduction in LDL-C at 2 weeks was 62.5% (P<0.0001) from baseline, 53.7% compared with placebo-treated PCSK9 GOF mutation patients (P=0.0009; primary end point). After all subjects received 8 weeks of alirocumab treatment, LDL-C was reduced by 73% from baseline (P<0.0001). CONCLUSIONS: PCSK9 GOF mutation carriers have elevated LDL-C levels and are at high risk of premature cardiovascular disease. Alirocumab, a PCSK9 antibody, markedly lowers LDL-C levels and seems to be well tolerated in these patients. CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov. Unique Identifier: NCT01604824.

Endogenous ghrelin attenuates pressure overload-induced cardiac hypertrophy via a cholinergic anti-inflammatory pathway.

Cardiac hypertrophy, which is commonly caused by hypertension, is a major risk factor for heart failure and sudden death. Endogenous ghrelin has been shown to exert a beneficial effect on cardiac dysfunction and postinfarction remodeling via modulation of the autonomic nervous system. However, ghrelin's ability to attenuate cardiac hypertrophy and its potential mechanism of action are unknown. In this study, cardiac hypertrophy was induced by transverse aortic constriction in ghrelin knockout mice and their wild-type littermates. After 12 weeks, the ghrelin knockout mice showed significantly increased cardiac hypertrophy compared with wild-type mice, as evidenced by their significantly greater heart weight/tibial length ratios (9.2±1.9 versus 7.9±0.8 mg/mm), left ventricular anterior wall thickness (1.3±0.2 versus 1.0±0.2 mm), and posterior wall thickness (1.1±0.3 versus 0.9±0.1 mm). Furthermore, compared with wild-type mice, ghrelin knockout mice showed suppression of the cholinergic anti-inflammatory pathway, as indicated by reduced parasympathetic nerve activity and higher plasma interleukin-1ß and interleukin-6 levels. The administration of either nicotine or ghrelin activated the cholinergic anti-inflammatory pathway and attenuated cardiac hypertrophy in ghrelin knockout mice. In conclusion, our results show that endogenous ghrelin plays a crucial role in the progression of pressure overload-induced cardiac hypertrophy via a mechanism that involves the activation of the cholinergic anti-inflammatory pathway.

Atrial natriuretic peptide prevents cancer metastasis through vascular endothelial cells.

Most patients suffering from cancer die of metastatic disease. Surgical removal of solid tumors is performed as an initial attempt to cure patients; however, surgery is often accompanied with trauma, which can promote early recurrence by provoking detachment of tumor cells into the blood stream or inducing systemic inflammation or both. We have previously reported that administration of atrial natriuretic peptide (ANP) during the perioperative period reduces inflammatory response and has a prophylactic effect on postoperative cardiopulmonary complications in lung cancer surgery. Here we demonstrate that cancer recurrence after curative surgery was significantly lower in ANP-treated patients than in control patients (surgery alone). ANP is known to bind specifically to NPR1 [also called guanylyl cyclase-A (GC-A) receptor]. In mouse models, we found that metastasis of GC-A-nonexpressing tumor cells (i.e., B16 mouse melanoma cells) to the lung was increased in vascular endothelium-specific GC-A knockout mice and decreased in vascular endothelium-specific GC-A transgenic mice compared with control mice. We examined the effect of ANP on tumor metastasis in mice treated with lipopolysaccharide, which mimics systemic inflammation induced by surgical stress. ANP inhibited the adhesion of cancer cells to pulmonary arterial and micro-vascular endothelial cells by suppressing the E-selectin expression that is promoted by inflammation. These results suggest that ANP prevents cancer metastasis by inhibiting the adhesion of tumor cells to inflamed endothelial cells.

Removal of plasma mature and furin-cleaved proprotein convertase subtilisin/kexin 9 by low-density lipoprotein-apheresis in familial hypercholesterolemia: development and application of a new assay for PCSK9.

CONTEXT: Proprotein convertase subtilisin/kexin 9 (PCSK9) is known to be a good target to decrease LDL cholesterol (LDL-C) and two forms of PCSK9, mature and furin-cleaved PCSK9, circulate in blood. However, it has not been clarified whether and how the levels of each PCSK9 are affected by LDL-apheresis (LDL-A) treatment, a standard therapy in patients with severe forms of familial hypercholesterolemia (FH). OBJECTIVE: Our objective was to investigate the differences in LDL-A-induced reduction of mature and furin-cleaved PCSK9 between homozygous and heterozygous FH, and between dextran sulfate (DS) cellulose adsorption and double membrane (DM) columns and to clarify the mechanism of their removal. DESIGN: A sandwich ELISA to measure two forms of PCSK9s using monoclonal antibodies was developed. Using the ELISA, PCSK9 levels were quantified before and after LDL-A with DS columns in 7 homozygous and 11 heterozygous FH patients. A crossover study between the two column types was performed. The profiles of PCSK9s were analyzed after fractionation by gel filtration chromatography. Immunoprecipitation of apolipoprotein B (apoB) in FH plasma was performed. RESULTS: Both mature and furin-cleaved PCSK9s were significantly decreased by 55-56% in FH homozygotes after a single LDL-A treatment with DS columns, and by 46-48% or 48-56% in FH heterozygotes after treatment with DS or DM columns. The reduction ratios of LDL-C were strongly correlated with that of PCSK9 in both FH homozygotes and heterozygotes. In addition, more than 80% of plasma PCSK9s were in the apoB-deficient fraction and a significant portion of mature PCSK9 was bound to apoB, as shown by immunoprecipitation. CONCLUSIONS: Both mature and furin-cleaved PCSK9s were removed by LDL-A in homozygous and heterozygous FH either by binding to apoB or by other mechanisms. The ELISA method to measure both forms of plasma PCSK9 would be useful for investigating physiological or pathological roles of PCSK9.

Aggravated renal tubular damage and interstitial fibrosis in mice lacking guanylyl cyclase-A (GC-A), a receptor for atrial and B-type natriuretic peptides.

BACKGROUND AND AIM: The infusion of chronic angiotensin II (Ang II) has been shown to promote renal interstitial fibrosis. To evaluate the pathophysiological significance of the natriuretic peptide-GC-A system, we infused Ang II (1.0 mg/kg/day) in GC-A-deficient mice (GC-A-KO). METHODS: We used 5 groups (Wild-Saline n = 12, Wild-Ang II n = 14, GC-A-KO-Saline n = 11, GC-A-KO-Ang II n = 13, and GC-A-KO-Ang II-Hydralazine n = 10). Saline or Ang II was infused subcutaneously using an osmotic minipump for 3 weeks. Hydralazine was administered orally (0.05 g/L in drinking water). RESULTS: Systolic blood pressure was significantly higher in the GC-A-KO-Saline group (130 ± 12 mmHg) than in the Wild-Saline group (105 ± 30 mmHg), and was similar to that in the Wild-Ang II (141 ± 17 mmHg) and GC-A-KO-Ang II-Hydralazine (140 ± 20 mmHg) groups. Systolic blood pressure was significantly higher in the GC-A-KO-Ang II group (159 ± 21 mmHg) than in the 4 other groups. Renal tubular atrophy and interstitial fibrosis were significantly more severe in the GC-A-KO-Ang II group (atrophy 13.4 %, fibrosis 12.0 %) than in the Wild-Saline (0, 2.0 %), Wild-Ang II (2.9, 4.4 %), and GC-A-KO-Saline (0, 2.6 %) groups. Hydralazine could not inhibit this aggravation (GC-A-KO-Ang II-Hydralazine 13.5, 11.3 %). The expression of monocyte chemotactic protein-1 in tubular cells, and F4/80 and alpha-smooth muscle actin in the interstitium was clearly detected in the Ang II-infused wild and GC-A-KO groups and was associated with renal tubular atrophy and interstitial fibrosis. The expression of E-cadherin in tubular cells was absent in the Ang II-infused wild and GC-A-KO groups and was associated with renal tubular atrophy. CONCLUSIONS: The natriuretic peptide-GC-A system may play an inhibitory role in Ang II-induced renal tubular atrophy, interstitial fibrosis, and phenotypic transformation in renal tubular cells and fibroblasts.

Renal insufficiency without albuminuria is associated with peripheral artery atherosclerosis and lipid metabolism disorders in patients with type 2 diabetes.

AIMS: A high prevalence of a low glomerular filtration rate (GFR) has recently been reported in patients with diabetes without albuminuria. We aimed to clarify the clinical characteristics of such patients, including the associations between these characteristics and atherosclerosis. METHODS: We investigated the correlations between the estimated GFR (eGFR) and lipid profiles, the ankle-brachial index (ABI) and the intima-media thickness (IMT) in 450 patients with type 2 diabetes without macroalbuminuria. RESULTS: The prevalence of renal insufficiency (RI) (GFR ＜60 mL/min/1.73 m(2)) in the patients without albuminuria was 19.1%. The ABI values of the patients with RI were significantly lower than those of the patients without RI, regardless of the presence of microalbuminuria, while there were no significant differences in IMT between the patients with and without RI. In a multivariate analysis, a low ABI was found to be significantly associated with a low eGFR, independent of age, sex, smoking, history of hypertension and/or dyslipidemia and duration of diabetes (ß=0.134, p=0.013), whereas no significant associations were observed between the ABI and the urinary albumin excretion rate (UAER). The ApoB/LDL-C ratios and levels of ApoC3 were significantly higher in the patients with RI than those observed in the patients without RI, regardless of the presence of albuminuria. CONCLUSIONS: RI without albuminuria is closely associated with atherosclerosis of the peripheral arteries in diabetic patients. Furthermore, alterations in lipid metabolism may underlie this association.

Excessive sympathoactivation and deteriorated heart function after myocardial infarction in male ghrelin knockout mice.

We have previously demonstrated the protective role of endogenous ghrelin against malignant arrhythmias in the very acute phase of myocardial infarction (MI). However, the role of endogenous ghrelin in the chronic phase is unknown. Therefore, the aim of the current study was to focus on the effects of endogenous ghrelin on cardiac function and sympathetic activation after acute MI. In 46 ghrelin-knockout (KO) and 41 wild-type (WT) male mice, MI was produced by left coronary artery ligation. The mortality due to heart failure within 2 weeks was 0% in WT and 10.9% in KO (P < 0.05). At the end of this period, lung weight/tibial length, atrial natriuretic peptide and brain natriuretic peptide transcripts, end-systolic and end-diastolic volumes were all significantly greater in KO mice, whereas systolic function, represented by ejection fraction (16.4 ± 4.7% vs 25.3 ± 5.1%), end-systolic elastance, and preload-recruitable stroke work, was significantly inferior to that in WT mice (P < 0.05). Telemetry recording and heart rate variability analysis showed that KO mice had stronger sympathetic activation after MI than did WT mice. Metoprolol treatment and ghrelin treatment in KO mice prevented excessive sympathetic activation, decreased plasma epinephrine and norepinephrine levels, and improved heart function and survival rate after MI. Our data demonstrate that endogenous ghrelin plays a crucial role in protecting heart function and reducing mortality after myocardial infarction, and that these effects seem to be partly the result of sympathetic inhibition.

Ghrelin prevents incidence of malignant arrhythmia after acute myocardial infarction through vagal afferent nerves.

Ghrelin is a GH-releasing peptide mainly excreted from the stomach. Ghrelin administration has been shown to inhibit cardiac sympathetic nerve activity (CSNA), reduce malignant arrhythmia, and improve prognosis after acute myocardial infarction (MI). We therefore investigated the effects and potential mechanisms of the action of endogenous ghrelin on survival rate and CSNA after MI by using ghrelin-knockout (KO) mice. MI was induced by left coronary artery ligation in 46 KO mice and 41 wild-type mice. On the first day, malignant arrhythmia-induced mortality was observed within 30 min of the ligation and had an incidence of 2.4% in wild-type and 17.4% in KO mice (P < 0.05). We next evaluated CSNA by spectral analysis of heart rate variability. CSNA, represented by the low frequency/high frequency ratio, was higher in KO mice at baseline (2.18 ± 0.43 vs. 0.98 ± 0.09; P < 0.05), and especially after MI (25.5 ± 11.8 vs. 1.4 ± 0.3; P < 0.05), than in wild-type mice. Ghrelin (150 µg/kg, s.c.) 15 min before ligation suppressed the activation of CSNA and reduced mortality in KO mice. Further, this effect of ghrelin was inhibited by methylatropine bromide (1 mg/kg, i.p.) or by perineural treatment of both cervical vagal trunks with capsaicin (a specific afferent neurotoxin). Our data demonstrated that both exogenous and endogenous ghrelin suppressed CSNA, prevented the incidence of malignant arrhythmia, and improved the prognosis after acute MI. These effects are likely to be via the vagal afferent nerves.

Defining patients at extremely high risk for coronary artery disease in heterozygous familial hypercholesterolemia.

AIM: Heterozygous patients with familial hypercholesterolemia (FH) are known to be associated with a high risk of coronary artery disease (CAD), which is a major determinant of their clinical outcome. The prognosis of heterozygous FH patients substantially varies, being dependent on the level of their CAD risk, and their therapeutic regimen should be individualized. We assessed critical levels of LDL-cholesterol (LDL-C) and Achilles tendon thickness (ATT) to identify heterozygous FH patients at "very high" risk for CAD. METHODS: One hundred and nine heterozygous FH patients who had no history of CAD and had had their plasma lipid profile and ATT assessed before treatment were followed up until their first CAD event or 31 December 2010. Multivariable logistic regression models were used to analyze the correlation of LDL-C and/or ATT levels with the risk of developing CAD. RESULTS: During the follow-up period, 21 of the 109 patients had a CAD event, diagnosed by coronary angiogram. Individuals in the highest tertile of LDL-C had a CAD risk 8.29-fold higher than those in the lowest tertile. Individuals in the highest tertile of the ATT group had a 7.82-fold higher CAD risk than those in the lowest tertile. Those who had either LDL-C ≥ 260 mg/dL or ATT ≥ 14.5 had a 23.94-fold higher CAD risk than those with LDL-C < 260 mg/dL and ATT <14.5 mm. CONCLUSIONS: In heterozygous FH patients, LDL-C 260 mg/dL or higher and/or ATT 14.5 mm or thicker are useful markers for extracting patients at "very high" risk for CAD.

Role for p16(INK4a) in progression of gastrointestinal stromal tumors of the stomach: alteration of p16(INK4a) network members.

Gastrointestinal stromal tumors feature a wide spectrum of biologic behavior, ranging from benign to extremely malignant. To determine the role of p16(INK4a) alteration in progression of gastrointestinal stromal tumors of the stomach, we have investigated protein expression and gene methylation in correlation with clinicopathologic factors and survival. In addition to immunohistochemical analysis of p16(INK4a) in a series of 95 cases, real-time quantitative methylation specific polymerase chain reaction for p16(INK4a) and immunostaining for cyclin D1, cyclin E, pRb, DP-1, E2F-1, and Ki-67 were also evaluated in randomly selected samples. The p16(INK4a) labeling indices ranged from 0% to 74% (median, 21%), demonstrating a significant inverse correlation with size (P = .046). On univariate (P = .003) and multivariate (P = .067) analyses, loss of p16(INK4a) expression increased the likelihood of a poor tumor-related survival. In addition, size (P = .036) and the mitotic index (P = .005) had independent prognostic influence. The p16(INK4a) methylation index, which ranged from 0% to 100% (median, 17%), was significantly higher in larger tumors (P < .001) and in high-risk category lesions (P = .001) and inversely correlated with protein expression. Hierarchical cluster analysis based on expression of p16(INK4a) network members identified 2 clusters in 27 randomly selected tumor samples, containing 11 and 16 tumors each. Former cluster samples demonstrated higher risk category (P = .022), higher p16(INK4a) methylation (P < .001), and more reduced pRb expression (P < .018). In addition, p16(INK4a) network members clustered into 2 groups: (1) showing down-regulated p16(INK4a) protein and up-regulating of both cyclin D1 and DP-1 and (2) down-regulated pRb and up-regulated E2F-1. We conclude that p16(INK4a) alteration has an important role in progression of gastrointestinal stromal tumors of the stomach. Furthermore, the study provides a possible link between regulation of p16(INK4a) network members and gastrointestinal stromal tumors.

Impaired recovery of blood flow after hind-limb ischemia in mice lacking guanylyl cyclase-A, a receptor for atrial and brain natriuretic peptides.

OBJECTIVE: Atrial and brain natriuretic peptides (ANP and BNP, respectively) function via guanylyl cyclase (GC)-A, resulting in diuresis, natriuresis, and blood vessel dilation. Here, we investigated the role of endogenous ANP/BNP-GC-A signaling on reparative vascular remodeling using a hind-limb ischemia model. METHODS AND RESULTS: In GC-A-deficient mice (GC-A-KO), hind-limb ischemia resulted in autoamputation or severe ulcers in 60% of mice (6/10) during the 28-day observation period. In wild-type (WT) mice, partial amputation or mild ulcers were detected in only 20% of mice (2/10). Laser Doppler perfusion imaging revealed that the recovery of blood flow in the ischemic limb was significantly inhibited in GC-A-KO mice compared with WT mice. Immunostainings with anti-PECAM-1 antibody demonstrated that, in GC-A-KO, the capillary density of the ischemic tissue was significantly diminished compared to WT. Furthermore, bone marrow transplantation showed the predominant role of GC-A on local ischemic tissue rather than on vascular progenitor cells mobilized from bone marrow during vascular remodeling. In cultured human endothelial cells, ANP treatment significantly stimulated mRNA expressions of vascular endothelial growth factor and endothelial nitric oxide synthase via Erk1/2-dependent mechanism. CONCLUSIONS: These results suggest that endogenous ANP and BNP play important roles in reparative vascular remodeling in ischemic tissue.

Advanced gastric cancer showing long-term complete remission in response to S-1 monotherapy: two case reports.

We herein report two cases showing long-term complete remission (CR) in response to S-1 monotherapy. Case 1 was a 65-year-old male diagnosed with an advanced poorly differentiated adenocarcinoma of the stomach with paraaortic lymph node metastases, which disappeared after S-1 monotherapy. Subsequently a total gastrectomy was performed, and histological CR was evident. His progress is presently uneventful without recurrence 50 months after surgery. Case 2 was a 59-year-old female who underwent a total gastrectomy with a jejunal pouch. The resected tumor was a medullary type poorly differentiated adenocarcinoma infiltrating the serosa and involving the regional lymph nodes. One year after surgery, endoscopy revealed a recurrent tumor in the jejunal pouch. After the administration of S-1, this recurrent tumor completely disappeared, and she has since maintained CR for 39 months. These cases suggest that a subgroup of patients with advanced gastric cancer may attain CR with S-1 monotherapy.

Abnormal expression of p16(INK4a), cyclin D1, cyclin-dependent kinase 4 and retinoblastoma protein in gastric carcinomas.

BACKGROUND AND OBJECTIVES: The p16(INK4a) (p16), cyclin D1, cyclin-dependent kinase (CDK) 4 and retinoblastoma (Rb) genes are components of the Rb pathway that controls the G1-S checkpoint of the cell cycle. The aim of this study was to assess the relationship between their abnormalities and clinicopathological features in gastric carcinomas. METHODS: Immunohistochemical analysis of the encoded proteins was performed on a series of 158 cases. RESULTS: Loss of p16/Rb protein (pRb) expression and overexpression of cyclin D1/CDK4 were observed in 49%/40% and 37%/37% of gastric carcinomas, respectively. At least 1 of these abnormalities was found in 86% of the cases and a positive correlation was noted between p16 and pRb (P = 0.009). Cyclin D1 (P = 0.042) and CDK4 (P = 0.008) overexpession was inversely associated with lymph node metastasis and depth of invasion, respectively. Loss of pRb expression was more frequently in diffuse type lesions than in the intestinal type (P = 0.022). The patients with p16+/pRb-/cyclin D1-/CDK4- or p16-/pRb+/cyclin D1-/CDK4- tumors demonstrated particularly poor survival. With multivariate survival analysis, only depth of invasion and TNM stage could be proven as independent predictors. CONCLUSIONS: The Rb pathway is disrupted in the vast majority of gastric carcinomas. This study also identified specific immunohistochemical marker profiles for prognosis.

Exogenous ghrelin attenuates the progression of chronic hypoxia-induced pulmonary hypertension in conscious rats.

Chronic exposure to hypoxia, a common adverse consequence of most pulmonary disorders, can lead to a sustained increase in pulmonary arterial pressure (PAP), right ventricular hypertrophy, and is, therefore, closely associated with heart failure and increased mortality. Ghrelin, originally identified as an endogenous GH secretagogue, has recently been shown to possess potent vasodilator properties, likely involving modulation of the vascular endothelium and its associated vasoactive peptides. In this study we hypothesized that ghrelin would impede the pathogenesis of pulmonary arterial hypertension during chronic hypoxia (CH). PAP was continuously measured using radiotelemetry, in conscious male Sprague Dawley rats, in normoxia and during 2-wk CH (10% O(2)). During this hypoxic period, rats received a daily sc injection of either saline or ghrelin (150 microg/kg). Subsequently, heart and lung samples were collected for morphological, histological, and molecular analyses. CH significantly elevated PAP in saline-treated rats, increased wall thickness of peripheral pulmonary arteries, and, consequently, induced right ventricular hypertrophy. In these rats, CH also led to the overexpression of endothelial nitric oxide synthase mRNA and protein, as well as endothelin-1 mRNA within the lung. Exogenous ghrelin administration attenuated the CH-induced overexpression of endothelial nitric oxide synthase mRNA and protein, as well as endothelin-1 mRNA. Consequently, ghrelin significantly attenuated the development of pulmonary arterial hypertension, pulmonary vascular remodeling, and right ventricular hypertrophy. These results demonstrate the therapeutic benefits of ghrelin for impeding the pathogenesis of pulmonary hypertension and right ventricular hypertrophy, particularly in subjects prone to CH (e.g. pulmonary disorders).

P21WAF1/CIP1 expression in colorectal carcinomas is related to Kras mutations and prognosis.

BACKGROUND/AIM: P21WAF1/CIP1 is a cyclin-dependent kinase inhibitor activated by p53 to produce cell cycle arrest. A consensus has not been reached concerning the prognostic value of p21WAF1/CIP1 expression in colorectal cancers. PATIENTS/METHODS: P21WAF1/CIP1 expression was determined immunohistochemically in a series of 211 cases of colorectal carcinomas, together with its relation to p53, bcl-2, cell turnover (as assessed by Ki67 expression and apoptotic counts) and the Kras gene status. The expression of p21WAF1/CIP1 was also compared with reference to clinicopathological parameters and patient survival. RESULTS: The median value for nuclear p21WAF1/CIP1 expression was 31% (interquartile range, 13-47%) and the fraction of cases considered to be high expressers (>20%) was 66%. Expression of p21WAF1/CIP1 was not associated with immunoreactivity for p53 or bcl-2, or cell turnover. P21WAF1/CIP1 high-expressing tumors were more often well differentiated (P<0.001), node-negative (P=0.037), Dukes' B (P=0.027) and Kras gene-mutated cases (P=0.04). On univariate analysis, low p21WAF1/CIP1 expressers (