Characterization of Prolylcarboxypeptidase Expression Pattern in Rat Cardiomyocytes in Nutrition Overload Conditions

Prolylcarboxypeptidase (PRCP) is a serine protease that cleaves the last amino acid at the carboxy-terminus of peptides with a penultimate proline such as angiotensin II (Ang II), angiotensin III (Ang III), prekallikrein (PK) and α-melanocyte-stimulating-hormone (1-13) (α-MSH1-13). By inactivating Ang II and Ang III, PRCP promotes vasodilation and insulin sensitivity. As well by activating PK, PRCP increases nitric oxide (NO) generation and vasorelaxation through bradykinin (BK) liberation from high molecular weight kininogen (HK). PK is activated to kallikrein by PRCP, implicating a role for PRCP in the coagulation pathway. The resultant kallikrein will activate FXII which in a reciprocal manner leads to the activation of FXI. Lastly, by metabolizing α-MSH1-13 to an inactive metabolite (α- MSH1-12), PRCP inhibits the anorexigenic response to the endogenous α-MSH1-13, leading to an increase in appetite. Thus, by means of vasodilation, insulin sensitization (via direct stabilization of IRS-1 and indirect reduction of Ang II), and suppressing reactive oxygen species (ROS) generation (via direct increase in NO generation and indirect reduction of Ang II) PRCP functions to protect cardiovascular system. Moreover, PRCP mutation is related to acute coronary syndrome in men, and individuals with obesity, diabetes and arteriosclerosis have elevated plasma PRCP concentration and activity. Since cardiovascular disease is the leading cause of mortality in diabetes and obesity, PRCP overexpression in nutrition excess conditions represents an important target for studying metabolic syndrome- related cardiac dysfunction. The overall goal of this study was to elucidate the hormetic effects of major nutrients on PRCP-dependent pathways. Therefore, the following methods were exploited to address our goals: cell culture of rat H9c2 cardiomyocytes, enzymatic assays for detecting plasma and cardiomyocyte PRCP activity, PRCP RNA and protein studies in high glucose and high fatty acid (saturated and unsaturated) conditions, enzymatic assays to detect PRCP-stimulated activation of PK and plasma protein studies of PK and its downstream target in the coagulation cascade, FXI. Using in vitro biochemical assays, plasma PRCP and kallikrein activity were significantly increased in uncontrolled diabetic patients; however metformin and insulin treated diabetic patients had reduced plasma PRCP and kallikrein activity. As well, uncontrolled diabetic patients had markedly elevated plasma prekallikrein (PK), PRCP and FXI protein which was reduced in insulin and metformin treated groups. In a rat cardiomyocyte model of nutrition overload, saturated fatty acid palmitate, unlike glucose, suppressed PRCP levels by 60% in a dose-dependent and time-dependent manner without affecting cell viability, while other tested saturated and unsaturated fatty acids did not alter the basal cardiomyocyte PRCP expression. Thyroxine and insulin, but not metformin, restored palmitate-induced cardiac PRCP depletion. Lastly, although fatty acid uptake inhibition by the CD36 antagonist increased palmitate- induced PRCP depletion by 2-fold, NFκB inhibition did not restore palmitate-stimulated PRCP suppression. Our results indicate that fatty acid-accelerated cardiomyocyte PRCP depletion along with diabetes-stimulated increase in plasma PK and FXI concentration, may contribute to nutrition overload-stimulated cardiovascular dysfunction. The details of fatty acid-induced PRCP down regulation in the heart could open new therapeutic avenues to protect against metabolic syndrome-related cardiovascular complications

Dietary Walnuts Protect Against Obesity-Driven Intestinal Stem Cell Decline and Tumorigenesis

Obesity can negatively impact intestinal homeostasis, and increase colon cancer risk and related mortality. Thus, given the alarmingly high rates of obesity in the US and globally, it is critical to identify practical strategies that can break the obesity-cancer link. Walnuts have been increasingly recognized to mitigate cancer risk, and contain many bioactive constituents with antioxidant and anti-inflammatory properties that could potentially counteract pathways thought to be initiators of obesity-related cancer. Therefore, the purpose of this study was to determine if walnuts could preserve intestinal homeostasis, and attenuate tumorigenesis and growth in the context of obesity and a high calorie diet. To this end, we studied effects of walnuts on these parameters under different dietary conditions in wildtype mice, two independent Apc models (Apc1638N/+ and ApcΔ14), and in MC38 colon cancer cells in vivo, respectively. Walnuts did not alter the metabolic phenotype or intestinal morphology in normal mice fed either a low-fat diet (LFD), LFD with 6% walnuts (LFD+W), high-fat diet (HFD), or HFD with 7.6% walnuts (HFD+W). However, walnuts did lead to a significant reduction in circulating CCL5 and preserved intestinal stem cell (ISC) function under HFD-fed conditions. Furthermore, walnuts reduced tumor multiplicity in Apc1638N/+ male HFD+W animals, as compared to HFD controls (3.7 ± 0.5 vs. 2.5 ± 0.3; P = 0.015), tended to reduce the number of adenocarcinomas (0.67 ± 0.16 vs. 0.29 ± 0.12; P = 0.07), and preferentially limited tumor growth in ApcΔ14 male mice (P = 0.019) fed a high-calorie western-style diet. In summary, these data demonstrate that walnuts confer significant protection against intestinal tumorigenesis and growth and preserve ISC function in the context of a high-calorie diet and obesity. Thus, these data add to the accumulating evidence connecting walnuts as a potentially effective dietary strategy to break the obesity-colon cancer link

Image_2_Dietary Walnuts Protect Against Obesity-Driven Intestinal Stem Cell Decline and Tumorigenesis.pdf

<p>Obesity can negatively impact intestinal homeostasis, and increase colon cancer risk and related mortality. Thus, given the alarmingly high rates of obesity in the US and globally, it is critical to identify practical strategies that can break the obesity-cancer link. Walnuts have been increasingly recognized to mitigate cancer risk, and contain many bioactive constituents with antioxidant and anti-inflammatory properties that could potentially counteract pathways thought to be initiators of obesity-related cancer. Therefore, the purpose of this study was to determine if walnuts could preserve intestinal homeostasis, and attenuate tumorigenesis and growth in the context of obesity and a high calorie diet. To this end, we studied effects of walnuts on these parameters under different dietary conditions in wildtype mice, two independent Apc models (Apc^1638N/+ and Apc^Δ14), and in MC38 colon cancer cells in vivo, respectively. Walnuts did not alter the metabolic phenotype or intestinal morphology in normal mice fed either a low-fat diet (LFD), LFD with 6% walnuts (LFD+W), high-fat diet (HFD), or HFD with 7.6% walnuts (HFD+W). However, walnuts did lead to a significant reduction in circulating CCL5 and preserved intestinal stem cell (ISC) function under HFD-fed conditions. Furthermore, walnuts reduced tumor multiplicity in Apc^1638N/+ male HFD+W animals, as compared to HFD controls (3.7 ± 0.5 vs. 2.5 ± 0.3; P = 0.015), tended to reduce the number of adenocarcinomas (0.67 ± 0.16 vs. 0.29 ± 0.12; P = 0.07), and preferentially limited tumor growth in Apc^Δ14 male mice (P = 0.019) fed a high-calorie western-style diet. In summary, these data demonstrate that walnuts confer significant protection against intestinal tumorigenesis and growth and preserve ISC function in the context of a high-calorie diet and obesity. Thus, these data add to the accumulating evidence connecting walnuts as a potentially effective dietary strategy to break the obesity-colon cancer link.</p

Sarcosine Is Uniquely Modulated by Aging and Dietary Restriction in Rodents and Humans

Summary: A hallmark of aging is a decline in metabolic homeostasis, which is attenuated by dietary restriction (DR). However, the interaction of aging and DR with the metabolome is not well understood. We report that DR is a stronger modulator of the rat metabolome than age in plasma and tissues. A comparative metabolomic screen in rodents and humans identified circulating sarcosine as being similarly reduced with aging and increased by DR, while sarcosine is also elevated in long-lived Ames dwarf mice. Pathway analysis in aged sarcosine-replete rats identify this biogenic amine as an integral node in the metabolome network. Finally, we show that sarcosine can activate autophagy in cultured cells and enhances autophagic flux in vivo, suggesting a potential role in autophagy induction by DR. Thus, these data identify circulating sarcosine as a biomarker of aging and DR in mammalians and may contribute to age-related alterations in the metabolome and in proteostasis. : In a comparative metabolic screen of rodents and humans, Walters et al. show that circulating sarcosine is similarly reduced with aging and increased by dietary restriction. They demonstrate that sarcosine activates macroautophagy in cultured cells and in vivo, suggesting a role in improved proteostasis via dietary restriction. Keywords: sarcosine, aging, metabolomics, dietary restriction, autophagy, GNMT, glycerophospholipids, amino acids, glycine, methionin