Tim Lahm

Human mesenchymal stem cells stimulated by TNF-α, LPS, or hypoxia produce growth factors by an NFκB- but not JNK-dependent mechanism

Understanding the mechanisms by which adult stem cells produce growth factors may represent an important way to optimize their beneficial paracrine and autocrine effects. Components of the wound milieu may stimulate growth factor production to promote stem cell-mediated repair. We hypothesized that tumor necrosis factor-alpha (TNF-alpha), endotoxin (LPS), or hypoxia may activate human mesenchymal stem cells (MSCs) to increase release of vascular endothelial growth factor (VEGF), fibroblast growth factor 2 (FGF2), insulin-like growth factor 1 (IGF-1), or hepatocyte growth factor (HGF) and that nuclear factor-kappa B (NF kappa B), c-Jun NH2-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK) mediates growth factor production from human MSCs. To study this, human MSCs were harvested, passaged, divided into four groups (100,000 cells, triplicates) and treated as follows: 1) with vehicle; 2) with stimulant alone [24 h LPS (200 ng/ml), 24 h TNF-alpha (50 ng/ml), or 24 h hypoxia (1% O2)]; 3) with inhibitor alone [NF kappa B (PDTC, 1 mM), JNK (TI-JIP, 10 microM), or ERK (ERK Inhibitor II, 25 microM)]; and 4) with stimulant and the various inhibitors. After 24 h incubation, MSC activation was determined by measuring supernatants for VEGF, FGF2, IGF-1, or HGF (ELISA). TNF-alpha, LPS, and hypoxia significantly increased human MSC VEGF, FGF2, HGF, and IGF-1 production versus controls. Stem cells exposed to injury demonstrated increased activation of NF kappa B, ERK, and JNK. VEGF, FGF2, and HGF expression was significantly reduced by NF kappa B inhibition (50% decrease) but not ERK or JNK inhibition. Moreover, ERK, JNK, and NF kappa B inhibitor alone did not activate MSC VEGF expression over controls. Various stressors activate human MSCs to increase VEGF, FGF2, HGF, and IGF-1 expression, which depends on an NFkB mechanism.

VEGF is critical for stem cell-mediated cardioprotection and a crucial paracrine factor for defining the age threshold in adult and neonatal stem cell function

Bone marrow mesenchymal stem cells (MSCs) may be a novel treatment modality for organ ischemia, possibly through the release of beneficial paracrine factors. However, an age threshold likely exists as to when MSCs gain their beneficial protective properties. We hypothesized that 1) VEGF would be a crucial stem cell paracrine mediator in providing postischemic myocardial protection and 2) small-interfering (si)RNA ablation of VEGF in adult MSCs (aMSCs) would equalize the differences observed between aMSC- and neonatal stem cell (nMSC)-mediated cardioprotection. Female adult Sprague-Dawley rat hearts were subjected to ischemia-reperfusion injury via Langendorff-isolated heart preparation (15 min equilibration, 25 min ischemia, and 60 min reperfusion). MSCs were harvested from adult and 2.5-wk-old neonatal mice and cultured under normal conditions. VEGF was knocked down in both cell lines by VEGF siRNA. Immediately before ischemia, one million aMSCs or nMSCs with or without VEGF knockdown were infused into the coronary circulation. The cardiac functional parameters were recorded. VEGF in cell supernatants was measured via ELISA. aMSCs produced significantly more VEGF than nMSCs and were noted to increase postischemic myocardial recovery compared with nMSCs. The knockdown of VEGF significantly decreased VEGF production in both cell lines, and the pretreatment of these cells impaired stem cell-mediated myocardial function. The knockdown of VEGF in adult stem cells equalized the myocardial functional differences observed between adult and neonatal stem cells. Therefore, VEGF is a critical paracrine mediator in facilitating postischemic myocardial recovery and likely plays a role in mediating the observed age threshold during stem cell therapy.

Medical and Surgical Treatment of Acute Right Ventricular Failure

Acute right ventricular (RV) failure is a frequent and serious clinical challenge in the intensive care unit. It is usually seen as a consequence of left ventricular failure, pulmonary embolism, pulmonary hypertension, sepsis, acute lung injury or after cardiothoracic surgery. The presence of acute RV failure not only carries substantial morbidity and mortality, but also complicates the use of commonly used treatment strategies in critically ill patients. In contrast to the left ventricle, the RV remains relatively understudied, and investigations of the treatment of isolated RV failure are rare and usually limited to nonrandomized observations. We searched PubMed for papers in the English language by using the search words right ventricle, right ventricular failure, pulmonary hypertension, sepsis, shock, acute lung injury, cardiothoracic surgery, mechanical ventilation, vasopressors, inotropes, and pulmonary vasodilators. These were used in various combinations. We read the abstracts of the relevant titles to confirm their relevance, and the full papers were then extracted. References from extracted papers were checked for any additional relevant papers. This review summarizes the general measures, ventilation strategies, vasoactive substances, and surgical as well as mechanical approaches that are currently used or actively investigated in the treatment of the acutely failing RV.

Sex steroids and stem cell function.

Gender dimorphisms exist in the pathogenesis of a variety of cardiovascular, cardiopulmonary, neurodegenerative, and endocrine disorders. Estrogens exert immense influence on myocardial remodeling following ischemic insult, partially through paracrine growth hormone production by bone marrow mesenchymal stem cells (MSCs) and endothelial progenitor cells. Estrogens also facilitate the mobilization of endothelial progenitor cells to the ischemic myocardium and enhance neovascularization at the ischemic border zone. Moreover, estrogens limit pathological myocardial remodeling through the inhibitory effects on the proliferation of the cardiac fibroblasts. Androgens also may stimulate endothelial progenitor cell migration from the bone marrow, yet the larger role of androgens in disease pathogenesis is not well characterized. The beneficial effects of sex steroids include alteration of lipid metabolism in preadipocytes, modulation of bone metabolism and skeletal maturation, and prevention of osteoporosis through their effects on osteogenic precursors. In an example of sex steroid-specific effects, neural stem cells exhibit enhanced proliferation in response to estrogens, whereas androgens mediate inhibitory effects on their proliferation. Although stem cells can offer significant therapeutic benefits in various cardiovascular, neurodegenerative, endocrine disorders, and disorders of bone metabolism, a greater understanding of sex hormones on diverse stem cell populations is required to improve their ultimate clinical efficacy. In this review, we focus on the effects of estrogen and testosterone on various stem and progenitor cell types, and their relevant intracellular mechanisms.

17β-Estradiol Attenuates Hypoxic Pulmonary Hypertension via Estrogen Receptor–mediated Effects

RATIONALE 17β-Estradiol (E2) attenuates hypoxic pulmonary vasoconstriction and hypoxic pulmonary hypertension (HPH) through an unknown mechanism that may involve estrogen receptors (ER) or E2 conversion to catecholestradiols and methoxyestradiols with previously unrecognized effects on cardiopulmonary vascular remodeling. OBJECTIVES To determine the mechanism by which E2 exerts protective effects in HPH. METHODS Male rats were exposed to hypobaric hypoxia while treated with E2 (75 μg/kg/d) or vehicle. Subgroups were cotreated with pharmacologic ER-antagonist or with inhibitors of E2-metabolite conversion. Complementary studies were performed in rats cotreated with selective ERα- or ERβ-antagonist. Hemodynamic and pulmonary artery (PA) and right ventricular (RV) remodeling parameters, including cell proliferation, cell cycle, and autophagy, were measured in vivo and in cultured primary rat PA endothelial cells. MEASUREMENTS AND MAIN RESULTS E2 significantly attenuated HPH endpoints. Hypoxia increased ERβ but not ERα lung vascular expression. Co-treatment with nonselective ER inhibitor or ERα-specific antagonist rendered hypoxic animals resistant to the beneficial effects of E2 on cardiopulmonary hemodynamics, whereas ERα- and ERβ-specific antagonists opposed the remodeling effects of E2. In contrast, inhibition of E2-metabolite conversion did not abolish E2 protection. E2-treated hypoxic animals exhibited reduced ERK1/2 activation and increased expression of cell-cycle inhibitor p27(Kip1) in lungs and RV, with up-regulation of lung autophagy. E2-induced signaling was recapitulated in hypoxic but not normoxic endothelial cells, and was associated with decreased vascular endothelial growth factor secretion and cell proliferation. CONCLUSIONS E2 attenuates hemodynamic and remodeling parameters in HPH in an ER-dependent manner, through direct antiproliferative mechanisms on vascular cells, which may provide novel nonhormonal therapeutic targets for HPH.

Embryonic stem cells attenuate myocardial dysfunction and inflammation after surgical global ischemia via paracrine actions

Stem cell treatment may positively influence recovery and inflammation after shock by multiple mechanisms, including the paracrine release of protective growth factors. Embryonic stem cells (ESCs) are understudied and may have greater protective power than adult bone marrow stem cells (BMSCs). We hypothesized that ESC paracrine protective mechanisms in the heart (decreased injury by enhanced growth factor-mediated reduction of proinflammatory cytokines) would be superior to the paracrine protective mechanisms of the adult stem cell population in a model of surgically induced global ischemia. Adult Sprague-Dawley rat hearts were isolated and perfused via Langendorff model. Hearts were subjected to 25 min of warm global ischemia and 40 min of reperfusion and were randomly assigned into one of four groups: 1) vehicle treated; 2) BMSC or ESC preischemic treatment; 3) BMSC or ESC postischemic treatment; and 4) BMSC- or ESC-conditioned media treatment. Myocardial function was recorded, and hearts were analyzed for expression of tissue cytokines and growth factors (ELISA). Additionally, ESCs and BMSCs in culture were assessed for growth factor production (ELISA). ESC-treated hearts demonstrated significantly greater postischemic recovery of function (left ventricular developed pressure, end-diastolic pressure, and maximal positive and negative values of the first derivative of pressure) than BMSC-treated hearts or controls at end reperfusion. ESC-conditioned media (without cells) also conferred cardioprotection at end reperfusion. ESC-infused hearts demonstrated increased VEGF and IL-10 production compared with BMSC hearts. ESC hearts also exhibited decreased proinflammatory cytokine expression compared with MSC hearts. Moreover, ESCs in cell culture demonstrated greater pluripotency than MSCs. ESC paracrine protective mechanisms in surgical ischemia are superior to those of adult stem cells.

Neonatal Iron Deficiency Causes Abnormal Phosphate Metabolism by Elevating FGF23 in Normal and ADHR Mice

Fibroblast growth factor 23 (FGF23) gain of function mutations can lead to autosomal dominant hypophosphatemic rickets (ADHR) disease onset at birth, or delayed onset following puberty or pregnancy. We previously demonstrated that the combination of iron deficiency and a knock‐in R176Q FGF23 mutation in mature mice induced FGF23 expression and hypophosphatemia that paralleled the late‐onset ADHR phenotype. Because anemia in pregnancy and in premature infants is common, the goal of this study was to test whether iron deficiency alters phosphate handling in neonatal life. Wild‐type (WT) and ADHR female breeder mice were provided control or iron‐deficient diets during pregnancy and nursing. Iron‐deficient breeders were also made iron replete. Iron‐deficient WT and ADHR pups were hypophosphatemic, with ADHR pups having significantly lower serum phosphate (p < 0.01) and widened growth plates. Both genotypes increased bone FGF23 mRNA (>50 fold; p < 0.01). WT and ADHR pups receiving low iron had elevated intact serum FGF23; ADHR mice were affected to a greater degree (p < 0.01). Iron‐deficient mice also showed increased Cyp24a1 and reduced Cyp27b1, and low serum 1,25‐dihydroxyvitamin D (1,25D). Iron repletion normalized most abnormalities. Because iron deficiency can induce tissue hypoxia, oxygen deprivation was tested as a regulator of FGF23, and was shown to stimulate FGF23 mRNA in vitro and serum C‐terminal FGF23 in normal rats in vivo. These studies demonstrate that FGF23 is modulated by iron status in young WT and ADHR mice and that hypoxia independently controls FGF23 expression in situations of normal iron. Therefore, disturbed iron and oxygen metabolism in neonatal life may have important effects on skeletal function and structure through FGF23 activity on phosphate regulation.

The effects of estrogen on pulmonary artery vasoreactivity and hypoxic pulmonary vasoconstriction: Potential new clinical implications for an old hormone

Background and Objectives:Recent research recognizes gender as a major factor determining the outcomes in trauma, ischemia/reperfusion, shock, and sepsis. In particular, estrogen has been demonstrated to exert protective effects in these settings. The effects of estrogens on the pulmonary vasculature are potent and complex yet not fully understood. A better mechanistic understanding may allow for future therapeutic interventions in pulmonary hypertensive crises after cardiac surgery and during acute lung injury as well as in patients with pulmonary arterial hypertension. Data Sources and Study Selection:We searched PubMed for articles in the English language by using the search words pulmonary hypertension, hypoxic pulmonary vasoconstriction, estrogen, estradiol, inflammation, acute injury, ischemia reperfusion, sepsis, trauma, and burns. These were used in various combinations. We read the abstracts of the relevant titles to confirm their relevance, and the full articles were then extracted. References from extracted articles were checked for any additional relevant articles. Data Extraction and Synthesis:Estrogen plays a critical role in the improved outcomes in the settings of trauma, shock, sepsis, myocardial ischemia/reperfusion, and acute lung injury. Several new mechanisms of action have been identified. In the pulmonary vasculature, estrogen causes vasodilation and attenuates the vasoconstrictor response to various stimuli, including hypoxia. This is mediated by increased levels of prostacyclin and nitric oxide as well as decreased levels of endothelin-1. In addition, effects on intracellular signaling pathways and several kinases as well as anti-inflammatory mechanisms may contribute as well. Recent studies suggest the importance of acute, nongenomic effects. Conclusion:Estrogen exerts a variety of nongenomic actions, which may allow for future therapeutic interventions in pulmonary vascular disease.

Stem cell mechanisms and paracrine effects: potential in cardiac surgery.

Heart disease remains the leading cause of death in the industrialized world. Stem cell therapy is a promising treatment modality for injured cardiac tissue. A novel mechanism for this cardioprotection may include paracrine actions. Cardiac surgery represents the unique situation where preischemia and postischemia treatment modalities exist that may use stem cell paracrine protection. This review (1) recalls the history of stem cells in cardiac disease and the unraveling of its mechanistic basis for protection, (2) outlines the pathways for stem cell-mediated paracrine protection, (3) highlights the signaling factors expressed, (4) explores the potential of using stem cells clinically in cardiac surgery, and (5) summarizes all human stem cell studies in cardiac disease to date.ABBREVIATIONS-MSC- mesenchymal stem cell; VEGF- vascular endothelial growth factor; HGF- hepatocyte growth factor; FGF- fibroblast growth factor; TGF- transforming growth factor; LV- left ventricular; LVEF- left ventricular ejection fraction; MI- myocardial infarction; PCI- percutaneous coronary intervention

β-Blockers in sepsis: Reexamining the evidence

Sepsis remains the leading cause for noncardiac intensive care unit deaths in the United States. Despite recent advances in the treatment of this devastating condition, mortality and morbidity remain unacceptably high. Sepsis is characterized by a multitude of pathophysiological changes that include inflammation, metabolic derangements, hemodynamic alterations, and multiorgan dysfunction. Unfortunately, several studies of treatment modalities aimed at correcting one or more of the underlying derangements have led to disappointing results. New treatment modalities are needed. &bgr;-Receptor blockers have long been used for a variety of conditions such as coronary artery disease, congestive heart failure, and arterial hypertension. Recent data suggest that &bgr;-blocker effects on metabolism, glucose homeostasis, cytokine expression, and myocardial function may be beneficial in the setting of sepsis. Although treating a potentially hypotensive condition with a drug with antihypertensive properties may initially seem counterintuitive, the metabolic and immunomodulatory properties of &bgr;-blockers may be of benefit. It is the purpose of this review to discuss the effects of &bgr;-blockers on the following: (1) metabolism, (2) glucose regulation, (3) the inflammatory response, (4) cardiac function, and (5) mortality in sepsis.