Insulin-like growth factor I improves intestinal barrier function in cirrhotic rats

BACKGROUND AND AIMS: In liver cirrhosis, disruption of the intestinal barrier facilitates bacterial translocation and spontaneous bacterial peritonitis. Insulin-like growth factor I (IGF-I) is an anabolic hormone synthesised by hepatocytes that displays hepatoprotective activities and trophic effects on the intestine. The aim of this study was to investigate the effect of IGF-I on intestinal barrier function in cirrhotic rats. METHODS: In rats with carbon tetrachloride induced cirrhosis, we investigated the effect of IGF-I therapy on: (a) portal pressure; (b) intestinal histology and permeability to endotoxin and bacteria; (c) intestinal expression of cyclooxygenase 2 (COX-2) and tumour necrosis factor alpha (TNF-alpha), two factors that influence in a positive and negative manner, respectively, the integrity of the intestinal barrier; (d) intestinal permeability to 3H-mannitol in rats with bile duct ligation (BDL); and (e) transepithelial electrical resistance (TER) of polarised monolayers of rat small intestine epithelial cells. RESULTS: IGF-I therapy reduced liver collagen expression and portal pressure in cirrhotic rats, induced improvement in intestinal histology, and caused a reduction in bacterial translocation and endotoxaemia. These changes were associated with diminished TNF-alpha expression and elevated COX-2 levels in the intestine. IGF-I reduced intestinal permeability in BDL rats and enhanced barrier function of the monolayers of epithelial intestinal cells where lipopolysaccharide (LPS) caused a decrease in TER that was reversed by IGF-I. This effect of IGF-I was associated with upregulation of COX-2 in LPS treated enterocytes. CONCLUSIONS: IGF-I enhances intestinal barrier function and reduces endotoxaemia and bacterial translocation in cirrhotic rats. IGF-I therapy might be useful in the prevention of spontaneous bacterial peritonitis in liver cirrhosis.     

Insulin-like growth factor I and growth hormone in canine starvation

The roles of plasma insulin-like growth factor I (IGF I) and growth hormone (GH) were studied in 7 beagle dogs before and during starvation and during refeeding. IGF I levels significantly decreased from 75.2 +/- 5.9 ng/ml at 7 days prior to the start of starvation to 9 +/- 1.7 ng/ml at 19 days after the commencement of starvation (mean +/- SEM; P less than 0.0001). During refeeding IGF I significantly rose from 9 +/- 1.7 ng/ml to 55.5 +/- 7.5 ng/ml within 9 days (mean +/- SEM; P less than 0.002). During starvation plasma GH levels significantly increased (P less than 0.05) and these elevated levels returned to normal during refeeding. The dogs' GH secretory capacity significantly increased during starvation (P = 0.012) and became normal again during refeeding. The following conclusions can be drawn from this study: 1) starvation in the dog leads to a significant and drastic reduction of the circulating levels of IGF I, and 2) starvation in the dog, as in man, leads to increased circulating GH levels and to an increased GH-secretory capacity possibly brought about by a lack of a negative feedback normally exerted by IGF I.     

Insulin-like growth factor I stimulates erythropoiesis in hypophysectomized rats.

Stimulation of erythropoiesis during growth is necessary to ensure proportionality between erythrocyte mass and body mass. However, the way by which erythrocyte formation is adapted to body growth is still unknown. Growth arrest in hypophysectomized rats is accompanied by decreased erythropoiesis. We have, therefore, examined whether insulin-like growth factor I (IGF-I), the mediator of growth hormone effects on body growth, is able to restore erythropoiesis in these animals. Subcutaneous infusions of 120 micrograms of recombinant human IGF-I per day in hypophysectomized rats led to increases in body weight, 59Fe incorporation into erythrocytes, and the number of reticulocytes that were similar to increases caused by infusions of 28 milliunits of human growth hormone per day. Body weight gain and 59Fe incorporation were linearly correlated. Like growth hormone, IGF-I also caused a significant rise in serum erythropoietin concentrations. However, the stimulatory effect on erythropoiesis occurred before serum erythropoietin levels had risen. These results demonstrate that IGF-I mediates the stimulatory effect of growth hormone on erythropoiesis in vivo and thus further support the somatomedin concept. They also show that IGF-I can stimulate erythropoiesis in an endocrine manner, and they suggest two possible routes of action: a direct one and an indirect one by means of enhanced erythropoietin production.     

Insulin-like growth factor I stimulates myofibril development and decreases smooth muscle alpha-actin of adult cardiomyocytes.

Adult rat cardiomyocytes in long-term cultureexpress type 1 insulin-like growth factor (IGF) receptors. In contrast toinsulin receptors, type 1 IGF receptors are up-regulated during culturing. IGF-Iadded to the cells at plating increased granular density and pseudopodia numberper cell after 7 days. After 16 days, IGF-I-treated cells showed, as comparedwith controls, a dramatic increase of the number of newly built sarcomeres andwere packed with myofibrils. At the same time, IGF-I suppressed the accumulationof smooth muscle alpha-actin (sm-alpha-actin) in a dose-dependent manner. Underthe conditions of this in vitro system, growth hormone had no effect on cellmorphology or sm-alpha-actin. sm-alpha-Actin, a nonsarcomeric isoform of actinexpressed in early fetal cardiac development, reappears both during long-termculture of adult rat cardiomyocytes and during heart hypertrophy. This studyshows that type 1 IGF receptors are up-regulated in adult rat cardiomyocytes inlong-term culture and that IGF-I enhances myofibril development andconcomitantly down-regulates sm-alpha-actin. This protein formsstress-fiber-like structures and may temporarily serve as a scaffold for theformation of new sarcomeres until myofibrils have developed throughout the celland the scaffold is no longer needed. Our findings thus allow us to proposeanother hypothesis for the mechanism leading to overload hearthypertrophy.     

Connective tissue growth factor regulates cardiac function and tissue remodeling in a mouse model of dilated cardiomyopathy

Cardiac structural changes associated with dilated cardiomyopathy (DCM) include cardiomyocyte hypertrophy and myocardial fibrosis. Connective tissue growth factor (CTGF) has been associated with tissue remodeling and is highly expressed in failing hearts. Our aim was to test if inhibition of CTGF would alter the course of cardiac remodeling and preserve cardiac function in the protein kinase Cε (PKCε) mouse model of DCM. Transgenic mice expressing constitutively active PKCε in cardiomyocytes develop cardiac dysfunction that was evident by 3 months of age, and that progressed to cardiac fibrosis, heart failure, and increased mortality. Beginning at 3 months of age, PKCε mice were treated with a neutralizing monoclonal antibody to CTGF (FG-3149) for an additional 3 months. CTGF inhibition significantly improved left ventricular (LV) systolic and diastolic functions in PKCε mice, and slowed the progression of LV dilatation. Using gene arrays and quantitative PCR, the expression of many genes associated with tissue remodeling was elevated in PKCε mice, but significantly decreased by CTGF inhibition. However total collagen deposition was not attenuated. The observation of significantly improved LV function by CTGF inhibition in PKCε mice suggests that CTGF inhibition may benefit patients with DCM. Additional studies to explore this potential are warranted.     

The role of apoptosis in dilated cardiomyopathy

Dilated cardiomyopathy is a cardiac disease of unknown origin which is characterized by the gradual development of cardiac failure. Apoptosis, i.e. suicidal programmed cell death, may play a role in the development of heart failure. Only few studies have been carried out until now that describe the rate of apoptosis in human hearts with dilated cardiomyopathy. The numbers reported vary widely. This is also true for studies in different other cardiac diseases such as myocardial infarction or hibernating myocardium. The methods used to identify apoptosis include electron microscopy, labeling of the DNA fragments (TUNEL), staining with the Hoechst dye, annexin V labeling and documentation of DNA fragmentation using gel electrophoresis (laddering). None of these methods are totally reliable in tissue sections in which apoptosis is not a frequent event when they are not combined with another technique, e.g. TUNEL with electron microscopy or laddering. This has, however, only rarely been done. These technical difficulties may be the reason for the wide variation in the rate of apoptosis reported. From our own data we conclude that apoptosis plays a significant role in acute ischemia and in hibernating myocardium but its significance in the progression to heart failure in dilated cardiomyopathy has still to be established.     

Fish oil supplementation improves left ventricular function in children with idiopathic dilated cardiomyopathy

Fish oil has a cardioprotective effect in adults with ischemic heart disease. The authors examined the effects of fish oil in children with idiopathic dilated cardiomyopathy (DCM). Eighteen DCM patients (group I) and 12 healthy children (group III) were given fish oil (10 mL/d). Their cardiac findings were compared with those of 11 patients with DCM who did not receive fish oil (group II). After 6.62+/-1.70 months, left ventricular ejection fraction had increased by 8.44%+/-3.80% (P<.05), in group I; 2.48%+/-3.85% (not statistically significant) in group II; and 0.84%+/-2.34% (not statistically significant) in group III. Left ventricular internal diastolic diameter (mm) was reduced by 4.36+/-4.86 (P=.001) in group I and 1.92+/-5.37 (P=.263) in group II, but increased by 0.22+/-2.54 (not statistically significant) in group III. The results suggest that fish oil leads to accelerated improvement of left ventricular function. The authors believe that if these results are confirmed in larger studies, fish oil should be added to the standard anticongestive therapy of children with DCM.     

Insulin-like growth factor I is essential for postnatal growth in response to growth hormone.

Insulin-like growth factor I (IGF-I) is essential for cell growth and intrauterine development while both IGF-I and GH are required for postnatal growth. To explore the possibility of direct GH action on body growth, independent of IGF-I production, we have studied the effects of GH in an IGF-I-deficient mouse line created by the Cre/loxP system. The IGF-I null mice are born with 35% growth retardation and show delayed onset of peripubertal growth, grow significantly slower, and do not attain puberty. Their adult body weight was approximately one third and body length about two thirds that of their wild-type litter mates. Injection of recombinant human GH (rhGH, 3 mg/kg, twice daily, sc) between postnatal day 14 (P14) to P56 failed to stimulate their growth as measured as both body weight and length. In contrast, wild-type mice receiving the same doses of rhGH exhibited accelerated growth starting at P21 that continued until P56, when their body weight was increased by 30% and length by 12% compared with control mice treated with diluent. Despite the lack of response in growth, IGF-I null mice have normal levels of GH receptor expression in the liver and increased liver Jun B expression and liver size in response to rhGH treatment. Our results support an essential role for IGF-I in GH-induced postnatal body growth in mice.     

Insulin-like growth factor I in suckling rat gastric contents

The small intestinal mucosa of the neonatal rat expresses primarily lactase activity until just prior to weaning when lactase falls to low levels and a full complement of adult digestive enzymes appears. Insulin-like growth factor 1 (IGF-I) is a normal component of maternal milk of humans and experimental animals. Experiments were performed to examine the concentrations of IGF-I in dam milk and the gastric content of suckling pups. Lactase activity in 1-day-old neonates was 0.66 µmol glucose formed/mg protein/hr (unit) and fell progressively until day 25, whereas sucrase activity at day 1 postpartum was 0.07 units and rose progressively to 0.21 units at day 25. The IGF-I content of dam milk was measured at 1, 5, 10, 15, 18, and 20 days postpartum by radioreceptor assay (RRA). Milk contained 1.02 pmol IGF-I/ml milk at one day postpartum, peaked at day 18 with 5.08 pmol IGF-I/ml, and fell to 2.31 pmol/ml at day 20. By day 25, dams were dry. The IGF-I content of the neonate gastric lumen was also measured by RRA. At day 1 the gastric lumen contained 2.63 pmol IGF-I/ml of luminal contents, fell to 1.06 pmol IGF-I/ml at day 5, and then rose again to peak at 3.37 pmol/ml at day 15 just prior to weaning. Two days after weaning, the level of luminal IGF-I had fallen to 1.15 pmol/ml. These data demonstrate the concentration of IGF-I in maternal milk is reflected in the concentration of the peptide in gastric contents of suckling pups and that the concentration in the gastric lumen may be high enough to affect epithelial cell proliferation and differentiation.This work was supported by grants to ER Seidel from the NIH (DK 34110) and the North Carolina Institute of Nutrition.     

Insulin-like growth factor I receptors on erythrocytes in NIDDM

We examined 125I-insulin-like growth factor I (125I-IGF-I) binding to erythrocytes from 24 normal and 21 non-insulin-dependent diabetic (NIDDM) subjects. 125I-IGF-I binding to human erythrocytes was specifically inhibited by unlabeled IGF-I, and Scatchard analysis indicated a curvilinear plot. There was a significant difference in IGF-I binding between normal and diabetic subjects (7.78 +/- 0.42 vs. 5.80 +/- 0.33%/2.4 x 10(9) cells/ml, P less than 0.001). Among diabetic patients, IGF-I binding to erythrocytes from those with retinopathy and those without retinopathy was comparable. These results suggested that decreased IGF-I binding might be a factor responsible for some pathological features such as delayed wound healing in diabetic patients.     

Insulin-like growth factor binding protein-3 mediates cytokine-induced mesangial cell apoptosis.

Mesangial cells are critical for glomerular filtration. Mesangial cell dysfunction, the hallmark of diabetic nephropathy, results from disordered mesangial growth induced by cytokines, abnormal hemodynamic influence, and metabolic factors associated with chronic hyperglycemia. Insulin-like growth factors (IGFs) and their high affinity binding proteins (IGFBPs) exert major actions on mesangial cell survival, but their underlying mechanisms remain unclear. In light of emerging IGF-independent roles for IGFBP-3, we investigated IGFBP-3 actions during mesangial cell apoptosis induced by cytokine or high glucose concentration. Quantified by DNA fragmentation ELISA and Annexin V flow cytometry, apoptosis occurred in rat mesangial cells (RMC) exposed to 2 microg/mL IGFBP-3 for 24 h under high ambient or standard glucose. Anti-sense IGFBP-3 oligo at 10 microg/mL significantly inhibited apoptosis induced by 100 ng/mL TNF-alpha, serum-free conditions, or high (25 mM) glucose. Increased IGFBP-3 release associated with high ambient glucose or TNF-alpha was inhibited by pre-treatment with anti-sense oligo. Under serum-free conditions, recombinant human IGFBP-3 blocked Akt phosphorylation at threonine 308 (pThr308), whereas anti-sense oligo treatment was associated with enhanced pThr308 activity. In summary, these data support a novel mechanism for TNF-alpha-induced mesangial cell apoptosis mediated by IGFBP-3 and present regulation of pThr308 activity as a novel mechanism underlying IGFBP-3 action.     

Insulin-like growth factor II and transforming growth factor beta 1 regulate insulin-like growth factor I secretion in mouse bone cells.

Bone cells in culture produce and respond to growth factors, suggesting that local as well as systemic factors regulate bone volume. Previous studies have shown that IGF-I is the major mitogen produced by mouse bone cells and that its production is regulated by systemic agents such as PTH and estrogen. Because IGF-II and transforming growth factor beta 1 have been shown, respectively, to increase and decrease MC3T3-E1 cell proliferation, we tested the hypothesis that these two growth factors modulate the production of IGF-I in this cell line. In order to eliminate artifacts owing to IGF binding proteins, conditioned media samples were pretreated with IGF-II before measurement of IGF-I by RIA. After 24 h treatment at a density of 2.5 x 10(4) cells/cm2, IGF-II (10 micrograms/l) induced a 2.2-fold increase compared with untreated control (9.5 +/- 1.5 vs 4.2 +/- 0.44 pg/micrograms protein, p less than 0.001), whereas transforming growth factor beta 1 (1 microgram/l) caused a 66% decrease in IGF-I production (1.5 +/- 0.3 vs 4.2 +/- 0.44 pg/micrograms protein, p less than 0.001). Both IGF-II and transforming growth factor beta 1 regulated IGF-I production in a dose-, time- and cell density-dependent manner. The lowest effective doses for IGF-II and transforming growth factor beta 1 were 1 and 0.01 microgram/l, respectively. These results support a role for IGF-II and transforming growth factor beta 1 as potent modulators of IGF-I secretion in mouse bone cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Right ventricular function in dilated cardiomyopathy

The right ventricular (RV) function was comprehensively examined in 18 patients with dilated cardiomyopathy (DCMP) and compared with RV function in 10 controls. In most cases of DCMP, the RV function is affected simultaneously with a disturbance of the left ventricular (LV) function. However, the degree of its affection is usually less pronounced and, with less severe LV dysfunction, the right ventricle can work even normally. There can be substantial individual differences in the degree of affection of the right ventricle in DCMP. However, severe LV dysfunction is invariably associated with a marked involvement of the right ventricle. A disturbance of the RV function results (besides the influence of the decreased LV function) from both a decrease in its own contractility and a decrease in RV compliance. There exist significant relations between the RV systolic and diastolic function. The degree of RV disturbance in DCMP both of the systolic and diastolic function) can be approximately estimated from the level of diastolic pressure.     

PDGF-BB protects cardiomyocytes from apoptosis and improves contractile function of engineered heart tissue

Platelet-derived-growth-factor-BB (PDGF-BB) can protect various cell types from apoptotic cell death, and induce hypertrophic growth and proliferation, but little is known about its direct or indirect effects on cardiomyocytes. Cardiac muscle engineering is compromised by a particularly high rate of cardiomyocyte death. Here we hypothesized that PDGF-BB stimulation can (1) protect cardiomyocytes from apoptosis, (2) enhance myocyte content in and (3) consequently optimize contractile performance of engineered heart tissue (EHT). We investigated the effects of PDGF-receptor activation in neonatal rat heart monolayer- and EHT-cultures by isometric contraction experiments, cytomorphometry, ³H-thymidine and ³H-phenylalanine incorporation assays, quantitative PCR (calsequestrin 2, α-cardiac and skeletal actin, atrial natriuretic factor, α- and β-myosin heavy chain), immunoblotting (activated caspase 3, Akt-phosphorylation), and ELISA (cell death detection). PDGF-BB did not induce hypertrophy or proliferation in cardiomyocytes, but enhanced contractile performance of EHT. This effect was concentration-dependent (E_max 10 ng/ml) and maximal only after transient PDGF-BB stimulation (culture days 0–7; total culture duration: 12 days). Improvement of contractile function was associated with higher cardiomyocyte content, as a consequence of PDGF-BB mediated protection from apoptosis (lower caspase-3 activity particularly in cardiomyocytes in PDGF-BB treated vs. untreated EHTs). We confirmed the anti-apoptotic effect of PDGF-BB in monolayer cultures and observed that PI3-kinase inhibition with LY294002 attenuated PDGF-BB-mediated cardiomyocyte protection. We conclude that PDGF-BB does not induce hypertrophy or proliferation, but confers an anti-apoptotic effect on cardiomyocytes. Our findings suggest a further exploitation of PDGF-BB in cardiomyocyte protection in vivo and in vitro.     

Insulin-like growth factor I, growth hormone and insulin in white adipose tissue

Maturation of adipose tissue results from both the expansion of mature adipocytes and the formation of new adipocytes from adipocyte precursor cells. A variety of hormones related to adipogenesis have been identified recently. Both growth hormone (GH) and insulin-like growth factor I (IGF-I) are of major significance in adipocyte differentiation. IGF-I has been suggested to be a major regulator of cell proliferation, differentiation and metabolism, thus regulating, among other biological processes, adipose tissue growth and differentiation of pre-adipocytes into adipocytes. GH exerts its effects by increasing the pool of adipocyte precursor cells capable of differentiating into mature adipocytes. In addition, GH seems to have the potential to reduce the volume of mature adipocytes, thus inhibiting the expansion of adipose tissue and reducing body fat. This chapter gives an overview of studies that have investigated the roles of insulin, GH and IGF-I in adipogenesis.