Elisabeth Svanberg

The effect of recombinant human IGF-I on protein metabolism in post-operative patients without nutrition compared to effects in experimental animals

Abstract. This study has evaluated the effects of recombinant human insulin‐like growth factor I (rhIGF‐I) to moderately stressed post‐operative patients provided with dextrose as the only exo‐geneous substrate. Thirty patients who underwent elective colorectal surgery were randomized to receive either rhIGF‐I (80μg kg‐1 bw) subcutaneously twice daily or placebo injections in a double‐blind parallel group design. Nitrogen balance, urinary 3‐methyl‐histidine excretion plasma growth hormone (GH), serum cortisol, IGF‐I binding proteins (IGFBP‐1,3), glomerular filtration rate, plasma amino acid concentrations and whole‐body energy expenditures were measured as effector variables during days 1–5 post‐operatively. Animal and isolated tissue experiments were performed as additional control experiments to confirm cellular effectiveness of the recombinant material. rhIGF‐I increased significantly the glomerular filtration rate and prevented the adaptive decrease in whole‐body energy expenditure in response to partial starvation in the postoperative period. Serum and plasma concentrations of IGFBP‐1,3 cortisol, blood glucose and amino acids were not significantly influenced by rhIGF‐I administration, while plasma GH levels decreased significantly as expected. rhIGF‐I had no effect on either nitrogen balance or protein breakdown (3‐methylhistidine excretion) in post‐operative patients on dextrose supplementation only, although plasma concentrations of IGF‐I increased from 130 140 ngmL‐1 to a range of 300–450 ngmL‐1. In contrast, IGF‐I stimulated the synthesis of both globular and myofibrillar proteins (+50%, P<0.01), when given as a single dose (100μgkg‐1) 2 h before measurements of protein synthesis in skeletal muscles of overnight fasted adult mice. This stimulatory effect by IGF‐I (1μgmL‐1) was also confirmed by measurements of skeletal muscle protein synthesis in vitro (+40%, P<0.05). Orally re‐fed mice had a normal transcription of IGF‐I mRNA in skeletal muscle cells, while overnight fasted mice showed a trend to down‐regulated transcription. Our results demonstrate that rhIGF‐I has several significant physiological effects, without major side‐effects, when supplied to partially starved patients in the post‐operative phase. The lack of a whole‐body nitrogen sparing effect by rhIGF‐I alone to postoperative patients is not clear, but was most likely explained by subnormal plasma concentrations of amino acids.

p53 Mutations in primary tumors and subsequent liver metastases are related to survival in patients with colorectal carcinoma who undergo liver resection

The appearance of p53 mutations in colorectal carcinoma was determined, independent of differentiation and tumor stage of the primary tumors, in relation to the survival of patients who were scheduled to undergo liver resection.

Cyclooxygenase inhibition in early onset of tumor growth and related angiogenesis evaluated in EP1 and EP3 knockout tumor-bearing mice

It is well established that prostanoids are essential for local inflammation including cell proliferation and apoptosis. Accordingly, prostaglandin E2 (PGE2) is a critical factor in wound healing, tumor invasiveness and progression. Therefore, the aim of the present work was to evaluate effects by PGE2 on tumor vascular density at early onset of tumor growth where hypoxia is limited. Wild-type mice (C57Bl, C3H/HeN) bearing either MCG-101 tumors or a malignant melanoma (K1735-M2) with either high or insignificant PGE2 production and subsequently different in sensitivity to cyclooxygenase (COX) inhibition were used. Tumor angiogenesis was estimated by intravital microscopy and immune histochemical analysis in wild type and EP1 or EP3 subtype receptor knockout mice (C57Bl). Both MCG-101 and K1735-M2 tumor cells stimulated early outgrowth of tumor vessels in proportion to intrinsic growth rate of tumor cells. Indomethacin had no effects on tumor growth or tumor related vascular area in K1735-M2 bearing mice. By contrast, indomethacin decreased tumor cell proliferation and increased apoptosis in MCG-101 tumors with subsequent adaptation in tumor vascular density. Effects of indomethacin on early growth of MCG-101 tumors were not related to tumor content of bFGF protein, while our earlier studies on long-term tumor growth have shown decreased mRNA levels of bFGF during indomethacin treatment. Early onset of tumor growth was significantly promoted in EP3- but not in EP1-knockouts, although long-term tumor growth is attenuated in EP1-knockouts as reported elsewhere. Our results demonstrate that tumor production of PGE2 promotes primarily net growth of tumor cells with subsequent adaptations in development of the tumor vasculature. Therefore, it is likely that angiogenesis is not a limiting step at the early onset of tumor growth.

Prostaglandin E and prostacyclin receptor expression in tumor and host tissues from MCG 101-bearing mice: A model with prostanoid-related cachexia

Preclinical and clinical studies in our laboratory have suggested that prostaglandin (PG) E2 is involved in anorexia and cachexia development, although the role of COX pathways on the pathogenesis of cancer cachexia remains to be clarified. Expressions of PGE (EP1, EP2, EP3α,β,γ and EP4) and PGI (IP) receptors in the central nervous system (brain cortex, hypothalamus and brain stem), in peripheral (liver, white adipose tissue and skeletal muscle) and tumor tissue from MCG‐101‐bearing mice with and without indomethacin treatment were investigated by RT‐PCR and immunohistochemistry. Expression of EP1 in the liver and EP4 receptor in white adipose tissue were upregulated and responded to indomethacin treatment, while downregulated expression of EP3 in skeletal muscle from tumor‐bearing mice was unresponsive to indomethacin treatment despite improved carcass weight. Expression of EP and IP receptors in brain and tumor tissue from tumor‐bearing mice were neither related nor responsive to systemic PGE2 levels including increased IL‐1bβ, IL‐6 and TNF‐aα host activities. The expression IP receptor in CNS, peripheral tissue and tumor tissue was unchanged by cachexia development. Our results suggest that transcription of EP receptors in liver, fat and skeletal muscle tissue may be a control level for host metabolic alterations during tumor progression, while overall EP and IP receptor expression in CNS did not indicate an important control level for appetite regulation in MCG 101‐bearing mice despite prostanoid related anorexia.

Postprandial resynthesis of myofibrillar proteins is translationally rather than transcriptionally regulated in human skeletal muscle.

Feeding stimulates protein synthesis in skeletal muscles, although the regulatory mechanisms are incompletely understood. The aim of this study was to determine whether this could be detected at the gene transcription level for postprandial stimulation of the synthesis of muscle proteins. Healthy male volunteers were investigated after an overnight fast. Open muscle biopsies were performed in the starved state and 3 h after meal intake, consisting of 0.15 gN/kg, 12 kcal/kg. Blood samples were drawn every 15 to 30 min for 5 h. Myosin mRNA and insulin growth factor-I (IGF-I) mRNA were measured by solution hybridization assay in homogenized muscle specimens. After food intake, plasma glucose concentrations increased from 5.0 +/- 0.1 to 7.3 +/- 0.3 (P < or = 0.001), and insulin concentration rose from 3.8 +/- 0.5 mU/L before to 75.3 +/- 11.4 15 min after the meal (P < or = 0.001). Plasma concentration of free fatty acids declined after food intake (P < or = 0.001). Plasma concentrations of amino acids increased from basal values (2864 +/- 128 microM) to 4419 +/- 262 microM (P < or = 0.05) 90 min after meal ingestion. Myosin mRNA concentration in the biopsied muscle tissue was higher during starvation and was reduced by 20% after food intake: 10.8 +/- 1.3 amol mRNA/microgram DNA in the starved state and 8.5 +/- 1.3 amol mRNA/microgram DNA after food intake (P < or = 0.05). Feeding did not alter IGF-I mRNA concentrations in muscle: 0.51 +/- 0.05 and 0.55 +/- 0.06 amol/microgram DNA in the starved and fed state, respectively (P < or = 0.48). Improved protein balance by stimulation of protein synthesis has been related to increased plasma amino acids. Interestingly, in the short term, this was not related to increases in gene transcription of either myofibrillar proteins (myosin) or muscle IGF-I. Thus, postprandial stimulation of protein synthesis appears not to be regulated by increased gene transcription but by increased translation using the increased concentrations of amino acids. In contrast, as far as the 2X myosin mRNA level is concerned, this is enhanced during starvation, which facilitates rapid recovery once the availability of substrate is resumed.

Lack of effects by tricyclic antidepressant and serotonin inhibitors on anorexia in MCG 101 tumor-bearing mice with eicosanoid-related cachexia ☆

OBJECTIVES Anorexia is a major clinical problem in large number of patients with advanced cancer disease. Serotonergic mechanisms are assumed to play a role in the process of feeding behavior during normal and pathologic circumstances, which may also involve cancer anorexia according to previous experimental and clinical studies. METHODS In the present study, we evaluated the effect of the tricyclic antidepressants desipramine (7.5 mg x kg(-1) x d(-1), intraperitoneal) and imipramine (2 to 5 mg. kg(-1) x d(-1), intraperitoneal) the serotonin synthesis inhibitor para-chlorophenylalanine (300 mg x kg(-1) x d(-1), intraperitoneal), the serotonin receptor 5-HT(2C) antagonist cyproheptadine (5 mg x kg(-1) x d(-1), intraperitoneal) and the selective serotonin reuptake inhibitor citalopram (20 mg x kg(-1) x d(-1), intraperitoneal) on anorexia in MCG-101 tumor-bearing mice, a model with significant anorexia and cachexia sensitive to cyclooxygenase inhibition. Also, MCG 101-bearing mice develop well-recognized alterations in brain tryptophan/serotonin metabolism as increased Trp, 5-HPT, and 5-HIAA during tumor progression. RESULTS Daily provision of desipramine, imipramine, para-chloropheylalanine, cyproheptadine, and citalopram at doses that cause behavioral and metabolic alterations in normal mice did not alter food intake or body weight in tumor-bearing and healthy control mice. Also, the treatments did not decrease elevated plasma concentrations of interleukin-6 and prostaglandin E(2) in the tumor-bearing mice. CONCLUSIONS Thus, our results do not support previous observations that serotonin metabolism itself is a major factor behind anorexia in tumor-bearing animals in general. Rather, other mechanisms, such as eicosanoid and nitric oxide-dependent pathways, seem to be more important for induction of anorexia along tumor progression in the present model.

Semi‐starvation alters myofibrillar mRNA concentrations to expedite rapid recovery of muscle protein stores following feeding

Protein synthesis in skeletal muscle is reduced following starvation and restored by feeding. The mediators and mechanisms are incompletely understood. The aim of this study was to evaluate whether prolongation of undernutrition induced changes in muscle gene expression at the level of mRNA and protein.

Anabolic effects of rhIGF‐I/IGFBP‐3 in vivo are influenced by thyroid status

Background There are close interrelationships between hormones that regulate bone formation and protein biosynthesis. For example, growth hormone and thyroid hormones can influence plasma levels of insulin‐like growth factor‐I (IGF‐I). A patients status regarding hormones other than IGF‐I may thus indirectly modify the efficacy of IGF‐I treatment. The aim of the current study was to determine if a statistical method could be used to identify key endocrine variables controlling an individuals response to IGF‐I treatment.

A One-Piece Plexiglass Access Chamber for Subcutaneous Implantation in the Dorsal Skin Fold of the Mouse

This technical report describes the production and installation of a newly developed, one-piece, light-weight (0.6 g) access plexiglass chamber for the dorsal skin fold of the mouse.

Mouse Extensor Digitorum Longus Muscle Preparation as a Tool in Nutrition Research: A Quantitative Comparison to In Vivo and Cell Culture Experiments

Incubated restrained and unrestrained extensor digitorum longus (EDL) muscles from adult non-growing mice were evaluated as a tool in non-steady state nutrition experiments. Energy state was determined by nucleotide determinations in muscles. Protein synthesis was estimated by the amount of L-[U-14C]phenylalanine incorporated into proteins, and protein balance was measured by tyrosine release from muscle proteins. Confluent cultured L6 rat muscle cells served as a reference system in steady state without hypoxia being sensitive to growth factors and regulatory peptides at physiologic concentrations. Irrespective of medium composition, incubated EDL muscles remained in negative protein balance, being unrelated to the resting tension of the incubated muscles. Energy-rich phosphates were not restored to normal levels during incubation, but protein synthesis was not attenuated by the decline in energy state. Fractional protein synthesis (0.05-0.15%/h) remained constant for up to 6 h of EDL incubation, and was comparable to protein synthesis in cultured confluent non-proliferating myocytes (0.20-0.30%/h) and to mixed leg muscles measured in vivo (0.10-0.20%/h). Protein synthesis in incubated EDL muscles reflected alterations in muscle peptide formation in vivo following either oral provision of food or parenteral injection of insulin. EDL muscles were sensitive to in vitro exposure to both insulin (60-125 microU/mL) and insulin-like growth factor 1 (IGF-1) (1000 ng/mL). The sensitivity to insulin seemed to be modified by the nutritional state (starved/fed) of the animals before sacrifice. Protein synthesis in EDL muscles was less responsive to serum-containing growth factors (IGF-1, epidermal growth factor [EGF], platelet-derived growth factor [PDGF]) compared to confluent L6 muscle cells, which probably reflected different receptor expression. Our results demonstrate that protein metabolism in incubated unrestrained mouse EDL muscles reflects in vivo protein metabolism.