A randomized translational study on protein- and glucose metabolism in skeletal muscles evaluated by gene-ontology, following preoperative oral carbohydrate loading compared to overnight peripheral parenteral nutrition (PPN) before major cancer surgery.

BACKGROUND: Effects of preoperative drinks on muscle metabolism are unclear despite general recommendations. The aim of the present study was therefore to compare metabolic effects of a preoperative oral nutrition drink, recommended by protocols for enhanced recovery after surgery (ERAS), compared to overnight preoperative peripheral total parenteral nutrition (PPN) on skeletal muscle metabolism in patients aimed at major gastrointestinal cancer surgery. METHODS: Patients were randomized, based on diagnosis and clinical characteristics, to receive either a commercial carbohydrate-rich nutrition drink (Drink); or overnight (12 h) peripheral parenteral nutrition (PPN) as study regimens; compared to isotone Ringer-acetate as Control regimen. Arterial blood- and abdominal muscle tissue specimens were collected at start of surgery. Blood chemistry included substrate- and hormone concentrations. Muscle mRNA transcript analyses were performed by microarray and evaluated for changes in gene activities by Gene Ontology algorithms. RESULTS: Patient groups were comparable in all measured preoperative assessments. The Nutrition Drink had significant metabolic alterations on muscle glucose metabolism (p < 0.05), without any significant effects on amino acid- and protein metabolism. PPN showed similar significant effects on glucose metabolism as Drinks (p < 0.05), but indicated also major positive effects on amino acid- (p < 0.001) and protein anabolism (p < 0.05), particularly by inhibition of muscle protein degradation, related to both ubiquitination of proteins and autophagy/lysosome pathways (p < 0.05). CONCLUSION: Conventional overnight preoperative PPN seems effective to induce and support improved muscle protein metabolism in patients aimed at major cancer surgery while preoperative oral carbohydrate loading, according to ERAS-protocols, was ineffective to improve skeletal muscle catabolism and should therefore not be recommended before major cancer surgery. Trial registration Clinical trials.gov: NCT05080816, Registered June 10th 2021- Retrospectively registered. https://clinicaltrials.gov/study/NCT05080816.

Surgical Morbidity and Mortality From the Multicenter Randomized Controlled NeoRes II Trial: Standard Versus Prolonged Time to Surgery After Neoadjuvant Chemoradiotherapy for Esophageal Cancer.

OBJECTIVE: To investigate if prolonged TTS after completed nCRT improves postoperative outcomes for esophageal and esophagogastric junction cancer. SUMMARY OF BACKGROUND DATA: TTS has traditionally been 4-6 weeks after completed nCRT. However, the optimal timing is not known. METHODS: A multicenter clinical trial was performed with randomized allocation of TTS of 4-6 or 10-12 weeks. The primary endpoint of this sub-study was overall postoperative complications defined as Clavien-Dindo grade II-V. Secondary endpoints included complication severity according to Clavien-Dindo grade IIIb-V, postoperative 90-day mortality, and length of hospital stay. The study was registered in Clinicaltrials.gov (NCT02415101). RESULTS: In total 249 patients were randomized. There were no significant differences between standard TTS and prolonged TTS with regard to overall incidence of complications Clavien-Dindo grade II-V (63.2% vs 72.6%, P = 0.134) or regarding Clavien-Dindo grade IIIb-V complications (31.6% vs 34.9%, P = 0.603). There were no statistically significant differences between standard and prolonged TTS regarding anastomotic leak (P = 0.596), conduit necrosis (P = 0.524), chyle leak (P = 0.427), pneumonia (P = 0.548), and respiratory failure (P = 0.723). In the standard TTS arm 5 patients (4.3%) died within 90 days of surgery, compared to 4 patients (3.8%) in the prolonged TTS arm (P = 1.0). Median length of hospital stay was 15 days in the standard TTS arm and 17 days in the prolonged TTS arm (P = 0.234). CONCLUSION: The timing of surgery after completed nCRT for carcinoma of the esophagus or esophagogastric junction, is not of major importance with regard to short-term postoperative outcomes.

Pretreatment with a CRF antagonist amplifies feeding inhibition induced by fourth ventricular cocaine- and amphetamine-regulated transcript peptide.

BACKGROUND: Pre-treatment with the corticotropin-releasing factor antagonist α-helical CRF9-41 prevents inhibition of gastric emptying by cocaine-and amphetamine-regulated transcript peptide at a dorsal hindbrain level, but its inhibition of sucrose intake is not affected. This is suggestive of separable underlying mechanisms of action in the caudal brainstem for cocaine-and amphetamine-regulated transcript peptide with regard to food intake and gastrointestinal functions. Here we further examine cocaine-and amphetamine-regulated transcript peptide-corticotropin-releasing factor receptor interactions in caudal brainstem controls of solid food intake. Injections of combinations of vehicle, cocaine-and amphetamine-regulated transcript peptide (0.5 µg or 1 µg) or α-helical CRF9-41 were given into the fourth cerebral ventricle of rats. Nocturnal solid food intake was recorded over 22 h. RESULTS: Pre-treatment with α-helical CRF9-41 into the fourth ventricle significantly increased the responsivity to cocaine-and amphetamine-regulated transcript peptide on hypophagia. In a separate control experiment, α-helical CRF9-41 pre-treatment blocked CRF-induced food intake inhibition indicative of its antagonistic effectiveness. CONCLUSIONS: We conclude that an endogenous Corticotropin-releasing factor agonist may modulate suppression of food intake caused by cocaine-and amphetamine-regulated transcript peptide at a dorsal hindbrain level in the absence of stress. A potential caudal brainstem mechanism whereby cocaine-and amphetamine-regulated transcript peptide effects on food intake is attenuated via corticotropin-releasing factor receptor activity causing tonic inhibition, is suggested.

LPS immune challenge reduces arcuate nucleus TSHR and CART mRNA and elevates plasma CART peptides.

BACKGROUND: The aim was to examine the impact of lipopolysaccharide-induced systemic inflammation on expression of mRNA for cocaine- and amphetamine-regulated transcript (CART) and the thyrotropin receptor (TSHR) and its ligands in CNS areas of relevance for feeding controls and metabolism. Lipopolysaccharide effects on plasma levels of TSH and CART peptides were also examined. METHODS: Lipopolysaccharide (150-200 µg/mouse) was injected in C57BL/6J mice and tissue and plasma samples taken after 24 h. To establish if plasma increase in CART peptide levels were prostanoid dependent, indomethacin was given via the drinking water beginning 48 h prior to LPS. We evaluated mRNA expression for CART, TSHR, TSHß, and thyrostimulin in brain and pituitary extracts. Plasma levels of TSH, CARTp, and serum amyloid P component were analyzed by ELISA. RESULTS: Lipopolysaccharide suppressed TSHR mRNA expression in the arcuate nucleus and the pituitary. CART mRNA expression was reduced in the arcuate nucleus but elevated in the pituitary of mice treated with Lipopolysaccharide, whereas plasma TSH remained unchanged. Plasma CART peptide concentration increased after LPS treatment in a prostanoid-independent manner, and CART peptide levels correlated positively to degree of inflammation. CONCLUSIONS: Our findings suggest that central and peripheral CART is affected by acute inflammation. Considering the role of the arcuate nucleus in feeding controls, our data highlight TSHR and CART as putative neuroendocrine signaling components that respond to inflammation, perhaps to maintain weight and metabolic homeostasis during states of disease.

Validation of instruments for the assessment of dysphagia due to malignancy of the esophagus.

The aim of the study was to validate the Watson scale, the Ogilvie scale, and the Goldschmid scale for assessment of dysphagia due to malignancy of the esophagus. After translation of the scales to Swedish, 35 patients with dysphagia due to esophageal malignancy were asked to participate. On day 1, patients were asked to fill in the questionnaires. The patients also kept a food diary for 4 consecutive days, for assessment of actual swallowing ability. On day 10, the patients were asked to fill in the scales again, to control for individual variability. As an external control group, 29 healthy volunteers were asked to fill in the questionnaires once. External validation was done against actual swallowing ability, and against the European Organization for Research and Treatment of Cancer scales QLQ-C30 and QLQ-OG25, which are already validated quality of life scales for malignancy. Reliability in the categorical variables (Ogilvie and Goldschmid) showed weighted kappa values of 0.52 and 0.54, respectively. For the Watson scale and the Dysphagia module of QLQ-OG25, the intraclass correlation coefficients were 0.68 and 0.80, respectively. Correlations between all scales were good to excellent with values of correlation coefficients (rs) between 0.69 and 0.88, with the strongest correlations between the Ogilvie score and the dysphagia module in QLQ-OG25. These latter two scales had the strongest correlation to the food diary (rs = 0.72). Although the Ogilvie scale was superior, all the three scales showed good reliability and are thus judged to have good validity for assessment of dysphagia due to esophageal malignancy.

Fourth ventricular thyrotropin induces satiety and increases body temperature in rats.

Besides its well-known action to stimulate thyroid hormone release, thyrotropin mRNA is expressed within the brain, and thyrotropin and its receptor have been shown to be present in brain areas that control feeding and gastrointestinal function. Here, the hypothesis that thyrotropin acts on receptors in the hindbrain to alter food intake and/or gastric function was tested. Fourth ventricular injections of thyrotropin (0.06, 0.60, and 6.00 µg) were given to rats with chronic intracerebroventricular cannulas aimed at the fourth ventricle. Thyrotropin produced an acute reduction of sucrose intake (30 min). The highest dose of thyrotropin caused inhibition of overnight solid food intake (22 h). In contrast, subcutaneous administration of corresponding thyrotropin doses had no effect on nutrient intake. The highest effective dose of fourth ventricular thyrotropin (6 µg) did not produce a conditioned flavor avoidance in a standardized two-bottle test, nor did it affect water intake or gastric emptying of glucose. Thyrotropin injected in the fourth ventricle produced a small but significant increase in rectal temperature and lowered plasma levels of tri-iodothyronin but did not affect plasma levels of thyroxine. In addition, there was a tendency toward a reduction in blood glucose 2 h after fourth ventricular thyrotropin injection ( P = 0.056). In conclusion, fourth ventricular thyrotropin specifically inhibits food intake, increases core temperature, and lowers plasma levels of tri-iodothyronin but does not affect gastromotor function.

Fully covered stents are similar to semi-covered stents with regard to migration in palliative treatment of malignant strictures of the esophagus and gastric cardia: results of a randomized controlled trial.

INTRODUCTION: Stent migration is a significant clinical problem in palliation of malignant strictures in the esophagus and gastro-esophageal junction (GEJ). We have compared a newer design of a fully-covered stent to a widely used semi-covered stent using migration >20 mm as the primary outcome variable. Effects on dysphagia, quality of life (QoL) and re-intervention frequency were also investigated. METHODS: Patients with dysphagia due to non-curable esophagus/GEJ cancer were randomized to receive either a more recent design of a fully-covered stent (n = 48) or a conventional semi-covered stent (n = 47). Chest x-ray, dysphagia and QoL were studied at baseline, one week, four weeks and three months thereafter. RESULTS: There were no significant differences either in stent migration distance or in the migration frequency. Stent migration during the total study period occurred in 37.2 % in the semi-covered group compared to 20.0 % for the fully-covered group. Dysphagia was measured with Watson and Ogilvie scores and with the dysphagia module in the QoL scale (QLQ-OG25). On average, there was a tendency to better dysphagia relief for the fully-covered design as scored with the two latter dysphagia instruments (p= 0.081 and p= 0.067) at three months and towards more re-interventions in the semi-covered group (p= 0.083). CONCLUSION: In spite of its somewhat lower intrinsic radial force, the fully-covered stent was comparable to the conventional semi-covered stent with regard to stent migration. The data further suggest a potential benefit of the fully-covered stent in improving dysphagia in patients with longer life expectancy.

Validation of the paracetamol absorption test for measuring gastric tube emptying in esophagectomized patients versus gold standard scintigraphy.

OBJECTIVE: Scintigraphy is the gold standard for objective measurement of delayed gastric tube emptying after esophagectomy. The aim of this pilot study is to validate, by reference to scintigraphy, the paracetamol absorption test for measuring gastric tube emptying in esophagectomized patients. MATERIAL AND METHODS: The paracetamol absorption test and scintigraphy were performed simultaneously in 13 patients who had undergone an esophagectomy with gastric tube reconstruction. Emptying was calculated for both methods and compared. Post-esophagectomy symptoms and quality of life (QoL) were assessed by European Organization on Research and Treatment of Cancer questionnaires. RESULTS: Mean time to 50% emptying was 17 min measured with the paracetamol absorption test and 23 min with scintigraphy. For time to 25% emptying, Bland-Altman calculation gave a bias of 1.6 min and 95% limits of agreement (LoA) of -6.3 to 9.5 min. For time to 50% emptying, there was one outlier resulting in a bias of -6.33 min and 95% LoA of -36.4 to 23.8 min. For time to 75% emptying, bias was -11.6 min and 95% LoA of -38.5 to 15.4 min. Post-esophagectomy symptoms were similar to those reported previously, and QoL was comparable to the general Swedish population. CONCLUSIONS: There was reasonably close correlation between the paracetamol absorption test and scintigraphy for time to 25% and 50% emptying, except for one outlier. For time to 75% emptying the methods were in less accordance. The results indicate that the paracetamol absorption test may be a useful screening tool for identifying delayed gastric tube emptying in this patient group.

Treatment of achalasia with laparoscopic myotomy or pneumatic dilatation: long-term results of a prospective, randomized study.

BACKGROUND: This study compares the long-term results of pneumatic dilatations versus laparoscopic myotomy using treatment failure as the primary outcome. The frequency and degree of dysphagia, the effects on quality of life (QoL), and health economy were also examined. METHODS: Fifty-three patients with achalasia were randomized to laparoscopic myotomy with a posterior partial fundoplication [laparoscopic myotomy (LM) n = 25] or repetitive pneumatic dilatation [pneumatic dilatation (PD) n = 28]. The median observation period was 81.5 months (range 12-131). RESULTS: At the minimal follow-up of 5 years, ten patients (36%) in the dilatation group and two patients (8%) in the myotomy group, including two patients lost to follow-up (one in each arm), were classified as failures (p = 0.016). The cumulative incidence of treatment failures was analyzed by survival statistics. Taking the entire follow-up period into account, a significant difference was observed in favor of the LM strategy (p = 0.02). Although both treatments resulted in significant improvements in dysphagia scores, LM was significantly favored over PD after 1 and 3 years, but not after 5 years. Health-related QoL assessed by the personal general well being score was higher in the LM group after 3 years, but the difference was not fully statistically significant at 5 years. Direct medical costs during the entire follow-up period were in median $13,421 for LM as compared to $5,558 for PD (p = 0.001). CONCLUSIONS: This long-term follow-up of a randomized clinical study shows that LM is superior to repetitive PD treatment of newly diagnosed achalasia, albeit that this surgical strategy is burdened by high initial direct medical costs. www.ClinicalTrials.gov NCT 02086669.

Long-Term Changes to the Microbiome, Blood Lipid Profiles and IL-6 in Female and Male Swedish Patients in Response to Bariatric Roux-en-Y Gastric Bypass.

Lipid metabolism dysregulation is a critical factor contributing to obesity. To counteract obesity-associated disorders, bariatric surgery is implemented as a very effective method. However, surgery such as Roux-en-Y gastric bypass (RYGB) is irreversible, resulting in life-long changes to the digestive tract. The aim of the present study was to elucidate changes in the fecal microbiota before and after RYGB in relation to blood lipid profiles and proinflammatory IL-6. Here, we studied the long-term effects, up to six years after the RYGB procedure, on 15 patients' gut microbiomes and their post-surgery well-being, emphasizing the biological sex of the patients. The results showed improved health among the patients after surgery, which coincided with weight loss and improved lipid metabolism. Health changes were associated with decreased inflammation and significant alterations in the gut microbiome after surgery that differed between females and males. The Actinobacteriota phylum decreased in females and increased in males. Overall increases in the genera Prevotella, Paraprevotella, Gemella, Streptococcus, and Veillonella_A, and decreases in Bacteroides_H, Anaerostipes, Lachnoclostridium_B, Hydrogeniiclostridium, Lawsonibacter, Paludicola, and Rothia were observed. In conclusion, our findings indicate that there were long-term changes in the gut microbiota after RYGB, and shifts in the microbial taxa appeared to differ depending on sex, which should be investigated further in a larger cohort.

Reduced tumor growth in EP2 knockout mice is related to signaling pathways favoring an increased local anti­tumor immunity in the tumor stroma.

Inflammatory signaling through prostaglandin E2 receptor subtype 2 (EP2) is associated with malignant tumor growth in both experimental models and cancer patients. Thus, the absence of EP2 receptors in host tissues appears to reduce tumor growth and systemic inflammation by inducing major alterations in gene expression levels across tumor tissue compartments. However, it is not yet well­established how signaling pathways in tumor tissue relate to simultaneous signaling alterations in the surrounding tumor­stroma, at conditions of reduced disease progression due to decreased host inflammation. In the present study, wild­type tumor cells, producing high levels of prostaglandin E2 (MCG 101 cells, EP2+/+), were inoculated into EP2 knockout (EP2­/­) and EP2 wild­type (EP2+/+) mice. Solid tumors were dissected into tumor­ and tumor­stroma tissue compartments for RNA expression microarray screening, followed by metabolic pathway analyses. Immunohistochemistry was used to confirm adequate dissections of tissue compartments, and to assess cell proliferation (Ki­67), prostaglandin enzymes (cyclooxygenase 2) and immunity biomarkers (CD4 and CD8) at the protein level. Microarray analyses revealed statistically significant alterations in gene expression in the tumor­stroma compartment, while significantly less pathway alterations occurred in the tumor tissue compartment. The host knockout of EP2 receptors led to a significant downregulation of cell cycle regulatory factors in the tumor­stroma compartment, while interferon γ­related pathways, chemokine signaling pathways and anti­tumor chemokines [chemokine (C­X­C motif) ligand 9 and 10] were upregulated in the tumor compartment. Thus, such gene alterations were likely related to reduced tumor growth in EP2­deficient hosts. On the whole, pathway analyses of both tumor­ and tumor­stroma compartments suggested that absence of host EP2 receptor signaling reduces 'remodeling' of tumor microenvironments and increase local immunity, probably by decreased productions of stimulating growth factors, perhaps similar to well­recognized physiological observations in wound healing.

Esophageal perforation in South of Sweden: results of surgical treatment in 125 consecutive patients.

BACKGROUND: For many years there has been a debate as to which is the method of choice in treating patients with esophageal perforation. The literature consists mainly of small case series. Strategies for aiding patients struck with this disease is changing as new and less traumatic treatment options are developing. We studied a relatively large consecutive material of esophageal perforations in an effort to evaluate prognostic factors, diagnostic efforts and treatment strategy in these patients. METHODS: 125 consecutive patients treated at the University Hospital of Lund from 1970 to 2006 were studied retrospectively. Prognostic factors were evaluated using the Cox proportional hazards model. RESULTS: Pre-operative ASA score was the only factor that significantly influenced outcome. Neck incision for cervical perforation (n = 8) and treatment with a covered stent with or without open drainage for a thoracic perforation (n = 6) had the lowest mortality. Esophageal resection (n = 8) had the highest mortality. A CAT scan or an oesophageal X-ray with oral contrast were the most efficient diagnostic tools. The preferred treatment strategy changed over the course of the study period, from a more aggressive surgical approach towards using covered stents to seal the perforation. CONCLUSION: Pre-operative ASA score was the only factor that significantly influenced outcome in this study. Treatment strategies are changing as less traumatic options have become available. Sealing an esophageal perforation with a covered stent, in combination with open or closed drainage when necessary, is a promising treatment strategy.

Total gastrectomy causes a sustained, long-term elevation of somatostatin in plasma, independent of the mode of reconstruction in pigs.

AIM: The long-term effects of gastrectomy and various reconstructions of the gastrointestinal tract on fasting plasma levels of gastrointestinal hormones known to contribute to the control of gastrointestinal motor function were evaluated in pigs. MATERIALS AND METHODS: Domestic pigs were randomly selected to sham surgery or total gastrectomy (TG) followed by reconstruction with oesophago-jejunostomy on a Roux-en-Y loop (OJRY), jejunal interposition between the oesophagus and the duodenum (OJD), or an oesophagojejunostomy with a proximal jejunal pouch reservoir (J-pouch) on a Roux-en-Y loop. Blood was collected just before surgery and ten weeks later and peptide levels were analysed by radioimmunoassay. RESULTS: Somatostatin levels were sustained at a high level after TG, regardless of the mode of reconstruction, but were significantly lower in sham-operated animals. Levels of vasoactive intestinal peptide (VIP), neurotensin and motilin were unchanged. CONCLUSION: TG by itself leads to high levels of somatostatin long term, however, somatostatin, motilin, neurotensin and VIP are unaffected by the mode of reconstruction.

Jejunal pouch reconstruction but not preservation of duodenal passage after total gastrectomy reduces plasma cholecystokinin and pancreatic polypeptide long term in pigs.

AIM: The long-term effects of reconstructions of the gastrointestinal tract after gastrectomy on plasma levels of gastrointestinal hormones that contribute to food intake controls were evaluated. MATERIALS AND METHODS: Domestic pigs were randomly assigned to sham-surgery or total gastrectomy followed by reconstruction with oesophagojejunostomy on a Roux-en-Y loop (OJRY), jejunal interposition between the oesophagus and the duodenum (OJD), or an oesophagojejunostomy with a jejunal pouch reservoir (J-pouch) on a Roux-en-Y loop. Plasma levels of peptides were analysed by radioimmunoassay (RIA). RESULTS: Ten weeks after surgery, levels of cholecystokinin (CCK) and pancreatic polypeptide (PP) were significantly lowered (79.6% and 67.0%, respectively) in animals with a J-pouch, but not in sham-operated animals or animals with OJRY or OJD, as compared to preoperative levels. The levels of neuropeptide Y (NPY) and peptide YY (PYY) remained unchanged, irrespective of the mode of reconstruction. CONCLUSION: J-pouch, but not preservation of duodenal passage after total gastrectomy, lowers levels of CCK and PP, peptides that reduce food intake.

Overnight Steady-State Infusions of Parenteral Nutrition on Myosin Heavy Chain Transcripts in Rectus Abdominis Muscle Related to Amino Acid Transporters, Insulin-like Growth Factor 1, and Blood Amino Acids in Patients Aimed at Major Surgery.

BACKGROUND: Evaluation of improvements by nutrition support to severely ill patients requires sensitive methods to demonstrate activation of protein synthesis in various tissues from groups with a limited number of patients to be statistically efficient. This study examines effects of standard parenteral nutrition (PN) on abdominal muscle transcripts of amino acid (AA) transporters, myosin heavy chains (MHCs), and the insulin-like growth factor 1 and its receptor (IGF-1/IGF-1R) in patients aimed at major surgery. METHODS: Twenty-two randomized patients received steady-state PN (0.16 gN/kg/d, 30 kcal/kg/d) or saline infusions for 12 hours before operation. Blood samples and muscle biopsies were obtained at operation start. Muscle messenger RNA (mRNA) levels of AA transporters (solute carrier family members SNAT2, LAT1, LAT3, LAT4, TAUT, PAT1, CD98), IGF-1, IGF-1R, MHC isoforms (MHC1, MHC2A, MHC2X), and LAT3 protein were quantified and related to concentrations of AA, IGF-1, insulin, and metabolic substrates in blood. RESULTS: Muscle mRNA LAT3, LAT4, IGF-1R, and MHC2A increased by PN infusion, with correlations to specific AA transporters and MHC isoforms (P < .01-.05). TAUT and LAT3 correlated to slow (MHC1) and fast (MHC2A, MHC2X) isoforms (P < .001-.02). Muscle IGF-1 mRNA correlated to plasma essential AAs, whereas IGF-1R mRNA was related to LAT3, MHC2A, and serum IGF-1 (P < .001-.03). CONCLUSIONS: The results confirm that short-term preoperative PN activates transcription of AA transporters and myosin isoforms. Thus, combinations of methods on gene transcription and translation of muscle proteins can be applied to define efficient combinations of nutrition and hormones to catabolic patients in preoperative and postoperative settings.

Host knockout of E-prostanoid 2 receptors reduces tumor growth and causes major alterations of gene expression in prostaglandin E2-producing tumors.

Prostaglandin E2 (PGE2) is elevated in a variety of malignant tumors and has been shown to affect several hallmarks of cancer. Accordingly, the PGE2 receptor, E-prostanoid 2 (EP2), has been reported to be associated with patient survival and reduced tumor growth in EP2-knockout mice. Thus, the aim of the present study was to screen for major gene expression alterations in tumor tissue growing in EP2-knockout mice. EP2-knockout mice were bred and implanted with EP2 receptor-expressing and PGE2-producing epithelial-like tumors. Tumor tissue and plasma were collected and used for analyses with gene expression microarrays and multiplex enzyme-linked immunosorbent assays. Tumor growth, acute phase reactions/systemic inflammation and the expression of interleukin-6 were reduced in EP2-knockout tumor-bearing mice. Several hundreds of genes displayed major changes of expression in the tumor tissue when grown in EP2-knockout mice. Such gene alterations involved several different cellular functions, including stemness, migration and cell signaling. Besides gene expression, several long non-coding RNAs were downregulated in the tumors from the EP2-knockout mice. Overall, PGE2 signaling via host EP2 receptors affected a large number of different genes involved in tumor progression based on signaling between host stroma and tumor cells, which caused reduced tumor growth.

MCG101-induced cancer anorexia-cachexia features altered expression of hypothalamic Nucb2 and Cartpt and increased plasma levels of cocaine- and amphetamine-regulated transcript peptides.

The aim of the present study was to explore central and peripheral host responses to an anorexia-cachexia producing tumor. We focused on neuroendocrine anorexigenic signals in the hypothalamus, brainstem, pituitary and from the tumor per se. Expression of mRNA for corticotropin-releasing hormone (CRH), cocaine- and amphetamine-regulated transcript (CART), nesfatin-1, thyrotropin (TSH) and the TSH receptor were explored. In addition, we examined changes in plasma TSH, CART peptides (CARTp) and serum amyloid P component (SAP). C57BL/6 mice were implanted with MCG101 tumors or sham-treated. A sham-implanted, pair­fed (PF) group was included to delineate between primary tumor and secondary effects from reduced feeding. Food intake and body weight were measured daily. mRNA levels from microdissected mouse brain samples were assayed using qPCR, and plasma levels were determined using ELISA. MCG101 tumors expectedly induced anorexia and loss of body weight. Tumor-bearing (TB) mice exhibited an increase in nesfatin-1 mRNA as well as a decrease in CART mRNA in the paraventricular area (PVN). The CART mRNA response was secondary to reduced caloric intake whereas nesfatin-1 mRNA appeared to be tumor-specifically induced. In the pituitary, CART and TSH mRNA were upregulated in the TB and PF animals compared to the freely fed controls. Plasma levels for CARTp were significantly elevated in TB but not PF mice whereas levels of TSH were unaffected. The plasma CARTp response was correlated to the degree of inflammation represented by SAP. The increase in nesfatin-1 mRNA in the PVN highlights nesfatin-1 as a plausible candidate for causing tumor-induced anorexia. CART mRNA expression in the PVN is likely an adaptation to reduced caloric intake secondary to a cancer anorexia-cachexia syndrome (CACS)­inducing tumor. The MCG101 tumor did not express CART mRNA, thus the elevation of plasma CARTp is host derived and likely driven by inflammation.

Presence of TSH receptors in discrete areas of the hypothalamus and caudal brainstem with relevance for feeding controls-Support for functional significance.

AIMS: Previous studies have shown that brain-derived thyroid-stimulating hormone (TSH) and its receptor (TSHr) are present in hypothalamic extracts. No studies investigating both the anatomical location and functional significance of putative TSHr proteins in specific central nervous system (CNS) nuclei involved in feeding controls have yet been conducted. The aim was thus to determine whether TSHr are present in nuclei associated with feeding behavior, and if such receptors may be functional. METHODS: Brain tissue from adult rats was analyzed for gene expression and receptor protein expression was investigated with immunohistochemistry and western blotting. To investigate whether putative TSHr may be functional, we evaluated food intake of rats given intraparenchymal nanoinjections of TSH into the nucleus of the solitary tract (NTS). RESULTS: RT-qPCR confirmed previous reports that TSHr mRNA is expressed in CNS tissues of the adult rat. Immunohistochemistry showed TSHr-immunoreactivity in the arcuate, the ventromedial, the dorsomedial, and the paraventricular hypothalamic nuclei. We also found TSHr-ir in the dorsal hindbrain to be localized to the area postrema, NTS, dorsal motor nucleus of the vagus, and the hypoglossal motor nucleus. Further protein analysis with western blotting showed 120kDa TSHr-ir proteins present in the hypothalamus and brainstem. Injections of TSH into the NTS reduced food intake similar to the positive control, urocortin. CONCLUSIONS: These data suggest that functional TSHr are present in the caudal brainstem and hypothalamic nuclei of relevance for feeding control as a possibly uncleaved holoreceptor, and highlights a hindbrain component to central TSH inhibition of food intake.

Fourth ventricular CART peptide induces c-fos in the area postrema and nucleus of the solitary tract via a CRF-receptor dependent mechanism.

Cocaine-and amphetamine-regulated transcript peptides (CARTp) suppress gastric emptying and nutritional intake following 4th icv administration. Whereas, the CARTp inhibition of gastric emptying was blocked by pre-treatment with a non-selective corticotropin releasing factor (CRF) antagonist, sucrose drinking was not, suggesting that CARTp- and CRF controls for food intake and gastric emptying are operated through separable dorsal hindbrain mechanisms. The aim of the study was to explore CARTp-CRF brainstem interactions on patterns of neuronal activation in areas of the brainstem and midbrain relevant to gastrointestinal control and feeding regulation. Rats received 4th icv injections of combinations of vehicle, CARTp (1µg), or the nonselective CRF antagonist, α-helical CRF9-41 (αCRF), in a randomized order. Brain sections were processed for c-fos by immunohistochemistry followed by image analysis at defined levels of the brain. CARTp (1µg, 4th icv) induced a robust c-fos response in the nucleus of the solitary tract (NTS) and area postrema (AP), whereas, no c-fos could be detected in the parabrachial nucleus (PBN), the paraventricular nucleus of the hypothalamus (PVN) or the arcuate nucleus of the hypothalamus (ARC). The c-fos expression in the structures of the dorsal vagal complex (DVC) was completely blocked by pre-treatment with the CRF antagonist, which did not by itself induce c-fos at any examined level. After CARTp and αCRF in combination, there was a tendency towards an increased c-fos response in the ARC. We conclude that CARTp activates cells of the area postrema and NTS via a downstream, CRF-dependent mechanism.

Food intake, tumor growth, and weight loss in EP2 receptor subtype knockout mice bearing PGE2-producing tumors.

Previous studies in our laboratory have demonstrated that prostaglandin (PG) E2 is involved in anorexia/cachexia development in MCG 101 tumor-bearing mice. In the present study, we investigate the role of PGE receptor subtype EP2 in the development of anorexia after MCG 101 implantation in wild-type (EP2 (+/+)) or EP2-receptor knockout (EP2(-/-)) mice. Our results showed that host absence of EP2 receptors attenuated tumor growth and development of anorexia in tumor-bearing EP2 knockout mice compared to tumor-bearing wild-type animals. Microarray profiling of the hypothalamus revealed a relative twofold change in expression of around 35 genes including mRNA transcripts coding for Phospholipase A2 and Prostaglandin D2 synthase (Ptgds) in EP2 receptor knockout mice compared to wild-type mice. Prostaglandin D2 synthase levels were increased significantly in EP2 receptor knockouts, suggesting that improved food intake may depend on altered balance of prostaglandin production in hypothalamus since PGE2 and PGD2 display opposing effects in feeding control.