The Regulation Roles Of Corticotropin Releasing Factor Receptors In Decreasing Hypermetabolism By Early Enteral Nutrition In Severely Burned Rats

The hypermetabolism is an important part of the stess response to burn injury.Suitable response of hypermetabolism is beneficial,and excessive response is harmful to burn organism.The central nervous system play an important role in burn hypermetabolism response.More evidences showed that there were intercommunications among nervous,immune and endocrine system,which gave us new idea of burn hypermetabolism mechanism and metabolism intervention.The regulation mechanism of energy balance is more complicated.The hypothalamus is a center of energy balance regulation.The burn injury hypermetabolism is also regulated by hypothalamus.The hypothalamus play a critical role in energy balance.Along with neurobiology advancement in twenty years,many hypothalamic neuropeptides and neurotransmitters related to energy balance were discovered gradually,which provided us with more importance rationale to search for burn injury hypermetabolism mechanism.The anabolic neuropeptides provoke ingestion,lower energy expenditure,and catabolic neuropeptides restrain ingestion,increase energy expenditure.The corticotrophin releasing factor(CRF) system belongs to catabolic neuropeptides.There were some evidences of CRF related peptides and its receptor roles in stress and energy metabolism. Roles of many other neuropeptides may go through CRF system.We presumed that the hypothalamic CRF related peptides and its receptors may play an important role in burn stress and hypermetabolism response. The hypothalamic CRF related peptides include CRF ,Urocortin1(UCN1),UCN2 and UCN3. Corticotropin-releasing factor (CRF) is a 41-residue peptide characterized from ovine hypothalamus by Vale et al in 1981.Within the brain, CRF binds to two receptor types,referred to as the CRFR1 and the CRFR2 receptors.The corticotrophin releasing factor (CRF) system could play diverse pathophysiologic role in stress response. The CRFR1 is the primary receptor that activates the secretion of ACTH and cortisone of hypothalamic-pituitary-adrenal(HPA) axis in response to a stressor,CRFR2 receptor regulates energy balance.Urocortin1 is a 40-amino acid peptide expressed in the mammalian brain.Urocortins show a particularly high specificity for CRFR2 receptor.Recent investigations have demonstrated that urocortins could play a significant role in the regulation of burn injury hypermetabolism.Recently,CRFR1,CRFR2 receptor specific antagonist had been discovered successively,astressin2-B was specific antagonist of CRFR2 receptor,antalarmin was specific antagonist of CRFR1 receptor.Meanwhile,antisense nucleic acids technique was a good way which can lower gene expression.This specific antagonist of CRF receptors and antisense nucleic acid technique had become a valuable tool for in vivo characterization of CRFR2 receptor function.Hypermetabolism of burn injury is mainly from wound,the gut is another importance source.The intestinal tract is a central organ to manifest surgical stress.The view that hypermetabolism is regulated by gut has gained acceptance in recent years.On the basis of many animal experiments and clinical studies in our institute,we concluded that early enteral nutrition can lower burn injury hypermeatbolism,and proposed"Enterogenous Hypermetabolism"theory.Early enteral nutrition is an essential stimulant to the growth and renewal of the mucosal cells of gastrointestinal tract.It could regulate the gastrointestinal tract movement,secretion and cell metabolism,lower intestinal ischemia injury,preserve the structure and function of gastrointestinal tract,reduce intestinal bacteria/endotoxin translocation,decrease hypermetabolism response.But,studies now only refer to the effects of early enteral nutrition on REE and the structure and function of gastrointestinal tract.As is known to all that hypermetabolism is regulated by central nervous system,there were no any data of central neuropeptides related to burn hypermetabolism by gut and the effect of early enteral nutrition on burn hypermetabolism.We suppose that the effect of early enteral nutrition on REE is through central neuropeptides related to metabolism.CRF related peptides and its receptors were also discovered expressing in gastrointestinal tract.Functional studies demonstrated that peripheral injection of CRF or Ucn 1 can also stimulate colonic transit and defecation,inhibit gastric emptying in rats.CRF or UCN 1 didn't pass through blood-brain barrier,it indicated that there were peripheral CRF,UCN 1,CRFR1 and CRFR2 receptors in gastrointestinal tract.Recently,some published data show that CRF related peptides and its receptors in gastrointestinal tract mucosa,submucous layer play an important role in gut stress response,inflammatory reaction and mucosal barrier function.But until now,there is no data showing that the changes of CRF related peptides and its receptors and effects of early enteral nutrition on CRF related peptides and its receptors in gastrointestinal tract after burn injury.ObjectivesTo investigate the relation between CRF receptors and burned hypermetabolism response,and observe the effects of early enteral nutrition on hypermetabolism,hypothalamus and ileum mucosa CRF related peptides and its receptors.Methods1. In this study we employed a stainless-steel cannula implanted into the 3rd ventricle animal model, after at least one week to recover from these operations, SD rats were inflicted with 30% TBSA full thickness flame burn on the back,to observe the effects of CRFR1ASODN,CRFR2ASODN and astressin2-B,antalarmin on resting energy expenditure(REE) at PBD7.The hypothalamic CRFR1mRNA,CRFR2mRNA expression level were determined by RT-PCR at PBD7 after CRFR1ASODN,CRFR2ASODN injection.The changes of hypothalamic CRFR1,CRFR2 were determined at PBD7 after CRFR1ASODN, CRFR2ASODN injection by Western blot and immunohistochemistry.2. The animal model of early enteral nutrition and parenteral nutrition was employed,SD rats were divided randomly into control group,parenteral nutrition(PN) group and early enteral nutrition(EN) group,and inflicted with 30% TBSA full thickness flame burn on the back.The resting energy expenditure(REE) were determined at PBD1, 3, 5, 7 respectively,and plasma ACTH,CORT,CA,and serum TNFα,IL-1β,IL-6 were detected at 1d, 3d, 5d, 7d postburn respectively.The hypothalamic CRFR2mRNA,CRFmRNA and UCN1mRNA were detected respectively by RT-PCR.The hypothalamic CRFR2 and CRF were detected respectively by Western blot.3. We employed early enteral nutrition and parenteral nutrition animal model, SD rats were divided randomly into control group,parenteral nutrition(PN) group and early enteral nutrition(EN) group,and inflicted with 30% TBSA full thickness flame burn on the back.The ileum mucosal CRFR1mRNA,CRFR2mRNA,CRFmRNA and UCN1mRNA were detected respectively by RT-PCR.The ileum mucosal CRFR1,CRFR2, CRF were detected respectively by Western blot.The ileum CRFR1 and CRFR2 immunohistochemistry expression were investigated.Results1. REE in CRFR2ASODN group were significantly lower than that of CRFR2ODN group.There was no significant difference of REE between CRFR1ASODN group and CRFR1ODN group.Compared with burn control group,astressin2-B lowered REE significantly.There was no significant difference of REE between antalarmin group and burned control group.CRFR1ASODN,CRFR2ASODN significantly lowered hypothalamic CRFR1mRNA,CRFR2 mRNA and protein level expression in burned rats.2. There was an obvious increasing trend of REE after severe burn injury.REE in EN group was significantly lower than that of PN group on PBD3,5,7 respectively (P<0.05).Plasma ACTH and CORT increased and peaked at PBD3,and subsided gradually after PBD3.Plasma ACTH and CORT in EN group were significantly lower than that of PN group on 3d,5d postburn respectively(P<0.05).Plasma CA increased and peaked at PBD1,and subsided gradually after PBD1.Plasma CA in EN group was significantly lower than that of PN group on 3d,5d postburn respectively(P<0.05).Serum TNFα,IL-1β,IL-6 increased gradually after burn injury, Serum TNFα,IL-1β,IL-6 in EN group were significantly lower than that of PN group on 3d,5d,7d postburn respectively(P<0.05).3. The hypothalamic CRFR2mRNA and protein expression increased gradually after burn injury. The hypothalamic CRFR2mRNA and protein expression in EN group were obviously lower than that of PN on 5d,7d postburn respectively.The hypothalamic CRFmRNA and protein expression decreased gradually after burn injury and peaked at PBD3.The hypothalamic CRFmRNA and protein expression in EN group were obviously higher than that of PN on 3d 5d,7d postburn respectively.The hypothalamic UCN1mRNA expression increased gradually after burn injury ,and UCN1mRNA expressions in EN group were obviously lower than that of PN on 5d,7d postburn respectively.4. The ileum mucosa CRFmRNA,UCN1mRNA,CRFR1mRNA,CRFR2mRNA expression in normal control group rats were lower and increased gradually after burn injury.Early enteral nutrition lowered obviously ileum mucosa CRFmRNA, CRFR1mRNA and protein expressions, and increased UCN1mRNA,CRFR2mRNA and protein expressions.5. The immunohistochemistry results also indicated that CRFR1 and CRFR2 receptor expressed in myenteric nerve plexus,submucous layer and mucosa of normal rats and increased significantly after burn injury.Early enteral nutrition lowered CRFR1 expression and increased CRFR2 expression in ileum mucosa.Conclusions1. The hypothalamic CRFR2 receptor is a main regulator and CRFR1 receptor may be of no importance in severely burned hypermetabolism response.2. Early enteral nutrition can lower severely burned hypermetabolism and plays very important roles in mediating hypermetabolism by decreasing plasma ACTH, CORT,CA and serum TNFα,IL-1β,IL-6.3. Early enteral nutrition plays important roles in lowering hypermetabolism by decreasing hypothalamic CRFR2mRNA and protein expression.The hypothalamic UCN1 may take part in hypermetabolism response, hypothalamic CRF may relate to burn stress response.4. CRFR1,CRFR2 expression in rat myenteric nerve plexus, submucous layer and mucosa were discovered.Early enteral nutrition lowered obviously ileum mucosa CRFmRNA,CRFR1mRNA and protein expressions,and increased UCN1mRNA, CRFR2mRNA and protein expressions.