Lipid-rich enteral nutrition regulates mucosal mast cell activation via the vagal anti-inflammatory reflex.

Nutritional stimulation of the cholecystokinin-1 receptor (CCK-1R) and nicotinic acetylcholine receptor (nAChR)-mediated vagal reflex was shown to reduce inflammation and preserve intestinal integrity. Mast cells are important early effectors of the innate immune response; therefore modulation of mucosal mast cells is a potential therapeutic target to control the acute inflammatory response in the intestine. The present study investigates intestinal mast cell responsiveness upon nutritional activation of the vagal anti-inflammatory reflex during acute inflammation. Mucosal mast cell degranulation was induced in C57/Bl6 mice by administration of Salmonella enterica LPS. Lipid-rich enteral feeding prior to LPS significantly decreased circulatory levels of mouse mast cell protease at 30 min post-LPS compared with isocaloric low-lipid nutrition or fasting. CCK-1R blockage reversed the inhibitory effects of lipid-rich feeding, whereas stimulation of the peripheral CCK-1R mimicked nutritional mast cell inhibition. The effects of lipid-rich nutrition were negated by nAChR blockers chlorisondamine and α-bungarotoxin and vagal intestinal denervation. Accordingly, release of ß-hexosaminidase by MC/9 mast cells following LPS or IgE-ovalbumin complexes was dose dependently inhibited by acetylcholine and nicotine. Application of GSK1345038A, a specific agonist of the nAChR α7, in bone marrow-derived mast cells from nAChR ß2-/- and wild types indicated that cholinergic inhibition of mast cells is mediated by the nAChR α7 and is independent of the nAChR ß2. Together, the present study reveals mucosal mast cells as a previously unknown target of the nutritional anti-inflammatory vagal reflex.

Cholecystokinin/Cholecystokinin-1 receptor-mediated peripheral activation of the afferent vagus by enteral nutrients attenuates inflammation in rats.

OBJECTIVE: The current study investigates activation of the nutritional anti-inflammatory pathway by lipid-rich nutrition. BACKGROUND: Enteral nutrition activates humoral and neural pathways to regulate food intake and sustain energy balance. Recently, we demonstrated that enteral nutrition and in particular lipid-rich nutrition modulates inflammation and prevents organ damage. METHODS: Male rats were fasted or fed lipid-rich nutrition before hemorrhagic shock. Disruption of afferent vagal fibers with capsaicin (deafferentation) was used to investigate involvement of afferent fibers. Peripheral activation of afferent vagal fibers via cholecystokinin (CCK)-mediated activation of CCK-1 receptors was investigated using administration of the selectively peripheral acting CCK-1 receptor antagonist, A70104 and PEGylated-CCK9. Tissue and blood were collected 90 minutes after shock to assess systemic inflammation and intestinal integrity. RESULTS: Deafferentation reversed the inhibitory effect of lipid-rich nutrition on systemic levels of tumor necrosis factor-alpha and interleukin-6, and on intestinal leakage of horseradish peroxidase and bacterial translocation. Furthermore, the protective effects of lipid-rich nutrition were negated by A70104, indicating that lipid-rich nutrition triggers peripheral CCK-1 receptors on vagal afferents to modulate inflammation. These findings were substantiated by the fact that pretreatment of fasted rats with PEGylated-CCK9, which acts on peripheral CCK-1 receptors, attenuated systemic inflammation, and loss of intestinal integrity. CONCLUSION: These data demonstrate that enteral lipid-rich nutrition modulates inflammation and preserves intestinal integrity via CCK release which activates CCK-1 receptors located on afferent vagal fibers. Taken together, the current study reveals a novel gut-brain-immune axis and provides new insight into the applicability of enteral nutrition to treat inflammatory conditions.

Disruption of the behavioral satiety sequence by simmondsin.

Simmondsin, a cyanoglycoside from jojoba meal, reduces food intake after oral administration. To diagnose if it acts by inducing satiation or by creating abnormal physiological effects, an observational study was undertaken to investigate the effects of simmondsin on feeding and other behaviors. Particular attention was paid to the behavioral sequence associated with satiety (BSS). At first contact, simmondsin non-significantly reduced food intake by 17% and had little effect on feeding and associated behaviors. The behavioral structure was preserved and a small shift of the onset of resting to the left was observed, suggesting a small satiative action of simmondsin at first contact. Simmondsin given for the second time caused a more pronounced food intake reduction of 52% due to a reduction in eating duration, mean bout intake and mean bout length, and to an increase in latency to eat. At second contact, simmondsin caused a strong switching in active behaviors, disrupting the BSS. The simmondsin-induced hyperactivity suggests that simmondsin produces aversiveness with second contact. Our results indicate that simmondsin exerts multiple effects. It probably facilitates a small natural process of satiation/satiety at first contact, but creates abnormal physiological effects resulting in aversive reactions from second contact on.

PEGylation of cholecystokinin prolongs its anorectic effect in rats.

The anorectic compound CCK-9 was coupled to polyethylene glycol 5 kDa, 10 kDa, 20 kDa and 30 kDa, under different reaction conditions. Conjugates were purified by HPLC and characterized by MALDI-TOF MS. A 96% PEGylation yield was obtained in buffer pH 7.5 after 6h reaction at 20 degrees C. The anorectic activity was tested in vivo in rats. A single bolus intra-peritoneal injection of non-modified CCK-9 resulted in a significant initial food intake reduction 30 min after food presentation (87% compared to paired control group). When PEG-CCK-9 conjugates modified with polymers of molecular weight up to 20 kDa were injected, lower but statistically significant initial food intake reductions were obtained (76% for PEG 10 kDa-CCK-9 conjugate compared to control group). The cumulative food intake reduction of non-modified CCK-9 is normalized within 1-2h, whereas the PEG-CCK-9 molecules showed a prolonged anorectic activity lasting for 6h for PEG 5 kDa-CCK-9; 23 h for PEG 10 kDa-CCK-9 and between 8h and 23 h for PEG 20 kDa-CCK-9. For PEG 30 kDa-CCK-9 conjugate, neither an initial nor a cumulative FI reduction was observed. PEG-CCK-9 conjugates show a significantly prolonged anorectic activity in comparison to the non-modified peptide. This effect is most evident for the PEG 10 kDa-CCK-9 conjugate.

Biodistribution and pharmacokinetics of PEG-10kDa-cholecystokinin-10 in rats after different routes of administration.

Cholecystokinin, produced in the proximal small intestine, is a short acting satiating peptide hormone. CCK-10, before and after mono-iodination, was previously coupled to 10kDa polyethylene glycol (PEG). The formed conjugates PEG10kDa-CCK-10 and PEG10kDa-[(127)I]-CCK-10 show after i.p. administration to rats a sustained food intake reduction during 8h in comparison to 2h for free CCK-10. The present study examined the blood pharmacokinetics of this pharmacological interesting molecule by means of PEG10kDa-[(123)I]-CCK-10 following intravenous, intraperitoneal, intramuscular and nasal administration and the biodistribution after i.p. administration. HPLC analysis with radiometric detection allowed the differentiation between inorganic iodide and the intact tracer in blood. Blood kinetics after i.v. injection was fitted to a bi-exponential with a distribution half-life of 15 min and with an elimination half-life of 8 hours for intact PEG10kDa-[(123)I]-CCK-10. The biodistribution studies showed a higher accumulation of the tracer for all administration routes in organs expressing CCK receptors localized in the gastrointestinal tract such as pancreas, duodenum and small intestine. No indication of blood brain barrier crossing for the conjugate could be observed independently of the administration route. Main clearance was via the urinary pathway.

Lack of tolerance development with long-term administration of PEGylated cholecystokinin.

Cholecystokinin (CCK) is a short acting satiating peptide hormone produced in the proximal small intestine. Daily CCK injection in rats initially inhibits food intake, but after several days, food intake is no longer affected, suggesting development of tolerance. Previously, we covalently coupled CCK to a 10kDa polyethylene glycol (mPEG-OH) and showed that this conjugate, PEG-CCK(9), produced a significantly longer anorectic effect than unmodified CCK(9). The present study examined whether tolerance to the anorectic effect develops during long-term administration of PEG-CCK(9). For 14 consecutive days, male Wistar rats (n=12) received a daily i.p injection of 8microgkg(-1) of PEG-CCK(9) and a control group received a daily control injection of mPEG-OH. Body weight and food intake were monitored daily during the experiment. Effects on the pancreas were investigated. On each day, injection of PEG-CCK(9) induced an anorectic effect lasting 3-6h, but failed to significantly reduce daily total food intake compared to controls. The body weight gain of the PEG-CCK(9)-treated animals was not different from controls. The PEG-CCK(9)-treated group had a significantly higher pancreas weight, mainly due to hyperplasia. In conclusion, PEG-CCK(9) continued to have a daily suppressive effect on food intake when administered for 14 consecutive days, showing there was no development of tolerance.