Neurotensin receptor type 1: Escherichia coli expression, purification, characterization and biophysical study.

NT (neurotensin) is an endogenous tridecapeptide neurotransmitter found in the central nervous system and gastrointestinal tract. One receptor for NT, NTS1, belongs to the GPCR (G-protein-coupled receptor) superfamily, has seven putative transmembrane domains, and is being studied by a range of single-molecule, functional and structural approaches. To enable biophysical characterization, sufficient quantities of the receptor need to be expressed and purified in an active form. To this end, rat NTS1 has been expressed in Escherichia coli in an active ligand-binding form at the cell membrane and purified in sufficient amounts for structural biology studies either with or without fluorescent protein [YFP (yellow fluorescent protein) and CFP (cyan fluorescent protein)] fusions. Ligand binding has been demonstrated in a novel SPR (surface plasmon resonance) approach, as well as by conventional radioligand binding measurements. These improvements in production of NTS1 now open up the possibility of direct structural studies, such as solid-state NMR to interrogate the NT-binding site, EM (electron microscopy), and X-ray crystallography and NMR.

Negative regulation of leucocyte functions by CD33-related siglecs.

The siglecs (sialic acid-binding Ig-like lectins) are a family of transmembrane receptors expressed in the haemopoietic, immune and nervous systems. The CD33-related siglecs are a distinct subset mostly expressed in the innate immune system where they can function as inhibitory receptors by suppressing the signalling mediated by receptors coupled with ITAMs (immunoreceptor tyrosine-based activation motifs). CD33-related siglecs contain ITIMs (immunoreceptor tyrosine-based inhibitory motifs) that recruit and activate SHP-1 [SH2 (Src homology 2) domain-containing phosphatase-1] and SHP-2. In addition, the ITIMs of CD33-related siglecs can suppress siglec-dependent adhesion of sialylated ligands and mediate endocytosis. Siglec-H is a recently characterized murine CD33-related endocytic receptor that lacks intrinsic tyrosine-based signalling motifs and is expressed selectively on PDCs (plasmacytoid dendritic cells). Siglec-H depends on DAP12 (DNAX-activating protein of 12 kDa) for surface expression and cross-linking with anti-siglec-H antibodies can selectively inhibit interferon-alpha production by PDCs following TLR9 (Toll-like receptor 9) ligation. Thus CD33-related siglecs are able to mediate diverse inhibitory functions of leucocytes in the innate immune system via both ITIM-dependent and -independent pathways.

Clinical evaluation of two-liter prepacked enteral diet delivery system: a controlled trial.

Following a recent study in our Unit which showed that significantly more of prescribed enteral diet was administered from 2-liter bags than 0.51 bottles, a controlled clinical trial was undertaken to investigate further the effectiveness of the 2-liter prepackaged enteral delivery system. Forty patients with normal gastrointestinal function requiring enteral nutrition were randomized to receive nasogastrically 2-liter/day of polymeric diet (2000 kcal, 12 g N) either from a presterilized 2-liter PVC bag (Express EFI Ltd, UK) or from two 1-liter rigid plastic containers filled with cans of Clinifeed 400 (Roussel Laboratories Ltd, UK). The actual volumes of feed infused were monitored in patients fed for 5 days or more. Both nursing and dietetic staff preferred the 2-liter bags which were more convenient to prepare and use. Significantly larger quantities of prescribed diet were delivered from the 2-liter bags than from the 1-liter containers (p less than 0.001). It was concluded that the 2-liter bag system was potentially more convenient and nutritionally efficacious than 1-liter containers. Although the incidence of microbial contamination associated with the 2-liter bags was low, measures to maintain the sterility of this system may need to be more stringent.

Clinical efficacy and design changes of "fine bore" nasogastric feeding tubes: a seven-year experience involving 809 intubations in 403 patients.

We report here our clinical experiences of "fine bore" nasogastric feeding tubes. Data have been collated over a 7-year period (1978-1985). A total of 403 patients were intubated on 809 occasions. In the first retrospective study, the clinical use of 491 unweighted tubes was compared with that of fifty 3.5-g weighted tubes. No advantage was found in the use of the weighted tubes. In the second prospective controlled clinical trial, these results were confirmed. Forty-six patients were intubated on 76 occasions with an 85-cm open-ended, unweighted nasogastric feeding tube (Prima, Portex UK), and 57 patients were intubated on 79 occasions with a 91-cm 3.0-g weighted tube (Entriflex, Biosearch, Raritan, NJ). Mean duration of placement was similar in each case, and 62% of both types of tubes were inadvertently removed. Without exception, all the tubes remained in the stomach throughout. Disappointed with the similar and overall performance of both types of tubes, we initiated a design program which resulted in the development of two new nasogastric tubes, one weighted and one unweighted. The tubes were manufactured with polyurethane, rather than polyvinylchloride (PVC), which permitted an increase in diameter of the internal lumen which, in turn, was coated with water-activated lubricant to ease removal of the introducer wire. A specially modeled outflow port was incorporated into the tips of both tubes. The performance of the two new polyurethane nasogastric feeding tubes was assessed under controlled trial conditions; as a reference, a widely used PVC unweighted open-ended tube was used.(ABSTRACT TRUNCATED AT 250 WORDS)

Improved design of nasogastric feeding tubes.

Disappointed with the overall performance of weighted and unweighted nasogastric feeding tubes, a design programme was initiated which resulted in the development of two new nasogastric tubes, one weighted and one unweighted. The tubes were manufactured with polyurethane rather than polyvinylchloride (PVC) which permitted an increase in diameter of the internal lumen which in turn was coated with water activated lubricant to ease removal of the introducer wire. A specially modelled outflow port was incorporated into the tips of both tubes. The performance of the two new polyurethane nasogastric feeding tubes was assessed under controlled trial condition using as a reference a widely used PVC unweighted open ended tube. While intubation times were similar in patients without concurrent endotracheal intubation, it took a significantly shorter time to intubate patients with concurrent endotracheal intubation with the new weighted tube. Following tube intubation, it was possible to aspirate gastric contents significantly more often through the new polyurethane tubes (p < 0.001) than through the PVC tube, and the unweighted polyurethane tube stayed in situ longer (p < 0.05) than the PVC tube. The newly designed polyurethane nasogastric feeding tubes are the first tubes that have been shown to have advantages over the simpler type of open ended, unweighted PVC nasogastric feeding tubes.

Elemental diet administered nasogastrically without starter regimens to patients with inflammatory bowel disease.

The present study questions the concept of routinely using 'starter regimens' at the outset of enteral feeding with chemically defined elemental diets. A hypertonic elemental diet with an osmolality of 630 mOsm/kg was administered by 24-hr nasogastric infusion to 12 patients with exacerbations of inflammatory bowel disease and to two patients with short bowel syndrome. Starter regimens were not used. Upper gastrointestinal symptoms of nausea, abdominal bloating, and colicky pain occurred transiently in only five of 14 patients. Stool frequency did not increase during full-strength feeding, and daily stool weights decreased significantly (p less than 0.01). These findings show that it is safe to administer undiluted hypertonic elemental diets by constant nasogastric infusion to patients with inflammatory bowel disease. Avoiding starter regimens leads to increased nutrient intake and improved nitrogen balance.

Administration of fat emulsions with nutritional mixtures from the 3-liter delivery system in total parenteral nutrition.

A series of studies was performed to test the efficacy and safety of a parenteral lipid emulsion, Lipofundin S, when given as part of a complete nutritive mixture from the three-liter bag total parenteral nutrition (TPN) delivery system. In vitro stability studies with mixtures corresponding to high and low nutritional intakes showed the fat emulsion to be stable during refrigerated storage for at least 6 days. The clinical use of Lipofundin S in 3-liter TPN bags was studied in 39 consecutive patients requiring TPN, and there were no untoward side-effects. Nitrogen balance was maintained in patients with pancreatitis, those recovering postoperatively, and those with miscellaneous conditions. However, patients with multiple trauma remained in negative balance. The ability of sera, from patients on TPN to agglutinate Lipofundin S was compared to that from healthy controls, and acutely ill patients not on TPN. Patients on TPN showed a higher degree of in vitro creaming than acutely ill controls, and this may have been related to the severity of the underlying illness. These studies suggest that this parenteral lipid emulsion can be safely administered to patients requiring TPN when given from the 3-liter bag delivery system.

Do fibre containing enteral diets have advantages over existing low residue diets?

This study investigated the influence of fibre on the pattern of absorption of protein and carbohydrate following administration of polymeric enteral diet and also the effect of added fibre on frequency of bowel action, stool weight and gastrointestinal side effects during enteral nutrition. No difference was seen in frequency of bowel action, stool weight or gastrointestinal side effects in five patients fed with either a fibre free polymeric diet or with the same diet augmented with 24 g fibre/24 h. Addition of fibre did not significantly alter breath hydrogen excretion. In an oral tolerance test on six normal subjects, the post prandial rises in blood glucose and levels of 17 amino acids were similar on ingestion of a fibre containing or fibre free test meal.

Relation between osmolality of diet and gastrointestinal side effects in enteral nutrition.

One hundred and eighteen patients with normal gastrointestinal function were randomly allocated to one of three feeding regimens in a double blind study to determine the relation between the tonicity of the diet and gastrointestinal side effects related to the diet and to evaluate the efficacy of "starter" regimens in reducing gastrointestinal side effects during enteral nutrition. Patients received a hypertonic diet with an osmolality of 430 mmol (mosmol)/kg (group 1), the same diet but with the osmolality increasing from 145 to 430 mmol/kg over the first four days (group 2), or an isotonic diet (300 mmol/kg) (group 3). All diets were prepared aseptically and administered by 24 hour nasogastric infusion. The mean daily nitrogen intake in group 1 was significantly greater (p less than 0.05) than that in both groups 2 and 3, and the mean overall daily nitrogen balance was significantly better (p less than 0.05) in group 1 than groups 2 and 3. The incidence of side effects related to the diet was similar in all three groups, but diarrhoea was significantly (p less than 0.001) associated with concurrent treatment with antibiotics. These findings show that undiluted hypertonic diet results in significantly better nitrogen intake and balance, that starter regimens reduce nutrient intake but not symptoms, and that diarrhoea is significantly related to treatment with antibiotics and not to administration of an undiluted hypertonic polymeric diet.

Effect of catheter tunnelling and a nutrition nurse on catheter sepsis during parenteral nutrition. A controlled trial.

In a three-year controlled trial of subcutaneous catheter tunnelling as a method of reducing total parenteral nutrition (TPN) catheter sepsis 99 silicone catheters (52 tunnelled, 47 untunnelled) were inserted into the subclavian (94%) or jugular (6%) veins under aseptic conditions. The influence of a nutrition nurse, who joined the nutrition team after 18 months, on catheter sepsis rate was also documented. Catheter sepsis was confirmed in 13 of 47 (28%) untunnelled catheters and only 6 of 52 (11.5%) tunnelled catheters (p less than 0.05). A nutrition nurse reduced sepsis rate from 33% (tunnelled 6, untunnelled 11) to 4% (0 tunnelled; 2 untunnelled) (p less than 0.001). There was no significant difference between tunnelled and untunnelled catheters in sepsis rates after the arrival of the nutrition nurse. Although 85% patients had concurrent internal sepsis, the pathogens implicated in catheter sepsis came from superficial sites in 16 of 19 cases (p less than 0.01). Rigorous aseptic nursing care is thus the most significant factor in the reduction of TPN catheter sepsis, but tunnelling can reduce sepsis rate when nursing care is suboptimum.

Enteral nutrition in malnourished patients with hepatic cirrhosis and acute encephalopathy.

Ten patients with histologically proven cirrhosis, admitted in grade I to II acute hepatic coma, received in addition to a standard dietary protein free "anticoma" regime, a continuous nasogastric infusion of a branched-chain amino acid enriched chemically defined enteral diet (Hepaticaid) containing an equivalent of 70 g protein/day for a mean of 7.3 days (range 3-23). No complications of therapy were observed and, specifically, the use of fine bore tubes did not provoke variceal hemorrhage. Overall positive nitrogen balance (4.3 +/- 1.7 g N/day) was observed in patients fed 7 or more days. Improvement to coma grade 0 was seen in all patients save one, who died from hepatorenal failure. While serum ammonia levels remained elevated during the study period, there was a significant increase in the branched-chain/aromatic ratio (p less than 0.01) as plasma branched-chain amino acids rose (p less than 0.05) and tyrosine levels fell (p less than 0.05).

Limitations and drawbacks of 'fine bore' nasogastric feeding tubes.

The clinical use of unweighted nasogastric feeding tubes (n = 491) was compared with that of weighted nasogastric feeding tubes. No advantage was found in the use of the weighted tubes.

A controlled trial of aseptic enteral diet preparation--significant effects on bacterial contamination and nitrogen balance.

This study was performed to evaluate the clinical effectiveness of the 'standard' and aseptic enteral diet preparation techniques and to investigate the relationship between bacterial contamination of diets and symptoms during enteral nutrition. In this controlled study patients were prescribed isonitrogenous polymeric diets prepared using either 'standard' or aseptic techniques, administered by 24 nasogastric infusion. 'Standard' diet preparation resulted in significant bacterial contamination (p<0.01), but this did not increase incidence of diet related symptoms. Use of sterile techniques permitted safe use of large volumes (1.5-2 litre) containers, and this resulted in significantly better nitrogen intake (p<0.05) and balance (p<0.05).

Clinical indications for weighted enteral feeding tubes.

Simple unweighted fine bore feeding tubes have been used by our Nutritional Support Team for routine nasogastric feeding with success in large numbers of patients. Three clinical situations where mercury or tungsten weighted tubes offer advantages over fine bore tubes have been defined. Significant advantages in patients with concurrent endotracheal intubation, gastric atony and severe oesophageal stricturing are described.

The roles of lactose and Clostridium difficile in the pathogenesis of enteral feeding associated diarrhoea.

This study investigated the influence of the disaccharide lactose on the incidence of clinically significant enteral feeding associated diarrhoea. In this double blind study both groups each of 25 patients were randomised to receive either a lactose containing diet Clinifeed 400 in 25 patients or a lactose free diet Ensure in 25 patients. Diarrhoea occurred with equal frequency in both treatment groups, even in those patients with symptomatic and biochemical evidence of impaired lactose handling. Although the onset of diarrhoea was significantly associated with antibiotic administration (p<0.01), Cl. difficile was not isolated from the stools of any patient with diarrhoea.

Endoscopic placement of fine bore nasogastric and nasoenteric feeding tubes.

In a small proportion of patients requiring enteral nutrition it may not be possible to site nasogastric or nasoenteric feeding tubes using standard intubation techniques. We describe an endoscopic method of tube placement applicable not only for positioning nasogastric feeding tubes in patients with coexisting oesophageal pathology, but also for placement of nasoenteric feeding tubes when disordered gastric emptying is present.