The dietary form of choline during lactation affects maternal immune function in rats.

PURPOSE: The present study was designed to determine the effects of both choline form and availability on maternal immune function during lactation. METHODS: Sprague-Dawley rats were randomized to one of the three diets 24-48 h before parturition and fed ad libitum until 21 days postnatal: 1 g/kg choline as free choline (C, n = 11), the current form, and amount of choline in commercial diets; 1 g/kg choline as phosphatidylcholine (PC1, n = 11); or 2.5 g/kg choline as PC (PC2.5, n = 8). Choline metabolites in offspring stomach contents were quantified. At 21 days, lymphocytes from mothers' mesenteric lymph nodes and spleens were isolated and phenotypes and ex vivo cytokine production after mitogen exposure were determined. RESULTS: There was a higher proportion of choline and a lower proportion of lyso-PC in stomach contents (representing dam's milk) of C pups compared to PC1. In the mesenteric lymph nodes, feeding PC1 compared to C led to a higher IL-2 production after Concanavalin A (ConA) stimulation and a higher proportion of T cells (CD3+) and a lower proportion of B cells [immunoglobulin (Ig)κ, CD45RA+, and IgM+; P < 0.05]. Splenocytes from the PC1 group produced more IL-6 and TNF-α after lipopolysaccharides stimulation compared to C (P < 0.05). Splenocytes from the PC2.5 group produced more IL-2 and IL-6 after ConA stimulation compared to PC1 (P < 0.05). CONCLUSIONS: Feeding choline as PC in the maternal diet improved the ability of immune cells to respond ex vivo to mitogens and increasing the amount of PC in the diet further improved T cell proliferation.

Allergen exposure chambers: harmonizing current concepts and projecting the needs for the future - an EAACI Position Paper.

BACKGROUND: Allergen exposure chambers (AECs) are clinical facilities allowing for controlled exposure of subjects to allergens in an enclosed environment. AECs have contributed towards characterizing the pathophysiology of respiratory allergic diseases and the pharmacological properties of new therapies. In addition, they are complementary to and offer some advantages over traditional multicentre field trials for evaluation of novel therapeutics. To date, AEC studies conducted have been monocentric and have followed protocols unique to each centre. Because there are technical differences among AECs, it may be necessary to define parameters to standardize the AECs so that studies may be extrapolated for driving basic immunological research and for marketing authorization purposes by regulatory authorities. METHODS: For this task force initiative of the European Academy of Allergy and Clinical Immunology (EAACI), experts from academia and regulatory agencies met with chamber operators to list technical, clinical and regulatory unmet needs as well as the prerequisites for clinical validation. RESULTS: The latter covered the validation process, standardization of challenges and outcomes, intra- and interchamber variability and reproducibility, in addition to comparability with field trials and specifics of paediatric trials and regulatory issues. CONCLUSION: This EAACI Position Paper aims to harmonize current concepts in AECs and to project unmet needs with the intent to enhance progress towards use of these facilities in determining safety and efficacy of new therapeutics in the future.

Feeding a diet devoid of choline to lactating rodents restricts growth and lymphocyte development in offspring.

The nutrient choline is necessary for membrane synthesis and methyl donation, with increased requirements during lactation. The majority of immune development occurs postnatally, but the importance of choline supply for immune development during this critical period is unknown. The objective of this study was to determine the importance of maternal supply of choline during suckling on immune function in their offspring among rodents. At parturition, Sprague-Dawley dams were randomised to either a choline-devoid (ChD; n 7) or choline-sufficient (ChS, 1 g/kg choline; n 10) diet with their offspring euthanised at 3 weeks of age. In a second experiment, offspring were weaned to a ChS diet until 10 weeks of age (ChD-ChS, n 5 and ChS-ChS, n 9). Splenocytes were isolated, and parameters of immune function were measured. The ChD offspring received less choline in breast milk and had lower final body and organ weight compared with ChS offspring (P<0·05), but this effect disappeared by week 10 with choline supplementation from weaning. ChD offspring had a higher proportion of T cells expressing activation markers (CD71 or CD28) and a lower proportion of total B cells (CD45RA+) and responded less to T cell stimulation (lower stimulation index and less IFN-γ production) ex vivo (P<0·05). ChD-ChS offspring had a lower proportion of total and activated CD4+ T cells, and produced less IL-6 after mitogen stimulation compared with cells from ChS-ChS (P<0·05). Our study suggests that choline is required in the suckling diet to facilitate immune development, and choline deprivation during this critical period has lasting effects on T cell function later in life.

Differential expression of hypothalamic, metabolic and inflammatory genes in response to short-term calorie restriction in juvenile obese- and lean-prone JCR rats.

BACKGROUND: Childhood obesity is an important early predictor of adult obesity and associated comorbidities. Common forms of obesity are underpinned by both environmental and genetic factors. However, the rising prevalence of obesity in genetically stable populations strongly suggests that contemporary lifestyle is a premier factor to the disease. In pediatric population, the current treatment/prevention options for obesity are lifestyle interventions such as caloric restriction (CR) and increase physical activity. In obese individuals, CR improves many metabolic parameters in peripheral tissues. Little is known about the effect of CR on the hypothalamus. This study aimed to assess the effect of CR on hypothalamic metabolic gene expression of young obese- and lean-prone animals. METHODS: Male juvenile JCR:LA-cp obese-prone rats were freely fed (Obese-FF) or pair fed (Obese-FR) to lean-prone, free-feeding animals (Lean-FF). A group of lean-prone rats (Lean-FR) were matched for relative average degree of CR to Obese-FR rats. RESULTS: In free-feeding conditions, obese-prone rats consumed more energy than lean-prone rats (P<0.001) and showed greater increases in body weight, fat mass, plasma glucose, insulin and lipids (P<0.01). These metabolic differences were associated with alterations of feeding-related neuropeptides expression in the hypothalamus, as well as pro-inflammatory cytokines and oxidative stress markers. When submitted to the same degree of CR, the two genotypes responded differently; hypothalamic inflammatory and oxidative stress gene expression was improved in Obese-FR, while it was worsened in Lean-FR rats. CONCLUSIONS: We demonstrate in JCR rats that the metabolic and inflammatory response of the brain to CR is genotype dependent.

Methylation demand and homocysteine metabolism: effects of dietary provision of creatine and guanidinoacetate.

S-adenosylmethionine, formed by the adenylation of methionine via S-adenosylmethionine synthase, is the methyl donor in virtually all known biological methylations. These methylation reactions produce a methylated substrate and S-adenosylhomocysteine, which is subsequently metabolized to homocysteine. The methylation of guanidinoacetate to form creatine consumes more methyl groups than all other methylation reactions combined. Therefore, we examined the effects of increased or decreased methyl demand by these physiological substrates on plasma homocysteine by feeding rats guanidinoacetate- or creatine-supplemented diets for 2 wk. Plasma homocysteine was significantly increased (~50%) in rats maintained on guanidinoacetate-supplemented diets, whereas rats maintained on creatine-supplemented diets exhibited a significantly lower (~25%) plasma homocysteine level. Plasma creatine and muscle total creatine were significantly increased in rats fed the creatine-supplemented or guanidinoacetate-supplemented diets. The activity of kidney L-arginine:glycine amidinotransferase, the enzyme catalyzing the synthesis of guanidinoacetate, was significantly decreased in both supplementation groups. To examine the role of the liver in mediating these changes in plasma homocysteine, isolated rat hepatocytes were incubated with methionine in the presence and absence of guanidinoacetate and creatine, and homocysteine export was measured. Homocysteine export was significantly increased in the presence of guanidinoacetate. Creatine, however, was without effect. These results suggest that homocysteine metabolism is sensitive to methylation demand imposed by physiological substrates.

Hyperglucagonemia in rats results in decreased plasma homocysteine and increased flux through the transsulfuration pathway in liver.

An elevated plasma level of homocysteine is a risk factor for the development of cardiovascular disease. The purpose of this study was to investigate the effect of glucagon on homocysteine metabolism in the rat. Male Sprague-Dawley rats were treated with 4 mg/kg/day (3 injections per day) glucagon for 2 days while control rats received vehicle injections. Glucagon treatment resulted in a 30% decrease in total plasma homocysteine and increased hepatic activities of glycine N-methyltransferase, cystathionine beta-synthase, and cystathionine gamma-lyase. Enzyme activities of the remethylation pathway were unaffected. The 90% elevation in activity of cystathionine beta-synthase was accompanied by a 2-fold increase in its mRNA level. Hepatocytes prepared from glucagon-injected rats exported less homocysteine, when incubated with methionine, than did hepatocytes of saline-treated rats. Flux through cystathionine beta-synthase was increased 5-fold in hepatocytes isolated from glucagon-treated rats as determined by production of (14)CO(2) and alpha-[1-(14)C]ketobutyrate from l-[1-(14)C]methionine. Methionine transport was elevated 2-fold in hepatocytes isolated from glucagon-treated rats resulting in increased hepatic methionine levels. Hepatic concentrations of S-adenosylmethionine and S-adenosylhomocysteine, allosteric activators of cystathionine beta-synthase, were also increased following glucagon treatment. These results indicate that glucagon can regulate plasma homocysteine through its effects on the hepatic transsulfuration pathway.

Plasma homocysteine is decreased in the hypothyroid rat.

Recent clinical studies have indicated that plasma homocysteine was significantly increased in hypothyroid patients. Since hyperhomocysteinemia is an independent risk factor for cardiovascular disease we investigated homocysteine metabolism in hypothyroid rats. Hypothyroidism was induced in one study by addition of propylthiouracil (PTU) to the drinking water for 2 weeks. In a second study, thyroidectomized and sham-operated rats were used with thyroid hormone replacement via mini-osmotic pumps. Unlike the human hypothyroid patients, both groups of hypothyroid rats exhibited decreased total plasma homocysteine (30% in PTU rats, 50% in thyroidectomized rats) versus their respective controls. Thyroid replacement normalised homocysteine levels in the thyroidectomized rat. Increased activities of the hepatic trans-sulfuration enzymes were found in both models of hypothyroidism. These results provide a possible explanation for the decreased plasma homocysteine concentrations. The hypothyroid rat cannot be used as a model to study homocysteine metabolism in hypothyroid patients.

Regulation of homocysteine metabolism.

We have used a combination of in vivo and in vitro techniques to measure factors regulating homocysteine metabolism and the plasma concentration of this atherogenic amino acid. The germane findings include: 1. Homocysteine metabolism in rat kidney proceeds predominantly through the transsulfuration pathway, whose enzymes are enriched within the proximal cells of kidney tubules. Furthermore, the rat kidney possesses significant reserve capacity to handle both acute and chronic elevations in plasma homocysteine concentrations. 2. Plasma homocysteine concentrations are lower in diabetic rats. Insulin administration corrects this perturbation. Therefore, insulin and/or one of its counter-regulatory hormones affects homocysteine metabolism, possibly through an increased flux in the hepatic transsulfuration pathway. In support of these data, glucagon administration to rats produced similar results. Further support was provided by studies with isolated rat hepatocytes, from which homocysteine export was reduced when incubated in the presence of glucagon.

Effects of streptozotocin-induced diabetes and of insulin treatment on homocysteine metabolism in the rat.

An elevation in the concentration of total plasma homocysteine is known to be an independent risk factor for the development of vascular disease. Alterations in homocysteine metabolism have also been observed clinically in diabetic patients. Patients with either type 1 or type 2 diabetes who have signs of renal dysfunction tend to exhibit elevated total plasma homocysteine levels, whereas type 1 diabetic patients who have no clinical signs of renal dysfunction have lower than normal plasma homocysteine levels. The purpose of this study was to investigate homocysteine metabolism in a type 1 diabetic animal model and to examine whether insulin plays a role in its regulation. Diabetes was induced by intravenous administration of 100 mg/kg streptozotocin to Sprague-Dawley rats. We observed a 30% reduction in plasma homocysteine in the untreated diabetic rat. This decrease in homocysteine was prevented when diabetic rats received insulin. Transsulfuration and remethylation enzymes were measured in both the liver and the kidney. We observed an increase in the activities of the hepatic transsulfuration enzymes (cystathionine beta-synthase and cystathionine gamma-lyase) in the untreated diabetic rat. Insulin treatment normalized the activities of these enzymes. The renal activities of these enzymes were unchanged. These results suggest that insulin is involved in the regulation of plasma homocysteine concentrations by affecting the hepatic transsulfuration pathway, which is involved in the catabolism of homocysteine.

Improving performance through an organizational culture of employee expertise.

Managers can do many things to improve organizational performance, but the accomplishments of the most skillful employees often are most important. This article makes the point that managers should be aware of employee expertise and its relationship to organizational performance. The article also describes the components of an organizational culture of employee expertise. An organizational culture of employee expertise builds on the learning organization metaphor that has frequently appeared in the management literature. How employees develop expertise to do their jobs is emerging as a critical issue for organizations, and managers will likely play a key role in that process.

Hearing in hospitalised geriatric patients. A descriptive study.

A descriptive audiometric study was performed on 394 patients hospitalised in the geriatric department of our hospital. The mean age of these patients was 82 years. No significant correlation could be found between hearing thresholds and age of the patients. Eighty-six percent of our patients had a socially handicapping hearing loss.

Foot amputations.

When presented with an ischemic limb with forefoot necrosis of varying amounts, the surgeon often categorizes the need for amputation into toe, ray, transmetatarsal, below-knee, and above-knee. Adherence to this type of algorithm ensures a primary above- or below-knee amputation rate of 10% to 20%. The utility of the more uncommon amputations advocated here is an increase of limbs deemed eligible for revascularization and limb salvage. Furthermore, delaying the amputations until the vascular supply is normalized maximizes tissue salvage and minimizes prolonged hospitalizations with multiple amputations performed as a prelude to major amputation. Although these amputations are often looked upon as an afterthought by many vascular surgeons, careful execution here is as important to effective limb salvage as any distal bypass procedure.

Increased limb salvage by the use of unconventional foot amputations.

PURPOSE: Limb salvage in the presence of ischemic foot necrosis requires revascularization followed by debridement or partial foot amputation. Necrosis extending beyond the toes and metatarsal heads may require the use of unconventional types of amputations. METHODS: Over a 15-year period 2105 ischemic limbs were treated with infrainguinal revascularization. In 98 cases, extensive foot necrosis was than managed with amputations, including 59 modified Chopart, 14 Lisfranc, 17 Pirogoff and 8 Syme amputations. Patients were not allowed to bear weight for several days to weeks. RESULTS: Skin flap necrosis in 14 cases was managed successfully by debridement and skin grafting. Ambulation required the use of a "clamshell" prosthesis and foot spacer. The overall limb salvage rate in this group was 84% (82 of 98). In general, the modified Chopart amputation most frequently produced ambulatory limb salvage and is technically easier to perform than a Syme amputation. Patient satisfaction and long-term ambulatory function was highest with the modified Chopart. CONCLUSION: Ischemic foot necrosis extending beyond the limits of conventional transmetatarsal amputation need not be treated with major amputation. This requires the surgeon to be well versed in the use of less common types of partial foot amputations. Acceptable limb salvage and good functional results may be attained by the motivated patient and surgeon with the use of these procedures in the revascularized limb.

Surgical arthrodesis of the neuropathic foot. A salvage procedure.

Reconstructive foot and ankle surgery is a salvage procedure in the deformed neuropathic foot, despite condemnation by some authors. Clinical union and stability was achieved in 91% of the patients, and soft-tissue coverage without skin breakdown was achieved in 100% of the cases. Although one patient had moderate to severe pain in her ankle after operation and was able to do only bed-to-wheelchair transfers, she had good clinical stability, no skin ulceration, and was satisfied overall with the procedure. In addition, a significant component of her pain was believed to be from diabetic neuropathy and not pain that was directly attributable to her reconstructive surgery. All other patients were able to ambulate to some degree. More than half had unlimited use of the affected lower extremity. More than half of the patients had mild or no pain, and all patients had a functional limb. Surgical arthrodesis of the deformed neuropathic foot as a salvage procedure can preserve the limb as a stable functional unit, and create an acceptable alignment of the ankle-foot complex that will promote viability of the overlying soft-tissue structures.

Salvage of the diabetic foot with exposed os calcis.

Fifty consecutive heel ulcers were managed in three groups by debridement, split-thickness skin graft (STSG), bypass procedures, and orthotics. Group I consisted of 24 ulcers in patients with diabetes (DM) and peripheral vascular disease (PVD), 14 patients in Group II with DM only, and 12 patients with PVD only (Group III). Healing occurred in 56.5%, 64.3%, and 83.3%, respectively. An average of 2.2 procedures were performed per patient. Follow-up periods were for a minimum of two years or until amputation. Time for complete healing and the number of amputations performed were similar in all groups. Of the diabetics (combined from Groups I and II), a subgroup of 27% required partial excision of the os calcis to facilitate closure. After saline dressing changes, STSG was accomplished over thin granulation tissue. Forty percent of this subgroup healed, 30% remained open, and 30% were amputated. Aggressive management, soft-tissue coverage, and orthotic use can lead to a functional weight-bearing extremity.

Chopart amputation with percutaneous heel cord lengthening.

When amputation becomes necessary in a patient with peripheral vascular disease, it is important to preserve as much tissue as possible to preserve maximum function. This is especially important because the other extremity may have similar involvement in the future. With appropriate care, an amputation at the Chopart (calcaneocuboid-talonavicular) level can give a good functional result.