Fecal IgE Analyses Reveal a Role for Stratifying Peanut-Allergic Patients.

BACKGROUND AND OBJECTIVE: Peanut allergy (PA) is an IgE-mediated food allergy with variable clinical outcomes. Mild-to-severe symptoms affect various organs and, often, the gastrointestinal tract. The role of intestine-derived IgE antibodies in astrointestinal PA symptoms is poorly understood. This study aimed to examine fecal IgE responses in PA as a novel approach to patient endotyping. METHODS: Feces and serum samples were collected from peanut-allergic and healthy children (n=26) to identify IgE and cytokines using multiplex assays. Shotgun metagenomics DNA sequencing and allergen database comparisons made it possible to identify microbial peptides with homology to known allergens. RESULTS: Compared to controls, fecal IgE signatures showed broad diversity and increased levels for 13 allergens, including food, venom, contact, and respiratory allergens (P<.01-.0001). Overall, fecal IgE patterns were negatively correlated compared to sera IgE patterns in PA patients, with the greatest differences recorded for peanut allergens (P<.0001). For 83% of the allergens recognized by fecal IgE, we found bacterial homologs from PA patients' gut microbiome (eg, thaumatin-like protein Acinetobacter baumannii vs Act d 2, 109/124 aa identical). Compared to controls, PA patients had higher levels of fecal IgA, IL-22, and auto-IgE binding to their own fecal proteins (P<.001). Finally, levels of fecal IgE correlated with abdominal pain scores (P<.0001), suggesting a link between local IgE production and clinical outcomes. CONCLUSION: Fecal IgE release from the intestinal mucosa could be an underlying mechanism of severe abdominal pain through the association between leaky gut epithelia and anticommensal TH2 responses in PA.

Isolated thyrotropin deficiency with thyrotropin-releasing-hormone induced TSH secretion and thyroidal release.

A patient with isolated thyrotropin (TSH) deficiency was studied. Prior to treatment with thyroid hormone, administration of thyrotropin releasing hormone (TRH) produced no increment in serum TSH and a normal increase in plasma prolactin (PRL). In order to explore whether physiologic increases in serum TSH might be occurring below the limits of detectability of TSH by radioimmunoassay, a double isotope technique of assessing thyroidal secretion secondary to release of TSH was employed. The patient was restudied seven months later, after discontinuing thyroid hormone replacement therapy for two months, and on this occasion repeat TRH administration produced small increments in serum TSH. After administration of 125I and 131I-T4 to assess thyroid hormone secretion, TRH was infused continuously for 6 h. Small increases in serum TSH were again observed, along with significant increases in PG125I/PB131I and urinary 125I/131I, reflecting increased thyroidal iodine secretion, although serum T3 and T4 did not change. These studies indicate that: 1) isolated TSH deficiency need not be complete and may be associated with detectable levels of immunoassayable TSH; 2) the TSH released possesses in vivo biological activity; and 3) therapy with thyroid hormone may have facilitated TSH release.

Measurement of serum T4 concentration by high performance liquid chromatography.

Although RIA techniques for the measurement of serum T4 have been extremely useful, this methodology has several disadvantages, including the requirement for the use of radioisotopes, various levels of thyronine cross-reactivity, and the ability to measure only a single iodothyronine in one assay. We have developed a high performance liquid chromatography (HPLC) method for quantitating serum T4 that utilizes the detection of dansyl-T4 compounds and obviates the problems described for RIA techniques. Serum samples were extracted with ethanol and then chloroform, reacted with dansyl chloride, and, after n-heptane extraction, placed directly on column. Utilizing this technique, dansyl-T4 was easily separated and identified. The sensitivity of detection of the dansylated-T4 in serum was 1 microgram/dl, and linearity was observed when increasing standard T4 concentrations were employed. Sensitivity to 10 ng/dl (100 fmol on column) was achieved when T4 was added to buffer. The coefficients of variation were 4.8% and 2.1% for normal and high serum samples, respectively. When 39 random serum samples were anayzed both by HPLC and RIA, there was concordance of these techniques, since the derived correlation coefficient was 0.94. In summary, the present study demonstrates that serum T4 concentrations can be measured by HPLC and that these measurements agree remarkably well with those obtained by RIA. Because of the inherent advantages of HPLC methodology over that of RIA, this technique of measurement of T4 may have wide applicability to the measurement of iodothyronines.

Normal valine disposal in obese subjects with impaired glucose disposal: evidence for selective insulin resistance.

Insulin has major effects on both glucose and branched chain amino acid metabolism. To determine whether the insulin resistance of obesity equally affects both glucose and branched chain amino acid metabolism, we measured the ability of obese and normal subjects to dispose of intravenous bolus dose of glucose (25 g) or L-valine (4 g). Basal plasma glucose levels were the same in the 18 normal and 17 obese (163 plus or minus 8% of ideal body weight) subjects, but basal plasma insulin levels were higher in the obese group (15 plus or minus 2 vs 6 plus or minus 1 microU/ml; p less than 0.001). The obese group had a slower glucose disappearance rate after glucose challenge (0.84 plus or minus 0.06 vs. 1.11 plus or minus 0.07 hr(-1); p less than 0.01) despite having a greater serum insulin response to the glucose load (26 plus or minus 4 vs 11 plus or minus 1 insulin area units; p less than 0.01), confirming insulin resistance. In contrast, disposal of a valine load was the same in normal and obese subjects, as assessed by initial and second phase exponential disappearance rates, metabolic clearance rates of valine, and volumes of distribution. In normal men, disposal rates of glucose and valine after simultaneous administration of both substances were slower than corresponding disposal rates determined when each substance was given alone. We conclude that obese subjects with impaired glucose disposal have normal valine disposal, suggesting that the insulin resistance of obesity can be selective in its effect on different metabolic systems. Glucose and valine also appear to mutually antagonize each other's disposal.

Resolution and quantitation of apolipoproteins A-I and A-II from human high-density lipoprotein by size exclusion high-performance liquid chromatography.

Apolipoproteins A-I and A-II, extracted from human high-density lipoprotein (HDL), were resolved and quantified by size exclusion high-performance liquid chromatography on TSK 125 and TSK 250 analytical columns connected in series without the use of chemical denaturants or detergents in the eluent buffer. The columns were pre-equilibrated with a solution containing 0.1 M sodium phosphate, pH 7.2, 0.2 M sodium chloride at a flow-rate of 1 ml/min. Delipidated HDL (1 mg protein per ml) was resolved into two populations of apolipoprotein (apo) A-I: one representing the apo A-I monomer and the other, a self-associated form with a molecular weight of approximately 120,000 daltons. The column eluates were screened for immunoreactivity to apo A peptides, and the identity of each peak was confirmed by sodium dodecyl sulfate--polyacrylamide gel electrophoresis followed by immunoblot analysis. Apo A-I peptides isolated by high-performance liquid chromatography disrupted unilamellar phospholipid vesicles to form smaller phospholipid particles that eluted on gel filtration columns within the size range of HDL. Thus, a rapid method for the isolation and quantitation of non-denatured apolipoproteins from HDL has been developed using size exclusion high-performance liquid chromatography.

Dermal hypersensitivity reactions to insulin: correlations of three patterns to their histopathology.

Fifteen diabetics with recurrent painful local reactions to insulin were studied. Reactions occurring after intradermal insulin injection were observed in nine patients over 48 hr and biopsies were taken at intervals for microscopic study using the 1 mu Giemsa technique. Insulin-specific IgE and IgG levels were measured on all patients. Five patients had biphasic reactions in which wheal and flare (WFR) were followed by an indurated lesion 4 to 6 hr later. These reactions lasted up to 24 hr and were histopathologically identical to similar "late-phase reactions" seen with ragweed. They were transferable with Prausnitz-Küstner (P-K) testing. Three patients had reactions that developed 8 to 12 hr after injection, peaked around 24 hr, and were not preceded by WFR. These reactions were morphologically delayed hypersensitivity reactions and were not transferred by P-K testing. One patient had a reaction that developed in 4 to 6 hr after injection and peaked by 12 hr. Histologically, this reaction was "Arthus" in type and was not transferred by P-K testing. Specific insulin antibody determinations were not helpful in distinguishing patients with different types of reactivity. These data show that recurrent local reactions to insulin may be of three distinct types: "late-phase reactions" (which are IgE dependent), "Arthus" local vasculitic reactions, or tuberculin-type delayed hypersensitivity reactions. These findings may influence the approach to management of these reactions.

Iodine treatment of iodine-induced thyrotoxicosis.

A 62-year-old female who had received prolonged iodine therapy for asthma presented with severe thyrotoxicosis and severe asthma. Her history, elevated serum thyroxine and triiodothyronine, low 131I uptake, and elevated intrathyroidal iodine content by fluorescent scan were most consistent wiht a diagnosis of iodine-induced thyrotoxicosis (IITT). The clinical course of her thyrotoxicosis was protracted, and in spite of its etiologic role in the precipitaton of thyrotoxicosos, iodine was therapeutically efficacious, although combined treatment with methimazole was required to ultimately restore euthyroidism. Therapy with lithium was also employed but appeared to be only transiently effective and combined no additional decrement in serum T4 than that seen with iodine alone. The case exemplifies the heterogeneity of what is considered "iodine-induced" thyrotoxicosis, the complexities inherent in establishing a diagnosis of IITT, and the use of other rapid acting pharmacologic agents in IITT when beta blockade is contraindicated by asthma.