Baseline insulin/glucose ratio as a marker for the clinical course of hyperglycemic critically ill children treated with insulin

Objective

The objective of this study was to investigate the relations of baseline insulin/glucose ratio to the clinical course of critically ill children. Such information will provide insight into the pathophysiologic mechanisms leading to hyperglycemia and will optimize preventive and therapeutic measures for hyperglycemia in critically ill children.

Methods

Sixty-four consecutively admitted critically ill children with hyperglycemia, defined as a blood glucose level higher than 8 mmol/L (>145 mg/dL) and treated with insulin according to a glucose-control protocol, were included. Demographic data and clinical and laboratory parameters were collected. Insulin sensitivity was investigated by calculating the ratio of insulin to the blood glucose level just before the start of insulin administration. Results are expressed as median (range).

Results

Sixty-four children (24 girls) 7.0 y of age (0.3-16.9 y) with various diagnoses were included. A hyperinsulinemic response, indicated by an increased insulin/glucose ratio (>18 pmol/mmol), was seen in 55% of children. The durations of insulin therapy, mechanical ventilation, and pediatric intensive care unit length of stay in children with a hyperinsulinemic response were longer than in children with a hypoinsulinemic response.

Conclusion

Hyper- and hypoinsulinemic responses play a role in the occurrence of hyperglycemia in critically ill children. Each is associated with a particular clinical course after the initiation of insulin therapy. It would be worthwhile to further investigate if the insulinemic response to hyperglycemia, determined by the insulin/glucose ratio in combination with the type of organ dysfunction, could be used in clinical practice to determine the need for insulin therapy.     

111In-exendin Uptake in the Pancreas Correlates With the β-Cell Mass and Not With the α-Cell Mass

Targeting of the GLP-1 receptor with ¹¹¹In-labeled exendin is an attractive approach to determine the β-cell mass (BCM). Preclinical studies as well as a proof-of-concept study in type 1 diabetic patients and healthy subjects showed a direct correlation between BCM and radiotracer uptake. Despite these promising initial results, the influence of α-cells on the uptake of the radiotracer remains a matter of debate. In this study, we determined the correlation between pancreatic tracer uptake and β- and α-cell mass in a rat model for β-cell loss. The uptake of ¹¹¹In-exendin (% ID/g) showed a strong positive linear correlation with the BCM (Pearson r = 0.82). The fraction of glucagon-positive cells in the total endocrine mass was increased after alloxan treatment (26% ± 4%, 43% ± 8%, and 69% ± 21% for 0, 45, and 60 mg/kg alloxan, respectively). The uptake of ¹¹¹In-exendin showed a negative linear correlation with the α-cell fraction (Pearson r = −0.76). These data clearly indicate toward specificity of ¹¹¹In-exendin for β-cells and that the influence of the α-cells on ¹¹¹In-exendin uptake is negligible.     

Human IL-32 expression protects mice against a hypervirulent strain of Mycobacterium tuberculosis

Silencing of interleukin-32 (IL-32) in a differentiated human promonocytic cell line impairs killing of Mycobacterium tuberculosis (MTB) but the role of IL-32 in vivo against MTB remains unknown. To study the effects of IL-32 in vivo, a transgenic mouse was generated in which the human IL-32γ gene is expressed using the surfactant protein C promoter (SPC-IL-32γTg). Wild-type and SPC-IL-32γTg mice were infected with a low-dose aerosol of a hypervirulent strain of MTB (W-Beijing HN878). At 30 and 60 d after infection, the transgenic mice had 66% and 85% fewer MTB in the lungs and 49% and 68% fewer MTB in the spleens, respectively; the transgenic mice also exhibited greater survival. Increased numbers of host-protective innate and adaptive immune cells were present in SPC-IL-32γTg mice, including tumor necrosis factor-alpha (TNFα) positive lung macrophages and dendritic cells, and IFN-gamma (IFNγ) and TNFα positive CD4⁺ and CD8⁺ T cells in the lungs and mediastinal lymph nodes. Alveolar macrophages from transgenic mice infected with MTB ex vivo had reduced bacterial burden and increased colocalization of green fluorescent protein-labeled MTB with lysosomes. Furthermore, mouse macrophages made to express IL-32γ but not the splice variant IL-32β were better able to limit MTB growth than macrophages capable of producing both. The lungs of patients with tuberculosis showed increased IL-32 expression, particularly in macrophages of granulomas and airway epithelial cells but also B cells and T cells. We conclude that IL-32γ enhances host immunity to MTB.Tuberculosis (TB) is a leading cause of morbidity and mortality in the world with nearly 9 million new cases and over 1 million deaths per year. Mycobacterium tuberculosis (MTB) strains possessing high levels of drug resistance such as multidrug resistant or extensively drug resistant MTB threaten to make TB into an incurable disease (1). Whereas development of new classes of effective anti-TB antibiotics is important to treat existing cases (2), history teaches that MTB will eventually develop resistance. Thus, it is important to continue to seek previously undiscovered immune responses to MTB in hopes that new approaches may be applied to treat human TB.First described in 2005, interleukin-32 (IL-32) is a pleiotropic cytokine that induces proinflammatory cytokines such as tumor necrosis factor-alpha (TNFα) and IL-1β (3). IL-32 protects against infections with HIV (4), influenza (5), and Mycobacterium avium (6). This protective effect is likely due to the proinflammatory nature of IL-32, which is also implicated in the pathogenesis of a number of inflammatory disorders (3, 7, 8). We previously demonstrated that infection of human macrophages or peripheral blood mononuclear cells (PBMCs) with MTB H37Rv induced IL-32 (9, 10). Silencing endogenous IL-32 by siRNA in THP-1 macrophages, differentiated from a human promonocytic cell line, increased the intracellular burden of MTB, indicating that IL-32 plays a host-protective role (9). However, the role of IL-32 in the response to TB in vivo remains unknown.IL-32 is composed of six isoforms (α, β, γ, δ, ε, and ζ) due to alternatively spliced mRNA variants (3). IL-32γ is biologically the most active, likely due to the lack of exonic deletions (11). Because a full-length mouse homolog of IL-32 is not present in the databases, studying a role for endogenous IL-32 in mice is not possible. Because murine macrophages do respond to IL-32 as measured by TNFα production (3), we developed a transgenic (Tg) mouse strain in which human IL-32γ is expressed in type II alveolar epithelial cells under the control of the surfactant protein C (SPC) gene promoter (SPC-IL-32γTg). Wild-type (WT) C57BL/6 and SPC-IL-32γTg mice were infected with a low-dose aerosol of MTB W-Beijing HN878, a hypervirulent MTB strain isolated from a patient with TB (12). We hypothesized that expression of human IL-32 in the lungs of mice would protect against MTB infection. If borne out, IL-32 may represent a target for previously unidentified immunotherapy to treat TB.     

The first step of peptide selection in antigen presentation by MHC class I molecules

MHC class I molecules present a variable but limited repertoire of antigenic peptides for T-cell recognition. Understanding how peptide selection is achieved requires mechanistic insights into the interactions between the MHC I and candidate peptides. We find that, at first encounter, MHC I H-2K^b considers a wide range of peptides, including those with expanded N termini and unfitting anchor residues. Discrimination occurs in the second step, when noncanonical peptides dissociate with faster exchange rates. This second step exhibits remarkable temperature sensitivity, as illustrated by numerous noncanonical peptides presented by H-2K^b in cells cultured at 26 °C relative to 37 °C. Crystallographic analyses of H-2K^b–peptide complexes suggest that a conformational adaptation of H-2K^b drives the decisive step in peptide selection. We propose that MHC class I molecules consider initially a large peptide pool, subsequently refined by a temperature-sensitive induced-fit mechanism to retain the canonical peptide repertoire.MHC class I molecules present a wide array of peptides to cytotoxic T lymphocytes, allowing the immune system to scan for intracellular pathogens and mutated proteins. These peptides are not chosen at random, but rather are selected for their ability to bind to the polymorphic MHC class I peptide-binding groove. Antigenic peptide precursors are produced by the proteasome and further trimmed by cytosolic aminopeptidases. They are translocated into the endoplasmic reticulum (ER) lumen by the peptide transporter TAP that has broad peptide specificity. Peptides can be further trimmed in the ER lumen by ER aminopeptidases before selection by a defined MHC class I allele (reviewed in ref. 1). Only few peptides from a broad peptidome are presented by a given MHC class I allele. How a defined MHC I allele selects the correct peptides for presentation out of a large and diverse peptide pool is unclear.In principle, MHC class I molecules could consider only “optimal” peptides and ignore the remainder of the TAP-translocated peptidome. Alternatively, MHC class I could bind all available peptides followed by a selection step for the optimal candidates for presentation. To discriminate between these scenarios, we studied peptide association (on-rates) and dissociation (off-rates) from the mouse MHC class I molecule H-2K^b. To characterize the biophysical basis of discrimination between candidate peptides we complemented the kinetic data with five crystal structures of H-2K^b with peptide variants. A comprehensive analysis of our in vitro data indicated that discrimination against suboptimal peptides by MHC class I may exhibit a strong temperature dependency, as illustrated by the H-2K^b peptidome from cells cultured at 26 °C versus 37 °C. We thus arrive at a two-step model for peptide selection by MHC class I molecules, which explains how not so “empty MHC class I molecules come out in the cold” (2).     

Prospective study of the effect of gastrectomy with and without bile reflux on serum pepsinogens

In order to prospectively determine the effect of gastrectomy with or without enterogastric reflux on serum pepsinogen concentrations, serum pepsinogen A, serum pepsinogen C and the pepsinogen A:C ratio were measured before, and 10 days and 6, 15 and 24 months after 2/3-3/4 distal gastrectomy in peptic ulcer patients with primary Roux-en-Y diversion (n = 11) or Billroth II reconstruction (n = 11). Gastrectomy induced early decreases in serum pepsinogen A from 100 +/- 12 to 66 +/- 7 micrograms/l (p less than 0.05) and from 111 +/- 11 to 82 +/- 20 micrograms/l (p = 0.05), serum pepsinogen C from 49 +/- 6 to 29 +/- 5 micrograms/l (p less than 0.05) and from 54 +/- 9 to 40 +/- 11 micrograms/l (p = 0.10) in patients with Roux-en-Y and Billroth II gastrectomy, respectively, but did not influence the pepsinogen A:C ratios. Serum pepsinogen A and the pepsinogen A:C ratio continued to decrease 6 months after surgery but no further significant reductions were observed 15 and 24 months postoperatively. Serum pepsinogen C concentrations did not significantly change during postoperative follow-up. Analyses of variances of serum pepsinogen A and the pepsinogen A:C ratio showed that the type of operation had no significant effect on the postoperative course. It is concluded that gastrectomy leads to early decreases in serum pepsinogen A and pepsinogen C levels followed by progressive further reductions of serum pepsinogen A and the pepsinogen A:C ratio for a period of 6 months postoperatively. These postoperative changes of serum pepsinogens are not due to increased enterogastric biliary reflux.     

Cytotoxic T-lymphocyte precursor frequency (CTLp-f) as a tool for distinguishing permissible from non-permissible class I mismatches in T-cell-depleted allogeneic bone marrow transplantation

Matching for HLA has been the gold standard in bone marrow donor selection. But, with the ever increasing number of identified HLA alleles, it is becoming more difficult to find a fully HLA-identical donor other than a sibling. Retrospective analysis revealed that HLA mismatches do not necessarily give rise to acute graft-versus-host-disease (GVHD). However, we have no means of defining these 'permissible' mismatches before bone marrow transplantation (BMT). Thus, we set out to establish whether functional matching by means of helper and cytotoxic T-lymphocyte precursor frequency analysis (HTLp-f and CTLp-f respectively) can be applied to this end. Fifty-five recipient-donor pairs other than HLA-identical siblings, the recipient of which received a T-cell-depleted graft, were analysed by high-resolution HLA typing and/or HTLp-f/CTLp-f analysis. The predictive value of the CTLp-f assay for development of acute GVHD was confirmed. More importantly, our data indicate that the CTLp-f assay was able to discriminate permissible from non-permissible HLA-A, -B or -Cw mismatches, but not for DRB/DQB mismatches. The absolute number of alloreactive CTLs present in the graft correlated with the risk of acute GVHD. Although HTLp-f and CTLp-f together had a high negative predictive value, HTLp-f outcome by itself was not correlated with acute GVHD. As we have no evidence yet that HTLp-f or CTLp-f can identify permissible DRB/DQB mismatches, high-resolution matching for these antigens remains the best option. The combination of high-resolution DRB/DQB typing and the CTLp-f assay would enable the accurate prediction of the risk of acute GVHD while extending the pool of potential donors. Furthermore, it would enable adjustment of the number of T- cells in the graft accordingly to improve clinical outcome.     

Effect of partial gastrectomy on serum anti-Helicobacter pylori immunoglobulins in peptic ulcer patients

Since biliary enterogastric reflux is suggested to eradicate gastric infection withHelicobacter pylori (HP), we have investigated in a prospective randomized study the effect of partial gastrectomy with either Billroth II or Roux-en-Y anastomosis on infection with HP as assessed by the titers of IgG and IgA antibodies against HP in serum. These antibodies were measured by ELISA in serum of 22 patients before and at 10 days and 6, 15, and 24 months after either Billroth II (N=11) or Roux-en-Y (N=11) gastrectomy for peptic ulcer. All patients had HP demonstrated in their preoperative endoscopic gastric biopsies. The preoperative serum IgA antibodies against HP (anti-HP IgA) were increased in 20 of the 22 patients (range 0.21–1.69) while the IgG antibodies (anti-HP IgG) were increased in all 22 patients (range 0.38–1.31). Four of the Billroth II patients had clearance of HP from gastric biopsies accompanied by rapid and pronounced decrease of anti-HP IgA. In contrast, the patients with Roux-en-Y gastrectomy and the Billroth II patients with persistent HP infection had no change in anti-HP IgA after surgery. Anti-HP IgG showed variable results in the four patients without gastric HP infection and was not affected by gastrectomy in the patients with persistent HP infection. We concluded that serum anti-HP IgA, but not anti-HP IgG, is helpful in identifying those patients in whom HP is no longer demonstrable after Billroth II gastrectomy. Gastrectomy with Roux-en-Y anastomosis had no effect on gastric HP infection.     

The functional variant (Asp299gly) of toll-like receptor 4 (TLR4) influences TLR4-mediated cytokine production in rheumatoid arthritis

OBJECTIVE: To investigate functional consequences of the Toll-like receptor 4 (TLR4) variant (Asp299Gly) in rheumatoid arthritis (RA). METHODS: Peripheral blood mononuclear cells from 28 patients with RA carrying or not carrying the TLR4 variant were incubated with lipopolysaccharide (LPS) and heat shock protein B8 (HSPB8). Concentrations of interleukin 6 (IL-6), tumor necrosis factor-alpha(TNF-alpha), and IL-10 were determined along with TLR4 and CD14 expression. RESULTS: TLR4 expression was similar in patients carrying or not carrying the variant. In contrast, both LPS and HSPB8 resulted in significantly lower secretion of IL-6, TNF-alpha, and IL-10 in those who carried the variant, whereas the frequency of CD14+ cells was higher in these individuals. CONCLUSION: TLR4 variant clearly reduces its potency to mediate signaling. Correction for CD14+ cells is necessary in comparable experiments.     

Urinary albumin excretion rate and puberty in non-diabetic children and adolescents

Slightly elevated urinary albumin excretion rate (microalbuminuria) is a marker of early diabetic nephropathy, but it is unclear if the established definition of microalbuminuria (20–200 μg/min) is correct for children and adolescents. We investigated the albumin excretion rate, albumin/creatinine ratio and urinary albumin concentration in 150 healthy schoolchildren and adolescents to (a) obtain a reference value for albumin excretion rate, (b) relate albumin excretion to pubertal stages and (c) evaluate albumin/creatinine ratio and morning albumin concentration as screening methods for elevatcd albumin excretion rate. Albumin concentration was measured by immunoturbidimetry in timed overnight urine samples. The albumin excretion showed a skewed distribution (geometric mean 3.2 μg/min, 95 percentile 15.1 μg/min). In girls, a peak in the albumin excretion rate was found at the pubertal stage 4 (Tanner) and in boys at stage 5. Albumin/creatinine ratio of 2.5 mg/mmol as a scrccning level for elevatcd albumin cxcrction (15 μg/min) showed a high positivc (0.88) and negative (0.99) predictive value.