Why hyperglycemia in sepsis

After obtaining approval by the Henry Ford Health System Institutional Review Board IRB we conducted a prospective observational study in a tertiary care, university affiliated hospital. Patients included in the study presented to the emergency department ED from to and were diagnosed with severe sepsis or septic shock.

Patients were grouped by glucose levels based on recommendations in the literature for glycemic control in critical illness during the study period. Biomarker assays were independently performed by Biosite, Inc. Alere , San Diego, Calif. Assays were performed using immunometric sandwich assays performed with NeutrAvidin-coated well block microtiter plates Pierce Biotechnology, Inc.

Each sample was tested in duplicate. The concentration of biotinylated antibody was predetermined by titration. The primary antibody 10 2 L per well was added to the plates and incubated.

Next, the plates were washed three times with borate-buffered saline containing 0. Ten-microliter aliquots of each sample were added to each plate well, and the plates were incubated. Following this incubation, the plates were washed three times and alkaline phosphatase-conjugated antibody 10 2 L per well was added to each plate well and further incubated.

The concentration of the alkaline phosphatase-conjugated antibody was predetermined to ensure a linear profile in the dynamic range of interest. After additional incubation, the plates were washed nine times with BBS-Tween.

AttoPhos substrate S; Promega Corporation, Madison, Wis , a fluorescence-enhancing substrate previously diluted in AttoPhos buffer S; Promega , was then added to aid in the measurement of the activity of antibody-conjugated alkaline phosphatase bound in each well.

Each well as scanned six times at second intervals, and the rate of fluorescence generation was calculated. Descriptive statistics were used to analyze comorbidities, baseline clinical data, organ dysfunction scores, and mortality.

We also performed a univariate test for means and median distribution using Spearman correlation and analysis of variance ANOVA. A total of patients with severe sepsis and septic shock presenting to the ED were included in the study. The baseline demographics for each BG group are shown in Table 1. The age, race, and gender distributions were similar among the BG groupings. The rates of pre-existing conditions such as hypertension, chronic obstructive pulmonary disease COPD , and congestive heart failure CHF were similar among the BG groups.

The prevalence of DM was 6. In addition, patients in the low BG group were noted to have a higher incidence of pre-existing renal failure Table 2 summarizes baseline vital signs and routine laboratory findings. An increase in lactate levels were noted in the subsequent BG groupings range 5.

To our knowledge this is the first study to establish a significant association between BG levels and MMP-9 levels in patients with severe sepsis or septic shock. Currently, it is well-known that in severe sepsis and septic shock patients, hyperglycemia develops due to a combination of several factors: 1 Insulin clearance is increased leading to a reduction in insulin-mediated glucose uptake; 2 Stress induced elevation in plasma levels of counter-regulatory hormones, such as catecholamines, glucagon, cortisol, and growth hormone.

It has been demonstrated via various significant in vivo and in vitro experimental studies that in the setting of acute or chronic hyperglycemia diabetes mellitus , glucose can induce the increase of pro-inflammatory transcription factors such as NF-kB, AP-1, and EGR The MMP-9 promoter region has been described to contain responsive elements to the aforementioned transcription factors and thus up-regulating MMP-9 transcription and activity [ 5 , 10 , 19 , 20 ].

Our results may suggest a possible up-regulation of MMP-9 in the setting of both acute and chronic hyperglycemia. High MMP-9 levels have been identified in patients in severe sepsis or septic shock.

In addition to the up-regulation of MMP-9 gene transcription secondary to pro-inflammatory factors, MMP-9 levels are increased in the setting of sepsis due to the release of MMP-9 by activated PMNs at the site of infection causing tissue injury and further perpetuating the vicious cycle of inflammation and tissue destruction [ 15 , 21 ]. The significant rise of MMP-9 across the BG groupings in this study cannot solely be attributed to acute stress or chronic hyperglycemia.

The rise in MMP-9 levels in patients with severe sepsis or septic shock has been described to occur very early in the disease process. In an experimental study involving human subjects Albert et al. Nakamura et al. They were able to demonstrate that plasma MMP-9 concentrations were significantly higher in the non-surviving patients with septic shock than in the surviving patients with septic shock [ 24 ].

Since then, various studies have corroborated the increase of MMP-9 levels in patients with severe sepsis or septic shock and their poor outcomes [ 11 , 12 , 25 — 27 ]. In one of our previous study in which we described the natural history of circulatory biomarker activity in the most proximal phases of severe sepsis and septic shock we also noted that MMP-9 levels peaked early 6 h after presentation [ 28 ].

The rise of MMP-9 in septic patients can be due to: 1 genetic up-regulation and 2 release by activated neutrophils. IL-8, a member of the CXC chemokine family has been reported to be elevated and play a role in the pathophysiology of sepsis because of its ability to attract, activate and degranulate neutrophils [ 29 , 30 ]. In addition, the expression of ICAM-1 a cell surface glycoprotein is increased on the cell surface of neutrophils allowing it to attach and migrate through the endothelium during sepsis [ 31 , 32 ].

A possibility behind the inverse relationship noted between MMP-9 and IL-8 is that MMP-9 is known to cleave a specific site of IL-8 molecular structure rendering a variant IL-8 molecule which is 10 to 30 fold more potent in neutrophil activation [ 33 , 34 ]. We theorize that our assay provided us with the measurement of the native IL-8 molecule and not the variant IL-8 form which we conceptually believe to be elevated. The aforementioned observation describes the relationship between IL-8 and MMP-9 from a proteonomic and metabolomic point of view.

To explain the relationship between ICAM-1 and MMP-9 is more challenging as there is a scarcity of studies describing this relationship. In the field of oncology MMP-9 has been shown to proteolytically cleave the extracellular domain of ICAM-1 leading to its release from the cell surface [ 35 ]. Our observation contrast this finding.

A possible explanation to the rise of MMP-9 and decrease of ICAM-1 levels may be due to the direct effect of insulin on such biomarkers. Aljada et al. On the other hand, Fischoeder et al. The early pathogenic link between elevated MMP-9 levels and elevated BG levels make it plausible to consider that early inhibition or removal of this biomarker may have therapeutic potential. Antibiotics, such as the tetracyclines inhibit MMPs, not only by chelating the zinc and calcium ions but also by affecting the induction of the MMP genes.

The therapeutic administration of tetracyclines over prolonged periods of time may cause undesirable side effects, such as gastrointestinal disturbances and emergence of antibiotic resistance.

A recent pilot study reported a low side effect profile of doxycycline tetracycline based on sub-antimicrobial concentrations dosing mg intravenous IV daily, followed by 50 mg IV once daily for 2 days. However, their results showed no effect on MMP-9 levels [ 38 ]. There were no significant differences in demographic characteristics or disease characteristics between patients with and without stress hyperglycemia. This unexpected result might be explained by improved cellular uptake of glucose when present at moderate levels and by their patient population which did not include patients with major trauma, or neurosurgical patients, or cardiac surgery patients.

All studies investigating the relationship between hyperglycemia and outcomes in patients with sepsis need to consider patient characteristics and co-morbidity at presentation, adverse events during the hospitalization, and, in particular, the presence or absence of diabetes prior to hospitalization 1.

In addition, the duration of diabetes and the quality of outpatient management HbA1c levels prior to acute illness might be important factors in these patients. This information is likely available in larger prospective databases and allows for adjustment in statistical models which should reduce the possibility of spurious conclusions about glucose effects. However, we should not assume that the loss of statistical significance in a multivariable model indicates that hyperglycemia is unimportant in complex patients.

Multiple factors influence glucose levels in patients with sepsis. Sepsis stimulates gluconeogenesis using recycled pyruvate and lactate, using glycerol from mobilized fatty acids, and using glucogenic amino acids mobilized by proteolysis 5 - 7. Glycogenolysis can also increase glucose levels, and relative insulin resistance reduces glucose uptake in some organs and prevents insulin modulation of gluconeogenesis in the liver.

However, glucose uptake in some tissues is increased through insulin independent glucose transporter, such as GLUT 1, 2, and 3. Multiple hormones associated with stress stimulate gluconeogenesis, and these include glucagon, epinephrine, and cortisol. Lactate production requires glycolysis and the production of pyruvate and then lactate. Increased lactate levels can occur when large fluxes of glucose exceed the capacity of mitochondria to take up pyruvate and when mitochondrial dysfunction impairs the uptake of pyruvate and its metabolism in the citric acid cycle 8.

Consequently, the number and complexity of factors which influence glucose levels probably exceed simple interpretation. However, glucose levels reflect, in part, the degree of stress associated with an acute illness and indicate abnormal metabolic responses during acute illness. Lactate levels are potentially more easily interpreted since they should reflect pyruvate formation and mitochondrial function.

However, lactate is also recycled into glucose in the liver and kidney, and lactate levels necessarily reflect formation and metabolism. Classifying patients according to glucose levels and lactate levels or according to glucose-lactate ratios might help identify patients with increased mortality in sepsis.

In macrophages and dendritic cells stimulated by LPS, a variety of metabolic changes occur, all of which lead to the inflammatory response. Inducible nitric oxide synthase iNOS increases the levels of nitric oxide NO , which in turn nitrosylates iron-sulfur proteins in the mitochondrial electron transport chain, leading to inhibition of oxidative phosphorylation. A large number of studies have also shown that one of three pyruvate kinases, PKM2, controls the level of oxidative phosphorylation in cells and that inhibiting PKM2 activity in macrophages reduces LPS-induced inflammation [reviewed in 31 ].

A component of traditional Chinese medicine, shikonin, is an inhibitor of PKM2 and protects mice from LPS-induced shock and sepsis 32 , Recent developments in understanding the Warburg Effect in critical illness suggest other, newer possibilities.

A quantitative modeling of the glycolytic pathway using metabolomics and known kinetic parameters for the very well-studied enzymatic steps that control this pathway has been carried out Variations in fructose 1,6-bisphosphate FBP led these authors to look for different rate-limiting steps as a function of FBP. They have deduced, and experimentally tested, the idea that during aerobic glycolysis the enzyme glyceraldehyde 3-phosphate dehydrogenase GAPDH , which oxidizes glyceraldehyde 3-phosphate GA3P to yield NADH and 1,3-bisphospho-glycerate 1,3-BPG , becomes rate limiting for glycolysis it is not rate limiting when FBP concentrations are low.

Although the context for this work and the work to be described below was the Warburg Effect in cancer cells, all available evidence suggests that the aerobic glycolysis seen in sepsis should follow the same metabolic rules. DPP4 activity has been reported on the cell surface of immune and endothelial cells 36 , as well as in blood serum as a soluble form The main function of DPP4 is thought to be the modification of biologically active peptides, cytokines, and other cell-surface proteins for the purpose of regulating the immune response and cell differentiation Of specific importance to the hyperglycemia encountered in critically ill patients is the action of DPP4 on incretins, specifically on glucagon-like peptide 1 GLP-1 The same applies to inflammation.

Figure 2. Importance of DPP4 in hyperglycemia. Dipeptidyl peptidase IV DPP4 , or CD26, is a cell surface and soluble peptidase that can cleave the first two N-terminal amino acids from specific proteins and peptides.

DPP4 is up-regulated in immune and endothelial cells as a result of inflammation seen in critically ill patients. GLP-1 is important in the storage and regulation of blood glucose by promoting insulin and limiting glucagon release. Inhibitors of DPP4 could be important in glucose control in critically ill patients with hyperglycemia. The receptors for incretins are widely expressed in endothelial cells, vascular smooth muscle cells, monocytes, macrophages, and lymphocytes suggesting that GLP-1 could have direct effects on inflammation Among the various effects of GLP-1, an important one is reduction of inflammation by reducing the levels of inflammatory mediators, decreased monocyte adhesion, reduction in the proliferation of macrophages and macrophage foam cell formation, and smooth muscle cell proliferation 38 , To that effect, hyperglycemia is at least partially the result of increased DPP4 activity in these conditions.

Stress hyperglycemia in critical illness may have some beneficial effects by supplying much-needed glucose to affected and hypo perfused tissue. However, this hyperglycemia has been associated with increased mortality and morbidity in surgical and medical patients.

It appears that the combination of hyperglycemia and hyperlactatemia deserves specific attention in view of the clinical and molecular mechanisms described here. DB-O and EB made contributions to conception and design, participated in drafting the article, and revised it critically for important intellectual content.

The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Multiple organ dysfunction syndrome in humans and animals. J Vet Intern Med. Stress hyperglycaemia and increased risk of death after myocardial infarction in patients with and without diabetes: a systematic overview. Impact of admission hyperglycemia on hospital mortality in various intensive care unit populations. Crit Care Med. Hyperglycaemia in critically ill patients: the immune system's sweet tooth. Crit Care. Glycemic control in the ICU.

Adv Surg. Review: traumatic brain injury and hyperglycemia, a potentially modifiable risk factor. Early hyperglycemia predicts multiple organ failure and mortality but not infection. J Trauma. Phagocyte-derived catecholamines enhance acute inflammatory injury. It is likely that severe stress hyperglycemia may occur more frequently in patients with underlying impaired glucose tolerance [ 43 ].

The results of this single-center, investigator-initiated and unblinded study have yet to be reproduced. This study has a number of serious limitations with concern regarding the biological plausibility of the findings [ 8 , 44 ].

Following the above study, tight glycemic control became rapidly adopted as the standard of care in ICUs throughout the world. Tight glycemic control then spread outside the ICU to the step-down unit, regular floor and even operating room.

Without any credible evidence that intensive glycemic control improves the outcome of hospitalized patients, this has become a world-wide preoccupation and 'compliance' with glycemic control is used as a marker of the quality of care provided.

Indeed, as recently as , the Endocrine Society Clinical Practice Guideline on the management of hyperglycemia in hospitalized patients stated that 'observational and randomized controlled studies indicate that improvement in glycemic control results in lower rates of hospital complications' and they provide strong recommendations for glycemic control [ 45 ].

We believe the evidence demonstrates that these assertions and recommendations are without a scientific basis and may be potentially detrimental to patients. Although an association between the degree of hyperglycemia and poor clinical outcomes exist in the hospitalized patient, there are few data demonstrating causation. Randomized, controlled studies do not support intensive insulin therapy. Furthermore, improving care through the acute treatment of mild or moderate hyperglycemia in the acutely ill hospitalized patient lacks biologic plausibility.

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