Patterns and Effects of Admission Hyperglycemia And In�ammatory Response in Trauma Patients: A Prospective Study

Background: Hyperglycemia following trauma could be a response to stress. The constellation of the initial hyperglycemia, proin�ammatory cytokines and severity of injury among trauma patients is understudied. We aimed to evaluate the patterns and effects of on-admission hyperglycemia and in�ammatory response in a level 1 trauma center admissions. Methods: A prospective, observational study was conducted for adult trauma patients who were admitted and tested for on-admission blood glucose, hemoglobin A1c, interleukin (IL)-6 ,Il-18 and hs-CRP. Patients were categorized into 4 groups (non-diabetic normoglycemic, diabetic normoglycemic, diabetic hyperglycemic (DH) and stress-induced hyperglycemic (SIH)). The in�ammatory markers were measured on 3 time points (admission, 24 h, and 48 h). Pearson’s correlation test and logistic regression analysis were performed. We hypothesized that higher initial readings of blood glucose and cytokines are associated with severe injuries and worse in-hospital outcomes in trauma patients. Results: During the study period, 250 adult trauma patients were enrolled. Almost 13% of patients presented with hyperglycemia (SIH&DH); of whom 50% had SIH. Compared to the other 3 groups; SIH patients were younger, had signi�cantly higher ISS, higher IL-6 readings, prolonged hospital length of stay and higher mortality. The SIH group had lower Revised Trauma Score (p=0.005), lower Trauma Injury Severity Score (p=0.01) and lower GCS (p=0.001). IL-18 and hs-CRP were comparable among the study groups. Compared to the normoglycemia groups, patients with hyperglycemia had elevated high-sensitive troponin T (p=0.001) and required more blood transfusion (p=0.03). Patients with hyperglycemia had 3-times higher in-hospital mortality than the normoglycemia groups (p=0.02). A signi�cant correlation was identi�ed between initial blood glucose and serum lactate, IL-6, ISS and hospital length of stay. IL-6 correlated well with ISS (r=0.40, p=0.001). On-admission blood glucose had age-sex-GCS adjusted odd ratio 1.20(95% CI 1.06-1.33, p=0.003) for severe injury (ISS ≥ 16). Conclusions: On-admission hyperglycemia is associated with a signi�cant severer injury than normoglycemia patients. Initial blood glucose correlates with serum IL-6 which indicates a potential role of the systemic in�ammatory response in the disease pathogenesis among the injured patients. On-admission glucose level could be a useful marker of injury severity, triage and risk assessment in trauma patients


Introduction
Hyperglycemia following trauma is a hypermetabolic response to stress which can be associated with a signi cant morbidity and mortality [1].Some investigators have suggested that admission glucose levels re ect the physiological stress reaction to injury and or severe bleeding and thus could be used as a potential predictor of outcomes [2][3][4].
The trauma-related metabolic surge and associated stress-induced hyperglycemia (SIH) were found to correlate with serum cortisol and catecholamine levels [5].It has been suggested that insulin production is suppressed in trauma patients due to systemic stress response secondary to elevated serum glucagon, catecholamine and cytokines [6][7][8].
Assessment of glycosylated hemoglobin (HbA1c) level is considered as a useful tool to distinguish occult (not known before) diabetes mellitus (DM) from SIH [9].Kopelman et al., [10] reported that 18% of trauma patients presented with hyperglycemia, of whom 22% had occult DM which represented 4% of the total screened trauma patients.The possible mechanism of the adverse effects of hyperglycemia may differ in patients with SIH as compared to DH. SIH is an acute process, initiated by the release of stress hormones and cytokines, while DH is a chronic process associated with subsequent microvascular changes [11].
Moreover, there is a relationship between hyperglycemia and altered cellular metabolism in critically ill patients that results in insulin resistance and release of systemic in ammatory mediators [6].Earlier studies suggested that proin ammatory cytokines such as interleukin (IL)-6 and IL-18 are involved in glucose metabolism and insulin action; therefore, hypercytokinemia may have a potential role in increased glucose levels [6,12,13].This indicates that immunoneuroendocrine alterations might be involved in the pathophysiology of SIH during acute illnesses [14].As most observations on the association of cytokines with hyperglycemia are based on experimental studies, there is a need to explore such relationship with respect to the clinical outcome in trauma patients.Herein, this prospective study aims to evaluate the patterns and effects of on-admission hyperglycemia, pro-in ammatory cytokine and severity of injury in trauma patients.
We hypothesized that higher initial readings of blood glucose and cytokines are associated with severe injuries and worse in-hospital outcomes in trauma patients.

Materials And Methods
A prospective observational study was conducted for trauma patients who were admitted to level 1 trauma center at Hamad General Hospital (HGH) between October 2016 and July 2019.Inclusion criteria were adult (≥ 18 years) trauma patients (both genders) presented to the emergency department who were investigated for random blood glucose level and HbA1C within 5 hours of hospital admission.Exclusion criteria included patients declined to participate or in whom random glucose level and HbA1C were not measured on time, vulnerable populations (children, pregnant women) and alcohol consumers.All trauma patients underwent thorough clinical assessment and resuscitation according to the Advanced Trauma Life Support (ATLS) guidelines.Potential subjects were enrolled after obtaining written informed consent either by subject or his/her next-of-kin or deferred consent for blood investigations and use of data with secured con dentiality of personal information.The Institutional Review Board (IRB# 14471/14) of Hamad Medical Corporation has approved this study.This study is registered at the ClinicalTrials.gov(Identi er: NCT02999386).Sample size sample size was calculated considering the prevalence of SIH in trauma patients that ranges from 10-17% for all trauma admissions [15,16] with a precision of estimate (margin of error) of 5% and a 95% level of con dence.Using the single proportion equation for dichotomous variables in the nMaster 2.0 sample size software package, the required sample size was 250 consecutive trauma patients.

Study variables
Data included patients' demographics, (age, gender, nationality), mechanism of injury, initial vitals (heart rate, respiratory rate, systolic blood pressure, diastolic blood pressure and shock index), routine laboratory ndings such as hemoglobin, base de cit, serum lactate, and blood glucose levels were recorded at the baseline, after 24 h and 48 h.Other investigations included white blood cell count (WBC), platelet count, high-sensitive troponin T (hs-TnT), HbA1C, and blood ethanol levels.We have collected information about history of DM, anti-diabetic medications, associated injuries, injury severity score (ISS), Glasgow Coma Score (GCS), Revised Trauma Score (RTS), Trauma Injury Severity Score (TRISS), surgical intervention, blood transfusion, hospital length of stay, intensive care unit stay, in-hospital complications (pneumonia, acute respiratory distress syndrome, renal failure, and sepsis) and hospital mortality.Shock index (SI) was de ned as initial heart rate divided by the initial systolic blood pressure.
The main exposure was hyperglycemia, de ned as random serum glucose 200 mg/dL (11.1 mmol/l) or more.This cutoff level of glucose was previously used to de ne hyperglycemia by earlier studies in trauma patients [15,17,18].DM was determined by patient history and/or admission HbA1c ≥ 6.5%.This level of HbA1c is based on current recommendations for the diagnosis of DM from the American Diabetes Association [19].Stress-induced hyperglycemia (SIH) was de ned as hyperglycemia on admission in patients with normal HbA1c in the index admission [20].
Serum levels of CRP, IL-6 and IL-18 On admission, 5 ml blood specimen was drawn in red vacutainer for proin ammatory cytokines and hs-CRP assay after recruitment in the study followed by 5 ml blood sampling repeated after 24 and 48 h post admission.The blood specimens were allowed to clot for half an hour and then centrifuged at 4500 rpm for 5 min to separate serum which was aliquoted in duplicates and stored at -80 °C in cryovials until analysis.
Brie y, to measure the concentrations of serum hs-CRP, IL-6 and IL-18, an enzyme-linked immunosorbent assay (ELISA) was performed using commercially available kits for cytokine detection (R&D Systems. The preparation of all reagents, the working standards and protocol were followed according to the manufacturer's instructions.The absorbance was read using ELISA reader (TECAN) at 450 nm and 570 nm dual lters.The minimum detectable dose for hs-CRP was 0.005 ng/ml, for IL-6 was 0.7 pg/ml and for IL-18 was 1.25 pg/ml.All the samples were thawed only once and assayed in duplicate.In addition, we recruited 50 age-sex matched healthy volunteers as controls to identify the reference values of in ammatory markers in our community; the mean serum levels of IL-6, IL-18 and hs-CRP were 1.95, 22.9 and 3.0, respectively.

Statistical analysis
Data were reported as proportion, mean (± standard deviation), con dence intervals, median, and range, when applicable.The levels of blood glucose, serum lactate, base excess, IL-6, IL-18, hs-CRP, complications and outcome were compared based on ISS (mild, moderate & severe).Trauma patients were rst compared as 2 groups based on the initial glucose levels (normoglycemic vs hyperglycemic).
Then trauma patients were divided into 4 subgroups based on the initial glucose and HbA1c values (nondiabetic normoglycemic, stress-induced hyperglycemia (SIH), diabetic normoglycemic, diabetic hyperglycemic (DH)) (Fig. 1).The study groups were compared using χ 2 test for categorical variables and One-way ANOVA or Student-T test for comparison of continuous variables.A signi cant difference was considered when the 2-tailed p-value was less than 0.05.Moreover, the percentages of change in the mean values of IL-6, IL-18, and hs-CRP from the baseline values were expressed as mean and 95% con dence interval (95% CI).Logistic regression analysis was performed to determine the age-sex-GCS adjusted odd ratio of on-admission blood glucose for the injury severity (ISS ≥ 16) and hospital mortality.
Also, correlation of the average rate of change of in ammatory markers with respect to initial glucose level, ISS, TRISS and hospital length of stay was performed using Pearson's correlation.Data analysis was carried out using the SPSS version 18 (SPSS Inc., Chicago, Illinois).

Analysis of the 4 subgroups
The 4 trauma patients' groups were DH (n = 16), SIH (n = 16), diabetic normoglycemia (n = 11), and nondiabetic normoglycemia (n = 207) as shown in Table 2. Compared to the other 3 groups, patients with SIH were signi cantly younger (mean age 32 years), more obese, had severe injuries (mean ISS 24.5 ± 12.3); higher IL-6 levels at the 3 time points, prolonged hospital length of stay and higher mortality (p = 0.005).However, the 4 groups were comparable in terms of serum IL-18 and C-reactive protein levels at the different time points.Patients with diabetic hyperglycemia had higher hs-TnT and needed more units of blood transfusion.Higher shock index (> 0.8) was observed in both types of hyperglycemia compared to normoglycemic groups.Figure 2 demonstrates the trend of in ammatory markers and blood glucose levels in trauma patients.
The serum levels of IL-6, IL-18 and blood glucose increased after injury then showed a slowly decreasing trend, but did not reach baseline after 48 h.

Discussion
This is a prospective study to identify the patterns and effect of initial hyperglycemia and in ammatory biomarkers in trauma patients.Up to our knowledge, the constellation of on-admission random blood glucose, proin ammatory cytokines and injury severity among trauma patients is understudied.There are several key ndings of this study.Almost 13% of trauma patients had on-admission hyperglycemia; half of them had SIH.Hyperglycemic patients showed higher association with elevated IL-6, IL-18 and hs-CRP compared to the normoglycemic patients.Also, patients with hyperglycemia were more likely to have severe injuries, prolonged hospitalization and higher mortality than normoglycemic patients.Moreover, IL-6 level was greater in SIH (non-diabetic) compared to DH group.Logistic regression analysis showed that adjusted on-admission blood glucose was predictor for injury severity and not for hospital mortality.
Initial readings of IL-6 correlated signi cantly with the ISS.The blood glucose levels showed signi cant correlation with serum IL-6, serum lactate, ISS, and length of hospitalization.Also, we found an association between hyperglycemia and hs-TnT.Prior works revealed a signi cant association between the severity of trauma and positivity of hs-TnT as a re ection of traumatic stress [21,22].Also, trauma patients with hyperglycemia presented with a higher shock index (> 0.8) which indicates worse presentation and outcome [23,24].
The current study demonstrated that patients with SIH had a 3-fold higher rate of mortality as compared to those with DH.These ndings are in accordance with earlier studies which showed signi cantly greater risk of mortality in patients with SIH as opposed to those who had DH [11,15,17,25].
A recent study on thoracoabdominal injury patients demonstrated a higher rate of mortality in nondiabetic patients with on-admission hyperglycemia as compared to those with initial normoglycemia [26].Furthermore, a prospective observational study of traumatic brain injury (TBI) reported a marked hyperglycemia in patients with severe TBI which independently predicted the poor short-term neurological outcome [27].
Prior studies showed a relationship between serum cortisol, catecholamine levels and severity of injury.Patients with severe injuries were more likely to develop SIH [5,[28][29][30].Consistent with these observations, hyperglycemia patients in our cohort had a higher injury severity as compared to normoglycemic patients.
In our study, the overall complications were higher in both types of hyperglycemia compared to normoglycemia groups, but the difference did not reach statistical signi cance.A prior study reported that initial hyperglycemia in trauma patients correlated with serum lactate and ISS and was associated with higher mortality; however, the rate of infection was not signi cantly higher [31].
In our patients, the increase in hs-CRP was detected after 24 h of trauma and reached its peak value at 48 h.Giannoudis et al., [32] reported that serum CRP levels were within the normal range on the initial presentation which then gradually increased and reached the peak value on the third day post-trauma.
The authors also found an association between ISS and IL-6 levels but such association was not observed with CRP.Consistent with our study, earlier studies reported a signi cant correlation between higher ISS and IL-6 level on the initial presentation [33][34][35].We also observed a higher level of Il-18 in DH in comparison to the other groups including SIH (but statistically non-signi cant); a nding that needs further explanation [36].Our study performed serial measurements of blood glucose and cytokines to understand the complex relationship between hyperglycemia and in ammatory response in trauma.These ndings indicated that immunoneuroendocrine alterations might be involved in the pathophysiology of trauma patients [16].

Limitations
The rst limitation is that patients with DH may have also some degree of stress response which was underestimated.Second, we could not measure the levels of stress response hormones or catecholamines.Third, selection bias cannot be ignored along the study period.In the emergency department, we were using sliding scale insulin for patients with hyperglycemia; however, we did not measure the effect of exogenous insulin on the cytokine's levels.Apart from the signi cant difference in age, we could not explain the mortality rate in the normoglycemic diabetic group which was relatively similar to that of the SIH.Finally, the HbA1c level might not be accurate in patients necessitate early blood transfusion; as transfusion may alter the HbA1c reading [37][38].
In conclusion, patients with on-admission hyperglycemia have more severe injury and worse hospital outcome compared to normoglycemia patients.The initial blood glucose correlates with serum IL-6 which indicates a potential role of the systemic in ammatory response in the disease pathogenesis among severely injured patients.On-admission glucose level could be a useful marker of injury severity, triage and risk assessment in trauma patients.These observations warrant further evaluation in larger multicenter studies.

Table 1
compares the demographics, clinical presentation and outcome of trauma patients according to the initial blood glucose levels (hyperglycemia versus normoglycemia).The two groups were comparable for age, gender and BMI.Compared with the normoglycemia group, hyperglycemic patients had higher HbA1c and positive hsTnT (51.6% vs. 16.2%;p=0.001).In the hyperglycemia group, serum concentrations of IL-6 at baseline, at 24 h and 48 h post trauma were signi cantly higher than those of the normoglycemia group.Moreover, the hyperglycemia group had a signi cantly higher mean ISS(22.6*random blood glucose ≥ 11.1 mmol/l at baseline, TRISS: trauma injury severity score

Table 3
Bivariate correlation between blood glucose and other factors * baseline; revised trauma score; TRISS: trauma injury severity score; ISS: injury severity score; HsCRP: high sensitive protein; ED: emergency department Table

Table 4
Presentation of in ammatory markers and glucose level based on injury severity score (ISS)