Insulin response and changes in composition of non-esterified fatty acids in blood plasma of middle-aged men following isoenergetic fatty and carbohydrate breakfasts

It was previously shown that a high plasma concentration of non-esterified fatty acids (NEFA) persisted after a fatty breakfast, but not after an isoenergetic carbohydrate breakfast, adversely affecting glucose tolerance. The higher concentration after the fatty breakfast may in part have been a result of different mobilization rates of fatty acids. This factor can be investigated as NEFA mobilized from tissues are monounsaturated to a greater extent than those deposited from a typical meal. Twenty-four middle-aged healthy Caucasian men were given oral glucose tolerance tests (OGTT), and for 28 d isoenergetic breakfasts of similar fat composition but of low (L) or moderate (M) fat content. The composition of NEFA in fasting and postprandial plasma was determined on days 1 and 29. No significant treatment differences in fasting NEFA composition occurred on day 29. During the OGTT and 0-1 h following breakfast there was an increase in plasma long-chain saturated NEFA but a decrease in monounsaturated NEFA (mug/100 mug total NEFA; Pg/100 mug total NEFA; P<0.05), expressed as an increase in 18:1 and decreases in 16:0 and 17:0 in treatment M relative to treatment L (P<0.05). Serum insulin attained 35 and 65 mU/l in treatments M and L respectively during this period. Negative correlations were found between 16:0 in fasting plasma and both waist:hip circumference (P=0.0009) and insulin response curve area during OGTT (within treatment M, P=0.0001). It is concluded that a normal postprandial insulin response is associated with a rapid change in plasma saturated:monounsaturated NEFA. It is proposed that this change is the result of a variable suppression of fat mobilization, which may partly account for a large difference in postprandial total plasma NEFA between fatty and carbohydrate meals

Advanced monitoring in traumatic brain injury: microdialysis

Purpose of review: Here, we review the present state-of-the-art of microdialysis for monitoring patients with severe traumatic brain injury, highlighting the newest developments. Microdialysis has evolved in neurocritical care to become an established bedside monitoring modality that can reveal unique information on brain chemistry. Recent findings: A major advance is recent consensus guidelines for microdialysis use and interpretation. Other advances include insight obtained from microdialysis into the complex, interlinked traumatic brain injury disorders of electrophysiological changes, white matter injury, inflammation and metabolism. Summary: Microdialysis has matured into being a standard clinical monitoring modality that takes its place alongside intracranial pressure and brain tissue oxygen tension measurement in specialist neurocritical care centres, as well as being a research tool able to shed light on brain metabolism, inflammation, therapeutic approaches, blood–brain barrier transit and drug effects on downstream targets. Recent consensus on microdialysis monitoring is paving the way for improved neurocritical care protocols. Furthermore, there is scope for future improvements both in terms of the catheters and microdialysate analyser technology, which may further enhance its applicability.K.L.H.C. receives support from National Institute for Health Research Biomedical Research Centre, Cambridge (Neuroscience Theme; Brain Injury and Repair Theme); A.M.H.Y., National Institute for Health Research Academic Clinical Fellowship; P.J.H., National Institute for Health Research Professorship, Academy of Medical Sciences/Health Foundation Senior Surgical Scientist Fellowship and the National Institute for Health Research Biomedical Research Centre, Cambridge

Microdialysis Monitoring in Clinical Traumatic Brain Injury and Its Role in Neuroprotective Drug Development

Injuries to the central nervous system continue to be vast contributors to morbidity and mortality; specifically, traumatic brain injury (TBI) is the most common cause of death during the first four decades of life. Several modalities are used to monitor patients suffering from TBI in order to prevent detrimental secondary injuries. The microdialysis (MD) technique, introduced during the 1990s, presents the treating physician with a robust monitoring tool for brain chemistry in addition to conventional intracranial pressure monitoring. Nevertheless, some limitations remain, such as limited spatial resolution. Moreover, while there have been several attempts to develop new potential pharmacological therapies in TBI, there are currently no available drugs which have shown clinical efficacy that targets the underlying pathophysiology, despite various trials investigating a plethora of pharmaceuticals. Specifically in the brain, MD is able to demonstrate penetration of the drug through the blood-brain barrier into the brain extracellular space at potential site of action. In addition, the downstream effects of drug action can be monitored directly. In the future, clinical MD, together with other monitoring modalities, can identify specific pathological substrates which require tailored treatment strategies for patients suffering from TBI.The author(s) gratefully acknowledge receipt of the following financial support. Medical Research Council (Grant nos. G0600986 ID79068 and G1002277 ID98489) and National Institute for Health Research Biomedical Research Centre, Cambridge (Neuroscience Theme; Brain Injury and Repair Theme). Authors’ support: EPT—the Swedish Society of Medicine (Grant no. SLS-587221) and the Swedish Brain Foundation; KLHC—the National Institute for Health Research Biomedical Research Centre, Cambridge (Neuroscience Theme; Brain Injury and Repair Theme); PJH—the National Institute for Health Research Professorship, the Academy of Medical Sciences/Health Foundation Senior Surgical Scientist Fellowship and the National Institute for Health Research Biomedical Research Centre, Cambridge; AH—the Medical Research Council/Royal College of Surgeons of England Clinical Research Training Fellowship (Grant no. G0802251)

Heparin-gold nanoparticles for enhanced microdialysis sampling

Cerebral microdialysis is a sampling technique which offers much potential for understanding inflammatory pathophysiology following traumatic brain injury (TBI). At present, the recovery of cytokines via microdialysis in clinical studies is not straightforward primarily due to their size, steric properties and low concentrations. Heparin, and heparin-coated microspheres, have previously shown promise as cytokine binding agents for enhanced microdialysis sampling in animal models [1, 2]. However, there are several factors limiting application for microdialysis in patients. The aim of this study was to produce heparin-coated gold nanoparticles as cytokine capture agents for enhanced microdialysis sampling, potentially applicable to a clinical setting. Gold nanoparticles (AuNP) were chemically conjugated to heparin via a bifunctional polyethylene glycol (PEG) linker. The heparin AuNP (AuNP-Hep) were characterised, demonstrating the successful addition of heparin to the gold surface. The performance of the AuNP-Hep during in vitro testing was compared both to current methodology [3], and to the heparin-coated microspheres developed by Duo and Stenken [1, 2]. The AuNP-Hep yielded a higher recovery of cytokines compared to current methodology and heparin-coated microspheres, during in vitro testing designed to mimic the human environment and the intensive care unit. In this study, AuNP-Hep were developed for enhanced microdialysis sampling of cytokines, potentially applicable in a clinical setting. Based on the success of the AuNP-Hep in vitro, the proposed method offers a candidate alternative to the use of current protocols that rely on a blood product (albumin) for microdialysis sampling of cytokines in patients.The work was supported by funding for SGC and KLHC from the National Institute for Health Research Biomedical Research Centre, Cambridge (Neuroscience Theme; Brain Injury and Repair Theme). HBF was supported by a University of Melbourne Global Scholar’s Award and a University of Melbourne Mobility Financial Assistance Grant for travel to Cambridge. PJH is funded by a National Institute for Health Research Professorship, Academy of Medical Sciences/Health Foundation Senior Surgical Scientist Fellowship and the National Institute for Health Research Biomedical Research Centre, Cambridge

Assessing metabolism and injury in acute human traumatic brain injury with magnetic resonance spectroscopy: current and future applications

Traumatic brain injury triggers a series of complex pathophysiological processes. These include abnormalities in brain energy metabolism; consequent to reduced tissue pO₂ arising from ischaemia or abnormal tissue oxygen diffusion, or due to a failure of mitochondrial function. In-vivo magnetic resonance spectroscopy (MRS) allows non-invasive interrogation of brain tissue metabolism in patients with acute brain injury. Nuclei with ‘spin’ e.g. ¹H, ³¹P and ¹³C, are detectable using MRS and are found in metabolites at various stages of energy metabolism, possessing unique signatures due to their chemical shift or spin-spin interactions (J-coupling). The most commonly used clinical MRS technique, ¹H MRS, uses the great abundance of hydrogen atoms within molecules in brain tissue. Spectra acquired with longer echo-times include N-acetylaspartate, creatine and choline. N-acetylaspartate, a marker of neuronal mitochondrial activity related to ATP, is reported to be lower in patients with TBI than healthy controls, and the ratio of N-acetylaspartate/creatine at early time points may correlate with clinical outcome. ¹H MRS acquired with shorter echo-times produces a more complex spectrum, allowing detection of a wider range of metabolites. ³¹P MRS detects high energy phosphate species, which are the end-products of cellular respiration: adenosine triphosphate (ATP) and phosphocreatine. ATP is the principal form of chemical energy in living organisms, and phosphocreatine (PCr) is regarded as a readily mobilised reserve for its replenishment during periods of high utilisation. The ratios of high energy phosphates are thought to represent a balance between energy generation, reserve and use in the brain Additionally, the chemical shift difference between Pi and PCr enables calculation of intracellular pH. ¹³C MRS detects the ¹³C-isotope of carbon in brain metabolites. As the natural abundance of ¹³C is low (1.1%), ¹³C MRS is typically performed following administration of ¹³C-enriched substrates which permits tracking of the metabolic fate of the infused ¹³C in the brain over time, and calculation of metabolic rates in a range of biochemical pathways, including glycolysis, the tricarboxylic acid (TCA) cycle, and glutamate-glutamine cycling. The advent of new hyperpolarization techniques to transiently boost signal in ¹³C-enriched MRS in-vivo studies shows promise in this field and further developments are expected.The funding bodies acknowledged on the paper are: PJAH is supported by a National Institute for Health Research (NIHR) Research Professorship, Academy of Medical Sciences/Health Foundation Senior Surgical Scientist Fellowship and the National Institute for Health Research Biomedical Research Centre, Cambridge. PJAH and KLHC are supported by the NIHR Biomedical Research Centre, Cambridge. MGS is supported by PH’s NIHR Research Professorship. AS is funded by the NIHR via an award to the Cambridge NIHR/Wellcome Trust Clinical Research Facility

Succinate supplementation improves metabolic performance of mixed glial cell cultures with mitochondrial dysfunction

Mitochondrial dysfunction, the inability to efficiently utilise metabolic fuels and oxygen, contributes to pathological changes following traumatic spinal cord or traumatic brain injury (TBI). In the present study, we tested the hypothesis that succinate supplementation can improve cellular energy state under metabolically stressed conditions in a robust, reductionist in vitro model of mitochondrial dysfunction in which primary mixed glial cultures (astrocytes, microglia and oligodendrocytes) were exposed to the mitochondrial complex I inhibitor rotenone. Cellular response was determined by measuring intracellular ATP, extracellular metabolites (glucose, lactate, pyruvate), and oxygen consumption rate (OCR). Rotenone produced no significant changes in glial ATP levels. However, it induced metabolic deficits as evidenced by lactate/pyruvate ratio (LPR) elevation (a clinically-established biomarker for poor outcome in TBI) and decrease in OCR. Succinate addition partially ameliorated these metabolic deficits. We conclude that succinate can improve glial oxidative metabolism, consistent our previous findings in TBI patients' brains. The mixed glial cellular model may be useful in developing therapeutic strategies for conditions involving mitochondrial dysfunction, such as TBI

Delineating Astrocytic Cytokine Responses in a Human Stem Cell Model of Neural Trauma

Neuroinflammation has been shown to mediate the pathophysiological response following traumatic brain injury (TBI). Accumulating evidence implicates astrocytes as key immune cells within the central nervous system (CNS), displaying both pro- and anti-inflammatory properties. The aim of this study was to investigate how in vitro human astrocyte cultures respond to cytokines across a concentration range that approximates the aftermath of human TBI. To this end, enriched cultures of human induced pluripotent stem cell (iPSC)-derived astrocytes were exposed to interleukin-1β (IL-1β) (1–10,000 pg/mL), IL-4 (1–10,000 pg/mL), IL-6 (100–1,000,000 pg/mL), IL-10 (1–10,000 pg/mL) and tumor necrosis factor (TNF)-α (1–10,000 pg/mL). After 1, 24, 48 and 72 h, cultures were fixed and immunolabeled, and the secretome/supernatant was analyzed at 24, 48, and 72 h using a human cytokine/chemokine 39-plex Luminex assay. Data were compared to previous in vitro studies of neuronal cultures and clinical TBI studies. The secretome revealed concentration-, time- and/or both concentration- and time-dependent production of downstream cytokines (29, 21, and 17 cytokines, respectively, p<0.05). IL-1β exposure generated the most profound downstream response (27 cytokines), IL-6 and TNF had intermediate responses (13 and 11 cytokines, respectively), whereas IL-4 and IL-10 only led to weak responses over time or in escalating concentration (8 and 8 cytokines, respectively). Notably, expression of IL-1β, IL-6, and TNF cytokine receptor mRNA was higher in astrocyte cultures than in neuronal cultures. Several secreted cytokines had temporal trajectories, which corresponded to those seen in the aftermath of human TBI. In summary, iPSC-derived astrocyte cultures exposed to cytokine concentrations reflecting those in TBI generated an increased downstream cytokine production, particularly IL-1β. Although more work is needed to better understand how different cells in the CNS respond to the neuroinflammatory milieu after TBI, our data shows that iPSC-derived astrocytes represent a tractable model to study cytokine stimulation in a cell type-specific manner

Monitoring the Neuroinflammatory Response Following Acute Brain injury

Traumatic brain injury (TBI) and subarachnoid hemorrhage (SAH) are major contributors to morbidity and mortality. Following the initial insult, patients may deteriorate due to secondary brain damage. The underlying molecular and cellular cascades incorporate components of the innate immune system. There are different approaches to assess and monitor cerebral inflammation in the neuro intensive care unit. The aim of this narrative review is to describe techniques to monitor inflammatory activity in patients with TBI and SAH in the acute setting. The analysis of pro- and anti-inflammatory cytokines in compartments of the central nervous system (CNS), including the cerebrospinal fluid and the extracellular fluid, represent the most common approaches to monitor surrogate markers of cerebral inflammatory activity. Each of these compartments has a distinct biology that reflects local processes and the cross-talk between systemic and CNS inflammation. Cytokines have been correlated to outcomes as well as ongoing, secondary injury progression. Alongside the dynamic, focal assay of humoral mediators, imaging, through positron emission tomography, can provide a global in vivo measurement of inflammatory cell activity, which reveals long-lasting processes following the initial injury. Compared to the innate immune system activated acutely after brain injury, the adaptive immune system is likely to play a greater role in the chronic phase as evidenced by T-cell-mediated autoreactivity toward brain-specific proteins. The most difficult aspect of assessing neuroinflammation is to determine whether the processes monitored are harmful or beneficial to the brain as accumulating data indicate a dual role for these inflammatory cascades following injury. In summary, the inflammatory component of the complex injury cascade following brain injury may be monitored using different modalities. Using a multimodal monitoring approach can potentially aid in the development of therapeutics targeting different aspects of the inflammatory cascade and improve the outcome following TBI and SAH