Systematic review regarding metabolic profiling for improved pathophysiological understanding of disease and outcome prediction in respiratory infections

Metabolic profiling through targeted quantification of a predefined subset of metabolites, performed by mass spectrometric analytical techniques, allows detailed investigation of biological pathways and thus may provide information about the interaction of different organic systems, ultimately improving understanding of disease risk and prognosis in a variety of diseases. Early risk assessment, in turn, may improve patient management in regard to cite-of-care decisions and treatment modalities. Within this review, we focus on the potential of metabolic profiling to improve our pathophysiological understanding of disease and management of patients. We focus thereby on lower respiratory tract infections (LRTI) including community-acquired pneumonia (CAP) and chronic obstructive pulmonary disease (COPD), an important disease responsible for high mortality, morbidity and costs worldwide. Observational data from numerous clinical and experimental studies have provided convincing data linking metabolic blood biomarkers such as lactate, glucose or cortisol to patient outcomes. Also, identified through metabolomic studies, novel innovative metabolic markers such as steroid hormones, biogenic amines, members of the oxidative status, sphingo- and glycerophospholipids, and trimethylamine-N-oxide (TMAO) have shown promising results. Since many uncertainties remain in predicting mortality in these patients, further prospective and retrospective observational studies are needed to uncover metabolic pathways responsible for mortality associated with LRTI. Improved understanding of outcome-specific metabolite signatures in LRTIs may optimize patient management strategies, provide potential new targets for future individual therapy, and thereby improve patients’ chances for survival.


Metabolomics
The metabolome can be considered the "quantitative complement of all the low molecular weight molecules (<1500 amu) in a particular physiological or developmental state" of a cell, tissue or organism [1]. Metabolomics is defined as "the analysis of the whole metabolome under a given set of physiological, environmental and/or clinical conditions" [2]. Metabolite profiling through targeted quantification of a predefined subset of metabolites of the metabolome [3] allows analysis of metabolic pathways, thereby clarifying the interaction of different organic systems. While proteomics is the study of proteins made by the organism and the conditions under which the organism makes them, metabolomics is a more systematic study of the biochemical fingerprints that result of cellular processes.
Mass spectrometry (MS) is a sensitive tool that characterizes and quantifies metabolites in a biological sample [4]. Since metabolomics requires proper separation of the different compounds to be analyzed, chemical separation techniques such as gas chromatography (GC) and liquid chromatography (LC) or capillary electrophoresis (CE) can be combined with MS detection. Analytical techniques such as LC-MS, GC-MS, CE-MS, and matrixassisted laser desorption ionization-MS (MALDI-MS) may therefore expand coverage of the metabolome [5]. MS is now a routine diagnostic instrument in clinical laboratories [6].
Biological information derived from these techniques can provide accurate and clinically useful diagnostic capability for the management of diseases [7,8] by mapping disease risk to metabolic pathways [9]. Previously, many studies have focused on risk prediction in patients with lower respiratory tract infections (LRTI) including community-acquired pneumonia (CAP) and case (COPD) because such information may have direct consequences in patient management [10][11][12][13]. Early risk prediction may improve site-of-care decisions (i.e., ICU treatment vs. hospital ward vs. outpatient) as well as treatment modalities (i.e., antibiotic drugs) [11,14,15]. Indeed, improved understanding of metabolic interactions in LRTI patients may enhance prediction of outcomes and provide targets for individual therapy, which in turn could facilitate personalized patient management (Fig. 2) [16][17][18].

Prediction of mortality in community-acquired pneumonia
CAP is the leading cause of infectious death [19], and of high mortality, morbidity, and costs worldwide [20][21][22]. Patients with CAP manifest high short-term mortality [23,24] but also substantial long-term morbiditiy [25][26][27][28]. To predict all-cause mortality within 30 days, the pneumonia severity index (PSI) is a validated tool that categorizes CAP patients within distinct risk classes (I-V) [29]. In Europe, the CURB-65 score (Confusion, Urea, Respiratory rate, Blood pressure, Age >65 years) is used for the same purpose [30]. Several clinical studies show that measuring specific inflammatory or metabolic blood biomarkers, including among others proadrenomedullin (proADM) [14,15,[31][32][33] and procalcitonin (PCT) [26,[34][35][36], provides prognostic information and possibly improves short-term risk stratification and management decisions in CAP compared to the use of clinical risk scores alone [37]. Pneumonia not only increases short-term mortality; clinical trials also found greater long-term mortality rates in patients who survived an initial CAP episode, compared to the general population and to patients who have other infections [28,38,39]. These observations suggest that CAP is a poor prognostic indicator of long-term outcomes.
Particularly, it has been hypothesized that cardiovascular events that are triggered by an episode of respiratory infection may be responsible for the excess mortality observed in large long-term CAP cohorts. For this reason, cardiovascular biomarkers (such as natriuretic peptides) may also help in the identification of high risk LRTI patients who need close monitoring and follow-up [28]. The metabolomics approach for LRTI thus may also look into metabolites that have been linked to cardiovascular disease as these conditions are being linked very closely.
Prediction of mortality in patients hospitalized with exacerbated chronic obstructive pulmonary disease Prediction of outcomes in chronic obstructive pulmonary disease (COPD) is challenging. Three published observational studies suggest that the inflammatory blood biomarker ProADM independently predicts mediumterm all-cause mortality in patients with stable [40,41] or exacerbated COPD. Additionally, measurement of ProADM combined with the clinical parameters of body mass index, airflow obstruction, dyspnea and exercise capacity index (BODE) [42] improves the predictive power of medium-term all-cause mortality compared to the use of BODE alone [40]. A recent observational clinical cohort study demonstrated that ProADM levels are independent predictors of 5-to 7-year all-cause mortality, and may enhance long-term prognostic accuracy of demographic and clinical variables in patients hospitalized for exacerbation of pneumonic or non-pneumonic COPD [32]. Similar to CAP, in COPD cardiovascular disease may be responsible for large portions of the high mortality associated with this disease. Again, the metabolomic approach in COPD may therefore also consider interesting metabolites associated with adverse cardiovascular outcomes.

Aims of the review and methodology
The goal of the present review is to investigate wether new metabolic blood biomarkers help optimizing identification of patients at risk for poor outcome in CAP and exacerbated COPD. We discuss nine metabolite classes: steroid hormones, biogenic amines, intermediate energy metabolism, monosaccharides, trimethylamines, oxidative status, sphingolipids, glycerophospholipids, and acylcarnitines. Each section will begin with a brief description of pathways and known physiological effects, followed by a summary of observational data regarding the analyte's role in predicting all-cause mortality. We focus on patients with LRTI which is responsible for high mortality, morbidity and costs worldwide. Yet, if no data are available in strictly LRTI patients, we also discuss these markers in other diseases namely cardiovascular disease which may account for a large proportion of deaths attributable to LRTI.

Steroid hormones Pathway and physiological role
Only few tissues contribute substantially to de novo steroid biosynthesis: primarily the adrenal glands, the gonads, and the placenta [43]. All steroid hormones are derived from cholesterol. Cholesterol enters the cell via lipoprotein-binding and is stored in cytoplasmic vacuoles. After binding of adrenocorticotropic hormone (ACTH) to its receptor (melanocortin receptor type 2, MC2R) on adrenal cells, cholesterol is mobilized and thus available for steroid synthesis. Follicle-stimulating hormone (FSH) is critical in estradiol synthesis before and during ovulation and Luteinizing hormone (LH) for synthesis of progesterone. In the placenta, steroid biosynthesis is dependent on human chorionic gonadotropin (hCG) in early gestation [43].
One of the first measurable physiopathological reactions to infection is the activation of the hypothalamopituitary-adrenal (HPA) axis via stimulation of the central noradrenergic stress system by cytokines and other mediators that are released in response to inflammation [44][45][46]. Under healthy conditions, cortisol secretion is synchronized with dehydroepiandrosterone (DHEA) in response to the release of ACTH and corticotropinreleasing hormone (CRH), whereas in critically ill patients the circadian pattern of cortisol is lost [47] and DHEA and dehydroepiandrosterone-sulfate (DHEA-S) decrease [48]. Although the physiological role of DHEA and DHEA-S is not completely understood, there are clear indications that they modulate the immune response and influence proinflammatory and anti-inflammatory cytokine release [48][49][50][51][52]. In severely ill pneumonia patients, an intra-adrenal shift from DHEA-S to cortisol production occurs, which might be a potentially life-saving process during critical illness and thus important for survival in CAP [53]. Estradiol induces activation of macrophages and production of proinflammatory cytokines and chemokines from inflammatory cells [54][55][56], and testosterone has suppressive effects on immune responses, thereby inducing increased suceptibility to infection [57]. In autoimmune diseases progesterone shows proinflammatory and anti-inflammatory effects [58].
Current research (Table 1)   Table 1 lists several studies reporting that increased cortisol (free and total) levels measured at initial hospital presentation are independent predictors of in-hospital and short-term all-cause mortality in hospitalized patients with mild to severe CAP [53,[59][60][61][62]. Interestingly,      -Only 32 deaths and thus relatively low statistical power ACTH adrenocorticotropic hormone, APACHE II Acute Physiology And Chronic Health Evaluation, ApoE apolipoprotein E, AUC area under the receiver operating characteristic curve, CAD coronary artery disease, CAP community-acquired pneumonia, CAPNETZ network of excellence community-acquired pneumonia, Germany, cFDHT calculated free dihydrotestosterone, cFT calculated free testosterone, CHS Cardiovascular Health Study, CI confidence interval, CKD chronic kidney disease, COPD chronic obstructive pulmonary disease, CRP C-reactive protein, CURB-65 score new-onset confusion urea >7 mmol/L respiratory rate ≥30 breaths per minute systolic or diastolic blood pressure <90 mmHg or ≤60 mmHg respectively age ≥65 years (pneumonia/LRTI risk scoring system), CVD cardiovascular disease, DETECT Diabetes Cardiovascular Risk-Evaluation: Targets and Essential Data for Commitment of Treatment, DHEA dehydroepiandrosterone, DHEA-S dehydroepiandrosterone-sulfate, DHT dihydrotestosterone, FC free cortisol, FT free testosterone, HD hemodialysis, HR hazard ratio, hsCRP high-sensitive C-reactive protein, ICU intensive care unit, IHD ischemic heart disease, IL-1beta interleukin-1 beta, KTR kidney transplant recipients, LDL low-density lipoprotein, MMAS Massachusetts Male Aging Study, OR odds ratio, p p-value are statistically significant at p < 0.05, PSI pneumonia severity index, Q quartile, ROC receiver operationg characteristic, RR relative risk, sd-LDL small dense low-density lipoprotein, SHBG sex hormone binding globuline, SHIP Study of Health in Pomerania, TC total cortisol, TT total testosterone, VLDL very-low-density lipoprotein the predictive accuracy of free cortisol (FC) is equal to that of total cortisol (TC) in pneumonia, independent of serum albumin levels [59]. The prognostic accuracy of cortisol in predicting mortality is equal to that of the PSI, but higher compared to routinely measured laboratory parameters such as CRP, PCT or leukocytes, which do not show significant differences (p > 0.05) [59][60][61]. Kolditz et al. observed that serum cortisol levels may noticeably improve the predictive power of the CURB-65 score alone as regards short-term mortality [60,61]. Salluh et al. demonstrated that compared to baseline serum cortisol levels, which are proven independent mortality predictors, neither postcorticotropin cortisol nor Δ-cortisol has notable predictive power in severe CAP [62]. A recent animal study of canines suffering from staphylococcus aureus pneumonia showed that ACTH stimulation induces a pronounced correlation between all-cause mortality and cortisol (free and total) levels and Δ-cortisol [63].
Conflicting results mark studies evaluating the role of serum DHEA as a mortality predictor. A prospective clinical trial conducted by Mueller et al. demonstrated that increased DHEA predicts 6-week mortality in CAP, whereas blood levels of DHEA-S as well as different ratios between cortisol and DHEA, cortisol and DHEA-S, and DHEA and DHEA-S shows no significant correlation with all-cause mortality [53]. Contrary to these findings, Kolditz et al. found no difference in DHEA levels in non-survivors compared to survivors, whereas the lack of significance for the ratio between DHEA and DHEA-S as a mortality predictor accords with the results of Mueller et al. [53,61]. Data on patients suffering from sepsis reveal a noticeable dissociation between cortisol and DHEA in non-survivors compared to survivors, and from this observation it can be inferred that an increased cortisol/DHEA ratio is a prognostic marker in sepsis [64]. However, although the cortisol/DHEA ratio in clinical studies of CAP patients is not significant in the prediction of mortality, nevertheless, in nonsurvivors an increased cortisol/DHEA ratio occurs as a tendency [53]. In elderly Swedish men, an inverse association between serum DHEA and DHEA-S is evident, whereas low serum DHEA and high DHEA-S levels are independent predictors of all-cause mortality [65]. Furthermore Hsu et al. demonstrated a marked and independent association between low levels of plasma DHEA-S and higher mid-term all-cause mortality in men with chronic kidney disease (CKD) who were undergoing hemodialysis [66]. Table 1 depicts the strong association between increased serum estradiol and progesterone levels and 28-day mortality in patients suffering from pneumoniarelated septic shock, whereas serum testosterone levels are not noticeably different between survivors and non-survivors after a 28-day follow-up [67]. A prospective clinical study of elderly men who were free of cardiovascular disease at the time of blood sample collection, revealed a significant inverse association between dehydrotestosterone (DHT) and calculated free DHT levels with all-cause mortality during a follow-up period of 9 years, whereas total testosterone and calculated free testosterone showed no significant correlation with mortality [68]. Contrary to these findings, Friedrich et al. and Grossmann et al. observed that low serum total testosterone levels in men with and without CKD significantly and independently predict all-cause mortality [69,70], whereas another population-based clinical study of 40-to 70-yearold men [71] observed no significant association with either total or free testosterone, or with DHT in all-cause mortality in multivariate-adjusted analysis.
In postmenopausal women low estrone levels independently predict long-term all-cause mortality [72].
Biogenic amines I. L-arginine metabolites: Asymmetric dimethylarginine and symmetric dimethylarginine Pathway and physiological role Endothelial nitric oxide synthases (NOS) synthesize nitric oxide (NO) from L-arginine (Arg). NO is responsible for vasodilatation and acts as a neurotransmitter with several functions, including memory formation [73]. NO promotes relaxation of airway smooth muscle in the respiratory system, but NO deficiency results in hyperreactivity of the airway [74]. Asymmetric dimethylarginine (ADMA) and its isomer symmetric dimethylarginine (SDMA) are endogenous compounds of methylated protein turnover. ADMA-but not SDMA-is a competitive inhibitor of NOS and thus indirectly of NO [75]. One study shows that ADMA is present in the sputum of COPD patients [76].

Current research (Table 2)
A large community-based study of middle-aged participants revealed a positive association between ADMA levels and long-term mortality, whereas the Arg/ADMA ratio was inversely associated with death rate [77]. In people >65 years, ADMA is the strongest predictor of all-cause mortality, while some traditional risk factors-e.g., body mass index, systolic and diastolic blood pressure, male sex or triglycerides-lose their predictive power in this age group [78]. Additionally, elevated ADMA and SDMA serum levels correlate noticeably with higher all-cause mortality in patients with stable CHD [79] and in critically ill patients [80,81]. Contrary to these findings, another study-after adjustment for NT-proBNP, hsCRP, troponin T and cardiorenal indices-reveaed that SDMA, but not ADMA, is an independent predictor of all-cause mortality [82].  -Relatively small size study -Single-center study -High 3-NT/Tyr was associated with higher 90-day mortality (p = 0.025) 3-NT 3-nitrotyrosine, ACS acute coronary syndrome, ADMA asymmetric dimethylarginine, AKI acute kidney injury, CAP community-acquired pneumonia, CHD coronary heart disease, CI confidence interval, CURB-65 score new-onset confusion, urea >7 mmol/L, respiratory rate ≥30 breaths per minute, systolic or diastolic blood pressure <90 mmHg or ≤60 mmHg, respectively, age ≥65 years (pneumonia/LRTI risk scoring system), HR hazard ratio, hsCRP high-sensitivity C-reactive protein, ICU intensive care unit, KAROLA Langzeitfolge der KARdiOLogischen Anschlussheilbehandlung, Germany, Kyn kynurenine, NT-proBNP pro-B-type natriuretic peptide, p p-value are statistically significant at p < 0.05, PSI pneumonia severity index, SDMA symmetric dimethylarginine, Trp tryptophan, Tyr tyrosine
Current research ( Table 2) Subjects with CAP [85] and sepsis [86] show significantly higher Kyn levels and lower Trp levels compared to controls. As a result, the Kyn/Trp ratio, which represents the IDO activity, is elevated in CAP and sepsis [85,86]. Increased severity of CAP (defined by PSI and CURB-65 score) [85] and of sepsis [86] are associated with higher Kyn levels, higher Kyn/Trp ratios and lower Trp levels. In patients with CAP [85] or bacteremia [87], short-term non-survivors have higher Kyn/Trp ratios than do survivors, whereas the sepsis group manifests no marked difference in Kyn/ Trp ratio between non-survivors and survivors in the short-term [86].

III. Nitrotyrosine
Pathway and physiological role Homeostatic balance is usually maintained between the formation of reactive oxygen species (ROS) and its removal by endogenous antioxidants. Oxidative stress arises from an imbalance towards ROS production [88]. Peroxynitrite (ONOO − ) is a potent oxidant of this ROS group and catalyzes nitration of the non-essential amino acid tyrosine (Tyr) to the non-proteinogenic amino acid 3-nitrotyrosine (3-NT) [89]. Accordingly, nitrotyrosine is a characteristic marker of oxidative stress (e.g., inflammation) [90]. Protein-associated nitrotyrosine is lower in smokers than in non-smokers, evidence consistent with a lower production of endothelial NO in cigarette smokers. In contrast, smokers with COPD show higher average nitrotyrosine levels in plasma proteins compared with smokers without COPD. Therefore, it seems that chronic inflammatory processes in COPD patients increase the nitration of Tyr [91]. Table 2, subjects with hospital-acquired acute kidney injury (AKI) have higher 3-nitrotyrosin/tyrosine ratios (3-NT/Tyr) than do critically ill subjects without AKI and healthy controls. In the AKI group, the 3-NT/Tyr ratio is positively associated with 90-day mortality independent of the severity of illness [92].

Trimethylamine-N-oxide, betaine, choline
Pathway and physiological role Synthesis of phospholipids to build cell membranes, acetylcholine for neurotransmission just as the methyl group metabolism depend on dietary phosphatidylcholine (PC)/choline intake [93]. The liver and kidney can both oxidize choline to betaine. Betaine is an important metabolite in methyl group metabolism; it functions as a methyl group donor to form methionine [94,95]. Eggs, red meat, beef and chicken liver, wheat germ, bacon, dried soy beans and pork contain the highest concentration of total choline. Betaine levels are high in wheat bread, wheat bran, wheat germ, spinach and shrimp [96]. Intestinal microbes (gut microflora) transform choline and other quaternary ammonium compounds (e.g., betaine) to the gas trimethylamine (TMA) [97]. This gas is reabsorbed efficiently into the circulation and then into human liver. Hepatic flavin-containing monooxygenases (FMOs) rapidly oxidize TMA to trimethylamine-N-oxide (TMAO) [98]. Thus TMA is not produced via intermediary metabolism; rather, it depends on intestinal microbial breakdown of choline and other precursors. Thus for the production of TMA and its oxidation product TMAO, intact and functional intestinal microflora are essential [97,99]. (Table 3) Several animal studies show the effects of elevated TMAO levels. An increase in the concentration of plasma TMAO was observed in mice, which were fed with either choline or TMAO [99,100]. After 16 weeks of feeding there was a marked increase in tubulointerstitial fibrosis and serum cystatin C [100]. A significant positive correlation between plasma levels of TMAO and atherosclerotic plaque size after 20 weeks of feeding in both male and female mice was observed. Furthermore, macrophages from mice supplemented with either choline, betaine or TMAO showed augmented lipid-loaded macrophages and higher levels of the macrophage scavenger receptors CD36 and SR-A1. These receptors implicate atherosclerosis. In a mouse model, application of broad-spectrum antibiotics for 3 weeks suppressed dietary choline-induced formation of macrophage foam cells and cholinemediated enhancement in atherosclerosis [99].

Current research
In humans (3), higher plasma NT-proBNP [101,102] and advanced left ventricular diastolic dysfunction are associated with elevated plasma betaine, choline and TMAO. TMAO levels are markedly higher in patients with heart failure NYHA III/IV or diabetes mellitus. Baseline plasma levels of TMAO have marked prognostic value for major adverse cardiovascular events in lowrisk subgroups after adjustments for age and cardiorenal indices [102,103]. Since cardiovascular disease (CVD) remains the major cause of deaths [104], these findings might be predictive of poor outcome. In addition, ACS acute coronary syndrome, BNP brain natriuretic peptide, CI confidence interval, CKD chronic kidney disease, CVD cardiovascular disease, eGFR estimated glomerular filtration rate, HF heart failure, HR hazard ratio, NT-proBNP pro-B-type natriuretic peptide, NYHA New York Heart Association functional classification, p p-value are statistically significant at p < 0.05; TMAO trimethylamine-N-oxide elevated TMAO levels are predictive of 5-year mortality risk in subjects with heart failure after adjustment for cardiovascular risk factors and renal function [101]. In healthy adults a dietary PC challenge raises TMAO level. In six human participants this elevation was suppressed with oral broad-spectrum antibiotics [103].
A high correlation is evident between plasma and urine levels of TMAO, which indicates an effective urinary clearance of TMAO [103]. Median TMAO blood levels in subjects with CKD stage 3 or beyond are markedly higher than in non-CKD subjects. Moreover, there is a modest correlation between elevated TMAO with low estimated glomerular filtration rate (eGFR) and high cystatin C. After adjustment for traditional cardiovascular risk factors an increased all-cause mortality rate at 5 years for CKD subjects with a higher TMAO level is evident. In subjects with preserved eGFR (>60 mL/min per 1.73 m 2 ) higher TMAO levels are associated with higher 5-year mortality, particularly in patients who have concomitant high cystatin C levels [100].

Intermediate energy metabolism Lactate
Pathway and physiological role Glycolysis produces the metabolite pyruvate. Lactate dehydrogenase (LDH) converts pyruvate to lactic acid under anaerobic conditions. Lactate acid dissociates spontaneously to lactate and H + in aqueous solutions. Thus lactate is the end product of anaerobic glucose metabolism in erythrocytes, perivenous hepatocytes, skeletal myocytes and skin cells. A normal plasma lactate concentration is 0.3-1.3 mmol/L and the liver removes 70 % of the lactate. An imbalance between systemic oxygen demand and oxygen availability (e.g., hypoperfusion in patients with sepsis) heightens anaerobic metabolism and results in lactic acidosis [105,106]. (Table 4) In children with pneumonia, relative risk for in-hospital mortality is higher if the lactate level is above 2 mmol/L [107]. In adults with pneumonia, lactate level is the better predictor of 28-day mortality than the CURB-65 score, whereas a combination of CURB-65 with lactate level improves the predictive value of CURB-65 alone (Table 4) [108].

Current research
Lactate >3.5 mmol/L in combination with a P a O 2 /FiO 2 (PF) ratio <170 is associated with a higher 28-day mortality rate; accordingly, a combination of serum lactate and the PF ratio may be a useful predictor of mortality in patients with sepsis, including pneumonia [109]. Metabolic profiling of plasma from 90 critically ill intensive care unit (ICU) patients revealed significantly higher lactate levels in non-survivors compared with survivors in 28-day mortality [110].

Monosaccharides
Pathway and physiological role Monosaccharides are the simplest form of sugar and therefore the most basic units of carbohydrates; they are important as an energy resource. The most important monosaccharides in human metabolism are the three aldohexoses (Dglucose, D-mannose, and D-galactose), and D-fructose, a sugar monomer of the ketohexose series of carbohydrates [111].

Current research (Table 5)
In patients with CAP (Table  5) , hypoglycemia [112] and hyperglycemia are associated with a higher rate of in-hospital mortality [112,113] and in-hospital complications [113]. Furthermore, hyperglycemia is a predictor of death at 28 and 90 days after admission to hospital in patients with CAP and no pre-existing diabetes [114].

Oxidative status
Pathway and physiological role Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are important in regulating the proliferation of cells and their survival. However, a sudden and prolonged surge of ROS and RNS may contribute to cell death. The redox homeostasis of cells, which maintains a balance between the generation and elimination of ROS and RNS ensures that endogenous and exogenous stimuli are modulated. Altered redox homeostasis leads to oxidative stress, which may cause aberrant cell death and the development of disease [115]. Glutathione (gamma-glutamylcysteinyl-glycine, GSH) is the predominant antioxidant non-protein cysteine-containing thiol, which is found in all animal cells. It is important in maintaining the status of cellular redox, which is determined by the ratio of the concentration of oxidizing equivalents to reducing equivalents [116], and it is the principal intracellular defense against oxidative stress. GSH has two redox forms: reduced GSH and glutathione disulfide (GSSG, the oxidized form) [117]. Cells can excrete GSSG or restore it to GSH through the action of GSH reductase. De novo synthesis of GSH from its amino-acid constituents-a process that involves two enzymatic steps catalyzed by glutamate cysteine ligase and GSH synthetase- [118] is essential for the adaptive elevation of GSH during oxidative stress [117]. The rate-limiting step in the de novo synthesis of GSH occurs at the cellular level of the amino acid cysteine [119]. Additionally, reduction of cystine to cysteine is an important mechanism for intracellular GSH elevation in vivo in lungs [120]. During oxidative stress and inflammation, synthesis of GSH is upregulated, possibly providing a protective or adaptive response to subsequent oxidative and inflammatory stress. An imbalance of antioxidant AUC area under the receiver operating characteristic curve, CI confidence interval, CURB-65 score new-onset confusion, urea >7 mmol/L, respiratory rate ≥30 breaths per minute, systolic or diastolic blood pressure <90 mmHg or ≤60 mmHg, respectively, age ≥65 years (pneumonia/LRTI risk scoring system), p p-value are statistically significant at p < 0.05 CAP community-acquired pneumonia, CAPNETZ network of excellence community-acquired pneumonia, Germany, CI confidence interval, HbA1c glycated hemoglobin, HR hazard ratio, OR odds ratio, p, p-value are statistically significant at p < 0.05 and proinflammatory genes can lead to chronic inflammation [117]. Homocysteine is built from the breakdown of the essential amino acid methionine and can subsequently be converted to cystathionine with vitamin B6 and further to cysteine. Conversely, vitamin B12 promotes remethylation of homocysteine to methionine [121,122]. The latter reaction is catalyzed by the enzyme methionine synthase, requiring 5-methyltetrahydrofolate, the main circulating form of folate, and vitamin B12 in its cofactor form methylcobalamin [122].
Current research ( Table 6) Table 6 outlines a clinical study that ascertained the status of glutathione-reflected by the reduced ratio of glutathione (GSH) to total glutathione in whole blood-as an independent mid-term predictor of all-cause mortality in post-ICU patients, whereas the total concentration of glutathione as well as the concentration of GSH alone revealed no association with 6-month mortality [118]. Although total glutathione and GSH concentrations in ICU patients have been observed as low, the quartile with the highest redox ratio of glutathione (GSH/total glutathione) carries the highest post-ICU all-cause mortality risk [118]. Lower levels of glutathione-S-bimane (GSB)-that is, intracellular GSH measured by fluorescence-activated cell sorter (FACS)-in CD4 T cells are prominent long-term mortality predictors in HIV-infected individuals, independent of CD4 T cell count [123].
Several clinical studies have demonstrated that elevated levels of plasma homocysteine independently predict long-term all-cause mortality in the elderly [124][125][126][127] as well as in younger adults [128,129]. In contrast, in patients after ischemic stroke a marked association is evident between high plasma homocysteine and overall mortality only after exclusion of patients with dissection [129]. Another study showed a correlation between higher homocysteine levels and increased long-term mortality in the subgroup of patients with malnutrition and inflammation [130]. Additionally, elevated plasma homocysteine levels are discernibly related to a higher death rate only in older women [126]. Besides their relation to all-cause mortality, high homocysteine concentrations are independently associated with cardiovascular mortality [128].

Glycerophospholipids
Pathway and physiological role Glycerophospholipids are glycerol-based phospholipids that form the bilayer in biological membranes. Their basic structure is composed of glycerol, to which a phosphoric acid and two fatty acids are attached as esters. An additional alcohol group (e.g., serine, choline, ethanolamine or inositol) attached to the phosphate allows the formation of distinct phosphoglycerides. The five classes of glycerophospholipids are phosphatidylcholine (PC, lecithin), phosphatidylethanolamine (PE), phosphatidylserine (PS), phosphatidylinositol (PI) and diphosphatidylgylcerol (cardiolipin) [131].

Current research (Table 7)
In lipid profiling of patients undergoing coronary angiography, 19 PC and 3 lysophosphatidylcholine (lysoPC) species reveal a noticeable association with mortality due to total and coronary artery disease. Ten of 19 PC and all 3 analyzed lysoPC species are inversely associated with mortality, whereas PC 32:0 reveals the strongest positive correlation with mortality [132]. Four lysoPC species-C16:0, C18:0, C18:1, and C18:2-account for over 90 % of total plasma lysoPC concentration. The levels of these main lysoPC species as well as total lysoPC concentrations are markedly reduced in sepsis, including pulmonary infections, compared to healthy controls (Table 7). Moreover, 30-day non-survivors show higher ratios of lysoPC/PC compared with survivors [133].

Sphingolipids
Pathway and physiological role Sphingolipids are essential components of cell membranes, and in neural tissue they are important in signal transduction and cell recognition. Fundamental building blocks of sphingolipids are long-chain bases (sphingoid bases). Their amine group is linked to an acyl group (e.g., fatty acid) and the backbone of the sphingoid base is linked with phosphate to an alcohol group (serine, ethanolamine or choline). The 3 principal forms of sphingolipids are ceramide and its derivatives sphingomyelin (SM) and glycosphingolipids [134].
Current research (Table 7) Elevated plasma SM levels are an independent risk factor for CHD and an outcome predictor in patients with acute coronary syndrome [135,136]. In lipid profiling of patients undergoing coronary angiography, 9 SM species are significantly associated with total and CHD long-term mortality (Table 7). Of 9 SM species, 5 showed an inverse association and 4 (specifically SM 16:0, 16:1, 24:1, and 24:2) displayed a positive association with total and cardiovascular mortality [132].

Acylcarnitines
Pathway and physiological role Carnitine (3-Hydroxy-4-N-trimethylammoniobutanoate) is biosynthesized in the liver and kidneys from the amino acids lysine and methionine. Nutrition is an important, but not essential, source of carnitine. The highest concentrations of carnitine are found in red meat, but carnitine is also found in other foods such as nuts, seeds, legumes, vegetables, fruits and cereals. Fatty acid oxidation and thus fatty acid energy metabolism occur in the cellular mitochondria. In the cytosol, free fatty acids are attached with a     19, p = 0.0001) quartiles were higher than the first quartile ACS acute coronary syndrome, CAD coronary artery disease, CI confidence interval, HR hazard ratio, LURIC Ludwigshafen Risk and Cardiovascular Health study, Germany, lysoPC lysophosphatidylcholine, OR odds ratio, p p-value are statistically significant at p < 0.05, PC phosphatidylcholine, SM sphingomyelin thioester bond to coenzyme A (CoA). Carnitine transports long-chain acyl groups of fatty acids from the cellular cytosol through the mitochondrial membrane. On the outer mitochondrial membrane, carnitine acyltransferase l transfers acyl-CoA to the hydroxyl group of carnitine, resulting in acylcarnitine. The enzyme carnitineacylcarnitine translocase catalyzes the transport of acylcarnitine to the inner mitochondrial membrane where carnitine acyltransferase II re-converts acylcarnitine to acyl-CoA [137].
Current research (Table8 ) Metabolic profiling of 165 polar metabolites was performed in hemodialysis patients. Individuals who died of a cardiovascular cause within 1 year of initiating hemodialysis revealed markedly higher concentrations of the 4 long-chain acylcarnitines oleoylcarnitine, linoleylcarnitine, palmitoylcarnitine, and stearoylcarnitine compared with survivors who had undergone hemodialysis for 1 year. Of these metabolites, oleoylcarnitine showed the strongest association with cardiovascular mortality after multivariable adjustment (Table 8) [138].

Conclusions
Since many uncertainties remain in predicting all-cause mortality in patients hospitalized for CAP or for exacerbated COPD, additional prospective and retrospective observational studies are needed to ascertain other potential contributors to all-cause mortality in order to improve the prediction of initial high risk. Metabolomic analysis using MS may facilitate identification of outcome-specific metabolite signatures that could potentially generate new metabolic biomarkers of various organic systems as all-cause mortality predictors in CAP and in exacerbated COPD. Improvement in early and accurate identification of high-risk patients can help optimize short-and long-term management strategies in LRTI and as a result, potentially improve survival, leading to a more cost-effective allocation of medical  ArMORR The Accelerated Mortality on Renal Replacement Study, United States, CI confidence interval, HD hemodialysis, ICD-9 International Classification of Diseases, OR odds ratio, p p-value are statistically significant at p < 0.05; SD, standard deviation resources. In addition, deeper knowledge of metabolic interactions in LRTI may provide new targets for future individual therapy and also be beneficial in predicting responses to therapy.