Validation of S100B use in a cohort of Spanish patients with mild traumatic brain injury: a multicentre study.

INTRODUCTION: The aim of this study was to validate the S100B protein as a diagnostic tool for ruling out the presence of intracranial lesion (IL) after mild traumatic brain injury (mTBI). Subjects with a Glasgow Coma Scale (GCS) score of 15 and at least one neurological symptom post-trauma were selected from a large Spanish cohort. METHODS: A number of 260 patients with mTBI were enrolled. Blood samples were extracted within 6 h and CT scan performed within 24 h post-injury. Blood samples were also drawn from 18 healthy subjects. RESULTS: CT scan revealed the presence of IL in 22 patients (8.5%). Patients with mTBI had higher S100B serum levels (p = 0.008) than the healthy subjects (p < 0.001). The ROC analysis of S100B discriminated between patients with and without IL (AUC: 0.671; 95%CI: 0.574-0.769; p = 0.008). The multivariate analysis identified male gender (OR: 5.39; 95%CI: 1.45-20.10; p = 0.012), age > 65 (OR: 2.97; 95%CI: 1.04-8.44; p = 0.041) and S100B level >0.10 µg/L (OR: 7.93; 95%CI: 1.03-60.76; p = 0.046) as independent risk factors for IL in patients with mTBI. CONCLUSION: Measurement of S100B within 6 h of mTBI accurately predicts risk of IL in patients with a GCS score of 15 and at least one neurological symptom.

LDH/ADA ratio in pleural fluid for the diagnosis of infectious pleurisy.

Pleural effusion (PE) is a common medical concern, often requiring thoracentesis for a definitive diagnosis. An elevated pleural fluid adenosine deaminase (ADA) may indicate tuberculosis, but this is not always the case. This study aimed to evaluate the accuracy of biomarkers determined in pleural fluid and propose a new diagnostic strategy for PE in patients with high levels of ADA in pleural fluid. This retrospective analysis studied patients with PE who received thoracentesis for the first time with an ADA level of > 33 U/L in the pleural fluid analysis at two tertiary hospitals from March 2019 to March 2023. Demographic and clinical data, as well as pleural fluid biomarkers and their ratios, were studied and compared between different PE groups, and a decision tree was developed. During the study period, 259 patients were enrolled, with four different types of PE: parapneumonic (PPE) 155, tuberculosis (TPE) 41, malignant (MPE) 50, and miscellaneous 13. Biomarkers and their ratios performed well in the differential diagnosis of PE, with the LDH/ADA ratio distinguishing between PPE and non-PPE with sensitivity and specificity of 98.06% and 98.08%, respectively. The combination of LDH/ADA ratio, ADA, and mononuclear cell percentage was identified as important factors for creating a decision tree with an overall accuracy of 89.96%. The pleural fluid LDH/ADA ratio was a useful diagnostic for distinguishing PPE from non-PPE, and a decision tree with an accuracy of 89.96% was created to differentiate the four forms of PE in clinical situations.

Donor-Specific Cell-Free DNA qPCR Quantification as a Noninvasive Accurate Biomarker for Early Rejection Detection in Liver Transplantation.

(1) Background: Graft-cell-free DNA (cfDNA) in the circulation of liver transplant recipients has been proposed as a noninvasive biomarker of organ rejection. The aim of this study was to detect donor-specific cfDNA (ds-cfDNA) in the recipient's serum after either liver damage or rejection using a qPCR-based method. (2) Methods: We proposed a qPCR method based on the amplification of 10 specific insertion-deletion (InDel) polymorphisms to detect donor-specific circulating DNA diluted in the recipient cfDNA. ds-cfDNA from 67 patients was evaluated during the first month post-transplantation. (3) Results: Graft rejection in the first month post-transplantation was reported in 13 patients. Patients without liver complications showed a transitory increase in ds-cfDNA levels at transplantation. Patients with rejection showed significant differences in ds-cfDNA increase over basal levels at both the rejection time point and several days before rejection. Receiver operator characteristic (ROC) analysis showed that ds-cfDNA levels discriminated rejection, with an AUC of 0.96. Maximizing both sensitivity and specificity, a threshold cutoff of 8.6% provided an estimated positive and negative predictive value of 99% and 60%, respectively. (4) Conclusions: These results suggest that ds-cfDNA may be a useful marker of graft integrity in liver transplant patients to screen for rejection and liver damage.

GPX3 Overexpression in Cumulus Cells Entails a Poor Prognosis for Uterine Implantation of Morphotype A Embryos.

Morphological embryo quality is an accurate prognostic tool for the success of assisted reproduction implantation, although complete certainty cannot be guaranteed. The transcriptome of the cumulus cells could be monitored as a faithful reflex of the physiological state of the oocytes, given the molecular crosstalk between both types of cells. Here, we compare the expression of specific genes related to oocyte competence, such as hyaluronic acid synthase 2 (HAS2), cell division control protein 42 (CDC42), connexin 43 (CX43), and glutathione peroxidase 3 (GPX3), in cumulus cells from implanted versus non-implanted embryos in 25 women, using RT-qPCR. After embryo transfer, two cohorts were differentiated: the pregnant group (women with the implantation of 100% of embryos transferred) versus the non-pregnant group (with an absence of embryo implantation), aiming to compare the possible differential expression of the selected genes in the cumulus cells of embryos from each group. HAS2, CDC42 and CX43 did not reveal differential expression between the two cohorts. However, GPX3 showed significantly reduced expression in the cumulus belonging to the pregnant group. Interestingly, even cumulus cells belonging only to morphotype A embryos showed a significantly lower expression of GPX3 in the pregnancy group. GPX3 overexpression in cumulus cells could be a poor prognostic indicator of implantation, discriminating beyond the capacity of the morphokinetic score. Unveiling the cumulus transcriptome could improve successful implantation in assisted reproduction treatments.

Safety and Efficacy of a Beverage Containing Lupine Protein Hydrolysates on the Immune, Oxidative and Lipid Status in Healthy Subjects: An Intervention Study (the Lupine-1 Trial).

SCOPE: We have previously demonstrated the anti-inflammatory and antioxidant properties of in vitro administered Lupinus angustifolius protein hydrolysates (LPHs) on human peripheral blood mononuclear cells (PBMCs). This study aims to evaluate the safety and efficacy of a beverage containing LPHs (LPHb) on the immune, oxidative and metabolic status of healthy subjects. METHODS AND RESULTS: In this open-label intervention, 33 participants daily ingest a LPHb containing 1 g LPHs for 28 days. Biochemical parameters are assayed in fasting peripheral blood and urine samples before, during (14 days) and after LPHb ingestion. Participants' health status and the immune and antioxidant responses of PBMCs are also evaluated throughout the trial. The LPHb ingestion is safe and effective in both increasing the anti-/pro-inflammatory response of PBMCs and improving the cellular anti-oxidant capacity. LPHb also reduces the low-density lipoprotein-cholesterol (LDL-C)/high-density lipoprotein-cholesterol (HDL-C) atherogenic index. LPHb effect is particularly beneficial on decreasing not only the LDL-C/HDL-C index but also serum total cholesterol levels in the male cohort that shows the highest baseline levels of well-known cardiovascular risk factors. CONCLUSION: This is the first study to show the pleiotropic actions of a lupine bioactive peptides-based functional food on key steps of atherosclerosis including inflammation, oxidative stress, and cholesterol metabolism.

Lupinus angustifolius Protein Hydrolysates Reduce Abdominal Adiposity and Ameliorate Metabolic Associated Fatty Liver Disease (MAFLD) in Western Diet Fed-ApoE-/- Mice.

Metabolic-associated fatty liver disease (MAFLD) is the most important cause of liver disease worldwide. It is characterized by the accumulation of fat in the liver and is closely associated with abdominal obesity. In addition, oxidative stress and inflammation are significant features involved in MAFLD. Recently, our group demonstrated that lupin protein hydrolysates (LPHs) had lipid lowering, antioxidant, and anti-inflammatory effects. Sixty male mice fed with a Western diet were intragastrically treated with LPHs (or vehicle) for 12 weeks. Liver and adipose tissue lipid accumulation and hepatic inflammatory and oxidant status were evaluated. A significant decrease in steatosis was observed in LPHs-treated mice, which presented a decreased gene expression of CD36 and LDL-R, crucial markers in MAFLD. In addition, LPHs increased the hepatic total antioxidant capacity and reduced the hepatic inflammatory status. Moreover, LPHs-treated mice showed a significant reduction in abdominal adiposity. This is the first study to show that the supplementation with LPHs markedly ameliorates the generation of the steatotic liver caused by the intake of a Western diet and reduces abdominal obesity in ApoE-/- mice. Future clinical trials should shed light on the effects of LPHs on MAFLD.

Immunomodulatory and Antioxidant Properties of Wheat Gluten Protein Hydrolysates in Human Peripheral Blood Mononuclear Cells.

Peptides from several plant food proteins not only maintain the nutritional values of the original protein and decrease the environmental impact of animal agriculture, but also exert biological activities with significant health-beneficial effects. Wheat is the most important food grain source in the world. However, negative attention on wheat-based products has arose due to the role of gluten in celiac disease. A controlled enzymatic hydrolysis could reduce the antigenicity of wheat gluten protein hydrolysates (WGPHs). Therefore, the aims of the present study were to evaluate the effects of the in vitro administration of Alcalase-generated WGPHs on the immunological and antioxidant responses of human peripheral blood mononuclear cells (PBMCs) from 39 healthy subjects. WGPH treatment reduced cell proliferation and the production of the Type 1 T helper (Th1) and Th17 pro-inflammatory cytokines IFN-γ and IL-17, respectively. WPGHs also improved the cellular anti-inflammatory microenvironment, increasing Th2/Th1 and Th2/Th17 balances. Additionally, WGPHs improved global antioxidant capacity, increased levels of the reduced form of glutathione and reduced nitric oxide production. These findings, not previously reported, highlight the beneficial capacity of these vegetable protein hydrolysates, which might represent an effective alternative in functional food generation.

Homocysteine and C-Reactive Protein Levels Are Associated With Frailty in Older Spaniards: The Toledo Study for Healthy Aging.

High-sensitivity C-reactive protein (hsCRP) and homocysteine (Hcy) are inflammation markers but are also related to cardiovascular diseases, disability, or higher risk of death. Although inflammation is considered to be associated with frailty, data regarding the association between hsCRP or Hcy and frailty are controversial or scarce, especially with respect to their association with prefrailty. Thus, our objective was to study the association of hsCRP and Hcy with prefrailty and frailty in 1,211 Spanish men and women aged 65-98 years from the Toledo Study for Healthy Aging (TSHA) cohort, classified according to Fried's criteria. Hcy was independently associated with frailty (odds ratio [OR] = 1.06; 95% confidence interval [CI]: 1.01-1.12), whereas hsCRP was independently associated with both prefrailty (OR = 1.03; 95% CI: 1.01-1.06) and frailty (OR = 1.07; 95% CI: 1.02-1.12). Furthermore, both markers were positively correlated with the number of Fried's criteria that were met and were independently associated with the criteria of exhaustion (Hcy: OR = 1.03, 95% CI: 1.00-1.06), weakness (hsCRP: OR = 1.03, 95% CI: 1.01-1.05), and low physical activity (hsCRP: OR = 1.04, 95% CI: 1.02-1.06). Thus, our results highlight the importance of inflammation in age-related physical decline and, in particular, its association with fatigue, low strength, and decreased physical activity.

Evidence of immune system melatonin production by two pineal melatonin deficient mice, C57BL/6 and Swiss strains.

We evaluated two pineal melatonin deficient mice described in the literature, i.e., C57BL/6 and Swiss mice, as animal models for studying the immunomodulatory action of melatonin. Plasma melatonin levels in C57BL/6 and Swiss strains were detectable, but lower than levels in control C3H/HENHSD mice. Since these strains are suppose to be pineal melatonin deficient an extrapineal melatonin synthesis may contribute to plasma levels. Regarding cells and tissues from the immune system, all of them were found to synthesize melatonin although at low levels. N-acetyltransferase (AANAT) mRNA was also amplified in order to analyze the alternative splicing between exons 3-4 described for pineal C57BL/6 mice which generates an inclusion of a pseudoexon of 102 bp. For the pineal gland, both the wild type and the mutant isoforms were present in all mice strains although in different proportions. We observed a predominant wild type AANAT mature RNA in thymus, spleen and bone marrow cells. Peripheral blood mononuclear cells (PBMC) culture shown an evident AANAT amplification in all strains studied. Although the bands detected were less intense in melatonin deficient mice, the amplification almost reached the control cell intensity after stimulation with phytohemaglutinin (PHA). In summary, melatonin detection and AANAT mRNA expression in inbred and outbred mice clearly indicate that different cells and tissues from the immune system are able to synthesize melatonin. Thus, the pineal defect seems not to be generalized to all tissues, suggesting that other cells may compensate the low pineal melatonin production contributing to the measurable plasma melatonin level.

Peripheral CD39-expressing T regulatory cells are increased and associated with relapsing-remitting multiple sclerosis in relapsing patients.

CD39, an ectonucleotidase that hydrolyses pro-inflammatory ATP, is a marker of highly active and suppressive T regulatory cells (Tregs). Although CD39 has a role in Treg suppression and might be important in the control of neuroinflammation in relapsing-remitting multiple sclerosis (RR-MS), to date, there are contradictory reports concerning the Tregs expression of CD39 in RR-MS patients. Thus, our objectives were to assess the activity and expression of CD39, especially in Tregs from peripheral blood mononuclear cells (PBMCs) of relapsing RR-MS patients compared with control subjects and to evaluate the association of CD39+ Tregs with disability and the odds of RR-MS. The activity and expression of CD39 and the CD39+ Treg frequency were measured in PBMCs from 55 relapsing RR-MS patients (19 untreated and 36 receiving immunomodulatory treatment) and 55 age- and sex-paired controls. Moreover, the association between CD39+ Tregs and RR-MS was assessed by multivariate logistic regression. CD39 activity and the frequency of CD39-expressing Tregs were elevated in relapsing RR-MS patients. Moreover, CD39+ Tregs were significantly correlated with the EDSS score and were independently associated with the odds of RR-MS. Our results highlight the relevance of CD39+ Treg subset in the clinical outcomes of RR-MS.

Lupine protein hydrolysates decrease the inflammatory response and improve the oxidative status in human peripheral lymphocytes.

Although cell-free systems and immortalized cell lines have been used to demonstrate the potential health benefits of lupine proteins and peptides, no study has examined the effects of lupine protein hydrolysates (LPHs) on the immune and oxidative responses of non-immortalized human cells. Therefore, the aims of this study were to evaluate the effects of the in vitro administration of LPHs from Lupinus angustifolius on the immunological and oxidative statuses of human peripheral blood mononuclear cells (PBMCs) from 53 healthy donors. LPHs reduced PBMCs proliferation and the levels of Th1, Th9 and Th17 pro-inflammatory cytokines without being cytotoxic. LPHs also skewed the pro-/anti-inflammatory balance towards a Th2 protective response. Additionally, LPHs increased superoxide dismutase and catalase activities, and the total antioxidant capacity (TAC). This study is the first to show that LPHs reduce T cell inflammatory responses and improve the anti-inflammatory/pro-inflammatory cytokine balance and the TAC by PBMCs. Thus, LPHs may represent an effective option for developing nutritional strategies to prevent pathologies with underlying inflammation and oxidative stress.

Temporal expression patterns of the melatoninergic system in the human thymus of children.

OBJECTIVES: To obtain greater knowledge of the extra-pineal sources of melatonin during development, the amount of indolamine and the expression levels of the last two enzymes involved in its biosynthesis, Arylalkylamine N-acetyltransferase (AANAT) and acetylserotonin O-methyltransferase (ASMT), were analyzed in the human thymus from children from three different age groups (from days to years). The melatonin membrane and nuclear receptor expression levels also were studied. METHODS: Quantitative reverse transcriptase PCR and western blot were performed to investigate the receptor and enzyme expression levels. The results were examined and correlated with the ages of the thymuses. RESULTS: We found high levels of indolamine in the thymuses of newborns (younger than 1 month), which decreased during development; thymuses from the months (from 2 to 11 months) and years (from 1 to 12 years) groups showed lower levels. A similar decline was also observed in the mRNA of the AANAT enzyme and the expression levels of melatonin receptors. However, ASMT expression was exactly the opposite, with low levels in the newborn group and higher levels in the years group. Our results show that the thymic synthesis of melatonin occurs very early in childhood. Additionally, this is the first report that is focused on melatonin receptors expression in the human thymus. CONCLUSION: Considering the limited melatonin synthesis performed by the newborn pineal gland, we suggest that the high levels of melatonin found in human thymus in this experimental group arise from synthesis in the tissue itself, which could be contributing to the immune efficiency at the thymic level.

Donor-specific circulating cell free DNA as a noninvasive biomarker of graft injury in heart transplantation.

BACKGROUND: Considerable effort has been exerted to develop noninvasive diagnostic biomarkers that might replace or reduce the need to perform endomyocardial biopsies. In this context, graft DNA circulating on transplant recipients has been proposed as a potential biomarker of organ rejection or cellular graft injury. METHODS: We propose a digital PCR (dPCR) method based on the amplification of ten specific InDels sufficiently sensitive to detect small amounts of specific donor circulating DNA diluted on the host cell free DNA (cfDNA). We obtained 23 informative mismatches from 30 host and donor organ biopsy pairs. RESULTS: Patients without heart-related complications showed a high increase in the specific genomic marker levels during the first 24 h after transplantation that dropped to the basal levels on days 3-4 post-surgery. In contrast, patients with complications presented a significantly lagged decay pattern from day one after transplantation. A specific donor cfDNA increase was detected in one patient two days before rejection diagnosis, diminishing the basal levels after successful immunotherapy. A cfDNA increase was also observed during graft injury due to heart damage. CONCLUSION: These results suggest that cfDNA monitoring of transplanted patients may be a useful tool to detect and probably anticipate graft rejection.

[Treatment indication with endothoracic drainage tube in parapneumonic effusions by partial pressure of carbon dioxide measurement in pleural fluid]. / Indicación de tratamiento con tubo de drenaje endotorácico en derrames pleurales paraneumónicos, según la presión parcial de anhídrido carbónico de líquido pleural.

BACKGROUND AND OBJECTIVE: Parapneumonic effusions with pH < 7.20 or glucose < 0.40 g/l or lactate dehydrogenase (LDH) > 1000 U/l have indication of treatment with endothoracic drainage tube (EDT). The aim of the present study was to determine the accuracy of partial pressure of carbon dioxide (pCO2) measurement in pleural fluid for the subsequent treatment indication with EDT in parapneumonic effusions, by analyzing the area under curve ROC (AUC) and determining the optimal cut off value. PATIENTS AND METHOD: 207 pleural fluids were studied. Glucose, LDH, pCO2, and pH were measured, and data concerning the etiology of pleural effusion and whether EDT treatment was needed were collected after patients were discharged from hospital. RESULTS: Forty-six out of 207 pleural fluids studied were parapneumonic effusions. Thirty-two were treated with EDT. AUC values were 0.888 (p < 0.0001), 0.890 (p < 0.0001), 0.816 (p < 0.0001), and 0.801 (p < 0.0001) for pCO2, pH, glucose, and LDH, respectively. No significant differences were found among them. Optimal cut off value for pCO2 was 48.6 mmHg, exhibiting 90.6% sensitivity and 78.6% specificity. All parapneumonic effusions showing pCO2 > 60.9 mmHg were treated with EDT. Remarkably, 3 out of 46 parapneumonic effusions (6.5%) that had been improperly treated following pH, glucose or LDH values, were correctly treated following pCO2. CONCLUSIONS: pCO2 determination in pleural fluid appears to be the best way to decide the indication of EDT in parapneumonic effusions.

Homocysteine levels are associated with bone resorption in pre-frail and frail Spanish women: The Toledo Study for Healthy Aging.

BACKGROUND: Homocysteine (Hcy) high levels are associated with fractures, bone resorption and an early onset of osteoporosis in elderly persons; a relationship between Hcy and bone formation has also been suggested but is still controversial. Frailty, an independent predictor of fractures and decreased bone mineral density is associated with altered bone metabolism in women. However, no previous works have studied the relationship among frailty, Hcy levels and bone turnover. METHODS: We studied the association among Hcy, osteoporosis and N-terminal propeptide of type I procollagen (PINP), C-terminal telopeptide of type I collagen (ß-CTX), parathyroid hormone (PTH), calcium and 25-hydroxyvitamin D (25(OH)D) in 631 Spanish women between the ages of 65-78 from the Toledo Study for Healthy Aging (TSHA) cohort, who were classified as highly functional (robust subjects) or non-robust (pre-frail or frail subjects) according to Fried's criteria. RESULTS: Hcy was independently associated with ß-CTX in the entire population (B = 0.22; 95% CI, 0.09-0.34; p = 0.001) and in the non-robust group (B = 0.24; 95% CI, 0.09-0.39; p = 0.002). Hcy was also associated with PINP in the entire and non-robust populations, but the association was lost after including the levels of ß-CTX, but not the other bone biomarkers, in the multivariate analysis. This suggests that the controversial relationship between Hcy and bone formation might be explained, at least to a certain extent, by the confounding effects of ß-CTX. CONCLUSIONS: This work highlights the important implication of frailty status in the association between Hcy and increased bone turnover in older women.

The modulatory role of melatonin on immune responsiveness.

Afterhe successful discovery of the melatonin molecule by Aaron B Lerner et al at Yale University in 1958, melatonin and the pineal gland, a tiny endocrine gland situated at the center of the human brain, have primarily been considered in terms of their effects on the endocrine and reproductive systems. During the last decade, a substantial body of research has defined melatonin as a remarkable molecule with pleiotropic effects on the immune system. Moreover, its synthesis cannot be considered as exclusively endocrine; key immunocompetent cells have the functional enzymatic machinery for melatonin synthesis, paving the wayfo r complex intracrine, autocrine and paracrine regulatory loops. The immunomodulatory role of melatonin, with regard to infection, inflammation and autoimmunity, is outlined here, and the evidence discussed in this review strengthens the notion that the nature of an immune response may be modified, and therefore therapeutically manipulated, by circadian effector signals.

S100B and Neuron-Specific Enolase as mortality predictors in patients with severe traumatic brain injury.

OBJECTIVE: To determine temporal profile and prognostic ability of S100B protein and neuron-specific enolase (NSE) for prediction of short/long-term mortality in patients suffering from severe traumatic brain injury (sTBI). METHODS: Ninety-nine patients with sTBI were included in the study. Blood samples were drawn on admission and on subsequent 24, 48, 72, and 96 h. RESULTS: 15.2% of patients died in NeuroCritical Care Unit, and 19.2% died within 6 months of the accident. S100B concentrations were significantly higher in patients who died compared to survivors. NSE levels were different between groups just at 48 h. In the survival group, S100B levels decreased from 1st to 5th sample (p < 0.001); NSE just from 1st to 3rd (p < 0.001) and then stabilized. Values of S100B and NSE in non-survival patients did not significantly vary over the four days post sTBI. ROC-analysis showed that all S100B samples were useful tools for predicting mortality, the best the 72 h sample (AUC 0.848 for discharge mortality, 0.855 for six-month mortality). NSE ROC-analysis indicated that just the 48-h sample predicted mortality (AUC 0.733 for discharge mortality, 0.720 for six-month mortality). CONCLUSION: S100B protein showed higher prognostic capacity than NSE to predict short/long-term mortality in sTBI patients.

[Impact of thymic function in age-related immune deterioration]. / Impacto de la función tímica en el deterioro inmunológico asociado a la edad.

Age-related biological deterioration also includes immune system deterioration and, in consequence, a rise in the incidence and prevalence of infections and cancers, as well as low responses to vaccination strategies. Out of all immune cell subsets, T-lymphocytes seem to be involved in most of the age-related defects. Since T-lymphocytes mature during their passage through the thymus, and the thymus shows an age-related process of atrophy, thymic regression has been proposed as the triggering event of this immune deterioration in elderly people. Historically, it has been accepted that the young thymus sets the T-lymphocyte repertoire during the childhood, whereupon atrophy begins until the elderly thymus is a non-functional evolutionary trace. However, a rising body of knowledge points toward the thymus functioning during adulthood. In the elderly, higher thymic function is associated with a younger immune system, while thymic function failure is associated with all-cause mortality. Therefore, any new strategy focused on the improvement of the elderly quality of life, especially those trying to influence the immune system, should take into account, together with peripheral homeostasis, thymus function as a key element in slowing down age-related decline.