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.

Secondary infections in mechanically ventilated patients with COVID-19: An overlooked matter?

Objective: The susceptibility to infection probably increases in COVID-19 patients due to a combination of virusand drug-induced immunosuppression. The reported rate of secondary infections was quite low in previous studies. The objectives of our study were to investigate the rate of secondary infections, risk factors for secondary infections and risk factors for mortality in COVID-19 critically ill patients.

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.

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).

Noninvasive prenatal diagnosis by cell-free DNA screening for fetomaternal HPA-1a platelet incompatibility.

Background: The development of new noninvasive approaches for the diagnosis of human platelet antigen (HPA)-1 fetomaternal incompatibility has become of great interest. These approaches allow determination of whether the fetus is incompatible or not with the mother and a decision on antenatal therapy to avoid fetal or neonatal alloimmune thrombocytopenia (FNAIT). The objective of this work was to perform rapid, noninvasive prenatal test for HPA-1ab fetal antigen detection after the detection of an HPA-1-homozygous mother by using plasma cell-free DNA (cfDNA).

Evaluation of the State of Transplanted Liver Health by Monitoring of Organ-Specific Genomic Marker in Circulating DNA from Receptor.

The evaluation of the transplanted liver health by non-invasive approaches may offer an improvement in early clinical intervention. As transplanted organs have genomes that are distinct from the host's genome, the quantification of the specific DNA of the donated liver in the patient serum will allow us to obtain information about its damage. We evaluated the state of transplanted liver health by monitoring the RH gene in serum circulating DNA (cirDNA) from 17 recipient and donor mismatched for this gene.

Screening of KRAS Mutation in Pre- and Post-Surgery Serum of Patients Suffering from Colon Cancer by COLD-PCR HRM.

Genomic characterization of cell-free circulating tumour DNA (ctDNA) may offer an opportunity to assess clonal dynamics throughout the course of a patient's illness. The existence of KRAS driver mutations in colon cancer patients is determinant to decide their treatment and to predict their outcome. DNA is extracted automatically from 400 μL of serum using the MagNa Pure Compact with the Nucleic Acid Isolation Kit I.

Detection of p53 Mutations in Circulating DNA of Transplanted Hepatocellular Carcinoma Patients as a Biomarker of Tumor Recurrence.

p53 is the most commonly mutated gene in malignant human cancers. To detect p53 mutations in circulating DNA (cirDNA) of transplanted hepatocellular carcinoma (HCC) patients could be an interesting approach to know of any tumor recurrence. In this study, our objective was to determine the utility of this method in the diagnosis and the prognosis of HCC tumor recurrence.

Role of early cell-free DNA levels decrease as a predictive marker of fatal outcome after severe traumatic brain injury.

Introduction: Circulating cell-free DNA levels are increased after trauma injury. This increase is higher since the first hours after trauma and may be related with primary outcome. A sensitive and reliable biomarker for patients at higher risk is needed to identify these patients to initiate early intervention.

Blocking of melatonin synthesis and MT(1) receptor impairs the activation of Jurkat T cells.

Melatonin has been proposed as regulating the immune system by affecting cytokine production in immunocompetent cells, enhancing the production of several T helper (Th)1 cytokines. To further investigate the melatonin's role in IL-2/IL-2R system, we established an inducible T-REx expression system in Jurkat cells in which the protein levels of HIOMT enzyme or MT(1) receptor were significantly down-regulated upon tetracycline incubation. We found that T-REx Jurkat cells with lower levels of HIOMT activity, and consequently lower content of endogenous melatonin, showed IL-2 production decrease after activation with lectin.

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.

A novel interplay between membrane and nuclear melatonin receptors in human lymphocytes: significance in IL-2 production.

Human lymphocyte melatonin, through membrane and nuclear receptors binding, acts as an activator in IL-2 production. Antagonism of membrane melatonin receptors using luzindole exacerbates the drop of the IL-2 production induced by PGE(2) in peripheral blood mononuclear and Jurkat cells. This paper studies the melatonin membrane and nuclear receptors interplay in PGE(2)-diminished IL-2 production.

Treatment with testosterone or estradiol in melatonin treated females and males MRL/MpJ-Faslpr mice induces negative effects in developing systemic lupus erythematosus.

MRL/MpJ-Fas(lpr) mice is widely accepted as a valuable model of systemic lupus erythematosus. As described in a previous work, the incidence of lupus in this strain is determined by sex hormones, i.e.

Evidence for melatonin synthesis in the rat brain during development.

Melatonin production is not restricted to the pineal gland. Several extrapineal sources of this indole such as retina, Harderian gland, and immune system are well documented. Melatonin of pineal origin is not present in the rat at early stages of development.

Melatonin biosynthesis in the thymus of humans and rats.

Melatonin is an indoleamine widely distributed in the evolution that shows a great functional versatility, playing an important role as a transmitter of photoperiodic information and exhibiting antioxidant, oncostatic, anti-aging and immunomodulatory properties. In vertebrates, this molecule is produced by the pineal gland and other extrapineal sites. The present study was carried out to investigate the presence of melatonin in thymus and the possibility of an endogenous melatonin synthesis in this organ, in which T cells are matured.

Sex-dependent effect of melatonin on systemic erythematosus lupus developed in Mrl/Mpj-Faslpr mice: it ameliorates the disease course in females, whereas it exacerbates it in males.

In this study, the effect of chronic administration of melatonin on MRL/MpJ-Fas(lpr) mice has been studied. These mice spontaneously develop an autoimmune disease that has many features resembling human systemic lupus erythematosus. In fact, histological studies showed that all female mice and most male mice exhibited glomerular abnormalities, arteritic lesions, and cellular interstitial inflammatory infiltrate ranging from mild to severe patterns.

Melatonin prevents hyperhomocysteinemia and neural lipid peroxidation induced by methionine intake.

This study was designed to determine the protective effect of melatonin treatment against oxidative damage in rat brain induced by hyperhomocysteinemia (Hhcy). Oral administration of methionine and its degradation product, homocysteine (hcy), causes mild to moderate Hhcy. The major end-point of oxidative damage measured in this report was lipid peroxidation (LPO).

Melatonin synthesis and melatonin-membrane receptor (MT1) expression during rat thymus development: role of the pineal gland.

To gain insight into the relationship between thymus and pineal gland during rat development, the melatonin content as well as the activity and expression of the two key enzymes for melatonin biosynthesis, i.e. N-acetyltransferase (NAT) and hydroxyindole-O-methyltransferase (HIOMT), were studied in the thymus at fetal and postnatal stages.

Dual effect of melatonin as proinflammatory and antioxidant in collagen-induced arthritis in rats.

The aim of this study was to determine the effects of melatonin on proinflammatory status of rats with collagen-induced arthritis (CIA). CIA was induced in male Wistar rats with an emulsion of type II collagen in Freund's Incomplete Adjuvant (C-II/FIA). For 14 days, control and pinealectomized rats received a subcutaneous injection of 100 microL melatonin (30 microg) or vehicle (saline on 1% ethanol).

Involvement of nuclear receptors in the enhanced IL-2 production by melatonin in Jurkat cells.

This report shows that melatonin enhances IL-2 production by Jurkat cells via a nuclear receptor-mediated mechanism. Jurkat cells express nuclear (RZR alpha, ROR alpha 1, and ROR alpha 2) and membrane (mt1) melatonin receptors, and melatonin binds to Jurkat nuclei and membranes with the same affinity described for human peripheral blood mononuclear cells (PBMCs). Melatonin enhances IL-2 production by Jurkat cells activated by either phytohemagglutinin (PHA) or phorbol myristate acetate (PMA).

Nuclear receptors are involved in the enhanced IL-6 production by melatonin in U937 cells.

This report shows that melatonin enhances IL-6 production by U937 cells via a nuclear receptor-mediated mechanism. Resting U937 cells only express membrane (mt1) melatonin receptors. In these cells, melatonin did not modify basal production of IL-6 or when activated by PMA plus lipopolysaccharide, a treatment that downregulates the expression of mt1 receptor.

Melatonin is responsible for the nocturnal increase observed in serum and thymus of thymosin alpha1 and thymulin concentrations: observations in rats and humans.

This paper shows that melatonin regulates both thymosin alpha1 and thymulin production as well as the expression of the prothymosin alpha gene. The results revealed the following facts: (a) The concentrations of thymosin alpha1 in both serum and thymus of rat showed a nyctohemeral profile with peak values late at night and basal values during the day. The concentrations of thymulin in rat serum also showed a 24-h rhythm with an increase in their values at night.

Circadian variations in the rat serum total antioxidant status: correlation with melatonin levels.

In this paper, we show for the first time, a nyctohemeral rhythm in serum total antioxidant status (TAS) in rats which parallels the 24-H melatonin cycle. Both TAS and melatonin in rat serum exhibited 24 hr variations with nocturnal peak values at 05.00 hr and low basal values during the day.