Characterization of serotonin transporter expression in human T lymphocytes

Serotonin transporter (SERT) expression has been demonstrated in human lymphocytes, including B lymphocytes, NK cells and other immune cells. However, discussion remains on whether human T lymphocytes express SERT. Given the potentially important role of serotonin in lymphocyte activation and proliferation, we investigated SERT expression in purified human T lymphocytes both in resting and activated state. Blood samples were collected from 9 healthy volunteers. PBMCs were isolated using Ficoll density centrifugation and T lymphocytes were further purified with magnetic activated cell sorting. T cells were either processed for mRNA and protein isolation immediately, or activated using anti-CD3/CD28 coated magnetic beads and allowed to proliferate for 72h at 37°C and 5% CO2. SERT mRNA expression was measured using droplet digital PCR to allow for increased sensitivity in comparison with qRT-PCR. SERT protein was detected on western blot. SERT expression was detected both on mRNA and protein level, although expression levels were very low. On mRNA level, SERT was expressed in both resting and activated cells. On the protein level however, only activated cells displayed SERT expression. This observation might point to a ‘translational readiness’ were resting T lymphocytes already produce SERT mRNA, but translation is only induced after activation of the cell

Fluoxetine reduces murine graft-versus-host disease by induction of T cell immunosuppression

Serotonin reuptake inhibitors (SRIs) are widely used drugs in the treatment of depression and anxiety disorders. Although SRIs are generally regarded as safe drugs with relatively few side effects, literature suggests that high concentrations of SRIs may alter immune function. We investigated whether high-dose treatment with fluoxetine was able to suppress acute graft-versus-host disease (GvHD) in a MHC-matched, minor histocompatibility antigen mismatched murine bone marrow transplantation model. We found that high doses fluoxetine induce a significant reduction of clinical symptoms and increase survival of these animals. The amelioration of clinical GvHD was accompanied by a reduced expansion of alloreactive T cells. We further analyzed the direct in vitro effect of six SRIs on the viability and proliferation of human T cells and found an anti-proliferative and pro-apoptotic effect that was significantly larger in activated than in resting T cells. We discuss these results in the light of potential future exploration of SRIs as a novel class of T cell immunosuppressive drugs

Fluoxetine suppresses calcium signaling in human T lymphocytes through depletion of intracellular calcium stores

Selective serotonin reuptake inhibitors, such as fluoxetine, have recently been shown to exert anti-inflammatory and immunosuppressive effects. Although the effects on cytokine secretion, proliferation and viability of T lymphocytes have been extensively characterized, little is known about the mechanism behind these effects. It is well known that Ca2+ signaling is an important step in the signaling transduction pathway following T cell receptor activation. Therefore, we investigated if fluoxetine interferes with Ca2+ signaling in Jurkat T lymphocytes. Fluoxetine was found to suppress Ca2+ signaling in response to T cell receptor activation. Moreover, fluoxetine was found to deplete intracellular Ca2+ stores, thereby leaving less Ca2+ available for release upon IP3- and ryanodine-receptor activation. The Ca2+-modifying effects of fluoxetine are not related to its capability to block the serotonin transporter, as even a large excess of 5HT did not abolish the effects. In conclusion, these data show that fluoxetine decreases IP3- and ryanodine-receptor mediated Ca2+ release in Jurkat T lymphocytes, an effect likely to be at the basis of the observed immunosuppression

Detailed method description for noninvasive monitoring of differentiation status of human embryonic stem cells

The (non)differentiation status of human embryonic stem cells (hESCs) is usually analyzed by determination of key pluripotency defining markers (e.g., OCT4, Nanog, SOX2) by means of reverse transcription quantitative polymerase chain reaction (RT-qPCR), flow cytometry (FC), and immunostaining. Despite proven usefulness of these techniques, their destructive nature makes it impossible to follow up on the same hESC colonies for several days, leading to a loss of information. In 2003, an OCT4-eGFP knock-in hESC line to monitor OCT4 expression was developed and commercialized. However, to the best of our knowledge, the use of fluorescence microscopy (FM) for monitoring the OCT4-eGFP expression of these cells without sacrificing them has not been described to date. Here, we describe such a method in detail, emphasizing both its resolving power and its complementary nature to FC as well as the potential pitfalls in standardizing the output of the FM measurements. The potential of the method is demonstrated by comparison of hESCs cultured in several conditions, both feeder free (vitronectin, VN) and grown on feeder cells (mouse embryonic fibroblasts, MEFs)

Fluoxetine onderdrukt graft-versus-hostziekte bij muizen door T-celimmunosuppressie

samenvatting van: Gobin V, Van Steendam K, Fevery S, Tilleman K, Billiau AD, Denys D, Deforce DI. Fluoxetine reduces murine graft-versus-host disease by induction of T cell immunosupression. J Neuroimmune Pharmacol 2013; doi: 10.1007/s11481-013-9463-

Selective serotonin reuptake inhibitors as a novel class of immunosuppressants

In the past decades, selective serotonin reuptake inhibitors (SSRIs) have been shown to exert several immunological effects, such as reduced lymphocyte proliferation, alteration of cytokine secretion and induction of apoptosis. Based on these effects, SSRIs were proposed as drugs for the treatment of autoimmune pathologies and graft-versus-host disease. This review summarizes preclinical and clinical evidence supporting a role for SSRIs in autoimmune diseases and graft-versus-host disease, and discusses what is known about the mechanism underlying these effects

Does shifting from conventional to zero tillage in combination with a cover crop offers opportunities for silage maize cultivation in Flanders?

Tillage is an important agricultural practice, influencing the physical, chemical and biological soil characteristics. In this paper the influence of various tillage systems combined with or without a cover crop under different nitrogen fertilization levels on silage maize yield and soil fertility was investigated. Based on a field trial in Bottelare (Belgium), during the period 2007-2015, it was concluded that for each tillage system higher nitrogen levels resulted in a higher yield. In addition, the highest yield was achieved for the conventional tillage system, the yield gain for mouldboard ploughing varied between 13% (2015) and 71% (2012) compared to zero tillage. In case reduced tillage was adopted, the yield loss compared to mouldboard ploughing varied between 6% (2013 and 2015) and 24% (2012). Furthermore, it seemed that the accumulated temperature during the growing season and rainfall around flowering were decisive in determining maize yield. Additionally, rainfall in the period 60 days post sowing was significantly negatively correlated with the yield from the zero tillage plots, whereas in case tillage was adopted no correlations with rainfall 60 days post sowing were detected. Concerning the soil organic carbon content and the amount of earthworms, no clear trends could be observed. Zero tillage resulted in high weed pressure and caused soil compaction. So, in this trial, under humid conditions, the less labor intensive zero tillage system did not result in competitive maize yields. In conclusion, reduced tillage methods offer opportunities for maize cultivation in Belgium. This method of farming resulted in a lower yield, however, the difference with mouldboard ploughing was not significant. Therefore, adopting a reduced tillage system can be seen as a valid alternative for ploughing as this tillage system ensures a sustainable environment