Jorge M. Tolosa

The endogenous retroviral envelope protein syncytin-1 inhibits LPS/PHA-stimulated cytokine responses in human blood and is sorted into placental exosomes

OBJECTIVES To examine whether syncytin-1 has immune regulatory functions and is carried by human placental exosomes. Further, to examine whether corticotropin-releasing hormone (CRH) can induce the production of syncytin-1. STUDY DESIGN Human placental exosomes were isolated from placental explant, primary trophoblast and BeWo cell cultures. The presence of exosomes was confirmed by transmission electron microscopy and western blotting. Exosomal protein was probed with 3 separate antibodies targeting syncytin-1. Syncytin-1 immunosuppression was tested, using either a syncytin-1 recombinant ectodomain protein or a synthetic peptide with the human syncytin-1 immunosuppressive domain sequence, in an in vitro human blood culture system immune challenged with LPS or PHA. The inhibition of cytokine production by syncytin-1 was determined by ELISA of TNF-α, IFN-γ and CXCL10. BeWo cells were stimulated with CRH or vehicle for 24 h. mRNA and Protein was extracted from the cells for real-time PCR and western blotting analysis while exosomes were extracted from conditioned media for analysis by western blotting. RESULTS Protein expression of syncytin-1 was detected in exosomes isolated from placental explants, primary trophoblast and BeWo cell cultures. Syncytin-1 recombinant ectodomain was also shown to inhibit the production of the Th1 cytokines TNF-α and IFN-γ as well as the chemokine, CXCL10 in human blood cells. Finally, this study showed that syncytin-1 can be stimulated by CRH. CONCLUSIONS The presence of syncytin-1 in placental exosomes provides a mechanism for syncytin-1 to reach and interact with target cells of the maternal immune system and represents a novel mechanism of endogenous retroviral mediated immunosuppression that may be relevant for maternal immune tolerance.

Column-based method to simultaneously extract DNA, RNA, and proteins from the same sample

We describe a procedure for the simultaneous extraction of proteins and nucleic acids from the same experimental sample allowing for direct correlations between genetic, genomic, and proteomic data. This approach, using commercially available column-based nucleic acid extraction kits, requires no hazardous chemicals and is a quick, reliable, and consistent method for concomitant protein extraction. Buffer choice is critical to completely solubilize all proteins in the sample. Proteins solubilized in radioimmunoprecipitation assay (RIPA) buffer did not represent the entire profile when compared with conventionally extracted proteins using the same buffer at the one-dimensional (1-D) sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) level, however proteins extracted from the columns and solubilized in a two-dimensional (2-D) electrophoresis lysis buffer showed a similar profile to conventionally extracted proteins when analyzed at both the 1-D and the 2-D level. We further showed that proteins extracted using these methods were compatible with Western blot analysis. This technique provides a simple and effective way to analyze protein and nucleic acids simultaneously from the same sample without affecting yield and quality.

Recent advances in understanding the endocrinology of human birth.

The timing of human birth has a crucial impact upon the survival of the fetus. New knowledge on the regulation of human birth includes the role of endogenous retroviruses in the formation of the syncytiotrophoblast cells and consequently the secretion of corticotrophin releasing hormone, a hormone linked to gestational length determination. miRNAs have been identified that mediate progesterone withdrawal at labor by suppressing progesterone-induced transcription factors. Progress has also been made in understanding how the contractile machinery of the uterine myocytes is activated at labor and the role of small heat-shock proteins in this process. From this work, new therapeutic targets have been identified that may be used to regulate the onset of labor and improve neonatal mortality.

The placental protein syncytin-1 impairs antiviral responses and exaggerates inflammatory responses to influenza.

Background Pregnancy increases susceptibility to influenza. The placenta releases an immunosuppressive endogenous retroviral protein syncytin-1. We hypothesised that exposure of peripheral monocytes (PBMCs) to syncytin-1 would impair responses to H1N1pdm09 influenza. Methods and Findings Recombinant syncytin-1 was produced. PBMCs from non-pregnant women (n=10) were exposed to H1N1pdm09 in the presence and absence of syncytin-1 and compared to responses of PBMCs from pregnant women (n=12). PBMCs were characterised using flow cytometry, release of interferon (IFN)-α, IFN-λ, IFN-γ, IL-10, IL-2, IL-6 and IL-1β were measured by cytometric bead array or ELISA. Exposure of PBMCs to H1N1pdm09 resulted in the release of IFN-α, (14,787 pg/mL, 95% CI 7311-22,264 pg/mL) IFN-λ (1486 pg/mL, 95% CI 756-2216 pg/mL) and IFN-γ (852 pg/mL, 95% CI 193-1511 pg/mL) after 48 hours. This was significantly impaired in pregnant women (IFN-α; p<0.0001 and IFN-λ; p<0.001). Furthermore, in the presence of syncytin-1, PBMCs demonstrated marked reductions in IFN-α and IFN-λ, while enhanced release of IL-10 as well as IL-6 and IL-1β. Conclusions Our data indicates that a placental derived protein, syncytin-1 may be responsible for the heightened vulnerability of pregnant women to influenza.

Mechanisms of maternal immune tolerance during pregnancy

Throughout their evolution, animals have developed mechanisms which protect them against parasites or infections by detecting and destroying foreign biological material within their own bodies. These mechanisms for excluding “non-self” biological materials whilst at the same time maintaining the integrity of the “self” have evolved for hundreds of millions of years into a highly complex body system, the immune system (Bainbridge, 2000). Any disbalance or alteration of the mechanisms maintaining the dynamic equilibrium between the “self” and “non-self” recognition could translate into a pathological state (or condition) like autoimmune disease at one end of the scale or immunodeficiency at the other.

Targeting the TSH receptor in thyroid cancer

Recent advances in the arena of theranostics have necessitated a re-examining of previously established fields. The existing paradigm of therapeutic thyroid-stimulating hormone receptor (TSHR) targeting in the post-surgical management of differentiated thyroid cancer using levothyroxine and recombinant human thyroid-stimulating hormone (TSH) is well understood. However, in an era of personalized medicine, and with an increasing awareness of the risk profile of longstanding pharmacological hyperthyroidism, it is imperative clinicians understand the molecular basis and magnitude of benefit for individual patients. Furthermore, TSHR has been recently re-conceived as a selective target for residual metastatic thyroid cancer, with pilot data demonstrating effective targeting of nanoparticles to thyroid cancers using this receptor as a target. This review examines the evidence for TSHR signaling as an oncogenic pathway and assesses the evidence for ongoing TSHR expression in thyroid cancer metastases. Priorities for further research are highlighted.