December 1 is World AIDS Day, a global awareness day to bring attention to the disease, the research being conducted in relation to it, and the many people living with HIV/AIDS. In honor of World AIDS Day this year, we spoke with Gildas Loussouarn, University of Nantes, about his research on cardiac channels dysfunction in Long QT Syndrome, a disorder seen much more frequently in HIV patients as compared to the general population.
What is the connection between your research and HIV/AIDS?
With the worldwide development of antiretroviral therapies, HIV patients now live longer. As a result, they encounter additional pathologies and these pathologies are over-represented as compared to the general population. Among these pathologies, the Long QT syndrome, a heart rhythm condition associated with arrhythmias. In the general population, this syndrome is a rare disorder, characterized by a delayed ventricular repolarization leading to tachycardia and/or sudden cardiac death. HIV patients present with a higher prevalence of Long QT syndrome, as compared to the general population suggesting a higher risk of sudden cardiac death. Here at the institute du thorax, the global objective of our team is to decipher molecular mechanisms of cardiac ion channels and their dysfunction in cardiac arrhythmias, in order to identify new therapeutic targets. We are thus interested in cardiac channels dysfunction in the context of HIV.
Why is your research important to those concerned about HIV/AIDS?
More than 10 studies lead to the conclusion HIV patients have a higher prevalence of Long QT syndrome, as compared to the general population. Despite that, it is still difficult to address whether the LQT syndrome is due to the virus itself or to drugs that are proposed to HIV patients, which are known to prolong cardiac repolarization. Sorting this out is essential, in order to limit arrhythmias and the potential sudden cardiac death in HIV patients. By directly looking at the effect of HIV proteins on cardiac repolarization, we aim at addressing if the virus itself participate to cardiac repolarization prolongation. Importantly, we have already observed that one of the viral protein, Tat, can delay repolarization in human cardiomyocytes generated from induced pluripotent stem cells.
This figure shows that HIV-Tat is detected intracellularly in human cardiomyocytes but not simian fibroblasts (COS-7), after a 24h external application (200 ng/ml). Tat immunostaining is shown in red, plasma membrane is identified by hERG channel immunostaining (green), nucleus is in blue (DAPI). Tat remains in the extracellular compartment of COS-7 cells (red arrow) while in human cardiomyocytes, Tat is located inside the cytoplasm (asterisks) and colocalizes with hERG at the plasma membrane (yellow arrows).* (Adapted from Es-Salah-Lamoureux et al, 2016, JMCC 99:1-13, with permission.)
How did you get into this area of research and how long have you been working on it?
I was first contacted by a colleague, Dr. Bruno Beaumelle (CNRS Research Director, Montpellier). Dr. Beaumelle had previously observed that due to its high affinity to the membrane phospholipid PIP2 (phosphatitylinositol 4,5-bisphosphate), HIV-Tat could interfere with some PIP2-dependent mechanisms of neurosecretion. Bruno Beaumelle had read our previous works showing the impact of a decrease in available PIP2 on cardiac repolarization through specific cardiac channels KCNQ1 (Kv7.1) and hERG (KV11.1). Our previous works indeed suggested that a decrease in PIP2 or a decrease in KCNQ1 / hERG channels affinity for PIP2 leads to a decrease in the activity of the repolarizing channels and hence leads to Long QT syndrome. Bruno Beaumelle anticipated that we will be interested in looking at a potential effect of Tat on these channels. The hypothesis was that Tat may be a link between HIV and LQT syndrome. Our results seems to confirm this hypothesis since expression or extracellular application of Tat leads to a decrease in the activity of the repolarizing channels KCNQ1 and hERG. In human cardiomyocytes, HIV-Tat leads to a delayed repolarization and other Action Potential alterations, which are common triggers of cardiac arrhythmias.
Do you receive public funding for this work? If so, from what agency?
Our work that was just published in the Journal of Molecular and Cellular Cardiology was partially funded by the Fédération Française de Cardiologie, the Fondation Genavie, the Marie Curie European Actions, the French Regional Council of Pays-de-la-Loire, the National Research Agency, the Fondation Lefoulon Delalande, the Fondation pour la Recherche Médicale, the Association of Scientific Orientation and Specialization and Campus France. We hope to get specific funding from AIDS focused agencies allowing us to study the effects of Tat in further details.
Have you had any surprise findings thus far?
Yes! Our first surprise was that external application of Tat, which is known to penetrate cells, had an effect on hERG channels depending only in specific cell type! Tat did not have any effect on hERG channels expressed in COS-7 cells. Paradoxically, the same application halved the activity of the same channels in cardiomyocytes! The absence of Tat effect in COS-7 cells is surprising since Tat is a promiscuous ligand that binds a plethora of receptors (such as Heparan Sulfate Proteoglycans), internalization of which supposedly allowing Tat entry. To identify the reason of this cell-specific effect, we looked for intracellular Tat in both models, after its extracellular application. We observed that intracellular Tat could be detected by immunofluorescence, in cardiomyocytes but not in COS-7 cells (cf. also figure). We suppose that Tat requires specific cellular components to be internalized in sufficient amount to have an effect on the potassium channels. It has been shown that Tat interacts tightly with some receptors such as LRP and CXCR4 receptors, which represent a cell-specific way for internalization. Such receptors may lack in COS-7 cells. In addition, these observations illustrate the great value of induced pluripotent stem (iPS) cells derived cardiomyocytes, a model closer to mature human cardiomyocytes as compared to COS-7 cells. Human iPS cells are quite easy to get: they were obtained by reprogramming renal cells contained in the urine of a patient. This is clearly a non-invasive (but long!) way of getting human cardiomyocytes.
What is particularly interesting about the work from the perspective of other researchers?
Tat is present in the patient’s serum and seems to target different organs and interfere with various PIP2-dependent processes in these organs: Bruno Beaumelle showed that Tat can interfere with the secretory activity of neuroendocrine cells, by sequestering PIP2. We then showed that Tat can interfere with cardiac repolarization, also by sequestering PIP2. Now, we could test if Tat is also a link between HIV and epilepsy, which is also overrepresented in HIV patients. This hypothesis is founded on the observation that PIP2 activates neuronal channels (KCNQ2/KCNQ3), alteration of which leads to epilepsy. We can hence speculate that AIDS may be seen as a “PIP2-pathie” with multiple organs targeted by Tat.
What is interesting about the work from the perspective of the public?
I think it is important for the public to bear in mind that despite huge progresses made in treatments against HIV, living with AIDS is associated to many other severe pathologies, including cardiac diseases. It is thus important to remain aware of risk- behaviors. Regarding potentially new therapies, an anti-Tat vaccine may represent a new therapeutic strategy, as tested currently by another French laboratory (ETRAV laboratory, Aix Marseille University/CNRS).
*Image scale= 5μm.