A recent study by CONICET scientists reveals the existence of a circuit that involves extracellular vesicles (nanometric-sized structures present in all body fluids, such as blood), immune system cells known as macrophages, and Galectin-1. (a protein that plays an important role in different pathologies), which reverses the latency of the virus and, in this way, modulates the activity and repopulation of the HIV viral reservoir. The results of this work were published in the journal mBio of the American Society for Microbiology.
The human immunodeficiency virus (HIV) is a pathogen that causes the progressive loss of CD4 lymphocytes - cells of the immune system involved in the response to infections - and, if not treated, leads to acquired immunodeficiency syndrome (AIDS). Although currently, treatment with antiretrovirals allows, in most cases, to control the replication of the virus, stabilize the CD4 number, prevent immunodeficiency and reduce mortality, until now it has not been possible to develop a therapy that makes it possible completely eradicate the virus. Even in cases in which the treatment is successful and makes the viral load in the blood undetectable by standard tests, the virus persists in a state of latency in a small group of cells and, if therapy is interrupted, it can return. to replicate.
The development of therapies that allow achieving a sterilizing cure (the complete eradication of the viral load and the body’s reservoirs) is one of the central objectives of those who research in the field of HIV. To achieve this goal, it is essential to better understand the dynamics of HIV reservoirs. In this sense, experts are exploring both possible treatments consisting of attacking the cells in which the infection persists by first reactivating the virus (which would allow infected lymphocytes to be distinguished from non-infected ones), as well as others that are based on achieving greater control and silencing of the viral reservoir.
“Many individuals living with HIV, even when they are under effective antiretroviral treatment, have persistent activation of their immune system and a state of chronic inflammation. This situation is considered one of the keys that explain the persistence of the viral reservoir in people under treatment. Infected individuals, even though they may have a very long survival thanks to antiretroviral therapy, suffer from different health complications - cardiovascular, bone and metabolic - related to chronic inflammation. In previous work we showed that, in HIV patients, extracellular vesicles stimulate macrophages to produce inflammation. In this research we saw that these extracellular vesicles also cause macrophages to express high levels of Galectin-1 (Gal-1) and secrete it into the blood. But in addition, we detected that this galectin, when interacting with infected lymphocytes, reverses latency," explained Matías Ostrowski , CONICET researcher at the Institute for Biomedical Research in Retroviruses and AIDS (INBIRS, CONICET-UBA) and one of the coordinators of the job.
Although the reactivation of the viral reservoir induced by Gal-1 is not dangerous in terms of leading a person under treatment to immunodeficiency, it could be one of the impediments to achieving a sterilizing cure, that is, to the eradication of latent HIV reservoirs. Achieving a sterilizing cure, on the other hand, could allow patients to interrupt treatment without the risk of the virus replicating again, as well as provide a solution to the pathogenesis linked to chronic inflammation.
“This leads to the question of whether blocking Gal-1 expression could contribute to controlling latent HIV reservoirs. Another alternative would be, on the contrary, to stimulate the expression of Gal-1 to ’wake up’ the infected cells and attack them using other drugs ," explained CONICET researcher Gabriel Rabinovich , director of the Glycomedicine Laboratory of the Institute of Biology and Experimental Medicine. (IBYME, CONICET- F-IBYME) and also coordinator of the study.
The interesting thing, in this sense, is that Rabinovich’s team at IBYME is working both on the development of an antibody that can block Gal-1 (which could be used, for example, to treat certain tumors), and on a agent that mimics or stimulates the production of this protein (which could serve to prevent autoimmune diseases).
This study arose as a collaboration between the Ostrowski laboratory, a specialist in the role of extracellular vesicles in HIV-associated inflammation, and Gabriel Rabinovich’s team, which has been dedicated to investigating the role of galectins for almost three decades – especially Gal-1- in different physiological and pathological scenarios. The work is part of the doctoral thesis of the first author of the article, Julia Rubione , who between 2015 and 2020 was a CONICET doctoral fellow at INBIRS.
“The goal of this collaboration was to study whether Gal-1 had a role in HIV infection. "Given that Gal-1 is an immunomodulator and has an anti-inflammatory function in many pathologies, we expected to find something on that side, but the results took us down a different path and we ended up discovering a role for Gal-1 that we were unaware of until now," comments Rabinovich, who is also a professor of Immunology at the Faculty of Exact and Natural Sciences of the University of Buenos Aires (FCEN, UBA).
The tests carried out for this study included both the analysis of samples from different groups of patients with HIV, as well as the performance of in vitro experiments in cell cultures that involved tissues taken from individuals with HIV.
Circuit that reprograms the latent HIV reservoir
“The analysis of serum from different groups of patients allowed us to determine that there is a considerable increase in the normal levels of Gal-1 in circulation in the blood of individuals with HIV. Furthermore, we were able to associate the increase in Gal-1 levels with the chronic inflammation of patients with HIV, as well as with an increase in the transcriptional activity of the viral reservoir ,” Rubione said.
On the other hand, the comparison between the different groups of patients and the follow-up of infected individuals - before and after treatment with antiretrovirals - allowed us to rule out the existence of a correlation between the increase in Gal-1 and the viral load , as well as with the CD4 lymphocyte count. No association was found between the increase in Gal-1 and the size of the reservoirs, although there was an association between the concentrations of the lectin and the transcriptional activity of the reservoirs.
“In a subsequent trial, in cell cultures, we studied the impact of extracellular vesicles on macrophages and we saw that vesicles from patients with HIV induced the secretion of Gal-1 in much greater quantities than vesicles from healthy donors,” Rubione noted. .
This experiment allowed the research team to confirm that the increase in Gal-1 in the blood of HIV patients is associated with the inflammatory phenomenon and the action of extracellular vesicles.
“Then we took the supernatant from the stimulated macrophages and exposed it to cells that are reporters of the activity of the latent virus. This cell line, when faced with a stimulus that reactivates the latent virus, expresses a fluorescent protein. In this way we were able to see that the supernatants that came from macrophages stimulated with HIV vesicles promoted the activation of the viral reservoir much more than the supernatants stimulated with vesicles from healthy patients,” explained Rubione.
In this way, the research team was able to reassemble, from in vitro tests, the circuit that the analyzes of patient samples seemed to suggest: GAL-1, when secreted by proinflammatory macrophages stimulated by extracellular vesicles, reverses the latency of the HIV.
“Something that I would like to highlight about this work is that after the in vitro experiments that allowed us to assemble the circuit and understand its mechanisms, we returned to patient samples to validate the effect that we had described. One of the latest experiments consisted of taking cells from patients with HIV and exposing them to Gal-1 and we saw that the virus was indeed reactivated,” Ostrowski concluded.
According to the researchers, the importance of the results obtained invites us to continue the study of this new circuit, which could be key both to unraveling the pathologies associated with chronic inflammation in HIV patients and to designing therapies that allow modulating viral reservoirs.
