In a proof-of-principle study, researchers at Johns Hopkins revealed that certain immune system cells found in the human liver, called liver macrophages, contain only inert HIV and aren't likely to reproduce infection on their own in HIV-infected people on long-term antiretroviral therapy (ART). ART is a regimen containing combinations of HIV-targeting drugs that prevents the growth of the virus but does not eradicate it.
The report on the findings, published in the October issue of the Journal of Clinical Investigation, revealed that although inactive, HIV can remain in liver immune cells after more than 10 years of ART. However, the researchers say there is no evidence that it can be considered an HIV reservoir in this state because the virus can't replicate at high levels.
This revelation supports the idea that when developing HIV treatments, liver macrophages can be ruled out as a cell type that may act as a reservoir of the virus. The researchers argue that it may not be important to address curing liver macrophage infection, thereby narrowing targets for treatment.
"Our study was the first, to our knowledge, that looked at whether liver macrophages also served as reservoirs, similar to CD4+ T cells, in ART-suppressed, HIV-infected people," says Ashwin Balagopal, M.D., associate professor of medicine in the Division of Infectious Diseases at the Johns Hopkins University School of Medicine and senior author of the study. "We zeroed in on the liver since liver macrophages comprise 80 to 90 percent of all macrophages in the body," notes Balagopal.
According to the Centers for Disease Control and Prevention, 36.7 million people worldwide and 1.1 million people in the United States are infected with HIV. Commonly, ART is used to suppress the replication of HIV and control the progression of AIDS in humans. The virus infects the body's immune system--the white blood cells also referred to as T cells and macrophages.
The most common reservoirs in humans are in immune cells called resting memory CD4+ T cells. Even when ART suppresses HIV, the virus can remain hidden in cellular reservoirs. The interruption or discontinuation of ART can spread HIV to new cells because the virus becomes active and begins replicating again.
"Although it is well-established that macrophages are a natural target for HIV infection, for many years researchers have not known whether macrophages also harbor HIV in a reservoir during long-term ART, similar to the resting memory CD4+ T cells," adds Balagopal.
The inability to wipe out reservoirs of infectious HIV has for decades frustrated efforts to completely cure the infection. In addition, it means that the interruption or discontinuation of ART at any time reactivates HIV replication, spreading the virus to new cells.
Balagopal and his team examined if any HIV reservoirs remained in populations of tissue macrophages that reside in livers; specifically, he looked for latent HIV-1, the most common strain of the virus, in liver macrophages.
"The other organs we could have examined that contain macrophages include the brain, heart, lungs and skin," says Abraham Kandathil, Ph.D., research associate in the Division of Infectious Diseases at the Johns Hopkins University School of Medicine, who performed all key experiments. "Macrophages can be scarce in these organs in comparison to the liver, and therefore are even more difficult to obtain than liver macrophages in sufficiently large numbers," concludes Kandathil.
To determine if liver macrophages serve as a reservoir of infection-capable HIV-1 after ART, liver tissue samples were taken from nine HIV-1 infected persons, seven of whom underwent liver transplantation at The Johns Hopkins Hospital and otherwise would have had their livers discarded because they were no longer functional and/or cancerous.
Eight of the nine persons were on ART for periods ranging from eight months to approximately 12 years.
Using lab techniques that separate out the liver macrophages, the researchers found HIV-1 to be present in the macrophages even after exposure to longstanding virus-suppressing ART.
"However, when we tried to simulate virus "rebound" by activating liver macrophages to see if the virus was infectious or noninfectious, we found HIV-1 at low levels, without the ability to replicate at high levels needed to re-establish infection," says Kandathil.
By examining the virus in these liver samples and separating out the liver macrophages, the researchers found HIV-1 to be present in the macrophages of one person among the group who took suppressive ART for almost 12 years. However, the virus was still determined to be inert, otherwise unable to replicate itself and spread.
The researchers conclude that while liver macrophages might harbor HIV-1 for a long time, it's unlikely these viruses could continue an infection on their own, and they are unlikely to function as a reservoir because the viruses were not able to replicate.
In the future, Balagopal says, more research is needed to determine if the inert HIV-1 infected liver macrophages have any functional significance in people taking ART because expression of defective HIV-1 proteins can confuse the immune system and cause tissue inflammation.
"While we have potentially ruled out the liver as an infectious reservoir, it's important to identify all of the relevant virus reservoirs in the body, such as the brain, since it's likely that the virus hides in the DNA of different cell types and will require different strategies to cure," says Balagopal. "Then we can move forward to finding a 'functional' HIV-1 cure that's comprehensive."
The researchers caution that their study is limited because of the small number of liver macrophages and human samples studied. In addition, the small number of CD4+ T cells (less than or equal to 1 percent) in the liver macrophage cultures may affect the researchers' ability to detect them, although they say it is virtually impossible for contamination of 1 percent or less to have confounded the findings.
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Other authors on this paper include Sho Sugawara, Christine M. Durand, Jeffrey Quinn, Jaiprasath Sachithanandham, Andrew M. Cameron and Justin R. Bailey of the Johns Hopkins University School of Medicine along with Ashish Goyal and Alan S. Perelson from Los Alamos National Laboratory.
This research is supported by grants from the National Institute of Allergy and Infectious Diseases (K08 AI 081544, R56 AI 118445, R01 AI024333, R01 AI116868, and P01 AI131365), the National Institute on Drug Abuse (R01 DA 016078), the National Institutes of Health Office of the Director (R01 OD 011095 and 108707-54-RKRL) and amfAR, the Foundation for AIDS Research (108814-55-RGRL).
The authors have declared no conflict of interest.
Journal
Journal of Clinical Investigation