Memory CD4 T cells and the neonatal gut

I have found a short paper on the potential mechanism of how HIV virus may be transmitted between mother and child. I think it is interesting because it not only provides the information which may be useful for a given pathology but it also poses some questions as to the basic immunology processes. The main theme of the paper is the quest for HIV targets among neonatal CD4 T cells. As it is known the virus tends to infect memory CD4 T cells but these cells are practically absent in the cord blood. Thus authors inspect neonatal CD4 T populations from various anatomical compartments and find that CD4 T cells bearing a memory marker and HIV co-receptor abound at the intestinal mucosa.

The link: http://bloodjournal.hematologylibrary.org/content/120/22/4383.abstract

CD4 T cells collected for this study derive from children born to healthy mothers therefore this report asks only about the potential mechanism of mother to child transmission. Authors follow CD4 T cells that bear also CD45RO (which is a marker of memory state) and CD5 (HIV uses this molecule as a co-receptor to infect an individual cell – only CD5-tropic strains tend to become transmitted form mother to child). The main conclusion of this publication is that the population of CD4+CD45RO+CD5+cells (the potential HIV target according to the current state of knowledge) exists at the neonatal gut mucosa but not in the lymph nodes, spleen or blood. Additionally, around half of this intestinal memory CD4+CD45RO+CD5population appears to be differentiated into Th17 phenotype since these cells express RORγt transcription factor and CCR6.  In an in vitro experiment investigators also show that neonatal CD4 T cells from the gut are more susceptible to HIV infection than CD4 T cells from the lymph nodes or blood.

Based on obtained data authors propose a model of how HIV gets transmitted from mother to child. According to them the virus may take the oral route of transmition by the ingestion of infected body fluids during the delivery or milk shortly afterwards. I lack the clinical knowledge to critically evaluate such proposal. But I have more basic question instead. This paper not only shows the presence of memory CD4 T cells population at the neonatal gut mucosa but it also provides the evidence that these memory cells underwent substantial clonal expansion that must have happened in utero. I would like to know more details on the nature of antigenic challenge that underlies such prenatal activation of the adaptive immune system.

Bunders MJ, van der Loos CM, Klarenbeek PL, van Hamme JL, Boer K, Wilde JC, de Vries N, van Lier RA, Kootstra N, Pals ST, & Kuijpers TW (2012). Memory CD4+CCR5+ T cells are abundantly present in the gut of newborn infants to facilitate mother-to-child transmission of HIV-1. Blood, 120 (22), 4383-90 PMID: 23033270

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First prime and then pull – the novel immunization approach

Some areas of our body enjoy a special status as far as the immune reaction is concerned. Anatomical entities like the gut or female genital tract as well as other mucosal surfaces do not support the same extend of protective response compared to many non-mucosal tissues. This exclusion is crucial to avoid the unwanted inflammation in places that are regularly exposed to the outer environment but sometimes it may present a problem when there is the need to elicit the strong protective response at such privileged site. I have found an interesting report which applies the novel vaccination strategy aimed to enhance the protection against herpes simplex virus 2 which being the virus transmitted through the contact with infected body fluids often enters the body through the genital organs. The innovation that this report introduces consists of double treatment (“prime and pull”) which bypasses the restrictive entry of memory T cells into the vaginal mucosa.

The link: http://www.nature.com/nature/journal/v491/n7424/full/nature11522.html

The mentioned “prime and pull” strategy is the subcutaneous immunization with an attenuated strain of HSV-2 (prime) which is followed by the topical application of chemokines CXCL9 and CXCL10 to the vaginal mucosa (pull). Authors follow the localization of CD8 T cells that recognize an epitope within one of HSV-2 glycoproteins and activated CD4 T cells to show that the distal immunization event plus the localized chemokine treatment provokes the significant recruitment of activated lymphocytes to the vagina whereas the immunization alone has much weaker effect. Interestingly, this recruitment is specific to CD4 and CD8 lymphocytes and does not encompass other cell types that express the relevant chemokine receptor CXCR3.

Is the “prime and pull” approach able to provide the longstanding and reliable protection? Data demonstrate that CD8 T cells (but not CD4 T cells) are retained at vaginal mucosa after the primary response period is over. Most importantly the “prime and pull” treatment may be indeed superior in enforcing the better protective immunity to HSV-2 challenge than the immunization alone. Investigators also ask about the mechanism by which the protection is delivered by the “prime and pull” strategy. It appears that this application can prevent the virus from entering the nervous system where HSV-2 propagates past the mucosal stage of infection.

What will be the future of “prime and pull”, though? The pros are obvious – there is the simple method to enhance the mucosal migration of protective lymphocytes without the “ugly face” of immunity which in this case would be the excessive inflammation at the sensitive anatomical location. Authors speculate about the future applications ranging from HIV protection to solid tumors treatment. The method itself may also be developed as in the discussed paper it provides the optimal protection only in conjunction with the adoptive transfer of virus-specific lymphocytes. The “pull” works as well with the endogenous population of CD8 T cells; however, the protection is suboptimal in such scenario. I will follow this story.

Shin H, & Iwasaki A (2012). A vaccine strategy that protects against genital herpes by establishing local memory T cells. Nature, 491 (7424), 463-467 PMID: 23075848

Non-pathogenic SIV infection and type-I interferon signaling

How monkeys or apes respond to the challenge of lentiviral immunodeficiency viruses varies across different primate species. Some primates like rhesuses are similar to humans because following SIV infection they develop the AIDS-like disease with all the characteristic features of progressive immune destruction. However, there are other species that do not display such aggravated pathology. African sooty mangabeys are the best studied example among these AIDS-refractory animals. Infected sooty mangabeys do not clear the virus but seem to have adapted to live with it. Such infection lasts for life but it is the relatively mild condition without the continuous depletion of memory CD4 T cells and the chronic immune activation that are associated with human or simian AIDS. The current clinical efforts in humans aim at the reduction of damage caused by the infection and slowing down the progression to AIDS. Thus the detailed knowledge of how AIDS-refractory species achieve their status might be instructive and there is the respective research avenue devoted to studying these species. I have found the publication that looks at the role of type-I interferon signaling during the chronic phase of SIV infection in a species that does not progress to AIDS.

The link: http://bloodjournal.hematologylibrary.org/content/119/24/5750.abstract

Authors attempt to clarify the interactions between the presence of the augmented type-I interferon signaling and the immune response in the chronic phase of SIV infection. Their rationale is simple – since the up-regulation of interferon signature genes correlates with HIV/SIV infections that progress to AIDS, so what may happen if artificially boost the expression of these genes during the non-pathogenic SIV infection? To this end they choose several naturally infected sooty mangabeys and subject them to the treatment with type-I IFN agonist which procures strong but transient enhancement in the expression of interferon signature genes.

To obtain the answer to their question investigators focus on how the increased type-I interferon signaling influences several relevant immune parameters. Acquired data are compared to the baseline values that were observed before the onset of treatment (no control group is included in this research due to availability reasons). The studied parameters comprise the range of CD4 T cells depletion (an indicator of the immune system impairment), the activation and proliferation levels of CD4 T cells (indicators of the chronic immune activation) as well as the intensity of anti-SIV CD8 response.

The take-home message from this report is that the administration of type-I interferon agonist does not impact any of immune parameters that were tested but it only brings down temporarily the viremia level (after all, type-I IFN is regarded as the anti-virus defensive molecule). What does it mean for the understanding of non-pathogenic SIV infection? The mechanisms responsible for the AIDS-refractory status are most probably complex, robust and might not depend on just one particular pathway.

Vanderford TH, Slichter C, Rogers KA, Lawson BO, Obaede R, Else J, Villinger F, Bosinger SE, & Silvestri G (2012). Treatment of SIV-infected sooty mangabeys with a type-I IFN agonist results in decreased virus replication without inducing hyperimmune activation. Blood, 119 (24), 5750-7 PMID: 22550346

The different chemokine profile in HIV-exposed seronegative persons

The studies on human population infected with or exposed to HIV have brought the description of several virus-refractory phenotypes. Among them are long-time non-progressors and HIV-exposed seronegative persons. The first group is able to control virus replication without the anti-retroviral therapy, maintain the normal CD4 T cell number and avoid the chronic immune activation that is normally associated with HIV infection. In consequence, long-time non-progressors can live with the presence of HIV for many years with minimal or without any ill effects. The second group is less widely known and comprises people who despite the persistent exposure to the virus do not become infected. In ordinary circumstances HIV gets into the body through mucosal surfaces at genital organs. HIV-exposed seronegative persons (HESN) appear to differ from the virus-sensitive population in terms of chemokine profile at the natural infection site.

The link: http://www.nature.com/mi/journal/v5/n3/abs/mi20127a.html

The study is conducted among Kenyan commercial sex workers divided into three categories. The experimental group includes HESN women, whereas control groups are HIV-1 negative women and HIV-1 infected patients. Authors analyze the cytokine/chemokine profile in the cervicovaginal lavage of each group. They detects that MIG and IP-10 (two IFN-γ inducible chemokines involved in the leukocyte trafficking) and cytokine IL-1α are expressed at lower level in the HESN cohort. Both chemokines are involved in the mucosal migration of activated CD4 T cells – the main HIV target. Hence one of HESN phenotype explanation might be that these people display the state of immune quiescence at their genital mucosa and simply do have enough number of activated CD4 T cells to become infected by virus.

To validate this point investigators examine plasma chemokine/cytokine profile in all studied groups as lymphocytes migrate to mucosal surfaces according to the chemokine gradient between blood and mucosa. They demonstrate that MIG levels in HESN group were higher in systemic compartments than at the genital mucosa whereas the HIV-1 negative population shows the opposite trend. Additionally, HESN subjects uniquely display the decreasing gradient for IP-10 from plasma to mucosal surfaces. The supportive data in this paper include the analysis of CXCR3 (the receptor bound by MIG and IP-10) on CD4 and CD8 T cells collected from the genital mucosa. Authors also study the expression of antiproteases in the genital tract as they are important factors regulating mucosal chemokine levels. It would be interesting to know what really drives the decreased chemokine levels in the genital tract of HESN.

J Lajoie, J Juno, A Burgener, S Rahman, K Mogk, C Wachihi, J Mwanjewe, F A Plummer, J Kimani, T B Ball, and K R Fowke (2012). A distinct cytokine and chemokine profile at the genital mucosa is associated with HIV-1 protection among HIV-exposed seronegative commercial sex workers Mucosal Immunology DOI: 10.1038/mi.2012.7

Memory compartment regeneration in SIV infection does not rely on naive CD4 T cells

Among mechanisms hold responsible for the severe CD4 T cell depletion in AIDS are those that contribute to increased apoptosis rates for CD4 T cells (either infected or non-infected) and decreased CD4 T cells regeneration capacity. Pathogenic HIV/SIV clones are in their majority CCR5-tropic. CCR5 is expressed on tissue-resident effector memory CD4 T cell populations and not surprisingly these subsets are decimated during the primary HIV/SIV infection. However, the CCR5-negative central memory subset which resides in secondary lymphoid organs is relatively spared and may form the reservoir for the subsequent effector memory subsets regeneration. The publication I have found investigates details of memory CD4 T cells renewal in non-human primate model infected with SIV and makes an intriguing point about what is not needed to bring back depleted memory compartments.

The link: http://jem.rupress.org/content/209/4/641.abstract

The chief aim of this paper is to assess whether naïve CD4 T cells presence is needed for the memory compartment replenishment during pathogenic SIV infection. Authors employed rhesus macaques and subjected them to a number of manipulations before proper infection. Briefly, experimental groups have been either thymectomized or left intact and subsequently CD4 T cells in each group were depleted with anti-CD4 antibody following which animals were left to regenerate CD4 T cell counts. Such approach allows comparing regeneration of memory subsets and general disease outcome between two different conditions – complete lack vs. normal level of naïve CD4 T cells. If you have read my previous post, note that this report assumes that the thymus is the only place of significant naïve CD4 T cell development.

After complete rebuilding of CD4 T cells counts took place animals from each group were infected with SIVmac239. Then they underwent the long data collection period which included initial untreated infection, anti-retroviral therapy phase and treatment discontinuation in order for animals to develop AIDS-like symptoms. Investigators amassed data relating to multiple parameters – most notably they followed plasma viral load, memory CD4 T cells depletion levels (in blood and at mucosal lung surfaces) and the presence of anti-SIV adaptive immunity over the entire span of experiment. They also observed the disease progression and appearance of opportunistic infections in both groups. The summary conclusion of this study is that in the course of pathogenic SIV infection naïve CD4 cells presence is not necessary for the regeneration of memory CD4 T cells subsets. Additionally, the depletion of naive CD4 T cells do not seem to influence the disease advancement.

Major findings of this publication are very convincing and well supported with data. However, I would like to pose one question – it looks like the depletion of naive CD4 T cells in thymectomized group impairs the scope of adaptive anti-SIV response. For example, anti-SIV CD4 T cell responses (defined as the percentage of memory compartment specific to gag+pol+env+nef) never really start working. Thymectomized animals show much slower seroconversion rate and blunted anti-SIV CD8 T cell reactions as well. Yet the general disease outcome between both groups is remarkably similar at all studied stages. How to explain it?

Okoye, A., Rohankhedkar, M., Abana, C., Pattenn, A., Reyes, M., Pexton, C., Lum, R., Sylwester, A., Planer, S., Legasse, A., Park, B., Piatak, M., Lifson, J., Axthelm, M., & Picker, L. (2012). Naive T cells are dispensable for memory CD4+ T cell homeostasis in progressive simian immunodeficiency virus infection Journal of Experimental Medicine, 209 (4), 641-651 DOI: 10.1084/jem.20112071

Germinal centers in pathogenic SIV infection

The characteristic feature of HIV and some SIV infections is the chronic immune activation that probably drives the continuous CD4 T cell depletion and progression towards AIDS. What exactly causes this aberrant activation is not clear; however, traits inherent to the immune response seem to be more responsible than the virus itself. For example, there are data showing that the same strain of virus (SIV) can induce lasting but non-pathogenic infection in one primate species (sooty mangabeys) and devastating disease which resembles AIDS in the other (rhesus macaques). It may be therefore instructive to study details of immune response against SIV in pathogenic conditions. B lymphocytes are not spared from overall activation during HIV/SIV infection – in fact, sick individuals display hypergammaglobulinemia, or elevated levels of IgG antibodies. Switched antibodies are generally produced by cells that derive from the germinal center reaction where B lymphocytes interact with the unique subset of CD4 T cells called follicular helpers (TFH). Follicular helpers can be distinguished from other CD4 T cell subsets by their expression of CXCR5, ICOS, PD-1 and the transcription factor Bcl-6 as well as the high level of IL-21 cytokine secretion. TFH have been shown to be crucial for the successful generation of plasma cell and memory compartments. The recent paper takes a look at germinal centers during SIV infection in monkeys that develop AIDS-like disease.

The link: http://www.jimmunol.org/content/188/7/3247.abstract

Authors use immunohistological staining and flow cytometry to prove that PD-1 positive CD4 T cells (according to what we know TFH population) accumulate in germinal centers along disease progression in rhesus macaques infected with SIVmac239. They also correlate the enhanced PD-1 expression on CD4 T cells with remodeling of B cell populations that reside in lymph nodes (the trend is towards reducing naïve subset and enriching memory subsets) and the increased IgG secretion. Additionally, it is demonstrated that CD8 T cells are excluded from germinal centers (both in naïve and infected monkeys). Investigators conclude that the buildup of TFH population during pathogenic SIV infection may play the important role in shifting B cell activation status. Apart from that they suggest that germinal centers could form anatomical sites of potential viral escape due to the lack of CD8 T cells presence (and alleged insufficient control of infected CD4 T cells there).

The experimental setting of this publication is restricted to recording several immune parameters at time points that represent either acute or chronic SIV infection. However, it is interesting because it attempts the systematic documentation of changes in the immune response that may underlie the pathologic immune activation distinctive for some primate lentiviruses. The obvious question for the future is whether there may be any significant changes in germinal center reaction between species that develop pathogenic or non-pathogenic SIV infection.

Hong, J., Amancha, P., Rogers, K., Ansari, A., & Villinger, F. (2012). Spatial Alterations between CD4+ T Follicular Helper, B, and CD8+ T Cells during Simian Immunodeficiency Virus Infection: T/B Cell Homeostasis, Activation, and Potential Mechanism for Viral Escape The Journal of Immunology, 188 (7), 3247-3256 DOI: 10.4049/jimmunol.1103138

The link between Th17 and HIV pathogenesis

The Th17 population is a CD4 T lymphocyte effector subset that in the simplest way can be defined as IL-17A releasing cells; although several other cytokines and transcription factors seem to be typical as well for cells that express IL-17A. These cells are protective in several bacterial and fungal diseases, particularly at mucosal parts of our body. They also are regarded as pathogenic during autoimmune conditions and the infection with Schistosoma mansoni parasite. Another interesting feature of Th17 population is the link to HIV pathogenesis. HIV virus replicates within Th17 subset and IL-17A expressing cells are preferentially depleted from the gut mucosa during HIV infection. The reduction in Th17 level has been linked to increased permeability of mucosal barrier in the digestive tract. This can lead to continuous leaking of microbial products into the bloodstream and maintains chronic immune activation – the HIV infection distinctive feature. Two recent publications reinforce the connection between Th17 and HIV.

The first link:

http://www.jimmunol.org/content/187/11/6032.abstract?etoc

This publication presents data collected in the region that is affected by AIDS pandemic. Subjects of the study are Kenyan women divided into two pools – virus-negative healthy controls and HIV-positive sex workers. HIV tends to infect activated CD4 T cells that express certain co-receptors facilitating the virus entry. Additionally, α4β7 integrin known to be involved in the mucosal homing binds HIV gp120 envelope protein. Accordingly, authors perform the comparative analysis of cervical and blood CD4 T lymphocytes from healthy donors. This analysis includes enumeration of mucosal homing markers (CCR9, α4β7 and CD103), the early activation marker CD69 and HIV co-receptors (CCR5 and CXCR4).

The research aims to estimate features of potential invasion targets in the female genital tract. As the first conclusion authors have found that cervical CD4 T lymphocytes are enriched for several indicators of HIV susceptibility with α4β7 integrin, CD69 and CCR5 often being co-expressed on individual cells. It has been also established that cervical CD4 T lymphocytes include cells positive for IL-17A. The majority of these IL-17A-positive cells express α4β7 integrin and CCR5 which makes them probable targets during the initial infection stage. Next, authors examine IL-17A expression levels in blood and cervical samples from infected women. The study concludes with the evidence that IL-17A-positive CD4 T cells are strikingly depleted from the cervix mucosa of diseased participants but still present in their blood.

The second link:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3248296/?tool=pubmed

Data reported in this article have been obtained from European healthy donors and patients infected with HIV that are treated with anti-retroviral therapy. People infected with HIV are continuously loosing their CD4 T cells and as a result develop the permanent immune deficiency. The anti-retroviral therapy helps rebuilding overall level of CD4 T cells but often proves ineffective in the restoration of intestinal CD4 T cell count. Investigating this matter authors focus on CD4 T lymphocytes from the small intestine and the blood to compare their gut-homing markers expression (α4β7 integrin and CCR9). Healthy controls harbor the vast majority of double positive β7+CCR9+ cells in the gut. However, infected subjects demonstrate the conspicuous reversal in proportions with most of β7+CCR9+ population being present in the blood. Additionally, HIV-positive individuals display the reduction of CCL25 (CCR9 ligand) expression level on intestinal epithelial cells.

This result implies that HIV infection may disturb the redistribution of CD4 T lymphocytes to the gut by unbalancing the CCR9-CCL25 interaction. Authors also associate the low level of intestinal β7+CCR9+ CD4 T cells with increased indicators of damage to the intestinal epithelium. Finally, it is demonstrated that β7+CCR9+ subset is richer in IL-17A secreting cells than β7+CCR9 population. However, in most of their experiments authors did not discriminate between Th17 and other CD4 T cell subsets.

I am following the Th17-HIV story because it emphasizes certain point, which I think is important from the immunologist perspective. Data seem to indicate that HIV infection drives the reduction of Th17 cells at mucosal surfaces. This reduction seems to be irreversible and it disturbs the intestinal homeostasis. Changes in mucosal environment may lead to the enhanced rate of microbial translocation which fuels the unique HIV trait – systemic immune activation. The link between Th17 and HIV pathogenesis makes me wonder whether the consequences of Th17 depletion from mucosal sites arise because of insufficient protection versus enteric pathogens or inadequate accommodation of intestinal commensals.

Our immune system is often metaphorically viewed as well trained and properly equipped coercive body that provides protection against foreign invaders. That’s right – the immune system does provide protection and it is very good at it. But such metaphor might be too narrow to adequately reflect what our immune system actually is, especially at mucosal surfaces. After all we are inhabited by diverse commensal organisms to be reckoned with but not to be disposed of.

Obviously our digestive tract needs protection as it is constantly exposed to harmful bacteria and sometimes the distinction between pathogen and commensal is not entirely clear. Additionally, pathogens are equipped with virulence tools like secretion systems that may provide them with possibilities to stand out from the crowd. But if you consider the sheer biomass argument, commensal organisms prevail over pathogens by such enormous margin that their proper accommodation definitely looks like more urgent matter than the efficient protection against few marauders. So returning to the insufficient protection versus inadequate accommodation dilemma – my gut feeling is that the second option might be more correct.