Rectal inflammation study establishes normal values, confirms efficacy of potent anti-HIV therapy

A study examining rectal biopsies from 16 men has established, for the first time, the typical immunological ‘climate’ inside the rectum including the usual immune-cell populations in non-inflamed tissues. It goes some way to establishing the most stable immune parameters to use in studies of candidate microbicides used rectally. The study is published in the October edition of the Journal of Acquired Immune Deficiency Syndromes.

The study also confirmed previous findings showing that HIV infection dramatically, but selectively, disorders the immune status of the gut, the body’s largest immune organ, decimating certain T-cell subsets while putting others in a state of hyperactivation. However it also found encouraging indications that long-term antiretroviral treatment can reverse the majority of this immune derangement.

In the last few years studies have established that HIV infection, far from being a slowly-worsening chronic depletion of the immune system, involves an immediate and drastic reduction in certain populations of CD4 cells in the gut, which occurs within the first few weeks of infection and from which, in untreated HIV, the body never recovers. While the exact causes are uncertain, this depletion is suspected of being the major driver of progression to AIDS as reported on aidsmap.com here.

It is also important to establish a better understanding of immune processes in the gut so that benchmarks can be developed to use in studies of microbicides or other substances or conditions that cause gut inflammation.

While a qualitative histological scoring system has existed for some time to measure inflammation and cell damage in conditions like ulcerative colitis, microbicide developers need to have a more precise set of baseline measurements so that they can accurately calibrate the inflammatory and cytotoxic (cell-killing) potential of candidate microbicides.

This has already been done in vaginal and cervical tissues. However, the researchers say, “It is important to assess the rectal safety of vaginal microbicides…[and] similar studies are required as rectal-specific microbicides are developed.”

Ian McGowan and his team from the University of California, Los Angeles, recruited 16 gay volunteers: four HIV-negative men who had been having (protected) anal sex at least once a week; four HIV-negative men who had not had anal sex for at least two months; four HIV-positive men not on antiretrovirals; and four HIV-positive men on antiretroviral drugs (ARVs) with viral loads under 50. All the HIV-positive men recorded having anal sex, protected or otherwise.

The volunteers varied in their age and clinical experience. In the HIV-negative men, those who had not been having anal sex were older than those who did (average ages 47 and 38).

In the HIV positive men, those on ARVs were older than those not on treatment (average ages 44 versus 27) and length of diagnosis. Those on ARVs had lived with HIV for an average of 14 years (range, 7-22 years) while of the four not on potent anti-HIV therapy, one had been diagnosed for nearly seven years but the other three for less than a year. The group not taking treatment had an average HIV viral load of 96,000 copies/ml (range, 16,000-262,000). The CD4 counts in the two groups of HIV positive men were similar (average 600 cells/mm³ for those not on ARVs and 680 cells/mm³ for those on ARVs).

The volunteers were described by the researchers as “dedicated” because the study involved taking 20 tissue biopsies from two different sites, ten at each site, 10cm and 30cm from the anal entrance, three times within a month. They also took samples of rectal secretions.

The team then evaluated the samples for:

• Qualitative histological inflammation score, as established by microscopic examination of tissues
• The number of leucocytes (‘white blood cells’) and their type in each tissue sample
• The amount of soluble antibody protein in rectal secretions
• The amount of messenger RNA (small lengths of ‘signalling’ nucleic acid) that were specific signals for cells to produce three substances that control the inflammatory response. These are the cytokines interferon-gamma and interleukin-10 and the chemokine RANTES.

The first piece of good news was that microscopic examination found almost no evidence of overt inflammation in any subject, finding only four examples of even mild inflammation in any subject (two in the negative men who had anal sex and two in the men on ARVs) and then only at one site each on one occasion. Thus there is no evidence that anal intercourse in itself produces inflammation (men with conditions like diarrhoea, herpes, gonorrhoea and chlamydia that would cause inflammation were excluded from the study).

The study found relatively few differences in most leukocyte populations between the four groups. The significant differences were largely restricted to the HIV-positive men not taking ARVs.

These were, relative to the other groups:

• A generally higher level of lymphocytes (T-cells) of all kinds in the rectal lining tissue
• More CD8 (T-suppressor) cells, which kill off other infected cells
• More dendritic cells, which transport foreign tissues to the lymph nodes
• Fewer Natural Killer cells, a part of the faster-acting but less specific innate branch of the immune system
• Fewer CD4 cells but a higher than normal fraction of CD4 cells that were activated, as indicated by another maker called CD38. This study is the first direct evidence of higher CD38 levels in gut cells in untreated people with HIV. CD38 activity has been seen as an indicator of fast progression to AIDS.
• Fewer CD4 cells with either the CCR5 HIV co-receptor on their surface or both co-receptors (CCR5 and CXCR4).

Most of these values had normalised in the group taking ARVs. They still had somewhat lower CD4 and higher CD8 counts than the HIV negative men, and somewhat more dendritic cells, but the differences were only of borderline statistical significance, and all other cell subsets had normalised to the levels seen in HIV-negative subjects.

The HIV-negative men who had anal sex had somewhat higher general levels of leukocytes in the site near the anal entrance but were otherwise exactly the same as those who did not have anal sex.

The balance of T-cells (which kill other cells) and B-cells (which secrete antibodies) changed according to site, with more B-cells at the ‘shallow’ site and more T-cells at the ‘deep’ one, indicating that colonic tissue may be more susceptible to HIV infection than anal tissue.

These results were interesting from the point of view of the pathology of AIDS and the effect of treatment, but there was too much inter-subject variability for most of them to be used as baselines for microbicide studies. The most consistent figures were those for the messenger RNAs that signal cytokine production and to a lesser extent the total amount of IgA and IgG antibodies in rectal secretions.

Within the four groups but not between them, the relative proportions of CD4 and CD8 cells, and their proportions relative to the more general lymphocyte marker called CD3, also varied little, and could be possibly used as an indicator of inflammation.

There was such heterogeneity in the detailed immune states of each subject, the researchers cautioned, that it may prove difficult to develop a set of baselines against which to measure tissue inflammation any more sophisticated than that that provided by microscopic examination. Different people seem to have different gut ‘climates’, so the effect of introducing a foreign substance like a microbicide may be difficult to gauge.

This study also only looked at men and results might be different in women. However a larger study might have found more homogeneity of response.

In terms of telling us more about HIV infection and AIDS, however, this study adds to our knowledge both of the pathogenesis of AIDS and the beneficial effects of HIV treatment, with the researchers commenting that it has “confirmed the mucosal CD4 lymphopenia [deficit] and partial reconstitution reported with treated HIV infection.”