Thomas Zippel

HIV-1 p24 but not proviral load is increased in the intestinal mucosa compared with the peripheral blood in HIV-infected patients

Objective: To investigate differences in viral and proviral load between the peripheral blood and the intestinal mucosal immune system in HIV-infected patients. Design: HIV-1 p24 and HIV DNA content were compared in blood samples and intestinal biopsies from HIV-infected patients. Methods: Intestinal biopsies and peripheral blood were simultanously obtained from 27 HIV-infected patients undergoing diagnostic endoscopy. The p24 concentrations were measured in serum and homogenized intestinal biopsies by enzyme-linked immunosorbent assay after acid-dissociation of immune complexes. Proviral load was determined in blood and intestinal biopsies by a quantitative competitive polymerase chain reaction amplifying the HIV-1 nef gene from genomic DNA. Results: No significant differences were found in proviral load comparing HIV copies per 1.5 x 105 cell equivalents in blood [2650 (600–44 000)] and intestinal biopsies [4200 (1325–19 625)]. Paired analysis revealed a strong positive correlation between serum and mucosal proviral load. In contrast, HIV core protein p24 was detected in intestinal biopsies from 18 patients in much higher concentrations than in serum [858 (262–4111) pg/g versus 34 (9–242) pg/g; P < 0.005]. The p24 concentrations in serum and intestinal biopsies did not correlate and no significant correlation was observed in serum or intestinal biopsies between proviral load and p24 concentrations. No clear correlations were observed between clinical parameters and HIV DNA or HIV p24 levels in blood or biopsies. Conclusions: Our findings demonstrate a homogenous distribution of HIV proviral load in the peripheral blood and the intestinal mucosal immune system. The high viral antigen load in the intestine therefore indicates that mucosal HIV production is upregulated at the transcriptional and/or translational level. The intestinal mucosa is a major reservoir for HIV in HIV-infected patients.

Mucosal HIV Infection

The gastrointestinal tract is not only a major site of clinical manifestations of the acquired immune deficiency syndrome but also an important compartment for HIV infection at all stages of the disease. In intestinal biopsies HIV has been detected so far mainly in mononuclear cells of the lamina propria but also in epithelial cells. Primary and permanent epithelial cell lines can be productively infected by HIV in vitro and epithelial cell monolayers can rapidly transport HIV by transcytosis. Both phenomena are most effective for cell-associated virus and do not require CD4 expression but seem to involve the interaction of gp120 with galactosyl ceramide. In vivo, productive HIV infection of epithelial cells appears to be rare if it occurs at all. Mucosal HIV infection is highly active at all stages of HIV infection and the gastrointestinal tract is probably a major source of HIV in the body. Highest mucosal HIV production is found rather early in HIV disease and is associated with histological abnormalities and gastrointestinal symptoms. The excessive production of HIV in the intestine appears to be due to transcriptional or translational upregulation and local variations in HIV production correlate with local cytokine levels.

Increased immunoglobulin G production by short term cultured duodenal biopsy samples from HIV infected patients

Background—Secretory immunity is a major defence mechanism against infections at mucosal surfaces which are common in HIV infected patients. Aims—To analyse intestinal immunoglobulin production in HIV infection in comparison with that in saliva and serum. Patients and methods—Immunoglobulin G (IgG), A (IgA), and M (IgM) concentrations were determined in supernatants of short term cultured duodenal biopsy samples, serum, and saliva from HIV infected patients (n = 28) and controls (n = 14) by radial immunodiffusion. Results—IgG was increased in the supernatants of short term cultured biopsy samples and saliva from HIV infected patients compared with controls (p<0.01), but IgA and IgM levels were normal. In contrast, both IgG and IgA concentrations in serum were higher in HIV infected patients than in controls (p<0.002). No correlation was found between IgA produced by duodenal biopsy specimens and serum IgA. Conclusion—Abnormalities in mucosal immunoglobulin production in HIV infection were suprisingly small, indicating that specific secretory immunity rather than quantitative immunoglobulin production may be impaired. However, increased production of IgG could contribute to mucosal inflammation by complement activation. Our findings of normal mucosal IgA production and the lack of correlation between serum and mucosal IgA argues against an intestinal origin for the increased serum IgA levels in HIV infected patients.

Cytokine Expression in the Colonic Mucosa of Human Immunodeficiency Virus-Infected Individuals before and during 9 Months of Antiretroviral Therapy

ABSTRACT High-level human immunodeficiency virus (HIV) replication and the rapid breakdown of the mucosal immune system are the hallmarks of HIV infection in the gut. Cytokine dysregulation may be related to both phenomena. Using real-time PCR we quantified the colonic mucosal mRNA expression of selected proinflammatory and regulatory (gamma interferon [IFN-gamma], tumor necrosis factor alpha [TNF-α], and interleukin-2 [IL-2], IL-4, IL-6, and IL-10) and HIV-inhibitory (IL-16, CCL3, and CCL5) cytokines for 10 HIV-infected patients before and during 9 months of highly active antiretroviral therapy (HAART). HIV RNA and T-cell dynamics were measured in the colonic mucosa and the blood. Seven HIV-negative individuals served as controls. The mucosal mRNA expression of TNF-α, IFN-gamma, IL-4, IL-6, and IL-10 was significantly higher in HIV-infected patients than in control patients and remained elevated during 9 months of HAART despite the decline in blood and mucosal HIV RNA levels and an increase in the level of CD4+ T lymphocytes. The mRNA levels of CCL3 and CCL5, both of which were elevated before treatment, returned to nearly normal during therapy. Despite reductions in levels of mucosal HIV RNA and the restoration of mucosal CD4+ T lymphocytes, antiretroviral therapy failed to restore the normal colonic immunologic environment.

Abnormal predominance of IgG in HIV-specific antibodies produced by short-term cultured duodenal biopsy specimens from HIV-infected patients

The aim of our study was to analyze HIV-specific humoral immunity in the intestinal mucosa at different stages of HIV infection in comparison with serum and saliva. Duodenal biopsy specimens from 30 AIDS patients and 9 HIV-infected patients without AIDS were cultured for 48 hours. Culture supernatants, as well as simultaneously obtained serum and saliva samples, were adjusted to the same immunoglobulin concentrations and tested for HIV-specific IgG and IgA by Western blot. The HIV antigen pattern differed clearly between IgA and IgG but was similar for each isotype independent of its origin (i.e., serum, saliva, or biopsy specimen supernatants). Short-term cultured duodenal biopsy specimens from HIV-infected patients at all stages produced predominantly IgG, which was broadly reactive with HIV antigens. Lower titers of HIV-specific IgA, which recognized few antigens, were found, mostly the glycoprotein gp160. At later stages of the disease compared with earlier stages, the reaction pattern of mucosal IgA from saliva and biopsy supernatants was even more restricted; secretory component was frequently absent. The abnormal predominance of HIV-specific IgG over IgA in mucosal secretions may result from abnormal antibody production in the mucosa rather than from serum leakage. Mucosal inflammation induced by HIV-IgG immune complexes and insufficient immune exclusion by secretory IgA may not only lead to increased mucosal HIV replication but may also contribute to gastrointestinal disease in HIV-infected patients.

Secretory Immunity in HIV Infection

Secretory IgA plays a crucial role in the defense of pathogens at mucosal surfaces. As CD4+ T cells are lost early in the mucosa of human immunodeficiency virus (HIV)-infected patients and as CD4+ T cells play an essential role in the regulation of specific IgA responses to pathogenic agents at mucosal sides, it could be expected that this first line of defense is impaired in HIV-infected patients. Therefore, several studies were undertaken to characterize the humoral immune response at mucosal surfaces. However, the results obtained so far are in part contradictory. For intestinal IgA, reduced, increased and no changes compared to controls were described. The different results may be due to different methods applied. In most studies an abnormal predominance of HIV-specific IgG over IgA response was found in the intestine of HIV-infected patients. Studies on cytomegalovirus-specific intestinal antibodies indicate a complete lack of a specific intestinal IgA response. However, in cryptosporidiosis of HIV-infected patients, diarrhea persists despite a secretory IgA response indicating that other factors are also important for the clearance of this pathogen.

SERUM ANTIBODIES TO DIETARY ANTIGENS IN PATIENTS WITH HIV-1 INFECTION

Diarrhoea is a common complication of HIV-1 infection [1], but in a considerable proportion of patients, especially in earlier stages of the disease, the cause of diarrhoea remains unclear despite extensive investigation [2]. Increased levels of serum antibodies against food proteins have been described in HIV-infected children [3], similar to patients with non-IgE-mediated gastrointestinal food hypersensitivity [4], coeliac disease [5], or chronic inflammatory bowel disease [6]. These abnormalities are thought to result from increased gut permeability leading to increased uptake of dietary antigens and an aberrant mucosal IgG response in these patients [7]. Increased intestinal permeability [8] and an increase of IgG in mucosal secretions [9] have also been found in patients infected with HIV-1. Since abnormal immunity to dietary antigens may contribute to the pathogenesis of diarrhoea, we investigated serum antibodies to food proteins in HIV-1-infected patients with and without diarrhoea.