Kristina Allers

Long-Term Control of HIV by CCR5 Delta32/Delta32 Stem-Cell Transplantation

Infection with the human immunodeficiency virus type 1 (HIV-1) requires the presence of a CD4 receptor and a chemokine receptor, principally chemokine receptor 5 (CCR5). Homozygosity for a 32-bp deletion in the CCR5 allele provides resistance against HIV-1 acquisition. We transplanted stem cells from a donor who was homozygous for CCR5 delta32 in a patient with acute myeloid leukemia and HIV-1 infection. The patient remained without viral rebound 20 months after transplantation and discontinuation of antiretroviral therapy. This outcome demonstrates the critical role CCR5 plays in maintaining HIV-1 infection.

Evidence for the cure of HIV infection by CCR5Δ32/Δ32 stem cell transplantation

HIV entry into CD4(+) cells requires interaction with a cellular receptor, generally either CCR5 or CXCR4. We have previously reported the case of an HIV-infected patient in whom viral replication remained absent despite discontinuation of antiretroviral therapy after transplantation with CCR5Δ32/Δ32 stem cells. However, it was expected that the long-lived viral reservoir would lead to HIV rebound and disease progression during the process of immune reconstitution. In the present study, we demonstrate successful reconstitution of CD4(+) T cells at the systemic level as well as in the gut mucosal immune system after CCR5Δ32/Δ32 stem cell transplantation, while the patient remains without any sign of HIV infection. This was observed although recovered CD4(+) T cells contain a high proportion of activated memory CD4(+) T cells, ie, the preferential targets of HIV, and are susceptible to productive infection with CXCR4-tropic HIV. Furthermore, during the process of immune reconstitution, we found evidence for the replacement of long-lived host tissue cells with donor-derived cells, indicating that the size of the viral reservoir has been reduced over time. In conclusion, our results strongly suggest that cure of HIV has been achieved in this patient.

Impairment of the intestinal barrier is evident in untreated but absent in suppressively treated HIV-infected patients

Background and aims: Impairment of the gastrointestinal mucosal barrier contributes to progression of HIV infection. The purpose of this study was to investigate the effect of highly active antiretroviral therapy (HAART) on the HIV-induced intestinal barrier defect and to identify underlying mechanisms. Methods: Epithelial barrier function was characterised by impedance spectroscopy and [3H]mannitol fluxes in duodenal biopsies from 11 untreated and 8 suppressively treated HIV-infected patients, and 9 HIV-seronegative controls. The villus/crypt ratio was determined microscopically. Epithelial apoptoses were analysed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labelling (TUNEL) and caspase-3 staining. Tight junction protein expression was quantified by densitometric analysis of immunoblots. Mucosal cytokine production was determined by cytometric bead array. Results: Only in untreated but not in treated HIV-infected patients, epithelial resistance was reduced (13 (1) vs 23 (2) Ω cm2, p<0.01) and mannitol permeability was increased compared with HIV-negative controls (19 (3) vs 9 (1) nm/s, p<0.05). As structural correlates, epithelial apoptoses and expression of the pore-forming claudin-2 were increased while expression of the sealing claudin-1 was reduced in untreated compared with treated patients and HIV-negative controls. Furthermore, villous atrophy was evident and mucosal production of interleukin 2 (IL2), IL4 and tumour necrosis factor α (TNFα) was increased in untreated but not in treated HIV-infected patients. Incubation with IL2, IL4, TNFα and IL13 reduced the transepithelial resistance of rat jejunal mucosa. Conclusions: Suppressive HAART abrogates HIV-induced intestinal barrier defect and villous atrophy. The HIV-induced barrier defect is due to altered tight junction protein composition and elevated epithelial apoptoses. Mucosal cytokines are mediators of the HIV-induced mucosal barrier defect and villous atrophy.

Emergence of Minor Populations of Human Immunodeficiency Virus Type 1 Carrying the M184V and L90M Mutations in Subjects Undergoing Structured Treatment Interruptions

The use of structured treatment interruption (STI) in human immunodeficiency virus (HIV)-infected subjects is currently being studied as an alternative therapeutic strategy for HIV-1. The potential risk for selection of drug-resistant HIV-1 variants during STI is unknown and remains a concern. Therefore, the emergence of drug resistance in sequential plasma samples obtained from 28 subjects with chronic HIV infection was studied. They underwent 4 cycles of 2-week STI, followed by 8-week retreatment with highly active antiretroviral therapy identical to that used before STI, and they had never failed treatment before undergoing STI. At week 40, treatment was stopped for a longer period. Minor populations of drug-resistant variants were detected by quantitative real-time polymerase chain reaction, by use of allele-discriminating oligonucleotides for 2 key resistance mutations: L90M (protease) and M184V (reverse transcriptase). The approximate discriminative power was 0.1%. In 14 of 25 and in 3 of 25 subjects, the M184V and the L90M mutations, respectively, were detected as minor populations, at different times during STI. Overall, these results indicate that, in subjects undergoing multiple STIs, HIV-1 variants carrying drug-resistance mutations can emerge during periods of increased HIV-1 replication.

Impaired Immune Functions of Monocytes and Macrophages in Whipple's Disease

BACKGROUND & AIMS Whipples disease is a chronic multisystemic infection caused by Tropheryma whipplei. Host factors likely predispose for the establishment of an infection, and macrophages seem to be involved in the pathogenesis of Whipples disease. However, macrophage activation in Whipples disease has not been studied systematically so far. METHODS Samples from 145 Whipples disease patients and 166 control subjects were investigated. We characterized duodenal macrophages and lymphocytes immunohistochemically and peripheral monocytes by flow cytometry and quantified mucosal and systemic cytokines and chemokines indicative for macrophage activation. In addition, we determined duodenal nitrite production and oxidative burst induced by T whipplei and by other bacteria. RESULTS Reduced numbers of duodenal lymphocytes, increased numbers of CD163(+) and stabilin-1(+), reduced numbers of inducible nitric synthase+ duodenal macrophages, and increased percentages of CD163(+) peripheral monocytes indicated a lack of inflammation and a M2/alternatively activated macrophage phenotype in Whipples disease. Incubation with T whipplei in vitro enhanced the expression of CD163 on monocytes from Whipples disease patients but not from control subjects. Chemokines and cytokines associated with M2/alternative macrophage activation were elevated in the duodenum and the peripheral blood from Whipples disease patients. Functionally, Whipples disease patients showed a reduced duodenal nitrite production and reduced oxidative burst upon incubation with T whipplei compared with healthy subjects. CONCLUSIONS The lack of excessive local inflammation and alternative activation of macrophages, triggered in part by the agent T whipplei itself, may explain the hallmark of Whipples disease: invasion of the intestinal mucosa with macrophages incompetent to degrade T whipplei.

The Immune Reconstitution Inflammatory Syndrome in Whipple Disease: A Cohort Study

BACKGROUND Whipple disease, which is caused by infection with Tropheryma whipplei, can be treated effectively with antimicrobials. Occasionally, inflammation reappears after initial improvement; this is often interpreted as refractory or recurrent disease. However, polymerase chain reaction for T. whipplei in tissue is sometimes negative during reinflammation, indicating absence of vital bacteria, and this reinflammation does not respond to antimicrobials but does respond to steroids. OBJECTIVE To demonstrate that the immune reconstitution inflammatory syndrome (IRIS) occurs in patients treated for Whipple disease. DESIGN Cohort study. (International Standard Randomised Controlled Trial Number Register registration number: ISRCTN45658456) SETTING 2 academic medical centers in Germany. METHODS 142 patients treated for Whipple disease out of a cohort of 187 were observed for reappearance of inflammatory signs after effective antibiotic therapy. Definitions of IRIS in HIV infection, tuberculosis, and leprosy were adapted for application to Whipple disease. RESULTS On the basis of study definitions, IRIS was diagnosed in 15 of 142 patients. Symptoms included fever, arthritis, pleurisy, erythema nodosum, inflammatory orbitopathy, small-bowel perforation, and a hypothalamic syndrome. Two patients died. There was a positive correlation with previous immunosuppressive treatment and a negative correlation with previous diarrhea and weight loss. LIMITATIONS The study was observational and thus has inherent weaknesses, such as incomplete and potentially selective data recording. CONCLUSION The immune reconstitution inflammatory syndrome was diagnosed in about 10% of patients with Whipple disease in the study cohort; the outcome varied from mild to fatal. Patients who had had previous immunosuppressive therapy were at particular risk. An immune reconstitution syndrome should be considered in patients with Whipple disease in whom inflammatory symptoms recur after effective treatment. Early diagnosis and treatment with steroids may be beneficial; prospective studies are needed. PRIMARY FUNDING SOURCE European Commission and Deutsche Forschungsgemeinschaft.

Rapid selection of drug-resistant HIV-1 during the first months of suppressive ART in treatment-naive patients

Objective: Efficient antiretroviral therapy (ART) of HIV-1 infection reduces the viral load to undetectable levels and restores the immune system. However, therapy failure appears in a substantial fraction of patients and is mostly associated with the appearance of drug-resistant viruses. It is still not clear when the drug pressure leads to the earliest selection and appearance of drug-resistant HIV-1 populations. In this study, we wanted to determine whether drug-resistant viruses are already selected during viral decline within the first months of ART. Design and methods: Fifteen mostly chronically HIV-1 infected patients were included. None had received ART prior to this study. The selection of three key resistance mutations, L90M (protease), K103N and M184V (reverse transcriptase), were measured by allele-specific real-time PCR allowing us to track minority quasispecies with a discriminative power of 0.01–0.2%. Results: Drug-resistant HIV-1 variants were found in 7/15 patients (46.7%) prior to ART. Rapid selection of drug resistance was detected in six patients (40%) independent of the presence of drug-resistant HIV-1 prior to ART. The risk for the selection of drug resistant viruses was correlated with the time until viral load became undetectable (P = 0.02). Besides the proportional increment of drug-resistant viruses, we observed in two patients a quantitative increase of this virus population while the total viral load decreased. Conclusions: Drug-resistant viruses can be selected and replicate even in the first weeks of suppressive ART, thus, intensification of ART during the initial treatment period should be considered and further evaluated in clinical studies.

Macrophages Accumulate in the Gut Mucosa of Untreated HIV-infected Patients

BACKGROUND Mucosal macrophages are involved in the maintenance of epithelial barrier integrity and the elimination of invading pathogens. Although an intestinal barrier defect and microbial translocation are hallmarks of human immunodeficiency virus (HIV) infection, recent data on gut mucosal macrophages in HIV infection are sparse. METHODS Treatment-naive and treated HIV-infected patients and healthy controls were studied for frequencies and functional parameters of blood monocytes and macrophages in duodenal mucosa. RESULTS We found mucosal enrichment of macrophages in untreated HIV infection associated with reduced monocyte counts in blood and increased monocyte expression of the gut-homing molecule integrin β7. Increased CCR2 density on integrin β7-expressing monocytes and mucosal secretion of CCL2 suggest that CCR2/CCL2-chemotaxis is involved in enhanced trafficking of blood monocytes to the gut. Secretion of macrophage-related proinflammatory molecules interleukin 1β, CCL5, CXCL9, and CXCL10 was increased in the gut mucosa of untreated patients. Moreover, mucosal macrophages of untreated patients showed reduced phagocytic activity. CONCLUSIONS These data suggest a role for gut mucosal macrophages in HIV immune pathogenesis: infiltrated macrophages in the intestinal mucosa may promote local inflammation and tissue injury, whereas their low phagocytic activity prevents the efficient elimination of luminal antigens that cross the damaged intestinal barrier.

Gut Mucosal FOXP3+ Regulatory CD4+ T Cells and Nonregulatory CD4+ T Cells Are Differentially Affected by Simian Immunodeficiency Virus Infection in Rhesus Macaques

ABSTRACT The gastrointestinal tract represents a major site for human and simian immunodeficiency virus (HIV and SIV) replication and CD4+ T-cell depletion. Despite severe depletion of mucosal CD4+ T cells, FOXP3+ regulatory CD4+ T cells (Treg) are highly increased in the gut mucosa of chronically HIV-infected individuals and may contribute to HIV pathogenesis, either by their immunosuppressive function or as a significant target cell population for virus production. Little is known about the susceptibility of mucosal Treg to viral infection and the longitudinal effect of HIV/SIV infection on Treg dynamics. In this study, we determined the level of SIV infection in Treg and nonregulatory CD4+ T cells (non-Treg) isolated from the colon of SIV-infected rhesus macaques. The dynamics of mucosal Treg and alterations in the mucosal CD4+ T-cell pool were examined longitudinally. Our findings indicate that mucosal Treg were less susceptible to productive SIV infection than non-Treg and thus were selectively spared from SIV-mediated cell death. In addition to improved survival, local expansion of Treg by SIV-induced proliferation of the mucosal CD4+ T-cell pool facilitated the accumulation of mucosal Treg during the course of infection. High frequency of mucosal Treg in chronic SIV infection was strongly related to a reduction of perforin-expressing cells. In conclusion, this study suggests that mucosal Treg are less affected by productive SIV infection than non-Treg and therefore spared from depletion. Although SIV production is limited in mucosal Treg, Treg accumulation may indirectly contribute to viral persistence by suppressing antiviral immune responses.

Modulation of the CD4+ T-Cell Response by Helicobacter pylori Depends on Known Virulence Factors and Bacterial Cholesterol and Cholesterol α-Glucoside Content

Helicobacter pylori blocks the proliferation of human CD4(+) T cells, facilitated by vacuolating exotoxin (VacA) and γ-glutamyl transpeptidase (GGT). H. pylori-triggered T-cell reactions in mice correlate with bacterial cholesterol and cholesterol α-glucoside content but their role in human cells is unclear. We characterized the effect of VacA, GGT, and cholesterol on T-helper 1, T-helper 2, T-regulatory and T-helper 17 associated cytokines and T-cell proliferation. VacA, GGT, and bacterial cholesterol content exhibited differential and synergistic inhibitory effects on the expression of activation markers CD25 and CD69 and on interleukin 2, interleukin 4, interleukin 10, and interferon γ production. These factors did not affect the H. pylori-mediated abrogation of transforming growth factor β secretion or increased interleukin 6 production. Cholesterol α-glucosyltransferase-deficient bacteria exerted strongly reduced antiproliferative effects on primary human CD4(+) T cells. In conclusion, H. pylori shapes rather than suppresses human CD4(+) T-cell responses, and glucosylated cholesterol is a relevant bacterial component involved in this modulation.