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Cell-free and cell-associated human immunodeficiency virus (HIV) may differently affect the immune system and the efficacy of prevention strategies. Here we examined mucosal events in simian immunodeficiency virus (SIV) infection, using infected cells together with cell-free virus and cell-free virus alone. Intravenously inoculated SIV-infected cells disseminated virus to the intestine within 16 hours. Infection with both virus forms accelerated viral dissemination in the intestinal mucosa and the loss of mucosal CD4+ T cells as compared to infection with cell-free virus only. As all natural sources of HIV infection contain both virus forms, future prevention studies should focus on efficacy against both cell-free and cell-associated virus.

Background: Understanding the mechanism(s) by which broadly neutralizing antibodies (bNAbs) emerge naturally following infection is crucial for the development of a protective vaccine against human immunodeficiency virus (HIV). Although previous studies have implicated high viremia and associated immune activation as potential drivers for the development of bNAbs, here we sought to unlink the effect of these 2 parameters by evaluating the key inflammatory predictors of bNAb development in HIV-infected individuals who spontaneously control HIV in the absence of antiretroviral therapy ("controllers")., Methods: The breadth of antibody-mediated neutralization against 11 tier 2 or 3 viruses was assessed in 163 clade B spontaneous controllers of HIV. Plasma levels of 17 cytokines were screened in the same set of subjects. The relationship of the inflammatory signature was assessed in the context of viral blips or viral RNA levels in peripheral blood or gastrointestinal biopsies from aviremic controllers (<50 copies RNA/mL) and in the context of viral sequence diversity analysis in the plasma of viremic controllers (<50-2000 copies RNA/mL)., Results: A unique inflammatory profile, including high plasma levels of CXCL13, sCD40L, IP10, RANTES, and TNFα, was observed in HIV controllers who developed bNAbs. Interestingly, viral load and tissue viremia, but not intermittent viral blips, were associated with these cytokine profiles. However, viral diversity was not significantly associated with increased breadth in controllers., Conclusion: These results suggest that low antigenic diversity in the setting of a unique inflammatory profile associated with antigen persistence may be linked to the evolution of neutralizing antibody breadth., (© The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Background: Inflammatory bowel disease (IBD) patients on thiopurine therapy are at increased risk of Epstein-Barr virus (EBV)-associated lymphomas. This virus is frequently detected in the intestinal mucosa of IBD patients and may cause a wide spectrum of lymphoproliferations similar to post-transplantation lymphoproliferative disorders (PTLDs). We aimed to assess whether histological aberrations aid in predicting EBV presence and to correlate histological assessment and EBV load with disease outcome in IBD., Methods: We included all IBD patients from our centre who underwent EBV testing of intestinal biopsies between January 2004 and October 2013. All biopsies were classified according to the WHO PTLD classification and the EBV load was scored per high-power field (HPF). Clinical data were collected from patient charts. Reported clinical outcomes included colectomy, need for chemotherapy and mortality., Results: Our cohort included 58 patients: 28 were EBV-positive and 30 EBV-negative. An atypical infiltrate was seen more frequently in EBV-positive than in EBV-negative patients (57.1 versus 3.3%; p < 0.001). A high EBV load occurred more frequently in EBV-positive patients undergoing colectomy than in EBV-positive patients without colectomy (50.0 versus 10.0%; p = 0.048). Monomorphic lymphoproliferative disorders, including two overt lymphomas, were present in 10 patients. Reduction of immunosuppression resulted in histological normalization and loss of EBV expression in seven of eight non-lymphoma patients., Conclusion: The presence of atypical infiltrate in the intestinal mucosa of IBD patients warrants EBV testing. Reduction of immunosuppression is an effective strategy to achieve morphological normalization and loss of EBV. Lymphoproliferation related to IBD appears to have less aggressive clinical behaviour than PTLDs., (Copyright © 2015 European Crohn’s and Colitis Organisation (ECCO). Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

We studied 3 patients with focal intra-anal tissue high-grade squamous intraepithelial lesions (HSILs). All had increased human immunodeficiency virus type 1 (HIV-1) RNA and DNA in lesions compared with that in healthy mucosa. HIV-1 RNA and HIV-1 episomal DNA were indicative of ongoing viral replication, more so in anal HSILs.

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.

Background and Aim: Cytomegalovirus (CMV) exacerbates ulcerative colitis (UC) refractory to immunosuppressive therapies. The conditions under which CMV reactivation occurs in patients with UC, however, is unclear. In addition, the diagnostic and treatment strategies for UC positive for CMV have not been established. Granulocyte and monocyte adsorptive apheresis (GMAA) is natural biological therapy for UC in which the granulocytes/macrophages producing inflammatory cytokines are removed. We investigated the rate of colonic CMV reactivation and the efficacy of GMAA in active UC patients positive for CMV without concomitant corticosteroid (CS) therapy., Methods: Fifty-one active UC patients without concomitant CS therapy were enrolled. Colonic CMV reactivation was examined by real-time polymerase chain reaction (PCR) using biopsy specimen and/or histological examination. All patients were treated with intensive GMAA (twice per week). Rates of clinical remission and mucosal healing were compared between UC patients positive and negative for CMV., Results: Of 51 patients, 15 (29.4%) were diagnosed as CMV positive. The clinical remission rates following intensive GMAA did not differ between UC patients positive and negative for CMV (73.3% vs 69.4%, p=0.781). Proportion of patients achieving mucosal healing was also similar between these two groups. CMV-DNA became negative in all UC patients positive for CMV who achieved clinical remission 1 week after completion of intensive GMAA., Conclusions: Intestinal inflammation might trigger CMV reactivation in a subpopulation of active UC patients without CS treatment. GMAA could be a promising option for active UC positive for CMV., (Copyright © 2012 European Crohn's and Colitis Organisation. Published by Elsevier B.V. All rights reserved.)

Objectives: Precise characterization of viruses present in reservoirs in long-term pretreated patients will be a major issue to consider in the context of viral eradication. We assessed the frequency of defective viruses present in cellular reservoirs., Methods: Peripheral blood mononuclear cells (PBMCs) and rectal biopsy samples were compared between five patients on successful long-term highly active antiretroviral therapy (HAART) (>7 years without blips) and five untreated patients. Molecular cloning and sequencing of the reverse transcriptase region were used to detect the presence of and quantify in-frame stop codons in HIV quasi-species. The relationship between the size of the reservoir and the frequency of defective genomes was assessed., Results: Defective genomes were systematically detected in all patients on long-term HAART in both compartments (PBMCs and rectal tissues), with a higher level of defective genomes per sample compared with PBMCs of untreated patients. A high level of defective genomes was correlated with a small size of HIV proviral DNA. Regarding the nucleotide context, guanine (G) to adenine (A) substitution at tryptophan positions was responsible for the appearance of 89% of all in-frame stop codons in the context of G-to-A hypermutation, likely reflecting APOBEC3 footprints on the viral genome., Conclusions: We propose a scenario whereby defective genomes accumulate during HAART treatment, eventually reaching a viral extinction threshold. In the context of viral eradication, measurement of the relative amounts of defective and non-defective viruses (by molecular cloning and ultradeep sequencing) should be used as a new criterion for eradicating HIV.

The progression of cervical intraepithelial lesions to invasive cancer is associated with corresponding reductions in human papillomavirus (HPV) L1 capsid antigen (L1) expression. We sought to determine whether a similar loss of L1 occurs during anal carcinogenesis using immunohistochemistry on paraffin-embedded sections as well as INNO-LiPA HPV Genotyping (Innogenetics, Gent, Belgium) technology to determine HPV infection status. We analyzed 31 squamous cell carcinomas (SCCs), 26 SCCs in situ (SCC-IS), and 11 normal anal mucosae from 36 patients. High-risk HPV subtypes were detected in all patients. L1 nuclear staining was identified in 38% of SCC-IS; however, there was no detection in normal anal mucosae, SCC, or recurrent SCC. Of those SCC-IS associated with a concomitant invasive SCC, only 15% demonstrated nuclear L1 expression as compared to 62% of isolated SCC-IS (P = .02). Nuclear expression of L1 is lost in the progression of anal SCC-IS to SCC and may serve as a possible prognostic marker of enhanced malignant potential.

The human respiratory tract is a major site of avian influenza A(H5N1) infection. However, many humans infected with H5N1 present with gastrointestinal tract symptoms, suggesting that this may also be a target for the virus. In this study, we demonstrated that the human gut expresses abundant avian H5N1 receptors, is readily infected ex vivo by the H5N1 virus, and produces infectious viral particles in organ culture. An autopsy colonic sample from an H5N1-infected patient showed evidence of viral antigen expression in the gut epithelium. Our results provide the first evidence, to our knowledge, that H5N1 can directly target human gut tissues.

Background: Worldwide, approximately 90% of all human immunodeficiency virus (HIV) transmissions occur mucosally; almost all involve R5 strains. Risks of sexual HIV acquisition are highest for rectal, then vaginal, and finally oral exposures., Methods: Mucosal lacerations may affect the rank order of susceptibility to HIV but cannot be assessed in humans. We measured relative virus transmissibility across intact mucosae in macaques using a single stock of SHIV-1157ipd3N4, a simian-human immunodeficiency virus encoding a primary R5 HIV clade C env (SHIV-C)., Results: The penetrability of rhesus macaque mucosae differed significantly, with rectal challenge requiring the least virus, followed by vaginal and then oral routes (P = .031, oral vs vaginal; P < .001 rectal vs vaginal). These findings imply that intrinsic mucosal properties are responsible for the differential mucosal permeability. The latter paralleled the rank order reported for humans, with relative risk estimates within the range of epidemiological human studies. To test whether inflammation facilitates virus transmission--as predicted from human studies--we established a macaque model of localized buccal inflammation. Systemic infection occurred across inflamed but not normal buccal mucosa., Conclusion: Our primate data recapitulate virus transmission risks observed in humans, thus establishing R5 SHIV-1157ipd3N4 in macaques as a robust model system to study cofactors involved in human mucosal HIV transmission and its prevention.

To dissect the early events in the transmission of HIV-1 from mother to child, we investigated whether DCs participate in HIV-1 entry into human small intestinal mucosa. We isolated human MNLs from jejunal lamina propria and identified a subpopulation of CD11c(+)HLA-DR(+) MNLs that expressed DC-SIGN, CD83, CD86, CD206, and CCR7, indicating a DC phenotype. Jejunal DCs also expressed the HIV-1 receptor CD4 and coreceptors CCR5 and CXCR4 and in suspension rapidly took up cell-free HIV-1. HIV-1 inoculated onto the apical surface of explanted jejunum was transported by lamina propria DCs through the mucosa and transmitted in trans to blood and intestinal lymphocytes. These findings indicate that in addition to intestinal epithelial cells, which we showed previously transcytose infectious HIV-1 to indicator cells, intestinal DCs play an important role in transporting HIV-1 through the intestinal mucosa and the subsequent transmission to T cells.

Despite suppression of human immunodeficiency virus (HIV) replication by antiretroviral therapy, reconstitution of CD4+ cells is variable and incomplete, particularly in gut-associated lymphatic tissues (GALT). We have previously shown that immune activation and inflammation in HIV-infected and simian immunodeficiency virus-infected lymph nodes results in collagen deposition and disruption of the lymphatic tissue architecture, and this damage contributes to CD4+ cell depletion before treatment and affects the extent of immune reconstitution after treatment. In the present study, we compared collagen deposition and the extent of depletion and reconstitution of total CD4+ cells and subsets in peripheral blood, lymph nodes, and inductive and effector sites in GALT. We show that CD4+ cell depletion in GALT correlates with the rapidity and greater magnitude of collagen deposition in this compartment, compared with that in peripheral lymph nodes, and that although treatment does not restore CD4+ cells to effector sites, treatment in the early stages of infection can increase CD4+ central memory cells in Peyer patches.

Background: Human herpesvirus 6 (HHV-6) infections are usually asymptomatic reactivations in adult liver transplant recipients, but they may also cause fever or graft dysfunction. HHV-6 infection can also present symptoms of gastroenteritis. In this study, we investigated the presence of HHV-6 in the gastroduodenal mucosa of liver transplant recipients and in immunocompetent patients undergoing gastroscopic examination because of dyspeptic symptoms., Methods: HHV-6 and cytomegalovirus (CMV) examinations were performed on gastroduodenal biopsy specimens obtained during upper gastrointestinal endoscopic examinations from 90 liver transplant recipients and from 31 immunocompetent patients with upper gastrointestinal symptoms. In the gastroduodenal mucosa, HHV-6 and CMV was demonstrated by immunohistochemistry in frozen sections using monoclonal antibodies against HHV-6- and CMV-specific antigens., Results: HHV-6-positive cells were found in biopsy specimens from 21 (23%) of the liver transplant recipients and 6 (19%) of the immunocompetent patients, CMV-positive cells were found in specimens from 55 (61%) of the transplant recipients and 7 (23%) of the immunocompetent patients, and 12 transplant recipients were found to have both HHV-6 and CMV infection. Fifteen transplant recipients with positive HHV-6 findings in the gastroduodenal mucosa also had HHV-6 antigenemia, whereas 30 patients with HHV-6 antigenemia did not have gastroduodenal involvement. Endoscopic findings in these patients included biliary complications in 10 patients and gastritis in 2 patients. Histopathological findings were nonspecific and included very mild inflammation. A total of 30 (94%) of the transplant recipients with biliary complications also had HHV-6 or CMV detected in the duodenal mucosa., Conclusions: HHV-6-positive cells and CMV-positive cells were frequently found in the gastroduodenal mucosa of liver transplant recipients and of immunocompetent patients undergoing gastroscopic examination because of dyspeptic symptoms.

The enteropathic manifestations of the human immunodeficiency virus (HIV) and the simian immunodeficiency virus (SIV) in late infection are usually due to infection by other microbes, but in early infection the viruses themselves cause an enteropathy by heretofore undetermined mechanisms. Here we report that SIV induces massive apoptosis of intestinal epithelial cells lining the small and large bowel, thus identifying apoptosis as the driving force behind the regenerative pathology of early infection. We found that apoptosis of gut epithelium paralleled the previously documented apoptosis and massive depletion of CD4 T cells in gut lamina propria, triggered by established mechanisms of gut epithelial cell apoptosis and, at peak, possibly by virus interactions with GPR15/Bob, an intestinal epithelial cell-associated alternative coreceptor for SIV and HIV-1. Apoptosis in early SIV infection is thus the common theme of the pathological processes that quickly afflict the innate as well as adaptive arms of the gut immune system.

Enterovirus infections have been diagnosed more frequently in type 1 diabetic patients than in the healthy population, and enteroviruses have also been found in the pancreas of diabetic patients. Primary replication of the virus occurs in the gut, but there are no previous studies evaluating possible presence of virus in the intestine of diabetic patients. The purpose of this study was to investigate if enteroviruses can be found in small intestinal tissue of type 1 diabetic patients. Formalin-fixed, paraffin-embedded upper intestinal biopsy samples were analysed for the presence of enterovirus using in situ hybridization and immunohistochemistry. Enterovirus was detected by in situ hybridization in six (50%) of the type 1 diabetic patients (n = 12) but in none of the control subjects (n = 10, P = 0.015). Immunohistochemistry identified enterovirus in nine (75%) of the patients and one (10%) control subject (P = 0.004). The presence of the virus was confirmed by reverse transcription-polymerase chain reaction in one of the four patients from whom a frozen and unfixed sample was available. Intestinal morphology was normal in all study subjects. The results suggest that a substantial proportion of type 1 diabetic patients have an ongoing enterovirus infection in gut mucosa, possibly reflecting persistent enterovirus infection. This observation opens new avenues for further studies on the possible role of enteroviruses in human type 1 diabetes.

Background: Studies suggest that cytomegalovirus (CMV) infection exacerbates ulcerative colitis (UC) refractory to immunosuppressive therapies. Early and accurate diagnosis of CMV infection is important for the treatment of UC. We evaluated the usefulness of quantitative real-time polymerase chain reaction (PCR) for detecting CMV infection in inflamed colonic mucosa of patients with UC refractory to immunosuppressive therapies., Methods: From 2003 to 2006, 30 patients (mean age: 41 +/- 18 years; 14 men, 16 women) with UC refractory to immunosuppressive therapies were enrolled in the study. We evaluated CMV infection by CMV antigenemia, histologic examination, and quantitative real-time PCR for CMV using colonic mucosa and investigated the clinical outcomes of antiviral therapy., Results: CMV-DNA was detected only in the inflamed colonic mucosa in 17 (56.7%) of 30 patients. Of the 17 CMV-DNA-positive patients, 4 were positive for CMV antigenemia or inclusion bodies on histologic examination; of the 13 CMV-DNA-negative patients none was positive for CMV antigenemia or inclusion bodies. Of the 17 CMV-DNA-positive patients, 12 (70.6%) were treated with ganciclovir for 2 weeks and 10 patients went into remission. Two other patients required colectomy after antiviral therapy. In contrast, of the 13 CMV-DNA-negative patients 12 (92.3%) achieved remission after intensifying their immunosuppressive therapies., Conclusions: Quantitative real-time PCR assay for detecting CMV-DNA is useful for early, accurate diagnosis of CMV infection in patients with UC refractory to immunosuppressive therapies, enabling prompt and appropriate treatment.

Excluding parenteral transmissions, virtually all vertical and homosexual transmission of human immunodeficiency virus type 1 (HIV-1) occurs via the gastrointestinal tract. Cellular routes implicated in the translocation of virus across the epithelium include M cells, dendritic cells, and epithelial cells. Intestinal epithelial cells express CCR5 and can selectively transfer CCR5-tropic HIV-1, the phenotype of the majority of transmitted viruses. In the lamina propria, virus encounters the largest reservoir of mononuclear cells in the body. Surprisingly, lamina propria lymphocytes, not macrophages, express CCR5 and CXCR4 and support HIV-1 replication, implicating intestinal lymphocytes as the initial target cell in the intestinal mucosa. From the mucosa, virus is disseminated to systemic sites, followed by profound depletion of CD4+ T cells, first in the intestinal lamina propria and subsequently in the blood. As mucosal and circulating CD4+ T cells are depleted, monocytes and macrophages assume an increasingly important role as target and reservoir cells for HIV-1. Blood monocytes, including HIV-1-infected cells, are recruited to the mucosa, where they differentiate into lamina propria macrophages in the presence of stroma-derived factors. Although the prevalence of HIV-1-infected macrophages in the mucosa is low (0.06% of lamina propria mononuclear cells), the extraordinary size of the gastrointestinal mucosa imparts to intestinal macrophages a prominent role as a HIV-1 reservoir. Elucidating the immunobiology of mucosal HIV-1 infection is critical for understanding disease pathogenesis and ultimately for devising an effective mucosal HIV-1 vaccine.

Primary prostate and cervical epithelial cells and epithelial cell lines were examined for human immunodeficiency virus type 1 (HIV-1) infection or transmission to peripheral blood mononuclear cells (PBMC). Neither cell-free nor cell-associated HIV-1 infected primary epithelial cells or cell lines. Pretreatment of HIV-1 to enhance CD4-independent entry did not augment infection. Cell surface expression was detected for galactosyl ceramide but not for CC-chemokine receptor 5, CXC-chemokine receptor 4, or CD4. The ability to transfer HIV-1 to resting or activated PBMC was tested by culturing with rinsed or trypsinized and replated HIV-1-exposed epithelial cells. Virus was not recovered from the rinsed or replated cocultures with resting PBMC; however, activated PBMC recovered HIV-1 from rinsed epithelial cells and rarely from replated epithelial cells. Although urogenital epithelial cells are not infected, these data suggest that they can transfer virus to activated immune cells and have implications for sexual transmission of HIV-1.

The gastrointestinal mucosa is a major lymphoid tissue reservoir for human immunodeficiency virus (HIV) replication. Genotypic and phenotypic resistance patterns of HIV type 1 (HIV-1) RNA isolated from colonic mucosa were compared with those from the plasma and peripheral blood mononuclear cells (PBMC) of 7 patients. Genotyping was performed using full-sequence analysis, and phenotyping was performed using a recombinant virus assay. Mutations in the reverse-transcriptase (kappa=.84) and protease (kappa=.73) genes were highly concordant among compartments. Similarly, phenotypic resistance patterns were highly concordant among compartments (intraclass correlation coefficient,.91). In 5 instances among 3 patients, a different genotypic result was observed between plasma and the other tissue compartments. Mixtures of wild-type and mutated HIV-1 RNA were present in the mucosa and PBMC but not in the plasma. Despite significant concordance among compartments, mucosal- and PBMC-derived viral RNA showed instances of discordance with plasma-derived virus that may suggest compartmentalization of virus.

Most human immunodeficiency virus type 1 (HIV-1) infections are acquired via mucosal surfaces, and transmitted viruses are nearly always macrophage-tropic, suggesting that mucosal macrophages participate in early HIV-1 infection. Mucosal lymphocytes isolated from normal human intestine expressed CD4 (14,530+/-7970 antibody-binding sites [ABSs]/cell), CCR5 (2730+/-1524 ABSs/cell), and CXCR4 (2507+/-1840 ABSs/cell), but intestinal macrophages, which also expressed CD4 (2959+/-2695 ABSs/cell), displayed no detectable CCR5 or CXCR4 ABS. The absence of CCR5 on intestinal macrophages was not due to expression of the Delta32 deletion allele because matched-blood monocytes expressed CCR5. CCR5(+)CXCR4(+) intestinal lymphocytes supported both R5 (BaL) and X4 (IIIB) HIV-1 replication, whereas the CCR5(-)CXCR4(-) macrophages were not permissive to either isolate or other laboratory isolates (ADA and DJV) and primary isolates (MDR 24 and JOEL). In the intestinal mucosa, lymphocytes, not macrophages, are the likely target cell for R5 (and X4) HIV-1 and are the major source of HIV-1 production during early infection.

During a 28-month period, endoscopic mucosal biopsy specimens from all HIV-infected patients were submitted for routine histologic evaluation. Immunoperoxidase staining for cytomegalovirus and herpesvirus antigens (esophagus), mycobacterial and fungal staining, and Gram staining of mucosal biopsy specimens were done. Special fungal and acid-fast stains were selectively performed in patients with absolute CD4 cell counts of less than 200 cells per microliter (200 x 10(6)/L) and/or with diarrhea and or wasting syndrome. Treatment was based on the endoscopic and histologic findings, and long-term follow-up was performed. The 121 symptomatic HIV-infected patients underwent 221 upper and/or lower endoscopies with 285 biopsy sites. The sensitivity and specificity of H&E staining for the diagnosis of gastrointestinal cytomegalovirus were 97% and 100%, respectively. The results of fungal and mycobacterial stains neither altered therapy nor identified previously undiagnosed infections in any patient. Long-term follow-up revealed no patient in whom an infection was missed on routine H&E, which affected outcome. Routine H&E staining is accurate for the diagnosis of gastrointestinal opportunistic infections in HIV-infected patients. Special histologic stains for fungal, mycobacterial, and viral infections did not increase the diagnostic yield or alter medical therapy but doubled the costs.

Mucosal surfaces are the portal of entry for most HIV-1 infections and play an important role in disease pathogenesis. To characterize the biological parameters of HIV-1 infection in mucosal cells, we used purified lamina propria lymphocytes and macrophages from normal human small intestine to determine the distribution of the HIV-1 receptor and coreceptors on intestinal mononuclear cells and the permissiveness of these cells to HIV-1 infection. Lamina propria lymphocytes expressed CD4, CCR5, and CXCR4. In contrast, lamina propria macrophages expressed CD4 but not CCR5 or CXCR4. Intestinal lymphocytes supported replication by R5 and X4 isolates of HIV-1, but lamina propria macrophages were permissive to neither. RANTES, macrophage inflammatory protein-1alpha (MIP-1alpha), and MIP-1beta inhibited infection of intestinal lymphocytes by BaL, indicating that R5 infection of the intestinal lymphocytes was mediated by CCR5. Thus, resident lamina propria lymphocytes, not macrophages, are the target mononuclear cell for HIV-1 infection in the intestinal mucosa during early HIV-1 infection.

In the rectal mucosa, specialized M cells of the lymphoid follicle-associated epithelium conduct vesicular transport of antigens from the mucosal surface into organized mucosal lymphoid tissues, where immune responses are induced. Bacteria and viruses may exploit this mechanism to initiate mucosal or systemic infection. Viral pathogens, including reovirus, poliovirus, and possibly human immunodeficiency virus (HIV), can enter the intestinal or rectal mucosa by adhering to apical membranes of M cells, but the membrane components involved in these interactions are unknown. Glycoprotein coats on the apical surfaces of epithelial cells act as diffusion barriers that limit access of particles and microorganisms to membrane glycolipids and to certain oligosaccharide epitopes of enterocytes but allow selective adherence to M cells. The accessibility of membrane glycolipids of M cells, along with their active endocytic activity, could promote entry of HIV into the rectal mucosa.

The interaction between human immunodeficiency virus type 1 (HIV-1) and primary mucosal cells isolated from normal human small intestine was investigated. Purified primary intestinal epithelial cells could transport cell-free HIV-1 to mononuclear cells, although the epithelial cells did not support viral replication. An unexpected finding was that primary intestinal macrophages were markedly less permissive to HIV-1 than were blood monocytes. The reduced permissiveness appeared to be due to the near absence of surface CCR5 on resident intestinal macrophages. Surface CCR5 could be up-regulated on the monocytes but not the intestinal macrophages by HIV-1 and gp120. Impaired permissiveness of intestinal macrophages to HIV-1 may play an important role in the low prevalence of HIV-1 mRNA-expressing macrophages in the lamina propria during HIV-1 infection in vivo. Characterization of the biologic properties of HIV-1 transport and infection in primary mucosal cells will be key to elucidating the pivotal role of mucosal surfaces in HIV-1 disease.

Inoculation of chimpanzees with human immunodeficiency virus type 1 (HIV-1) has been used as a model system to define mechanisms of pathogenesis and to test protective efficacy of candidate HIV-1 vaccines. In most of these studies, the animals were inoculated intravenously. However, because HIV-1 is transmitted primarily across mucosal surfaces, future evaluations of vaccines should employ mucosal routes for administering infectious virus to immunized animals. To develop a model of rectal transmission of HIV-1, chimpanzees were exposed without trauma to 4 different HIV-1 strains at doses ranging from 200 to 10,000 TCIDs. Infection, characterized by seroconversion and repeated isolation of virus from lymphocytes, was established in 1 of 5 animals. This animal was sequentially inoculated with a subtype B and then an E strain and was infected with both strains. The results show that rectal exposure of adult chimpanzees to cell-free HIV-1 was not an efficient mode of transmission in this cohort.

Human immunodeficiency virus (HIV) can infect human colon epithelial cell lines by both CD4-dependent and -independent mechanisms. The present studies assessed cellular factors that are important for HIV-1 transcription in human colon epithelial cells. The HIV-1 long terminal repeat (LTR) was shown to contain functional DNA cis-regulatory elements downstream of the viral transactivator-responsive element in the transcribed noncoding 5' leader sequence. These downstream regulatory elements, termed DSE, can bind c-Fos and JunD and transmit protein kinase C activation signals to the HIV LTR. Moreover, specific Jun and Fos transcription factors can transactivate HIV-1 provirus in human colon epithelial cells. The DSE also bind related proteins of the CREB/ATF family. In this regard, the DSE behave as 12-0-tetradecanoylphorbol 13-acetate responder element-like cAMP-responsive elements because they bind both AP-1 and CREB/ATF transcription factors, thereby permitting induction of the HIV-1 LTR by both protein kinase C and A activation signals.

Mucosal surfaces are the main natural site of entry into the body for human immunodeficiency virus (HIV). Herein, an alternative mechanism for virus spread is described. The mechanism, which involves transcytosis of endosome-internalized HIV-particles, was generated by contact of HIV-infected cells with the apical surface of an epithelial cell line. Transcytosed viruses rapidly (in 20-30 min) access the serosal side of the epithelial barrier without infecting the epithelium itself. In turn, transcytosed HIV could infect host submucosal mononucleated target cells, and thus the infection could spread. An investigation was done to determine whether mucosal antibodies could block HIV transcytosis. Both secretory IgA (S-IgA) and IgG that were purified from colostrum from HIV-seropositive women impaired HIV transcytosis, irrespective of the level of the recombinant HIV envelope anti-gp160-specific activities in an ELISA. However, specific S-IgAs were more efficient than IgG. Therefore, mucosal-specific S-IgA to HIV-1 could be relevant to reducing infectivity of HIV-1 in corporeal fluids.

In macaques, the route of immunization has a profound effect on the immune response. Augmenting rectal or vaginal immunization with oral or nasal immunization enhanced the secretory IgA, serum IgG, and T cell responses. However, targeted iliac lymph node (TILN) immunization with recombinant simian immunodeficiency virus (SIV) gp120 and p27 elicited the most consistent mucosal antibody responses in the rectum, vagina, urine, seminal fluid, and blood. Both mucosal and TILN immunization induced specific CD4+ T cell proliferative responses in the iliac lymph nodes, which drain these mucosal surfaces, and in the splenic and circulating T cells. Rectal mucosal challenge with cell-free SIV induced total protection in 4 of 7 macaques that were immunized by the TILN route, and, compared with unimmunized macaques or those immunized by the mucosal route (P<.001), it induced a >90% decrease in virus load in 3 of them. Protection from mucosal rectal infection with SIV was significantly associated with an increase in the CD8 suppressor factor (which was generated by the iliac lymph node), RANTES, and MIP-1beta (P<.01).

Combination antiretroviral therapies suppress human immunodeficiency virus (HIV) in peripheral blood, but the effect in gastrointestinal mucosa is uncertain. The occurrence of pathogen-negative diarrhea led to speculation that local HIV infection is etiologic. Mucosal cellular reservoirs for HIV were documented by use of several techniques. Correlations were found among gastrointestinal symptoms, histopathologic findings, cytokine expression, lymphoid apoptosis, and HIV RNA and protein expression in rectal mucosa. Disproportionate depletion of mucosal CD4+ lymphocytes also was found. The short-term effects of antiretroviral therapies were examined to test the hypothesis that these changes are directly related to mucosal HIV infection. Therapy was associated with decreased symptoms, with comparable drops in peripheral blood and mucosal HIV RNA contents, and by increases in blood and mucosal CD4+ lymphocyte contents. In addition, the number of apoptotic cells also declined during therapy. These results suggest that HIV plays a direct role in producing intestinal dysfunction.

Previously, a mucosal model of immunization against human immunodeficiency virus type 1 (HIV-1) was established by using influenza virus as a vector for the neutralizing gp41 epitope ELDKWA. Whether replication of this chimeric influenza virus in the upper respiratory tract of mice is sufficient for inducing mucosal immune responses in the genital tract was investigated. An immunization strategy was established that permits the virus to replicate in the murine upper respiratory tracts but not in the lungs. Intranasal application of the chimeric virus induced HIV-1-specific antibodies in sera and genital tract. In addition, chimeric virus-specific antibody-secreting cells were detected in lymphocyte populations obtained from lungs, spleens, and urogenital tracts. These results indicate that replication of the chimeric influenza/ELDKWA virus in the upper respiratory tract is sufficient to induce systemic immune responses as well as local immune responses in the genital tract.

To define the determinants of anal-rectal shedding of human immunodeficiency virus (HIV) DNA and RNA, 374 HIV-seropositive homosexual men were tested. Factors independently associated with detection of anal-rectal HIV DNA included anal-rectal inflammation and detection of anal human papillomavirus DNA; predictors of HIV RNA included detection of anal-rectal HIV DNA, anal-rectal inflammation, and high plasma HIV RNA levels. The latter (>10,000 copies/mL) was the main determinant of anal-rectal HIV RNA shedding when HIV DNA (e.g., HIV-infected cells) was not detected in the anal-rectal sample. The local presence of HIV-infected cells and local inflammation were the principal determinants of HIV RNA among those with low (<10,000 copies/mL) plasma HIV RNA load. Among those with anal-rectal HIV DNA present, increased HIV RNA plasma load did not increase the risk of shedding of HIV RNA into the anal-rectal canal.

As the largest lymphoid organ and the largest reservoir of macrophages in the body, the gastrointestinal tract mucosa is probably the largest organ reservoir of macrophages infected with HIV-1. To elucidate the biology of HIV-1 infection of intestinal macrophages, we isolated lamina propria macrophages from normal human jejunum by neutral protease digestion, purified the cells by counterflow centrifugal elutriation, and then infected the cells with HIV-1. The lamina propria macrophages were permissive to macrophagetropic isolates of HIV-1 and substantially less permissive to lymphocyte-tropic isolates. Compared with blood monocytes, mucosal macrophages produced 2-3 logs less p24 antigen at peak infection. The reduced level of infection was not due to impaired macrophage viability, reduced CD4 expression, or the isolation procedure. These results confirm that macrophages isolated from the gastrointestinal tract mucosa can support HIV-1 production, albeit at a lower level than blood monocytes. The reduced level of virus production may reflect the unique biology of intestinal lamina propria macrophages.

Mucosal antibody induction by otherwise identical replication-competent and replication-incompetent rhesus monkey rotaviruses was compared. Although psoralen-inactivated virus induced diarrhea, the magnitude of the intestinal antibody response was severely attenuated compared with that of replicating rotavirus, as determined by assay of mucosal antibody-secreting cells. Rotavirus that was neutralized by monoclonal antibodies (anti-VP4 and -VP7) prior to inoculation was similarly ineffective at induction of specific antibodies in intestinal secretions. In contrast to genetically inactivated virus, antibody-neutralized virus did not induce diarrhea. In this murine model, viral replication is an important determinant of antibody induction. The diarrhea response is blocked by neutralizing antibodies, but the mechanism of action is not exclusively the inhibition of viral replication.

Patients with symptomatic gastrointestinal disease due to cytomegalovirus (CMV) were randomized to receive open-label ganciclovir (22) or foscarnet (26). Patients were stratified by disease site and concurrent gut infection. Response was assessed by a visual analogue score of symptoms, endoscopic appearances, histologic inflammation, and numbers of CMV inclusions. In each treatment group, 73% had a complete or good clinical response; 83% of foscarnet-treated and 85% of ganciclovir-treated patients showed response by endoscopy, and inclusion bodies disappeared from follow-up biopsies in 73% of these. Most patients (35) developed further evidence of CMV disease during follow-up. The time to progression was not significantly different between recipients (16 weeks) and nonrecipients (13 weeks) of maintenance therapy, although patients were not randomized to receive maintenance or not. Survival in both treatment groups was < 40 weeks and was unaffected by maintenance treatment. Both ganciclovir and foscarnet are effective first-line treatments for gastrointestinal (GI) CMV infection. Maintenance therapy does not prevent progression of disease.