Delayed vaginal SHIV infection in VRC01 and anti-α4ß7 treated rhesus macaques.

VRC01 protects macaques from vaginal SHIV infection after a single high-dose challenge. Infusion of a simianized anti-α4ß7 mAb (Rh-α4ß7) just prior to, and during repeated vaginal exposures to SIVmac251 partially protected macaques from vaginal SIV infection and rescued CD4+ T cells. To investigate the impact of combining VRC01 and Rh-α4ß7 on SHIV infection, 3 groups of macaques were treated with a suboptimal dosing of VRC01 alone or in combination with Rh-α4ß7 or with control antibodies prior to the initiation of weekly vaginal exposures to a high dose (1000 TCID50) of SHIVAD8-EO. The combination Rh-α4ß7-VRC01 significantly delayed SHIVAD8-EO vaginal infection. Following infection, VRC01-Rh-α4ß7-treated macaques maintained higher CD4+ T cell counts and exhibited lower rectal SIV-DNA loads compared to controls. Interestingly, VRC01-Rh-α4ß7-treated macaques had fewer IL-17-producing cells in the blood and the gut during the acute phase of infection. Moreover, higher T cell responses to the V2-loop of the SHIVAD8-EO envelope in the VRC01-Rh-α4ß7 group inversely correlated with set point viremia. The combination of suboptimal amounts of VRC01 and Rh-α4ß7 delayed infection, altered antiviral immune responses and minimized CD4+ T cell loss. Further exploration of the effect of combining bNAbs with Rh-α4ß7 on SIV/HIV infection and antiviral immune responses is warranted and may lead to novel preventive and therapeutic strategies.

Integrin α4ß7 Blockade Preferentially Impacts CCR6+ Lymphocyte Subsets in Blood and Mucosal Tissues of Naive Rhesus Macaques.

Infusion of a simianized anti-α4ß7 mAb (Rh-α4ß7) just before and following SIV infection protected rhesus macaques from developing AIDS and partially from vaginal SIV acquisition. Recently, short-term treatment with Rh-α4ß7 in combination with cART was found to lead to prolonged viral suppression after withdrawal of all therapeutic interventions. The humanized form of Rh-α4ß7, vedolizumab, is a highly effective treatment for inflammatory bowel disease. To clarify the mechanism of action of Rh-α4ß7, naive macaques were infused with Rh-α4ß7 and sampled in blood and tissues before and after treatment to monitor several immune cell subsets. In blood, Rh-α4ß7 increased the CD4+ and CD8+ T cell counts, but not B cell counts, and preferentially increased CCR6+ subsets while decreasing CD103+ and CD69+ lymphocytes. In mucosal tissues, surprisingly, Rh-α4ß7 did not impact integrin α4+ cells, but decreased the frequencies of CCR6+ and CD69+ CD4+ T cells and, in the gut, Rh-α4ß7 transiently decreased the frequency of memory and IgA+ B cells. In summary, even in the absence of inflammation, Rh-α4ß7 impacted selected immune cell subsets in different tissues. These data provide new insights into the mechanisms by which Rh-α4ß7 may mediate its effect in SIV-infected macaques with implications for understanding the effect of treatment with vedolizumab in patients with inflammatory bowel disease.

A Small Molecule, Which Competes with MAdCAM-1, Activates Integrin α4ß7 and Fails to Prevent Mucosal Transmission of SHIV-SF162P3.

Mucosal HIV-1 transmission is inefficient. However, certain viral and host characteristics may play a role in facilitating HIV acquisition and systemic expansion. Cells expressing high levels of integrin α4ß7 have been implicated in favoring the transmission process and the infusion of an anti-α4ß7 mAb (RM-Act-1) prior to, and during a repeated low-dose vaginal challenge (RLDC) regimen with SIVmac251 reduced SIV acquisition and protected the gut-associated lymphoid tissues (GALT) in the macaques that acquired SIV. α4ß7 expression is required for lymphocyte trafficking to the gut lamina propria and gut inductive sites. Several therapeutic strategies that target α4ß7 have been shown to be effective in treating inflammatory conditions of the intestine, such as inflammatory bowel disease (IBD). To determine if blocking α4ß7 with ELN, an orally available anti-α4 small molecule, would inhibit SHIV-SF162P3 acquisition, we tested its ability to block MAdCAM-1 (α4ß7 natural ligand) and HIV-gp120 binding in vitro. We studied the pharmacokinetic profile of ELN after oral and vaginal delivery in macaques. Twenty-six macaques were divided into 3 groups: 9 animals were treated with ELN orally, 9 orally and vaginally and 8 were used as controls. All animals were challenged intra-vaginally with SHIV-SF162P3 using the RLDC regimen. We found that ELN did not protect macaques from SHIV acquisition although it reduced the SHIV-induced inflammatory status during the acute phase of infection. Notably, integrins can exist in different activation states and, comparing the effect of ELN and the anti-α4ß7 mAb RM-Act-1 that reduced susceptibility to SIV infection, we determined that ELN induces the active conformation of α4ß7, while RM-Act-1 inhibits its activation through an allosteric mechanism. These results suggest that inhibition of α4ß7 activation may be necessary to reduce susceptibility to SIV/SHIV infection and highlight the complexity of anti-integrins therapeutic approach in HIV as well as in IBD and other autoimmune diseases.

Correction: A Small Molecule, Which Competes with MAdCAM-1, Activates Integrin α4ß7 and Fails to Prevent Mucosal Transmission of SHIV-SF162P3.

[This corrects the article DOI: 10.1371/journal.ppat.1005720.].

A model of genital herpes simplex virus Type 1 infection in Rhesus Macaques.

BACKGROUND: Although HSV-2 is the major cause of genital lesions, HSV-1 accounts for half of new cases in developed countries. METHODS: Three healthy SHIV-SF162P3-infected Indian rhesus macaques were inoculated with 4×108 pfu of HSV-1 twice, with the second inoculation performed after the vaginal mucosa was gently abraded with a cytobrush. RESULTS: HSV-1 DNA was detected in vaginal swabs 5 days after the second but not the first inoculation in all three macaques. An increase in inflammatory cytokines was detected in the vaginal fluids of the animals with no or intermittent shedding. Higher frequency of blood α4 ß7high CD4+ T cells was measured in the animals with consistent and intermitted shedding, while a decrease in the frequency of CD69+ CD4+ T cells was present in all animals. CONCLUSIONS: This macaque model of genital HSV-1 could be useful to study the impact of the growing epidemic of genital HSV-1 on HIV infection.

Retinoic acid imprints a mucosal-like phenotype on dendritic cells with an increased ability to fuel HIV-1 infection.

The tissue microenvironment shapes the characteristics and functions of dendritic cells (DCs), which are important players in HIV infection and dissemination. Notably, DCs in the gut have the daunting task of orchestrating the balance between immune response and tolerance. They produce retinoic acid (RA), which imprints a gut-homing phenotype and influences surrounding DCs. To investigate how the gut microenvironment impacts the ability of DCs to drive HIV infection, we conditioned human immature monocyte-derived DCs (moDCs) with RA (RA-DCs), before pulsing them with HIV and mixing them with autologous T cells. RA-DCs showed a semimature, mucosal-like phenotype and released higher amounts of TGF-ß1 and CCL2. Using flow cytometry, Western blot, and microscopy, we determined that moDCs express the cell adhesion molecule mucosal vascular addressin cell adhesion molecule-1 (MAdCAM-1) and that RA increases its expression. MAdCAM-1 was also detected on a small population of DCs in rhesus macaque (Macaca mulata) mesenteric lymph node. RA-DCs formed more DC-T cell conjugates and promoted significantly higher HIV replication in DC-T cell mixtures compared with moDCs. This correlated with the increase in MAdCAM-1 expression. Blocking MAdCAM-1 partially inhibited the enhanced HIV replication. In summary, RA influences DC phenotype, increasing their ability to exacerbate HIV infection. We describe a previously unknown mechanism that may contribute to rapid HIV spread in the gut, a major site of HIV replication after mucosal exposure.

HSV-2 infection of dendritic cells amplifies a highly susceptible HIV-1 cell target.

Herpes simplex virus type 2 (HSV-2) increases the risk of HIV-1 infection and, although several reports describe the interaction between these two viruses, the exact mechanism for this increased susceptibility remains unclear. Dendritic cells (DCs) at the site of entry of HSV-2 and HIV-1 contribute to viral spread in the mucosa. Specialized DCs present in the gut-associated lymphoid tissues produce retinoic acid (RA), an important immunomodulator, able to influence HIV-1 replication and a key mediator of integrin α4ß7 on lymphocytes. α4ß7 can be engaged by HIV-1 on the cell-surface and CD4⁺ T cells expressing high levels of this integrin (α4ß7 (high)) are particularly susceptible to HIV-1 infection. Herein we provide in-vivo data in macaques showing an increased percentage of α4ß7 (high) CD4⁺ T cells in rectal mucosa, iliac lymph nodes and blood within 6 days of rectal exposure to live (n = 11), but not UV-treated (n = 8), HSV-2. We found that CD11c⁺ DCs are a major target of HSV-2 infection in in-vitro exposed PBMCs. We determined that immature monocyte-derived DCs (moDCs) express aldehyde dehydrogenase ALDH1A1, an enzyme essential for RA production, which increases upon HSV-2 infection. Moreover, HSV-2-infected moDCs significantly increase α4ß7 expression on CD4⁺ T lymphocytes and HIV-1 infection in DC-T cell mixtures in a RA-dependent manner. Thus, we propose that HSV-2 modulates its microenviroment, influencing DC function, increasing RA production capability and amplifying a α4ß7 (high)CD4⁺ T cells. These factors may play a role in increasing the susceptibility to HIV-1.

Blockade of TGF-ß signaling reactivates HIV-1/SIV reservoirs and immune responses in vivo.

TGF-ß plays a critical role in maintaining immune cells in a resting state by inhibiting cell activation and proliferation. Resting HIV-1 target cells represent the main cellular reservoir after long-term antiretroviral therapy (ART). We hypothesized that releasing cells from TGF-ß-driven signaling would promote latency reversal. To test our hypothesis, we compared HIV-1 latency models with and without TGF-ß and a TGF-ß type 1 receptor inhibitor, galunisertib. We tested the effect of galunisertib in SIV-infected, ART-treated macaques by monitoring SIV-env expression via PET/CT using the 64Cu-DOTA-F(ab')2 p7D3 probe, along with plasma and tissue viral loads (VLs). Exogenous TGF-ß reduced HIV-1 reactivation in U1 and ACH-2 models. Galunisertib increased HIV-1 latency reversal ex vivo and in PBMCs from HIV-1-infected, ART-treated, aviremic donors. In vivo, oral galunisertib promoted increased total standardized uptake values in PET/CT images in gut and lymph nodes of 5 out of 7 aviremic, long-term ART-treated, SIV-infected macaques. This increase correlated with an increase in SIV RNA in the gut. Two of the 7 animals also exhibited increases in plasma VLs. Higher anti-SIV T cell responses and antibody titers were detected after galunisertib treatment. In summary, our data suggest that blocking TGF-ß signaling simultaneously increases retroviral reactivation events and enhances anti-SIV immune responses.

HIV-1 Establishes a Sanctuary Site in the Testis by Permeating the BTB Through Changes in Cytoskeletal Organization.

Studies suggest that HIV-1 invades the testis through initial permeation of the blood-testis barrier (BTB). The selectivity of the BTB to antiretroviral drugs makes this site a sanctuary for the virus. Little is known about how HIV-1 crosses the BTB and invades the testis. Herein, we used 2 approaches to examine the underlying mechanism(s) by which HIV-1 permeates the BTB and gains entry into the seminiferous epithelium. First, we examined if recombinant Tat protein was capable of perturbing the BTB and making the barrier leaky, using the primary rat Sertoli cell in vitro model that mimics the BTB in vivo. Second, we used HIV-1-infected Sup-T1 cells to investigate the activity of HIV-1 infection on cocultured Sertoli cells. Using both approaches, we found that the Sertoli cell tight junction permeability barrier was considerably perturbed and that HIV-1 effectively permeates the BTB by inducing actin-, microtubule-, vimentin-, and septin-based cytoskeletal changes in Sertoli cells. These studies suggest that HIV-1 directly perturbs BTB function, potentially through the activity of the Tat protein.

Blocking α4ß7 integrin delays viral rebound in SHIVSF162P3-infected macaques treated with anti-HIV broadly neutralizing antibodies.

Anti-HIV broadly neutralizing antibodies (bNAbs) may favor development of antiviral immunity by engaging the immune system during immunotherapy. Targeting integrin α4ß7 with an anti-α4ß7 monoclonal antibody (Rh-α4ß7) affects immune responses in SIV/SHIV-infected macaques. To explore the therapeutic potential of combining bNAbs with α4ß7 integrin blockade, SHIVSF162P3-infected, viremic rhesus macaques were treated with bNAbs only (VRC07-523LS and PGT128 anti-HIV antibodies) or a combination of bNAbs and Rh-α4ß7 or were left untreated as a control. Treatment with bNAbs alone decreased viremia below 200 copies/ml in all macaques, but seven of eight macaques (87.5%) in the bNAbs-only group rebounded within a median of 3 weeks (95% CI: 2 to 9). In contrast, three of six macaques treated with a combination of Rh-α4ß7 and bNAbs (50%) maintained a viremia below 200 copies/ml until the end of the follow-up period; viremia in the other three macaques rebounded within a median of 6 weeks (95% CI: 5 to 11). Thus, there was a modest delay in viral rebound in the macaques treated with the combination antibody therapy compared to bNAbs alone. Our study suggests that α4ß7 integrin blockade may prolong virologic control by bNAbs in SHIVSF162P3-infected macaques.

Blockade of attachment and fusion receptors inhibits HIV-1 infection of human cervical tissue.

Identification of cellular factors involved in HIV-1 entry and transmission at mucosal surfaces is critical for understanding viral pathogenesis and development of effective prevention strategies. Here we describe the evaluation of HIV-1 entry inhibitors for their ability to prevent infection of, and dissemination from, human cervical tissue ex vivo. Blockade of CD4 alone or CCR5 and CXCR4 together inhibited localized mucosal infection. However, simultaneous blockade of CD4 and mannose-binding C-type lectin receptors including dendritic cell-specific intercellular adhesion molecule-grabbing integrin was required to inhibit HIV-1 uptake and dissemination by migratory cells. In contrast, direct targeting of HIV-1 by neutralizing mAb b12 and CD4-IgG2 (PRO-542) blocked both localized infection and viral dissemination pathways. Flow cytometric analysis and immunostaining of migratory cells revealed two major populations, CD3(+)HLA-DR(-) and CD3(-)HLA-DR(+) cells, with a significant proportion of the latter also expressing dendritic cell-specific intercellular adhesion molecule-grabbing integrin. Bead depletion studies demonstrated that such HLA-DR(+) cells accounted for as much as 90% of HIV-1 dissemination. Additional studies using immature monocyte-derived dendritic cells demonstrated that although mannose-binding C-type lectin receptors and CD4 are the principal receptors for gp120, other mechanisms may account for virus capture. Our identification of the predominant receptors involved in HIV-1 infection and dissemination within human cervical tissue highlight important targets for microbicide development.

Anti-α4ß7 monoclonal antibody-conjugated nanoparticles block integrin α4ß7 on intravaginal T cells in rhesus macaques.

Intravenous administration of anti-α4ß7 monoclonal antibody in macaques decreases simian immunodeficiency virus (SIV) vaginal infection and reduces gut SIV loads. Because of potential side effects of systemic administration, a prophylactic strategy based on mucosal administration of anti-α4ß7 antibody may be safer and more effective. With this in mind, we developed a novel intravaginal formulation consisting of anti-α4ß7 monoclonal antibody-conjugated nanoparticles (NPs) loaded in a 1% hydroxyethylcellulose (HEC) gel (NP-α4ß7 gel). When intravaginally administered as a single dose in a rhesus macaque model, the formulation preferentially bound to CD4+ or CD3+ T cells expressing high levels of α4ß7, and occupied ~40% of α4ß7 expressed by these subsets and ~25% of all cells expressing α4ß7 Blocking of the α4ß7 was restricted to the vaginal tract without any changes detected systemically.

A Tat/Rev Induced Limiting Dilution Assay to Measure Viral Reservoirs in Non-Human Primate Models of HIV Infection.

The establishment of latent infection and poorly characterized viral reservoirs in tissues represent major obstacles to a definitive cure for HIV. Non-human primate (NHP) models of HIV infection are critical to elucidate pathogenic processes and an essential tool to test novel therapeutic strategies. Thus, the availability of novel assays to measure residual viral replication and reservoirs in NHP models may increase their utility in the search for an HIV cure. We developed a tat/rev induced limiting dilution assay to measure the frequency of CD4+ T cells that express multiply-spliced(ms)_SIV RNA in presence and absence of stimulation. We validated the assay using cell lines and cells from blood and lymph nodes of SIV infected macaques. In vitro, SIV/SHIV TILDA detects only cells expressing viral proteins. In SIV/SHIV-infected macaques, CD4+ T cells that express msSIV/SHIV RNA (TILDA data) were detected also in the setting of very low/undetectable viremia. TILDA data were significantly higher after stimulation and correlated with plasma viral load (pVL). Interestingly, TILDA data from early cART initiation correlated with peak and AUC pVL post-cART interruption. In summary, we developed an assay that may be useful in characterizing viral reservoirs and determining the effect of HIV interventions in NHP models.

Experimental Oral Herpes Simplex Virus-1 (HSV-1) Co-infection in Simian Immunodeficiency Virus (SIV)-Infected Rhesus Macaques.

Herpes simplex virus 1 and 2 (HSV-1/2) similarly initiate infection in mucosal epithelia and establish lifelong neuronal latency. Anogenital HSV-2 infection augments the risk for sexual human immunodeficiency virus (HIV) transmission and is associated with higher HIV viral loads. However, whether oral HSV-1 infection contributes to oral HIV susceptibility, viremia, or oral complications of HIV infection is unknown. Appropriate non-human primate (NHP) models would facilitate this investigation, yet there are no published studies of HSV-1/SIV co-infection in NHPs. Thus, we performed a pilot study for an oral HSV-1 infection model in SIV-infected rhesus macaques to describe the feasibility of the modeling and resultant immunological changes. Three SIV-infected, clinically healthy macaques became HSV-1-infected by inoculation with 4 × 108 pfu HSV-1 McKrae on buccal, tongue, gingiva, and tonsils after gentle abrasion. HSV-1 DNA was shed in oral swabs for up to 21 days, and shedding recurred in association with intra-oral lesions after periods of no shedding during 56 days of follow up. HSV-1 DNA was detected in explant cultures of trigeminal ganglia collected at euthanasia on day 56. In the macaque with lowest baseline SIV viremia, SIV plasma RNA increased following HSV-1 infection. One macaque exhibited an acute pro-inflammatory response, and all three animals experienced T cell activation and mobilization in blood. However, T cell and antibody responses to HSV-1 were low and atypical. Through rigorous assessesments, this study finds that the virulent HSV-1 strain McKrae resulted in a low level HSV-1 infection that elicited modest immune responses and transiently modulated SIV infection.

PolyICLC Exerts Pro- and Anti-HIV Effects on the DC-T Cell Milieu In Vitro and In Vivo.

Myeloid dendritic cells (mDCs) contribute to both HIV pathogenesis and elicitation of antiviral immunity. Understanding how mDC responses to stimuli shape HIV infection outcomes will inform HIV prevention and treatment strategies. The long double-stranded RNA (dsRNA) viral mimic, polyinosinic polycytidylic acid (polyIC, PIC) potently stimulates DCs to focus Th1 responses, triggers direct antiviral activity in vitro, and boosts anti-HIV responses in vivo. Stabilized polyICLC (PICLC) is being developed for vaccine adjuvant applications in humans, making it critical to understand how mDC sensing of PICLC influences HIV infection. Using the monocyte-derived DC (moDC) model, we sought to describe how PICLC (vs. other dsRNAs) impacts HIV infection within DCs and DC-T cell mixtures. We extended this work to in vivo macaque rectal transmission studies by administering PICLC with or before rectal SIVmac239 (SIVwt) or SIVmac239ΔNef (SIVΔNef) challenge. Like PIC, PICLC activated DCs and T cells, increased expression of α4ß7 and CD169, and induced type I IFN responses in vitro. The type of dsRNA and timing of dsRNA exposure differentially impacted in vitro DC-driven HIV infection. Rectal PICLC treatment similarly induced DC and T cell activation and pro- and anti-HIV factors locally and systemically. Importantly, this did not enhance SIV transmission in vivo. Instead, SIV acquisition was marginally reduced after a single high dose challenge. Interestingly, in the PICLC-treated, SIVΔNef-infected animals, SIVΔNef viremia was higher, in line with the importance of DC and T cell activation in SIVΔNef replication. In the right combination anti-HIV strategy, PICLC has the potential to limit HIV infection and boost HIV immunity.

Immunodeficiency virus exploitation of dendritic cells in the early steps of infection.

The unique capacity of dendritic cells (DCs) to capture and process pathogens for presentation to the immune system, combined with their capacity to express costimulatory and adhesion molecules as well as cytokines and chemokines, renders them powerful antigen-presenting cells. However, immunodeficiency viruses hijack DCs to facilitate virus dissemination while subverting effective immune activation. Depending on the activation level of the DC subset, human immunodeficiency virus can use different receptors (CD4, chemokine, and C-type lectin receptors) to bind to DCs. These aspects likely impact whether a DC is productively infected by or simply carries virus for transmission to more permissive targets. DCs efficiently transmit virus to CD4+ T cells, driving virus growth as well as providing signals to trigger virus expansion in virus-bearing CD4+ T cells. There is accumulating evidence that viral determinants (nef, tat) selectively modulate immature DC biology, fostering DC-T cell interactions and virus replication without up-regulating costimulatory molecules for effective immune function. In addition, virus-loaded, immature DCs activate CD4+ virus-specific T cells, and mature DCs stimulate CD4+ and CD8+ T cells. Thus, even if immature DCs entrap virus as it crosses the mucosae and initiate a CD4+ T cell response, this is likely insufficient to control infection. Appreciating how virus modulates DC function and what determines whether virus is processed for immune stimulation or transmitted between cells will unveil the exact role of these cells in the onset of infection and advance preventative microbicide and vaccine/therapeutic approaches.

Enhanced in vitro stimulation of rhesus macaque dendritic cells for activation of SIV-specific T cell responses.

The macaque-simian immunodeficiency virus (SIV) system is one of the best animal models available to study the role of dendritic cells (DCs) in transmission and pathogenesis of HIV, as well as to test DC-based vaccine and therapeutic strategies. To better define and optimize this system, the responsiveness of macaque monocyte-derived DCs to a variety of maturation stimuli was examined. Characteristic immunophenotypic and functional DC maturation induced by standard monocyte conditioned medium (MCM) was compared to the activation induced by a panel of stimuli including soluble CD40L, LPS, Poly I:C, PGE(2)/TNFalpha, and a cocktail mixture of PGE(2)/TNFalpha/IL-1beta/IL-6. Immunophenotypic analysis confirmed that all stimuli induced stable up-regulation of CD25, CD40, CD80, CD83, CD86, HLA-DR, DC-LAMP (CD208), and DEC-205 (CD205). In general, macaque DCs exhibited weaker responses to LPS and Poly I:C than human DCs, and soluble CD40L stimulation induced variable expression of CD25. Interestingly, while the endocytic capacity of CD40L-matured cells was down-modulated comparably to DCs matured with MCM or the cocktail, the T cell stimulatory activity was not enhanced to the same extent. The particularly reproducible and potent T cell stimulatory capacity of cocktail-treated DCs correlated with a more homogenous mature DC phenotype, consistently high levels of IL-12 production, and better viability upon reculture compared to DCs activated by other stimuli. Furthermore, cocktail-matured DCs efficiently captured and presented inactivated SIV to SIV-primed T cells in vitro. Thus, the cocktail represents a particularly potent and useful stimulus for the generation of efficacious immunostimulatory macaque DCs.

Entry inhibitors SCH-C, RANTES, and T-20 block HIV type 1 replication in multiple cell types.

The small-molecule CCR5 antagonist SCH-C (SCH 351125) was tested for its ability to inhibit HIV-1 replication in peripheral blood mononuclear cells (PBMCs), cord blood mononuclear cells, immature dendritic cells (DCs), and macrophages. Inhibition of infection of PBMCs by virus associated with mature DC in trans was also studied. For comparison, the peptide-based fusion inhibitor T-20 and the CC-chemokine RANTES were also evaluated. Although some cell type-dependent differences in potency were observed, each of the three entry inhibitors was active against the replication of three different CCR5-using primary isolates in each cell type. CCR5-dependent HIV-1 infectivity, whether DC associated or not, is thus vulnerable to inhibitors that block the virus-cell fusion process by different mechanisms. Together, these results suggest that SCH-C and other entry inhibitors should be evaluated for their clinical potential as inhibitors of HIV-1 replication in several settings, including the prevention of maternal-infant transmission and the prevention of sexual transmission by topical application as a microbicide.

Hypergravity and dehydration-induced shifts of interstitial fluid in the skin monitored by ultrasound.

INTRODUCTION: Spaceflight is associated with a shift of interstitial fluid toward the skin of the head and a fluid loss that can decrease superficial tissue thickness (TT). To collect the physiological data needed to develop techniques for monitoring the hydration status of astronauts during spaceflight, we investigated the changes in TT induced by a 12-h nil-by-mouth period and a 30-min period of +2 Gz hypergravity measured at forehead and tibia. METHODS: There were 16 male volunteers who were twice subjected to 30 min of +2 Gz acceleration in a human centrifuge, once following an oral fluid load of 250 ml x h(-1) (procedure A), and once following a nil-by-mouth period of 12 h (procedure B). The TT at the forehead (TT-f) and tibia (TT-t) was measured before (t0), after 10 min (t10), and after 30 min (t30) of hypergravity using a miniature 10 MHz A-mode ultrasound device. The bodyweight, hematocrit (HCT), and plasma viscosity (PVS) were measured at tO and t0. RESULTS: The nil-by-mouth period induced a significant weight loss and increase in HCT and PVS. A significant increase was noted in TT-t, but not in TT-f (p < 0.05). Following both procedures, exposure to hypergravity produced declines in TT-f at t10 and t30 compared with baseline (p < 0.05), whereas TT-t was not influenced. CONCLUSIONS: Changes in interstitial fluid load of superficial tissues can be tracked by A-mode ultrasonography. Fluid loss-induced changes are best detected at the tibia, hypergravity-induced changes at the forehead.