Run-Hong Zhou

Exosomes contribute to the transmission of anti-HIV activity from TLR3-activated brain microvascular endothelial cells to macrophages.

Human brain microvascular endothelial cells (HBMECs), the major cell type in the blood-brain barrier (BBB), play a key role in maintaining brain homeostasis. However, their role in the BBB innate immunity against HIV invasion of the central nervous system (CNS) remains to be determined. Our early work showed that TLR3 signaling of HBMECs could produce the antiviral factors that inhibit HIV replication in macrophages. The present study examined whether exosomes from TLR3-activated HBMECs mediate the intercellular transfer of antiviral factors to macrophages. Primary human macrophages could take up exosomes from TLR3-activated HBMECs. HBMECs-derived exosomes contained multiple antiviral factors, including several key IFN-stimulated genes (ISGs; ISG15, ISG56, and Mx2) at mRNA and protein levels. The depletion of exosomes from TLR3-activated HBMECs culture supernatant diminished HBMECs-mediated anti-HIV activity in macrophages. In conclusion, we demonstrate that exosomes shed by HBMECs are able to transport the antiviral molecules to macrophages. This finding suggests the possibility that HIV nonpermissive BBB cells (HBMECs) can help to restore the antiviral state in HIV-infected macrophages, which may be a defense mechanism against HIV neuroinvasion.

Comparative Analysis of Immune Activation Markers of CD8+ T Cells in Lymph Nodes of Different Origins in SIV-Infected Chinese Rhesus Macaques

Altered T-cell homeostasis, such as expansion of CD8+ T cells to the secondary lymphatic compartments, has been suggested as a mechanism of HIV/simian immunodeficiency virus (SIV)-pathogenesis. However, the role of immune activation of CD8+ T cells in the CD4/CD8 turnover and viral replication in these tissues is not completely understood. In this study, we compared the expression of immune activation markers (CD69 and HLA-DR) on CD8+ T cells in the peripheral blood and lymph nodes (LNs) of SIV-infected/uninfected Chinese rhesus macaques. SIV-infected macaques had significantly higher percentages of CD8+CD69+ and CD8+HLA-DR+ T cells in all these anatomical compartments than uninfected macaques. LNs that located close to the gastrointestinal (GI) tract (colon, mesenteric, and iliac LNs) of SIV-infected macaques had profoundly lower numbers of CD4+ T cells, but no significant difference in expression of activation marker (CD8+CD69+ and CD8+HLA-DR+) as compared with the peripheral lymphatic tissues (axillary and inguinal LNs). The CD4/CD8 ratios were negatively correlated with the activation of CD8+ T cells in the overall LNs, with further associations with CD8+HLA-DR+ in GI LNs while CD8+CD69+ in peripheral LNs. These observations demonstrate that the increase of CD8+ T cell activation is a contributing factor for the decline of CD4/CD8 ratios in GI system.

Soybean-derived Bowman-Birk Inhibitor (BBI) Inhibits HIV Replication in Macrophages

The Bowman-Birk inhibitor (BBI), a soybean-derived protease inhibitor, is known to have anti-inflammatory effect in both in vitro and in vivo systems. Macrophages play a key role in inflammation and immune activation, which is implicated in HIV disease progression. Here, we investigated the effect of BBI on HIV infection of peripheral blood monocyte-derived macrophages. We demonstrated that BBI could potently inhibit HIV replication in macrophages without cytotoxicity. Investigation of the mechanism(s) of BBI action on HIV showed that BBI induced the expression of IFN-β and multiple IFN stimulated genes (ISGs), including Myxovirus resistance protein 2 (Mx2), 2′,5′-oligoadenylate synthetase (OAS-1), Virus inhibitory protein (viperin), ISG15 and ISG56. BBI treatment of macrophages also increased the expression of several known HIV restriction factors, including APOBEC3F, APOBEC3G and tetherin. Furthermore, BBI enhanced the phosphorylation of IRF3, a key regulator of IFN-β. The inhibition of IFN-β pathway by the neutralization antibody to type I IFN receptor (Anti-IFNAR) abolished BBI-mediated induction of the anti-HIV factors and inhibition of HIV in macrophages. These findings that BBI could activate IFN-β-mediated signaling pathway, initialize the intracellular innate immunity in macrophages and potently inhibit HIV at multiple steps of viral replication cycle indicate the necessity to further investigate BBI as an alternative and cost-effective anti-HIV natural product.

Human Intestinal Epithelial Cells Release Antiviral Factors That Inhibit HIV Infection of Macrophages

As a rich source of CD4+ T cells and macrophages, the gastrointestinal (GI) tract is a major target site for HIV infection. The interplay between GI-resident macrophages and intestinal epithelial cells (IECs) constitutes an important element of GI innate immunity against pathogens. In this study, we investigated whether human IECs have the ability to produce antiviral factors that can inhibit HIV infection of macrophages. We demonstrated that IECs possess functional toll-like receptor 3 (TLR3), the activation of which resulted in induction of key interferon (IFN) regulatory factors (IRF3 and IRF7), IFN-β, IFN-λ, and CC chemokines (MIP-1α, MIP-1β, RANTES), the ligands of HIV entry co-receptor CCR5. In addition, TLR3-activated IECs release exosomes that contained the anti-HIV factors, including IFN-stimulated genes (ISGs: ISG15, ISG56, MxB, OAS-1, GBP5, and Viperin) and HIV restriction miRNAs (miRNA-17, miRNA-20, miRNA-28, miRNA-29 family members, and miRNA-125b). Importantly, treatment of macrophages with supernatant (SN) from the activated IEC cultures inhibited HIV replication. Further studies showed that IEC SN could also induce the expression of antiviral ISGs and cellular HIV restriction factors (Tetherin and APOBEC3G/3F) in HIV-infected macrophages. These findings indicated that IECs might act as an important element in GI innate immunity against HIV infection/replication.

Epigallocatechin gallate inhibits macaque SEVI-mediated enhancement of SIV or SHIV infection.

Background: Human semen contains a factor that can enhance HIV infection up to 105-fold in cultures. This factor is termed semen-derived enhancer of virus infection (SEVI) and is composed of proteolytic fragments (PAP248-286) from prostatic acid phosphatase in semen. In this study, we examined whether macaque SEVI can facilitate simian immunodeficiency virus (SIV) or chimeric simian/human immunodeficiency virus (SHIV) infection. We also studied the effect of epigallocatechin gallate (EGCG) on macaque SEVI-mediated SIV or SHIV enhancement. Methods: SIV or SHIV was mixed with different concentrations of macaque SEVI in the presence or absence of EGCG. The mixture was added to cultures of TZM-bl cells or macaque PBMCs. The effect of EGCG on macaque SEVI was measured by Congo-red staining assay and thioflavin T (ThT) fluorescence assay and was visualized by a transmission electron microscope. Results: We identified that there is one amino acid difference at the site of 277 between human PAP248-286 and macaque PAP248-286. Macaque SEVI significantly enhanced SIV or SHIV infection of TZM-bl cells and macaque PBMCs. EGCG could block macaque SEVI-mediated enhancement of SIV or SHIV infection. Mechanistically, EGCG could degrade the formation of macaque SEVI amyloid fibrils that facilitates HIV attachment to the target cells. Conclusions: The finding that macaque SEVI could enhance SIV or SHIV infection indicates the possibility to use the macaque SEVI in vivo studies with the macaque models. In addition, future studies are necessary to examine whether EGCG can be used as an effective microbicide for preventing SIV or SHIV mucosal transmission.

Soybean-derived Bowman-Birk inhibitor (BBI) blocks HIV entry into macrophages

Bowman-Birk inhibitor (BBI) is a soybean-derived protease inhibitor that has anti-inflammation and anti-HIV effect. Here, we further investigated the anti-HIV action of BBI in macrophages, focusing on its effect on viral entry. We found that BBI could significantly block HIV entry into macrophages. Investigation of the mechanism(s) of the BBI action on HIV inhibition showed that BBI down-regulated the expression of CD4 receptor (as much as 80%) and induced the production of the CC chemokines (up to 60 folds at protein level) in macrophages. This inhibitory effect of BBI on HIV entry could be blocked by the neutralization antibodies to CC chemokines. These findings indicate that BBI may have therapeutic potential as a viral entry inhibitor for the prevention and treatment of HIV infection.

Human Cervical Epithelial Cells Release Antiviral Factors and Inhibit HIV Replication in Macrophages

The female reproductive tract is a major site of HIV sexual transmission. We here examined whether human cervical epithelial cells (HCEs) can be immunologically activated and produce antiviral factors against HIV. We demonstrated that HCEs (End1/E6E7 cells) possess the functional toll-like receptor (TLR)3 signaling system, which could be activated by Poly I:C and induce multiple cellular HIV restriction factors. The treatment of primary human macrophages with supernatant (SN) from TLR3-activated End1/E6E7 cell cultures resulted in HIV inhibition. This SN-mediated HIV inhibition was mainly through the induction of interferons (IFN)-β and IFN-λs, as the antibodies to IFN-β or IFN-λs receptor could effectively block the SN-mediated anti-HIV effect. Further studies showed that the incubation of macrophages with SN from the activated cervical epithelial cell cultures induced the expression of a number of IFN-stimulated genes (ISGs), including IFN-stimulated gene (ISG15), ISG56, 2′, 5′-oligoadenylate synthetase 1 (OAS 1), OAS 2, Myxovirus Resistance A (MxA), MxB, and Guanylate-binding protein 5 (GBP5). In addition, TLR3-activated cells produced the CC chemokines [regulated on activation, normal T cell expressed and secreted (RANTES), Human macrophage inflammatory protein 1 alpha (MIP-1α), MIP-1β] the ligands of HIV entry co-receptor CCR5. These observations support further studies on HCEs as potentially crucial and alternative targets for immunological intervention to control and prevent HIV sexual transmission.

IFN-λ4 inhibits HIV infection of macrophages through signalling of IFN-λR1/IL-10R2 receptor complex

The recently discovered IFN‐λ4 has been found to have antiviral activity against several viruses. However, its unknown whether IFN‐λ4 can inhibit HIV infection. Here, we show that IFN‐λ4 could suppress HIV infection of macrophages. This IFN‐λ4‐mediated HIV inhibition was compromised by the antibodies against IFN‐λ receptor complex, IFN‐λR1/IL‐10R2. IFN‐λ4 enhanced the phosphorylation of STAT1, and induced antiviral interferon‐stimulated genes. These findings indicated that IFN‐λ4 can inhibit HIV via JAK/STAT signalling pathway.

Epigallocatechin-3-gallate local pre-exposure application prevents SHIV rectal infection of macaques

Epigallocatechin-3-gallate (EGCG), a natural and major ingredient of green tea, has been shown to have anti-inflammation and anti-HIV-1 properties. We demonstrated that the intrarectal administration of EGCG could protect rhesus macaques from repetitive, intrarectal challenges with low-dose SHIVSF162P3N. This protection has a per-exposure risk reduction of 91.5% (P = 0.0009; log-rank test) and a complete protection of 87.5% (P < 0.001; Fisher’s exact test). All protected animals showed no evidence of systemic and mucosal SHIV infection as demonstrated by the absence of viral RNA, DNA and antibodies. In contrast, all controls became infected after repeated SHIV challenges (a median of 2.5 times, range of 1–8 times). Mechanistically, EGCG could block the binding of HIV-1 gp120 to CD4 receptor and suppress the macrophage infiltration/activation in the rectal mucosa of macaques. These data support further clinical evaluation and development of EGCG as a novel, safe and cost-effective microbicide for preventing sexual transmission of HIV-1.