Study on the Interaction of Algal Peptides on Virulence Factors of Helicobacter pylori: In Silico Approach.

In the Asian region, Helicobacter pylori infects about 80% populations, which is most leading cause of peptic ulcers, and it is an asymptomatic infection. Studies reported that the particular bacteria carry specific virulence factors that leads to severe complications. These virulence factors can be used as a drug targets to inhibit their growth and pathogenicity. Chronic infection with H. pylori virulence factors are CagA, VacA and HtrA positive strains the risk factor of gastric cancer. In this study, we aimed to study the antagonistic interaction pattern between the potential eight algal peptides against the virulence factors of H. pylori through in silico analysis intended to treat peptic ulcer and prevent the further complications such as cancer. The proteins of virulent factors are docked using C-Docker algorithm and calculated the bind energy of the complexes. The results showed that the peptide derived from a green alga, Tetradesmus sp. are active against the three virulent factors such as cag-A, vac-A, and Htr-A with multiple hydrogen, vdW, electrostatic interactions, and mild π-hydrophobic bindings with the libdock energy score for CagA, VacA and HtrA are 175.625, 158.603 and 89.397 kcal/mol. These primes and the peptide lead to develop a better and potential inhibitors against H. pylori infection.

Pyropia yezoensis protein protects against TNF­α­induced myotube atrophy in C2C12 myotubes via the NF­κB signaling pathway.

The protein extracted from red algae Pyropia yezoensis has various biological activities, including anti­inflammatory, anticancer, antioxidant, and antiobesity properties. However, the effects of P. yezoensis protein (PYCP) on tumor necrosis factor­α (TNF­α)­induced muscle atrophy are unknown. Therefore, the present study investigated the protective effects and related mechanisms of PYCP against TNF­α­induced myotube atrophy in C2C12 myotubes. Treatment with TNF­α (20 ng/ml) for 48 h significantly reduced myotube viability and diameter and increased intracellular reactive oxygen species levels; these effects were significantly reversed in a dose­dependent manner following treatment with 25­100 µg/ml PYCP. PYCP inhibited the expression of TNF receptor­1 in TNF­α­induced myotubes. In addition, PYCP markedly downregulated the nuclear translocation of nuclear factor­κB (NF­κB) by inhibiting the phosphorylation of inhibitor of κB. Furthermore, PYCP treatment suppressed 20S proteasome activity, IL­6 production, and the expression of the E3 ubiquitin ligases, atrogin­1/muscle atrophy F­box and muscle RING­finger protein­1. Finally, PYCP treatment increased the protein expression levels of myoblast determination protein 1 and myogenin in TNF­α­induced myotubes. The present findings indicate that PYCP may protect against TNF­α­induced myotube atrophy by inhibiting the proinflammatory NF­κB pathway.

Cyanobacteria and Eukaryotic Microalgae as Emerging Sources of Antibacterial Peptides.

Cyanobacteria and microalgae are oxygen-producing photosynthetic unicellular organisms encompassing a great diversity of species, which are able to grow under all types of extreme environments and exposed to a wide variety of predators and microbial pathogens. The antibacterial compounds described for these organisms include alkaloids, fatty acids, indoles, macrolides, peptides, phenols, pigments and terpenes, among others. This review presents an overview of antibacterial peptides isolated from cyanobacteria and microalgae, as well as their synergism and mechanisms of action described so far. Antibacterial cyanopeptides belong to different orders, but mainly from Oscillatoriales and Nostocales. Cyanopeptides have different structures but are mainly cyclic peptides. This vast peptide repertoire includes ribosomal and abundant non-ribosomal peptides, evaluated by standard conventional methodologies against pathogenic Gram-negative and Gram-positive bacteria. The antibacterial activity described for microalgal peptides is considerably scarcer, and limited to protein hydrolysates from two Chlorella species, and few peptides from Tetraselmis suecica. Despite the promising applications of antibacterial peptides and the importance of searching for new natural sources of antibiotics, limitations still persist for their pharmaceutical applications.

Dietary osteopontin-enriched algal protein as nutritional support in weaned pigs infected with F18-fimbriated enterotoxigenic Escherichia coli.

This study investigated the effects of dietary osteopontin (OPN)-enriched algal protein on growth, immune status, and fecal fermentation profiles of weaned pigs challenged with a live infection of F18-fimbriated enterotoxigenic E. coli (ETEC). At 21 d of age, 54 pigs (5.95 ± 0.28 kg BW; blocked by BW) were allotted to 1 of 3 experimental groups combining dietary and health statuses. A control diet, containing 1% wild-type algal protein, was fed to both sham-inoculated (NC) and ETEC-inoculated (PC) pigs, while the test diet contained 1% OPN-enriched algal protein as fed only to ETEC-inoculated pigs (OA). All pigs received their assigned dietary treatment starting at study initiation to permit a 10-d acclimation period prior to inoculation. Growth performance, fecal dry matter, as well as hematological, histopathological, immune, and microbiota outcomes were analyzed by ANOVA, where treatment and time were considered as fixed effects and pig as a random effect; significance was accepted at P < 0.05. Overall, ETEC-inoculated pigs (PC and OA) exhibited decreased (P < 0.05) ADG and G:F, as well as increased (P < 0.05) peripheral blood helper T-cells and total leukocyte counts, compared with NC pigs during the postinoculation period. The OA treatment also elicited the highest (P < 0.05) concentrations of circulating tumor necrosis factor-α and volatile fatty acid concentrations in luminal contents at various postinoculation time-points, compared with other treatments. A principal coordinate analysis based on Unifrac weighted distances indicated that NC and OA groups had similar overall bacterial community structures, while PC pigs exhibited greater diversity, but infection status had no impact on α-diversity. Osteopontin-specific effects on microbial community structure included enrichment within Streptococcus and Blautia genera and decreased abundance of 12 other genera as compared with PC pigs. Overall, ETEC-infected pigs receiving 1% OPN-enriched algal protein exhibited changes immunity, inflammatory status, and colonic microbial community structure that may benefit weanling pigs experiencing F18 ETEC infection.

In Silico Analysis of ACE Inhibitory Peptides from Chloroplast Proteins of Red Alga Grateloupia asiatica.

Inhibition of angiotensin I-converting enzyme (ACE) is one of the key factors to repress high blood pressure. Although many studies have been reported that seaweed protein hydrolysates showed the ACE inhibitory activity, the comprehensive understanding of the relationship was still unclear. In this study, we employed chloroplast genome for in silico analysis and compared it with in vitro experiments. We first extracted water-soluble proteins (WSP) from red alga Grateloupia asiatica, which contained mainly PE, PC, APC, and Rbc, and prepared WSP hydrolysate by thermolysin, resulting that the hydrolysate showed ACE inhibitory activity. Then, we determined the complete chloroplast genome of G. asiatica (187,518 bp: 206 protein-coding genes, 29 tRNA, and 3 rRNA) and clarified the amino acid sequences of main WSP, i.e., phycobiliproteins and Rubisco, to perform in silico analysis. Consequently, 190 potential ACE inhibitory peptides existed in the main WSP sequences, and 21 peptides were obtained by in silico thermolysin digestion. By comparing in vitro and in silico analyses, in vitro ACE inhibitory activity was correlated to the IC50 value from in silico digestion. Therefore, in silico approach provides insight into the comprehensive understanding of the potential bioactive peptides from seaweed proteins.

Characterization of a Novel Mannose-Binding Lectin with Antiviral Activities from Red Alga, Grateloupia chiangii.

Lectins have the ability to bind specific carbohydrates and they have potential applications as medical and pharmacological agents. The unique structure and usefulness of red algal lectin have been reported, but these lectins are limited to a few marine algal groups. In this study, a novel mannose-binding lectin from Grateloupia chiangii (G. chiangii lectin, GCL) was purified using antiviral screens and affinity chromatography. We characterized the molecular weight, agglutination activity, hemagglutination activity, and heat stability of GCL. To determine the carbohydrate specificity, a glycan microarray was performed. GCL showed strong binding affinity for Maltohexaose-ß-Sp1 and Maltoheptaose-ß-Sp1 with weak affinity for other monosaccharides and preferred binding to high-mannan structures. The N-terminal sequence and peptide sequence of GCL were determined using an Edman degradation method and LC-MS/MS, and the cDNA and peptide sequences were deduced. GCL was shown to consist of 231 amino acids (24.9 kDa) and the N-terminus methionine was eliminated after translation. GCL possessed a tandem repeat structure of six domains, similar to the other red algal lectins. The mannose binding properties and tandem repeat structure of GCL may confer it the potential to act as an antiviral agent for protection against viral infection.

Phycocyanin Extracted from Oscillatoria minima Shows Antimicrobial, Algicidal, and Antiradical Activities: In silico and In vitro Analysis.

BACKGROUND: Phycocyanin is an algae-derived protein, which binds to pigment for harvesting light. It has been reported in various different species, including that of red algae, dinoflagellates, and cryptophyta. Importantly, phycocyanin has enormous applications, including cosmetic colorant, food additive, biotechnology, diagnostics, fluorescence detection probe, an anticancer agent, anti-inflammatory, immune enhancer, etc. In addition, several different algae were utilized for the isolation of cyano-phycocyanin (C-PC), but most of the purification methods consist of several steps of crude extraction. AIM: To isolate C-PC from a new source of microalgae with better purity level and to evaluate its antimicrobial, algicidal, and antiradical activities. METHODS: Biological activity, permeability, pharmacokinetics, and toxicity profile of C-PC were predicted by in silico studies. C-PC was purified and isolated by using ammonium sulphate precipitation, ion-exchange chromatography and gel-filtration chromatography. C-PC was characterized by SDS-PAGE and elution profile (purity ratio) analysis. Antimicrobial and algicial activities of C-PC were evaluated by the microtitre plate based assays. Antiradical activity of C-PC was evaluated by DPPH- and ABTS*+ radical scavenging assays. CONCLUSION: C-PC was extracted from Oscillatoria minima for the first time, followed by its quantitative as well qualitative evaluation, indicating a new alternative source of this important protein. Furthermore, the antimicrobial, algicidal, and antiradical activities of the isolated C-PC extract have been demonstrated by both in silico as well as in vitro methods.

Spirulina Crude Protein Promotes the Migration and Proliferation in IEC-6 Cells by Activating EGFR/MAPK Signaling Pathway.

Spirulina is a type of filamentous blue-green microalgae known to be rich in nutrients and to have pharmacological effects, but the effect of spirulina on the small intestine epithelium is not well understood. Therefore, this study aims to investigate the proliferative effects of spirulina crude protein (SPCP) on a rat intestinal epithelial cells IEC-6 to elucidate the mechanisms underlying its effect. First, the results of wound-healing and cell viability assays demonstrated that SPCP promoted migration and proliferation in a dose-dependent manner. Subsequently, when the mechanisms of migration and proliferation promotion by SPCP were confirmed, we found that the epidermal growth factor receptor (EGFR) and mitogen-activated protein (MAPK) signaling pathways were activated by phosphorylation. Cell cycle progression from G0/G1 to S phase was also promoted by SPCP through upregulation of the expression levels of cyclins and cyclin-dependent kinases (Cdks), which regulate cell cycle progression to the S phase. Meanwhile, the expression of cyclin-dependent kinase inhibitors (CKIs), such as p21 and p27, decreased with SPCP. In conclusion, our results indicate that activation of EGFR and its downstream signaling pathway by SPCP treatment regulates cell cycle progression. Therefore, these results contribute to the research on the molecular mechanism for SPCP promoting the migration and proliferation of rat intestinal epithelial cells.

Unravelling the inhibitory activity of Chlamydomonas reinhardtii sulfated polysaccharides against α-Synuclein fibrillation.

α-Synuclein (α-Syn) is an intrinsically disordered presynaptic protein, whose aggregation is critically involved in Parkinson's disease (PD). Many of the currently available drugs for the treatment of PD are not sufficiently effective in preventing progress of the disease and have multiple side-effects. With this background, efficient drug candidates, sulfated polysaccharides from Chlamydomonas reinhardtii (Cr-SPs) were isolated and investigated for their effect on inhibition of α-Syn fibrillation and dissolution of preformed α-Syn fibrillar structures through a combination of spectroscopic and microscopic techniques. The kinetics of α-Syn fibrillation demonstrates that Cr-SPs are very effective in inhibiting α-Syn fibrillation. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis gel-image shows presence of soluble protein in the presence of Cr-SPs after completion of the fibrillation process. The morphological changes associated with fibrillation monitored by transmission electron microscopy showed that Cr-SPs efficiently bind with α-Syn and delay the conversion of α-helical intermediate into ß-sheet rich structures. Cr-SPs are also effective even if onset of α-Syn fibrillation has already started and they also have the ability to dissolve pre-formed fibrils. Thus, the current work has substantial therapeutic implications towards unlocking the immense potential of algal products to function as alternative therapeutic agents against PD and other protein aggregation related disorders.

Isolation and identification of anti-proliferative peptides from Spirulina platensis using three-step hydrolysis.

BACKGROUND: Spirulina platensis is an excellent source of proteins (>60%) that can be hydrolyzed into bioactive peptides. RESULTS: In this study, whole proteins of Spirulina platensis were extracted and hydrolyzed using three gastrointestinal endopeptidases (pepsin, trypsin and chymotrypsin). Subsequently, gel filtration chromatography was employed to separate hydrolysates, and four fractions (Tr1-Tr4) were obtained. Among them, Tr2 showed the strongest anti-proliferation activities on three cancer cells (MCF-7, HepG-2 and SGC-7901), with IC50 values of <31.25, 36.42 and 48.25 µg mL-1 , respectively. Furthermore, a new peptide, HVLSRAPR, was identified from fraction Tr1. This peptide exhibited strong inhibition on HT-29 cancer cells with an IC50 value of 99.88 µg mL-1 . CONCLUSION: Taken together, these peptides possessed anti-proliferation activities on cancer cells and low cytotoxicity on normal cells, suggesting that they might serve as a natural anticancer agent for nutraceutical and pharmaceutical industries. © 2016 Society of Chemical Industry.

Griffithsin-Modified Electrospun Fibers as a Delivery Scaffold To Prevent HIV Infection.

Despite current prophylactic strategies, sexually transmitted infections (STIs) remain significant contributors to global health challenges, spurring the development of new multipurpose delivery technologies to protect individuals from and treat virus infections. However, there are few methods currently available to prevent and no method to date that cures human immunodeficiency virus (HIV) infection or combinations of STIs. While current oral and topical preexposure prophylaxes have protected against HIV infection, they have primarily relied on antiretrovirals (ARVs) to inhibit infection. Yet continued challenges with ARVs include user adherence to daily treatment regimens and the potential toxicity and antiviral resistance associated with chronic use. The integration of new biological agents may avert some of these adverse effects while also providing new mechanisms to prevent infection. Of the biologic-based antivirals, griffithsin (GRFT) has demonstrated potent inhibition of HIV-1 (and a multitude of other viruses) by adhering to and inactivating HIV-1 immediately upon contact. In parallel with the development of GRFT, electrospun fibers (EFs) have emerged as a promising platform for the delivery of agents active against HIV infection. In the study described here, our goal was to extend the mechanistic diversity of active agents and electrospun fibers by incorporating the biologic GRFT on the EF surface rather than within the EFs to inactivate HIV prior to cellular entry. We fabricated and characterized GRFT-modified EFs (GRFT-EFs) with different surface modification densities of GRFT and demonstrated their safety and efficacy against HIV-1 infection in vitro We believe that EFs are a unique platform that may be enhanced by incorporation of additional antiviral agents to prevent STIs via multiple mechanisms.

High-Mannose Specific Lectin and Its Recombinants from a Carrageenophyta Kappaphycus alvarezii Represent a Potent Anti-HIV Activity Through High-Affinity Binding to the Viral Envelope Glycoprotein gp120.

We previously reported that a high-mannose binding lectin KAA-2 from the red alga Kappaphycus alvarezii, which is an economically important species and widely cultivated as a source of carrageenans, had a potent anti-influenza virus activity. In this study, the full-length sequences of two KAA isoforms, KAA-1 and KAA-2, were elucidated by a combination of peptide mapping and cDNA cloning. They consisted of four internal tandem-repeated domains, which are conserved in high-mannose specific lectins from lower organisms, including a cyanobacterium Oscillatoria agardhii and a red alga Eucheuma serra. Using an Escherichia coli expression system, an active recombinant form of KAA-1 (His-tagged rKAA-1) was successfully generated in the yield of 115 mg per a litter of culture. In a detailed oligosaccharide binding analysis by a centrifugal ultrafiltration-HPLC method with 27 pyridylaminated oligosaccharides, His-tagged rKAA-1 and rKAA-1 specifically bound to high-mannose N-glycans with an exposed α1-3 mannose in the D2 arm as the native lectin did. Predicted from oligosaccharide-binding specificity, a surface plasmon resonance analysis revealed that the recombinants exhibit strong interaction with gp120, a heavily glycosylated envelope glycoprotein of HIV with high association constants (1.48-1.61 × 10(9) M(-1)). Native KAAs and the recombinants inhibited the HIV-1 entry at IC50s of low nanomolar levels (7.3-12.9 nM). Thus, the recombinant proteins would be useful as antiviral reagents targeting the viral surface glycoproteins with high-mannose N-glycans, and the cultivated alga K. alvarezii could also be a good source of not only carrageenans but also this functional lectin(s).

Enzyme-assisted extraction of bioactive material from Chondrus crispus and Codium fragile and its effect on herpes simplex virus (HSV-1).

Codium fragile and Chondrus crispus are, respectively, green and red seaweeds which are abundant along the North Atlantic coasts. We investigated the chemical composition and antiviral activity of enzymatic extracts of C. fragile (CF) and C. crispus (CC). On a dry weight basis, CF consisted of 11% protein, 31% neutral sugars, 0.8% sulfate, 0.6% uronic acids, and 49% ash, while CC contained 27% protein, 28% neutral sugars, 17% sulfate, 1.8% uronic acids, and 25% ash. Enzyme-assisted hydrolysis improved the extraction efficiency of bioactive materials. Commercial proteases and carbohydrases significantly improved (p ≤ 0.001) biomass yield (40%-70% dry matter) as compared to aqueous extraction (20%-25% dry matter). Moreover, enzymatic hydrolysis enhanced the recovery of protein, neutral sugars, uronic acids, and sulfates. The enzymatic hydrolysates exhibited significant activity against Herpes simplex virus (HSV-1) with EC50 of 77.6-126.8 µg/mL for CC and 36.5-41.3 µg/mL for CF, at a multiplicity of infection (MOI) of 0.001 ID50/cells without cytotoxity (1-200 µg/mL). The extracts obtained from proteases (P1) and carbohydrases (C3) were also effective at higher virus MOI of 0.01 ID50/cells without cytotoxity. Taken together, these results indicate the potential application of enzymatic hydrolysates of C. fragile and C. crispus in functional food and antiviral drug discovery.

An S-type anion channel SLAC1 is involved in cryptogein-induced ion fluxes and modulates hypersensitive responses in tobacco BY-2 cells.

Pharmacological evidence suggests that anion channel-mediated plasma membrane anion effluxes are crucial in early defense signaling to induce immune responses and hypersensitive cell death in plants. However, their molecular bases and regulation remain largely unknown. We overexpressed Arabidopsis SLAC1, an S-type anion channel involved in stomatal closure, in cultured tobacco BY-2 cells and analyzed the effect on cryptogein-induced defense responses including fluxes of Cl(-) and other ions, production of reactive oxygen species (ROS), gene expression and hypersensitive responses. The SLAC1-GFP fusion protein was localized at the plasma membrane in BY-2 cells. Overexpression of SLAC1 enhanced cryptogein-induced Cl(-) efflux and extracellular alkalinization as well as rapid/transient and slow/prolonged phases of NADPH oxidase-mediated ROS production, which was suppressed by an anion channel inhibitor, DIDS. The overexpressor also showed enhanced sensitivity to cryptogein to induce downstream immune responses, including the induction of defense marker genes and the hypersensitive cell death. These results suggest that SLAC1 expressed in BY-2 cells mediates cryptogein-induced plasma membrane Cl(-) efflux to positively modulate the elicitor-triggered activation of other ion fluxes, ROS as well as a wide range of defense signaling pathways. These findings shed light on the possible involvement of the SLAC/SLAH family anion channels in cryptogein signaling to trigger the plasma membrane ion channel cascade in the plant defense signal transduction network.

Molecular characteristics and biological activities of anionic macromolecules from Codium fragile.

Water-soluble anionic macromolecules isolated from Codium fragile and fractionated using ion-exchange chromatography were investigated to determine their molecular characteristics and immunostimulating activity. The crude molecules and fractions (F1, F2, and F3) consisted mostly of carbohydrates (44.1-80.5%), sulfates (3.2-22.2%) and proteins (3.0-15.7%) with small amounts of uronic acids (1.1-4.2%), and included different levels of mannose (91.3-18.7%), glucose (62.7-8.6%) and galactose (37.5-59.5%). These molecules contained one or two subfractions with molecular weights (Mw) ranging from 148×10(3) to 4879×10(3)g/mol. The crude, F1 and F2 stimulated RAW264.7 cells to produce considerable amounts of pro-inflammatory mediator nitric oxide (NO) and cytokines. The treatment of sample molecules facilitated the degradation of Iκ-B and phosphorylation of MAPK in RAW264.7 cells, suggesting that they might stimulate RAW264.7 cells through the activation of NF-κB and MAPK pathway. Proteins in fraction F2 were essential to possess its bioactivity and its main backbone was composed of mixed linkages of (1→3)-α and ß-d-mannan.

Algal lectins as potential HIV microbicide candidates.

The development and use of topical microbicides potentially offers an additional strategy to reduce the spread of the Human Immunodeficiency Virus (HIV). Carbohydrate-binding agents (CBAs) that show specificity for high mannose carbohydrates on the surface of the heavily glycosylated envelope of HIV are endowed with potent anti-HIV activity. In fact, a number of algal lectins such as cyanovirin-N, microvirin, microcystis viridis lectin, scytovirin, Oscillatoria agardhii agglutinin and griffithsin are considered as potential microbicide candidates to prevent the sexual transmission of HIV through topical applications. They not only inhibit infection of cells by cell-free virus but they can also efficiently prevent virus transmission from virus-infected cells to uninfected CD4(+) target T-lymphocytes and DC-SIGN-directed capture of HIV-1 and transmission to CD4(+) T lymphocytes. This review focuses on the structural properties and carbohydrate specificity of these algal lectins, their antiviral activity against HIV and several other enveloped viruses, their safety profile and viral resistance patterns.

Induction of apoptosis signaling by glycoprotein of Capsosiphon fulvescens in human gastric cancer (AGS) cells.

Capsosiphon fulvescens is a well-known green sea algae that has been touted in recent years as a potential anticancer drug. In this study, C. fulvescens glycoprotein (Cf-GP) showed proapoptotic signaling in AGS cells. An MTS assay indicated that Cf-GP inhibited the proliferation of AGS cell lines in a dose-dependent manner. Cells were treated with Cf-GP and the expression of proteins associated with apoptosis was examined by Western blotting. Based on the Western blot, expression of Cf-GP-activated caspase-cascade and PARP, which is a substrate of caspase-3 and -8, and proteins of the Bcl-2 family was observed. Cf-GP treatment stimulated the release of cytochrome C and apoptotic protease activating factor-1 from mitochondria to the cytosol. Cf-GP inhibited the growth of AGS cells through induction of sub-G1 phase arrest. We confirmed that sub-G1-phase arrest was associated with a decrease in the expression of cyclin D, cyclin E, Cdk2, Cdk4, and Cdk6, and an increase in the protein levels of p21 and p27. As a result, the increased sub-G1 ratio appears to be inhibited by cell proliferation. Therefore, we can confirm apoptosis in the AGS cells. Our results suggest that Cf-GP could be a potential source of biofunctional material that shows anticancer effects in human gastrointestinal cancer.

The lectins griffithsin, cyanovirin-N and scytovirin inhibit HIV-1 binding to the DC-SIGN receptor and transfer to CD4(+) cells.

It is generally believed that during the sexual transmission of HIV-1, the glycan-specific DC-SIGN receptor binds the virus and mediates its transfer to CD4(+) cells. The lectins griffithsin (GRFT), cyanovirin-N (CV-N) and scytovirin (SVN) inhibit HIV-1 infection by binding to mannose-rich glycans on gp120. We measured the ability of these lectins to inhibit both the HIV-1 binding to DC-SIGN and the DC-SIGN-mediated HIV-1 infection of CD4(+) cells. While GRFT, CV-N and SVN were moderately inhibitory to DC-SIGN binding, they potently inhibited DC-SIGN-transfer of HIV-1. The introduction of the 234 glycosylation site abolished HIV-1 sensitivity to lectin inhibition of binding to DC-SIGN and virus transfer to susceptible cells. However, the addition of the 295 glycosylation site increased the inhibition of transfer. Our data suggest that GRFT, CV-N and SVN can block two important stages of the sexual transmission of HIV-1, DC-SIGN binding and transfer, supporting their further development as microbicides.

Occluding the mannose moieties on human immunodeficiency virus type 1 gp120 with griffithsin improves the antibody responses to both proteins in mice.

To assess the influence of mannosylated glycans on the immunogenicity of human immunodeficiency virus type 1 (HIV-1) Env proteins, we immunized mice with monomeric gp120 in the presence and absence of the mannose-binding protein, griffithsin (GRFT). For comparison, other groups of mice received the nonglycosylated HIV-1 Gag protein, with and without GRFT. Coimmunization with GRFT increased the anti-gp120 IgG reactivity significantly, but had no effect on the anti-Gag response. We also investigated the IgG response to GRFT and found that gp120, but not Gag, enhanced its immunogenicity. For both proteins, IgG1 antibodies dominated the IgG response, with IgG2b as the next most prevalent subclass. We conclude that gp120-GRFT complexes are more immunogenic than the free proteins, for both components, and that occluding the mannose moieties on monomeric gp120 can improve the humoral immune response to this protein.

Griffithsin has antiviral activity against hepatitis C virus.

Hepatitis C virus (HCV)-infected patients undergoing liver transplantation universally experience rapid reinfection of their new liver graft. Current treatment protocols do not prevent graft reinfection and, in addition, an accelerated disease progression is observed. In the present study, we have evaluated a novel strategy to prevent HCV infection using a lectin, griffithsin (GRFT) that specifically binds N-linked high-mannose oligosaccharides that are present on the viral envelope. The antiviral effect of GRFT was evaluated in vitro using the HCV pseudoparticle (HCVpp) and HCV cell culture (HCVcc) systems. We show here that preincubation of HCVpp and HCVcc with GRFT prevents infection of Huh-7 hepatoma cells. Furthermore, GRFT interferes with direct cell-to-cell transmission of HCV. GRFT acts at an early phase of the viral life cycle by interfering in a genotype-independent fashion with the interaction between the viral envelope proteins and the viral receptor CD81. The capacity of GRFT to prevent infection in vivo was evaluated using uPA(+/+)-SCID mice (uPA stands for urokinase-type plasminogen activator) that harbor human primary hepatocytes in their liver (chimeric mice). In this proof-of-concept trial, we demonstrated that GRFT can mitigate HCV infection of chimeric mice. Treated animals that did become infected demonstrated a considerable delay in the kinetics of the viral infection. Our data demonstrate that GRFT can prevent HCV infection in vitro and mitigate HCV infection in vivo. GRFT treatment of chronically infected HCV patients undergoing liver transplantation may be a suitable strategy to prevent infection of the liver allograft.