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Amyloid‐β oligomers (Aβo) are the most pathologically relevant Aβ species in Alzheimer's disease (AD), because they induce early synaptic dysfunction that leads to learning and memory impairments. In contrast, increasing VEGF (Vascular Endothelial Growth Factor) brain levels have been shown to improve learning and memory processes, and to alleviate Aβ‐mediated synapse dysfunction. Here, we designed a new peptide, the blocking peptide (BP), which is derived from an Aβo‐targeted domain of the VEGF protein, and investigated its effect on Aβ‐associated toxicity. Using a combination of biochemical, 3D and ultrastructural imaging, and electrophysiological approaches, we demonstrated that BP strongly interacts with Aβo and blocks Aβ fibrillar aggregation process, leading to the formation of Aβ amorphous aggregates. BP further impedes the formation of structured Aβo and prevents their pathogenic binding to synapses. Importantly, acute BP treatment successfully rescues long‐term potentiation (LTP) in the APP/PS1 mouse model of AD, at an age when LTP is highly impaired in hippocampal slices. Moreover, BP is also able to block the interaction between Aβo and VEGF, which suggests a dual mechanism aimed at both trapping Aβo and releasing VEGF to alleviate Aβo‐induced synaptic damage. Our findings provide evidence for a neutralizing effect of the BP on Aβ aggregation process and pathogenic action, highlighting a potential new therapeutic strategy. [ABSTRACT FROM AUTHOR]

Tetraspanins, a transmembrane protein is a 'molecular organiser' with diverse functions that promote a network of signalling involved in the cellular and pathological processes. Tetraspanins-enriched microdomains (TEMs) are the crucial feature for the protein-protein interactions in the cell membrane and are vulnerable to pathogens exploitation. Targetting the tetraspanins has shown to be effective against microbial infections although more research is needed to be performed. This article will review previous evidence that has successfully demonstrated the potential mechanism to target tetraspanins as an antimicrobial agent. [ABSTRACT FROM AUTHOR]

Filopodia are sensors which, along with microtubules, regulate the persistence of locomotion. To determine whether protrusions were involved in sensing adhesion, epithelial cells were cultured on platinum and tantalum gradients. Protrusions were defined by an unbiased statistical method of classification as factors 4 (filopodia), 5 (mass distribution), and 7 (nascent neurites). When the prevalence of protrusions was measured in zones of high (H), middle (M), and low (L) adhesiveness, the main differences were in factor 4. Its values were highest at H and declined at M and L regardless of the gradient composition. The significance of the differences was enhanced when T (top/adhesive end) and B (bottom/nonadhesive end) sides of cells were analyzed separately. Since information about sidedness increased the statistical power of the test, this result suggested that cells pointed more filopodia toward the adhesive end. Trends occurred in factors 5 and 7 only when conditions allowed for a marked trend in factor 4. The data showed that gradient sensing is proportional to the prevalence of filopodia, and filopodia are the only protrusions engaged in comparing adhesiveness across a cell. The probability (P) of the significance of a trend was then used to determine how cells sense the gradient. Binding peptides (BPs) were introduced representing sequences critical for Cdc42 docking on a specific partner. BPs for IQGAP (IQ(calmodulin-binding domain)-containing GTPase-activating protein) and ACK (Cdc42-associated kinase) reduced factor 4 values and prevented cell orientation on the gradient. Micrographs showed attenuated or stubby filopodia. These effectors may be implicated in gradient sensing. Another IQGAP BP increased filopodia prevalence and enhanced orientation on the gradient (P < 0.00015). A Wiskott–Aldrich syndrome protein (WASP) BP had no effect. When sensing and orientation were abolished, they both failed at the level of filopodia, indicating that filopodia are both sensors and implementers of signals transduced by adhesion. [ABSTRACT FROM AUTHOR]

Transmigration of neutrophils through the microvascular endothelium is a cardinal event of acute inflammation. It has been suggested that gap junctions made of connexin43 (Cx43) may serve as a conducting pathway to spread inflammatory signals within the lung capillary network. To determine whether Cx43 contributes to neutrophil transmigration in vivo, the number of transmigrated neutrophils was monitored in lungs of Cx43 mouse models subjected to inflammation by intratracheal instillations of Pseudomonas aeruginosa lipopolysaccharide (LPS). Cx43 was detected in inflamed lungs independently of neutrophil recruitment, whereas Cx43 up-regulation was not detected in mice genetically protected from inflammation. Mice heterozygous for the Cx43 gene ( gja1) showed a 56% ( P < 0.01) reduction in airway neutrophil count. In contrast, increased ( P < 0.05) neutrophil recruitment in response to LPS was observed in a mouse model expressing a mutant Cx43 with enhanced channel conductivity. In vitro adhesion assays showed that reduced conductivity of Cx43 channels with 43Gap26, a Cx43 blocking peptide, decreased adhesion of neutrophils to endothelial cells. Finally, we found that instillation of 43Gap26 in inflamed lungs reduced neutrophil transmigration by 65% ( P < 0.05). These results indicate that inflammatory mediators up-regulate alveolar Cx43 to promote neutrophil recruitment to the airspace. Cx43 may therefore represent a pharmacological target in lung diseases characterized by excessive neutrophil recruitment to the airways. [ABSTRACT FROM AUTHOR]

Disruption of the CD40–CD154 interaction was found to be effective in the prevention and treatment of several immune-mediated diseases. The antibody-based strategy of inhibition was in humans limited by platelet activation leading to thrombotic effects. Other strategies different from antibody technology may be useful to create tools to interfere with CD40–CD154 pathway. In the present study, we selected and characterized from a phage display library, cyclic hepta-peptides specific for human CD154 through biopanning against plate-immobilized recombinant hCD154-muCD8. Nine phage clones were selected for the ability to bind CD154 expressed on the surface of J558L cells transfected with human CD154. From the nine selected phage clones, we obtained seven different amino acidic sequences, and the corresponding hepta-peptides rendered cyclic by two cysteines were synthesized. All the peptides specifically bound CD154 expressed on J558L. However, only the peptide 4.10 (CLPTRHMAC) was found to recognize the active binding site of CD154, as it competed with the blocking anti-CD154 antibody. When changes in the amino acid composition were introduced in the sequence of 4.10 peptide, the binding to CD154 was abrogated, suggesting that the amino acid sequence was critical for its specificity. This peptide was found to inhibit the CD40–CD154 interaction, preventing CD40-dependent activation of B lymphocytes in vitro as it was able, as the blocking anti-human CD154 mAb, to prevent the expression of CD80 and CD86 costimulatory molecules and switching of Ig isotype induced by CD154. Moreover, the peptide 4.10 inhibited the in vitro endothelial cell motility and organization into capillary-like structures, and the in vivo angiogenesis of human umbilical cord-derived endothelial cells implanted in Matrigel in severe combined immunodeficiency mice. In vitro studies on platelet activation demonstrated that the 4.10 peptide, at variance of the anti-CD154 mAb, was unable to prime human platelet activation and aggregation. In conclusion, we identify a cyclic hepta-peptide able to displace the binding of human CD154 to CD40 expressed on cell surface and to abrogate some biological effects related to the CD40 stimulation, such as B cell activation and endothelial triggered angiogenesis. [ABSTRACT FROM AUTHOR]

To assess the merits of PEGylated poly (lactic-co-glycolic acid) (PEG-PLGA) nanoparticles as drug carriers for tumor necrosis factor-α receptor blocking peptide (TNFR-BP), PEG-PLGA copolymer, which could be used to prepare the stealth nanoparticles, was synthesized with methoxypolyethyleneglycol, DL-lactide and glycolide. The structure of PEG-PLGA was confirmed with 1H-NMR and FT-IR spectroscopy, and the molecular weight (MW) was determined by gel permeation chromatography. Fluorescent FITC-TNFRBP was chosen as model protein and encapsulated within PEG-PLGA nanoparticles using the double emulsion method. Atomic force microscopy and photon correlation spectroscopy were employed to characterize the stealth nanoparticles fabricated for morphology, size with polydispersity index and zeta potential. Encapsulation efficiency (EE) and the release of FITC-TNFR-BP in nanoparticles in vitro were measured by the fluorescence measurement. The stealth nanoparticles were found to have the mean diameter less than 270 nm and zeta potential less than −20 mV. In all nanoparticle formulations, more than 45% of EE were obtained. FITC-TNFRBP release from the PEG-PLGA nanoparticles exhibited a biphasic pattern, initial burst release and consequently sustained release. The experimental results show that PEG-PLGA nanoparticles possess the potential to develop as drug carriers for controlled release applications of TNFR-BP. [ABSTRACT FROM AUTHOR]

Fibrosis is characterized by excessive production of disorganized collagen- and fibronectin-rich extracellular matrices (ECMs) and is driven by the persistence of myofibroblasts within tissues. A key protein contributing to myofibroblast differentiation is extra domain A fibronectin (EDA-FN). We sought to target and interfere with interactions between EDA-FN and its integrin receptors to effectively inhibit profibrotic activity and myofibroblast formation. Molecular docking was used to assist in the design of a blocking polypeptide (antifibrotic 38-amino-acid polypeptide, AF38Pep) for specific inhibition of EDA-FN associations with the fibroblast-expressed integrins α4β1 and α4β7. Blocking peptides were designed and evaluated in silico before synthesis, confirmation of binding specificity, and evaluation in vitro. We identified the high-affinity EDA-FN C-C′ loop binding cleft within integrins α4β1 and α4β7. The polypeptide with the highest predicted binding affinity, AF38Pep, was synthesized and could achieve specific binding to myofibroblast fibronectin-rich ECM and EDA-FN C-C′ loop peptides. AF38Pep demonstrated potent myofibroblast inhibitory activity at 10 µg/mL and was not cytotoxic. Treatment with AF38Pep prevented integrin α4β1-mediated focal adhesion kinase (FAK) activation and early signaling through extracellular-signal-regulated kinases 1 and 2 (ERK1/2), attenuated the expression of pro-matrix metalloproteinase 9 (MMP9) and pro-MMP2, and inhibited collagen synthesis and deposition. Immunocytochemistry staining revealed an inhibition of α-smooth muscle actin (α-SMA) incorporation into actin stress fibers and attenuated cell contraction. Increases in the expression of mRNA associated with fibrosis and downstream from integrin signaling were inhibited by treatment with AF38Pep. Our study suggested that AF38Pep could successfully interfere with EDA-FN C-C′ loop-specific integrin interactions and could act as an effective inhibitor of fibroblast of myofibroblast differentiation. [ABSTRACT FROM AUTHOR]

G protein-coupled receptors regulate a variety of cellular responses and have been considered as therapeutic targets for human diseases. Lysophosphatidic acid receptor 1 (LPA1) is a receptor for bioactive lysophospholipid, LPA. LPA/LPA1-mediated signaling contributes to inflammatory and fibrotic responses in lung diseases; thus understanding regulation of LPA1 stability is important for modulating LPA/LPA1 signaling. Our previous study has shown that LPA1 is degraded in the Nedd4 like (Nedd4L) E3 ubiquitin ligase-mediated ubiquitin-proteasome system. In the current study, we attempt to identify a peptide that stabilizes LPA1 through disrupting LPA1 association with Nedd4L. LPA treatment induces both endogenous and overexpressed LPA1 degradation, which is attenuated by a proteasome inhibitor, suggesting that LPA1 is degraded in the proteasome. LPA increases phosphorylation of extracellular signal-regulated kinase 1/2 (Erk1/2) and I-κB kinase in lung epithelial cells, and this effect is promoted by overexpression of a peptide (P1) that mimics C-terminal of LPA1. P1, not a control peptide, attenuates LPA-induced LPA1 ubiquitination and degradation, suggesting that P1 stabilizes LPA1. Further, P1 diminishes Nedd4L-mediated degradation of LPA1 and Nedd4L/LPA1 association. In addition to increasing LPA1 signaling, P1 enhances LPA-induced cell migration and gene expression of Elafin, matrix metallopeptidase 1, and serpin family B member 2 in lung epithelial cells. These data suggest that disruption of LPA1 interaction with Nedd4L by P1 increases LPA1 stability and LPA/LPA1 signaling., (© 2021 Wiley Periodicals LLC.)

• We find that there are highly homology in HR region of SARS-CoV-2 and SARS-CoV. • We firstly used molecular dynamics method in designing antivirus peptide. • Our antivirus peptide can prevent virus membrane fuse with host cell. • Furthermore, we first calculate the binding energy of HR1 and HR2 in SARS-CoV-2 spike protein. An outbreak caused by 2019 novel coronavirus (2019-nCoV) was first identified in Wuhan City, Hubei Province, China. The new virus was later named SARS-CoV-2. The virus has affected tens of thousands of patients in the world. The infection of SARS-CoV-2 causes severe pneumonia and even death. It is urgently needed to find a therapeutic method to treat patients with SARS-CoV-2 infection. Studies showed that the surface spike (S) protein is essential for the coronavirus binding and entry of host cells. The heptad repeats 1 and 2 (HR1 and HR2) in the S protein play a decisive role in the fusion of the viral membrane with the host cell membrane. We predicted the HR1 and HR2 regions in S protein by sequence alignment. We simulated a computational model of HR1/2 regions and the fusion core. The binding energy of HR1 and HR2 of the fusion core was –33.4 kcal/mol. We then designed antivirus peptides by molecular dynamics simulation of the fusion core. The binding energy of HR2-based antiviral peptide to HR1 was −43.0 kcal/mol, which was stronger than the natural stage of the fusion core, suggesting that the predicted antiviral peptide can competitively bind with HR1 to prevent forming of the fusion core. The antiviral peptides can prevent SARS-CoV-2 membrane fusion and can potentially be used for the prevention and treatment of infections. [ABSTRACT FROM AUTHOR]

The interaction between apoptosis-inducing factor (AIF) and cyclophilin A (CypA) has been shown to contribute to caspase-independent apoptosis. Blocking the AIF/CypA interaction protects against glutamate-induced neuronal cell death in vitro , and the purpose of this study was to determine the in vivo effect of an AIF/CypA interaction blocking peptide (AIF(370-394)-TAT) on neonatal mouse brain injury after hypoxia-ischemia (HI). The pups were treated with AIF (370-394)-TAT peptide intranasally prior to HI. Brain injury was significantly reduced at 72 h after HI in the AIF(370-394)-TAT peptide treatment group compared to vehicle-only treatment for both the gray matter and the subcortical white matter, and the neuroprotection was more pronounced in males than in females. Neuronal cell death was evaluated in males at 8 h and 24 h post-HI, and it was decreased significantly in the CA1 region of the hippocampus and the nucleus habenularis region after AIF(370-394)-TAT treatment. Caspase-independent apoptosis was decreased in the cortex, striatum, and nucleus habenularis after AIF(370-394)-TAT treatment, but no significant change was found on caspase-dependent apoptosis as indicated by the number of active caspase-3-labeled cells. Further analysis showed that both AIF and CypA nuclear accumulation were decreased after treatment with the AIF(370-394)-TAT peptide. These results suggest that AIF(370-394)-TAT inhibited AIF/CypA translocation to the nucleus and reduced HI-induced caspase-independent apoptosis and brain injury in young male mice, suggesting that blocking AIF/CypA might be a potential therapeutic target for neonatal brain injury. • Apoptotic cell death is more prominent in the immature brain after insult. • Apoptosis inducing factor (AIF) plays a crucial role in the process of apoptosis in the immature brain after injury. • AIF induces apoptosis require interaction with cyclophilin A. • Blocking interaction of AIF and cyclophilin A reduces caspase-independent cell death and brain injury only in males. [ABSTRACT FROM AUTHOR]

Tetraspanins are suggested to regulate the composition of cell membrane components and control intracellular transport, which leaves them vulnerable to utilization by pathogens such as human papillomaviruses (HPV) and cytomegaloviruses (HCMV) to facilitate host cell entry and subsequent infection. In this study, by means of cellular depletion, the cluster of differentiation (CD) tetraspanins CD9, CD63, and CD151 were found to reduce HPV16 infection in HeLa cells by 50 to 80%. Moreover, we tested recombinant proteins or peptides of specific tetraspanin domains on their effect on the most oncogenic HPV type, HPV16, and HCMV. We found that the C-terminal tails of CD63 and CD151 significantly inhibited infections of both HPV16 and HCMV. Although CD9 was newly identified as a key cellular factor for HPV16 infection, the recombinant CD9 C-terminal peptide had no effect on infection. Based on the determined half-maximal inhibitory concentration (IC 50 ), we classified CD63 and CD151 C-terminal peptides as moderate to potent inhibitors of HPV16 infection in HeLa and HaCaT cells, and in EA.hy926, HFF (human foreskin fibroblast) cells, and HEC-LTT (human endothelial cell-large T antigen and telomerase) cells for HCMV, respectively. These results indicate that HPV16 and HCMV share similar cellular requirements for their entry into host cells and reveal the necessity of the cytoplasmic CD151 and CD63 C-termini in virus infections. Furthermore, this highlights the suitability of these peptides for functional investigation of tetraspanin domains and as inhibitors of pathogen infections.

Ornithine decarboxylase (ODC) is the sentinel enzyme in polyamine biosynthesis. Both ODC and polyamines regulate cell division, proliferation, and apoptosis. Sepiapterin reductase (SPR) catalyzes the last step in the biosynthesis of tetrahydrobiopterin (BH4), an essential cofactor of nitric oxide synthase, and has been implicated in neurological diseases but not yet in cancer. In this study, we present compelling evidence that native ODC and SPR physically interact, and we defined the individual amino acid residues involved in both enzymes using in silico protein-protein docking simulations. The resulting heterocomplex is a surprisingly compact structure, featuring two energetically and structurally equivalent binding modes both in monomer and in dimer conformations. The novel interaction between ODC and SPR proteins was confirmed under physiological conditions by co-immunoprecipitation and co-localization in neuroblastoma (NB) cells. Importantly, we showed that siRNA (small interfering RNA)-mediated knockdown of SPR expression significantly reduced endogenous ODC enzyme activity in NB cells, thus demonstrating the biological relevance of the ODC-SPR interaction. Finally, in a cohort of 88 human NB tumors, we found that high SPR mRNA expression correlated significantly with poor survival prognosis using a Kaplan-Meier analysis (log-rank test, P=5 × 10(-4)), suggesting an oncogenic role for SPR in NB tumorigenesis. In conclusion, we showed that ODC binds SPR and thus propose a new concept in which two well-characterized biochemical pathways converge via the interaction of two enzymes. We identified SPR as a novel regulator of ODC enzyme activity and, based on clinical evidence, present a model in which SPR drives ODC-mediated malignant progression in NB., (© 2013.)