Sulfation at Glycopolymer Side Chains Switches Activity at the Macrophage Mannose Receptor (CD206) In Vitro and In Vivo.

The mannose receptor (CD206) is an endocytic receptor expressed by selected innate immune cells and nonvascular endothelium, which plays a critical role in both homeostasis and pathogen recognition. Although its involvement in the development of several diseases and viral infections is well established, molecular tools able to both provide insight on the chemistry of CD206-ligand interactions and, importantly, effectively modulate its activity are currently lacking. Using novel SO4-3-Gal-glycopolymers targeting its cysteine-rich lectin ectodomain, this study uncovers and elucidates a previously unknown mechanism of CD206 blockade involving the formation of stable intracellular SO4-3-Gal-glycopolymer-CD206 complexes that prevents receptor recycling to the cell membrane. Further, we show that SO4-3-Gal glycopolymers inhibit CD206 both in vitro and in vivo, revealing hitherto unknown receptor function and demonstrating their potential as CD206 modulators within future immunotherapies.

Polymer Coated Oncolytic Adenovirus to Selectively Target Hepatocellular Carcinoma Cells.

Despite significant advances in chemotherapy, the overall prognosis of hepatocellular carcinoma (HCC) remains extremely poor. HCC targeting strategies were combined with the tumor cell cytotoxicity of oncolytic viruses (OVs) to develop a more efficient and selective therapeutic system. OVs were coated with a polygalactosyl-b-agmatyl diblock copolymer (Gal32-b-Agm29), with high affinity for the asialoglycoprotein receptor (ASGPR) expressed on the liver cell surface, exploiting the electrostatic interaction of the positively charged agmatine block with the negatively charged adenoviral capsid surface. The polymer coating altered the viral particle diameter (from 192 to 287 nm) and zeta-potential (from -24.7 to 23.3 mV) while hiding the peculiar icosahedral symmetrical OV structure, as observed by TEM. Coated OVs showed high potential therapeutic value on the human hepatoma cell line HepG2 (cytotoxicity of 72.4% ± 4.96), expressing a high level of ASGPRs, while a lower effect was attained with ASPGR-negative A549 cell line (cytotoxicity of 54.4% ± 1.59). Conversely, naked OVs showed very similar effects in both tested cell lines. Gal32-b-Agm29 OV coating enhanced the infectivity and immunogenic cell death program in HepG2 cells as compared to the naked OV. This strategy provides a rationale for future studies utilizing oncolytic viruses complexed with polymers toward effective treatment of hepatocellular carcinoma.

Schwann cells are activated by ATP released from neurons in an in vitro cellular model of Miller Fisher syndrome.

The neuromuscular junction is exposed to different types of insult, including mechanical trauma, toxins and autoimmune antibodies and, accordingly, has retained through evolution a remarkable ability to regenerate. Regeneration is driven by multiple signals that are exchanged among the cellular components of the junction. These signals are largely unknown. Miller Fisher syndrome is a variant of Guillain-Barré syndrome caused by autoimmune antibodies specific for epitopes of peripheral axon terminals. Using an animal model of Miller Fisher syndrome, we recently reported that a monoclonal anti-polysialoganglioside GQ1b antibody plus complement damages nerve terminals with production of mitochondrial hydrogen peroxide, which activates Schwann cells. Several additional signaling molecules are likely to be involved in the activation of the regeneration program in these cells. Using an in vitro cellular model consisting of co-cultured primary neurons and Schwann cells, we found that ATP is released by neurons injured by the anti-GQ1b antibody plus complement. Neuron-derived ATP acts as an alarm messenger for Schwann cells, where it induces the activation of intracellular pathways, including calcium signaling, cAMP and CREB, which, in turn, produce signals that promote nerve regeneration. These results contribute to defining the cross-talk taking place at the neuromuscular junction when it is attacked by anti-gangliosides autoantibodies plus complement, which is crucial for nerve regeneration and is also likely to be important in other peripheral neuropathies.

ATP Released by Injured Neurons Activates Schwann Cells.

Injured nerve terminals of neuromuscular junctions (NMJs) can regenerate. This remarkable and complex response is governed by molecular signals that are exchanged among the cellular components of this synapse: motor axon nerve terminal (MAT), perisynaptic Schwann cells (PSCs), and muscle fiber. The nature of signals that govern MAT regeneration is ill-known. In the present study the spider toxin α-latrotoxin has been used as tool to investigate the mechanisms underlying peripheral neuroregeneration. Indeed this neurotoxin induces an acute, specific, localized and fully reversible damage of the presynaptic nerve terminal, and its action mimics the cascade of events that leads to nerve terminal degeneration in injured patients and in many neurodegenerative conditions. Here we provide evidence of an early release by degenerating neurons of adenosine triphosphate as alarm messenger, that contributes to the activation of a series of intracellular pathways within Schwann cells that are crucial for nerve regeneration: Ca(2+), cAMP, ERK1/2, and CREB. These results contribute to define the cross-talk taking place among degenerating nerve terminals and PSCs, involved in the functional recovery of the NMJ.