Overcoming HBV immune tolerance to eliminate HBsAg-positive hepatocytes via pre-administration of GM-CSF as a novel adjuvant for a hepatitis B vaccine in HBV transgenic mice

Granulocyte-macrophage colony-stimulating factor （GM-CSF） is known to be a potential vaccine adjuvant despite contradictory results from animal and human studies. The discrepancies may be due to the different doses and regimens of GM-CSF that were used, given that either mature or immature dendritic cells （DCs） could be induced under different conditions. To test the hypothesis that GM-CSF can be used as a novel adjuvant for a hepatitis B virus （HBV） therapeutic vaccine, we administered GM-CSF once per day for three days prior to vaccination with recombinant HBV vaccine （rHBVvac） in mice. We observed greater DC maturation in these pre-treated animals at day 3 as compared to day 1 or day 2 of daily GM-CSF administration. This strategy was further investigated for its ability to break the immune tolerance established in hepatitis B surface antigen-transgenic （HBsAg-Tg） animals. We found that the levels of induced anti-HBsAg antibodies were significantly higher in animals following three days of GM-CSF pre-treatment before rHBV vaccination after the third immunization. In addition to the increase in anti-HBsAg antibody levels, cell-mediated anti-HBsAg responses, including delayed-type hypersensitivity, T-cell proliferation, interferon-y production, and cytotoxic T lymphocytes, were dramatically enhanced in the three-day GM-CSF pre-treated group. After adoptive transfers of CD8＋ T cells from immunized animals, antigen-specific CD8＋ T cells were observed in the livers of recipient HBsAg-Tg animals. Moreover, the three-day pre-treatments with GM-CSF prior to rHBVvac vaccination could significantly eliminate HBsAg-positive hepatocytes, suggesting beneficial therapeutic effects. Therefore, this protocol utilizing GM-CSF as an adjuvant in combination with the rHBVvac vaccine has the potential to become a novel immunotherapy for chronic hepatitis B patients.

Activation of the Melanocortin-4 receptor signaling byα-MSH stimulates nervedependent mouse digit regeneration

Background:Expression of Mc4r in peripheral organs indicates it has broader roles in organ homeostasis and regeneration.However,the expression and function of Mc4r in the mouse limb and digit has not been fully investigated.Our previous work showed that Mc4r−/−mice fail to regenerate the digit,but whether activation of MC4R signaling could rescue digit regeneration,or stimulate proximal digit regeneration is not clear.Results:We analyzed the expression dynamics of Mc4r in the embryonic and postnatal mouse limb and digit using the Mc4r-gfp mice.We found that Mc4r-GFP is mainly expressed in the limb nerves,and in the limb muscles that are undergoing secondary myogenesis.Expression of Mc4r-GFP in the adult mouse digit is restricted to the nail matrix.We also examined the effect ofα-MSH on mouse digit regeneration.We found that administration ofα-MSH in the Mc4r+/−mice rescue the delayed regeneration of distal digit tip.α-MSH could rescue distal digit regeneration in denervated hindlimbs.In addition,α-MSH could stimulate regeneration of the proximally amputated digit,which is non-regenerative.Conclusions:Mc4r expression in the mouse limb and digit is closely related to nerve tissues,andα-MSH/MC4R signaling has a neurotrophic role in mouse digit tip regeneration.