The mechanisms of action of vaccines containing aluminum adjuvants: an in vitro vs in vivo paradigm

Abstract

Adjuvants such as the aluminum compounds (alum) have been dominantly used in many vaccines due to their immunopotentiation and safety records since 1920s. However, how these mineral agents influence the immune response to vaccination remains elusive. Many hypotheses exist as to the mode of action of these adjuvants, such as depot formation, antigen (Ag) targeting, and the induction of inflammation. These hypotheses are based on many in vitro and few in vivo studies. Understanding how cells interact with adjuvants in vivo will be crucial to fully understanding the mechanisms of action of these adjuvants. Interestingly, how alum influences the target cell at both the cellular and molecular level, and the consequent innate and adaptive responses, will be critical in the rational design of effective vaccines against many diseases. Thus, in this review, mechanisms of action of alum have been discussed based on available in vitro vs in vivo evidences to date.

Keywords: Alum internalization; Antigen targeting; Depot; Inflammasome; Innate and adaptive.

Figure 1

Current understanding of immunology of vaccines containing alum adjuvants in vivo. Alum immunization leads to recruitment of neutrophil, natural killer cell, macrophage, eosinophil, and immature DC at the injection site. Immature DCs take up soluble Ag released from alum or particulate Ags mixed in alum in the subcutaneous areas and migrate towards draining lymph node (DLN). Soluble Ag can reach to DLN without help of DCs. In T cell area (paracortex), soluble Ags leaked out of conduits are taken up by resident DCs. DCs present Ag to the naïve T cells or transfer Ag to the resident DCs that present Ag to those T cells. In addition, B cells are capable of binding to this Ag with their surface immunoglobulins. B cells undergo activation, produce effector B cells (eB cells), and rapidly differentiate in plasma cells (PCs). Plasma cells produce low-affinity antibodies (LAb). B cells also migrate to B cell follicle. On the other hand, as a result of CD8 and CD4 T cell activation, effector CD8 (eCD8) T cells and effector CD4 (eCD4) T cells are produced. The eCD4 polarizes into T helper (Th) 1, 2, 17 or T follicular helper (Tfh) cells though alum induces development of mostly Th2 and Tfh cells. Tfh or Th2 may reach to the border of B cell follicle to activate B cells that produce eB cells and then PCs. The PCs consequently produce and secrete high-affinity antibody (HAb). Few alum-fed DCs and eCD4 may travel to efferent lymph vessels to reach into the distant LN in vivo. Ag: antigen, HEV: high endothelial venule, PVS: perivenular space (see text for explanation).

Figure 2

EαGFP:YAe system can be used to assess antigen (Ag) uptake, processing, and presentation by DCs in the presence of alum in vitro and in vivo. (A): When EαGFP is taken up by DCs, it is processed and presented in the context of Eα(52–68):MHCII complexes. (B): The Eα(52–68):MHCII complexes can be bound by YAe antibody (Ab). (C): T cell receptor (TCR) of TEa mice sees the same complexes what YAe sees. Alum enhances Ag uptake, reduces degradation, and eventually increases presentation by DCs in vitro. This adjuvant also enhances the expression of CD86, CD80, and CD40 molecules by DCs in vitro. Arrow shows Ag processing path.

Figure 3

Hypotheses of NALP3 activation via inflammatory pathway. (A1): Alum activates XOR and induces the secretion of uric acid (UA). (A2): Alum-adsorbed antigen (Ag) is internalized by DC via actin-polymerization pathway. (A3): Alum interacts with lipid raft of DC membrane and activates ITAM-Syk pathway to internalize Ag. (A4): Alum-induced production of monosodium urate (MSU) crystals may be internalized by DC in CD16/32-dependent pathway. (B): Alum in phagosome enhances maturation into phagolysosome and release of Al³⁺ and OH⁻. (C): Al³⁺ inhibits the cathepsin L activity and consequently the generated stress induces the rupture of phagolysosomes. (C1): The leaked Ag may bind with MHCI molecule in cytoplasm. (C2): Nanometer-sized Al³⁺ ion precipitates in the cytosol. (C3): The released K⁺ will be exported outside and extracellular adenosine tri-phosphate (ATP) will enter inside via P2X7 molecule. (C4): Cathepsin molecule will be released in the cytosol. (C5): MSU is released in cytosol. (D): Reactive oxygen species (ROS) is generated. The ROS will activate various molecules such as (D1) unknown or (D2) Heat shock protein (HSP)70 that can trigger nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling and induces the secretion of IL-1β (D3). (E): Mitochondrion becomes activated and generates ROS (E1) and mitochondrial deoxyribonucleic acid (DNA) (E2). (F): C3, C4, C5, D1, D2, E1, or E2 may activate nucleotide-binding oligomerization domain family-like receptor pyrin domain containing 3 (NALP3) and consequently AC-1 (G). (H): Necrosis-induced release of double stranded (ds)DNA enters cytoplasm and initiates toll-like receptor (TLR)9–interferon regulatory factor (IRF)-3 signaling (I). (J): MSU molecules directly activate myeloid differentiation (MYD)88 signaling. (K): Pro-IL-1β and IL-18 may be transcripted from nucleus. (L): Active caspase (AC)-1 induces the secretion of their matured forms. (M): immunoreceptor tyrosine-based activation motif (ITAM)-Syk-PI3Kδ pathway activates secretion of prostaglandin E2 (PGE2). (N): Alum activates complement.