Complement, named because it assists antibodies in removing pathogens, is a group of about 20 proteins found in plasma. Complement is mainly derived from the liver or certain cell membrane proteins and is a class of soluble pattern recognition molecules in intrinsic immunity. It has a variety of effector functions and is the body’s first line of defense in removing foreign pathogens or mutated self-cells.
Complement can be activated through three different pathways: the classical, lectin, and alternative pathways. Each of these three pathways leads to the production of C3 convertases that cleave C3 into C3a and C3b, which are then involved in the formation of different complexes. The complement cascade then leads to C5 activation, initiating the formation of the C5b-7 complex and finally the membrane attack complex (MAC), which leads to cell lysis.
Complement activation mediates the removal of microorganisms and the clearance of aberrant self-cells (e.g., apoptotic cells). Complement regulatory factors (e.g., CD46 and CD55) control the spontaneous activation of the complement cascade response. Disruption of this delicate balance may lead to tissue damage and autoimmune disease.
Mesenchymal stem cells (MSCs) are widely present in the human body and interact very closely with the immune system. Most studies have focused on the interactions between MSCs and immune T cells, B cells, and NK cells, with very few articles published on the interactions between MSCs and complement.
MSC Inhibits Complement Activation
MSCs express soluble factor H and complement regulatory proteins CD46, CD55, and CD59, enabling them to inhibit the activation of the complement system to some extent. Coating and adsorption of soluble factor H on the surface of MSCs reduce the attack of MAC formed by complement activation.
Not only do MSCs produce soluble factor H to inhibit complement activation, but the inflammatory cytokines TNF-α and interferon-γ (IFN-γ) also increase the amount of soluble factor H secreted by MSCs in a dose- and time-dependent manner. Inhibitors of prostaglandin E2 (PGE2) synthesis and inhibitors of tryptophan dioxygenase (IDO) inhibit soluble factor H secretion.
In fungal or yeast peritonitis, mesothelial tissues show strong expression of membrane complement regulators such as Crry, CD55, and CD59. Administration of localized injections of adipose MSCs into the peritoneum blocks the deposition of complement-activated products, suppressing peritoneal inflammation and ameliorating peritoneal injury. Complement activation complex C5b-9 activates neutrophils to produce interleukin-17 (IL-17). However, cellular experiments revealed that MSC exosomes inhibited the activation of neutrophils by complement C5b-9, thereby limiting the exacerbation and worsening of inflammation.
Animal experiments found that after intravenous infusion of MSCs for the treatment of transient focal cerebral ischemia in mice, the content of complement C3 in the brain and blood of mice was reduced, suggesting that MSCs inhibited the activation of complement C3. Some foreign studies have shown that the mild inflammation detected after intravenous infusion of MSCs is likely related to complement activation.
Effect of Complement on MSCs
The complement protein C3b/iC3b is deposited on MSCs when they come into contact with blood, enhancing the phagocytosis of MSCs by monocytes. About 80% of MSCs are phagocytosed by C3-mediated monocytes. The addition of a C3 inhibitor reduces the deposition of C3b/iC3b on the surface of MSCs, thereby reducing the damage to MSCs by activated complement.
Although MSCs express three known complement regulators, the killing function of activated complement exceeds the protection of these regulators upon contact of MSCs with serum. The complement activation pathways are all involved in the production of MAC, which directly damages MSCs, leading to the emergence of MSC cytotoxicity and dysfunction. However, the sera currently used to culture MSCs have been treated with complement inactivation, so there is no need to worry about the cytotoxicity of complement to MSCs during their culture.
An MSC team at Karolinska University Hospital in Sweden, led by Prof. Katarina Le Blanc, found that their own cultured MSCs had low expression of the complement regulatory proteins CD46, CD55, and CD59. However, a team at Wake Forest School of Medicine in North Carolina, USA, found that the expression of CD46, CD55, and CD59 in MSCs was not low, and that the expression of CD59 was as high as 50%. Moreover, the use of gene editing to modify MSCs to overexpress US2 protein, thus promoting the high expression of CD59, CD55 and CD46 in MSCs, directly reduced the cytotoxic damage of MSCs by 59% due to complement activation.
MSCs cultured by different teams secrete different small molecules of proteins that inhibit complement activation, leading to different survival times in vivo, which, in turn, affect the therapeutic efficacy of MSCs. This may also partly explain why the MSC team at Karolinska University Hospital, Sweden, has always struggled to obtain significant positive therapeutic results in the field of MSC clinical research, i.e., there are certain problems with the culture of MSCs.
Not only can complement damage MSCs, but it has also been experimentally found that complement C1q can mediate MSC migration as chemotactic agents, enhancing the chemotactic response of MSCs to SDF-1. Complements C3a and C5a are also chemotactic agents for MSCs and both cause persistent phosphorylation of the signaling molecules ERK1/2 and Akt.
In summary, there is no strong experimental data demonstrating that MSCs activate the complement system after entering the body. Numerous clinical applications have demonstrated that MSC treatment does not cause inflammation. In addition, after MSCs enter the body, whether they are autologous or allogeneic, they must be removed by the body. It is likely, then, that the activation of complement is also one of the mechanisms by which the body removes MSCs, or that complement assists antibodies directed against MSCs to accelerate MSC removal.