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Macrophage activating factor (MAF)

Figure 1 Schematic illustration of formation of macrophage-activating factor (A) and deglycosylation of Gc protein (B). * lyso-Pc-treated B cells.

contact of Gc protein with the inducible 3-galacto-sidase of B cells and the Neu-1 sialidase of T cells. These glycosidases are present in limiting amounts on the plasma membranes. Native Gc protein must first interact with B cells and, through the action of (3-galactosidase, have a galactose moiety removed. This yields a proactivating factor whose structure allows it to then interact with T cells. The T cell sialidase cleaves off the sialic acid residue, producing a macrophage-activating factor. Thus, the specific oligosaccharide sugars of the native Gc protein and its derivatives would appear to recognize the respective lymphocyte receptors. The affinity studies of the membranous receptors for Gc derivatives revealed that the native Gc protein binds B lymphocytes while the proactivating factor binds T lymphocytes exclusively. Further, the macrophage-activating factor binds macrophage exclusively. However, there is no evidence that these Gc derivatives or macrophage-activating factors, transmit calcitriol or its precursors to lymphocytes and macrophages during these processes.

Enzymatic conversion of vitamin D-binding protein to MAF

In vitro incubation of Gc protein with a mixture of commercial (3-galactosidase and sialidase generates a potent macrophage-activating factor, termed GcMAF. For clinical application of GcMAF, immobilized glycosidases should be used to prevent contamination of enzymes. Stepwise incubation of Gc protein with immobilized 3-galactosidase and sialidase revealed that either sequence of hydrolysis of Gc glycoprotein by these glycosidases yields a remarkably potent GcMAF (Figure 2). A 3 h incubation of mouse peritoneal macrophages or human peripheral blood monocytes/macrophages with 4-10 pg ml-1 of GcMAF results in greatly enhanced (5- to 7-fold) phagocytic capacity and a 15-fold increase in superoxide generation. Administration of 4-10 pg/mouse (10-100 ng/human) of GcMAF also results in a greatly enhanced (5- to 7-fold) phagocytic and 15-fold increased superoxide-generating capacities of macrophages.

Administration to mice of GcMAF (3-50 pg/ mouse) and sheep red blood cells simultaneously produces a large number of antibody-secreting splenic B cells (8 x 104 to 24 xlO4 cells per spleen) in 2-4 days. Thus, the structurally well-defined macrophage-activating factor has a potent adjuvant activity for immunization of mice with sheep erythrocytes.

Gc protein

Macrophage activating factor (MAF)

Gc protein

Macrophage activating factor (MAF)

Gc protein

Gc protein

Figure 2 Two stepwise enzymatic pathways of Gc protein with immobilized glycosidases to yield the macrophage-activating factor (GcMAF). Stepwise hydrolysis of Gc protein by (3-galactosidase and sialidase (A). Stepwise hydrolysis of Gc protein by sialidase and (J-galactosidase (B).

GolNAc

Macrophage activating factor (MAF)

GolNAc

Macrophage activating factor (MAF)

Figure 2 Two stepwise enzymatic pathways of Gc protein with immobilized glycosidases to yield the macrophage-activating factor (GcMAF). Stepwise hydrolysis of Gc protein by (3-galactosidase and sialidase (A). Stepwise hydrolysis of Gc protein by sialidase and (J-galactosidase (B).

Clinical significance of GcMAF

Vitamin D-binding protein, the precursor protein for the macrophage-activating factor, is abundant in the serum because Gc protein circulates in remarkably high concentration (250-350 pg ml-1) in the bloodstream. Only 5% of Gc protein carries 25-hydroxy-vitamin D; which is a reservoir form of vitamin D. Gc protein has a strong affinity for 25-hydroxyvit-amin D, but a weak affinity for calcitriol. This differential affinity of vitamin D metabolites allows Gc protein to transport and transmit the vitamin D metabolites to target cells. However, purified Gc protein, free of 25-hydroxyvitamin D3, is enzymatically converted to GcMAF. Addition of vitamin D metabolites during the above GcMAF generation process has no effect on generation rate or potency of GcMAF.

The membranous glycosidases of inflammation-primed lymphocytes rapidly convert serum Gc protein to the potent macrophage-activating factor. This inflammation-primed macrophage-activation process appears to be the major macrophage-activation cascade which is shared by other phagocytes such as osteoclasts. A defect in the inducible (3-galactosidase of B lymphocytes in the macrophage-activation cascade produces dysfunctional macrophages and osteoclasts which manifest osteopetrosis in newborn. GcMAF can bypass the defective lymphocyte function and act directly on phagocytes. In vitro incubation of human osteoclasts with GcMAF efficiently activates osteoclasts as determined by superoxide generation. GcMAF has a capacity to rectify the osteopetrotic disorder as demonstrated by increased bone resorption and marrow formation when administered to osteopetrotic mice.

Another defect in the macrophage-activating cascade can be caused by deglycosylation of serum Gc protein by a-N-acetylgalactosaminidase secreted into the bloodstream from cancer cells or HIV-infected cells, as shown in Figure IB. Deglycosylated Gc protein can not be converted to macrophage-activating factor, leading to immunosuppression. An impairment of immune function of the host will enhance opportunities for survival and proliferation of neoplastic cells.

See also: Cytokines; Leukemia; Macrophage activation; Monocytes; Neuroendocrine regulation of immunity; Nutrition and the immune system.

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