Intracellular fate of Mycobacterium leprae in normal and activated mouse macrophages

Abstract

Mycobacterium leprae replicates within mononuclear phagocytes, reaching enormous numbers in the macrophage-rich granulomas of lepromatous leprosy. To examine the capability of macrophages to digest M. leprae, we studied the intracellular fate of M. leprae organisms in normal and activated mouse macrophages by using the electron-dense secondary lysosome tracer Thoria Sol. Intracellular M. leprae organisms, surrounded by a characteristic electron-transparent zone, were contained within phagosomal vacuoles of macrophages cultured in vitro for 1 to 6 days. In normal macrophages, a majority of phagosomes containing freshly isolated live M. leprae cells resisted fusion with Thoria Sol-labeled lysosomes. The extent of fusion was not significantly affected by pretreatment of M. leprae with human patient serum high in specific immunoglobulin G and M antibodies. In contrast, a majority of phagosomes containing gamma-irradiated M. leprae cells underwent lysosome fusion in normal macrophages. In addition, increased phagolysosome fusion was observed with live M. leprae-containing phagosomes in macrophages activated with gamma interferon. Increased fusion was associated with an increase in the number of fragmented and damaged bacilli, suggesting that increased digestion followed fusion. This study indicates that activated macrophages may have an increased capacity for clearance of normally resistant M. leprae.

Mycobacterium-containing phagosomes are accessible to early endosomes and reflect a transitional state in normal phagosome biogenesis.

“…It is unlikely, therefore, that pH alone is responsible for the death of intracellular bacilli. Once macrophages are activated, there is a marked increase in the fusion of mycobacterial vacuoles with lysosomes (Sibley et al, 1987;Orme et al, 1993). Activation of murine macrophages by interferon-y and tumor necrosis factor-a is accompanied by the up-regulation of superoxide production, and expression of the inducible nitric oxide synthase (iNOS) (Nathan and Hibbs, 1991).…”

“…Furthermore, the maintenance of this regulated fusion correlates directly with the intracellular survival of the bacilli. In Mycobacterium tuberculosis, M.avium and M.leprae infections, activation of the host macrophage leads to fusion of the bacilli-containing compartments with lysosomes and culminates in death of the bacteria (Sibley et al, 1987;de Chastellier et al, 1993).…”

Axenic incorporation of [U-14C]palmitic acid into the phenolic glycolipid-I of Mycobacterium leprae

“…In order to determine if residual footpad debris was contributing to this activity, the rate of [14C]PA incorporation into PGL-I of a suspension prepared by differential centrifugation (as described above) was compared to that of suspensions treated further by (a) brief suspension in 0.1 N NaOH and (b) exposure to trypsin, collagenase, DNase, and elastase for 1 h at 33°C followed by purification on 2-step discontinuous Percoll gradients as previously described [15]. Alkaline treatment [16][17][18] and Percoll gradient purification [13,15] are both known to be effective in removing contaminating host debris from tissue homogenates of M. leprae. Rates of PGL-I synthesis as well as ATP content of the treated, washed and density-adjusted suspensions were actually higher than the differentially centrifuged preparations (Table 1), suggesting an enrichment for viable bacilli [15] and confirming the bacterial nature of the activity.…”

“…Alkaline treatment [16][17][18] and Percoll gradient purification [13,15] are both known to be effective in removing contaminating host debris from tissue homogenates of M. leprae. Rates of PGL-I synthesis as well as ATP content of the treated, washed and density-adjusted suspensions were actually higher than the differentially centrifuged preparations (Table 1), suggesting an enrichment for viable bacilli [15] and confirming the bacterial nature of the activity. Removal of competing tissue-associated fatty acids may also have contributed to an enhanced rate of []4C]PA incorporation.…”

Down‐Regulatory Effect of Mycobacterium leprae Cell Wall Lipids on Phagocytosis, Oxidative Respiratory Burst and Tumour Cell Killing by Mouse Bone Marrow Derived Macrophages

“…The role of macrophages on the control of mycobacteria infection is still unclear despite the great efforts addressed to this issue [1,[3][4][5][6][7][8][9][10][15][16][17][18][19][20][21][22][23]. Since macrophages are the obligatory homing for pathogen mycobacteria in the tissues, we decided to investigate further the effects of dead M. leprae and its lipidic content on macrophage activation evaluating the phagocytic ability, superoxide production and tumour cell killing ability of these cells.…”