CD43 is required for optimal growth inhibition of Mycobacterium tuberculosis in macrophages and in mice.
ABSTRACT We explored the role of macrophage (Mphi) CD43, a transmembrane glycoprotein, in the pathogenesis of Mycobacterium tuberculosis. Using gene-deleted mice (CD43-/-), we assessed the association of the bacterium with distinct populations of Mphi and found that CD43-/- Mphi bound less M. tuberculosis than CD43+/+ Mphi. Increased infective doses did not abrogate this difference. However, reduced association due to the absence of CD43 could be overcome by serum components. Mphi from heterozygote mice, which express 50% of wild-type CD43, bound more bacteria than CD43-/- but less than CD43+/+, proving that the gene dose of CD43 correlates with binding of M. tuberculosis. Furthermore, the reduced ability of CD43-/- Mphi to bind bacteria was restricted to mycobacterial species. We also found that the survival and replication of M. tuberculosis within Mphi was enhanced significantly in the absence of CD43, making this the first demonstration that the mechanism of mycobacterial entry influences its subsequent growth. Most importantly, we show here that the absence of CD43 in mice aerogenically infected with M. tuberculosis results in an increased bacterial load during both the acute and chronic stages of infection and more rapid development of granulomas, with greater lung involvement and distinctive cellularity.
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ABSTRACT: Silicosis is considered to be among the occupational lung diseases and associated with sandblasting, mining, quarrying and tunneling. Acute silicosis is usually progressive diseaseand despite treatment with corticosteroids it leads to cardio-respiratory failure and death. Alveolar silicoproteinosis is one of it's acute presentations due to exposure to silica dust and lungs filling with proteinaceous material. Here, we have presented a 29 year old male sandblaster with the three conditions of acute silicosis, secondary alveolar proteinosis and pulmonary tuberculosis on four anti tuberculous medications who presented with respiratory distress.Medical journal of the Islamic Republic of Iran 01/2014; 28(1):23.
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ABSTRACT: Establishment of Tuberculosis infection begins with the successful entry and survival of the pathogen within macrophages. We previously showed that macrophage CD43 is required for optimal uptake and growth inhibition of Mycobacterium tuberculosis both in vitro and in vivo. Here, we explore the mechanisms by which CD43 restricts mycobacterial growth in murine macrophages. We found that although M. tuberculosis grows more readily in resting CD43-/- macrophages, priming of cells with IFN-gamma returns the bacterial growth rate to that seen in CD43+/+ cells. To discern the mechanisms by which M. tuberculosis exhibits enhanced growth within resting CD43-/- macrophages, we assessed the induction of inflammatory mediators in response to infection. We found that absence of CD43 resulted in reduced production of TNF-alpha, IL-12 and IL-6 by M. tuberculosis-infected macrophages. We also found that infected resting, but not activated CD43-/- macrophages, showed decreased apoptosis and increased necrosis. Exogenous addition of the pro-inflammatory cytokine TNF-alpha restored control of M. tuberculosis growth and induction of apoptosis to CD43+/+ levels. We propose that CD43 is involved in the inflammatory response to M. tuberculosis and, through the induction of pro-inflammatory mediators, can regulate apoptosis to control intracellular growth of the bacterium.Cellular Microbiology 10/2008; 10(10):2105-17. · 4.81 Impact Factor
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ABSTRACT: The development of effective immunoprophylaxis against tuberculosis (TB) remains a global priority, but is hampered by a partially protective Bacillus Calmette-Guérin (BCG) vaccine and an incomplete understanding of the mechanisms of immunity to Mycobacterium tuberculosis. Although host genetic factors may be a primary reason for BCG's variable and inadequate efficacy, this possibility has not been intensively examined. We hypothesized that Toll-like receptor (TLR) variation is associated with altered in vivo immune responses to BCG. We examined whether functionally defined TLR pathway polymorphisms were associated with T cell cytokine responses in whole blood stimulated ex vivo with BCG 10 weeks after newborn BCG vaccination of South African infants. In the primary analysis, polymorphism TLR6_C745T (P249S) was associated with increased BCG-induced IFN-γ in both discovery (n = 240) and validation (n = 240) cohorts. In secondary analyses of the combined cohort, TLR1_T1805G (I602S) and TLR6_G1083C (synonymous) were associated with increased IFN-γ, TLR6_G1083C and TLR6_C745T were associated with increased IL-2, and TLR1_A1188T was associated with increased IFN-γ and IL-2. For each of these polymorphisms, the hypo-responsive allele, as defined by innate immunity signaling assays, was associated with increased production of TH1-type T cell cytokines (IFN-γ or IL-2). After stimulation with TLR1/6 lipopeptide ligands, PBMCs from TLR1/6-deficient individuals (stratified by TLR1_T1805G and TLR6_C745T hyporesponsive genotypes) secreted lower amounts of IL-6 and IL-10 compared to those with responsive TLR1/6 genotypes. In contrast, no IL-12p70 was secreted by PBMCs or monocytes. These data support a mechanism where TLR1/6 polymorphisms modulate TH1 T-cell polarization through genetic regulation of monocyte IL-10 secretion in the absence of IL-12. These studies provide evidence that functionally defined innate immune gene variants are associated with the development of adaptive immune responses after in vivo vaccination against a bacterial pathogen in humans. These findings could potentially guide novel adjuvant vaccine strategies as well as have implications for IFN-γ-based diagnostic testing for TB.PLoS Pathogens 08/2011; 7(8):e1002174. · 8.14 Impact Factor
CD43 Is Required for Optimal Growth Inhibition of
Mycobacterium tuberculosis in Macrophages and in Mice1
April K. Randhawa,*¶Hermann J. Ziltener,†§Jasmeen S. Merzaban,*§and
Richard W. Stokes2*†‡¶
We explored the role of macrophage (M?) CD43, a transmembrane glycoprotein, in the pathogenesis of Mycobacterium tuber-
culosis. Using gene-deleted mice (CD43?/?), we assessed the association of the bacterium with distinct populations of M? and
found that CD43?/?M? bound less M. tuberculosis than CD43?/?M?. Increased infective doses did not abrogate this difference.
However, reduced association due to the absence of CD43 could be overcome by serum components. M? from heterozygote mice,
which express 50% of wild-type CD43, bound more bacteria than CD43?/?but less than CD43?/?, proving that the gene dose of
CD43 correlates with binding of M. tuberculosis. Furthermore, the reduced ability of CD43?/?M? to bind bacteria was restricted
to mycobacterial species. We also found that the survival and replication of M. tuberculosis within M? was enhanced significantly
in the absence of CD43, making this the first demonstration that the mechanism of mycobacterial entry influences its subsequent
growth. Most importantly, we show here that the absence of CD43 in mice aerogenically infected with M. tuberculosis results in
an increased bacterial load during both the acute and chronic stages of infection and more rapid development of granulomas, with
greater lung involvement and distinctive cellularity. The Journal of Immunology, 2005, 175: 1805–1812.
zation estimates that between 2002 and 2020, 1 billion people will
become infected with M. tuberculosis and ?36 million people will
die of tuberculosis if the rise in incidence is not controlled.
A critical step in the pathogenesis of M. tuberculosis is the ini-
tial interaction of the pathogen with the host macrophage (M?)3.
This interaction is mediated by several M? receptors in association
with ligands on the bacterium, including the complement receptors
CR1, CR3, and CR4, (1–6), Fc?Rs (7), mannose/glucan receptors
(1, 8), CD14 (9, 10), scavenger receptors (4), and surfactant pro-
tein receptors A (11, 12) and D (13). It has also been shown that
CD43 (leukosialin; sialophorin) may be important in promoting a
stable interaction of mycobacteria with M? (14).
CD43 is a negatively charged transmembrane sialoglycoprotein
expressed on most hemopoietic cells (15). The function of this
molecule has been the subject of debate; it has been shown that
ycobacterium tuberculosis infects ?8 million people
and causes ?2 million deaths annually, making it the
deadliest human pathogen. The World Health Organi-
CD43 on T and B cells acts as a barrier molecule restricting cell-
cell contact (16–19) but that it can also have a proadhesive quality
(20–22). Thus, it has been proposed that CD43 may play a dual
role in intercellular contact (23, 24). Involvement of CD43 in leu-
kocyte homing and tissue infiltration, possibly due to its adhesive
or anti-adhesive properties, has been shown in several studies (19,
25, 26). It has also been demonstrated that CD43 can regulate cell
survival (27, 28) and is involved in the apoptosis of T cells and
hemopoietic progenitor cells (29–32).
Fratazzi et al. (14) first described a role for CD43 in mycobac-
terial pathogenesis when they found that splenic M? (SpM?) from
CD43?/?mice could not bind M. tuberculosis or Mycobacterium
avium in vitro but that the ability to bind M. avium could be re-
stored by addition of the extracellular region of CD43. They also
found that CD43-transfected HeLa cells bound M. avium but not
other bacteria and that CD43 was required for TNF-? production
by M? in response to infection with M. avium (14).
In this study, we further explore the role of CD43 in the binding
and uptake of M. tuberculosis by M? to determine the role of
CD43 in M. tuberculosis pathogenesis using a gene-deleted mouse
model that lacks expression of CD43 (33).
Materials and Methods
M. tuberculosis (strain Erdman, TMC no. 107; ATCC no. 35801), M. tu-
berculosis (strain H37Rv, TMC no. 102, ATCC no. 27294), and M. avium
(TMC no. 724, ATCC no. 25291) were grown to late log phase in
Proskauer and Beck medium supplemented with 0.05% Tween 80. Cul-
tures were stored and tested for viability as described previously (2). Sal-
monella enterica serovar Typhimurium (S. typhimurium) and Listeria
monocytogenes were grown to mid-log phase in tryptic soy broth (Difco)
and washed in PBS before use.
Wild-type (WT) control mice (CD43?/?), CD43?/?, and CD43?/?mice
backcrossed seven generations on C57BL/6 background (33) were housed
in a specific pathogen-free animal facility in micro isolator cages. Exper-
iments were done in accordance with the standards set by the Canadian
Council on Animal Care. For all experiments, mice were age- and sex-
matched and controls were littermates.
*Department of Medicine,†Department of Pathology and Laboratory Medicine,‡De-
partment of Paediatrics, and§Biomedical Research Centre, University of British Co-
lumbia, Vancouver, British Columbia, Canada, University of British Columbia; and
¶Division of Infectious and Immunological Diseases, British Columbia’s Children’s
and Women’s Hospital, Vancouver, British Columbia, Canada
Received for publication February 9, 2005. Accepted for publication May 18, 2005.
The costs of publication of this article were defrayed in part by the payment of page
charges. This article must therefore be hereby marked advertisement in accordance
with 18 U.S.C. Section 1734 solely to indicate this fact.
1This work was supported by grants from the British Columbia Lung Association (to
R.W.S.), the Tuberculous and Chest Disabled Veterans’ Association of British Co-
lumbia and the Network Centres of Excellence (Canadian Bacterial Diseases Net-
work) (to R.W.S.), and by Canadian Institutes of Health Research Grant MOP-64267
(to H.J.Z.). R.W.S. is the recipient of a British Columbia Research Institute for Chil-
dren’s and Women’s Health Investigatorship Award.
2Address correspondence and reprint requests to Dr. Richard W. Stokes, Department
of Paediatrics, University of British Columbia, Room 304, British Columbia Research
Institute for Children’s and Women’s Health, 950 West 28th Avenue, Vancouver,
British Columbia V5Z 4H4, Canada. E-mail address: firstname.lastname@example.org
3Abbreviations used in this paper: M?, macrophage; SpM?, splenic M?; WT, wild
type; AM?, alveolar macrophage; PM?, peritoneal macrophage; BMM?, bone mar-
row-derived macrophage; MOI, multiplicity of infection.
The Journal of Immunology
Copyright © 2005 by The American Association of Immunologists, Inc. 0022-1767/05/$02.00
Resident alveolar, peritoneal, and bone marrow-derived M? (AM?, PM?,
and BMM?, respectively) were obtained from CD43?/?and WT mice as
described previously (2, 34, 35). SpM? were obtained by gently disrupting
spleens into single-cell suspensions, washing in PBS, and resuspending
cells in supplemented RPMI 1640 (RPMI 1640 medium with 10% FCS, 10
mM L-glutamine, and 10 mM sodium pyruvate; all from Invitrogen Life
Technologies) at a concentration of 2 ? 106cells/ml. Cells were plated in
1-ml aliquots onto 13-mm acid-washed, sterile glass coverslips in 24-well
plates and incubated at 37°C/5% CO2for 24 h, at which point the nonad-
herent cells were removed, and 1 ml of fresh medium was added to each
well. The cells were incubated for an additional 4 days before use.
Particles for probing macrophage receptors
The function of Fc?Rs was examined using SRBC coated with IgG (EIgG),
complement receptors were investigated using SRBC coated with IgM and
iC3b (EIgMC?), whereas lectin-like receptors were probed using zymosan
particles, all as described previously (2). Latex beads (diameter 1.07 ?m,
Polybead polystyrene microspheres; Polysciences) were used to investigate
AM?, PM?, BMM?, and SpM? from CD43?/?and control mice were
isolated as described above and plated onto sterile petri dishes (bacterio-
logic plastic was used to facilitate subsequent removal of adherent cells).
Following incubation at 37°C/5% CO2, nonadherent cells were removed by
washing with RPMI 1640 medium. Adherent cells were removed by cool-
ing and scraping, washed with DMEM (Invitrogen Life Technologies), and
processed for flow cytometry. Cells were stained with mAb S11-FITC
(rat-anti-pan-CD43; kindly supplied by Dr. J. Kemp, University of Iowa,
Iowa City, Iowa) (36, 37) and/or rat anti-mouse M? F4/80 (Caltag Labo-
ratories) at 2 mg/ml or with secondary Ab alone (streptavidin-Cychrome).
After staining, cells were washed twice with HBSS (Invitrogen Life Tech-
nologies) and analyzed on a FACScan IV flow cytometer (BD
In vitro assay for binding of particles to macrophages
M? monolayers on coverslips in 24-well plates were washed twice with
binding medium (138 mM NaCl, 8.1 mM Na2HPO4, 1.5 mM KH2PO4, 2.7
mM KCl, 0.6 mM CaCl2, 1 mM MgCl2, and 5.5 mM D-glucose) (38). A
500-?l aliquot of binding medium was added to each well, and the cells
were acclimatized for 10 min at 37°C/5% CO2. For nonopsonic studies, the
particles to be tested were diluted to the desired concentration in binding
medium and added to monolayers. In studies of opsonic binding, 1% nor-
mal or 1% heat-inactivated CD43?/?or WT mouse serum was added be-
fore addition of bacteria. For experiments with mycobacteria or particles,
monolayers were infected for 1 h of rocking (Nutator; BD Biosciences)
followed by 2 h stationary at 37°C/5% CO2, whereas in experiments with
S. typhimurium and L. monocytogenes, monolayers were infected for 40
min at 4°C. Monolayers were then washed three times, fixed, and stained
with Kinyoun’s Carbol Fuschin and malachite green for mycobacteria-
infected M? or Giemsa for other bacteria and control particles. Binding
was quantified microscopically by counting 100 M?/coverslip and assess-
ing the percentage of M? that bound at least one bacterium and the average
number of bacteria that were associated with each infected M?.
In vitro survival and replication of M. tuberculosis following
phagocytosis by CD43?/?and CD43?/?M?
Intracellular growth assays were conducted in BMM? from CD43?/?and
WT CD43?/?mice. Monolayers were infected with M. tuberculosis Erd-
man at a multiplicity of infection (MOI) of 20:1 (bacteria:M?). Because
preliminary studies showed that fewer bacteria were able to infect
CD43?/?M?, these were also infected at 30:1. Monolayers were infected
according to the procedures described above. To eliminate extracellular
bacteria from the infected M? monolayer, coverslips were washed three
times in binding medium after infection and transferred to new 24-well
plates containing 1 ml of supplemented RPMI 1640 medium/well. At this
time (day 0) and on days 1, 4, and 7 postinfection, coverslips and super-
natants were processed to assess the CFU, as described previously (39).
Growth and pathogenesis of M. tuberculosis in mice
CD43?/?and WT mice were infected with a low dose of M. tuberculosis
Erdman (50–100 CFU) using an inhalation exposure chamber (Glas-Col).
On specific days postinfection, four to six mice in each group were eutha-
nized, and lungs, livers, and spleens were aseptically removed and
weighed. A portion of each tissue was removed and fixed in 10% buffered
formalin for histopathological examination, while the remainder (lungs and
spleens only) was homogenized in PBS to assess bacterial loads. Serial
dilutions of tissue homogenates were plated on 7H10 agar supplemented
with Oleic Acid Dextrose Complex. Plates were incubated at 37°C and
CFU counted after 21–25 days. Another group of mice was infected i.v.
with M. tuberculosis by injecting 1 ? 105bacteria into the tail vein. Bac-
terial loads were determined as described above.
Livers, spleens, and lungs from mice infected with M. tuberculosis (as
described above) were fixed in 10% buffered formalin, embedded in par-
affin, sectioned at 5 ?m, and stained with H&E for histopathological ex-
amination using standard techniques. The population of cells in areas of
pulmonary inflammation was quantified by classifying cells as either 1)
typical M? (including epithelioid cells), 2) “foamy” M?—cells having
highly vacuolated cytoplasm, which gave it a foamy appearance, or 3)
lymphocytes. The proportion of each cell type in representative areas of
inflammation was determined by counting 100 cells, and the results ex-
pressed to the nearest 5% to allow for a degree of variation between areas.
Data are expressed as mean ? SEM. Student’s t test for independent means
was used; a value of p ? 0.05 was considered significant unless otherwise
CD43?/?M? of different origins bind M. tuberculosis less
readily than CD43?/?M? at various multiplicities of infection
We first determined whether CD43 affected the nonopsonic bind-
ing of M. tuberculosis by using SpM?, PM?, BMM?, or AM?
from CD43?/?and WT CD43?/?mice. SpM?, PM?, and
BMM? were infected at an MOI of 20:1, whereas AM? were
infected at a higher MOI of 500:1 as it has been shown that these
cells do not bind M. tuberculosis very well in vitro (35). After a
3-h infection, we found that SpM? and PM? bound bacteria more
efficiently than did BMM? and AM?, as assessed by both the
percentage of the M? population infected (data not shown) and the
number of bacteria binding to individual infected M? (Fig. 1A).
All CD43?/?M? phenotypes bound significantly less bacteria
than WT CD43?/?M? (p ? 0.01); WT SpM?, PM?, BMM?,
and AM? bound, respectively, 117, 31, 162, and 55% more bacilli
than did CD43?/?-derived M? (Fig. 1A). We chose to use BMM?
as a model in subsequent studies because they can be obtained in
large numbers and represent a recently differentiated M?, such as
those that may be found entering sites of infection during the
course of pathogenesis of M. tuberculosis.
Infection of BMM? with increasing numbers of M. tuberculosis
demonstrated that reduced association of the bacteria with
CD43?/?M? was consistent over a wide range of MOI. At ratios
of 20:1, 40:1, 60:1, 100:1, and 200:1 (Fig. 1B) bacteria:M?,
CD43?/?M? associated with significantly less bacteria than did
CD43?/?M? (p ? 0.05). The reduction in nonopsonic binding of
M. tuberculosis by CD43?/?BMM? is manifested not only as a
reduction in the number of bacteria binding to the M? population
but also a reduction in the percentage of M? binding at least one
bacillus (data not shown).
Opsonization of bacteria overcomes the impaired ability of
CD43?/?M? to bind M. tuberculosis
CD43?/?and CD43?/?BMM? were infected with M. tubercu-
losis in the absence or presence of 1% normal or heat-inactivated
(56°C for 30 min) mouse serum at a MOI of 40:1 (Fig. 2). For
these experiments, only serum autologous to the M? source was
used. In the unopsonized control, 27.8 ? 1.4% of CD43?/?M?
had associated with bacteria while only 9.3 ? 1.3% of CD43?/?
M? were infected (p ? 0.001). In the presence of serum opsonins,
a higher percentage of both M? populations bound bacteria, and
1806 INTERACTION OF M. tuberculosis WITH MACROPHAGE CD43
there was no significant difference between them (51.0 ? 1.6% of
CD43?/?and 47.3 ? 4.8% of CD43?/?M?-bound bacteria).
However, when monolayers were infected with M. tuberculosis in
the presence of heat-inactivated serum, the difference in binding of
bacteria to the two M? populations was restored (32.5 ? 2.5% of
CD43?/?M?; 20.8 ? 1.9% of CD43?/?M? associated with
bacteria, p ? 0.01). Interestingly, the level of binding observed
with CD43?/?M? in the presence of heat-inactivated serum was
higher than that seen in the absence of any serum.
CD43 is involved in binding other mycobacteria, but its absence
does not abrogate binding of S. typhimurium or L.
monocytogenes by BMM?
To determine whether the reduced ability of CD43?/?M? to bind
M. tuberculosis strain Erdman was unique, we infected BMM?
from CD43?/?and WT mice with the virulent strain, H37Rv, with
the opportunistic pathogen M. avium and with representative
Gram-positive (L. monocytogenes) or -negative (S. typhimurium)
intracellular bacteria (Fig. 3). For M. tuberculosis and M. avium,
BMM? were infected at MOIs of 40:1 and 20:1, respectively, for
3 h. For nonmycobacteria, monolayers were infected at a MOI of
10 bacteria/M? for 40 min at 4°C. Less CD43?/?M? were able
to bind the mycobacterial species than could CD43?/?M?. How-
ever, there was no difference between CD43?/?and CD43?/?M?
in their ability to bind the other bacteria.
CD43 deficiency does not affect M?-nonspecific uptake or
phagocytosis via Fc?Rs and complement receptors but does
enhance binding via lectin-like receptors
To investigate whether CD43 deficiency affects the function of
other M? receptors, we measured binding of several different con-
trol particles that are commonly used to study receptor-ligand in-
teractions of phagocytes (Fig. 4). EIgG were used to investigate
the function of Fc?Rs, and EIgMC? were used to examine com-
plement receptors. Zymosan, a yeast cell wall preparation contain-
ing polysaccharides, was used to probe for lectin-like receptors,
while latex beads were used to examine nonspecific interactions.
When assessing the actual number of particles associated with M?,
there was no difference in the ability of the CD43?/?M? to
phagocytose EIgG, EIgMC?, or latex spheres when compared with
the WT controls (Fig. 4). However, CD43?/?M? bound signifi-
cantly more zymosan particles than WT M? (20.4 ? 0.8 vs
14.13 ? 1.1, p ? 0.01, respectively).
The level of CD43 surface expression differs between M?
CD43 cell surface expression was monitored using mAb S11 (37).
The epitope recognized by anti-CD43 mAb S11 is not affected by
changes in CD43 glycosylation (40); thus, mAb S11 cell surface
binding reflects the level of CD43 protein expression. As expected,
all M? populations from CD43?/?mice did not express CD43
above background. Although all cell types from WT mice ex-
pressed low levels of CD43 compared with T cells (41), levels on
BMM? were higher than those on SpM?, PM?, or AM? (Fig. 5).
The mean fluorescence intensity for AM? was 20% of that seen in
species but not other intracellular bacteria. CD43?/?and WT BMM? were
infected with Mycobacterium tuberculosis H37Rv (H37Rv) or M. avium
(M. av) at MOI of 40:1 and 20:1, respectively, for 3 h or with S. typhi-
murium (S. ty) or L. monocytogenes (L. mo) at 10:1 for 40 min. The
mean ? SEM percentage of M? binding at least one bacillus is shown for
three independent experiments, each with three coverslips. ?, p ? 0.01
when compared with control.
CD43 is involved in M? binding of other Mycobacterial
CD43-deficient M?. A, SpM?, PM?, and BMM? from CD43?/?and WT
mice were infected with M. tuberculosis Erdman at 20:1 bacteria:M?,
whereas AM? were infected at 500:1. The average number of bacteria per
infected M? was assessed microscopically for 100 randomly chosen M?.
B, CD43?/?and WT BMM? were infected with M. tuberculosis Erdman
at MOIs of 20:1, 40:1, 60:1, 100:1, and 200:1, and associated bacteria were
quantified as above. A and B, The mean ? SEM from two independent
experiments, each with three coverslips, is shown. All CD43?/?values are
statistically less than WT values (p ? 0.05).
M. tuberculosis has a reduced ability to associate with
of M. tuberculosis to CD43?/?M?. BMM? from CD43?/?and CD43?/?
mice were infected with M. tuberculosis Erdman in the absence (serum-
free) or presence of 1% normal (? serum) or heat-inactivated (? heat-
inactivated (HI)-serum) mouse serum at a MOI of 40:1. The percentage of
M? binding at least one bacillus is shown. Values represent the mean ?
SEM from three independent experiments, each with three coverslips. ??,
p ? 0.001 and ?, p ? 0.01 when compared with WT control.
Heat-labile serum opsonins overcome the reduced binding
1807 The Journal of Immunology
BMM?. In addition, a much lower percentage of the AM? pop-
ulation expressed CD43.
CD43 gene dose correlates to the ability of BMM? to bind M.
To ascertain whether the amount of CD43 expressed by M? affects
their ability to bind M. tuberculosis, we compared the binding of
the bacterium to M? that were heterozygous (?/?) for the CD43
gene (and express 50% less CD43 than do CD43?/?) (33) with
CD43?/?and WT CD43?/?M? (Fig. 6). We found that the
CD43?/?M? population associated less with bacteria than did the
WT M? (19.0 ? 2.5 and 37.3 ? 2.11%, respectively). In contrast,
only 11.8 ? 1.6% CD43?/?M? had associated bacilli (Fig. 6A).
When the actual numbers of bacteria per M? were assessed, the
CD43 gene dose again correlated with the amount of associated
bacteria as WT cells bound 2.07 ? 0.24, whereas CD43?/?bound
1.08 ? 0.24 bacilli/M?, and CD43?/?bound 0.56 ? 0.11 bacil-
li/M? (Fig. 6B).
The survival and replication of M. tuberculosis within CD43?/?
M? is enhanced
BMM? from WT and CD43?/?mice were infected with M. tu-
berculosis, and the subsequent growth of the bacteria was mea-
sured by determining CFU over 7 days. As shown in Table I, at a
MOI of 20:1, CD43?/?M? phagocytosed more bacteria than
CD43?/?M? on day 0 (p ? 0.001), but by day 7, there were
comparable amounts in the two populations. However, when
CD43?/?M? were infected at a MOI of 30:1, the same amount of
bacteria was taken up as in CD43?/?M? at 20:1 on day 0, and by
day 7 postinfection, there were twice as many bacteria in the
CD43?/?population (p ? 0.001). Moreover, the doubling times
of M. tuberculosis in CD43?/?M? were significantly less than in
WT M?, where it took 27.69 ? 0.26 h for one doubling compared
with 24.04 ? 0.18 and 24.12 ? 0.39 h in CD43?/?M? infected
at 20:1 and 30:1, respectively (p ? 0.001).
CD43-deficient mice have a reduced ability to control M.
tuberculosis growth during the acute and chronic phases of
infection following aerosol inhalation of bacteria
To determine the role of CD43 on the in vivo growth and patho-
genesis of M. tuberculosis, we infected CD43?/?and WT mice
aerogenically with a low dose of the bacterium (50–100 bacilli).
There appeared to be no differences in bacterial load in either the
lung or spleen during the first 2 wk of infection (Fig. 7). However,
by day 28, there was a significantly higher bacterial load in both
the lungs and spleens of mice lacking CD43. After this initial peak,
there was a period of host control of growth in the CD43?/?mice,
resulting in the bacterial load being reduced to levels similar to
those in the WT. Subsequently, bacterial loads remained relatively
constant in WT mice, whereas in CD43?/?mice, bacterial loads
steadily increased until the termination of the experiment. There
were no differences in the survival of mice (data not shown). A
comparable experiment following infection over a shorter time pe-
riod (84 days) gave similar results (data not shown).
To determine whether the greater bacterial load in the spleens of
CD43?/?mice during the acute phase of infection (up to day 28)
was due to a greater susceptibility of splenic M? in CD43?/?mice
or to greater seeding of the spleen with bacteria from the lung, we
types. CD43?/?and WT SpM?, PM?, BMM?, and AM? were stained
with mAb S11 (anti-pan-CD43). The mean fluorescence intensity ? SEM
from two independent experiments is shown. Numbers above the bars rep-
resent the percentage of the M? population that expressed CD43. The
dotted line represents background levels of anti-CD43 mAb binding
Surface expression of CD43 varies on different M? pheno-
via complement receptors, Fc?Rs, or nonspecific uptake but enhances up-
take of zymosan. CD43?/?and WT BMM? were incubated with test par-
ticles at the following MOI: EIgG (50:1), EIgMC? (50:1), latex spheres
(25:1), or zymosan (25:1). After 3 h, the association of particles was as-
sessed microscopically. The average number of particles bound per M? is
shown. Results are expressed as the mean ? SEM of three independent
experiments, each with three coverslips. ?, p ? 0.01 when compared with
The absence of CD43 on ?? does not affect phagocytosis
CD43 gene dose. BMM? from WT (?/?), CD43-knockout (?/?), and
CD43-heterozygous (?/?) mice were incubated with M. tuberculosis Er-
dman at a ratio of 40 bacteria:M?. A, The percentage of M? binding at
least one bacillus is shown. B, The average number of bound bacteria per
infected M? is shown. Figures represent the mean ? SEM from two ex-
periments, each with three coverslips. ?, p ? 0.001 when compared with
CD43?/?; †, p ? 0.05 when compared with CD43?/?.
M. tuberculosis binding to BMM? is dependent on the
1808INTERACTION OF M. tuberculosis WITH MACROPHAGE CD43
infected CD43?/?and WT mice with M. tuberculosis via the i.v.
route. This ensured equal numbers of bacteria were deposited into
the spleens and lungs of both mouse strains. No differences were
seen in the growth rate of M. tuberculosis over 6 wk (Fig. 8).
Organ pathology is exacerbated in CD43-deficient mice
Histopathological assessment of organs from M. tuberculosis-in-
fected mice revealed that pathology in CD43?/?mice was more
severe and developed more rapidly than in WT mice. By day 1
postinfection, lymphoid hyperplasia was evident in the spleens of
CD43?/?mice, and by day 56, these mice displayed multifocal
granulomatous inflammation affecting ?50% of the lung (Table
II). This level of pathology did not appear in the WT mice until day
85. By the final experimental time point, CD43?/?mice had gran-
ulomatous inflammation affecting ?50% of the lung sections, se-
vere lymphoid hyperplasia in the spleen, and vascular, perivascu-
lar, and interstitial infiltrates of lymphocytes and neutrophils in the
liver. At various time points, CD43?/?mice also showed an in-
creased number of foamy M? in lung sections. Although granu-
lomas in WT mice also contained some foamy M?, these were
only seen during the chronic stages of infection and decreased in
numbers toward the end of the experiment, whereas in CD43?/?
mice, foamy M? were present from day 56 onward and in greater
numbers (Fig. 9). Overall granuloma formation in CD43?/?mice
occurred more rapidly and more extensively, affected a greater
proportion of the lung, and included more foamy M?.
Although recent studies have shown that the interaction of M. tu-
berculosis with M? does not necessarily result in uptake of the
bacteria, certain M? populations can and do ingest them (2, 35,
38). Understanding how M. tuberculosis enters, survives, and es-
tablishes an infection in these M? populations is crucial to com-
prehending the pathogenesis of mycobacterial infections.
Recently, Fratazzi et al. (14) described a role for CD43 in my-
cobacteria-M? interactions. Their results suggested that CD43
may play a role in promoting a stable interaction of mycobacteria
with receptors on host cells and that this interaction regulated
TNF-? production by the M?. To advance our understanding of
this interaction, we further characterized the role of CD43 in my-
cobacterial infections by analyzing the association of M. tubercu-
losis with different M? phenotypes, by using other bacterial spe-
cies and control particles, by studying the relationship between
CD43 expression and mycobacterial binding, and by monitoring
the growth of M. tuberculosis in M? monolayers and in
We confirm that CD43 is involved in the ability of M? to bind
and engulf mycobacteria because M? from CD43-knockout mice
were less able to phagocytose M. tuberculosis. Our studies extend
this observation and show that this reduction of M. tuberculosis
binding depends upon various factors. Firstly, M? of distinct or-
igins varied in their ability to bind M. tuberculosis and showed
different levels of reduction in binding when CD43 was absent.
during both the acute and chronic phase of infection in mice. CD43?/?(E)
and WT (F) mice were infected aerogenically with a low dose of M. tu-
berculosis. The bacterial loads in the lung (A) and spleen (B) are shown as
the mean CFU/organ ? 104? SEM for four to six mice per experimental
group at each time point. ?, p ? 0.05 and ??, p ? 0.01 when compared with
CD43 is necessary for the control of M. tuberculosis growth
not show impaired control of bacterial growth during the acute phase of
infection. CD43?/?(E) and WT (F) mice were infected i.v. with M. tu-
berculosis. The bacterial loads in the lung (A) and spleen (B) are shown as
the mean CFU/organ ? 103? SEM for five mice per group at each time
point. No significant difference was found between the experimental
CD43-deficient mice infected i.v. with M. tuberculosis do
Table I. Intracellular survival and replication of M. tuberculosis is enhanced in CD43?/?BMM?a
M? Type (MOI)Day 0 Day 1Day 4 Day 7Doubling Time (h)
4.76 ? 104(7.85 ? 102)
2.58 ? 104* (4.42 ? 102)
4.81 ? 104(8.08 ? 102)
4.01 ? 104(9.35 ? 102)
2.31 ? 104* (7.38 ? 102)
3.96 ? 104(9.30 ? 102)
1.63 ? 105(2.85 ? 103)
1.68 ? 105(3.85 ? 103)
2.20 ? 105* (4.25 ? 103)
3.19 ? 106(5.46 ? 104)
3.31 ? 106(4.91 ? 104)
6.13 ? 106* (9.59 ? 104)
aCD43?/?and WT BMM? were incubated with M. tuberculosis Erdman at 20:1 bacteria: M?, and CD43?/?M? were also infected at 30:1. Average CFU/ml and doubling
times are shown for day 0 and days 1, 4, and 7 postinfection. Results are expressed as mean ? SEM for three independent experiments, each with three coverslips, plated in
duplicate at each time point.
1809The Journal of Immunology
Although the previous study of CD43-tuberculosis interactions fo-
cused on SpM? (14), we show here that BMM?, PM?, and AM?
also had an impaired association with M. tuberculosis in the ab-
sence of CD43. Although AM? are the cell that first encounters M.
tuberculosis in vivo, new mononuclear phagocytes arrive at the
site of infection during the course of the disease, where they may
differentiate and encounter bacteria. BMM? are an acceptable
model for these elicited M? and are commonly used in studies of
M. tuberculosis. Therefore, we used these cells as a model for our
Within a single population of M? (BMM?), the level of ex-
pression of CD43 directly correlated with binding of M. tubercu-
losis (Fig. 6). However, flow cytometry of different M? popula-
tions showed that expression of CD43 did not directly correlate
with binding of M. tuberculosis. AM? expressed the lowest level
of CD43 and bound M. tuberculosis poorly. However, BMM?
bound M. tuberculosis at levels lower than did SpM? and PM? yet
expressed the highest levels of CD43 (Fig. 5). It is possible that the
expression of CD43 on the surface of M? may not reflect its func-
tional state, as has been seen with other M? receptors (2). Alter-
natively, CD43 may act in conjunction with other M? receptors to
mediate uptake of M. tuberculosis. Thus, variation in expression of
these other receptors would explain differences in binding capacity
of the various M? populations. This contention is supported by the
observation that binding of mycobacteria to CD43?/?M? can be
restored by the addition of the extracellular portion of CD43 (14).
That CD43 is critical for optimal association of M. tuberculosis
with these secondary receptors is shown by the direct correlation
of binding with CD43 expression within a single M? population.
Additional evidence that CD43 was not the only surface moiety
involved in binding M. tuberculosis was that as the MOI was in-
creased both CD43?/?and WT M? bound higher numbers of
bacteria. However, at no point did CD43?/?BMM? bind the same
number of bacteria as did CD43?/?M?, and it was calculated that
CD43 was accountable for up to 40–50% of M. tuberculosis bind-
ing by M?.
We show that a heat-labile component of serum can overcome
the reduction in binding due to the absence of CD43. It has been
demonstrated previously that the extracellular mucin region of
CD43 is present in, and can be isolated from, plasma (42) and that
this molecule can increase binding of mycobacteria by CD43?/?
M? (14). Therefore, it appears, in agreement with previous find-
ings, that soluble CD43 present in serum may potentiate myco-
bacteria-M? interactions. It is also very likely that complement is
responsible for at least some of the enhanced binding of the bac-
terium in the presence of serum, as it has been implicated in fa-
cilitating uptake of mycobacteria by M? (1, 3, 5, 6), and heat-
inactivated serum lacks the capacity for complement activation
(43). However, other heat-resistant opsonins may contribute to en-
hanced binding. Even in the presence of heat-inactivated serum,
both CD43?/?and WT M? showed increased binding of M. tu-
berculosis compared with nonopsonic binding. This suggests that
some component of serum that is heat stable can also mediate
binding to M?. Thus, our studies show that CD43-mediated bind-
ing of M. tuberculosis depends upon the M? phenotype, the num-
ber of infecting bacteria, the presence of serum opsonins, and the
amount of CD43 expression.
Binding studies with control particles demonstrated that the ab-
sence of CD43 does not affect nonspecific phagocytosis by M? or
the function of complement receptors and Fc?Rs. Interestingly,
CD43?/?M? had an increased affinity for zymosan. This could be
due to the removal of factors impeding the interaction of zymosan
binding, such as the large negative charge of CD43 sialic acid
residues or steric hindrance created by the large size of CD43. The
effect of CD43 on bacterial binding to M? also had a level of
specificity, as representative Gram-negative and -positive bacteria
did not require the presence of CD43 to bind to M?. However,
three strains of mycobacteria all required the presence of CD43 for
(B) mice 210 days postinfection with M. tuberculosis via aerosol exposure, ?200 magnification. A predominantly lymphocytic infiltration is seen in the
WT mouse, whereas numerous foamy macrophages can still be seen surrounding lymphocytes in the granuloma of the CD43?/?mouse. C, Foamy
macrophages are shown at ?600 magnification.
Lung pathology is exacerbated in CD43-deficient mice infected with M. tuberculosis. Representative granuloma from WT (A) and CD43?/?
Table II. Granuloma formation in CD43?/?mice is more severe and has altered morphologya
Time PostinfectionMouse Type
Area of Lung
Infiltrating Cell Types (%)
Day 56 CD43?/?
aAt the indicated times postinfection, H&E-stained sections of lung from CD43?/?and WT mice infected with M. tuberculosis were evaluated for the number and type of
granulomas present (multifocal ? numerous granulomas throughout the lung), the amount of the section affected (%), and the dominant cell types present in the granulomatous
region (%), assessed as described in Materials and Methods. Organ sections are from the same mice for which bacterial loads were assessed in Fig. 7.
1810INTERACTION OF M. tuberculosis WITH MACROPHAGE CD43
optimal binding. This supports the contention that CD43 binds
specifically to a mycobacterial moiety.
The intracellular growth of M. tuberculosis was significantly
enhanced in CD43?/?M?, even though the bacteria are less
readily phagocytosed by the M?. This increased growth rate was
independent of the number of bacteria initially ingested. The
higher rate of growth could be due to the fact that CD43?/?M?
have an impaired ability to initiate TNF-? production (14), which
is known to be involved in controlling intracellular growth of M.
tuberculosis (44–47). Other cytokines have also been shown to be
involved in stimulating the release of the chemokines RANTES
and M? inflammatory protein-1 (48), which could also affect the
intracellular growth of M. tuberculosis. Additionally, CD43?/?
M? could be selectively phagocytosing the more virulent bacteria
within the inoculum, or uptake of the bacterium in the absence of
CD43 may lead to altered phagosome maturation, which is known
to be associated with the survival of intracellular mycobacteria
(49–53). Moreover, the induction of killing mechanisms by M?
may differ in CD43?/?and CD43?/?cells. For example, it has
been demonstrated that CD43 is involved in apoptotic signaling
pathways that would affect the fate of mycobacteria-infected M?
CD43 appears to have a significant role in controlling the
growth of M. tuberculosis in the murine host. When infected via
aerosol, CD43?/?mice had increased bacterial loads in the lung
and spleen during the acute phase of infection up to day 28. This
increased growth of M. tuberculosis in CD43?/?mice may be
attributed to the enhanced growth in CD43?/?M? we demon-
strated in vitro. Alternatively, it may be due to differences in the
type and/or number of cells recruited to sites of infection. In H&E-
stained tissue samples, lymphoid hyperplasia was seen in the
spleens of CD43?/?mice as early as day 1 postinfection, and there
were more granulomas in the lungs by day 28 compared with WT.
The difference in bacterial growth during the acute phase of in-
fection in CD43?/?and CD43?/?mice was more pronounced in
the spleens. Because this effect was not seen in mice infected i.v.
with M. tuberculosis, we can conclude that this is not just because
of enhanced bacterial growth in CD43?/?SpM? but is more likely
due to increased dissemination from the lungs of CD43?/?mice.
Following the development of the adaptive immune response
(around day 28), the mice were able to control the infection for a
period of time. However, following this period the CD43?/?mice
also failed to control bacterial growth during the chronic stage of
infection. It is possible this difference can also be ascribed to the
increased susceptibility of CD43?/?M?. However, the adaptive
immune response was able to reduce the bacterial load in
CD43?/?mice between days 28 and 56. This suggested that
CD43?/?M? were capable of being activated to kill intracellular
M. tuberculosis just as effectively as CD43?/?M? during this
stage of the infection. During the chronic stage of the infection, an
effective immune response must be maintained with the corre-
sponding maintenance of granulomata to contain the bacteria. We
have shown that during this late stage of infection, bacterial growth
is not controlled in CD43?/?mice, and histological findings show
that normal granuloma formation is impaired in these mice and
may account for increased bacterial loads. Other published roles of
CD43, including involvement in T cell activation and differentia-
tion, and the recruitment of lymphocytes to sites of infection (19,
25, 26) could also explain the inadequacies in the immune re-
sponse against M. tuberculosis.
It is critical to note that the ability of CD43?/?mice to control
infection with M. tuberculosis depended upon the route of infec-
tion. Although significant differences were seen between CD43?/?
and WT mice infected aerogenically with M. tuberculosis, there
was no significant difference when mice were infected via an i.v.
injection with the same bacterium. It has been shown previously
that M. tuberculosis may have increased virulence when adminis-
tered aerogenically as opposed to i.v. (54) and that the pathogen-
esis of the organism is affected by route of delivery (55). This
emphasizes the importance of using experimental infection proce-
dures that most closely mimic natural exposure to obtain results
that are most physiologically relevant.
In summary, this study establishes that CD43 is involved in the
binding, uptake, and subsequent growth of M. tuberculosis in mu-
rine M? and in vivo. These results support the theory that CD43
has a dual function in cell-cell interactions (23, 24, 56) and that the
nature of particles interacting with CD43 can dictate its function.
Additional studies are necessary to determine the biology of M.
tuberculosis-CD43 interactions, to identify potential mycobacte-
rial ligands for CD43, and to understand mechanisms of cell re-
cruitment in CD43-knockout mice.
Amanda Rooyakkers for her assistance with in vitro growth assays, and to
Dr. Douglas Carlow for help with the FACS analysis. We also thank
Dr. Brett Finlay for providing Salmonella and Listeria strains, and Dr. P. N.
Nation at the University of Alberta, Edmonton, Alberta, Canada, for his-
toLisa Thorsonfor technicalassistance, to
The authors have no financial conflict of interest.
1. Schlesinger, L., C. Bellinger-Kawahara, N. Payne, and M. Horwitz. 1990. Phago-
cytosis of Mycobacterium tuberculosis is mediated by human monocyte comple-
ment receptors and complement component C3. J. Immunol. 144: 2771–2780.
2. Stokes, R. W., I. D. Haidl, W. A. Jefferies, and D. P. Speert. 1993. Macrophage
phenotype determines the nonopsonic binding of Mycobacterium tuberculosis to
murine macrophages. J. Immunol. 151: 7067–7076.
3. Schlesinger, L. 1993. Macrophage phagocytosis of virulent but not attenuated
strains of Mycobacterium tuberculosis is mediated by mannose receptors in ad-
dition to complement receptors. J. Immunol. 150: 2920–2930.
4. Zimmerli, S., S. Edwards, and J. Ernst. 1996. Selective receptor blockade during
phagocytosis does not alter the survival and growth of Mycobacterium tubercu-
losis in human macrophages. Am. J. Respir. Cell Mol. Biol. 15: 760–770.
5. Melo, M. D., I. R. Catchpole, G. Haggar, and R. W. Stokes. 2000. Utilization of
CD11b knockout mice to characterize the role of complement receptor 3 (CR3,
CD11b/CD18) in the growth of Mycobacterium tuberculosis in macrophages.
Cell. Immunol. 205: 13–23.
6. Velasco-Vela ´zquez, M. A., D. Barrera, A. Gonza ´lez-Arenas, C. Rosales, and
J. Agramonte-Hevia. 2003. Macrophage-Mycobacterium tuberculosis interac-
tions: role of compliment receptor-3. Microb. Pathog. 35: 125–131.
7. Armstrong, J., and P. Hart. 1975. Phagosome-lysosome interactions in cultured
macrophages infected with virulent tubercle bacilli: reversal of the usual nonfu-
sion pattern and observations on bacterial survival. J. Exp. Med. 142: 1–16.
8. Astarie-Dequeker, C., E.-N. N?Diaye, V. Le Cabec, M. G. Rittig, J. Prandi, and
I. Maridonneau-Parini. 1999. The mannose receptor mediates uptake of patho-
genic and nonpathogenic mycobacteria and bypasses bactericidal responses in
human macrophages. Infect. Immun. 67: 469–477.
9. Peterson, P., G. Gekker, S. Hu, W. Sheng, W. Anderson, R. Ulevitch, P. Tobias,
K. Gustafson, T. Molitor, and C. Chao. 1995. CD14 receptor-mediated uptake of
nonopsonized Mycobacterium tuberculosis by human microglia. Infect. Immun.
10. Reiling, N., K. Klug, U. Krallmann-Wenzel, R. Laves, S. Goyert, M. E. Taylor,
T. K. Lindhorst, and S. Ehlers. 2001. Complex encounters at the macrophage-
Mycobacterium interface: studies on the role of the mannose receptor and CD14
in experimental infection models with Mycobacterium avium. Immunobiology
11. Gaynor, C., F. McCormack, D. Voelker, S. McGowan, and L. Schlesinger. 1995.
Pulmonary surfactant protein A mediates enhanced phagocytosis of Mycobacte-
rium tuberculosis by a direct interaction with human macrophages. J. Immunol.
12. Pasula, R., J. F. Downing, J. R. Wright, D. L. Kachel, T. E. Davis, Jr., and
W. J. Martin, II. 1997. Surfactant protein A (SP-A) mediates attachment of My-
cobacterium tuberculosis to murine alveolar macrophages. Am. J. Respir. Cell
Mol. Biol. 17: 209–217.
13. Ferguson, J. S., D. R. Voelker, F. X. McCormack, and L. S. Schlesinger. 1999.
Surfactant protein D binds to Mycobacterium tuberculosis Bacilli and Lipoarabi-
nomannan via carbohydrate-lectin interactions resulting in reduced phagocytosis
of the bacteria by macrophages1. J. Immunol. 163: 312–321.
1811The Journal of Immunology
14. Fratazzi, C., N. Manjunath, R. D. Arbeit, C. Carini, T. A. Gerken, B. Ardman, E.
Remold-O’Donnell, and H. G. Remold. 2000. A macrophage invasion mecha-
nism for mycobacteria implicating the extracellular domain of CD43. J. Exp.
Med. 192: 183–192.
15. Fukuda, M. 1991. Leukosialin, a major O-glycan-containing sialoglycoprotein
defining leukocyte differentiation and malignancy. Glycobiology 1: 347–356.
16. Ardman, B., M. Sikorski, and D. Staunton. 1992. CD43 interferes with T lym-
phocyte adhesion. Proc. Natl. Acad. Sci. USA 89: 5001–5005.
17. Manjunath, N., R. Johnson, D. Staunton, R. Pasqualini, and B. Ardman. 1993.
Targeted disruption of CD43 gene enhances T lymphocyte adhesion. J. Immunol.
18. Manjunath, N., M. Correa, M. Ardman, and B. Ardman. 1995. Negative regula-
tion of T cell adhesion and activation by CD43. Nature 377: 535–538.
19. Stockton, B., G. Cheng, N. Manjunath, B. Ardman, and U. von Andrian. 1998.
Negative regulation of T cell homing by CD43. Immunity 8: 373–381.
20. Sanchez-Mateos, P., M. Campanero, M. del Pozo, and F. Sanchez-Madrid. 1995.
Regulatory role of CD43 leukosialin on integrin-mediated T cell adhesion to
endothelial and extracellular matrix ligands and its polar redistribution to a cel-
lular uropod. Blood 86: 2228–2239.
21. Stockl, J., O. Majdic, P. Kohl, W. Pickl, J. Menzel, and W. Knapp. 1996. Leu-
kosialin (CD43)-major histocompatibility class I molecule interactions involved
in spontaneous T cell conjugate formation. J. Exp. Med. 184: 1769–1779.
22. Savage, N. D. L., S. L. Kimzey, S. K. Bromley, K. G. Johnson, M. L. Dustin, and
J. M. Green. 2002. Polar redistribution of the sialoglycoprotein CD43: implica-
tions for T cell function. J. Immunol. 168: 3740–3746.
23. Ostberg, J. R., R. K. Barth, and J. G. Frelinger. 1998. The Roman god Janus: a
paradigm for the function of CD43. Immunol. Today 19: 546–550.
24. van den Berg, T. K., D. Nath, H. J. Ziltener, D. Vestweber, M. Fukuda, I. van Die,
and P. R. Crocker. 2001. Cutting edge: CD43 functions as a T cell counterre-
ceptor for the macrophage adhesion receptor sialoadhesin (Siglec-1). J. Immunol.
25. McEvoy, L. M., H. Sun, J. G. Frelinger, and E. C. Butcher. 1997. Anti-CD43
inhibition of T cell homing. J. Exp. Med. 185: 1493–1498.
26. Woodman, R. C., B. Johnston, M. J. Hickey, D. Teoh, P. Reinhardt, B. Y. Poon,
and P. Kubes. 1998. The functional paradox of CD43 in leukocyte recruitment:
a study using CD43-deficient mice. J. Exp. Med. 188: 2181–2186.
27. Dragone, L., R. Barth, K. Sitar, G. Disbrow, and J. Frelinger. 1995. Disregulation
of leukosialin (CD43, Ly48, sialophorin) expression in the B cell lineage of
transgenic mice increases splenic B cell number and survival. Proc. Natl. Acad.
Sci. USA 92: 626–630.
28. Ostberg, J., L. Dragone, T. Driskell, J. Moynihan, R. Phipps, R. Barth, and
J. Frelinger. 1996. Disregulated expression of CD43 (leukosialin, sialophorin) in
the B cell lineage leads to immunodeficiency. J. Immunol. 157: 4876–4884.
29. Bazil, V., J. Brandt, A. Tsukamoto, and R. Hoffman. 1995. Apoptosis of human
hematopoietic progenitor cells induced by cross-linking of surface CD43, the
major sialoglycoprotein of leukocytes. Blood 86: 502–511.
30. Bazil, V., J. Brandt, S. Chen, M. Roeding, K. Luens, A. Tsukamoto, and
R. Hoffman. 1996. A monoclonal antibody recognizing CD43 (leukosialin) ini-
tiates apoptosis of human hematopoietic progenitor cells but not stem cells. Blood
31. Brown, T. J., W. W. Shuford, W.-C. Wang, S. G. Nadler, T. S. Bailey,
H. Marquardt, and R. S. Mittler. 1996. Characterization of a CD43/leukosialin-
mediated pathway for inducing apoptosis in human T lymphoblastoid cells.
J. Biol. Chem. 271: 27686–27695.
32. Onami, T. M., L. E. Harrington, M. A. Williams, M. Galvan, C. P. Larsen,
T. C. Pearson, N. Manjunath, L. G. Baum, B. D. Pearce, and R. Ahmed. 2002.
Dynamic regulation of T cell immunity by CD43. J. Immunol. 168: 6022–6031.
33. Carlow, D. A., S. Y. Corbel, and H. J. Ziltener. 2001. Absence of CD43 fails to
alter T cell development and responsiveness. J. Immunol. 166: 256–261.
34. Furney, S., P. Skinner, A. Roberts, R. Appelberg, and I. Orme. 1992. Capacity of
Mycobacterium avium isolates to grow well or poorly in murine macrophages
resides in their ability to induce secretion of tumor necrosis factor. Infect. Immun.
35. Stokes, R. W., L. M. Thorson, and D. P. Speert. 1998. Nonopsonic and opsonic
association of Mycobacterium tuberculosis with resident alveolar macrophages is
inefficient. J. Immunol. 160: 5514–5521.
36. Gulley, M., L. Ogata, J. Thorson, M. Dailey, and J. Kemp. 1988. Identification of
a murine pan-T cell antigen which is also expressed during the terminal phases
of B cell differentiation. J. Immunol. 140: 3751–3757.
37. Baecher-Allan, C., J. Kemp, K. Dorfman, R. K. Barth, and J. G. Frelinger. 1993.
Differential epitope expression of Ly-48 (mouse leukosialin). Immunogenetics
38. Smith, R., and S. Iden. 1981. Properties of calcium ionophore-induced generation
of superoxide anion by human neutrophils. Inflammation 5: 177–192.
39. Stokes, R. W., and D. Doxsee. 1999. The receptor-mediated uptake, survival,
replication, and drug sensitivity of Mycobacterium tuberculosis within the mac-
rophage-like cell line THP-1: a comparison with human monocyte-derived mac-
rophages. Cell. Immunol. 197: 1–9.
40. Merzaban, J. S., J. Zuccolo, S. Y. Corbel, M. J. Williams, and H. J. Ziltener.
2005. An alternate core 2 ?1,6-N-acetylglucosaminyltransferase selectively con-
tributes to P-selectin ligand formation in activated CD8 T cells. J. Immunol. 174:
41. Jones, A., B. Federsppiel, L. Ellies, M. Williams, R. Burgener, V. Duronio,
C. Smith, F. Takei, and H. Ziltener. 1994. Characterization of the activation-
associated isoform of CD43 on murine T lymphocytes. J. Immunol. 153:
42. Schmid, K., S. Mao, A. Kimura, S. Hayashi, and J. Binette. 1980. Isolation and
characterization of a serine-threonine-rich galactoglycoprotein from normal hu-
man plasma. J. Biol. Chem. 255: 3221–3226.
43. Guckian, J., G. Christensen, J. Schweinle, and D. Fine. 1981. Opsonization of
pneumococci. I. Heat-labile serum activity other than complement is required for
killing by human polymorphonuclear leukocytes. J. Immunol. 127: 1659–1665.
44. Hirsch, C., J. Ellner, D. Russell, and E. Rich. 1994. Complement receptor-me-
diated uptake and tumor necrosis factor ?-mediated growth inhibition of Myco-
bacterium tuberculosis by human alveolar macrophages. J. Immunol. 152:
45. Aung, H., Z. Toossi, J. J. Wisnieski, R. S. Wallis, L. A. Culp, M. Phillips,
L. E. Averill, T. M. Daniel, and J. J. Ellner. 1996. Induction of monocyte ex-
pression of tumor necrosis factor ? by the 30-kDa antigen of Mycobacterium
tuberculosis and synergism with fibronectin. J. Clin. Invest. 98: 1261–1268.
46. Byrd, T. F. 1997. Tumor necrosis factor ? (TNF-?) promotes growth of virulent
Mycobacterium tuberculosis in human monocytes. J. Clin. Invest. 99:
47. Keane, J., B. Shurtleff, and H. Kornfeld. 2002. TNF-dependent BALB/c murine
macrophage apoptosis following Mycobacterium tuberculosis infection inhibits
bacillary growth in an IFN-? independent manner. Tuberculosis 82: 55–61.
48. Nieto, M., J. L. Rodrı ´guez-Ferna ´ndez, F. Navarro, D. Sancho, J. M. Frade,
M. Mellado, C. Martı ´nez-A, C. Caban ˜as, and F. Sa ´nchez-Madrid. 1999. Signaling
through CD43 induces natural killer cell activation, chemokine release, and
PYK-2 activation. Blood 94: 2767–2777.
49. Deretic, V., and R. A. Fratti. 1999. Mycobacterium tuberculosis phagosome. Mol.
Microbiol. 31: 1603–1609.
50. Teitelbaum, R., M. L. Maitland, N. E. Freitag, J. Condeelis, and B. R. Bloom.
1999. Mycobacterial infection of macrophages results in membrane-permeable
phagosomes. Proc. Natl. Acad. Sci. USA 96: 15190–15195.
51. Fratti, R. A., J. Chua, I. Vergne, and V. Deretic. 2003. Mycobacterium tubercu-
losis glycosylated phosphatidylinositol causes phagosome maturation arrest.
Proc. Natl. Acad. Sci. USA 100: 5437–5442.
52. Pieters, J., and J. Gatfield. 2002. Hijacking the host: survival of pathogenic my-
cobacteria inside macrophages. Trends Microbiol. 10: 142–146.
53. Clemens, D. L., B.-Y. Lee, and M. A. Horwitz. 2002. The Mycobacterium tu-
berculosis phagosome in human macrophages is isolated from the host cell cy-
toplasm. Infect. Immun. 70: 5800–5807.
54. North, R. J. 1995. Mycobacterium tuberculosis is strikingly more virulent for
mice when given via the respiratory than via the intravenous route. J. Infect. Dis.
55. McMurray, D. N. 2003. Hematogenous reseeding of the lung in low-dose, aero-
sol-infected guinea pigs: unique features of the host-pathogen interface in sec-
ondary tuberculosis. Tuberculosis 83: 131–134.
56. Rosenstein, Y., A. Santana, and G. Pedraza-Alva. 1999. CD43, a molecule with
multiple functions. Immunol. Res. 20: 89–99.
1812 INTERACTION OF M. tuberculosis WITH MACROPHAGE CD43
April Kaur Randhawa