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Production and Lipopolysaccharide-Induced Endotoxic Shock1
,§,
,§
*
Department of Microbiology and Immunology and the University of North Carolina Neuroscience Center;
Program for Molecular Biology and Biotechnology;
Department of Pharmacology and Lineberger Comprehensive Cancer Center; and
§
Department of Medicine, University of North Carolina, Chapel Hill, NC 27599
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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production during bacterial sepsis are critical in attenuating adverse
host responses to endotoxemia. To study the function of a novel
receptor tyrosine kinase, mer, that is expressed in
monocytes, we generated mice (merkd) that
lack the signaling tyrosine kinase domain. Upon LPS challenge,
merkd animals died of endotoxic shock
(15/17, 88.2%), whereas control wild-type mice survived (1/15, 6.7%
died). Susceptible merkd mice exhibited
edema, leukocyte infiltration, and signs of endotoxic shock that
correlated with higher levels of TNF- found in the serum of
merkd mice as compared with wild-type
control animals. Death due to LPS-induced endotoxic shock in
merkd mice was blocked by administration of
anti-TNF- Ab, suggesting that overproduction of this cytokine
was principally responsible for the heightened suseptibility. The
increase in TNF- production appeared to be the result of a
substantial increase in the LPS-dependent activation of NF-
B nuclear
translocation resulting in greater TNF- production by macrophages
from merkd mice. Thus, Mer receptor tyrosine
kinase signaling participates in a novel inhibitory pathway in
macrophages important for regulating TNF- secretion and attenuating
endotoxic shock. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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One major immune reaction mediated by monocytes is the response to
bacterial Ags during inflammation or sepsis 9 . Bacterial endotoxin is
one of the most potent initiators of the inflammatory response that
results in >100,000 deaths annually, ranking it 13th among all
causes of deaths in the United States 10, 11, 12 . LPS endotoxin is a
component of Gram-negative bacteria that activates monocytes and
macrophages by binding the glycosylphosphatidylinositol (GPI)-linked
glycoprotein, CD14 9, 13 . Subsequent to surface binding, LPS results
in the induction of NF-B and the initiation of proinflammatory
cascade of cytokines, including TNF-, IL-1, and IL-6 9, 14 in
macrophages. When unabated, the host response to sepsis causes cellular
damage and lethal tissue injury characteristic of endotoxic shock
syndrome 15, 16 . Reduction of NF-B in vivo by i.v. somatic gene
transfer with IB before lethal LPS challenge increased survival
of mice 17 . Thus, mechanisms that inhibit LPS induction of NF-B
presumably would attenuate host immune responses to sepsis.
To further investigate the physiological function of Mer, we generated
gene-targeted mice (merkd) in which the
cytoplasmic kinase domain was replaced with the neomycin-resistant
gene. The mice were without developmental or growth-related anomalies.
However, investigation of monocyte function showed that the
merkd mice were extremely sensitive to endotoxin
(LPS) treatment. These mice exhibited excessive TNF- production and
increased susceptibility to lethal septic shock. LPS stimulation of
macrophages elevated NF-B levels in macrophages from
merkd mice, suggesting that Mer may normally act
to inhibit the LPS pathway. These results delineate a novel inhibitory
pathway via a cell surface tyrosine kinase receptor, Mer, which appears
to be critical in regulating immune responses to LPS and endotoxic
shock.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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A mer cDNA probe was created by using RT-PCR
amplification from mouse fetal liver with the specific oligonucleotide
primers 5'-GTGGCAGTGAAGACCATGAAGTTG-3' and
5'-GAACTCCGGGATAGGGAGTCAT-3'. The resulting 574-bp murine
mer cDNA fragment was used to screen a 129/Sv mouse genomic
library to isolate genomic fragments (Stratagene, La Jolla, CA). The
tyrosine kinase-encoding region of the mer gene was replaced
with the neomycin resistance gene from the pJNS2 plasmid 18 (Fig. 1
A). The herpes simplex
virus-thymidine kinase gene from the targeting vector was
positioned 3' of the homologous arms to select against random
recombination events. Gene targeting into the E14TG2a embryonic
stem cells 19 was performed as previously described 20 . The
resulting chimeras were bred with (C57BL/6 x DBA/2)F1
mice, and heterozygous merkd mice were
intercrossed to produce mice homozygous for the mer gene
mutation or wild-type mer.
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In vivo, LPS challenge, and treatment with anti-TNF- Ab
The titration experiments used LPS (Escherichia coli O55:B5; Sigma, St. Louis, MO.) doses ranging from 25 mg/kg to 200 mg/kg delivered to each mouse by i.p. injection. merkd and wild-type mer animals (6–8 wk) were given LPS at 100 mg/kg i.p. and monitored for morbidity over the indicated time course. Control mer animals were given equivalent volumes of sterile PBS.
For in vivo neutralization of TNF-, 6-wk-old
merkd animals were given i.p. either 250 µg of
hamster anti-TNF- Ab (Genzyme, Cambridge, MA) or irrelevant
isotype-matched hamster Ab, L2-3D9 (kindly provided by Dr. Robert
Schreiber at Washington University School of Medicine, St. Louis, MO),
6 h before lethal challenge with LPS (100 mg/kg). Control
merkd animals were given LPS alone i.p., and
additional controls were given sterile PBS.
Tissues for histological analysis were removed from euthanized animals and placed in 10% buffered formalin. Samples were paraffin embedded, 5 µm serially sectioned, and stained with hematoxylin and eosin for microscopic observation.
Analysis of TNF- production
For in vivo serum TNF- concentrations, naive wild-type or
merkd 6- to 8-wk-old animals were challenged
with 100 mg/kg of LPS, and, at the indicated time points, serum samples
were obtained by tail-vein bleed. TNF- concentrations were
quantified using anti-TNF- Ab in a sandwich ELISA technique
(PharMingen, San Diego, CA). Recombinant TNF- as a standard control
and Ab to TNF- was purchased from Genzyme (Cambridge, MA).
Statistical analyses were performed using standard Student’s
t test with unequal variance.
TNF- secretion by cultured macrophages was assessed by ELISA. Mice
(6–8 wk) were given 3 ml of aged 3% thioglycolate (Difco, Detroit,
MI) i.p. and PECs recovered by sterile PBS lavage on the third day. A
total of 
2 x 105 cells/well were cultured in
96-well plates in standard RPMI 1640 medium with 1% prescreened
FCS. Following a 72-h adjustment period, the PECs were given 100 ng/ml
LPS (E. coli 055:B5; Sigma) in culture medium. Supernatants
were harvested at the indicated time points, and TNF- concentrations
were quantified by ELISA.
Northern blot analysis for TNF- transcripts
All RNA samples were extracted by standard methodologies from
PECs following 30 min of in vivo treatment i.p. with either 10 µg of
LPS or an equivalent volume of PBS as a control. Equivalent amounts of
15 µg of total RNA from each sample were analyzed as indicated by the
ribosomal band. For calculations, background cpm were subtracted from
TNF- and ribosomal bands. The count for each of the TNF-
transcripts was divided by the ribosomal counts to provide relative
amounts of transcripts. This number for TNF- corrected to ribosomal
levels in the LPS-treated macrophages from wild-type mice was divided
by the number from the untreated wild-type group to give the increase
of TNF- transcripts in response to LPS. Similar calculations were
done for counts from macrophages of merkd mice.
The overall fold induction denotes the magnitude of the TNF-
induction from merkd over the TNF- induction
observed in wild-type.
Electromobility shift assay (EMSA)
Thioglycolate-elicited PECs were isolated after LPS stimulation
(10 µg), and nuclear extract samples were prepared from 1.0–3.0
x 106 cells as previously described 22 , except omitting
dialysis of samples. EMSA were performed by incubating 3 µg of
extract with end-labeled B3 site of the 5' murine TNF- promoter
element (5'-AGCTCAAACAGGGGGCTTTCCCTCCTC-3') in
binding buffer (250 mM NaCl, 50 mM Tris (pH 7.6), 50% glycerol, 5 mM
DTT, 2.5 mM EDTA, and 2 µg poly(dI:dC) for 20 min at room
temperature. The underlined sequence is the consensus B binding
element. Supershifts were performed by incubating extracts with Ab to
p50 or p65 (Santa Cruz Biotechnology, Santa Cruz, CA) for 10 min before
addition of probe.
Statistical analysis
Significance for survival time plots between two groups were
calculated using a logrank analysis consulting the 
square
distribution with 1 df. Other statistical analyses utilized the
Student’s t test with unequal variance.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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merkd mice were generated from embryonic
stem cell lines in which the neomycin resistance gene replaced
the last exon encoding the 3' end of the Mer kinase domain by
homologous recombination (Fig. 1
A). The mutation involved
the loss of 53 residues in the activation-loop domain of the Mer kinase
region, including the recently described autophosphorylation sites on
the cytoplasmic tail of the Mer receptor, tyrosine residues 749, 753,
and 754 23 . This recombination introduced a novel EcoRI
site into the locus, allowing detection of the mutated
merkd allele as a 6-kb fragment by Southern blot
analysis from the wild-type allele, which is a 10-kb fragment (Fig. 1B). The expression of a mutated mer was verified
by RT-PCR of mRNA derived from macrophages contained in PECs isolated
from merkd animals (Fig. 1C,
lane 7). The extracellular fibronectin domain (FNIII) of
mer was expressed in both PECs from wild-type and
merkd mice (lanes 4 and5); however, only wild-type mice expressed the cytoplasmic kinase
domain (lane 6), and not the macrophages, from
merkd mice (lane 7).
The specific mutation in mer permitted functional analyses in signal transduction and avoided possible developmental anomalies that may have arisen from complete ablation of the gene. Although mer mRNA has been detected as early as the 8-cell embryo stage 8 , merkd mice exhibited no gross developmental anomalies. In addition, while mer mRNA was expressed at high levels in testis and ovaries, and mer-like sequences have been implicated in sperm function 24 , homozygous merkd mice were produced at the expected Mendelian frequency. In addition, mer mRNA expression was found in a number of tissues, including kidney, liver, brain, spleen, and bone marrow 8 ; however, organs of adult merkd animals were morphologically normal with the exception of enlarged spleens.
Increased sensitivity of merkd mice to LPS/endotoxin
We initially tested responses of macrophages from
merkd mice to LPS. Our observations, using ELISA
assay to detect TNF-, showed that in vitro LPS stimulation of PEC
resulted in significant heightened secretion of TNF- into the
supernatants, compared with macrophages from wild-type mice (data not
shown). To determine whether merkd animals
responded differently to bacterial endotoxin, we assessed in vivo
LPS-toxicity in wild-type control and merkd
mice. In a dose-response study of 25–200 mg/kg of LPS administered
i.p. into mice, it was determined that the LD100 for
merkd animals (100 mg/kg for
merkd animals) was half that of control mice
(200 mg/kg for wild-type animals; data not shown). At the 100-mg/kg
dose of LPS, the merkd animals rapidly succumbed
to septic shock with 52.9% alive by 24 h (9/17), and only 11.8%
(2/17) of the mice survived to 96 h (Fig. 2A). Thus, 88.2% of the
merkd mice (15/17) died from endotoxic shock
induced by LPS. In contrast, more than 93.3% (14/15) of the wild-type
control mice survived this dose of LPS. The increased sensitivity of
merkd mice to endotoxic shock
(p < 0.0001) suggests that Mer may
attenuate responses to LPS.
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production correlated with the heightened
morbidity in the merkd mice. Serum TNF- was
quantitated before and during LPS challenge (at 100-mg/kg dose) of
merkd mice and wild-type animals. Both
merkd and wild-type animals had a rapid
elevation in TNF- 1 h post LPS administration (Fig. 2B), consistent with published observations. However, the
merkd animals responded by secreting nearly
3-fold higher TNF- serum levels than wild-type mice within the
critical first hour (1882.8 pg/ml vs 610.5 pg/ml, p <
0.04) (Fig. 2B). This suggests that greater susceptibility
to endotoxic shock in merkd mice may be due to
excessive production of TNF-.
Evidence of TNF--mediated endotoxic shock was observed in the small
intestine by both gross morphology (bowel dilatation) and
histopathology. Compared with wild-type controls (Fig. 3, A and C), the
LPS-treated merkd mice exhibited increased
lesions and hemorrhagic necrosis in the bowel (Fig. 3, B and
D), indicated by greater fluid accumulation, and an
increased infiltration of red cells (arrowheads) and leukocytes (L)
into the villi (Fig. 3D). The morphology of the intestinal
villi was degenerated in the septic bowel of
merkd mice (Fig. 3D) and showed
numerous involutions and loss in the brush border of the villi
(arrows). In contrast, the small intestine from LPS-treated wild-type
animals demonstrated the maintenance of the smooth brush border and
structure of the villi (Fig. 3C), with only a slight
increase in leukocyte cells (no RBCs apparent in villi). This pattern
of morphological changes was typical of TNF--mediated destruction of
tissue 25 and was consistently greater in
merkd mice suggesting Mer functions to modulate
TNF- production in vivo.
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To confirm the role of TNF- in the endotoxic death of
merkd animals, we protected
merkd mice from a lethal dose of LPS by
pretreatment of merkd animals with an
anti-TNF- mAb. A total of 75% of the anti-TNF-
Ab-treated merkd mice recovered from this LPS
challenge compared with 13% of the merkd mice
(p < 0.005) given PBS only and none of the
merkd mice pretreated with an irrelevant
isotype-matched control Ab (Fig. 4). The
75% survival rate of anti-TNF- Ab-treated
merkd mice was also similar to survival values
(80% survival) previously published for protecting mice from endotoxic
shock with this Ab 26 . This observation suggested that overproduction
of TNF- was the key mediator of the observed sepsis and pathology in
the merkd mice, and merkd
mice did not have an alternative mechanism for the induction for
endotoxic shock.
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secretion from macrophages of
merkd mice
The increased level of TNF- in serum of merkd mice
(Fig. 2B) suggested that LPS could be altering TNF-
expression in macrophages. Indeed, macrophages from
merkd mice that were challenged with LPS in
vitro showed increased secretion of TNF- into supernatants when
compared with macrophages from wild-type mice (Fig. 5A). TNF- levels from
macrophages of merkd mice and wild-type mice
increased over a 9-h period before subsiding. Most importantly, TNF-
levels from macrophages of merkd mice were twice
that of the TNF- production from macrophages of wild-type mice. In
addition, TNF- transcripts from LPS-stimulated macrophages from
merkd mice were elevated compared with mRNA
levels from macrophages of wild-type mice. We isolated RNA from
macrophages of wild-type and merkd mice during
LPS challenge. Fig. 5B is a representative Northern blot
showing increased TNF- mRNA (6-fold) from macrophages of
merkd animals stimulated with LPS over untreated
macrophages. In contrast, macrophages from wild-type mice showed only a
2-fold increase in TNF- transcripts compared with unstimulated
macrophages. The 6-fold induction of TNF- transcripts from
macrophages of merkd mice is three times greater
than the 2-fold induction of TNF- transcripts from macrophages of
wild-type mice. This observation was consistent with the elevated
TNF- serum protein levels observed in vivo from
merkd samples (Fig. 2B) and
production of protein in vitro (Fig. 5A). These data suggest
that macrophages that lack a functional Mer have an increase in TNF-
expression.
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secretion and TNF- mRNA transcripts indicate
that Mer may be altering the transcriptional regulation of the TNF-
gene. Since the 5' promoter region of TNF- contains three
cis-acting NF-B sites 27, 28 , the increased sensitivity
to endotoxic shock and the overproduction of TNF- mRNA and
protein may be due to heightened levels of the transcriptional
activator, NF-B 29, 30 . We assessed the activation of NF-B in
vivo using an oligonucleotide probe of the B3 site in the 5'
promoter of the murine TNF- gene. Elevated (3.5-fold at 15 min) and
sustained NF-B activity was observed in the nuclear extracts of
macrophages from LPS-treated merkd mice (Fig. 6, band 3, lanes 10,and 13–15) at levels greatly exceeding those observed
in macrophages from wild-type mice (lanes 2, and
5–7). In LPS-stimulated macrophages from wild-type mice,
NF-B was barely detectable at 30 min and was absent at 45 and 60 min
post LPS stimulation. In contrast, macrophages from
merkd mice showed sustained NF-B induction
beyond 30 min. Ab supershifts indicated that the p65 and p50 subunits
were contained in the active NF-B heterodimeric complex (arrow
3, lanes 3, 4, 11, and 12). The p50/p50
homodimeric complex (arrow 4) appeared to fluctuate in
binding activity; however, this was not apparent for the p65/p50
NF-B (arrow 3) activity during the time course
(lanes 10, 13, 14, and 15). Furthermore,
unstimulated samples from macrophages of merkd
mice showed a low basal level of nuclear NF-B, while
wild-type samples did not. This may indicate that macrophages from
merkd mice may have a consistent low-level
translocation of NF-B. These observations strongly suggest that
mer influences LPS-dependent TNF- production in
macrophages by actively regulating NF-B activity.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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31 . Our report suggests an additional novel
pathway regulating TNF- and host immune response to endotoxic shock.
Loss of Mer signaling abrogates this check on TNF- secretion in
vivo. We suggest that merkd mice challenged with
a sublethal dose of LPS succumb to endotoxic shock due to
TNF--mediated tissue injury. First, excessive production of TNF-
in vivo (Fig. 2B) was corroborated in vitro using
LPS-stimulated macrophages from merkd and
wild-type mice (Fig. 5A). The macrophages from
merkd mice showed an average 2-fold greater
amount of TNF- secretion compared with PECs from wild-type animals
(Fig. 5A). This increase in TNF- in vivo and in vitro is
a consequence of an absent Mer signal. Second, the anti-TNF- Ab
attenuated death of LPS-challenged merkd mice,
again indicating that the hyperresponsiveness of
merkd animals to LPS is mediated by TNF-. In
addition, another cytokine, IL-1, is also regulated similarly by Mer as
it is hypersecreted in LPS-challenged merkd mice
(data not shown). This suggests that additional inflammatory cytokine
production may be modulated by mer signal transduction. The
mechanism for elevated TNF- production includes increased TNF-
mRNA as presumably due to an increased translocation of NF-B to the
nucleus. Taken together, these data suggest that the Mer receptor on
macrophages serves to attenuate cytokine responses to bacterial
endotoxin and may be a critical inhibitory pathway to guard against
excessive tissue damage in endotoxic shock.
The signal transduction pathway beyond the activation of the Mer
receptor tyrosine kinase is unknown; however, we showed greater p65/p50
NF-B-binding complex (Fig. 6) that correlated closely with the
greater production of TNF- transcripts (Fig. 5B). Our
data suggest that mer functions to modulate the
LPS-dependent TNF- cytokine production by attenuating LPS-dependent
NF-B activation. A detailed analysis of points for interaction
between Mer and LPS signaling will be needed to determine whether the
Mer signal attenuates all activities stimulated by LPS, or whether
putative inhibition of NF-B nuclear translocation is selective.
However, it is clear that a lack of a Mer inhibitory signal in
merkd mice results in elevated and prolonged
NF-B activation, causing excess macrophage activation and TNF-
production.
Receptor tyrosine kinases are often discussed in the context of growth
and/or differentiation, yet deletion of the kinase-signaling capacity
of Mer does not result in attenuation of growth or development in
merkd animals. This report is the first to
demonstrate that the Mer receptor tyrosine kinase is involved in
modulating or dampening cell activation in the response to LPS. Our
results indicated that merkd mice are
hyperresponsive to LPS and routinely died when challenged with doses of
LPS below the lethal dose for control animals (Fig. 2A). It
has been speculated previously that tyrosine kinase activity is
important in the LPS-signal transduction process 32, 33 ; however,
Meng and Lowell 34 have shown that the Src-like kinases (Fgr, Hck,
and Lyn) are not the obligatory kinase components of the LPS signal
cascade. In fact, lack of Hck and Fgr impaired inflammation due to an
inability of neutrophils to migrate and damage tissue 35 . It appears
that Mer is not utilizing such nonreceptor tyrosine kinases in signal
transduction. This suggests that additional intracellular signaling
mechanisms or receptors exist to regulate the LPS response in
monocytes. Furthermore, recent reports have demonstrated that
inhibiting tyrosine kinase activity by tyrphostins (AG 126) actually
prevented LPS-induced septic shock in mice and, additionally,
tyrphostins block the production of TNF- of cultured primary
macrophage 36 . These findings emphasize the importance of tyrosine
kinase activity in the positive aspects of LPS signal transduction. Our
results showed that deletion of the Mer cytoplasmic tyrosine kinase
domain heightened TNF- production implying that at least one
tyrosine kinase inhibits LPS-dependent signaling.
These findings establish the first in vivo function for Mer tyrosine
kinase activity and for the Axl/Tyro3/Mer/Rek family of tyrosine
kinase. We have established that Mer influences the amount of NF-B,
and we have observed abnormal Jun-N-terminal kinase activity but
not p38 in LPS-challenged macrophages from Merkd mice
(our unpublished observation). Thus, it appears that Mer may
affect specific signal transductions pathways and not a general
inhibition of cell activation. Further experimentation is focused at
identifying the control point at which Mer down-regulates signal
transduction. The results reported here also suggest a novel inhibitory
pathway regulating the response to LPS and endotoxic shock that may be
exploited clinically.
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Acknowledgments |
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B p50 and p65 subunits and Rat-1 nuclear extracts; Dr. Cindy
Lawler for providing support with statistical analyses; Dr. Robert
Bagnell, Teresa Bone-Turrentine, Brian Garges, Anne Latour, Dr. Marty
Mayo, and Dr. Elizabeth Hicks for their excellent technical expertise;
and Drs. Rona Scott, Philip Cohen, and Jenny Ting for helpful
discussion and critical review of this manuscript. |
Footnotes |
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2 H.S.M. and G.K.M. are joint senior authors.
3 Address correspondence and reprint request to Dr. Glenn K. Matsushima, University of North Carolina Neuroscience Center, CB# 7250, University of North Carolina, Chapel Hill, NC 27599-7250. E-mail Address:
4 Abbreviations used in this paper: G3PDH, glyceraldehyde 3-phosphate dehydrogenase; PEC, peritoneal exudate cells; EMSA, electromobility shift assay.
Received for publication September 17, 1998. Accepted for publication December 9, 1998.
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B in the mortality of sepsis. J. Clin. Invest. 100:972.[Medline]
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promoter in primary macrophages. J. Immunol. 147:1649.
transcription in macrophages: involvement of four B-like motifs of constitutive and inducible forms of NF-B. Mol. Cell. Biol. 10:1498.B subunit expression during differentiation of human peripheral blood monocytes to macrophages. Cell Growth Differ. 8:435.[Abstract]
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S. Sather, K. D. Kenyon, J. B. Lefkowitz, X. Liang, B. C. Varnum, P. M. Henson, and D. K. Graham A soluble form of the Mer receptor tyrosine kinase inhibits macrophage clearance of apoptotic cells and platelet aggregation Blood, February 1, 2007; 109(3): 1026 - 1033. [Abstract] [Full Text] [PDF]
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P. Sen, M. A. Wallet, Z. Yi, Y. Huang, M. Henderson, C. E. Mathews, H. S. Earp, G. Matsushima, A. S. Baldwin Jr, and R. M. Tisch Apoptotic cells induce Mer tyrosine kinase-dependent blockade of NF-{kappa}B activation in dendritic cells Blood, January 15, 2007; 109(2): 653 - 660. [Abstract] [Full Text] [PDF]
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M. N. Sharif, D. Sosic, C. V. Rothlin, E. Kelly, G. Lemke, E. N. Olson, and L. B. Ivashkiv Twist mediates suppression of inflammation by type I IFNs and Axl J. Exp. Med., August 7, 2006; 203(8): 1891 - 1901. [Abstract] [Full Text] [PDF]
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Y. Li, M.-C. Gerbod-Giannone, H. Seitz, D. Cui, E. Thorp, A. R. Tall, G. K. Matsushima, and I. Tabas Cholesterol-induced Apoptotic Macrophages Elicit an Inflammatory Response in Phagocytes, Which Is Partially Attenuated by the Mer Receptor J. Biol. Chem., March 10, 2006; 281(10): 6707 - 6717. [Abstract] [Full Text] [PDF]
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 Y. Li, N. P. Mahajan, J. Webster-Cyriaque, P. Bhende, G. K. Hong, H. S. Earp, and S. Kenney The C-Mer Gene Is Induced by Epstein-Barr Virus Immediate-Early Protein BRLF1 J. Virol., November 1, 2004; 78(21): 11778 - 11785. [Abstract] [Full Text] [PDF]
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 C. L. McHenry, Y. Liu, W. Feng, A. R. Nair, K. L. Feathers, X. Ding, A. Gal, D. Vollrath, P. A. Sieving, and D. A. Thompson MERTK Arginine-844-Cysteine in a Patient with Severe Rod-Cone Dystrophy: Loss of Mutant Protein Function in Transfected Cells Invest. Ophthalmol. Vis. Sci., May 1, 2004; 45(5): 1456 - 1463. [Abstract] [Full Text] [PDF]
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P. H. Correll, A. C. Morrison, and M. A. Lutz Receptor tyrosine kinases and the regulation of macrophage activation J. Leukoc. Biol., May 1, 2004; 75(5): 731 - 737. [Full Text] [PDF]
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A. C. Morrison, C. B. Wilson, M. Ray, and P. H. Correll Macrophage-Stimulating Protein, the Ligand for the Stem Cell-Derived Tyrosine Kinase/RON Receptor Tyrosine Kinase, Inhibits IL-12 Production by Primary Peritoneal Macrophages Stimulated with IFN-{gamma} and Lipopolysaccharide J. Immunol., February 1, 2004; 172(3): 1825 - 1832. [Abstract] [Full Text] [PDF]
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Y. Qian, H. Wang, and S. H. Clarke Impaired Clearance of Apoptotic Cells Induces the Activation of Autoreactive Anti-Sm Marginal Zone and B-1 B Cells J. Immunol., January 1, 2004; 172(1): 625 - 635. [Abstract] [Full Text] [PDF]
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N. P. Mahajan and H. S. Earp An SH2 Domain-dependent, Phosphotyrosine-independent Interaction between Vav1 and the Mer Receptor Tyrosine Kinase: A MECHANISM FOR LOCALIZING GUANINE NUCLEOTIDE-EXCHANGE FACTOR ACTION J. Biol. Chem., October 24, 2003; 278(43): 42596 - 42603. [Abstract] [Full Text] [PDF]
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J. L. Duncan, M. M. LaVail, D. Yasumura, M. T. Matthes, H. Yang, N. Trautmann, A. V. Chappelow, W. Feng, H. S. Earp, G. K. Matsushima, et al. An RCS-Like Retinal Dystrophy Phenotype in Mer Knockout Mice Invest. Ophthalmol. Vis. Sci., February 1, 2003; 44(2): 826 - 838. [Abstract] [Full Text] [PDF]
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P. L. Cohen, R. Caricchio, V. Abraham, T. D. Camenisch, J. C. Jennette, R. A.S. Roubey, H. S. Earp, G. Matsushima, and E. A. Reap Delayed Apoptotic Cell Clearance and Lupus-like Autoimmunity in Mice Lacking the c-mer Membrane Tyrosine Kinase J. Exp. Med., July 1, 2002; 196(1): 135 - 140. [Abstract] [Full Text] [PDF]
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K. L. Guttridge, J. C. Luft, T. L. Dawson, E. Kozlowska, N. P. Mahajan, B. Varnum, and H. S. Earp Mer Receptor Tyrosine Kinase Signaling. PREVENTION OF APOPTOSIS AND ALTERATION OF CYTOSKELETAL ARCHITECTURE WITHOUT STIMULATION OR PROLIFERATION J. Biol. Chem., June 28, 2002; 277(27): 24057 - 24066. [Abstract] [Full Text] [PDF]
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 Q. Lu and G. Lemke Homeostatic Regulation of the Immune System by Receptor Tyrosine Kinases of the Tyro 3 Family Science, July 13, 2001; 293(5528): 306 - 311. [Abstract] [Full Text]
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 P. M. D'Cruz, D. Yasumura, J. Weir, M. T. Matthes, H. Abderrahim, M. M. LaVail, and D. Vollrath Mutation of the receptor tyrosine kinase gene Mertk in the retinal dystrophic RCS rat Hum. Mol. Genet., March 1, 2000; 9(4): 645 - 651. [Abstract] [Full Text] [PDF]
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