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HEMATOPOIESIS
From the Department of Internal Medicine, Division of
Rheumatology, Allergy and Immunology, Virginia Commonwealth University,
Richmond, Virginia, and the Department of Metabolic Diseases,
Hoffman-La Roche, Nutley, New Jersey.
Human mast cells in adult tissues have been thought to have
limited, if any, proliferative potential. The current study examined mast cells obtained from adult skin and cultured in serum-free medium
with recombinant human stem cell factor. During the first 4 weeks of
culture, the percentages of mast cells increased from 10 to almost 100. After 8 weeks, a 150-fold increase in the number of mast cells was
observed. When freshly dispersed mast cells were individually sorted
onto human fibroblast monolayers and cultured for 3 weeks, one or more
mast cells were detected in about two thirds of the wells, and in about
two thirds of these wells the surviving mast cells showed evidence of
proliferation, indicating most mast cells in skin can proliferate. Such
mast cells all expressed high surface levels of Kit and Fc Multipotential progenitor cells, capable of
becoming mast cells, leave the bone marrow and enter the circulation,
but they complete their differentiation into mature mast cells only
after arriving in peripheral tissues such as lung, bowel, skin, and nasal and conjunctival mucosa. Mature human mast cells can be distinguished from other cell types by expression of high levels of
surface Fc In a number of human diseases such as asthma,7,8 allergic
rhinitis,9,10 rheumatoid arthritis,11-13 and
vernal keratoconjunctivitis,14 significant increases in
mast cell density at the local affected sites have been described and
mast cells were estimated to play a central role in the pathophysiology
of the associated inflammation. The principal mechanisms involved in
mast cell hyperplasia were thought to be either the recruitment of
progenitor cells and their differentiation to mast cells or the
migration of mast cells from other regions of the tissue. Proliferation
has been considered less likely in part because cells were considered
to be terminally differentiated and also because mitotic mast cells are
rarely appreciated at these sites.
The proliferative potential of most myelomonocytic multipotential
hematopoietic cells diminishes as they differentiate. However, mature
murine mast cells have been reported to proliferate both in vivo and in
vitro. About 6% of mouse peritoneal mast cells proliferate after being
injected into skin of W/Wv mast
cell-deficient mice.15,16 Purified peritoneal mast
cells17,18 or dispersed skin mast cells,19
when plated in methylcellulose containing pokeweed mitogen-stimulated
spleen cell-conditioned medium, produced mast cell colonies. However,
in humans, isolated tissue-derived mast cells have shown little
capacity for proliferation. For example, dispersed newborn foreskin or
adult skin mast cells placed into culture with
lymphocyte-conditioned media and various cytokines showed
bromodeoxyuridine incorporation in up to 15% mast
cells.20
The current study shows that a substantial portion of mast cells
dispersed from adult human skin can proliferate under serum-free culture conditions with recombinant human stem cell factor (rhuSCF), while retaining expression of chymase and tryptase in secretory granules, Kit, and Fc Antibodies and reagents
Dispersal purification and culture of human skin mast
cells
Percoll gradient-enriched cells were suspended at the concentration of
1 × 106 cells/mL in serum-free AIM-V medium (Life
Technologies, Rockville, MD). Some cells were cultured at the same
concentration in complete RPMI 1640 medium supplemented with 10%
controlled process serum replacement-3 (CPSR-3; Sigma), 2 mM
L-glutamine, 0.1 mM nonessential amino acids, 10 mM Hepes,
50 µM 2-mercaptoethanol, 200 U/mL penicillin, and 100 µg/mL
streptomycin. In both cases, 100 ng/mL rhuSCF was included, and half
the culture medium was changed every week. When the cell number reached
2 × 106 cells/mL, half the cells were split to another
well. Cell numbers per sample were adjusted to account for the cells
that were split. Total cell number and viability were assessed by
trypan blue staining. The percentages of mast cells were assessed
cytochemically by metachromatic staining with toluidine blue, and by
flow cytometry with anti-Kit and anti-Fc Flow cytometry Surface Kit and Fc RI expression were assessed by
flow cytometry with mAbs YB5.B8 and 22E7, respectively. Cells were
incubated in 10% human AB serum in phosphate-buffered saline (PBS).
After washing in PBS containing 1% FCS and 0.1% sodium azide, cells then were incubated with anti-FcRI mAb or control mouse
IgG for 30 minutes at 4°C. After washing as above, cells were
incubated with fluorescein isothiocyanate (FITC)-conjugated goat
antimouse immunoglobulins (BD Biosciences, San Jose, CA) for 15 minutes at 4°C, and washed as above. After incubation with normal mouse IgG
to saturate antimouse antibody binding sites, cells were incubated with
phycoerythrin (PE)-conjugated anti-Kit mAb (BD Pharmingen) or control
IgG for 30 minutes at 4°C, resuspended in sheath solution, and
analyzed with a FACScan flow cytometer (BD Biosciences). The percentage
of positive cells was calculated by subtracting the percentage of cells
labeled with a control antibody (< 3% positive cells) from that of
cells with experimental mAb. HMC-1 cells, a human mast cell line, and
KU812 cells, a human basophil leukemia cell line, were used as positive
controls for surface Kit and FcRI expression, respectively.
For the detection of cells undergoing apoptosis after withdrawal of rhuSCF, cells were washed in PBS and incubated in the binding buffer as provided and recommended by the manufacturer (R & D Systems, Minneapolis, MN) with FITC-conjugated annexin-V, which binds to phosphatidylserine, a phospholipid that transfers from the inner to the outer side of the lipid bilayer of the plasma membrane during an early stage of apoptosis, and with propidium iodide (PI) for 15 minutes at room temperature to detect dead cells. Samples were directly analyzed by flow cytometry. To estimate the relative DNA content of cultured mast cells, cells were washed with PBS, incubated for 10 minutes on ice in hypotonic fluorochrome solution, which contained 50 µg/mL PI and 0.1% Triton-X100 in 0.1% sodium citrate, and then directly analyzed by flow cytometry. Immunochemistry Mast cells were assessed in cytocentrifuge preparations subjected to indirect immunocytochemical labeling as described.23 For chymase detection, B7-biotin-labeled cells were incubated with peroxidase-conjugated streptavidin and visualized with 3-amino-9-ethylcarbazole in 0.01% H2O2 to stain positive cells reddish brown. For tryptase detection, alkaline phosphatase-conjugated goat antimouse IgG (Sigma) was added to G3-labeled cells and visualized with naphthol AS-BI/new fuchsin to stain positive cells red. Cells were lightly counterstained with hematoxylin.Immunoaffinity magnetic enrichment and individual sorting Enzymatically dispersed skin cells were further enriched with mAb R24 against GD3, which was recently reported to be selectively expressed on the surface of mast cells in normal skin.28 Briefly, cells were incubated in PBS containing 10% human AB serum for 30 minutes, washed with PBS, and incubated with R24 (3 µg/1 × 107 cells) at 4°C for 30 minutes. Cells were washed with PBS to remove unbound mAb, incubated with microbeads conjugated to antimouse IgG (Miltenyi Biotec, Auburn, CA) at 4°C for 15 minutes, washed, and applied to the separation column. The column was exposed to a magnetic field in the magnetic cell separation system (Miltenyi Biotec) and washed to remove cells without attached microbeads. Retained cells were then eluted after the column was withdrawn from the magnetic field. Eluted cells were incubated with normal mouse IgG to saturate the antimouse antibody, and then with PE-conjugated mAb anti-Kit or control antibody for 1 hour at 4°C. Cells were then washed, and resuspended in PBS containing 20% AIM-V medium. Kit+ cells were individually seeded onto confluent 1059SK human fibroblasts (American Type Culture Collection, CRL-2072, Manassas, VA) in 96-well plates using a Coulter Epics Cell Sorting System (Coulter, Miami, FL) at Virginia Commonwealth University's Flow and Imaging Cytometry Facility, which permits single-cell sorting with an accuracy of more than 99%. Sorted cells were then cultured in AIM-V medium supplemented with 100 ng/mL rhuSCF. Half the medium was changed every week. After 3 weeks mast cell numbers were checked by immunocytochemistry for tryptase.Activation of mast cells and tryptase immunoassay Cultured mast cells were washed in Tyrode buffer containing 10 mM Hepes, pH 7.4, 0.1% gelatin, and 100 ng/mL DNase (TGD /), then suspended in TGD/ buffer
containing 1 mM MgCl2 and 2.5 mM CaCl2
(TGD+/+) and preincubated for 5 minutes at 37°C. For
stimulation, 5 µL mAb 22E7 or control mouse IgG, or nonimmune
reagents such as substance P or compound 48/80 (Sigma) were added to 25 µL cell suspension (1 × 106 cells/mL) in a 96-well
plate, and incubated for a further 15 minutes at 37°C. The reaction
was stopped by adding 200 µL ice-cold TGD/ buffer.
The cells were separated by centrifugation at 300g for 7 minutes at 4°C, and supernatant was carefully removed and adjusted to
1.2 M NaCl by adding 40 µL 5 M NaCl. The cell pellet was resuspended in 200 µL ice-cold extraction buffer (10 mM MES, pH 6.5, with 2 M
NaCl), quickly frozen in liquid nitrogen, and thawed 4 times to release
all cellular tryptase. After centrifugation at 12 000g for
15 minutes at 4°C, the soluble extract was collected and stored at
 70°C until ready to be assayed. Total immunoreactive tryptase levels were measured by an enzyme-linked immunosorbent assay (ELISA) using mAb B12 to capture tryptase and biotinylated mAb G5 to detect captured tryptase as described29; purified human
lung-derived tryptase was used as standard.
Proliferation of human skin-derived mast cells in a serum-free culture Because serum may contain both beneficial and detrimental factors for mast cell growth in vitro, synthetic media are potentially advantageous. Serum-free AIM-V media permitted an earlier, more selective and greater expansion of fetal liver-derived mast cells than serum-containing media in the presence of rhuSCF.23 However, most tissue-derived human mast cells, though probably long-lived, are thought to be terminally differentiated with limited, if any, proliferative capacity. The growth of skin-derived mast cells in the presence of rhuSCF (100 ng/mL) was assessed with this serum-free media compared to serum-containing RPMI 1640 (Figure 1). As expected with RPMI 1640 medium with serum, after an 8-week culture period, total cell numbers diminished by 90% (Figure 1A), mast cell percentage by about 80% (Figure 1B), and mast cell number by over 90% (Figure 1C). At 1 week there was a slight increase in the percentage of mast cells due to a more rapid loss of other cell types, but there was no increase in mast cell number. In contrast to serum-containing media, results with AIM-V media were dramatically different. Total cell numbers diminished slightly after the first week, but then increased 16-fold by 8 weeks (Figure 1A). Under these serum-free culture conditions the mast cell percentage increased from about 10% at the initiation of culture to nearly 100% by 4 weeks, and remained so for the remaining 4 weeks (Figure 1B). Mast cell numbers increased progressively from 0.1 × 106 cells on day zero to 15.9 × 106 after 8 weeks. A similar growth pattern was recognized in another 5 independent specimens. Of the 4 specimens kept in culture under serum-free conditions beyond 8 weeks, one stopped proliferating at 8 to 9 weeks, 2 stopped at 9 to 10 weeks and one continued until the culture was harvested at 12 weeks.
Skin-derived mast cells were examined by light microscopy (Figure
2). A cytospin of freshly dispersed skin
cells, stained with toluidine blue, shows about 10% of the cells to be
metachromatic with oval, nonsegmented nuclei (Figure 2A). After 4 weeks
of culture in AIM-V medium, monolayers of tightly adherent cells were
not observed. Instead, almost all of the cells were metachromatic (Figure 2B), although the staining intensity appeared to be less than
for newly dispersed mast cells. In contrast, after 4 weeks of culture
in RPMI 1640 with CPSR-3, the culture well was completely covered by
fibroblast-like cells. Toluidine blue staining of cytocentrifuged cell
preparations revealed few metachromatic cells, many fibroblast-like cells, and occasional metachromatic bodies in fibroblast- or
macrophage-like cells, suggesting that ingestion of mast cells or mast
cell granules may have occurred (Figure 2C). Also, morphometric
measurements (mean ± SE, n = 4 separate skin samples, 30 mast
cells examined per preparation) of mast cell diameter revealed cells
cultured for 4 to 8 weeks in AIM-V media (25.7 ± 0.2 µm,) to be
larger (P < 0.001, t test) than those prior to
culture (13.2 ± 0.2 µm). Although most nuclei were oval and
nonsegmented, 4.6% ± 2.1% (n = 5 specimens) of the cultured mast
cells were binucleated or showed mitotic figures (Figure 2B), features
not observed before culture. Almost no fibroblast-like cells were
observed. Essentially all mast cells in normal adult skin contain both
tryptase and chymase (MCTC type of mast
cell).21 To test whether skin-derived mast cells retained
this protease phenotype after 4 to 8 weeks of culture in serum-free
AIM-V media containing rhuSCF, cytospins of these cells were stained
with antichymase (Figure 2 D) and antitryptase (Figure 2 E) mAbs, and
counterstained with hematoxylin. Essentially all cells were labeled
with each mAb, indicating the MCTC phenotype was preserved
under these culture conditions.
To further evaluate the proliferation activity of 4-week-old cultured
mast cells, DNA content was assessed by flow cytometry with PI. In a
DNA fluorescence histogram, the cells in S, G2, and M
phases are shown in the M1 region of Figure
3A, whereas those in G0 and
G1 phases reside in the peak that precedes M1. When
4-week-old mast cells grown in AIM-V media were examined before (left
panel, 7 days after prior feeding) and 24 hours after rhuSCF feeding
(right panel), 3.7% ± 1.8% and 9.4% ± 3.3% of the cells,
respectively, were distributed in M1 (n = 3, mean ± SD). Thus
proliferation rates appeared to be somewhat greater 24 hours than 7 days after treatment with rhuSCF. Also, to confirm that these cells
were mast cells, surface Kit (Figure 3B) and Fc
Skin-derived mast cells cultured in AIM-V media are dependent on exogenous SCF for survival Survival of human mast cells is acknowledged to be primarily dependent on SCF. To test whether proliferating skin-derived mast cells were dependent on this growth factor, 3 separate preparations of 6- to 8-week cultures of mast cells in AIM-V media were washed and placed into culture with or without rhuSCF (100 ng/mL). As shown in Figure 4A, viability (trypan blue) remained near 100% in the presence of rhuSCF, but steadily decreased in the absence of rhuSCF. By day 1 and beyond the percent viabilities of cells cultured with rhuSCF were significantly different from those cultured without rhuSCF; by day 3 and beyond the percent viabilities of cells cultured in the absence of rhuSCF were significantly lower than the day 0 cells. The percentages of apoptotic cells on day 2 were evaluated by annexin-V surface and PI labeling. Without rhuSCF 49% of the cells were labeled with annexin V, whereas less than 20% (as with trypan blue) were also labeled with PI. All of the annexin V+ cells were smaller than the viable unlabeled cells (forward side scatter by flow cytometry), consistent with apoptosis being their death pathway. With rhuSCF only about 6% of the cells were labeled with annexin V and about 5% with PI. As shown in Figure 4B, with rhuSCF the number of viable cells increased significantly by day 3, whereas without rhuSCF the cell numbers had significantly declined by day 3 when compared to day 0 cells. Also, viable cell numbers in the presence and absence of rhuSCF were significantly different by day 3.
Proliferative potential of human skin-derived mast cells From the experiments described thus far, it was not clear whether the proliferating mast cells in this serum-free culture condition were derived from mature mast cells or from progenitors that might reside in preparations of dispersed skin cells. To address this, mast cells were immunoaffinity purified with anti-GD3 mAb to at least 95% purity as described,28 and then subjected to cell sorting after labeling the cells with PE-conjugated anti-Kit mAb. A portion of these cells was placed directly onto glass slides that then were stained with toluidine blue. All exhibited metachromasia, indicating this population contained virtually 100% mature mast cells (Figure 5). Another portion of the cells was individually sorted into 96-well culture plates, each well containing a confluent monolayer of human fibroblasts to act as feeder cells. Fibroblasts together with rhuSCF were necessary for cell survival; in the absence of either fibroblasts or exogenous rhuSCF, no cells were recovered after 3 weeks. These confluent fibroblasts survived but did not proliferate under the serum-free culture condition used. After 3 weeks of culture with AIM-V medium containing 100 ng/mL rhuSCF, the number of mast cells per well was evaluated by immunocytochemistry with antitryptase mAb. Results are shown in Figure 5B and Table 1. Mean data taken from 4 separate experiments indicated only one tryptase-positive cell in 19% of the wells, and 2 or more tryptase-positive cells per well in 44% of the wells (11 cells/well on average). Thus, one or more mast cells were detected in 63% of the wells, and in about two thirds of the mast cell-containing wells there was evidence for mast cell proliferation. In 2 control experiments single Kit-sorted HMC-1 cells yielded detectable cells in 65% (58% and 71%) of the wells, the remainder presumably not surviving the sorting or the culture conditions. The percentages of wells with surviving mast cells are similar for skin-derived mast cells and HMC-1 cells. These results indicate that most mature skin-derived mast cells are capable of proliferating in serum-free media containing rhuSCF.
Proliferating skin-derived mast cells retain their characteristic functional phenotype Experiments were performed to determine whether cultured skin-derived mast cells degranulate in response to FcRI
cross-linking and to agents known to stimulate skin but not
lung-derived mast cells. As shown in Figure
6A, tryptase release occurred in a
dose-response fashion to anti-FcRI mAb, maximal release
being observed at 0.3 to 3 µg/mL. Also, the amount of tryptase per
mast cell was calculated to be 22 ± 4 pg/mast cell (n = 5),
slightly less than the amount measured per mast cell from skin tissue
shortly after their dispersion and density-dependent enrichment, which
was 29 ± 3 pg/mast cell (n = 3). Degranulation also was observed
in a dose-dependent manner to substance P (Figure 6B) and to compound 48/80 (Figure 6 C).
The current study shows that skin-derived mast cells are capable
of proliferation, which could contribute to mast cell hyperplasia in
tissues. However, human mast cells that reside in peripheral tissues of
adult hosts have been considered to be long-lived cells with limited,
if any, proliferative capacity. At sites where mast cell hyperplasia
occurs, such as urticaria pigmentosa in skin,30 rheumatoid
synovium,11 bronchopulmonary dysplasia and fibrotic lung
disease,31 and atopic
keratoconjunctivitis,14,32 the rare presence of mitotic
mast cells has been used to argue that proliferation of mature mast
cells does not account for their increased presence. Even when mast
cell hyperplasia is iatrogenically induced by administration of rhuSCF,
mitotic mast cells in skin biopsies were not reported.33
Consequently, expansion of mast cell populations at specific tissue
sites in humans has been predicted to occur by diverting a greater
portion of the pool of hematopoietic progenitor cells to a mast cell
lineage, by stimulating proliferation of mast cell progenitors, by
increasing recruitment of mast cells and their precursors into tissues,
or by increasing survival of mast cells. Recruitment of mast cell from
one region of a tissue to another or of precursors from the circulation
to tissue sites may be driven by various chemokines,34-37
transforming growth factor- The current study demonstrates that most mast cells derived from adult skin can proliferate in serum-free media containing rhuSCF. This suggests that SCF is a proliferation factor as well as a survival42,43 and priming or activation44,45 factor for mature human mast cells. Mast cell numbers in suspension cultures increased 150-fold by 8 weeks. At 4 weeks, mitotic or binucleated cells accounted for 4.6% of the mast cells. A DNA fluorescence histogram with PI revealed that 9.4% of the 4-week cells were distributed in the S, G2, and M phases of the cell cycle. To rule out the possibility that this represented expansion of a small progenitor cell population, mature mast cells were purified by density-dependent sedimentation and immunoaffinity purification with anti-GD3 mAb,28 and then were individually sorted using PE-labeled anti-Kit mAb onto fibroblast monolayers in wells of microtiter plates. At least two thirds of the mature mast cells that survived such Kit-dependent sorting had the ability to proliferate. Why this was observed in serum-free but not serum-containing media is not known, but is consistent with previous observations showing survival but not proliferation of mast cells in serum-containing media.26,27 With serum, a substantial increase in fibroblast-like cells and metachromatic bodies in nonmast cells were observed in our experiments. Consequently, the attenuating effects of serum on mast cell growth, in part, could be indirect by stimulating other cell types to grow and to phagocytose mast cells. However, when serum was added to AIM-V-cultured mast cells at 4 weeks, when mast cells predominated and few if any fibroblast-like cells were observed, proliferation of mast cells was curtailed, suggesting direct as well as indirect effects of serum on mast cell growth. Thus, mature mast cells in human skin have the capacity to proliferate when exposed to SCF, but factor(s) in serum or plasma may regulate this response. Whether the extracellular fluid to which tissue mast cells are exposed in vivo acts similarly to serum in vitro remains to be determined. Proliferating skin-derived mast cells appear to retain several key
phenotypic features of freshly dispersed cells. For example, nearly all
expressed high levels of cell-surface Kit and Fc The typical protease phenotype of skin-derived mast cells, namely, both tryptase and chymase expression by immunocytochemistry, also was maintained. After 4 to 8 weeks of culture, the tryptase content measured in cell extracts by an ELISA averaged 22 pg/mast cell, slightly less than the 29 pg/mast cell measured after their initial dispersion. These values are comparable to what has been reported for freshly dispersed skin-derived mast cells (35 pg/mast cell),25 and greater than the content of lung-derived mast cells (11 pg/mast cell), and fetal liver-derived mast cells (3.7 ± 0.1 pg/mast cell, n = 323). As for tryptase, essentially all skin-derived mast cells also express chymase when they are dispersed from skin, and then retain their chymase-positive phenotype during culture in serum-free AIM-V medium. In contrast, suspension cultures of fetal liver-derived cells with rhuSCF produce mast cells with abundant chymase messenger RNA, but few mast cells with detectable chymase protein when serum-containing media is used.50 About one third of the mast cells express chymase protein when they develop in suspension cultures with serum-free AIM-V medium.23 Although human mast cells from all tissues degranulate in response to
Fc
We are grateful to Frances K. White (Flow and Imaging Cytometry Facility, Virginia Commonwealth University, Richmond, VA) for technical support with cell sorting.
Submitted October 27, 2000; accepted December 7, 2000.
Supported by National Institutes of Health grants AI27517 and AI20487 (to L.B.S.).
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: Lawrence B. Schwartz, Virginia Commonwealth University, PO Box 980263, Richmond, VA 23298-0263; e-mail: lschwart{at}hsc.vcu.edu.
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© 2001 by The American Society of Hematology.
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  W. Zhao, G. Gomez, S.-H. Yu, J. J. Ryan, and L. B. Schwartz TGF-{beta}1 Attenuates Mediator Release and De Novo Kit Expression by Human Skin Mast Cells through a Smad-Dependent Pathway J. Immunol., November 15, 2008; 181(10): 7263 - 7272. [Abstract] [Full Text] [PDF]
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Y. Fukuoka, H.-Z. Xia, L. B. Sanchez-Munoz, A. L. Dellinger, L. Escribano, and L. B. Schwartz Generation of Anaphylatoxins by Human {beta}-Tryptase from C3, C4, and C5 J. Immunol., May 1, 2008; 180(9): 6307 - 6316. [Abstract] [Full Text] [PDF]
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G. Gomez, S. Jogie-Brahim, M. Shima, and L. B. Schwartz Omalizumab Reverses the Phenotypic and Functional Effects of IgE-Enhanced Fc{epsilon}RI on Human Skin Mast Cells J. Immunol., July 15, 2007; 179(2): 1353 - 1361. [Abstract] [Full Text] [PDF]
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 J. Tessier, C. Green, D. Padgett, W. Zhao, L. Schwartz, M. Hughes, and E. Hewlett Contributions of Histamine, Prostanoids, and Neurokinins to Edema Elicited by Edema Toxin from Bacillus anthracis Infect. Immun., April 1, 2007; 75(4): 1895 - 1903. [Abstract] [Full Text] [PDF]
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W. Zhao, C. L. Kepley, P. A. Morel, L. M. Okumoto, Y. Fukuoka, and L. B. Schwartz Fc{gamma}RIIa, Not Fc{gamma}RIIb, Is Constitutively and Functionally Expressed on Skin-Derived Human Mast Cells J. Immunol., July 1, 2006; 177(1): 694 - 701. [Abstract] [Full Text] [PDF]
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 F. Schiemann, T. A. Grimm, J. Hoch, R. Gross, B. Lindner, F. Petersen, S. Bulfone-Paus, and E. Brandt Mast cells and neutrophils proteolytically activate chemokine precursor CTAP-III and are subject to counterregulation by PF-4 through inhibition of chymase and cathepsin G Blood, March 15, 2006; 107(6): 2234 - 2242. [Abstract] [Full Text] [PDF]
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W. Zhao, C. A. Oskeritzian, A. L. Pozez, and L. B. Schwartz Cytokine Production by Skin-Derived Mast Cells: Endogenous Proteases Are Responsible for Degradation of Cytokines J. Immunol., August 15, 2005; 175(4): 2635 - 2642. [Abstract] [Full Text] [PDF]
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S. Bagga, K. S. Price, D. A. Lin, D. S. Friend, K. F. Austen, and J. A. Boyce Lysophosphatidic acid accelerates the development of human mast cells Blood, December 15, 2004; 104(13): 4080 - 4087. [Abstract] [Full Text] [PDF]
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C. A. Oskeritzian, W. Zhao, A. L. Pozez, N. M. Cohen, M. Grimes, and L. B. Schwartz Neutralizing Endogenous IL-6 Renders Mast Cells of the MCT Type from Lung, but Not the MCTC Type from Skin and Lung, Susceptible to Human Recombinant IL-4-Induced Apoptosis J. Immunol., January 1, 2004; 172(1): 593 - 600. [Abstract] [Full Text] [PDF]
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T. Kumamoto, D. Shalhevet, H. Matsue, M. E. Mummert, B. R. Ward, J. V. Jester, and A. Takashima Hair follicles serve as local reservoirs of skin mast cell precursors Blood, September 1, 2003; 102(5): 1654 - 1660. [Abstract] [Full Text] [PDF]
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L. B. Schwartz, H.-K. Min, S. Ren, H.-Z. Xia, J. Hu, W. Zhao, G. Moxley, and Y. Fukuoka Tryptase Precursors Are Preferentially and Spontaneously Released, Whereas Mature Tryptase Is Retained by HMC-1 Cells, Mono-Mac-6 Cells, and Human Skin-Derived Mast Cells J. Immunol., June 1, 2003; 170(11): 5667 - 5673. [Abstract] [Full Text] [PDF]
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 P. Bradding, Y. Okayama, N. Kambe, and H. Saito Ion channel gene expression in human lung, skin, and cord blood-derived mast cells J. Leukoc. Biol., May 1, 2003; 73(5): 614 - 620. [Abstract] [Full Text] [PDF]
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Top
Abstract
Introduction
) or AIM-V
(
) as described. Total viable cell numbers (A), percentages of mast
cells (metachromatic staining) (B), and numbers of mast cells (product
of total cell number and mast cell percentage) (C) are depicted. Each
data point is the mean value from 3 separate experiments, each
performed in triplicate. Error bars indicate the SD. When the total
cell concentration in a well exceeded 2 × 106 cells/mL,
half of the cells were either transferred to another well or discarded;
cell numbers were adjusted to account for the cells that were discarded
as well as those that were retained when cultures were
split.
(TGF-
association of the MCTC subset with matrix turnover and clinical progression.
Arthritis Rheum.
1997;40:479-489