Download The role of class II antigen-expressing cells in corneal

Investigative Ophthalmology & Visual Science, Vol. 31, No. 11, November 1990
Copyright © Association for Research in Vision and Ophthalmology
The Role of Closs II Antigen-Expressing Cells
in Corneol Allogroft Immunity
Bryan M. Gebhardr
The goal of this study was to investigate the relationship between the presence of Class II antigen-expressing cells (Class II+) cells in the cornea and the generation of allograft immunity. Wistar/Furth
(W/F) rat Class II+ cells were injected in various numbers (0.01, 0.1,1.0, 5.0, 10.0, and 20.0 X 106)
into the corneal stroma of Fischer 344 (F344) rats. For comparison, the same numbers of W/F Class
II+ cells were injected directly into the peritoneal cavity of F344 rats. Also, W/F cells were injected
into the corneas of F344 rats and the corneas were "grafted" intraperitoneally in F344 hosts. The
results showed that up to 20 X 106 class II+ cells injected in situ into the corneal stroma did not elicit a
serum cytotoxic antibody response or a splenic or blood cytotoxic T-cell response against donor Class
II antigens. In contrast, systemic immune responses were elicited by both direct intraperitoneal
injection of large numbers (10 or 20 X 106) of allogeneic Class II+ cells and by intraperitoneal grafting
of syngeneic corneas carrying similar numbers of allogeneic Class II+ cells. F344 recipients of a
syngeneic cornea containing 10 or 20 X 106 W/F Class II+ cells or a suspension of W/F cells exhibited
an accelerated rejection of W/F skin allografts (13.3 versus 9.0 days, first- versus second-set rejection). These results indicate that the number of Class II+ cells required to elicit a systemic immune
response is larger than the number of Class II+ cells present in the normal cornea. Invest Ophthalmol
Vis Sci 31:2254-2260, 1990
corneal allografts.2'5'6"101213 Although it has been
shown that both Class I and II histocompatibility antigens expressed on corneal cells may stimulate an
allograft reaction,67 it has been suggested that cells
expressing Class II antigens are more efficient in this
capacity.7'9'10
Cells in various tissues undergoing immunologic
attack are induced to express Class II antigens that
they normally do not express.14"22 Young and
McMillan23 showed that corneal stromal keratocytes
in culture can be induced to express Class II histocompatibility antigens when exposed to the lymphokine, gamma interferon. These investigators speculated that the induced expression of Class II histocompatibility antigens could be an important early
event in the initiation of the corneal allograft immune reaction.23
Our purpose was to determine the relationship between the presence of Class II+ cells in the cornea and
the stimulation of a host antigraft reaction. These
results demonstrate that the presence of large numbers of allogeneic Class II+ cells in the cornea does not
elicit a systemic immune reaction; this suggests that
the immunity engendered by Class II+ allogeneic cells
in the cornea is a local rather than a systemic phenomenon. The role of donor Class II+ cells in eliciting
immune rejection of corneal grafts in the human eye
requires further study.
The cells of the normal cornea consist almost exclusively of epithelial cells, stromal keratocytes, and
endothelial cells, with relatively few immune accessory cells expressing Class II antigens.1"9 The Class II
antigen-expressing (Class II+) Langerhans cells present in the normal cornea are distributed in a gradient
from the peripheral cornea and conjunctiva, where
the largest numbers are found, to the central cornea,
where there are few or none.3'4'6"8 Both resident and
infiltrating Class II+ cells are present in larger numbers in corneas subjected to various stresses;1"10 their
distribution changes markedly in various corneal disease states, including corneal inflammatory reactions,7"9 corneal infections,611 and the corneal allograft immune response.6'710
Several investigators proposed that Class II+ cells of
donor origin contribute to the immunogenicity of
From the Lions Eye Research Laboratories, LSU Eye Center,
Louisiana State University Medical Center School of Medicine,
New Orleans, Louisiana.
Supported in part by Public Health Service grants EY03150 and
EY02377 from the National Eye Institute, National Institutes of
Health, Bethesda, Maryland.
Presented in part at the annual meeting of the Association for
Research in Vision and Ophthalmology, May 5, 1989, Sarasota,
Florida.
Reprint requests: Bryan M. Gebhardt, PhD, LSU Eye Center
2020 Gravier Street, Suite B, New Orleans, LA 70112.
2254
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CLASS II ANTIGEN-EXPRESSING CELLS / Gebhordr
No. 11
Materials and Methods
Animals
Female rats of the inbred strains, Wistar/Furth
(W/F, Rtl u ) and Fischer 344 (F344, Rtlvl), weighing
approximately 120 g, were used (Harlan Sprague
Dawley, Indianapolis, IN). All animals were maintained and handled according to the ARVO Resolution on the Use of Animals in Research.
Cell Suspensions
The W/F and F344 rats were killed by sodium
pentobarbital euthanasia. The spleens were removed
using sterile technique, prepared as single cell suspensions, and freed of erythrocytes and dead cells on
Lympho-paque gradients (Accurate Chemical and
Scientific, Westbury, NY). Total cell number and cell
viability were determined by hemocytometer count
using the vital dye, trypan blue.
Cell Fractionation
Two micrograms of mouse monoclonal, rat antiClass II antibody were incubated with 1 mg of magnetic immunobeads (Dynabeads M-450; Robbins,
Mountainview, CA) for 4 hr at room temperature
with gentle rocking. After incubation the labeled
beads were suspended in 10 ml of RPMI-1640
(Grand Island Biological, Grand Island, NY), containing 0.3% bovine serum albumin (Sigma, St.
Louis, MO) (RPMI/BSA), and concentrated using a
Dynal Magnetic Particle Concentrator (Robbins).
The supernatant was discarded, and the labeled beads
were resuspended in an additional 10 ml of RPMI/
BSA and again collected with the particle concentrator. Washing and concentration of the beads were
repeated two additional times.
Ten microliters of the suspension containing approximately 4 X 106 antibody-coated beads was adequate for the isolation of 5 X 106 Class II+ cells. In
practice, 0.5 ml of antibody-coated beads was incubated with 1 X 108 rat spleen cells (SC) for 30 min at
4°C on a rocking platform. The RMPI/BSA was
added to a final volume of 10 ml, the beads with
attached cells were collected in the magnetic particle
concentrator, and the supernatant removed. The
washing and concentration procedure was repeated
three additional times.
The bead-cell complexes were incubated at 37°C
for 8 hr, during which time the cells separated from
the beads. At the end of the incubation, the detached
beads were collected on the magnetic particle concentrator and the cell suspension collected in the supernatant. Cell number and viability were determined by hemocytometer count. The viability of cells
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2255
purified by this method was uniformly greater than
97%. The cell suspensions were held at 4°C for use in
the experimental protocol.
The presence of Class II histocompatibility antigens on the cells was confirmed by immunoperoxidase staining as previously reported.24 At least 95% of
the immunobead-purified cells were Class II+.
Cell-Mediated Cytotoxicity
Peripheral blood lymphocytes (PBLs), peritoneal
exudate lymphocytes (PELs), and SCs were used as
the sources of cytotoxic effector cells. Animals that
had been injected with allogeneic Class II+ cells were
killed at the times indicated in the experimental design. To obtain PBLs, 5-7 ml of whole blood was
collected in heparinized syringes. For PELs, the peritoneal cavity was lavaged with 10 ml of RPMI/BSA.
For SCs, the spleen was removed with sterile dissecting instruments and reduced to a cell suspension.
The cell suspensions obtained from blood, peritoneal exudates, and spleens were fractionated over
Lympho-paque gradients. Next, each cell suspension
was separately enriched for T-cells by nylon-wool
fractionation. Approximately 0.5 g of scrubbed
nylon-wool (type 200L; Dupont, Wilmington, DE)
was packed in a 10-ml syringe and equilibrated with
RPMI/BSA at 37°C for 1 hr. The mononuclear cells
obtained from the peritoneal washout, peripheral
blood gradient, or approximately one third of a total
spleen cell suspension (30 X 106 cells) were added to 2
ml of RPMI/BSA and pipetted onto each column.
After 1 hr at 37°C the nonadherent T-lymphocytes
were eluted from the nylon column with 50 ml of
RPMI/BSA. The T-cells were washed, counted, and
resuspended in complete tissue culture medium consisting of RMPI-1640 supplemented with 10% fetal
bovine serum (Grand Island Biological).
Class II+ cells used as 51Cr-labeled target cells were
obtained from the spleens of W/F strain rats and putor. Washing and concentration of the beads were
suspended in culture medium containing 100 nC\ of
Na251CrO4 (ICN Radiochemicals, Irvine, CA) at a
concentration of 1 X 107 cells/ml and incubated at
37°C for 60 min. The cells were washed three times
in culture medium, incubated for an additional 30
min at 37°C, and washed once more. The effector
and target cells were mixed together in round-bottom
microcytotoxicity plates (Falcon Labware; Becton
Dickinson, Lincoln Park, NJ) in ratios of 1:1, 10:1,
20:1, 40:1, 80:1, and 160:1.
Groups of eight replicate wells were established for
each mixture of cells. The plates were incubated for 5
hr at 37°C, then centrifuged at 250 X g for 5 min.
Fifty-microliter aliquots of the supernatant from each
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INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / November 1990
well were collected individually in 12 X 75-mm plastic test tubes (Falcon; Becton Dickinson) and the
amount of radioactivity determined in a gamma
counter. Control wells for determining the amount of
spontaneous 5lCr release contained target cells alone.
The total releasable 5lCr was determined by adding
50 n\ of a 1% solution of Triton X-100 to wells containing 51Cr-labeled cells. The percentage of specific
cytotoxicity was calculated according to the formula:
Percent Specific Cytotoxicity
Experimental CPM - Background Release CPM
•X 100
Release CPM - Background Release CPM
Serum Alloantibody Assay
Endogenous complement in sera obtained from
F344 strain recipients was inactivated by heating at
56°C for 30 min. The anti-Class I antibody was removed from the sera by repeated absorption with
W/F strain erythrocytes. Removal of the anti-Class I
antibody was assured by testing the absorbed serum
in hemagglutination assay as previously reported.25
Spleen cells from a W/F donor were gradient purified
on Lympho-paque and the Class II+ cells enriched
using the magnetic immunobead technique. The purified cells were suspended at a concentration of 5
X 106 cells/ml in RPMI/BSA and dispensed in 25-/il
volumes into the wells of round-bottom microtiter
plates containing serial twofold dilutions of serum
samples obtained from F344 donors. Next, 25 fA of a
1:10 dilution of Low Tox M rabbit complement (Accurate Chemical and Scientific) was added to all
wells, and the plates were incubated for 45 min at
37°C. The percentage of cells killed was determined
by hemocytometer chamber counts using trypan blue
as the indicator. Control sera obtained from nonimmune F344 donors and control wells containing only
the complement were done with each assay.
Skin Grafting
The technique of skin grafting rats originally reported by Krapienis26 and used in previous studies25
was employed. Rats of the W/F and F344 strains were
anesthetized with an intramuscular injection of Ketaset 60 mg/kg (ketamine hydrochloride; Bristol, Syracuse, NY) and Rompun 0.6 mg/kg (xylazine; Butler,
Columbus, OH). The W/F skin was obtained from
the dorsal basal region of the tail and grafted onto the
dorsal cervical region of the F344 recipients. The
grafts were inspected daily after removal of the gauze
covering. Complete rejection of the skin graft was
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Vol. 31
recorded as the day on which the graft was dry to the
touch, discolored, and thought to be inviable.25'26
Experimental Design
To test the capacity of Class II+ cells to elicit humoral and cellular immune responses after introduction into allogeneic recipients, several experimental
groups were studied. The W/F Class II+ cells were
injected into the corneas or peritoneal cavities of
F344 rats. For corneal injection, six groups of 12
F344 rats each were anesthetized and given injections
of 0.01, 0.1, 1.0, 5.0, 10.0, or 20.0 X 106 purified
W/F strain Class II+ cells into the corneal stroma
using a previously described technique.24 Two other
groups of F344 rats received equivalent injections of
W/F Class II+ cells as a suspension or as corneal implants into which W/F cells had first been injected.
This latter group was included to determine the effect
of the corneal milieu on the immunogenicity of cells.
The technique of implanting corneas into the peritoneal cavity was done as previously described.25 Negative-control F344 rats received 1.0-ml volumes of
RPMI-1640.
The time course of the development of humoral
and cellular immunity was determined by bleeding
and killing two F344 recipients from each experimental and control group at 1,2, 3, 4, and 5 weeks after
grafting or injection.
To determine whether the introduction of large
numbers of Class II+ cells is capable of generating
second-set graft rejection, four groups of F344 rats
were grafted with W/F skin. One group consisted of
eight F344 rats that had been injected with 20 X 106
W/F Class II+ cells into the corneal stroma 5 weeks
earlier. A second group involved eight F344 rats that
had received a syngeneic cornea containing 20 X 106
W/F Class II+ cells implanted intraperitoneally 5
weeks earlier. A third group included eight F344 rats
that had received an intraperitoneal injection of 20
X 106 W/F Class II+ cells 5 weeks earlier. The control
group consisted of 12 normal F344 rats; this group
was used to determine the median survival time
(MST) of W/F skin on the allogeneic F344 recipients.
Data Analysis
Each experiment was repeated at least three times.
The data in the 51Cr release assays and the serum
alloantibody assays were reduced to means and standard deviations of the means, and results of the different experimental and control groups were compared for statistical significance by analysis of variance.
CLASS II ANTIGEN-EXPRESSING CELLS / Gebhordr
No. 11
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Results
Serum Alloantibody Response
None of the F344 rats that received W/F Class II+
cells injected into the corneal stroma developed a
serum alloantibody response over the 5-week period
of the study (Fig. 1A). In contrast, the animals that
received an intraperitoneal injection of either 10
X 106 or 20 X 106 W/F strain Class II+ cells showed a
serum alloantibody response measurable at both 1
and 2 weeks after injection (Fig. 1 A). Lower cell numbers did not elicit an antibody response.
The F344 corneas injected with W/F Class II+ cells
and implanted intraperitoneally in F344 rats also
elicited an antibody response. The corneas containing the largest number of cells (20 X 106) generated
the greatest response, which became evident 14 days
after transplantation and reached a peak at approximately 21 days (Fig. IB). Corneas carrying 10 X 106
Class II+ cells produced a small, but measurable response; lower numbers of cells did not elicit a response above control levels (Fig. 1B).
Cell-Mediated Immune Response
The W/F Class II+ cells injected into the corneal
stroma did not elicit a measurable cellular cytotoxic
response in the blood, spleen, or peritoneal cavity of
the F344 recipients over the 5 weeks of the study. The
Fig. 1. Serum cytotoxic antibody assay results. F344 rats were
injected intrastromally, intraperitoneally, or were "grafted" intraperitoneally with 0.01, 0.1, 1.0,5.0, 10.0, or 20.0 X 106 W/F Class
II + cells and their sera tested for cytotoxic antibody at five weekly
intervals. Positive cytotoxic responses were obtained in the recipients of 10 (circles) and 20 (squares) X 106 Class II + cells given either
as a suspension (A) or as a syngeneic cornea (B) containing the
allogenic cells placed intraperitoneally. The response to lower cell
numbers fell within the 0-10% range (shaded area). Similarly, the
recipients of intrastromal injections failed to produce serum alloantibody; all the serum titers of these animals fell within the
0-10% range (shaded area). By analysis of variance (ANOVA) the
differences between the cytotoxicity elicited by 10 and 20 X 106
cells and the lower cell numbers tested were statistically significant
(P < 0.0005).
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Fig. 2. Cell-mediated cytotoxic responses of F344 rats immunized with W/F Class II + cells. F344 rats were injected intrastromally or intraperitoneally with 0.01, 0.1, 1.0, 5.0, 10.0, or 20.0
X 106 Class II + cells and the presence of cytotoxic T cells in the
blood, spleen, and peritoneal cavity was assessed. No cytotoxic cells
were found in the blood. Ten (circles) and 20 (squares) X 106
allogeneic Class II + cells injected intraperitoneally elicited positive
peritoneal (A) or splenic (B) cytotoxic responses, whereas the response to lower cell numbers fell within the range (shaded area,
0-10%) of cytotoxicity obtained with peritoneal and spleen cells
from nonimmune animals. Intrastromal in situ injections of allogenic cells did not elicit a cellular cytotoxic response, and the percent specific 5 l Cr release fell within the range of the control values
(shaded areas). The results obtained with 10 and 20 X 106 cells 7
and 14 days after immunization were statistically significantly different from one another (P < 0.0001). Similarly, the results obtained with these cell concentrations at 7 and 14 days were statistically significantly different from the results with the lower cell
numbers tested (P < 0.0001).
F344 rats that received 10 X 106 or 20 X 106 Class II+
cells injected into the peritoneum had a peritoneal
and splenic cellular cytotoxic response by 7 days (Fig.
2). Fewer than 10 X 106 allogeneic Class II+ cells did
not elicit a significant response (Fig. 2). The peritoneal and splenic cellular responses differed from each
other in two respects: (1) the peritoneal response
peaked earlier than the splenic response and (2) the
peritoneal cytotoxic response was greater than the
splenic response (Figs. 2A-B).
The F344 corneas injected with either 10 or 20
X 106 W/F Class II+ cells and implanted intraperitoneally elicited both peritoneal and splenic cellular cytotoxic responses measurable at 21 and 28 days (Figs.
3A-B). The cellular cytotoxic response given by the
peritoneal cells was of greater magnitude than that
given by spleen cells, but the peak response occurred
at approximately 21 days in both cases.
Skin Allograft Rejection
The MST of W/F skin grafts on naive F344 recipients was 13.3 ± 1.6 days. The W/F skin grafts subjected to a second-set reaction became vascularized
by day 5, underwent rapid discoloration, shrinkage,
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INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / November 1990
Fig. 3. Cell-mediated cytotoxic responses of F344 rats "grafted"
intraperitoneally with syngeneic corneas carrying 0.01, 0.1, 1.0,
5.0, 10.0, or 20.0 X 106 W/F Class II+ cells. The presence of cytotoxic T cells in the blood, spleen, and peritoneal cavity was assessed. No cytotoxic cells were found in the blood. Ten (circles) and
20 (squares) X 106 cells elicited positive peritoneal (A) or splenic (B)
cytotoxic responses, whereas the response to lower cell numbers fell
within the range (shaded area, 0-10%) of cytotoxicity obtained with
peritoneal or spleen cells from nonimmune animals. The results
obtained with 10 and 20 X 106 cells at 21 and 28 days after immunization were statistically significantly different from one another (P
< 0.005). The results obtained with these cell concentrations at 21
and 28 days were statistically significantly different from the lower
cell numbers tested.
and became dry scabs within 3-5 days thereafter. The
W/F grafts which were rejected in 10 days or less were
considered to have been subjected to a second-set
response.
None of the F344 rats that received allogeneic
Class II+ cells injected into the corneal stroma had a
second-set skin allograft rejection (Table 1). In contrast, F344 recipients of intraperitoneally transplanted syngeneic corneas carrying either 10 X 106 or
20 X 106 allogeneic Class II+ cells rejected W/F strain
skin allografts in an accelerated fashion (Table 1).
Similarly, F344 animals injected with suspensions of
10 X 106 or 20 X 106 allogeneic Class H+ cells also
mounted second-set skin allograft reactions (Table 1).
Discussion
The cellular constituents of the normal cornea primarily express Class I histocompatibility anti1-7,10.11,27,28
Class II histocompatibility antigens
gens.
have restricted expression and occur primarily on the
plasma membranes of B lymphocytes, macrophages,
various dendritic cells, and Langerhans cells of the
skin and conjunctiva.29'30 Under ordinary conditions,
the corneal epithelial, stromal, and endothelial cells
do not express Class II antigens. Also, there are very
few migratory Class II+ cells in the normal cornea.
The normal human cornea and the corneas of
some animal species may contain a few Class II+
Langerhans cells in the periphery.1"13 The number of
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Vol. 31
such cells, however, is vanishingly small; in an extensive analysis, not reported here, it was found that
young adult rats of the strains used in this study have,
on the average, 500 or fewer Class II+ cells in their
corneas (Gebhardt, unpublished).
Rubsamen et al31 and Williams et al7>9 speculate
that cells expressing Class II antigens in the cornea
could be expected to be potent immunogens and that
the larger the number of these cells present in the
cornea, the more likely it is that an immune graft
reaction will occur. The results of this investigation
indicate that while Class II+ cells may elicit local immune reactions in the cornea they do not generate
systemic immunity in the recipient.
It is known that certain anatomic locations in the
body are immunologically privileged and that allografts in these sites are less likely to be subjected to
immunologic attack.32"37 The cornea is one such
site.38 In contrast, there are sites in the body, such as
the peritoneal cavity, in which immune recognition
of alloantigens is facilitated.25'33 In agreement with
this concept, the current study showed that large
numbers of allogeneic Class II+ cells injected directly
Table 1. Survival times of W/F skin allografts on
F344 hosts*
Graft
Number of
cells (XI 0s)
recipient
Normal F344
12
NA
Mean survival
time (days)
13.3 ± 1.6
F344 recipient/intrastromal injection of W/F Class 11+ cells
8
0.01
12.1 ± 1.9
8
0.1
13.4 ±0.8
8
1.0
11.9 ±1.5
8
5.0
13.9 ±2.0
8
10.0
12.6 ±1.7
8
20.0
12.7 ±1.6
F344 recipient/intraperitoneal graft of F344 cornea
W/F Class 11+ cells
8
0.01
8
0.1
8
1.0
8
5.0
8
10.0
8
20.0
containing
12.0 ±1.9
13.7 ±2.1
14.5 ±1.4
11.8 ±1.3
10.1 ±1.1*
8.9 ± 0.8J
F344 recipient/intraperitoneal injection of W/F Class 11+ cells
8
0.01
14.5 ± 1.5
8
0.1
14.3 ±1.5
8
1.0
13.2 ±2.1
8
5.0
12.1 ±1.8
8.7±0.7J
8
10.0
8
20.0
8.4 ± 0.9*
* F344 strain rats were grafted with W/F skin. The grafts were observed
daily after the dressing was removed. Graft rejection was recorded as the day
the graft was judged in viable.
t N, the number of skin allograft recipients in each group.
X Second set graft reactions were characterized by an earlier onset of graft
discoloration, hemostasis, and scab formation. The differences in mean survival time for these grafts, compared to the other groups, were statistically
significant, P < 0.005.
No. 11
2259
CLASS II ANTIGEN-EXPRESSING CELLS / Gebhordr
into the peritoneum or implanted in a syngeneic carrier cornea produce a measurable systemic cellular
and humoral immune response. In situ intrastromal
injection of allogeneic Class II+ cells resulted in no
measurable systemic response, regardless of the number of cells used.
The failure of allogeneic Class II+ cells in situ to
stimulate an immune response is not the result of the
death of the cells injected into the cornea. Previous
studies established that such cells must be viable in
order to mediate immunologic reactions.24 Furthermore, it has been shown that inviable cells are very
poorly antigenic.39
Mononuclear cells, including lymphocytes, migrate into the cornea under various circumstances,
and they emigrate with equal facility.7'91024 Thus, the
difference between the immunogenicity of corneas
bearing allogeneic Class II+ cells placed intraperitoneally or in situ would not appear to be due to the
failure of the cells to emigrate from the cornea in situ
and gain exposure to host immunocompetent cells.
Resident cornea cells may be induced to express
Class II antigens as a consequence of an alteration in
the homeostasis of the tissue. In particular, lymphokines such as gamma interferon may induce Class II
antigen expression in cornea cells.23 The syngeneic
F344 corneas used as carriers for the allogeneic Class
II+ cells in these experiments contained very few resident Class II+ cells. Immunohistochemical staining of
corneas retrieved from the peritoneal cavities of syngeneic recipients revealed that the expression of Class
II antigens is restricted to the injected passenger cells
and that the carrier corneal cells remain Class II antigen negative (Gebhardt, unpublished).
In the model used in this study spleen cells "present" their endogenous Class II antigens directly.
This model mimics the classic T-cell mediated recognition of alloantigens in which foreign major histocompatibility complex molecules act as foreign antigens and elicit an allograft reaction.40
Direct evidence for the theory that Class II+ cells
are a potent source of antigen in the generation of
corneal allograft immune reactions is lacking at the
present time. The results of this study suggest that
Class II+ cells are not an important source of the
antigens that generate an allograft reaction. It may be
that coexpression of Class I and II antigens is necessary to initiate such reactions. It is also possible that
the corneal allograft immune response that is initiated and carried out in the local ocular environment
does not engender systemic cellular and humoral antibody responses; previous work supports this idea.25
Corneal allograft rejection continues to be a major
clinical problem in patients whose graft beds are
highly vascularized before transplantation.27 In such
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grafts, host blood vessels provide a route by which
immunologically competent cells can enter the graft,
recognize graft alloantigens, and mount an immune
attack. These graft reactions may take place regardless of the Class II antigen content of the allograft.
The results of the present investigation indicate that
factors in addition to Class II antigen-expressing cells
may be operative in the initiation of corneal allograft
immune rejection.
Key words: allograft, Class II antigens, cornea, immunity,
inbred rats
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