Investigative Ophthalmology & Visual Science, Vol. 31, No. 1, January 1990
Copyright © Association for Research in Vision and Ophthalmology
Stimulation of Cell Division by Argon and Nd:YAG
Loser Trabeculoplasty in Cynomolgus Monkeys
David K. Dueker,* Michael Norberg,* Douglas H. Johnson,f Renee C. Tschumper,-(- and Lynette Feeney-Burns"
Although laser treatment of the trabecular meshwork is the most common form of surgery for glaucoma, the tissue response to this therapy is still incompletely understood. We applied argon or
Nd:YAG laser to the trabecular meshwork of six monkeys. Cell division was identified by injecting
tritiated thymidine into the anterior chamber 24 hr after laser application. Autoradiography of tissue
sections revealed significantly more labelled cells in eyes treated with laser than in the untreated
controls. In addition, cells in neighboring tissues such as iris, ciliary body and sclera showed labelling
in association with laser application. Furthermore, comparison of argon-induced lesions with those
caused by pulsed Nd.YAG suggests that there are quantitative and qualitative differences in the
response of trabecular meshwork and surrounding tissues to these two forms of laser energy. Invest
Ophthalmol Vis Sci 31:115-124,1990
The trabecular meshwork (TM) constitutes the
main pathway for conventional aqueous outflow.
Some intrinsic or imposed alteration in the function
of this tissue is generally considered to be the cause
for the open-angle types of glaucoma. Many of the
open-angle glaucomas can be successfully treated by
applying laser energy to the trabecular meshwork, a
procedure known as "trabeculoplasty," although it is
not known that a physical reshaping of the tissue is
responsible for the beneficial effects of the treatment.
Wise and Witter1 proposed that laser treatment
tightens an abnormally lax meshwork and thereby
improves the facility of aqueous outflow. The most
likely basis for such an effect would be heat-induced
collagen shrinkage; the trabecular meshwork, like
other collagen containing tissues, has been found to
shrink with heating.2 But Van Buskirk et al3 found no
improvement in outflow facility in enucleated
human eyes treated with laser trabeculoplasty, even
though other methods for physically increasing tension in the meshwork do improve the outflow facility
of enucleated eyes.4 Possibly the laser treatment
would have been effective if the eyes had been glaucomatous5; alternatively, it is possible that the laser
produces its effect through a more complex process in
which living tissues respond to laser wounds by altering their function.3
The current study was designed to extend our
knowledge of the response of trabecular tissue to laser
therapy through a study of acute effects on trabecular
cell division in vivo. Two clinical forms of laser therapy—argon laser trabeculoplasty (ALT) and
Nd:YAG trabeculoplasty—were studied in cynomolgus monkeys.
Materials and Methods
Animals
Six adult male or female cynomolgus (Macacafascicularis) monkeys were used in this study. All procedures involving the animals were in accordance with
the ARVO Resolution on the Use of Animals in Research.
The animals were anesthetized with intramuscular
ketamine (Ketaset, Bristol Lab., Syracuse, NY) 10
mg/kg and xylazine (Rompun, Mobay Corp., Shawnee, KS) 0.5 mg/kg for preoperative evaluation 24 hr
prior to laser therapy. A complete examination of the
eye was carried out including direct ophthalmoscopy,
slit-lamp biomicroscopy, applanation tonometry and
gonioscopy to ensure there were no preexisting abnormalities.
From the *Mason Institute of Ophthalmology, University of
Missouri-Columbia, Columbia, Missouri, and fThe Mayo Clinic,
Rochester, Minnesota.
Supported by NEI Grants EY-05792 and EY-07065, and an
unrestricted grant from Research to Prevent Blindness, Inc.
Submitted for publication: January 23, 1989; accepted May 16,
1989.
Reprint requests: David K. Dueker, MD, One Hospital Drive,
Columbia, MO 65212.
Laser Treatments
Laser treatments were administered using either an
argon blue-green laser (Synemed Inc., Berkeley, CA)
or a Nd:YAG pulsed laser (AMO, Irvine, CA) with
the animals anesthetized in the same manner. A
Kaufman/Wallow gonioscopy lens (Ocular Instru-
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INVESTIGATIVE OPHTHALMOLOGY 6 VISUAL SCIENCE / January 1990
ments, Inc., Bellevue, WA) was used to deliver the
applications in all cases. One eye of each animal was
randomly selected for treatment with either argon
laser (three animals) or Nd:YAG laser (three animals). The opposite eye was an untreated control and
was subjected only to gonioscopy.
The protocol for argon laser trabeculoplasty (ALT)
included the following specifications: spot size, 50
fim; exposure, 0.1 sec; applications, 50 spots evenly
distributed over 360° of the trabecular circumference. The power was adjusted to produce visible
blanching of the tissue with or without formation of a
single small gas bubble. Applications were made in
one of two locations. Following the description of
Melamed et al6'7 some spots were centered on the
pigmented band. But our histologic studies on other
similar animals showed that the pigmented band in
these eyes is somewhat posterior to its relative position in human eyes. We therefore also placed some
burns more anteriorly, in meshwork more directly
adjacent to Schlemm's canal.
The Nd:YAG laser trabeculoplasty protocol was as
follows: spot size, 20 ixm; exposure, 14 nanoseconds;
applications, eight to ten spots spaced approximately
200 nm apart in the superior trabecular meshwork;
power, 4 mJ. We used a power setting somewhat
lower than that used for Nd:YAG trabeculoplasty in
human beings8 because preliminary work had demonstrated that these power levels consistently produced an open channel through trabecular meshwork
connecting anterior chamber with Schlemm's canal.
The applications were placed just anterior to the dark
pigment band so that they would impact directly over
Schlemm's canal. The superior chamber angle was
treated so that any reflux bleeding would not settle on
the lesions.
3
H-Thymidine Administration
Twenty-four hours after a laser treatment, the animal was anesthetized and clinically examined. Both
treated and control eyes were then infused with 3Hthymidine (NET-027Z, Specific Activity 50 mCi/
mmole, New England Nuclear, Boston, MA); 25 /id
in a 50 ix\ volume were given to each eye. The solution was delivered to the anterior chamber via a 27gauge lymphduct cannulator (North American Instrument Corp., Hudson Falls, NY) inserted through
the peripheral cornea. A motorized syringe was used
to deliver the fluid slowly (about 8 min per eye) to
avoid extremes of pressure. After 5 hr the animal was
again anesthetized and given a second identical infusion. The dose of 3H-thymidine and the manner of
delivery were based on labelling studies of trabecular
cells in cat and rabbit eyes.14
Vol. 31
Twenty-four hours after the first thymidine infusion, the animal was anesthetized with ketamine and
then given a lethal intracardiac dose of pentobarbital
(Fort Dodge, Fort Dodge, IA). The eyes were rapidly
enucleated and wrapped loosely in moist gauze. The
eyes were then perfused with fixative at 15 mm Hg
via a 23-gauge needle in the anterior chamber. The
fixative was 2% glutaraldehyde, 1% formaldehyde,
buffered with 0.1 M sodium cacodylate to 7.2 pH.
After 30 min, the entire globe, still continuously perfused at 15 mm Hg, was immersed in the fixative
solution. After 20-24 hr, the perfusion needle was
withdrawn and the eye was placed in 0.1 M cacodylate buffer.
Dissection and Embedment
Each eye was positioned under an operating microscope, and the 12:00 limbus was marked for reference. Next a 5 to 6 mm central corneal button was
dissected free and removed. The anterior chamber
angle of lasered eyes was examined under high magnification (X40) to locate the laser-treated sites. A 1
mm wedge of chamber angle containing an identifiable burn was then dissected free from the eye. This
wedge of tissue also contained peripheral cornea,
sclera overlying the burn, peripheral iris and ciliary
body.
At least one wedge was removed from each control
eye; for two control eyes a wedge was removed from
each quadrant. This resulted in 12 tissue blocks from
the control eyes. Seven blocks containing acute
Nd:YAG laser lesions were prepared: four from one
eye, two from another, and one from the third. Ten
blocks containing argon laser lesions were prepared:
four from each of two eyes, and two from the third.
Prior to embedment, the blocks containing laser lesions were trimmed so that the edge destined for initial sectioning was already one-third to one-half way
into a visible burn.
After final trimming of the tissue, the blocks were
post-fixed in 2% buffered osmium tetroxide, dehydrated in graded ethanols, and embedded in EPOX
812 epoxy resin (E. Fullam, Inc., Schenectady, NY).
Serial 1 ixm sections were cut from each block and
mounted sequentially in groups of three to six sections on three or four glass slides. Our standard sections were all made with the plane of section carefully
oriented perpendicular to the canal of Schlemm. In
three instances the plane of section was oriented parallel to Schlemm's canal and the plane of the iris. The
slides with attached sections were coated with Ilford
K.5 Nuclear Track Emulsion under safelight conditions, placed in a light-tight container and stored
under refrigeration for 1 week. The exposed slides
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CELL DIVISION AFTER LASER TRADECULOPLASTY / Dueker er ol
were developed in Kodak D-19, fixed, rinsed and
dried, and then stained with 1% toluidine blue buffered with 1% boric acid for 60 sec. The stained slides
were rinsed, dried and coverslipped.
The identifying markers on the slides were covered,
so that tabulations were made without knowledge of
the treatment history. Independent readings confirmed the reproducibility of the counts. The only
source of potential confusion was melanin. But these
granules, being of different size and color, located in
the cytoplasm, and in a different focal plane from the
silver grains, were readily distinguishable from silver
grains on top of the section over the nucleus. A formal counting rule adopted for the initial study required a minimum grain count of ten over the cell
nucleus to consider a cell labelled. Later a smaller
sample was reassessed using a counting rule of 20
grains.
Labelled trabecular cells were recorded as being
juxtacanalicular, or anterior (to Schlemm's canal),
middle (adjacent to Schlemm's canal) or posterior (to
Schlemm's canal) corneoscleral TM. Cells were
counted as corneoscleral TM cells if they were in a
normal position on a trabecular beam, that is, flattened against the underlying beam and contiguous
with neighboring TM cells. If a cell were rounded or if
it were not in a typical TM cell relationship to a
trabecular beam, it was not counted. Although juxtacanalicular cells do not invest trabecular beams, they
were identified on the basis of their morphology, their
relationship to surrounding tissues, and, in some instances, a typical functional characteristic—giant
vacuoles. Inflammatory cells, which were found in
both treated and control eyes, were distinguished
from TM cells on the basis of their rounded shape,
characteristic cell surface or nuclear morphology, and
their limited physical contact with the meshwork,
and were not counted.
A mitotic index was calculated for each section by
dividing the total number of labelled trabecular cells
by the total number of trabecular cells present. Since
the total number of trabecular cells did not vary
greatly in these sections, we generally determined the
total cell count on one section per slide, and used that
count for the determination of all sections on the
slide. A mean mitotic index was calculated for each
tissue block and for each eye in the study. Because the
number of animals was small, and since we did not
know whether the data were normally distributed, we
used a nonparametric statistical test, the Wilcoxon
Signed Rank Test.
The primary interest of this study was the behavior
of trabecular meshwork cells after exposure to argon
or Nd:YAG laser. The preparation of the specimens
was therefore designed to give thorough and consis-
117
tent samples of this tissue. Adjacent tissues were of
course present in all sections, but the extent of their
representation depended on the dissection. Cornea,
sclera, ciliary body and iris, to the degree they were
present, were examined in every section, and any labelled cells found in these cells were recorded.
Results
Control eyes showed very little trabecular cell labelling. Eight of the 12 control tissue specimens contained no labelled TM cells on any of the sections cut
from these blocks. The remaining four control tissue
blocks showed infrequent labelling: only 14 of 61
sections from these blocks had one or more labelled
cells. And, when labelling was present in control sections, it was sparse. Generally only one cell was labelled; the exceptions were two sections, each with
two labelled cells.
Cell labelling was increased substantially in association with both argon (Fig. 1) and Nd:YAG (Fig. 2)
laser lesions. Every argon laser tissue block (n = 10)
produced some sections with labelled cells (Fig. 3);
altogether 62% of sections cut from these blocks had
at least one labelled cell. With argon laser-treated tissue, when labelling was present in a cross-section of
the TM, the average number of labelled cells was
2.86. All tissue blocks with Nd:YAG laser lesions (n
= 7) also contained labelled cells (Figs. 4, 5). Of the
total of 130 sections examined from these blocks, 113
(87%) contained labelled cells. In sections with labelling, the average number of labelled cells was 7.95.
Preferential labelling of the anterior TM9 was not evident in either group.
A mitotic index for each section was calculated,
and these data are presented in Table 1 (A and B). A
graphic display of the distribution of mitotic indices
for each section of each eye is presented in Figure 6.
For statistical comparison, the mean mitotic index
for each laser-treated eye was compared with the
mean index for the paired control. The mean mitotic
index was significantly different (P = 0.036), laser
greater than control.
Since individual specimens from a single eye are
subject to a common environment, considering all
specimens from an eye in aggregate (as above)
seemed the most appropriate method of comparison.
However, some observations also suggested that each
laser lesion may represent an independent response.
For example, if we continued to cut sections from
blocks with laser lesions, we eventually reached tissue
well away from the burn center. These adjacent tissues were less actively labelled (or even unlabelled)
when compared to the sections taken from the center
of the lesion. In the three blocks sectioned parallel to
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INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / January 1990
Vol. 31
Fig. 1. Cross-section of a
typical argon laser burn 24
hr after treatment. A fibrin
plug (F) is enmeshed in the
inner trabeculae. AC = anterior chamber. Bar = 50
Schlemm's canal, we found that labelled cells were
closely associated with the burn center. Finally, there
is a fairly wide variability in laser lesion labelling.
While some dramatically high mitotic indices were
observed (eg, Table IB, Observation 9), three of the
anterior argon laser burns (Table IB, Observations 8,
10, 13) produced indices no higher than the most
active control tissues—even though other lesions in
the same eyes were actively labelled.
Since these observations suggested some autonomy
of individual lesions within eyes, we also subjected
the data to statistical analysis considering each specimen as an independent observation. The mean mitotic index from each of 12 control blocks was paired
AC
Fig. 2. A typical Nd:YAG
laser burn 24 hr after treatment. A crater-like area
marks the region where
trabeculae have been disrupted and dislodged. AC
= anterior chamber, SC
= Schlemm's canal. Bar
= 200 urn.
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No 1
119
CELL DIVISION AFTER LASER TRADECULOPLASTY / Dueker er ol
r
Fig. 3. An argon laser
burn with dividing cells
(arrows) in the corneoscleral
meshwork. SC = Schlemm's
canal. Bar = 100 iim.
with a laser block from the opposite eye. When there
were more laser blocks than control blocks for a given
eye pair, the least active laser block was used to be
conservative. The laser and control treatment groups
were significantly different (P = 0.0025). In addition,
each type of laser burn was compared to tissues from
its paired control eye in the same manner. This analy-
sis showed that both lasers caused significantly
greater labelling than controls (both P = 0.036). Finally, the two lasers were compared, again using the
most conservative blocks, and the Nd:YAG showed
significantly more labeling than the argon (Wilcoxon
Rank Sum, P = 0.0163). A comparison of the two
lasers based on randomly selected blocks (rather than
Fig. 4. Juxtacanalicular
tissue from a Nd:YAG laser
burn showing giant vacuoles in juxtacanalicular cells
with labelled nuclei as well
as a labelled corneoscleral
meshwork cell, (arrows)
Scleralfibroblastswith label
are seen near Schlemm's
canal (•). SC = Schlemm's
canal. Bar = 100 *im.
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INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE / January 1990
Vol. 31
sc
Fig. 5. Nd:YAG laser lesion. Labelled cells are seen on the outer wall of Schlemm's canal (arrows) as well as in the trabecular meshwork. SC
Schlemm's canal. Bar = 100 nm.
least active blocks) showed a less significant difference {P = 0.0782).
Examination of the tissues adjacent to the trabecuTable 1. Specimens examined
Animal
Eye
Block
Treatment
Average
mitotic
index
A
7
7
7
7
8
8
8
OD
CD
OD
OD
OS
OS
OS
8
OS
10
11
12
14
OD
OD
OS
i
OS
8
10
OS
OS
OS
OS
OD
OD
OD
OD
OS
It
OS
tl
12
12
12
12
14
14
OS
OD
OD
OD
OD
OD
OD
77
7
7
8
8
8
I
II
III
IV
I
II
III
IV
I
I
I
I
I
II
III
IV
I
II
III
IV
I
I
II
I
II
III
IV
I
II
CTRL
CTRL
CTRL
CTRL
CTRL
CTRL
CTRL
CTRL
CTRL
CTRL
CTRL
CTRL
0.000
0.000
0.000
0.000
0.000
0.000
0.004
0.001
0.003
0.000
0.002
0.000
YAG
YAG
YAG
YAG
0.031
0.012
0.025
0.033
0.013
0.025
0.011
0.003
0.197
0.001
0.098
ARG(P)
ARG(P)
ARG(P)
ARG(A)
YAG
ARG(A)
ARG(P)
ARG(A)
ARG(A)
ARG(P)
ARG(P)
YAG
YAG
0.062
0.003
0.032
0.024
0.088
0.051
Multiple sections (average 12) were cut from each block of tissue. A mitotic index was calculated for each section; the average of those indices for
each block is presented in the righthand column of Table 1. (Table IA,
Control specimens; Table IB, Laser-treated Specimens.) The more anterior
argon burn placement is represented as ARG(A), the more posterior,
ARG(P). Comparison of the mean mitotic index for control eyes vs. lasertreated eyes revealed a significant difference between groups {P = 0.036).
lar meshwork also revealed different patterns of thymidine labelling depending on the treatment. The
corneal epithelium was a regular site for labelled cells
in all specimens regardless of treatment. Control tissues showed a near total absence of labelling in adjacent tissues other than corneal epithelium. In 136
sections cut from 12 different blocks of untreated
control tissue, only one labelled cell—a scleral fibroblast—was found in an adjacent tissue other than
corneal epithelium.
By contrast, the sclera external to a Nd:YAG laser
lesion commonly showed labelling of scleral fibroblasts (Figs. 4, 7); six of seven Nd:YAG blocks contained labelled cells with a rather uniform distribution of label—61 of 95 sections. Some sections were
densely labelled, with as many as ten positive cells.
Labelling of scleral fibroblasts in association with
argon lesions was less frequent (four of ten blocks)
and less dense.
Cell labelling in the iris and ciliary body adjacent to
argon lesions was considerably more pronounced
than in either controls or Nd:YAG lesions. No control tissues had labelling in the iris, and only one
labelled iris cell (a vascular endothelial cell) was
found in the Nd:YAG specimens. In contrast, all but
one of the ten argon laser blocks showed labelling of
iris vascular endothelial cells (Fig. 8); the labelling
was quite uniform (89 of 134 sections) and often
dense (several sections had more than ten labelled
cells).
Labelling of cells in the ciliary body was almost
exclusively associated with argon laser lesions. Control tissues had no ciliary body labelling, while three
sections from one Nd:YAG block had one labeled
cell each. However, five of the ten argon laser blocks
(38 of 67 sections) did show some labelling in the
ciliary body (Fig. 9), with as many as 14 cells labelled
in a section. The labelled cells appeared to be stromal
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CELL DIVISION AFTER LASER TRADECULOPLASTY / Dueker er ol
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121
SECTIONS
Fig. 6. Distribution of mitotic indices. The six bars at
each mitotic index range
represent the six animals in
the experiment, with control eyes plotted in the
upper graph immediately
above their paired lasertreated eyes in the lower
graph. The treatment history for the six animals is,
from left to right: YAG,
argon, YAG, argon, argon,
YAG. Thus, for example,
the two animals with no
sections showing a zero mitotic index (bars 3 and 6 absent, lower graph, zero position), are from Nd:YAGtreated eyes. Please note the
expansion of the mitotic
index scale at the rightmost
two positions.
0.01
0.02
0.03
0.04
0.05
0.06
0.07
0.08
0.09
0.1
0.2
0.3
0.2
0.3
MITOTIC INDEX
LASER TREATED
SECTIONS
10 _
0.01
0.02
0.03
0.04
0.05
0.06
0.07
0.08
0.09
MITOTIC INDEX
cells, and the most frequent location was somewhat
posterior to the scleral spur.
Finally, thymidine labelling was reassessed using a
counting rule of 20 instead often for reasons detailed
in the discussion. The proportion of cells scored as
labelled diminished in both the control (by 21%) and
lasered (by 39%) groups, however the statistical
difference between groups remained significant
(P = 0.036).
Discussion
Laser trabeculoplasty using either argon or
Nd:YAG laser stimulates uptake of tritiated thymi-
dine and presumed cell replication. This method is
widely used to mark dividing cells, though it is not
absolutely specific, since DNA repair can occur in
nondividing cells. We believe the dense labelling
found in this study is most adequately explained by
thymidine uptake in association with cell replication.
For our 1 /urn thick sections we selected, initially, a
counting rule often or more grains over a nucleus as
evidence of S phase DNA synthesis. Bylsma et al10
selected 20 grains or more for their 3 ^m thick sections, which calculates to 13 grains for a 1 ^m thick
section." Nuss et al,12 in a study specifically designed
to distinguish between scheduled and unscheduled
DNA synthesis, considered 15 grains over nuclei in
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INVESTIGATIVE OPHTHALMOLOGY b VISUAL SCIENCE / January 1990
Vol. 01
Fig. 7. Nd:YAG laser lesion. Two scleral fibroblasts just anterior to Schlemm's canal are labelled (arrows). S = sclera. Bar = 100 ^m.
2-3 ^m thick sections to indicate S phase DNA synthesis; this correlates with 10 grains in 1 jum thick
sections, and supports our use of this counting rule.
Nonetheless, following the initial analyses of the data
we wished to determine the effect produced by a
counting rule of 20. Using a small sample of representative sections it was found that this 2-fold change in
the minimum grain count did not change the P value
of the difference between the labeled nuclei of control
and lasered tissue.
In our study, argon laser treatment caused cell division to be maximal in the immediate vicinity of the
laser lesion. In a similar study in organ culture, Acott
et al9 also found stimulation of cell division by ALT,
but the distribution was more widespread—involving
both treated and untreated areas equally. They note
that such a widespread effect is consistent with a medium-born regulatory signal for cell replication. The
focal activity we found in our study is not inconsistent with this hypothesis. If the lesions themselves
produce a diffusible mitotic stimulant(s), their influence would be widely and evenly distributed in an
organ culture system. In vivo, on the other hand,
factors produced locally in the TM would be carried
out of the eye by the normal flow of aqueous humor,
allowing little impact on more distal tissues.
We noted no preferential labelling of the anterior
TM cells. We found all TM cells capable of incorporating label, and the distribution was highly variable.
In organ culture, the initial post-laser labelling was
concentrated in the anterior TM.9 This difference
may reflect cell migration, species variation or other
dissimilarities in the two experiments.
One of the advantages of in vitro culture is the
ability to control the environment and thereby eliminate some sources of variability, but clinical treatments such as ALT may trigger any of a wide range of
responses operative in the living organism. And while
several laboratories have demonstrated trabecular
cell replication in culture,1013 few have addressed the
question in vivo.14 For this reason, a further careful
comparison of TM response to laser in culture versus
in vivo should help clarify which aspects of the response are controlled by neuronal or secondary phys-
Fig. 8. Iris vessel in the vicinity of an argon laser burn showing labelling of vascular endothelial cells. Bar = 50 pm.
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CELL DIVISION AFTER LASER TRADECULOPLASTY / Dueker er ol
•V
123
1
Fig. 9. Ciliary body posterior to an argon laser burn on the trabecular meshwork; several stromal cells are labelled (arrows). Bar = 50 ^m.
iological factors as opposed to strictly local influences.
The pulsed Nd:YAG laser also produced focal
stimulation of trabecular cell division, at least as potently as the argon laser. Organ culture studies using
Nd:YAG have not been reported, so comparison is
not possible with our labelling results.
Qualitative comparisons of adjacent cornea, sclera,
iris and ciliary body were possible, although only a
variable amount of these tissues were present. The
corneal epithelium15 was a regular site of labelled cells
consistent with its rapid replication rate. Other tissues
differed in their labelling, depending upon the treatment the eye received. A strong tendency for labelling
of iris vascular endothelium in argon-treated tissues
raises the question of endothelial proliferation contributing to the breakdown of the blood-aqueous
barrier in ALT.16 A potential concern raised by this
finding is the possibility of long lasting change in the
iris vessels making the eye more prone to future
blood-aqueous barrier breakdown and/or vasoproliferation. Labelling of ciliary body stromal cells was a
prominent feature of some argon-treated eyes, and
was rare or nonexistent in the other groups. The consequences of such changes are unclear, but their exclusive association with the argon lesion underscores
the variable responses of these tissues to differing
stimuli.
The pulsed Nd:YAG laser produced an impressive
response in the trabecular meshwork cells. In addition, frequent labelling of scleral fibroblasts external
to the impact was observed. Since very few scleral
fibroblasts were labelled in the other groups, this response seems to be specific for the Nd:YAG lesion,
and may relate to the greater physical disruption
caused by this instrument. It seems that the pulsed
laser stimulated division almost entirely in its direct
path: the primary target, trabecular meshwork, and
the sclera behind it. The thermal effect of the argon
laser, on the other hand, stimulates the direct target,
the trabecular meshwork, but also stimulates iris and
ciliary body indirectly. The potential mechanisms for
indirect stimulation include heat transfer, neuronal
mechanisms, or local hormones or growth factors.
Our studies show that laser stimulation of trabecular cell replication occurs in vivo, and that the stimulus may be either an argon thermal lesion or a pulsed
Nd: YAG lesion. Considering the decrease in trabecular cell density reported with age and glaucoma,17"19
this response to laser application may be an important factor in producing the benefits of laser trabeculoplasty. Laser stimulation may produce a net increase in trabecular cellularity, and it may even select
a population which is inherently more healthy, as
Bylsma et al have suggested.10 On the other hand,
traoecular cell proliferation might take the form of a
membranous covering of the meshwork with potentially adverse effects on aqueous outflow.20 Further
studies of these replicating cells, especially regarding
long-term functional and/or morphologic characteristics, should reveal what role such cellular alterations
play in the mechanism of laser trabeculoplasty.
Key words: autoradiography, laser trabeculoplasty, trabecular meshwork, tritiated thymidine, wound healing
Acknowledgments
Statistical consultation was provided by John C. Hewett,
PhD and Sharon Anderson, MA, Department of Biostatistics, University of Missouri, School of Medicine, Columbia,
Missouri. Secretarial assistance was provided by Theresa
Stathem.
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