clinical results of implantation of the chirila keratoprosthesis in rabbits - polymer gel
Ideal artificial cornea (English)KPro)
In terms of surgical treatment, optics, it is very similar to the donor corneal button, and the ability to heal with the host tissue to avoid many complications associated with current clinical use of KPros.
This study was conducted to evaluate long-term clinical outcomes of core and skirt Poly Implants (2-
KPro for animals.
Methods twenty KPros were prepared and implanted in rabbits as a full-thickness corneal substitute and followed up for 21 months.
As a result, 80% of the prosthesis was retained, and the incidence of complications such as cataract, glaucoma and postprosthetic membrane formation was lower, which were often associated with KPro surgery.
Conclusion This type of KPros may provide a therapeutic prospect for poor prognosis in patients using donor materials for penetrating corneal transplants.
However, the improvement of KPro and further animal trials, including the implantation of abnormal cornea, were mandatory before human implants were planned.
Material and method production manufacturing method the manufacturing method has been described in detail before. 14-
The Chirila KPro is made entirely of a flexible gel PHEMA, including an optical transparent core, surrounded by an opaque sponge edge on the periphery, allowing the tissue to grow in (Fig 1).
First, by aggregating HEMA in the presence of 80% water, rim (DVG)
, And the initiator in the mold.
The PHEMA with lower water content then aggregates at the center of the sponge skirt, producing transparent optical elements that lack pores.
In this process, a polymer network that permeates each other (IPN)
Formed between the core and the skirt, a flexible but very strong connection.
When removed from the mold, the thick button is frozen and frozen to the required size (9 mm diameter)
And bending, and high pressure sterilization before use.
10 KPros made without MMA as comonomer will be called "DVG KPros" and implanted in rabbit D1 to D10.
KPros that add MMA will be called "DVG/MMA KPros" and implanted in rabbit M1 to M10.
Download the new tabDownload figureOpen powerpointFigure 1The KPro lira.
Surgical experiments were performed under general anesthesia (
20 half lop Rabbits, weight 2.
5 kg and in compliance with the Australian code of practice for animal care and use for scientific purposes 1990.
The prepared right eye was given 2% before surgery.
The cornea is off.
An upper skin surgery was performed, and a suturing of a corneal ring was performed on limbus.
The eyes were then supported with gentle pressure, while the center of the cornea was marked to three quarters of depth with a 8mm trephine.
Then the eyes go back to orbit and remove the cornea button with scissors.
When open, rinse the front room with 5000 U heparin sodium.
KPro is also rinsed with heparin, placed in the full thickness bed of the receptor, and stitched in place with an interrupted 10/0 nylon.
To prevent scratching or tearing, the KPro is fixed with a lens that introduces tweezers.
The seam line is turned in order to place the knot inside.
No sticky bullet was used.
Then a 360 degree resection of the membrane sac was performed with scissors, and the Tenon SAC and the membrane sac-
Drew a Tenon flap on the cornea
Stitch The KPro surface to the limbus below using 6/0 Vicryl (Fig 2).
1 mg of cortisol was given and 10 mg of late-night penicillin was injected under the eyeball, as well as a local cap ointment.
Download the new tabDownload figureOpen powerpointFigure monthly Peroperative appearance (rabbit D3)
During the formation of the flap
These animals are examined daily to demonstrate complications such as the withdrawal of the membrane sac, leakage of aqueous humor, infection or iritis.
Take amoxicillin ointment once a day for the first month.
2 months after implantation, all animals were sent back to the theater and under general anesthesia, the eye mask flap was opened on the optical core.
Once the flap is opened, they receive an antibiotic ointment with an anti-antibiotic cover and lubricant once a day.
There were three intraocular pressures at 1, 2 and 6 months after surgery (IOP)
Pen XL electronic eye pressure meter (
Ma o & O Company mentor in USA)
Examination under anesthesia
Results clinical observation 16 (80%)
Of the 20 KPros (
10 DVG, 10 DVG/MMA)
It has been successfully retained so far.
In turn, the rabbits D1 to D10 and M1 to M10 were operated to make the rabbits D1 to D10 (
KPros without MMA)
The rabbit M1 to M10 has been followed up for 21 months (
KPros containing MMA)
For 19 months.
Table 1 gives early complications before and after surgery.
In rabbits D1 to D10, KPros made without MMA as co-onomer are considered to be easier to handle and less prone to tearing during stitching placement.
However, the surgical learning curve of the two groups offset this and there was no significant difference in the incidence of surgical complications.
In addition to stretching around the stitching track of the sponge skirt, only one surgical complication was seen --
The expanded lens in rabbit M1 needs to be cut.
View this table: View the inline View pop-up table 1 Peroperative and early (2–4 weeks)
Found after surgery.
In most cases, the immediate chamber can be seen by recovering the flap from the lower rim (AC)
Four rabbits so far (20%)
There are complications that need to kill them.
In the M8 suture of the Rabbit, the tear causes the wound to crack, and the Pinghua Iris blocks the gap.
The animal was killed in 13 days.
The wound was cracked, and the death of rabbit 5 was also caused in 13 days.
Then two more rabbits were lost during follow-up.
D4 at 8 weeks, due to the accidental damage of KPro during flap opening, M1 at 20 weeks.
In the latter, it may be due to trauma, the flap of the cornea is from the lower and local KPro-
Corneal fissure associated with the broken suture.
There were fewer late postoperative complications.
There were no cases of infection, iritis, cataract, postprosthetic membrane formation or KPro extrusion.
Red fundal reflex is noticed in all rabbits (Fig 3)
None of the rabbits had a pupil defect that was relatively passed in.
Table 2 gives intraocular pressure measurements at 1, 2 and 6 months.
Figure 3 appearance after surgery (rabbit D6)at 6 months.
View this table: View the inline View pop-up table 2 postoperative intraocular pressure 1, 2 and 6 months after KPro implantation.
There is no reading in the place marked as NA.
Each number given represents an average of three readings (mm Hg)using a Tono-
The pen on the cornea around the KPro, under the calm of the gram/rompan, the membrane sac-
It is found that the Tenon layer is firmly attached to the skirt, and tissue infiltration will occur on the skirt, but it will not attach to the surface of the optical core. after 2 months of KPro implantation, it is easy to get from
Over time, the opening of the membrane sac was found to contract in the purse style, requiring re-stitching every few weeks.
Mild giant nipple vaginitis (
Mucus is produced in response to the exposed optics, but there is no significant nipple change on the tar plate)
During the follow-up period, about half of the animals had noticed and responded to the increased lubrication.
White deposits were observed in the KPro optical system of five animals, reducing red reflection.
These changes have been noted since about 4 months after implantation and are gradual.
None of the 5 animals had evidence of eye inflammation.
These deposits were subsequently shown to represent calcium deposits.
Histological findings of four eyes
From D4, M1 and M8)
Within 2 to 20 weeks after the operation, the wound was removed due to a split wound or a medical injury and placed in 4% gelatin.
The specimens were examined by an optical microscope to confirm that there was a fiber-vascular extension in the skirt area (Fig 4)
As we have seen in our previous studies18-
The 20 cells that invade the front of the KPro skirt from the Tenon sac seem to help KPro-cornea healing.
In either of these four KPros, there is no evidence that there is calcium deposition in the visual area, but a small amount of calcium is seen in the KPro skirt area of rabbit M1, which is at 20Fig 5).
Download figureOpen in the new tabDownload powerpoint figure 4 tissue slice, 2 months after the implantation of rabbit D4, stained with chlorhexidine blue on the KPro skirt× 400).
After 5 months of implantation of rabbit M1, the tissue slices of the KPro skirt were removed and dyed 2% red;
Spots of calcium can be seen (× 100).
Discussing mechanical factors is critical for KPro's strong enough mechanical ability to perform surgical procedures and suturing and to provide long-term structural integrity for the eye.
After the stitching loses the tensile strength, the cells inside the skirt must have enough long-entry and collagen deposition to ensure permanent safety.
A large number of preliminary work confirmed that the PHEMA sponge contains pores of the size suitable for the growth of cells, and the growth of 19 tissues occurs at 20-
22 it is possible to implant the plate layer of the complete KPro.
23 A pilot study was conducted in which our prototype KPro was placed as a full-thickness implant in 8 rabbit cornea and 18 have now been followed up for 2 years.
This indicates that the prototype's PHEMA sponge skirt was not mechanically strong enough to cause three of the eight rabbits to fail early.
In order to improve the mechanical strength of the skirt, the manufacturing method was changed.
Cross-linking agent divinyl glycol for KPros in this study (DVG)
, Different from the previous use (
Because it's more hydrophilic.
That is to say, it is easier to dissolve in water.
The utility of this cross-linking agent in high water content biological materials has been gathered (1-vinyl-2-pyrrolidinone)
Gel evaluated as a potential glass replacement. 24-
26 mechanical tests conducted using the Syntech 200/M material testing system showed that PHEMA sponges aggregated using DVG as a cross-linking agent showed better tensile properties than sponges formed using EDMA.
27 This is due to improved cross-linking efficiency as DVG is more compatible with the water reaction medium.
Although the mechanical strength is significantly improved, due to the fact that sponge stretching elements often occur during the stitching process, further improvement is needed.
However, this does not necessarily lead to long-term complications, presumably because the holes are sealed after the flap covers and the tissue grows into the sponge skirt.
Biological factors (50%)
One of the KPros of this study was the addition of MMA as a co-onom in a sponge skirt.
28 The addition of MMA is ideal during the invasion of corneal cells and adult fiber cells to improve cell adhesion to sponges.
Known as non-modified PHEMA
Adhesive for mammalian cells, 29 but the addition of MMA or other monomer by polymerization has shown a significant increase in cell adhesion.
3031 in addition, if some clinical cases of KPro are designed to surface along with the corneal epithelial rather than the corneal flap, in the process of allowing epithelial cells to adhere to the surface, HEMA polymers may prove essential.
However, due to the fact that MMA is not soluble in water, the mechanical strength of the DVG cross-linked sponge produced with MMA as a monomer has decreased.
Therefore, another purpose of this study is to evaluate which sponge formulation has the best overall performance
That is to say, whether MMA can lead to poor resistance to surgical treatment increases the risk of early postoperative failure to a certain extent, exceeding any benefit of improving bioceramic.
This is important because we have previously found that the perioperative problem of KPro stitching is the most important predictor of early postoperative failure.
From this study, the DVG cross-linked sponge seems to be much stronger than our previous EDMA formulation, and the addition of MMA does not significantly reduce the intensity.
Therefore, there is no mechanical taboo to add MMA as comonomer to the PHEMA KPro skirt.
Histological examination was performed only in four failures in this study and MMA-was found-
KPro skirts containing rabbit M1 and M8 contain more than KPro skirts lacking MMA for rabbit D4 and d2.
The purpose of this study was not to kill clinically satisfactory animals at a predetermined time interval to assess whether there was a significant difference in the cell length obtained in both KPro skirts (
With or without MMA)
Because this will prevent us from having a long-term follow-up to eliminate the goal of late complications.
Therefore, in order to analyze the nature and time process of biological synthesis in simple cases, a separate study involved another 20 rabbits, 10 of which made comonomer from MMA, tissue pathology assessment was performed at the intended end point sacrifice animals.
32 The most suitable formula allowing the growth of skin cells on skirt and optical surfaces is being evaluated in vitro as MMA is expected to improve epithelial cell adhesion;
This may be related to any future KPro model designed for the implantation of a no-SAC flap in a near-normal eye.
The current method of implantation excludes optical suede because flap skirt adhesion prevents any epithelial cells from entering from the peripheral cornea. Cornea-
The skirt on the edge of the KPro is healing rapidly, preventing the epithelial from growing down into the AC.
After implantation, white calcium deposition was formed in the optical core of a few KPros.
Since such a variable is found, we suspect that this may be due to the manufacturing method.
We found that the Sony extraction method (
Remove toxic monomer)
After polymerization, any initiator remaining in the solution can also be removed, leaving a small space.
These spaces may be oversaturated by calcium ions, resulting in the formation of nidi in the gel for calcification.
Non-histological and in vitro studies
The biological environment supports this hypothesis (Fig 6)
Subsequently, the manufacturing method was changed in order to prevent the production of this space.
Malnutrition calcification is considered to be a less plausible explanation of what is seen in this case, although it is considered to be an explanation for the individual calcification within the PHEMA artificial lens.
33 similar calcium stoves were not seen in extensive clinical trials of PHEMA IOLs34, but other biological materials such as silica gel, especially those associated with infection, were also noted.
35. there is no clinical evidence to suggest that any eye has a calcium stove in the KPro optic disc.
In subsequent studies, calcium deposition was not observed in the optical system as manufacturing techniques improved.
Download the new tabDownload powerpointFigure month (a)
The histological slice of PHEMA gel with pores can be seen (× 400). (b)
After 1 week of placement in an in vitro calcium solution, photos of the entire loading preparation of the PHEMA gel, stained with 2% red and stained with acetone to prove calcium deposition in the gel hole× 400).
We used two rabbits with the most significant turbidity in the optical core (M5 and M10)
To evaluate the convenience of the KPro exchange, because it is reassuring to know that the KPro implant can be repeated.
KPro was replaced with a new 218 days (M5)and 323 days (M10)
After the original KPro is inserted.
This includes stripping the flap from the lower edge and sponge skirt and folding it up to the front. The KPro-
Open the cornea wound with scissors to remove the KPro, and sew a new KPro in place with 10/0 nylon before reforming the flap.
Wash the anterior chamber with heparin when opening, and give postoperative antibiotics as with the initial operation.
Postoperative findings (
Up to 12 months after replacement surgery)
It is satisfactory that the optics show no evidence of calcium deposition.
The calcium spots were found on the histology of the KPro sponge skirt removed from rabbit m1.
The implanted PHEMA sponge and other biological materials usually present a calcium stove. 36-
38 in the long run, progressive calcium stoves are expected to reduce the stability of the implant.
This has been explained by cell death due to undernutrition or cell toxicity, as well as by the calcification of dead cells.
However, we have no evidence of this necrosis or toxicity in an in vitro test.
Calcification may be related to the critical level of calcium concentration in sponge pores.
Alternatively, this may be due to a low level of foreign body reaction.
Occasional macrophages may be associated with PHEMA sponges, and there is no evidence of multiform nuclear infiltration.
The size of the particle sponge material seems to make it easy to devour attack and cells-
Polymer interactions may lead to activation of macrophages.
This finding highlights the importance of further investigation of cell behavior and interaction within porous biological materials and factors affecting the calcium-based focus.
We are using histological techniques, including living organisms.
Over time, death cell analysis and accelerated in vitro calcium system for further study.
Strategies to reduce calcium include changing the physical/chemical properties of the polymer to reduce the crystal nuclear sites, adding "competitive" ions that inhibit crystal growth, or modifying early inflammatory responses.
This is a key area of ongoing research.
Currently, we are adjusting the diameter of the button to 9mm, where the diameter of the central optical element is 7mm.
The final thickness is 0.
5mm, radius of curvature of the front surface 8.
00mm and 8 of the rear surface. 50 mm.
The front and rear surfaces of the optics are treated as refraction surfaces, ignoring the lens thickness, and taking the refractive index of air, hydrated polymer gel and water as 1. 00, 1. 43, and 1.
33 the following approximation is allowed: front surface power = 1. 43 − 1. 00/0. 008 = 53.
Surface Power after 75 DS = 1. 33 − 1. 43/0. 0085 = −11.
76 DS refraction power of KPro optics = 41. 99 DS.
For human patients with crystal eyes, an optical system of about 42 DS is appropriate, and conventional lens extraction is not required for implant surgery.
In patients with crystal-free eyes or patients who need simultaneous cataract surgery, KPro can be used to provide more refractive power.
For example, the front radius is 6.
5mm and rear radius 9.
5mm will give the lens power of 55. 62 DS.
After scanning ultrasound measurements, KPros suitable for clinical use in humans can be selected according to individual requirements and a series of optical power can be provided.
In addition, we recently showed in the rabbit model (
After it's possible-
The implantation adjustment of the optical surface was ablated by excbur laser.
This makes it possible for optical fine-tuning and improves the surface quality, eliminating the fine ridges generated during freezing.
Ridges are expected to reduce the quality of the images and may cause some rabbits to have a slight giant nipple Vaginitis due to our non-complex laboratory lathe.
For KPros implanted in humans, higher manufacturing standards must be adopted.
There is no clinical or histological evidence of complications found clinically (RPM)formation.
This is partly attributed to the PHEMA that forms the rear surface of the KPro.
Adhesive for cells.
However, other common complications of KPro surgery (
Iritis, cataract, retinal detachment, glaucoma)
Neither is this series.
It seems that the risk of all these complications is relatively low.
Invasive surgery is involved.
No postoperative steroids are required.
The absence of corneal melting around the KPro and the resulting extrusion is satisfactory and is considered to reflect the lack of mechanical stress at the implant site, as well as the protective effect of the flap, exclude collagen decomposing enzyme from healed corneaKPro wound.
General intraocular pressure measurement, intraocular pressure assessment (IOP)
In the KPro recipient, due to the rigid KPro component that occupies the cornea, it is usually covered by the eyelid skin, so it is limited to digital estimation.
In this series, using a flexible KPro that only takes part of the large rabbit cornea means "quasi-
The KPro "measurement can be performed with an electronic eye pressure gauge under composure, and the intraocular pressure is more accurately estimated than with a digital evaluation alone.
The pressure of the right eye and the surgical eye can be compared with the pressure of the left eye, and the pressure of the left eye is considered to be the normal value of the eye pressure meter under the anesthesia scheme used.
The mean value of the unoperated left eye measured under these conditions was 14. 4, range 7–23.
The average value of intraocular pressure after right eye surgery was 11. 2, 15. 0, and 13.
6 of D1 to D10 and 15 in rabbits at 1, 2 and 6 months. 1, 24. 2, and 16.
Rabbit M1 to m10 is 1 at 1, 2 and 6 months.
There are four rabbits in total (
M1, M2, M7 and M10)
Of the 17 rabbits whose intraocular pressure was recorded at 2 months, the intraocular pressure was 25mm Hg or higher, of which 3 rabbits (75%)
Tears associated with complicated surgery or stitching.
In contrast, only 5 of the 13 rabbits that were "normal" IOPs for 2 months (38%)
There were stitches related tears in the KPro sponge.
There is no clinical indication of hongitis that higher intraocular pressure measurements may be due to the narrow angle before the tissue grows into the sponge and returns to normal anterior chamber angle.
Therefore, it seems that a clinically useful estimation of intraocular pressure can be obtained with this KPro.
The large diameter of the rabbit cornea allows the use of Tono-to bring the intraocular pressure through the periphery of the cornea to the 9mm implantPen.
Use Tono-read reading through the implant itself
Pen is found to be highly variable.
Therefore, in human patients, unless the size of the KPro is reduced to allow
KPro measurements, this electronic eye pressure gauge, cannot be used.
In order to evaluate the possibility of estimating intraocular pressure using the Schiotz flattening eye pressure gauge, Tono-
Pen and Schiotz eye pressure gauge in rabbit right eye 4 months after operation.
Using the Schiotz eye pressure gauge, 10g weight gives a scale reading of 4 and 5.
The 5g weight reading was 1, and the intraocular pressure estimate of 13mm Hg was given using the fridenwald line chart. The mean Tono-
The amount of Pen reading on the same occasion was 11mm Hg.
This suggests that Chirila KPro allows for clinically useful intraocular pressure estimation by any method, which depends on the relative diameter of the KPro and the cornea.
It is found that increasing lubrication can reduce the speed of regeneration of the flap.
A variant of the KPro that is currently being evaluated has optical elongation designed to prevent excessive growth of the membrane sac.
In most rabbits, it is necessary to trim the flap that is opened on the optics every few weeks.
The flap tissue is not thin enough to achieve reasonable vision without removal from the optical surface.
In some rabbits, it is possible to see the optical response of the KPro surface giant nipple rhinitis to the exposure, which may be aggravated by the surface ridge.
However, inflammation is mild, probably because the rabbit's excellent tear film provides good lubrication, and its low blink rate ensures that the eye film is rarely brought to the optical surface.
In addition, regular regeneration of the membrane sac can prevent continuous exposure.
More advanced manufacturing technologies with the option of light ablation will further reduce this problem.
Given that the PHEMA used in the optical system has a relatively high water content of about 38%, it is presumed that water and dissolved matter will pass through the optical system and evaporate from the exposed surface.
The similarity of the surgical and unsurgical eye intraocular pressure readings suggests that this is similar to the net movement speed of the liquid on the natural cornea.
It may be necessary to lubricate the optical surface exposed by human patients to prevent the gel surface from drying.
We are evaluating potential surface-active drugs such as metho-Poly (MW 750), 40% (v/v)
In a presence that inhibits adhesion through tear membrane protein and pathogens to the exposed optical surface of the chlorhexidine base.
We are also evaluating the provision of a stronger protective layer on the outer surface to make it more resistant to minor trauma.
We envision two very different groups of potential KPro recipients;
First of all, those severe eye abnormalities, including corneal marginal damage and dryness, KPro provides the only hope for visual recovery.
In most of these patients, the epithelial cells were not maintained and did not
It is appropriate to have an upper skin vision exposed through the mucosa or skin.
However, if a real "artificial donor button" can be designed to maintain a stable multi-layer epithelial cell, it may eventually have a wider application, especially in countries where organizations in the eye socket are in short supply.
We are now working on ways to achieve PHEMA skinning in the core and skirt areas.
Corneal epithelial growth on PHEMA polymer gel and modified sponge in vitro (Fig 7)
However, long-term epithelial maintenance in vivo, which depends on the interaction of the mesenchymal, may be difficult to achieve in optics.
Optical design may need to be modified.
Download figureOpen in the new tabDownload powerpoint Figure 7, grow rabbit corneal epithelial cells from the donor button to an in vitro PHEMA sponge specimen (a)
Part dyed with toluene-based Blue (×200)and (b)surface view (
The arrow indicates the edge of the forward cell).
Conclusion in a relatively simple, repeatable process, Chirila KPro can be implanted in a manner similar to PK.
When covered with a flap opened 2 months after surgery, the incidence of complications in this animal study was advantageous compared to other KPro models;
After 21 months of follow-up, cataract, iritis, postprosthetic membrane formation, glaucoma, retinal detachment and extrusion were not found.
Intraocular pressure can be estimated by intraocular pressure.
This PHEMA KPro, when DVG is used as a cross-linking agent, and whether MMA is added to the sponge as a co-monomer, is less prone to stitching-related tears than our earlier prototypes.
Even so, three. 15%)
KPros failed due to early wound rupture due to a sponge tear or presumed eyeball trauma, one failed due to a medical injury at 8 weeks.
Sponge tears may also lead to temporary changes in AC and elevated intraocular pressure in the early stage of surgery.
Subjectively, the various KPros differ greatly in the features of surgical treatment, and it is obviously important to improve the quality control of KPro manufacturing.
This study identifies specific areas for further research.
In particular, it is of interest to the cellular reactions and interactions within the sponge skirt against the porous particle PHEMA.
Animal trials continue, with special emphasis on determining the incidence of long-term complications and improving the optical properties and surface quality of KPro.
Pre-evaluation of KPro in animal eyes
Current pathological examinations are also under way.
The author gratefully thanks Annette DAOs and the staff for their anesthesia help and Chris Barry for their photographic help.
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