RU2824574C1 - Method for preparation of three corneal grafts from one donor cornea - Google Patents

Abstract

FIELD: medicine; ophthalmology.

SUBSTANCE: three corneal grafts are prepared from one donor cornea. Femtosecond laser is used to perform a first circular section of the corneal stroma with diameter of 8 mm, which is calculated so that the residual thickness of the cornea from endothelium is 125 mcm. Second section is in the lamellar plane at depth of 125 mcm from the endothelium, with diameter of 10 mm. Third section is circular and is made from depth of 125 mcm upwards towards the endothelium, with diameter of 10 mm. Then, a radial cut is made in a vertical plane, which divides the corneal graft into two equal parts. Further, the endothelial grafts are separated with a blunt spatula, after which the anterior graft is separated from the corneoscleral disc of the donor, which is then subjected to a corneal collagen crosslinking procedure with preliminary de-epithelization.

EFFECT: method provides obtaining three quality uniform grafts, enables to increase the number of transplants for patients with endothelial corneal dystrophies and keratoconus, restore visual rehabilitation of patients with keratoconus due to cross-linking of anterior graft after dissection of donor cornea and provides absence of risk of secondary transplant ectasia.

1 cl, 3 ex

Description

The invention relates to the field of medicine, namely to ophthalmology, and can be used for preparing three corneal transplants from one donor cornea for performing posterior lamellar keratoplasty (PLK) and deep anterior lamellar keratoplasty (DALC) using a femtosecond laser.

For many years, penetrating keratoplasty was the gold standard in the treatment of corneal diseases. However, today, methods have been developed for treating diseases of various corneal layers by replacing them. For example, in case of endothelial corneal dystrophy, a posterior graft with a thickness of 80-150 μm is used, including the endothelium, Descemet's membrane (DM) and a thin layer of corneal stroma. [Preliminary results of posterior lamellar keratoplasty using a high-energy femtosecond laser for harvesting an ultra-thin donor corneal graft Kuzmichev K.N., Pashtaev A.N., Izmailova S.B., Malyugin B.E. Modern technologies in ophthalmology. 2018, No. 4. P. 140-142]. In anterior lamellar keratoplasty, a graft is used, which is the epithelium, Bowman's membrane, anterior and middle corneal stroma. Anterior graft can be used for various types of keratectasia, in particular for keratoconus stage 3-4, which reduces visual function and leads to thinning of the cornea and the risk of its perforation. [Penetrating and anterior lamellar keratoplasty Bikbov M.M., Usubov E.L., Zainetdinov A.F. Point of view. East - West. 2019 No. 1 P. 89-92.

Mechanical harvesting of grafts carries a risk of rejecting donor material. Femtosecond laser assisted corneal surgery has largely solved this problem. Also, modern laser systems make it possible to use lower laser energy but a denser cutting frequency, which mitigates the negative impact on endothelial cells, reduces their loss and reduces the incidence of haze at the donor-recipient interface in the postoperative period. [Shilova NF, Livny E, Anisimova NS, Antonova OP, Malyugin BE. Refractive outcomes following cataract combined with lamellar keratoplasty: femtosecond-DSEK versus microkeratome-DSAEK. Int Ophthalmol. 2021 Feb; 41(2):639-647. doi: 10.1007/s10792-020-01619-7. Epub 2020 Oct 22. PMID: 33090312]. However, the issue of donor material shortage is acute. The current stage of socio-economic development of society, the experience of previous years demonstrating instability of access to donor tissue for legislative and legal reasons, an acute shortage of donor tissue, despite the "presumption of unsolicited consent", and other factors show that it is possible to influence the provision of a practicing clinician with donor tissue through its rational use [Rational use of donor material: own ten-year experience, possible development paths and literature data / O.G. Oganesyan, S.S. Yakovleva, M.P. Harlampidi, A.A. Grdikanyan // Russian Medical Journal. - 2016. - Vol. 22, No. 4. - P.193-197. - DOI 10.18821/0869-2106-2016-22-4-193-197. - EDN WDGOZT].

The closest analogue is the method of rehabilitation of patients with endothelial dystrophies of the cornea using one donor cornea for two recipients (patent No. 2801493), during which the cornea ready for work in the artificial anterior chamber (AAC) is placed under an applanation lens connected to a femtosecond laser Ziemer FEMTO LDV Z8. After ensuring centering and complete applanation of the cornea, the first cut is made in the lamellar plane at a depth of 125 μm, the second is circular, going from a depth of 125 μm upwards of a given diameter - 10 mm. Then a radial cut is made in the vertical plane, which divides the donor cornea into two equal parts. The resulting corneal transplant, after the femtosecond laser has finished working in the IPC, is placed under an operating microscope and the transplant is separated from the donor cornea using a blunt spatula, resulting in a finished transplant of the posterior layers of the cornea.

The disadvantage of this method, despite the high technological nature of the process, is the lack of involvement in the process of preparing the transplant of the anterior layers of the donor cornea for the preparation of an additional transplant that can be used to treat diseases such as keratoconus, which is an important aspect due to the high demand for donor material.

The objective of the invention is to develop a method for preparing three corneal transplants from one donor cornea.

The technical result is an increase in the number of rehabilitated patients while using a smaller amount of donor tissue.

The technical result is achieved by forming a donor ultra-thin endothelial transplant with a diameter of 10 mm, divided into two equal parts, as well as obtaining a transplant for the GPPK with a diameter of 8 mm at a time using a femtosecond laser. The obtained anterior transplant is then subjected to the corneal collagen crosslinking procedure, which prevents the risks of secondary ectasia when transplanting material to recipients with keratoconus.

The method is implemented as follows. The donor cornea, preserved, for example, in a solution for storing corneas manufactured by Scientific and Experimental Production Microsurgery of the Eye LLC (Russia, TU 9393-013-29039336-2007, registration certificate No. FSR2010106650), is mounted on the IPC with the endothelium facing upwards, which is connected to an infusion system inside which a pressure of 20 cm H2O is created, which is important for creating correct and complete applanation. The endothelial surface of the cornea is moistened with the above solution for storing corneas to minimize the loss of endothelial cells at the time of applanation. The cornea ready for work in the IPC is placed under an applanation lens connected to a Ziemer FEMTO LDV Z8 femtosecond laser. After ensuring centering and complete applanation of the cornea, a corneal section of a given profile is made: the first section is a circular section of the corneal stroma with a diameter of 8 mm, calculated so that the residual thickness of the cornea from the endothelium is 125 μm. The second section is in the lamellar plane at a depth of 125 μm from the endothelium, with a diameter of 10 mm. The third section is circular, made from a depth of 125 μm upwards towards the endothelium, with a diameter of 10 mm. Then a radial section is made in the vertical plane, which divides the corneal transplant into two equal parts. After the femtosecond laser has finished working, the donor cornea in the IPC is placed under an operating microscope. The separation of corneal grafts is performed using a blunt spatula in the lamellar and vertical plane along the circumference, thereby obtaining two ready-made endothelial grafts and one anterior graft, which is then subjected to the crosslinking procedure: scarification of the epithelium is performed, then the donor tissue is placed in a Petri dish pre-filled with a solution of 0.1% riboflavin for 30 minutes, after which it is irradiated with UV light with a wavelength of 375 nm and a power density of 3 mW/ ^cm2 for 30 minutes, after which the anterior graft is also ready for transplantation to the recipient.

The proposed method is illustrated by the following clinical examples.

Example 1.

Patient H., 70 years old, with a diagnosis of Fuchs' corneal endothelial dystrophy of the left eye. Pseudophakia.

Visual acuity of 0.01 does not correct (n/c). Keratometry ax 58 ⁰ 46.25 ^D ax148 ⁰ 43.20 ^D. Central pachymetry 667 µm. According to endothelial microscopy, endothelial cell density (ECD) is not determined. At the first stage of the operation, two grafts were obtained from the donor cornea with ECD 2834 cells/mm ² to perform ZPK and one for GPPK using the declared (proposed) method. The next stage is ZPK for the first patient, for which a 3 mm wide tunnel incision is made on the cornea of the recipient with Fuchs' endothelial corneal dystrophy on the temporal side with a dosed knife, for example, Mani (Japan) 2.0 mm, paracentesis is performed on the nasal side. Using a reverse Sinskey hook with a reverse profile, an oval descemetorhexis measuring 7 mm along the horizontal axis and 5 mm along the vertical axis is performed, which ensures complete overlap of the descemetorhexis zone with the finished transplant and its high-quality adhesion. One of the finished transplants of the posterior corneal layers is positioned in the Buzin glide with the endothelium upwards using collet tweezers, for example 25 Ga Alcon, USA. The glide tip is inserted into the anterior chamber through a tunnel corneal incision. Using collet tweezers inserted through corneal paracentesis from the nasal side, the transplant is brought out into the anterior chamber of the recipient and straightened with an irrigation flow. The operation is completed by introducing air or a gas-air mixture for transplant adhesion. Suture fixation is not required due to the 3 mm corneal incision.

On the day after the surgery, the transplant was transparent, the cornea was slightly edematous, multiple folds of Descemet's membrane were present, there was 1/2 air bubble in the anterior chamber, the transplant was fully adapted. Visual acuity was 0.01 n/c. Keratometry ax72 ⁰ 45.65 D, ax167 ⁰ 43.00 D. On the sixth day, the transplant was transparent, corneal edema persisted, visual acuity was 0.1 n/c, pachymetry in the center of the cornea was 606 μm, according to optical coherence tomography, the transplant thickness in the central zone was 103 μm. After 3 months, vision was 0.5 n/c, the transplant was transparent, PEC - 1256 cells/mm ² , spherical equivalent of refraction +0.5 D. After 1 year, the transplant remained transparent, KOZ 0.7 n/k. PEC - 1240 cells/ ^mm2 , keratometry ax72 ⁰ 43.45 D, ax174 ⁰ 43.05 D, spherical equivalent of refraction +0.5 D, according to OCT data, the minimum thickness of the transplant in the central zone was 80 μm, on the periphery - 95 μm.

Example 2.

Patient I., 56 years old, diagnosed with bullous keratopathy of the left eye, pseudophakia. Visual acuity 0.01 n/a. Keratometry ax86 ⁰ 44.50 D, ax176 ⁰ 43.75 D. Central pachymetry 765 µm. Endothelial microscopy does not reveal PEC. At the first stage of the surgery, two grafts were obtained from the donor cornea with PEC 2834 cells/ ^mm2 to perform ZPK and one for GPPK using the declared (proposed) method. One of the posterior corneal layer grafts was transplanted to a patient with Fuchs's endothelial corneal dystrophy (example 1). Then, the second patient undergoes ZPK, for which a 3 mm wide tunnel incision is made on the cornea of the recipient with bullous keratopathy on the temporal side with a dosed knife, for example, Mani (Japan) 2.0 mm, and paracentesis is performed on the nasal side. Using a reverse Sinskey hook with a reverse profile, an oval descemetorhexis is performed measuring 7 mm along the horizontal axis and 5 mm along the vertical axis, which ensures complete overlap of the descemetorhexis zone with the finished transplant and its high-quality adhesion. The second transplant of the posterior corneal layers is positioned in the Buzin glide with the endothelium upward using collet tweezers, for example, 25 Ga Alcon, USA. The tip of the glide is inserted into the anterior chamber through a tunnel corneal incision. Using a collet forceps inserted through a corneal paracentesis from the nasal side, the transplant is brought out into the anterior chamber of the recipient and straightened by an irrigation flow. The operation is completed by introducing air or a gas-air mixture for transplant adhesion. Suture fixation is not required due to the 3 mm corneal incision.

The day after the surgery, the graft was transparent, the cornea was slightly edematous, multiple folds of Descemet's membrane were present, there was 1/2 air bubble in the anterior chamber, the graft was fully adapted. Visual acuity was 0.05 n/c. Keratometry ax78 ⁰ 43.45 D, ax168 ⁰ 43.00 D. On the sixth day, the graft was transparent, corneal edema persisted, visual acuity was 0.1 n/c, pachymetry in the center of the cornea was 635 μm, according to optical coherence tomography, the graft thickness in the central zone was 100 μm. After 3 months, vision was 0.4 n/c, the graft was transparent, PEC - 1276 cells/mm ² , spherical equivalent of refraction +0.5 D. After 1 year, the graft remained transparent, KOZ 0.7 n/c. PEC - 1235 cells/ ^mm2 , keratometry ax60 ⁰ 43.74 D, ax150 ⁰ 43.30 D, spherical equivalent of refraction +0.5 D, according to OCT data, the minimum thickness of the transplant in the central zone was 87 µm, on the periphery - 103 µm.

Example 3.

Patient K., 35 years old, diagnosis: stage III keratoconus of the left eye. Visual acuity 0.05 n/a, keratometry ax65 ⁰ 56.25 D, ax 155 ⁰ 50.40 D. According to Confoscan 4, the PEC was 3156/mm ² . According to ORA, the viscoelastic properties of the cornea are reduced. The thickness of the cornea at the thinnest point according to optical coherence tomography is 335 μm. At the first stage of the operation, two transplants were obtained from the donor cornea with PEC 2834 cells/mm ² for performing ZPK and one for GPPK using the declared (proposed) method. Posterior corneal grafts were transplanted to a patient with Fuchs' endothelial corneal dystrophy (case 1) and to a patient with bullous keratopathy (case 2). The prepared anterior corneal graft will be transplanted to a patient with keratoconus, for which the bed is prepared in advance: a corneal flap is cut out concentrically to the limbus with a femtosecond laser FEMTO LDV Z8 (Ziemer Ophthalmic Systems AG, Switzerland) to a depth of up to 80% of the total corneal thickness at the thinnest point with a diameter of 7.8 mm. A femtosecond laser is used to make a circular cut with a diameter of 7.8 mm, then a lamellar cut, during which an intrastromal tunnel is formed, starting from the bottom of the formed bed at 11 o'clock at an angle of 45 towards the center of the cornea, 0.8 mm wide and 1 mm long. After finishing the femtosecond laser operation, the tunnel is opened with a spatula, into which sterile air is then introduced in a volume of about 1.5 ml using a 27G cannula until a "large bubble" is formed. Next, paracentesis is performed in the limbus area to check for the presence of a formed air bubble between the DM and the residual stroma and to further reduce the pressure in the anterior chamber. The next step is to introduce 1.0 ml of cohesive viscoelastic through the tunnel into the area of the formed air bubble, by means of which the final separation of the residual stroma and the DM is performed. The donor cornea is placed in the prepared surgical bed, fixing sutures of 8-0 silk are applied, then a continuous suture of 10-0 nylon. In the postoperative period, good adaptation of the graft to the bed was observed, the graft was transparent from the first days, complete epithelialization on the 3rd day after the surgery. Seven days after the surgery, visual acuity was 0.05 sph +3.0 cyl 3.5 ах87 ⁰ = 0.2. According to optical coherence tomography, good adaptation of the graft was observed. The thickness of the residual stroma of the recipient is 85 μm in the thinnest area. The thickness of the graft is 408 μm. Average pachymetry in the center of the cornea is 515 μm. Keratometry ах87 ⁰ 43.35 D, ах177 ⁰ 39.95 D. The sutures were removed 12 months after the surgery. At the 1-year follow-up, the following data were obtained: the eye is calm, the transplant is transparent, the interface zone is barely visualized using a slit lamp. Visual acuity is 0.5 sph-1.5 cyl 1.5 ах92 ⁰ = 0.7. Keratometry ах90 ⁰ 44.35 D, ах0 ⁰ 42.85 D. According to ORA, the viscoelastic properties of the cornea are within the normal range. According to Confoscan-4, the PEC was 2985/mm ² , no signs of fibrosis were found in the interface zone. According to optical coherence tomography, good adaptation of the transplant was observed. The interface zone differed little from the surrounding tissues. The thickness of the residual stroma of the recipient was 74 μm in the thinnest area. The thickness of the transplant is 398 μm. The minimum pachymetry in the center of the cornea is 472 μm. The average pachymetry value in the central zone of the cornea according to OST data is 509 µm.

Thus, the described method of preparing three corneal grafts from one donor cornea ensures:

- obtaining three high-quality, geometrically uniform transplants;

- increasing the number of transplants for patients with corneal endothelial dystrophies and keratoconus;

- rapid visual rehabilitation of patients with keratoconus due to crosslinking of the anterior transplant after dissection of the donor cornea and the absence of the risk of secondary ectasia of the transplant.

Claims (1)

A method for preparing three corneal grafts from one donor cornea, characterized in that using a femtosecond laser, the first circular cut of the corneal stroma with a diameter of 8 mm is made, calculated in such a way that the residual thickness of the cornea from the endothelium is 125 μm, the second cut is in the lamellar plane at a depth of 125 μm from the endothelium, with a diameter of 10 mm, the third cut is circular, made from a depth of 125 μm upwards towards the endothelium, with a diameter of 10 mm, then a radial cut is made in the vertical plane, which divides the corneal graft into two equal parts, then the endothelial grafts are separated with a blunt spatula, after which the anterior graft is separated in the vertical plane from the donor's corneoscleral disc, which is then subjected to a corneal collagen crosslinking procedure with preliminary deepithelialization.