FIELD OF THE INVENTION
[001] The present invention generally relates to ophthalmology.
[002] More particularly, the present invention relates to method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects.
BACKGROUND FOR THE INVENTION:
[003] By reference to US application no. US2011189301A1 by Bioland Ltd dated 2010-04-15, titled” Method for preparing contact lens-shaped amniotic dressing” discloses a method for preparing a contact lens-shaped amniotic dressing and a contact lens-shaped amniotic dressing prepared therefrom for treating ocular surface diseases, which does not require the use of sutures or an adhesion material. The inventive contact lens-shaped amniotic dressing is capable of solving the problems associated with suturing an amniotic membrane, e.g., highly delicate surgical techniques of suturing, long surgery time, stitch abscess, granuloma formation, tissue necrosis, and discomfort of patients; and the problems associated with the use of a support, e.g., the elimination of the support by eye blinking, breaking of the support, and discomfort.
[004] By reference to US application no. US2004181240A1 by Individual dated 2003-03-14, titled” Amniotic membrane covering for a tissue surface and devices facilitating fastening of membranes” discloses a biopolymer covering for a tissue surface including, for example, a dressing, a bandage, a drape such as a bandage contact lens, a composition or covering to protect tissue, a covering to prevent adhesions, to exclude bacteria, to inhibit bacterial activity, or to promote healing or growth of tissue. An example of such a composition is an amniotic membrane covering for an ocular surface. Use of a covering for a tissue surface according to the invention eliminates the need for suturing. The invention also includes devices facilitating the fastening of a membrane to a support, culture inserts, compositions, methods, and kits for making and using coverings for a tissue surface and culture inserts. Compositions according to the invention may include cells grown on a membrane or attached to a membrane, and such compositions may be used as scaffolds for tissue engineering or tissue grafts. A method of preparing and using an amniotic membrane covering for a tissue surface as a controlled release drug delivery vehicle is also disclosed.
[005] By reference to US application no. US2002035358A1 by Vanderbilt University dated Pulsed electromagnetic field therapy for treatment of corneal disorders and injuries, titled” Pulsed electromagnetic field therapy for treatment of corneal disorders and injuries” discloses a method for treating corneal ulcers and other corneal conditions with pulsed electromagnetic fields (PEMFs). Specifically, the present invention includes a two part treatment regimen having particular waveforms, intensities, durations, pulse delays, and stepped frequency modulations which maximize the intrinsic healing capacity of cornea.
[006] By reference to US application no. US2003235580A1 by Vanderbilt University dated 2001-05-09, titled” Pulsed electromagnetic field therapy for treatment of corneal disorders and injuries” discloses a method for treating corneal ulcers and other corneal conditions with pulsed electromagnetic fields (PEMFs). Specifically, the present invention includes a two part treatment regimen having particular waveforms, intensities, durations, pulse delays, and stepped frequency modulations which maximize the intrinsic healing capacity of cornea.
[007] By reference to CN application no. CN111330081A by SHANDONG EYE INSTITUTE dated 2020-05-09, titled” Preparation method of medicine-carrying amniotic membrane and influence of medicine-carrying amniotic membrane on corneal epithelium repair” discloses drug-loaded amnion comprises the following steps: and selecting the human amniotic membrane preserved by the glycerol to prepare the amniotic membrane. Soaking in recombinant human nerve growth factor (rhNGF) at 4 deg.C for certain concentration, taking out amnion at different time periods, transferring to small culture dish containing Phosphate Buffered Saline (PBS) at room temperature, and screening for appropriate soaking time and concentration. The drug-loaded amniotic membrane prepared by the method is transplanted at a corneal epithelial defect part, and can promote the regeneration of corneal epithelium, the recovery of corneal sensitivity and the regeneration of nerve fibers of a nerve plexus under the corneal epithelium. The invention has the beneficial effects that: not only can reduce the frequency of drug administration after transplantation, reduce the influence of eye drops on the amnion and prevent the amnion from falling off in advance, but also can slowly release rhNGF to play the role of promoting the growth of epithelium and nerve. Therefore, the prepared amnion drug-carrying system with double functions has important clinical application value.
[008] By reference to RU application no. RU2007109452A by Sellriserch Corporation Pte Ltd. dated 2005-06-03, titled” Stem/progenitor cell recovery from amniotic membrane of umbilical cord” discloses method for stem/progenitor cell recovery from the amniotic membrane of the umbilical cord involves separating in vitro the amniotic membrane from the other components of an umbilical cord. Further, the amniotic membrane tissue is cultivated in the conditions to provide cell proliferation. Finally, the stem/progenitor cells are recovered from tissue cultures.
[009] By reference to US application no. US6152142A by Scheffer C. G. Tseng dated 2000-07-25, titled” Grafts made from amniotic membrane; methods of separating, preserving, and using such grafts in surgeries” discloses method for making, storing and using a surgical graft from human amniotic membrane; the resulting graft; and the storage solution. The amniotic membrane is obtained from human placenta, from which the chorion has been separated. Sheets of the amniotic membrane are cut to size and mounted on filter paper. The cells of the amniotic membrane are killed, preferably while being frozen and thawed in the storage solution. The storage solution comprises a culture medium and a hyperosmotic agent, wherein the hydration of the amniotic membrane is maintained. The membrane can be impregnated with therapeutic agents, prior to storage, for use in post surgical healing or other therapies.
[010] By reference to CN application no. CN100353835C by Scheffer C. G. Tseng dated 2000-07-25, titled” Grafts made from amniotic membrane; methods of separating, preserving, and using such grafts in surgeries” discloses method for making, storing and using a surgical graft from human amniotic membrane; the resulting graft; and the storage solution. The amniotic membrane is obtained from human placenta, from which the chorion has been separated. Sheets of the amniotic membrane are cut to size and mounted on filter paper. The cells of the amniotic membrane are killed, preferably while being frozen and thawed in the storage solution. The storage solution comprises a culture medium and a hyperosmotic agent, wherein the hydration of the amniotic membrane is maintained. The membrane can be impregnated with therapeutic agents, prior to storage, for use in post surgical healing or other therapies.
[011] However, none of the above-discussed inventions provides such a method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects. The method comprising a plurality of steps.
OBJECTS OF THE INVENTION:
[012] Some of the objects of the present disclosure, which at least one embodiment herein satisfies, are as follows.
[013] The main object of the present invention is to provide a method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects.
[014] Another object of the invention is to evaluate the clinical outcomes of tarsorrhaphy and amniotic membrane transplant in the healing of persistent corneal epithelial defects and subsequent corneal vascularization if any thereby providing symptomatic relief.
[015] Another object of the invention is to provide corneal vascularization, as an indicator of healing, progressively increased along the entire course of postoperative follow up.
[016] Other objects and advantages of the present disclosure will be more apparent from the following description, which is not intended to limit the scope of the present disclosure.
SUMMARY OF THE INVENTION:
[017] According to one aspect of our invention, method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects comprising the steps of: a) The stromal surface of the amniotic membrane is identified by the presence of vitreous such as strands using a cotton bud; b) All the debris over the corneal surface is removed prior to the graft placement with the help of the cotton bud and loose epithelium over a PED is also removed using small fine forceps; c) The graft is placed over the defect and spread out with the help of an iris repositor or a rod and the size of the defect is measured with calipers and grafts of appropriate size are cut in accordance; d) The size of the graft is kept at least 1 mm larger than the defect and 10-0 nonabsorbable nylon sutures are placed circumferentially or parallel to the cut edge in an interrupted or continuous purse-string manner and a bandage contact lens is applied over the amniotic membrane; e) The total healing time taken is recorded along with relief in the prerecorded symptoms and preoperative and postoperative pictures are taken; and further any complications related to tarsorrhaphy or AMT are recorded.
[018] In another aspect of the invention, pain score index: Pain perceived is recorded on a scale from grade 0–4 as described by the patient. Grade 0 - No pain, Grade 1- Mild pain, Grade 2- Moderate pain, Grade 3- Severe pain, Grade 4- Unimaginable pain.
[019] In another aspect of the invention, procedure Tarsorrhaphy is performed by first injecting local anesthesia (2% xylocaine) in the upper and lower lids.
[020] In another aspect of the invention, a raw area is created on the intermarginal strip using the number 11 blade.
BRIEF DESCRIPTION OF DRAWINGS:
[021] Reference will be made to embodiments of the invention, examples of which may be illustrated in accompanying figures. These figures are intended to be illustrative, not limiting. Although the invention is generally described in the context of these embodiments, it should be understood that it is not intended to limit the scope of the invention to these particular embodiments.
[022] Figure 1: illustrates a method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects, as per an embodiment of the present invention.
[023] The referral numerals in the figures refer to: 1,2,3,4,5,6,7,8 - Method steps of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects.
BRIEF DESCRIPTION OF INVENTION:
[024] The present invention will now be described hereinafter with reference to the accompanying drawings, in which some, but not all embodiments of the invention are shown. While the following description details the preferred embodiments of the present invention is not limited in its application to the details of construction and arrangement of the parts illustrated in the accompanying drawings. With reference to the figures, the enclosed description and drawings are merely illustrative of preferred embodiments and represent several different ways of configuring the present invention. Although specific components, materials, configurations and uses of the present invention are illustrated and set forth in this disclosure, it should be understood that a number of variations to the components and to the configuration of those components described herein and in the accompanying figures can be made without changing the scope and function of the invention set forth herein.
[025] The present invention proposes a method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects comprising the steps of: a) The stromal surface of the amniotic membrane is identified by the presence of vitreous such as strands using a cotton bud; b) All the debris over the corneal surface is removed prior to the graft placement with the help of the cotton bud and loose epithelium over a PED is also removed using small fine forceps; c) The graft is placed over the defect and spread out with the help of an iris repositor or a rod and the size of the defect is measured with calipers and grafts of appropriate size are cut in accordance; d) The size of the graft is kept at least 1 mm larger than the defect and 10-0 nonabsorbable nylon sutures are placed circumferentially or parallel to the cut edge in an interrupted or continuous purse-string manner and a bandage contact lens is applied over the amniotic membrane; e) The total healing time taken is recorded along with relief in the prerecorded symptoms and preoperative and postoperative pictures are taken; and further any complications related to tarsorrhaphy or AMT are recorded.
[026] The present invention as illustrated in Figure 1 provides another embodiment of the invention which includes method steps of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects.
[027] In another embodiment, to compare and study the clinical outcome of tarsorrhaphy and amniotic membrane transplant in the healing of persistent corneal epithelial defects in terms of clinical improvement and symptomatic relief.
[028] In another embodiment, a total of 60 patients with persistent epithelial defects (PED’s), randomly divided into two groups of 30 patients each who underwent tarsorrhaphy (Group A) or amniotic membrane transplantation (Group B) with a 4-week-follow-up period, are included. The main parameters studied are the size of an epithelial defect, total healing time, pain score, and complications.
[029] In another embodiment, the healing time is 9.83 ± 6.51 days in Group A (median = 9.50 days, IQR = 1–7 days) vs. 18.33 ± 13.46 days (median = 19.50 days, IQR = 1–21 days) in Group B. A total of ten eyes (16.7%) did not heal at the end of 4 weeks.
[030] In another embodiment, there is a significant reduction in the area of epithelial defect at the end of the 1 week and 2 week follow up postoperatively, in both the treatment forms. The mean healing time in patients of Group A is less as compared to that of the patients in Group B.
[031] In another embodiment, the various etiologies responsible for the PEDs included exposure keratopathy, post penetrating keratoplasty (PK), and trauma. All these patients are taken up for the respective procedures after instituting a maximum medical therapy and patching or BCL application for at least 2 weeks. The epithelial defects which did not heal on the above-mentioned therapy are labeled as PEDs and are included in the study.
[032] In another embodiment, Exclusion criteria:all patients with duration of epithelial defects less than 2 weeks, patients who are healed through medical therapy, severe LSCD, acute anterior segment infection or associated lid pathology, ocular surface disorders, painful bullous keratopathy, cases where intraocular pressure (IOP) is >21 mmHg, patients on long-term immunosuppressant drugs and repeated PK surgeries are excluded.
[033] In another embodiment, written informed consent is taken from the patient (or the patient’s parents in case the patient is less than 18 years) before including them in the study. Permission from the ethical committee is also obtained. These patients are randomly divided into two groups using envelope technique (Group A and Group B) of 30 eyes each. Patients in group A underwent tarsorrhaphy and the patients in group B underwent AMT. Demographic indices are included in the case recording form. Clinical assessment of the patients are done against pre decided parameters. A detailed history of present illness and past history is recorded. The ocular examination included best-corrected visual acuity (BCVA) done on Snellen’s visual acuity chart and later converted to logarithm of minimum angle of resolution (logMAR), IOP readings with a non contact tonometer or tonopen are taken wherever possible. Anterior segment evaluation is done using a Carl Zeiss Meditec AG slit lamp and the central fundus is seen with the help of a Heine beta 200 direct ophthalmoscope or an indirect ophthalmoscope.
[034] In another embodiment, the size of the epithelial defect, pain score, and the extent of vascularization of cornea is assessed preoperatively and postoperative follow-up assessment is carried out on the postoperative day 1, week 1, week 2, and week 4. The size of epithelial defect is recorded as the area of the cornea affected by the epithelial defect. It is calculated by measuring the length and the breadth of the PED (in mm) using the slit beam lamp bio-microscope and later multiplying the two entities. The area of PED is calculated in mm².
[035] In another embodiment, the extent of vascularization is recorded in the number of quadrants and type as superficial or deep. The symptoms of photophobia, lacrimation, and foreign body sensation are recorded.
[036] In another embodiment, the type of surgical modality performed is recorded (tarsorrhaphy/AMT). Further, the type of tarsorrhaphy (permanent/temporary) and site of tarsorrhaphy (central/para-median) is noted.
[037] In another embodiment, procedure tarsorrhaphy is performed by first injecting local anesthesia (2% xylocaine) in the upper and lower lids. A raw area is created on the intermarginal strip using the number 11 blade. The raw area measured 2mm × 5mm both on the lower and upper lids. The location of the same (para-median, median or lateral) is decided as per case and is sutured using 5-0 non-absorbable suture along with bolsters.
[038] In another embodiment, AMT is performed, the type of amniotic membrane used (fresh, cryopreserved, and commercially available) is recorded. The technique used is mentioned as either inlay, graft/overlay, patch/filling, or layered technique. The AMT stay is recorded in a number of days.
[039] In another embodiment, the stromal surface of the amniotic membrane is identified by the presence of vitreous such as strands using a cotton bud. All the debris over the corneal surface are removed prior to the graft placement with the help of the cotton bud.
[040] In another embodiment, loose epithelium over a PED is also removed using small fine forceps. The graft is placed over the defect and spread out with the help of an iris repositor or a rod. The size of the defect is measured with calipers and graft of appropriate size are cut in accordance. The size of the graft is kept at least 1 mm larger than the defect and 10-0 nonabsorbable nylon sutures are placed circumferentially or parallel to the cut edge in an interrupted or continuous purse-string manner. A bandage contact lens is applied over the amniotic membrane. The total healing time taken is recorded along with relief in the prerecorded symptoms. Preoperative and postoperative pictures are taken. Any complications related to tarsorrhaphy or AMT are recorded.
[041] In another embodiment, pain score index: pain perceived is recorded on a scale from grade 0–4 as described by the patient. Grade 0 - No pain, Grade 1- Mild pain, Grade 2- Moderate pain, Grade 3- Severe pain, Grade 4- Unimaginable pain.
[042] In another embodiment, statistical analysis: all continuous variables are described as mean ± standard deviation or median with interquartile range (IQR) and categorical variables are expressed as proportions. Group differences in continuous variables are analyzed using the Student t-test for normally distributed variables and the Mann-Whitney U test for nonparametric variables. The normality of distribution is tested using the Konglomerat Smirnov test. Group differences in categorical variables are analyzed using the Chi-square or Fischer’s exact test. The vision is measured in Snellens equivalent and is converted to logarithm of minimum angle of resolution (logMAR) for statistical analysis.
[043] In another embodiment, the demographic indices and the preoperative parameters are described in Table 1 along with the comparison between the two groups in Table 2. In the present study, 15 (25%) eyes had PED due to exposure keratopathy, 32 (54%) secondary to PK, 5 (8%) due to trauma, and 5 (8%) are idiopathic. Out of 30 eyes that underwent tarsorrhaphy, 27 (90%) had a permanent tarsorrhaphy and 3 (10%) had a temporary tarsorrhaphy. The tarsorrhaphy is removed after a mean ± SD of 52.1 ± 47 days (median = 26 days, IQR- 18–78 days, range = 6–171 days). Out of the 30 eyes that had AMT, 20 (71%) had freshAMT, 5 had cryopreserved AMT, and 5 had commercial available AMT transplantation.

[044] In another embodiment, the sutures securing the AMT are removed after a mean ± SD of 15.7 ± 5 days (median = 15.5 days, IQR = 12–20 days) and the AMT lasted for a mean duration ± SD of 22.6 ± 7 days (median = 23.5 days, IQR = 17.5– 27.5 days).
[045] In another embodiment, the healing time is 9.83 ± 6.51 days in Group A (median = 9.50 days, IQR = 1–7 days) vs. 18.33 ± 13.46 days (median = 19.50 days, IQR = 1–21 days) in Group B.
[046] In another embodiment, the comparison of healing characteristics in the form of size of the epithelial defect and the corneal vascularization between the two groups at various follow-up periods have been described in Tables 3 and 4, respectively. A total of ten eyes (16.7%) did not heal at the end of 4 weeks, out of which seven eyes (23.3%) are from Group B and three eyes (10%) are from Group A.

[047] In another embodiment, moreover, smokers showed a trend of delayed healing. It is also observed that patients with associated systemic comorbidities such as diabetes (although controlled on medications) also showed delayed healing.
[048] In another embodiment, furthermore, the eyes that did not heal showed a positive association with smoking (P-value = 0.04). All the patients who did not heal are smokers and four of them are diabetic patients.
[049] In another embodiment, the mean area (± SD) of the epithelial defect preoperatively is 30.74 ± 27.34 mm². This varied from 26.57 ± 24.55 mm² for patients in Group A. This is to 34.90 ± 30.14 mm² for patients in Group B. The difference between the preoperative sizes of the epithelial defect is statistically nonsignificant between the two groups hence making them comparable with no bias. This preoperative size
[050] In another embodiment, there is a statistically significant reduction in the area of epithelial defect at the end of week 1 and week 2 follow-up postoperatively in both the treatment forms. The mean healing time in patients of Group A is less compared to that of patients in Group B and is statistically significant. Habituations such as smoking delayed the healing of the PED. It is also noted the patients with comorbidities such as uncontrolled diabetes mellitus also retarded the healing process. Corneal vascularization, as an indicator of healing, progressively increased along the entire course of postoperative follow up. Patients in Group A ended up developing only superficial vascularization whereas the patients in Group B had both deep as well as superficial entities. Pain score also reduced subsequently along the follow-up period with drastic reduction at first follow-up day and at the end of week 1 with both the treatment forms.
[051] In another embodiment, both the treatment modalities significantly reduced the symptomatology associated with the PED by the end of the study. Only 10 out of 60 patients did not heal at the end of the study. Three patients are from Group A and seven are from Group B.
[052] The present disclosure described herein above has several technical advantages including, but not limited to,
? amniotic membrane contains higher levels of growth factors such as hepatocyte and transforming growth factors which modulate proliferation and differentiation of stromal fibroblast and promotes deep vascularization.
[053] The disclosure has been described with reference to the accompanying embodiments herein and the various features and advantageous details thereof are explained with reference to the non-limiting embodiments in the following description. Descriptions of well-known components and processing techniques are omitted so as to not unnecessarily obscure the embodiments herein.
[054] The foregoing description of the specific embodiments so fully revealed the general nature of the embodiments herein that others can, by applying current knowledge, readily modify and/or adapt for various applications such specific embodiments without departing from the generic concept, and, therefore, such adaptations and modifications should and are intended to be comprehended within the meaning and range of equivalents of the disclosed embodiments. It is to be understood that the phraseology or terminology employed herein is for the purpose of description and not of limitation. Therefore, while the embodiments herein have been described in terms of preferred embodiments, those skilled in the art will recognize that the embodiments herein can be practiced with modification within the scope of the embodiments as described herein.

We Claim:

1. A method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects comprising the steps of: a) The stromal surface of the amniotic membrane is identified by the presence of vitreous such as strands using a cotton bud; b) All the debris over the corneal surface is removed prior to the graft placement with the help of the cotton bud and loose epithelium over a PED is also removed using small fine forceps; c) The graft is placed over the defect and spread out with the help of an iris repositor or a rod and the size of the defect is measured with calipers and grafts of appropriate size are cut in accordance; d) The size of the graft is kept at least 1 mm larger than the defect and 10-0 nonabsorbable nylon sutures are placed circumferentially or parallel to the cut edge in an interrupted or continuous purse-string manner and a bandage contact lens is applied over the amniotic membrane; e) The total healing time taken is recorded along with relief in the prerecorded symptoms and preoperative and postoperative pictures are taken; and further any complications related to tarsorrhaphy or AMT are recorded.
2. The method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects as claimed in claim 1, wherein pain score index: Pain perceived is recorded on a scale from grade 0–4 as described by the patient. Grade 0 - No pain, Grade 1- Mild pain, Grade 2- Moderate pain, Grade 3- Severe pain, Grade 4- Unimaginable pain.
3. The method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects as claimed in claim 1, wherein procedure Tarsorrhaphy is performed by first injecting local anesthesia (2% xylocaine) in the upper and lower lids.
4. The method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects as claimed in claim 1, wherein a raw area is created on the intermarginal strip using number 11 blade.
5. The method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects as claimed in claim 1, wherein the raw area measured 2mm × 5mm both on the lower and upper lids.
6. The method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects as claimed in claim 1, wherein the location of the same (para-median, median or lateral) is decided as per case and is sutured using 5-0 nonabsorbable suture along with bolsters.
7. The method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects as claimed in claim 1, wherein 15 (25%) eyes had PED due to exposure keratopathy, 32 (54%) secondary to PK, 5 (8%) due to trauma, and 5 (8%) are idiopathic.
8. The method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects as claimed in claim 1, wherein out of 30 eyes that underwent tarsorrhaphy, 27 (90%) had a permanent tarsorrhaphy and 3 (10%) had a temporary tarsorrhaphy and the tarsorrhaphy is removed after a mean ± SD of 52.1 ± 47 days (median = 26 days, IQR- 18–78 days, range = 6–171 days)
9. The method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects as claimed in claim 1, wherein out of the 30 eyes that hadAMT, 20 (71%) had freshAMT, 5 had cryopreservedAMT, and 5 had commercial availableAMT transplantation; wherein the sutures securing the AMT are removed after a mean ± SD of 15.7 ± 5 days (median = 15.5 days, IQR = 12–20 days) and the AMT lasted for a mean duration ± SD of 22.6 ± 7 days (median = 23.5 days, IQR = 17.5– 27.5 days).
10. The method of tarsorrhaphy and amniotic membrane transplantation in the healing of persistent corneal epithelial defects as claimed in claim 1, wherein the healing time is 9.83 ± 6.51 days in Group A (median = 9.50 days, IQR = 1–7 days) vs. 18.33 ± 13.46 days (median = 19.50 days, IQR = 1–21 days) in Group B.