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Urea-De-Epithelialized Human Amniotic Membrane for Ocular Surface Reconstruction.

ABSTRACT: The conjunctiva is a clear tissue covering the white part of the eye and lines the back of the eyelids. Conjunctival diseases, such as symblepharon, cause inflammation, discharges, and photophobia. The treatment often requires excision of large parts of conjunctiva. Tissue engineering of conjunctival cells using human amniotic membrane (HAM) denuded of its epithelium as a basement membrane scaffold has been shown to be effective for covering conjunctival defects. However, most epithelial denudation protocols are time-consuming and expensive or compromise HAM's basement membrane structure and matrix components. We have previously described a method to de-epithelialize HAM using ice-cold urea (uHAM). In this report, we used this method to provide tissue-engineered constructs with cultivated conjunctival epithelial cells on uHAM in two patients, one with a giant conjunctival nevus and the other with a large symblepharon. Autologous conjunctival epithelial cells harvested from incisional biopsies of these two patients were cultured on the uHAM scaffold. The transplantation of tissue-engineered constructs to patients' ocular surface immediately after the removal of lesions showed successful reconstruction of the ocular surface. Postoperatively, there were neither recurrence of lesions nor epithelial defects throughout the follow-up (up to 7 and 19 months, respectively). This report highlights the translational potential of an efficient and inexpensive method to prepare de-epithelialized HAM as a basement membrane scaffold for cell-based tissue-engineered treatments of ocular surface disorders. Stem Cells Translational Medicine 2019;8:620&626.

SUBMITTER: Bandeira F

PROVIDER: S-EPMC6591546 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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Urea-De-Epithelialized Human Amniotic Membrane for Ocular Surface Reconstruction.

Bandeira Francisco F Yam Gary Hin-Fai GH Fuest Matthias M Ong Hon Shing HS Liu Yu-Chi YC Seah Xin-Yi XY Shen Sunny Y SY Mehta Jodhbir S JS

Stem cells translational medicine 20190313 7

The conjunctiva is a clear tissue covering the white part of the eye and lines the back of the eyelids. Conjunctival diseases, such as symblepharon, cause inflammation, discharges, and photophobia. The treatment often requires excision of large parts of conjunctiva. Tissue engineering of conjunctival cells using human amniotic membrane (HAM) denuded of its epithelium as a basement membrane scaffold has been shown to be effective for covering conjunctival defects. However, most epithelial denudat ...[more]

PMID: 30868769

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Project description:BackgroundCorneal disease is the second most common cause of blindness in China. Clinically, treatment options for corneal diseases with limbal stem cell deficiency (LSCD) are limited due to a shortage of organ donors and inevitable immune rejection. This study aims to determine the efficacy of reconstructing the ocular surface using autologous cultivated adipose tissue-derived stem cells (ADSCs) and to develop a new clinical treatment for patients with LSCD.MethodsA rabbit LSCD model was first established. Two weeks later, the animals were divided into three groups, including the sham group, the amniotic membrane transplantation group, and the ADSC combined with amniotic membrane transplantation group, and underwent surgery. The efficacy of reconstructing the ocular surface using ADSCs was evaluated using immunofluorescent staining, confocal microscopy (CM) observation, H&E staining, immunohistochemical staining, and scanning transmission electron microscopy observation one, two and four weeks after surgery.ResultsEvaluations of immunofluorescent staining of the cornea pre- and post-surgery yielded significantly lower scores for the corneas in the ADSCs transplantation group than for those in the sham group (F=-7, P=0.002, <0.05) and the amniotic membrane transplantation group (F=-4.67, P=0.018, <0.05) two weeks after surgery. Four weeks after surgery, the corneas of the ADSC combined with amniotic membrane transplantation group were scored significantly lower than those in the sham group (F=-8, P=0.007, <0.05) and the amniotic membrane transplantation group (F=-5.33, P=0.046, <0.05). The data suggest that the use of ADSCs to treat LSCD showed greater efficacy than the other treatment methods. The growth of ADSCs on the corneal surface was examined using confocal and electron microscopes. K3/K12 expression in the corneal epithelium, which was reconstructed by ADSCs, was negative, as shown by immunohistochemical staining.ConclusionsOcular surface reconstruction can be improved by using ADSCs as seed cells and the amniotic membrane as a carrier, thus providing a new therapeutic strategy for patients with LSCD.

Project description:BackgroundOcular surface burns can be caused by chemicals (alkalis and acids) or by direct heat. Amniotic membrane transplantation (AMT) performed in the acute phase (day 0 to day 7) of an ocular surface burn is reported to relieve pain, accelerate healing and reduce scarring and blood vessel formation. The surgery involves applying a patch of amniotic membrane (AM) over the entire ocular surface up to the eyelid margins.ObjectivesTo assess the effects of AMT on the eyes of people having suffered acute ocular surface burns.Search methodsWe searched CENTRAL (which contains the Cochrane Eyes and Vision Group Trials Register) (The Cochrane Library 2012, Issue 6), MEDLINE (January 1946 to June 2012), EMBASE (January 1980 to June 2012), Latin American and Caribbean Literature on Health Sciences (LILACS) (January 1982 to June 2012), the metaRegister of Controlled Trials (mRCT) (www.controlled-trials.com), ClinicalTrials.gov (www.clinicaltrials.gov) and the WHO International Clinical Trials Registry Platform (ICTRP) (www.who.int/ictrp/search/en). We did not use any date or language restrictions in the electronic searches for trials. We last searched the electronic databases on 11 June 2012.Selection criteriaWe included randomised trials of medical therapy and AMT applied in the first seven days after an ocular surface burn compared to medical therapy alone.Data collection and analysisTwo authors independently assessed the risk of bias of included studies and extracted relevant data. We contacted trial investigators for missing information. We summarised data using risk ratios (RRs) and mean differences (MDs) as appropriate.Main resultsWe included one RCT of 100 participants with ocular burns that were randomised to treatment with AMT and medical therapy or medical therapy alone. A subset of patients (n = 68) who were treated within the first seven days of the injury met the inclusion criteria and were included in the analysis. The remaining 32 eyes were excluded. The included subset consisted of 36 moderate (Dua classification II-III) and 32 severe (Dua classification IV-VI) ocular burns from alkali, acid and thermal injuries. In the moderate category, 13/20 control eyes and 14/16 treatment eyes had complete epithelialisation by 21 days. The RR of failure of epithelialisation by day 21 was 0.18 in the treatment group (95% confidence interval (CI) 0.02 to 1.31; P = 0.09). Mean LogMAR final visual acuities were 0.06 (standard deviation (SD) 0.10) in the treatment group and 0.38 (SD 0.52) in the control group, representing a MD of -0.32 (95% CI -0.05 to -0.59). In the severe category, 1/17 treatment and 1/15 control eyes were epithelialised by day 21. The RR of failure of epithelialisation in the treatment group was 1.01 (95% CI 0.84 to 1.21; P = 0.93). Final visual acuity was 1.77 (SD 1.31) in the treated eyes and 1.64 (SD 1.48) in the control group (MD 0.13; 95% CI -0.88 to 1.14). The risks of performance and detection biases were high, because treating personnel and outcome assessors could not be masked to treatment. There was also a high risk of bias in the visual outcomes of the moderate category, since mean visual acuity was significantly worse at presentation in the control eyes. This reduced confidence in the study findings.Authors' conclusionsConclusive evidence supporting the treatment of acute ocular surface burns with AMT is lacking. Heterogeneity of disease presentation, variations in treatment, undefined criteria for treatment success and failure, and non-uniform outcome measures are some of the factors complicating the search for clear evidence regarding this treatment.

Project description:BackgroundMany studies have shown that mucosa-treated tracheas can be used for allotransplantation without the use of immunosuppressants. However, no literature has reported whether the transplanted trachea has normal biomechanics. In this study, fresh dog trachea was used as a control to determine the mechanical properties of allograft trachea after mucous membrane removal.MethodsCervical tracheas (~10 cm in length, consisting of the cartilage rings down to the tracheal carina) and the surrounding connective tissues were removed from 20 mixed-breed dogs and then divided into 4 groups with 5 samples in each group. The groups were defined as follows: group A, fresh canine tracheas without any treatment; group B, canine tracheas after sodium dodecyl sulphate (SDS) de-epithelialization; group C, canine tracheas implanted into the dorsal fascia of beagle dogs for 1 month after SDS de-epithelialization; group D, canine tracheas implanted into the dorsal fascia of beagle dogs for 6 months after SDS de-epithelialization. All heterotopically implanted beagle dogs were free of any immunosuppressive treatment. Uniaxial tensile (n=20) and compressive (n=20) experiments were performed on the 4 groups of trachea using a universal testing machine; the tensile load was recorded at 150% of the original tracheal length (50% displacement), and the compressive load was recorded at 50% of the initial anteroposterior diameter (50% displacement). The average difference in tensile and compressive loads between groups was compared by analysis of variance.ResultsNo significant difference was found in the mean compressive strength among groups (group A =2.38±0.87 N, group B =3.33±1.03 N, group C =5.39±2.44 N, group D =3.04±2.12 N; P=0.07 for all groups). The mean tensile strength of group C was 9.68±0.86 N, resulting in a significant difference in tensile strength compared to group A (22.51±3.91 N) and B (24.24±7.46 N) (P<0.001). No significant difference in tensile strength was observed among groups A, B, and D.ConclusionsThe de-epithelialized trachea showed no significant change in mechanical properties after 6 months of implantation.

Project description:PurposeTo propose an improved stem cell-based strategy for limbal stem cell deficiency (LSCD) treatment.DesignExperimental randomized or parallel-group animal study.SubjectsFifty adult male New Zealand white rabbits.MethodsHuman limbal stem/progenitor cells (LSCs) and limbal stromal stem/progenitor cells (LSSCs) were cultured in serum-free conditions and further differentiated into corneal epithelial cells and keratocytes, respectively. All cell types were characterized with lineage-specific markers. Gene expression analysis was performed to identify the potential function of LSSCs in corneal regeneration. Two LSCD models of rabbits for transplantations were used: transplantation performed at the time of limbal and corneal epithelial excision (LSCD model) and transplantation performed after clinical signs were induced in an LSCD model (pLSCD model). The pLSCD model better mimics the pathologic changes and symptoms of human LSCD. Rabbit models received LSC or LSC plus LSSC treatment. Corneal epithelial defects, neovascularization, and opacity were assessed every 3 weeks for 24 weeks. ZsGreen-labeled LSSCs were used for short-term tracking in vivo.Main outcome measuresRates of corneal epithelial defect area, corneal neovascularization and opacity scores, graft survival rate, and immunofluorescence staining of specific markers.ResultsBoth LSC transplantation and LSC plus LSSC cotransplantation effectively repaired the corneal surface in the LSCD model. These 2 strategies showed no significant differences in terms of graft survival rate or epithelial repair. However, corneal opacity was observed in the LSC group (in 3 of 8 rabbits), but not in the LSC plus LSSC group. Notably, when treating LSCD rabbits with distinguishable stromal opacification and neovascularization, cotransplantation of LSCs and LSSCs exhibited significantly better therapeutic effects than transplantation of LSCs alone, with graft survival rates of 87.5% and 37.5%, respectively. The implanted LSSCs could differentiate into keratocytes during the wound-healing process. RNA sequencing analysis showed that the stromal cells produced not only a collagen-rich extracellular matrix to facilitate reconstruction of the lamellar structure, but also niche factors that accelerated epithelial cell growth and inhibited angiogenesis and inflammation.ConclusionsThese findings highlight the support of stromal cells in niche homeostasis and tissue regeneration, providing LSC plus LSSC cotransplantation as a new treatment strategy for corneal blindness.

Dataset Information

Urea-De-Epithelialized Human Amniotic Membrane for Ocular Surface Reconstruction.

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Urea-De-Epithelialized Human Amniotic Membrane for Ocular Surface Reconstruction.

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