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Purpose: To investigate the effects of riboflavin and/or ultraviolet?A (UV?A) irradiation on the cell viability of ex?vivo?cultured rat limbal stem cells (LSCs). Methods: LSCs of male Wistar rats (N = 12 eyes) were cultured, and immunofluorescence staining was performed to evaluate them. After characterization, these cells were assigned to four groups of control (C), a group that was exposed to UV?A radiation (UV), a group that was treated with riboflavin (R), and a group that cotreated with both UV?A and riboflavin (UV+R). To determine the cell viability of LSCs, these cells were subjected to MTT assay on days 1, 3, and 7 after exposure to UV?A and/or riboflavin. The duration of exposure to UV?A and riboflavin was similar to levels used during the conventional corneal collagen cross?linking procedure. Results: Compared with the viable cells in the control group, there was a significant decrease (P < 0.0001) in the number of LSCs in the UV group during all study days. In the R group, the level of viable LSCs was as same as the level of viable LSCs in the C group. Combined treatment with UV?A plus riboflavin significantly decreased the survival of LSCs on days 1 and 3 (P < 0.0001, P < 0.001, respectively) compared with the control group. Interestingly, in the UV+R group, the photosensitizing effect of riboflavin significantly decreased the cytotoxic effect of UV irradiation 7 days after exposure. Conclusion: These results suggest that the administered UV energy in the presence or absence of riboflavin can damage LSCs. Likewise, riboflavin could decrease the toxic effect of UVA on LSCs
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Purpose: To report the clinical outcomes and histopathological and immunohistochemistry (IHC) features in eyes with the sequelae stage of vernal keratoconjunctivitis (VKC). Methods: Investigative study of corneal samples obtained following surgical intervention for vision restoration in four eyes of three patients with VKC. Patient 1 (an 11?year?old boy) had deep anterior lamellar keratoplasty in both eyes, Patient 2 (a 24?year?old male) underwent superficial keratectomy followed by penetrating keratoplasty, and Patient 3 (a 22?year?old male) underwent penetrating keratoplasty. The corneal samples retrieved after surgical intervention were assessed for histology features and immunohistochemistry (IHC) studies. Results: The grafts were clear till the follow?up of 2–18 months. Histopathology of all four corneal samples showed epithelial hyperplasia, absent Bowman layer, thick hyalinized stromal lamellae, vascularization, and chronic inflammatory cells such as lymphocytes and plasma cells. IHC showed strong expression of CK 3 in both eyes of Patient 1 and no expression in Patients 2 and 3. The marker for limbal stem cells, ABCG2, was absent in all four samples; however, p63? was expressed strongly in Patients 2 and 3, moderately in the right eye of Patient 1, and marginally expressed in the left eye of Patient 1. Conclusion: The eyes in the sequelae stage of VKC (having corneal scarring and 360° hypertrophied limbus) can be managed favorably with keratoplasty and amniotic membrane transplantation without allogenic/cadaveric stem cell transplantation. The expression of transient progenitor cells in the scarred corneas of VKC patients in the sequelae stage suggests that the limbal stem cell dysfunction is more likely partial and self?renewal of limbal stem cells is a plausibility in these eyes
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@#AIM:To investigate the clinical effect of modified free conjunctival flap transplantation with limbal stem cells in the treatment of pterygium and its impact on tear film function.<p>METHODS: A total of 60 pterygium patients admitted to our hospital from March 2017 to March 2021 were randomly divided into the control group and the observation group, with 30 cases in each group. The control group was treated with amniotic membrane transplantation, and the observation group was treated with modified free conjunctival flap transplantation with limbal stem cells. The treatment period of both groups was 21d. And then the clinical efficacy, operation duration, corneal wound repair time, xerophthalmia score, tear film rupture time value and basic tear secretion test value before and 1, 3mo after treatment, and the occurrence of adverse reactions were compared between the two groups.<p>RESULTS: The total effective rate of clinical treatment in the observation group was higher than that of the control group(χ2=5.963, P=0.015), the operation duration in the observation group was longer than that for the control group(t=-2.643, P<0.05), and the repair time of corneal wound was shorter than that of the control group(t=2.182, P<0.05). At 1 and 3mo after surgery, dry eye score in two groups decreased compared with that before surgery(all P<0.05), and there was a difference between observation group and control group(t=2.082, 3.956; all P<0.05). The time of tear film rupture at 1 and 3mo after surgery was increased compared with that before surgery(all P<0.05), and there was a difference between the observation group and the control group(t=4.245, 2.070; all P<0.05).The experimental value of basal tear secretion at 1 and 3mo after surgery increased compared with that before surgery(all P<0.05), and there was a difference between the observation group and the control group(t=2.076, 2.223; all P<0.05).There was no significant difference in the incidence of complications between the two groups(P=0.572).<p>CONCLUSION: The improved free conjunctival flap transplantation with limbal stem cells can effectively improve the clinical symptoms and tear film function of patients with pterygium, which is beneficial to the recovery of postoperative corneal wound, and can reduce the incidence of adverse reactions.
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@#AIM: To investigate the effect of L-carnitine(LC)on corneal epithelial repair and its regulatory molecular mechanism in the hypertonic and inflammatory environment caused by alkali burn.<p>METHODS: Ninety-six healthy C57/6J mice were randomly divided into blank control group, phosphate buffered solution(PBS)group and LC group. The blank control group did not receive any treatment, LC group and PBS group were prepared acute alkali burn models. LC group was given 60mmol/L LC eye drops, and PBS group was given PBS eye drops, 6 times/d, for continuous days from one day before alkali burn. The repair of corneal epithelium was observed by fluorescein sodium staining under slit lamp microscope at 0h, 3 and 7d. On the 3d, the expressions of Ki-67 and IL-1β proteins in cornea were detected by immunofluorescence, the total proteins of corneal epithelial were extracted for Western blot to detect the expression of P63, NLRP3, Caspase-1 and phosphorylation level of STAT3.<p>RESULTS: The results of corneal fluorescein sodium staining showed that on the 3 and 7d after alkali burn, the percentage of residual corneal epithelial defect area in PBS group compared with LC group was(29.38±6.83)% <i>vs</i>(17.78±4.11)% and(14.23±4.51)% <i>vs</i>(4.10±2.10)%, respectively(<i>P</i><0.01). The repair of corneal epithelium in LC group was faster than that in PBS group. On the 3d, compared with the blank control group, the expressions of pyroptosis related proteins NLRP3 and Caspase-1 in the corneal epithelium of the alkali burn treated mice were up-regulated, the expression of P63 was decreased, and the p-STAT3/STAT3 level was increased, all the differences were significant except cleaved Caspase-1 of blank control group <i>vs</i> LC group. Compared with PBS group, in LC group, the expression of NLRP3, pro Caspase-1 and cleaved Caspase-1 protein were decreased, P63 was up-regulated, and p-STAT3 /STAT3 was increased, all the differences were significant. Immunofluorescence showed that compared with the blank control group,the expressions of IL-1β and Ki-67 were up-regulated in the alkali burned group. Compared with PBS group, the expression of Ki-67 protein was up-regulated and IL-1β was decreased in LC group.<p>CONCLUSION: LC can promote the proliferation of stem/progenitor cells in the corneal epithelium of mice and further promote the repair of corneal epithelium after alkali burn by inhibiting the pyroptosis signaling pathway and promoting the activation of STAT3 signaling pathway.
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BACKGROUND: Wnt signaling pathway plays an important role in the regulation of stem cells, but its regulatory effect on limbal stem cells is not well defined. OBJECTIVE: To investigate the regulation of limbal stem cells via Wnt signaling pathway and its function in the treatment of limbal stem cell deficiency. METHODS: Rat limbal segments were digested with Dispase and trypsin/EDTA and then single limbal stem cells were seeded in 3D-Matrigel. In experimental group LiCl (500 μmol/L) was added to the culture system. Control group received no LiCl. The expression levels of p63α, CK12, CEBPδ and Ki67 in limbal stem cells were detected by qRT-PCR on the 7th day of culture. The expression level of β-catenin in limbal stem cells was detected by immunofluorescence staining. The rat model of limbal stem cell deficiency was made by alkali burn method. Rats in treatment group were treated with Wnt-activated limbal stem cells by subconjunctival injection. Rats in control group were treated with PBS. Rats were checked by slit lamp every day. On the 4th day after treatment, the immunofluorescence staining and hematoxylin-eosin staining were applied to evaluate the repair of limbus. RESULTS AND CONCLUSION: (1) The limbal stem cells aggregated in 3D-Matrigel. β-Catenin was negative in the control group. β-Catenin was found in the cytoplasm and nucleus of the limbal stem cells in the experimental group. (2) qRT-PCR results showed there was no significant difference in the levels of p63α, CK12 or CEBPδ between control and experimental groups (P > 0.05). The Ki67 level in the experimental group was significantly higher than that in the control group (P < 0.05). (3) The rat models of limbal stem cell deficiency were established. The degree of corneal opacity in the treatment group was significantly lower than that in the control group (P < 0.05). Hematoxylin-eosin staining results observed that the corneal epithelial cells in the treatment group were neatly arranged, the cell size was uniform, and repair was good. Immunofluorescence staining showed that β-catenin was found in the cytoplasm and nucleus of most corneal epithelial cells in the treatment group. While β-catenin was only weakly positive in the control group, and was invisible in the cytoplasm or nucleus. (4) These results indicate that activation of Wnt signaling pathway in limbal stem cells enhances their proliferation and keeps them in an undifferentiated self-renewal state. Wnt-activated limbal stem cells can promote the regeneration of corneal epithelium and reduce the degree of corneal opacity in the treatment of limbal stem cell deficiency. The regulation of limbal stem cells via Wnt signaling pathway is expected to provide new ideas for the treatment of limbal stem cell deficiency.
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@#Persistent corneal epithelial defect(PED/PCEDs)is an eye disease that fails to form corneal epithelium rapidly even after 10-14d of corneal injury. Corneal protective epithelial destruction and stromal layer damage can easily lead to eye infection, stromal ulcer, perforation, scar formation, and even blindness. At present, clinicians still face considerable challenges in treating PED patients. Standard treatments such as wearing bandaged contact lenses and using artificial tears, while newly developed drugs can promote the formation of various growth factors to re-form the cornea, and further cooperate with the corresponding surgery to provide innervation for the cornea. In order to achieve the effect of treatment. In addition, treatment should be carried out as soon as possible after the diagnosis of PED to avoid secondary complications. This article reviews the epidemiology, etiology, diagnosis, clinical manifestation, treatment and prognosis of persistent corneal epithelial defect.
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@#The continuous proliferation and differentiation of limbal stem cells(LSCs)maintain the integrity and homeostasis of the corneal epithelium, which plays an important role in protecting the cornea and maintaining corneal transparency. Currently limbal stem cell deficiency(LSCD)is one of major causes of blindness in corneal diseases, and transplantation of LSCs is the hot therapeutic option. The effectiveness of transplantation mainly depends on the proportion of LSCs to the total transplanted cells, so it is very important to clearly identify LSCs. There are many markers of LSCs, but their specificity is controversial. Therefore, one of the main challenges in LSCs transplantation is the lack of definitive cell markers. In this paper, the latest research progress of LSCs markers is reviewed.
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Simple limbal epithelial transplantation (SLET) is an innovative limbal stem cell transplantation technique that has gained increasing popularity over the last few years. Different groups from across the world have published the clinical results of SLET in large case series with varying types and severities of limbal stem cell deficiency (LSCD). This review attempts to place all the available knowledge on SLET together in one place for the benefit of not only cornea specialists and trainees but also for residents and general ophthalmologists. It follows a balanced approach of blending evidence with experience by providing an objective analysis of published results along with helpful insights from subject experts, starting from preoperative considerations including the role of newer imaging modalities to the technical aspects of the surgery itself and the management of possible complications. Original data and novel insights on allogeneic SLET for bilateral LSCD are included in the review to address the few remaining lacunae in the existing literature on this topic. This review intends to inform, educate, and empower all aspiring and practicing SLET surgeons to optimize their clinical outcomes and to have maximal positive impact on the lives of the individuals affected by unilateral or bilateral chronic LSCD.
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Background: Pterygium is a triangular wing shaped fiovascular growth of subconjunctival tissue on to the cornea. Surgical removal is the treatment of choice but no single technique is successful due to high recurrence rate. Aim: To evaluate the success and complications of sutureless glue–free conjunctival–limbal autografting in management of primary pterygium. Materials and Methods: A prospective interventional study was carried out in 60 patients to analyse the outcome of sutureless and glue–free conjunctival–limbal autograft for the management of primary nasal pterygium. The patients were followed up after 1 week, 3 weeks, 6 weeks and at 3 months postoperatively. The mean age of the patients was 38.28± 13.77 years (range 21–67), 55% of which were females. Graft retraction occurred in 3(5%) eyes. Haemorrhage was seen in 20(33.33%) eyes at 24 hours, which persisted in only 8(13.33%) eyes at 3 weeks and resolved completely in 100% of eyes at 6 weeks. Oedema was noted in 5(8.33%) eyes at 24 hours, and resolved completely by 1 week. Recurrence of pterygium was observed in 2(3.33%) eyes at three months of follow–up. Conclusion: Sutureless and glue–free conjunctival–limbal autograft following pterygium excision is an easy, quick, safe, effective, and economical option for the management of primary pterygium.
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@#AIM: To investigate the effect of cultured limbal stem cells on proliferating of vascular endothelial cells and validate the theory of limbal stem cells.<p>METHODS: The limbal stem cells and epithelium cells of bulbar conjunctiva were cultured and determined by immunohistochemistry staining with AE-5. The supernatant was collected and added into the cultured human umbilical vein endothelium cells(ECV-304). The negative control was set up. After 24h, MTT was done to detect endothelium cell proliferation. The statistical analysis was done by analysis of Variance.<p>RESULTS: Negative stain of AE-5 can be seen in limbal stem cells and positive stain can be seen in conjunctiva cells. The ratio of MTT added by limbal stem cell supernatant, conjunctiva epithelium cell and negative control were 2.097±0.079, 1.981±0.034 and 1.990±0.044, respectively. There were significant difference among the three groups(<i>F</i>=9.169, <i>P</i>=0.000). The proliferate activity of vascular endothelial cells added by supernatant of limbal stem cells was higher than the other two groups(<i>P</i>=0.005 and <i>P</i>=0.007, respectively).<p>CONCLUSION:The supernatant of limbal stem cells can improve the proliferation of vascular endothelium cell. This may be the unique characteristics of limbal stem cells. This study provides more evidences for the theory of limbal stem cells by determining the functional methods.
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@#LSCs(limbal stem cells)are one of the adult unipotent stem cells located in the Vogt palisade area of the limbal cornea. It can supplement the repair of corneal epithelium by self renewal and play an important role in maintaining normal corneal epithelial integrity, corneal transparency and maintaining normal vision. The lack of corneal limbus stem cells caused by various reasons will lead to corneal turbid, ocular surface vascularization, and final blindness. The main methord to treat related diseases is to cultivate corneal limbal stem cells <i>in vitro</i> and retransplantation. How to effectively expand the limbal stem cells <i>in vitro</i> is the key to the success of the treatment. The location, acquisition methods and expansion methods of limbal stem cells were reviewed.
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Limbal stem cells (LSCs) transplantation is an effective surgical treatment to address limbal stem cells deficiency ( LSCD) resulting from Stevens Johnson's syndrome or corneal chemical burn. LSCs transplantation has been reported as the successful stem cell therapy for bodybuilding, while it still faces a number of daunting challenges. Researchers should pay more attention to substitute seed cell, novel cell carrier materials, effective expansion method,niche modification and immune rejection,so that patients with ocular surface diseases would get fullly therapeutic benefit.
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Objeetive To explore the influence of extracellular high glucose on the proliferation,migration and biomarkers of corneal limbal stem cells.Methods Establishment of a model of high glucose in cultured human limbal stem cells to observe and investigate the effects of extracellular high glucose on the proliferation and migration of corneal limbal stem cells by immunoflurescence,CCK-8 and Transwell assay,respectively.Totally 16 SPF rats were collected and induced diabetic model by streptozotocin as the high-glucose group,and the normal rats of the same age served as the control group.Corneal epidermises of rats in both groups were scraped to observe the repair of corneal epithelium.And the corneas were treated with HE staining and immunohistochemical staining to detect the expression of biomarkers of corneal limbal stem cells and the modality changes of cells.Results The proliferation rate of human limbal epithelial cells was significantly decreased when exposed to high glucose,and the rate at 24 h,48 h and 72 h was 0.728,0.345 and 0.395,respectively,which was markedly lower than that in the control group,with a significant difference (P ＜ 0.05);meanwhile the cell migration rate of the high-glucose group was 17.6％ at 48 h,which was significantly slower than that of the control group (100％).And the inhibition was accompanied by the decreased expression of β-catenin and vimentin.Furthermore,the expression levels of β-catenin and vimentin mRNA and protein were down-regulated,with abnormal location,in the high-glucose group.And diabetic rats had poor corneal epithelial healing.The epithelial layer became thinner and the structures were disorganized in diabetic rats through HE staining.The immunohistochemical assay revealed the expression of β-catenin and vimentin of cornea limbal stem cells was down-regulated in high-glocose group when compared with the control group.Conclusion High glucose can significantly inhibit the proliferation and migration of cornea limbal stem cells,and its main damage mechanism is correlated with the abnormalities of β-catenin and vimentin.
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Objective To investgate the effect of nervous growth factor(NGF) on the proliferation of the limbal stem cells(LSCs) in vitro,and the relationship bewteen expression of its receptors and cell proliferation.Methods After primary cultured,LSCs were divided into the control group and the NGF group.Selected cells cultured of 1 d,3 d and 5 d in the two groups and examined the expression of p63,TrkA,p75 with immunohistochemistry.Results The average gray scale values of expression of p63,TrkA and p75 at 1 d,3 d and 5 d in NGF group were significant decreased compared with the corresponding data in the control group(P<0.05).Pearson's correlations analysis showed that the average gray scale values of expression of TrkA and p63 were of statistically significant differences(P<0.05).Conclusion These results highlight that NGF could maintain the stem cell properties of LSCs.LSCs could exepress the NGF receptors of TrkA and p75,and the expression of TrkA showed a correlation with LSCs proliferation.
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Objective To isolate and culture the rat limbal stem cells ( LSCs) in vitro,and investigate briefly their capacity for transdif-ferentiation into neural stem cells ( NSCs) with cytokine EGF,bFGF and RA.Methods LSCs derived from rats were cultured and identified by immunohistochemistry in vitro.LSCs were induced to differentiate into NSCs in the presence of EGF (20 ng/mL) ,bFGF (10 ng/mL) and with or without of RA(group 1 or group 2)(25 ng/mL) for seven days.Cultures without factors were used as control group.Then the neural marker Nestin of the coultured cells were measured by immunohistology staining.Furthermore,the positive cell rate was counted under micro-scope between the 2 groups and analyzed by statistical software.Results It showed that P63 was positive in LSCs.Nestin in both of the dif-ferentiation groups was positive at the rate of (77.01 ±6.32)%and (84.01 ±5.43)%respectively,of which the second group was higher than the first one (P<0.05).However,it was negative in the control group.A band of Nestin protein from cells was detected by western blot assay.Conclusion LSCs are successfully isolated and cultured in vitro.LSCs could be induced to differentiate into NSCs in the presence of EGF and bFGF.Moreover,the differentiation capability is enhanced in the condition of RA.
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Background The fate of adult stem cells is associated with its surrounding microenviroment.Our previous work found that embryonic stem cells (ESCs) micro-environment enhance the stemness of human limbal stem cells (LSCs),but its mechanism has not been elucidated.Objective This study was to explore the molecular mechanism of ESC micro-environment enhancing the stemness and inhibiting the apoptosis of LSCs.Methods Human LSCs were cultured by explant culture method with CnT-20 medium and CnT-20+20％ ES culture supernatant (ESC-CM),respectively.Colony formation assay was used to analyze the proliferation ability of cells.Telomerase reverse transcriptase (TERT) siRNA (19-25nt siRNA) or siRNA (sc-37007) was transfected into the cells of ESCCM group.Apoptosis and mitochondrial membrane potential were assayed by flow cytometry,and the expressions of telomerase and reactive oxygen species (ROS) in TERT siRNA-or siRNA-F-transfected cells by immunofluorescence and flow cytomery.RT-PCR,immunofluorescence staining and Western blot were employed to determine the expressions of p63,ATP-binding cassette transporer G2 (ABCG2),integrin β1 mRNA and proteins and cytokeratin 3 (C K3) in the cells.The levels of focal adhesion kinase (FAK),Akt,glycogen synthase kinase 3β (GSK3β) and p21 protein and phosphorylation proteins in the cells were detected by Western blot.Results The LSCs presented an increased proliferative capacity and passaged to the eighth generation with the colony-forming efficiency (CFE) of (7.6±0.6) ％ in ESC-CM group,but the cells to the sixth generation with the CFE of (5.6±0.6)％,showing a significant difference between them (t =4.454,P =0.011).The apoptotic rates of the cells from 2 through 6 generations were lower in the ESC-CM group than those in the CnT-20 group (all at P＜0.05).The apoptotic rate of the cells was (7.67± 1.31)％ in the siRNA-F transfected group,which was significantly lower than (32.33 ±3.13)％in the siRNA-TERT transfected group (t =-12.588,P =0.000).No significant differences were seen in the expression levels of p63,ABCG2,integrin β1 mRNA and proteins and TERT protein in the primary cells between the ESC-CM group and the CnT-20 group (all at P＞0.05),but significantly declined expressions of CK3 mRNA and protein were found in the ESC-CM group compared with the CnT-20 group (all at P＜0.01).However,the expressions of p63,ABCG2,integrin β1 mRNA and proteins and TERT protein in the second generation of the cells were significantly higher in the ESC-CM group compared with the CnT-20 group (all at P＜0.01).The telomerase activity was (4.83±0.67) ％ in the siRNA-TERT transfected group,which was significantly lower than (46.71±1.22) ％ of the siRNA-F transfected group (t =52.116,P =0.000).The expression of pFAK,pAkt,pGSK3β proteins were weakened,but the expression of p21 was increased in the ESC-CM group after addition of FAK inhibitor,GSK3β inhibitor and TERT-siRNA transfected group.Mitochondrial membrane potential in the second generation of cells was elevated in the ESC-CM group in comparison with the CnT-20 group and the siRNA-TERT transfected group (all at P＜0.01),and the rates of ROS positively reaction was lower in the ESC-CM group and the siRNA-F transfected group than those of the CnT-20 group and siRNA-TERT transfected group (all at P＜0.01).Conclusions ESC-CM culture system can effectively keep the stemness of LSCs and inhibit apoptosis.ESC-CM culture system plays functions probably via telomerase-p21-mitochondrial axis and the activation of the FAK/Wnt signaling pathways.
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Background: Cultivated limbal epithelium for reconstruction of corneal surface is a well-established procedure; however, it is not adequate for damage which also extensively involves the conjunctiva. In severe cases of ocular surface damage that warrant additional conjunctival transplantation apart from cultivated limbal stem cell transplantation, we describe the long-term survival of a novel method of cocultivating autologous limbal and conjunctival epithelium on a single substrate. Materials and Methods: Forty eyes of 39 patients with severe limbal stem cell deficiency and conjunctival scarring or symblepharon underwent transplantation of autologous cocultivated epithelium on human amniotic membrane. A ring barrier was used to segregate the central limbal and peripheral conjunctival epithelia in vitro. Patients were followed up at regular intervals to assess stability of the ocular surface, defined by absence of conjunctivalization into the central 4 mm of the cornea and absence of diffuse fluorescein staining. Penetrating keratoplasty (PKP) was subsequently performed, where indicated, in patients with surface stability. Results: The cumulative survival probability was 60% at 1 year and 45% at 4 years by Kaplan–Meier analysis (mean follow-up duration: 33 ± 29 months, range: 1–87 months). Best-corrected visual acuity improved to greater than 20/200 in 38% eyes at the last follow-up, compared with 5% eyes before surgery. Immunohistochemistry in five of the corneal buttons excised for PKP showed an epithelial phenotype similar to cornea in all five. Conclusions: Synchronous use of cultured limbal and conjunctival epithelium offers a feasible alternative and a simpler one-step surgical approach to treat severe ocular surface disorders involving limbus and conjunctiva.
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Background & objectives: Ex vivo expansion of the limbal epithelial cells activates the nerve growth factor (NGF) mediated downstream signal transduction pathway. It is not clear as to which factors control the stemness of the corneal limbal stem cells, i.e., the maintenance of stem cell properties. It is likely that various signaling pathways are involved, including Notch, Wnt and NGF signaling, etc. In the present study, we investigated the activation of phosphoinositide-3-kinase (PI3K) pathway on cells cultured over the chitosan matrix, chitosan silver matrix, chitosan gold matrix, intact and denuded human amniotic membrane (HAM). Methods: Human limbal biopsies obtained from the cadaveric donor eyes were used in this study. The cells cultured over different substrate and observed for the activation of the downstream signaling molecules of PI3K/Akt/FKHRL1 pathway. Western blotting was done to prove the results. Results: The cells cultured over the intact HAM showed the activation of the downstream signaling molecules of PI3K/Akt/JNK pathway compared to the cells grown over other substrates. On inhibition of the PI3K activity there was absence of phosphorylation of downstream effectors in the limbal epithelial cells from the explant culture over the intact HAM. Interpretation & conclusions: The ex vivo expansion of human limbal epithelial progenitor cells on intact HAM was mediated by PI3K/Akt/FKHRL1 pathway, which is known to govern cell survival, and the mitogen-activated protein kinase (MAPK) pathway, known to control the cell mitosis.
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A 62-year-old man presented with severe bilateral ocular surface chemical injury and history of failed penetrating keratoplasty of right eye in 1996. Visual acuity was hand movement in right eye and light perception in left eye. Staged procedures of limbal stem cells allograft followed by penetrating keratoplasty have been done and resulted in good ocular surface restoration and rehabilitation of vision in right eye.
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Objective To compare the therapeutic effects of recurrent pterygium treated by microsur- gical management.Methods Sixty-two cases(67 eyes)with recurrent pterygiumwere randomly divided into limbal stem cell autograft transplantation comblined with mitomycin C(34 cases38 eyesand limbal stem cell autograft transplantation(28 cases29 eyes)The post operative follow-up period was 6 to 30 months.Re- suits One eye recurrence was noted in the trial groupthe recurrent rate was 2.63%Three eyes recurrence was noted in the control groupthe recurrent rate was 10.34%There was statistical significant difference be- tween two groups(P0.05).Conclusion Limbal stem cells autograft transplantation combined with mitomycin C can decrease the recurrent rate.It is an ideal methods of recurrent pterygium surgical procedureis worth spreading.