Corneal stromal mesenchymal stem cells: reconstructing a bioactive cornea and repairing the corneal limbus and stromal microenvironment

Corneal stroma-derived mesenchymal stem cells(CS-MSCs) are mainly distributed in the anterior part of the corneal stroma near the corneal limbal stem cells(LSCs). CS-MSCs are stem cells with self-renewal and multidirectional differentiation potential. A large amount of data confirmed that CS-MSCs can be induced to differentiate into functional keratocytes in vitro, which is the motive force for maintaining corneal transparency and producing a normal corneal stroma. CS-MSCs are also an important component of the limbal microenvironment. Furthermore, they are of great significance in the reconstruction of ocular surface tissue and tissue engineering for active biocornea construction. In this paper, the localization and biological characteristics of CS-MSCs, the use of CS-MSCs to reconstruct a tissue-engineered active biocornea, and the repair of the limbal and matrix microenvironment by CS-MSCs are reviewed, and their application prospects are discussed.

The Limbal Niche and Its Role in Maintaining Corneal Regeneration

In recent years, stem cells have been a focal point in research designed to evaluate the efficacy of ophthalmologic therapies, specifically those for corneal conditions. The corneal epithelium is one of the few regions of the body that maintains itself using a residual stem cell population within the adjacent limbus. Stem cell movement has additionally captivated the minds of researchers due to its potential application in different body regions. The cornea is a viable model for varying methods to track stem cell migratory patterns, such as lineage tracing and live imaging from the limbus. These developments have the potential to pave the way for future therapies designed to ensure the continuous regeneration of the corneal epithelium following injury via the limbal stem cell niche. This literature review aims to analyze the various methods of imaging used to understand the limbal stem cell niche and possible future directions that might be useful to consider for the better treatment and prevention of disorders of the cornea and corneal epithelium. .

Ocular injuries by durian fruit

·AIM:To report various ocular injuries caused by durian fruit. ·METHODS:Three cases of ocular injuries were described in young patients, due to accidental fall of durian fruit on the forehead and face, while they were taking rest/sleeping /playing under the durian tree. ·RESULTS:The ocular injuries observed were lacerating injury of cornea with iris incarceration, hyphema, superficial penetrating injury of sclera and angle recession glaucoma in the right eye of first patient; lacerating injury of cornea with iris prolapse in the left eye of second patient; subconjunctival haemorrhage, traumatic mydriasis and superficial penetrating injury of sclera, commotion retinopathy and macular edema in the left eye of third patient. Vision improved to normal in all the eyes following surgical/ medical/optical treatment. ·CONCLUSION:Evidence of penetrating injury (because of thorns) and blunt injury (because of weight) can be seen in the eyes when durian fruit falls on the face. Vision can be recovered fully with immediate and appropriate treatment in these cases. The ocular injuries can be prevented by educating the public to wear protective metal frame wide goggles and not to sleep/take rest under the durian tree.

Research progress on animal models of corneal epithelial-stromal injury

A corneal epithelial-stromal defect is recognized as a major contributor to corneal scarring.Given the rising prevalence of blindness caused by corneal scarring,increasing attention has been focused on corneal epithelialstromal defects.Currently,the etiology and pathogenesis of these defects remain inadequately understood,necessitating further investigation through experimental research.Various modeling methods exist both domestically and internationally,each with distinct adaptive conditions,advantages,and disadvantages.This review primarily aims to summarize the techniques used to establish optimal animal models of corneal epithelial-stromal injury,including mechanical modeling,chemical alkali burns,post-refractive surgery infections,and genetic engineering.The intention is to provide valuable insights for studying the mechanisms underlying corneal epithelial-stromal injury and the development of corresponding therapeutic interventions.

角巩膜缘与角膜切口超声乳化术治疗老年白内障疗效对比

目的 对比角巩膜缘与角膜切口超声乳化术治疗老年白内障的临床疗效。方法 随机将80例老年白内障患者分为2组。2组均行超声乳化术,对照组采用角膜切口,观察组采用角巩膜缘切口。比较2组上皮修复时间、切口修复时间、裸眼视力、最佳矫正视力(BCVA)、角膜散光度、手术源性角膜散光度、泪囊破裂时间(BUT)、角膜荧光素染色评分(FLCs)、泪液分泌试验(SIt)水平以及术后并发症发生率。结果 观察组上皮修复、切口修复时间均早于对照组(P<0.05);观察组术后1周、2周、4周时裸眼视力、BCVA水平以及BUT、Sit水平均高于对照组(P<0.05);观察组术后1周、2周、4周时角膜散光度、手术源性角膜散光度以及FLCs水平水平均低于对照组(P<0.05);2组术后并发症发生率(18%与12%)差异无统计学意义(P>0.05)。结论 角巩膜缘切口超声乳化术治疗老年白内障临床疗效较好,患者术后角膜散光度较低,且视力、泪膜功能稳定性恢复较好。

Effects of amniotic membrane transplantation on cytokines expression in chemically burned rat corneas

AIM: To investigate the effect of amniotic membrane transplantation (AMT) on the expressions of inflammatory-related, angiogenic-related and growth-related cytokines in rat corneas after chemical injury. METHODS: Alkali wounds were inflicted on the central corneas of rats by applying a round filter paper soaked in 1mol/L NaOH for 40 seconds. One week after alkali burn, 12 rats were randomly divided into 2 groups: the AMT group and the control group, and AMT was performed on the rats in the AMT group. Corneal opacity and neovascularization were observed by slit-lamp microscopy. The protein levels of interleukin (IL)-2, interferon (IFN)-gamma, IL-10 and transforming growth factor (TGF)-beta were determined by enzyme-linked immunosorbent assay 2 weeks after AMT. The mRNA levels of matrix metalloproteinase-2 (MMP-2), vascular endothelial growth factor (VEGF), epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) were evaluated by real-time quantitative PCR. RESULTS: In the AMT group, the corneal opacity was improved (P =0.011) and the area of corneal neovascularization was significantly decreased (P=0.005) compared with the control group. The amount of IL-2 and IFN-gamma secreted by Th1 cells were decreased after AMT, whereas the amount of IL-10 and TGF-beta secreted by Th2 cells were increased ( P <0.05). The level of MMP-2 was significantly down-regulated (P=0.013) at the nnRNA level in the AMT group, while the expression of EGF was significantly higher (P= 0.022) compared with the control. CONCLUSION: AMT may suppress corneal neovascularization after chemical injury by modulating the expressions of soluble factors.

Expression of VEGF-C in Rat Cornea after Alkali Injury

The expression of VEGF-C and molecular mechanisms of lymphangiogenesis in rat cornea after alkali injury was studied. The rat alkali injured corneal models were made. Under electron microscopy, the lymphatic vessels in the rat injured corneas were examined. The expression of VEGF-C proteins was detected by using immunohistochemical assay at day 1, 3, 5, 7, 9 after injury. The expression levels of VEGF-C mRNA were quantified with reverse transcription polymerase chain reaction (RT-PCR). The results showed that the lymphatic vessels were found in the injured rat corneas 14 days after the injury. The VEGF-C protein was detectable 3 days after injury, reached the peak 5 days after injury, and gradually decreased. In the control group, no VEGF-C proteins were detected. The VEGF-C mRNA was minimally detected in the normal rat corneas, but it was highly expressed 5 days after the injury. The difference was statistically significant. It was concluded that VEGF-C might be one of the most important relevant factors in corneal lymphangiogenesis after alkali injury.

Anti-inflammatory potential of human corneal stroma-derived stem cells determined by a novel in vitro corneal epithelial injury model

BACKGROUND An in vitro injury model mimicking a corneal surface injury was optimised using human corneal epithelial cells(hCEC).AIM To investigate whether corneal-stroma derived stem cells(CSSC) seeded on an amniotic membrane(AM) construct manifests an anti-inflammatory, healing response.METHODS Treatment of hCEC with ethanol and pro-inflammatory cytokines were compared in terms of viability loss, cytotoxicity, and pro-inflammatory cytokine release, in order to generate the in vitro injury. This resulted in an optimal injury of 20%(v/v) ethanol for 30 s with 1 ng/mL interleukin-1(IL-1) beta. Co-culture experiments were performed with CSSC alone and with CSSC-AM constructs.The effect of injury and co-culture on viability, cytotoxicity, IL-6 and IL-8 production, and IL1 B, TNF, IL6, and CXCL8 mRNA expression were assessed.RESULTS Co-culture with CSSC inhibited loss of hCEC viability caused by injury. Enzyme linked immunosorbent assay and polymerase chain reaction showed a significant reduction in the production of IL-6 and IL-8 pro-inflammatory cytokines, and reduction in pro-inflammatory cytokine mRNA expression during co-culture with CSSC alone and with the AM construct. These results confirmed the therapeutic potential of the CSSC and the possible use of AM as a cell carrier for application to the ocular surface.CONCLUSION CSSC were shown to have a potentially therapeutic anti-inflammatory effectwhen treating injured hCEC, demonstrating an important role in corneal regeneration and wound healing, leading to an improved knowledge of their potential use for research and therapeutic purposes.

Effect of Seawater Soaking on Explosive Cornea Injury

Objective: To observe the changes in corneal tissue after explosive corneal injury and the effect of seawater soaking on the healing of cornea after explosive injury with the help ofoptical microscope. Methods: Make 10 similar explosive injury models of rabbit′s eyeball using 10 adult greyrabbits. For each rabbit, both eyes are artificially injured through explosion; its right eye is the comparison eye and after the injury the left eye is soaked in seawater for 30 minutes.Conduct slit lamp examinations, fluorescein dyeing, and ultrasonic cornea thickness metering on the corneas at intervals, that is, before the injury, 1 , 2, 3, 4, 5, 7, 9, 12, 15days after the injury respectively. Examine the corneas under the optical microscope andmake comparisons.Results: The corneal thickness of the experiment eye becomes apparently thicker than thatof the comparison eye after the injury. The corneal clouding of the former apparentlyaggravates compared with the latter. The healing of corneal epithelium in the injured eye isslower compared with that in the comparison eye. Comparative examination under the opticalmicroscope shows: after the injury, the corneal tissue of the experiment eye changesdistinctly, its healing is delayed and the vascularization degree becomes high in cornealstroma.Conclusion: Scars and vascularization of various degrees will appear in corneal stroma afterexplosive injuries. Seawater soaking will have bad effects on the healing of the corneainjured in explosions.

Riboflavin/ultraviolet A-induced collagen cross-linking in rabbit corneal scar

AIM: To evaluate the biomechanical stability of the corneal scar treating with riboflavin and ultraviolet A(UVA). METHODS: Totally 86 New Zeal rabbits were divided into control group(group A, n=8) and trauma groups [group B(n=27), group C(n=24) and group D(n=27)]. Then groups B, C and D were divided into three sub-groups according to the time points of sacrifice, i.e. groups Ba, Ca and Da(4 wk, n=8); Bb, Cb and Db(6 wk, n=8); Bc(n=11), Cc(n=8) and Dc(8 wk, n=11). The right corneas of these 78 rabbits in the trauma groups were penetrated. Group B were only sutured. Group C were treated with corneal cross-linking(CXL) immediately after suturing. Group D were treated with CXL seven days after suturing. The corneal scar strips of 4.0×10.0 mm2 were cut and the stress and Young's modulus at 10% strain were evaluated. Samples from the three rabbits of group Bc and three of group Dc were used to measure the expression of alpha smooth muscle action(α-SMA). RESULTS: The mechanical strength of the corneal scar increased with time, and was strongest at 8 wk after the injury. The ultimate stress of corneal scar(group D) were 2.17±0.52 MPa, 2.92±0.63 MPa, and 4.21±0.68 Mpa at 4 wk, 6 wk and 8 wk, respectively; Young's modulus were 10.94±1.57 MPa, 11.16±2.50 MPa, and 13.36±2.10 Mpa, which were higher than that of other groups except for normal control. The expression of α-SMA in group B and group D were 0.28±0.11 and 0.65±0.20, respectively, and the difference was statistically significant(P=0.048). CONCLUSION: CXL with riboflavin/UVA at seven days after suturing improved the biomechanical properties of corneal scars most effectively in the present study.

An Analysis of 1 001 Blinding Patients with Corneal Disease in 1960-1989

Purpose:To investigate the condition and change of corneal blindness (CB) in the past 30 years.Methods: 1 001 blinding patients of corneal disease were clinically analysed from 1960 to 1989,including etiology, sex, age, occupation, difference between the urban and rural areas etc.Results: The male predominated. Infection stood the first cause of blinding corneal diseases, followed by trauma,malnutrition and the others.HSV-1 keratitis was the most frequent infection in 1980s. The highest incidence of CB was 20-40 year -old. The number of CB caused by occupational truauma was more than that by ordinary trauma in 1960s,but from 1970s, the latter was more than the former. The number of CB in rural area was more than that in the urban in 1960s,but from 1970s,there was no significant difference between the two areas. The percentage of bilateral CB was gradually decreased.Conclusion: We should try to find more effective strategies to prevent and treat CB caused by HSV-1 keratitis, pay special attention to

角膜绷带镜不能按计划连续佩戴的原因及临床特点分析

目的 分析角膜绷带镜在临床使用中出现不能按计划连续佩戴的原因及临床特点。方法 回顾性分析2020年6月至2021年6月就诊于空军军医大学西京医院眼科角膜及眼表专科患者200例(223眼),因不同原因佩戴角膜绷带镜以辅助治疗的病历资料。分析角膜绷带镜佩戴后发生的不能按计划连续佩戴的原因、发生率及其与疾病的关系。结果 佩戴角膜绷带镜以辅助治疗的患者主要是翼状胬肉术后及多种非感染性眼表损伤患者,双眼23例,单眼177例,平均年龄(53.87±12.51)岁;不能按计划连续佩戴角膜绷带镜者共22眼(9.87%),包括翼状胬肉术后6眼和非感染性眼表损伤16眼。不能按计划佩戴角膜绷带镜的原因为神经营养性角膜病变和早期眼化学伤引起的镜片自行脱落(14眼,6.28%);广泛角膜上皮糜烂的药源性眼表损伤导致的戴镜后眼部不适感加重而要求摘镜(5眼,2.24%);戴镜后出现继发性角膜感染(3眼,1.35%)。结论 角膜绷带镜在治疗眼表角膜疾病中存在一定的不能按计划连续佩戴的现象,在使用后需要随访观察。

An updated narrative review of treatment for limbal epithelial stem cell deficiency

Background and Objective:Nearly 30 years have passed since limbal stem cell deficiency(LSCD)was first identified by pioneers and given clinical attention.LSCD remains a difficult disease to treat.It can potentially lead to blinding.At present,understanding of limbal stem cells(LSCs)has deepened and various treatment options for LSCD have been devised.The objective of this review is to summarize basic knowledge of LSCD and current treatment strategies.Methods:PubMed search was performed to find studies published in English on LSCs and LSCD including original reports and reviews.Literatures published from 1989 to 2022 were reviewed.Key Content and Findings:LSCs are enigmatic stem cells for which no specific marker has been discovered yet.Although LSCD is not difficult to diagnose,it is still challenging to treat.An important advancement in the treatment of LSCD is the provision of guidelines for selecting systematic surgical treatment according to the patient’s condition.It is also encouraging that stem cell technologies are being actively investigated for their potential usefulness in the treatment of LSCD.Conclusions:Although various treatment options for LSCD have been developed,it should be kept in mind that the best chance of treatment for LSCD is in the early stage of the disease.Every effort should be made to preserve as many LSCs as possible in the early treatment of LSCD.

An interferometric ex vivo study of corneal biomechanics under physiologically representative loading,highlighting the role of the limbus in pressure compensation

Background:The mechanical properties of the cornea are complex and regionally variable.This paper uses an original method to investigate the biomechanics of the cornea in response to hydrostatic loading over the typical physiological range of intraocular pressure(IOP)fluctuations thereby increasing understanding of clinically relevant corneal biomechanical properties and their contributions to the refractive properties of the cornea.Methods:Displacement speckle pattern interferometry(DSPl)was used to measure the total surface displacement of 40 porcine and 6 human corneal-scleral specimens in response to pressure variations up to 1 mmHg from a baseline of 16.5 mmHg.All specimens were mounted in a modified artificial anterior chamber(AAC)and loaded hydrostatically.Areas of high strain in response to loading were identified by comparing the displacements across different regions.Results:The nature of the response of the corneal surface to loading demonstrated high regional topographic variation.Mechanical properties were shown to be asymmetrical,and deformation of the limbal and pre-limbal regions dominated these responses respectively with over 90%(nasal-temporal)and 60%(superior-inferior)of the total maximum displacement occurring in these regions indicating high strain.In contrast,the curvature of the central cornea remained relatively unchanged merely translating in position.Conclusions:The limbal and pre-limbal regions of the cornea appear to be fundamental to the absorption of small pressure fluctuations facilitating the curvature of the central cornea to remain relatively unchanged.The differential mechanical properties of this region could have important implications for the application of corneal surgery and corneal crosslinking,warranting further investigation.

An interferometric ex vivo study of corneal biomechanics under physiologically representative loading,highlighting the role of the limbus in pressure compensation

Background:The mechanical properties of the cornea are complex and regionally variable.This paper uses an original method to investigate the biomechanics of the cornea in response to hydrostatic loading over the typical physiological range of intra-ocular pressure(IOP)fluctuations thereby increasing understanding of clinically relevant corneal biomechanical properties and their contributions to the refractive properties of the cornea.Methods:Displacement speckle pattern interferometry(DSPI)was used to measure the total surface displacement of 40 porcine and 6 human corneal-scleral specimens in response to pressure variations up to 1 mmHg from a baseline of 16.5 mmHg.All specimens were mounted in a modified artificial anterior chamber(AAC)and loaded hydrostatically.Areas of high strain in response to loading were identified by comparing the displacements across different regions.Results:The nature of the response of the corneal surface to loading demonstrated high regional topographic variation.Mechanical properties were shown to be asymmetrical,and deformation of the limbal and pre-limbal regions dominated these responses respectively with over 90%(N-T)and 60%(S-I)of the total maximum displacement occurring in these regions indicating high-strain.In contrast,the curvature of the central cornea remained relatively unchanged merely translating in position.Conclusions:The limbal and pre-limbal regions of the cornea appear to be fundamental to the absorption of small pressure fluctuations facilitating the curvature of the central cornea to remain relatively unchanged.The differential mechanical properties of this region could have important implications for the application of corneal surgery and corneal crosslinking,warranting further investigation.

自体角膜缘干细胞移植与常规翼状胬肉切除术疗效比较

目的通过翼状胬肉单纯切除术及翼状胬肉切除加自体角膜缘干细胞移植术2种手术方式治疗翼状胬肉,比较其治愈及复发情况及角膜上皮修复时间,评价其疗效。方法将86例(88眼)翼状胬肉患者随机分为翼状胬肉单纯切除术组(A组)43例44眼及翼状胬肉切除加自体角膜缘干细胞移植术组(B组)43例44眼,组间比较采用χ2检验比较其疗效。结果A组10例10眼复发,B组仅有1例1眼复发,2组比较差异有统计学意义(P<0.05);A组角膜上皮修复时间平均为(5.15±0.92)d,B组角膜上皮修复时间平均为(3.36±0.82)d,2组比较差异有统计学意义(P<0.05)。结论翼状胬肉切除加自体角膜缘干细胞移植术可以降低复发率,缩短角膜上皮修复时间,是治疗翼状胬肉较为安全有效的方法。

正常角膜缘组织中与血管生成相关的生长因子及其受体的表达

目的探查正常人角膜缘组织里与血管生成相关的生长因子及其受体的分布。方法用冰冻切片和链亲合素过氧化物酶染色系统，选择一组兔抗人多克隆抗体，即：血管内皮生长因子（ＶＥＧＦ）、碱性成纤维细胞生长因子（ｂＦＧＦ）、血小板源性生长因子（ＰＤＧＦ－Ａ）和转化生长因子－β １（ＴＧＦ－β １）及其受体对１１例角膜移植供体材料的角膜缘组织进行了检查。结果ＶＥＧＦ，ｂＦＧＦ，ＰＤＧＦ－Ａ和ＴＧＦ－β １ ４种生长因子仅在１或２例标本中不表达外，在大部分标本中表达阳性。４种生长因子的受体在角膜缘上皮、基底膜、上皮下组织和角巩膜组织内阳性表达分别是：ＶＥＧＦ ９例、７例、１０例和８例；ｂＦＧＦ ８例、９例、９例和８例；ＰＤＧＦ ５例、４例、１０例和７例；ＴＧＦ－β １３例、０例、１１例和６例。结论在正常角膜缘组织中，存在ＶＥＧＦ，ｂＦＧＦ，ＰＤＧＦ－Ａ和ＴＧＦ－β １及其受体，可能在维持正常的角膜功能和调节角膜缘组织的生长和增殖中起着重要作用。

HLA匹配角膜缘干细胞联合羊膜移植修复翼状胬肉

背景:HLA匹配的角膜缘干细胞联合羊膜移植治疗翼状胬肉是眼科医生治疗的新理念,该方法治疗后并发症少、康复快、复发率低,尤其是能减少移植排斥反应等已被广泛接受。目的:探讨HLA匹配的角膜缘干细胞联合羊膜移植治疗翼状胬肉疗效。方法:选取2013年3月1日至2014年4月30日行HLA匹配的角膜缘干细胞联合羊膜移植治疗的翼状胬肉患者47例(47眼)作为试验组;回顾性分析2010年3月1日至2012年12月1日采用单纯翼状胬肉切除方法治疗的翼状胬肉患者40例(47眼)作为对照组,对两组患者治愈和复发情况进行比较。结果与结论:试验组47例,全部顺利完成治疗,无并发症,其中1例切口感染,特殊对症治疗后均已好转出院,全部患者均无移植排斥反应。试验组治愈45眼,复发2眼,复发率为4.3%。对照组47眼中治愈32眼,复发15眼,复发率31.9%。结果表明HLA匹配的角膜缘干细胞联合羊膜移植治疗翼状胬肉并发症少、复发率低,是治疗翼状胬肉一种行之有效的方法。

翼状胬肉模型的建立

目的人为地造成局部角膜缘干细胞缺损以及慢性结膜炎症的眼表模型,探讨翼状胬肉的形成原因。方法40只新西兰大白兔随机分为A、B、C、D 4组,设右眼为实验眼,左眼为对照眼。采用手术切除1个象限角巩膜缘组织的方法产生角膜缘干细胞缺损模型,眼部以稀盐酸(A、B组)或滑石粉(C、D组)定期点眼产生慢性角结膜炎模型。实验眼在同一眼上观察同时产生角膜缘干细胞缺损和慢性角结膜炎模型后的形态学变化,对照眼分别采用两种模型之一的相应形态学改变。于2、4、8、12周每只眼记录角膜上皮缺损、球结膜充血和结膜新生物情况,评分,并随机每组取材2只作病理。结果切除角膜缘干细胞的兔眼,可以在短期内在切除角膜缘干细胞的方向上出现类胬肉增殖组织样改变。类胬肉增殖组织的形成、程度和是否有局部的炎症反应以及角膜上皮完整性的破坏呈正相关性。结论局限性角膜缘干细胞的缺损和长期慢性炎症刺激的同时作用可能是翼状胬肉形成的主要原因。

角膜缘干细胞自体移植联用羊膜治疗复发性翼状胬肉

目的:观察自体角膜缘干细胞移植联合羊膜移植治疗复发性翼状胬肉的临床疗效。方法:自体角膜缘干细胞移植联合羊膜移植治疗复发性翼状努肉30例,术后随访1年,观察胬肉复发及手术并发症情况。结果:无复发及睑球粘连等并发症发生。结论:自体角膜缘干细胞移植联合羊膜移植治疗复发性翼状胬肉是安全有效的治疗方法。