Ocular stem cells: a narrative review of current clinical trials

INTRODUCTION

According to the World Health Organization (WHO),globally, at least 2.2 billion people have vision impairment. In almost half of these cases, vision impairment could have been prevented or has yet to be addressed. More than 400 million people are visually impaired due to agerelated macular degeneration (AMD), glaucoma, diabetic retinopathy, Stargardt disease, and retinitis pigmentosa.People suffering from these visual disorders are destined to increase dramatically in the next decades, due to population growth and aging. One of the most promising and hopeful potentials for future treatment is stem cell therapy.

Stem cells are the future of cell-based therapies due to their property to regenerate. Stem cell therapy is considered the treatment option of the 21Century for many health conditions such as hematological disorders, Parkinson’s disease, diabetes,and myocardial failure. Nowadays the eye has been verified as an ideal target of stem cell-based therapies and the perfect ascertainment field of their efficacy and safety. This is due to the facility of administration and effects evaluation, using state of the art ophthalmic devices and machinery.

This review aims to summarize scientific evidence of the present and future prospects of stem cells for ocular therapy.

Stem cells, based on their origin, have been classified into embryonic stem cells (ESCs) and adult stem cells. These cells are characterized by their property of self-renewal, under appropriate conditions, and by their additional ability to differentiate into the originating tissues.The stem cells, because of their pluripotency, bear a huge potential for therapeutic use in numerous pathologies. ESCs are pluripotent, while adult stem cells are multipotent.However, the current technological advances in science have achieved to induce pluripotency to adult stem cells.

Pluripotent stem cells have the ability to evolve to any cell type. These can be classified into embryonic, perinatal or induced pluripotent stem cells (iPSCs), based on the tissue of origin. Human induced pluripotent stem cells (hiPSCs)are derived from somatic cells exposed to reverting pattern techniques of gene expression that eventually undergo reprogramming to different specialized cell types. The capability to generate hiPSCs using a patient’s own cells originated a brand-new field of patient-specific therapies,always keeping in mind that undifferentiated iPSCs showcase a potential risk of tumorigenesis to the recipient.

During the past 20y, significant progress has been made in the treatment of corneal endothelium dysfunction. The global shortage of donor corneas as well as corneal graft rejection and failure remain important limiting factors to corneal transplant surgery. Consequently, novel treatments have been developed including endothelial keratoplasty techniques as well as the recently introduced promising approaches of cellbased therapies and regenerative medicine. Cell-based therapies include thecultivation of human corneal endothelial cells (CECs) obtained either from the patient’s own endothelium or from donor corneas. The cultured CECs can subsequently be transferred to recipients’ diseased corneas.Rho-associated kinases (ROCK) inhibitors have also been used in clinical use, both as eye drops and as adjunct drugs in cell-based therapies, for the treatment of corneal endothelium dysfunction, as they enhance cell proliferation as well as the engraftment of CECs onto recipient corneas. Finally, recently introduced surgical techniques in regenerative medicine,for the treatment of corneal endothelial diseases, include descemetorrhexis without endothelial keratoplasty (DWEK)/Descemet’s stripping only (DSO) and Descemet’s membrane transplantation (DMT). Despite the theoretical advantages of these techniques, larger randomized comparative studies with long-term follow-up are required to establish their effectiveness.

試驗(yàn)結(jié)果表明，試驗(yàn)件在進(jìn)行5個(gè)階段試驗(yàn)后，每一次的無(wú)損檢測(cè)結(jié)果都為合格，試驗(yàn)全部完成后，關(guān)鍵尺寸ФA在試驗(yàn)前后變化量最大為0.01%，尺寸ФB在試驗(yàn)前、后基本不變，說(shuō)明該渦輪盤具有5 200次的低循環(huán)疲壽命。

Stem cells are currently being trialed in a variety of ocular conditions and diseases. Sheets of corneal-limbal stem cells cultivated on fibrin matricesare successfully employed in ocular surface impairment due to limbal cell loss. The potential role and contingent success of stem cells are also currently trialed in AMD and inherited retinal diseases, being currently in phase I and Ⅱ, for which safety has already been proven.

However, olfactory mucosa mesenchymal stem cells (OMMSCs) not only behave in a similar way to BMSCs with regard to pluripotent abilities, but also demonstrate convenient isolation from the nasal lamina propria as well as high proliferation rate and safety. Additional advantages of OMMSCs include the ability of autologous transplants to avoid immune rejection as well as lack of ethical, moral and societal issues and the absence of gene mutations.

AMD is the most common cause of blindness that usually affects people aged 55y and older.Additionally, inherited retinal diseases, such as Stargardt’s macular dystrophy and retinitis pigmentosa, have a noteworthy impact on the quality of life. They are characterized by loss of photoreceptors, neurons, retinal pigment epithelium (RPE),and choroidal alterations. For these conditions treatment options are very limited or nonexistent, resulting in irreversible vision loss.

Despite the lack of specific markers for the quantification of stem cells in donor cultures, it was initially thought that corneal clarity could be restored by simply replacing lost LESCs.Nevertheless, the stem cell replacement mechanism has been recently addressed. Follow-up studies, demonstrated the absence of donor’s cells detection, 9mo after transplantation.These results firstly indicate that long-term immunosuppression is unnecessary and secondly that the replacement theory is not the underlying mechanism. It is more likely that reconstruction of defective limbal epithelial crypts occurs by providing a microenvironment suitable for the resettlement of endogenous stem cells.

Clinical trials based on ocular surface stability and good vision indicated positive outcomes forexpansion of autologous LESCs to treat LSCD. These trials showed differences in follow-up periods, some of them reporting results of more than 2y after transplantation. Considering the transparency of corneal epithelium, scarcity of conjunctivalization and vascularization, as well as bestcorrected visual acuity, the overall success rate was more than 80%.

Sangwaninvestigated the results of these procedures on the ocular surface of donor eyes. Following autologous limbal biopsy procedures, all donor sites showed complete healing as well as total absence of ocular surface disease and impairment,within a maximum of two weeks. In a retrospective clinical trial by Fasoloon cases treated with autologous LESCs transplantation, indicated that 60% showed adverse events including inflammation and corneal epithelial defects.Nevertheless, these results were debatable given the relatively short duration of follow-up. On the other hand, Ramaperformed clinical trials on 113 eyes that had suffered chemical or thermal burns with resultant LSCD, and were consequently treated with stem cell transplantationa fibrin carrier substrate. The results reported no persevering adverse effects during a decade follow-up (Figure 1). Meanwhile,using a contact lens (CL) as a carrier substrate for LSCT may be complicated by the advent of CL displacement, that necessitates the use of a supplemental CL which is a minor impediment.

Ocata Therapeutics (formerly known as Advanced Cell Techonology) performed clinical trials using transplantation of hESC-derived RPE cells for Stargardt’s disease and AMD. Since ESC-derived RPE cell transplantation is an allograft transplantation, this clinical trial was combined with immunosuppressive drug administration for 3mo. One patient developed post-surgery bacterial endophthalmitis, which was relieved after 2mo with the infusion of antibiotics and eye drops. However, the causative organism was not detected in the transplanted cells. Other reported systemic adverse events were due to the combined use of immunosuppressive drugs. No abnormal proliferation or inflammatory reaction of the transplanted cells was observed. The results of transplantation revealed an increased pigmentation at the RPE cell transplantation site, which was thought to be due to the proliferation of RPE cells at the transplantation site.

Over the last decades several treatment options have been developed to manage patients with LSCD. In 2015,Holoclar, Italy (manufactured by Holostem Terapie Avanzate S.R.L. in Italy, and originally developed by Professors Graziella Pellegrini and Michele De Luca) is an advanced therapy medicinal product (ATMP) and became the first stem cell-based treatment registered and approved by a Regulatory Body. Whilst not yet available in the USA, its use in National Health Service in the UK is approved and strictly regulated based upon retrospective data. Holoclar is a cellbased tissue engineered therapy and uses the patient’s own LESCs (autologous) from the unaffected eye to form a graft grown outside the body () for the treatment of LSCD.Another interesting application of cell-based treatment is the use of autologous mesenchymal stem cells (MSCs), isolated from limbal biopsies to treat cases of corneal stromal scarring thus reducing the need for corneal grafts LESCs.

(1) 木楠村樣品點(diǎn)1-3、1-5、1-6表現(xiàn)為相對(duì)舒緩平滑的單峰，表明沉積物的分組簡(jiǎn)單，可能只表現(xiàn)為一種組分的粒度特征，或兩種不同地點(diǎn)的組分經(jīng)歷了穩(wěn)定持續(xù)的搬運(yùn)作用而充分混合，整體呈現(xiàn)均一平緩的粒度特征。

Furthermore, stem cells can be autologous or allogeneic.When the source is a donor tissue, additional factors must be considered regardless of whether stem cells are tissue-specific or pluripotent. These include infectious agents screening, and immunosuppression due to transplant rejection threatening.Another factor to be considered is the arithmetic adequacy of harvested material without affecting the functionality of donor ocular tissues. The objective is to ensure safe and functional stem cell engraftment, by delivering sufficient cell numbers and trying to eliminate oncogenic mutations. Moreover, it is extremely important to provide correct differentiation pathways to obtain the covetable cell type at the requested level of development.

管線點(diǎn)的測(cè)量精度，管線測(cè)量控制網(wǎng)宜分級(jí)布設(shè)導(dǎo)線網(wǎng)，管線點(diǎn)的解析坐標(biāo)中誤差(指測(cè)點(diǎn)相對(duì)鄰近解析控制點(diǎn))不大于±5cm，地下管線圖上測(cè)量點(diǎn)位中誤差不大于±0.5 mm。

The anterior lens surface is lined by a single layer of epithelial cells, which is located between the lens capsule and the lens fibers. The epithelial cells show metabolic properties that sustain the physiological health of the lens, and work as a source of stem cells. These cells are liable for lens growth throughout life. In cases of incomplete removal during cataract surgery, they often migrate to the posterior lens area causing posterior capsular bag opacification as a result of fibrotic changes they incur in. So far, the only treatment for this condition is posterior laser capsulotomy, resulting to be the second most common ophthalmic proceeding worldwide.Nevertheless, these residual cells can occasionally regenerate lens fiber at various stages of transparency, around the implanted intraocular lens. They may form globular masses of epithelial and cell material known as Elschnig pearls. In addition, if residual lens epithelial cells are ensnared between anterior and posterior lens capsules, subsequent proliferation and differentiation may occur generating lens-like material involving cells and fiber-like material. Linreported a ground-breaking pediatric cataract surgery that aimed to use this postoperative complication as an alternative therapy for children aged under 24mo. This novel surgical method of cataract removal preserves endogenous lens cells, thus enabling functional crystalline lens regeneration. This ground-breaking pediatric cataract surgery is still in early stages of evaluation and requires additional study to address possible complications such as cataract reoccurrence due to underlying genetic defects and the risk of developing amblyopia, impaired vision due to inappropriate visual stimulation during the months required for lens regeneration.

Clinical trials have determined that transplantable lens stem cells provided by human embryonic stem cells (hESCs) and hiPSCs can generate lens-like organoids of various sizes and scalable clarity. Several investigators have tried to define simple and practicable methods of producing lens epithelial cells using hiPSCs. These cells are similar to human fetal lens epithelial cells, and,, are capable of generating light-focusing micro-lenses. Using the appropriate signaling pathways able to induce functional lens regeneration, we could pass frommodels toapplications. The microlenses bear the potential to reduce the personal, social and economic consequences of cataract as they can be used to model individual or combined cataract risk factors as well as assist withdrug development and toxicity assays.

To avoid LSCD in the donor’s eye, specific methods are being developed to achieveexpansion of donor’s stem cells. By using this approach, various substrates are utilized, such as the human amniotic membrane. This substrate, unfortunately, showcases limited availability, high screening costs, and is susceptible to folding and creasing during transplantation. As an alternative fibrin matrix can be used, which exhibit high success rates at ten-year follow up. Another alternative is the use of a polymer that doesn’t necessitate complex procedures as a substrate. Moreover,silicone hydrogel contact lenses can be used as a carrier substrate, which is highly approachable and very costeffective. Recently other substrates are also considered, such as anterior lens capsules, collagen, and synthetic polymers.These alternatives are currently under investigation and validation to determine their sustainability and effectiveness.Pre-clinical models and human clinical trials are investigating the possibility of alternative tissue sources, such as allogeneic conjunctival cells, and autologous nasal and buccal mucosa.

蕭瓊堅(jiān)決地道：“我要與我羽弟生死與共?！庇蝹}(cāng)?？嘈Φ溃骸肮媚棠?，你怎如此固執(zhí)？這樣好不好？我傳你武功，候你有了自保之能我?guī)闳ヒ?jiàn)你羽弟。”

In AMD, RPE pathology constitutes the first clinically detectable sign, followed by photoreceptor loss and permanent impairment of visual function. In the past decades, several trials examined the conduct of the subretinal space by trying to transplant allogeneic RPE cells. Nevertheless, the results were frustrating due to the rejection mechanisms of these donor cells. Currently, new clinical trials are considering the eventuality of using autologous RPE cells by accomplishing macular translocation, and RPE choroid patch graft transplantation.

Macular translocation consists of detaching and rotating the retina to reposition the macula over a healthy area of the RPE. A retrospective study by Takeuchi, on macular translocation for AMD and myopic choroidal neovascularization proved its effectiveness by demonstrating visual acuity preservation for more than 5y post-surgery.

On the other hand, RPE-choroid patch graft transplantation consists of harvesting full-thickness RPE-choroid patches from other regions of the patient’s retina and transplanting them under the impaired macula. Relevant clinical trials results demonstrated visual acuity preservation for more than 3y posttransplantation. Unfortunately, these surgical procedures demonstrated high technical complexity and were also correlated with high risk of retinal detachment, which impeded its clinical implementation and improvement.

Clinical trials concerning macular translocation and RPEchoroid patch graft transplantation have established the efficacy of RPE replacement in AMD. The possibility of providing the macula with healthy RPE and fresh underlying choroid is very promising and should be taken into account in future clinical trials. New techniques are being developed to manufacture RPE replacement patches in the laboratory. Sharmadeveloped a treatment for macular degeneration that used RPE cells derived from a patient’s own healthy cells. They also designed a scaffold to support healthy RPE cells and promote their attachment inside the eye, in order to overcome the risk of retinal detachments carried by surgically harvesting autologous RPE-choroid patch grafts.

Regenerative patch technologies (RPT) is developing a stem cell-based implant technology. The CPCB-RPE1 implant consists of stem cell-derived RPE cells on an ultrathin synthetic parylene membrane creating a 6.25×3.5 mmpatch.The implant is designed to replace the RPE and Bruch’s membrane in AMD disease. The latest results from RPT’s Phase 1/2a clinical trial [National Library of Medicine (NLM),NCT02590692]demonstrate the safety and tolerability of the implant along with the providing signals of sight-improving activity. The data positions RPT to proceed to a Phase 2b clinical trial. On the other hand, the London Project to Cure Blindness has developed an hESC-derived RPE sheet grown on a biostable polyethylene terephthalate (PET) membrane,creating a 6×3 mmpatch. Both types of patches are delivered after a pars plana vitrectomy and a small retinotomy over the macula region.

Retinal organoids are derived from hiPSCs constituting autologous tissues organized in 3D structures, which structurally and functionally mimic the retina. In the last decade they have assisted enormously in studying several retinal diseases, that were difficult to reproduceor in rodent models. The mutation of CEP290, a primary ciliary protein, was studied in retinal organoids derived from CEP290-mutated Leber congenital amaurosis, to understand the inner mechanisms of these ciliopathies. In addition retinal organoids are extremely useful in the study of several retinitis pigmentosa mutations, such as TRNT1 mutation, and Stargardt’s disease. Furthermore, retinal organoids are being used to detect the protective effects of ophthalmic supplements,such as 4-hydroxytamoxifen and diethylstilbestrol, and to evaluate the pharmacological effects of moxifloxacin which is retinotoxic at high doses. They also emerged to be precious photoreceptor source for cell transplantation therapies, such as AMD and Stargardt’s disease.

從筆者20多年參加大中型水利水電工程建設(shè)項(xiàng)目的質(zhì)量監(jiān)督管理實(shí)踐中，感覺(jué)很多項(xiàng)目法人對(duì)招標(biāo)選擇的施工承建單位進(jìn)場(chǎng)后的表現(xiàn)并不滿意。施工承建單位本是項(xiàng)目法人通過(guò)招標(biāo)投標(biāo)方式擇優(yōu)選取的，之所以會(huì)出現(xiàn)這一現(xiàn)象，筆者認(rèn)為還是在于工程招標(biāo)投標(biāo)中沒(méi)有真正做到擇優(yōu)選取。項(xiàng)目法人通常認(rèn)為大型企業(yè)、央企實(shí)力雄厚，技術(shù)力量強(qiáng)，是一般中小型施工企業(yè)無(wú)法比擬的，因此在招標(biāo)投標(biāo)過(guò)程始終將雙眼盯在大型施工企業(yè)或央企上，認(rèn)為只要將工程建設(shè)施工任務(wù)交給它們，施工進(jìn)度、質(zhì)量就能完全得到保證，直到施工承建單位中標(biāo)進(jìn)場(chǎng)后，才發(fā)現(xiàn)事實(shí)并不像原來(lái)所期望的那樣，許多地方并不盡如人意。

Various clinical trials ascertained that transplantation of hESC-derived RPE resulted to reduced photoreceptors degeneration, but the safety and tolerability of this procedure had to be evaluated.Schwartzconducted two phase I/II studies involving patients with geographic atrophy in dry AMD and Stargardt disease. The 2-year resultswere very promising with no deployment of uncontrolled proliferation, adverse reactions, or rejection. An assessment at 4y demonstrated that it is possible to safely implant hESC-RPE in order to attempt to rescue photoreceptors and visual function. Patients experienced sustained improvement in visual acuity and no adverse events were noted. Indeed, there was no tumorigenicity or rejectionrelated inflammation. These results indicated the way to studies involving new strategies for RPE transplantation, such as sheets of cells with or without scaffolding to mimic Bruch’s membrane.

綜上所述,根據(jù)實(shí)驗(yàn)結(jié)果,腫瘤壞死對(duì)患者病情有較為嚴(yán)重的影響,并且隨著患者腫瘤T分期的增加,腫瘤實(shí)質(zhì)性區(qū)的強(qiáng)化程度逐漸增加;且在實(shí)驗(yàn)結(jié)果中,非小細(xì)胞癌的CT強(qiáng)化程度,高于小細(xì)胞癌。

Albeit hESC-RPE graft was safely tolerated, uncertainty was raised concerning the procedures with allogeneic cell grafts.Immunosuppression required in this case, caused inadmissible systemic side-effects forcing Songto retract it. Given the unpropitious immunosuppression effects and the failure of clinical trials using cultured fetal allogeneic RPE, effectiveness on human patients is yet to be confirmed.

Over a billion people worldwide,practically people aged 40y and older, are affected by presbyopia. Until today, the only efficacious treatment of presbyopia consists of non-surgical refractive correction. More than 100 million people suffer from vision impairment or even blindness due to cataracts. The reduction of lens translucency caused by cataract and consecutive loss of visual clarity and sharpness can only be treatedsurgical replacement of the cataractous lens, making cataract surgery the most common ophthalmic procedure worldwide.

DDC使用可變流量系統(tǒng)來(lái)控制空調(diào)水系統(tǒng)。空調(diào)在一定的水溫下工作，提高熱源機(jī)的工作效率，并通過(guò)水泵調(diào)節(jié)供水，節(jié)省能源。水系統(tǒng)冷水機(jī)組與冷凍水泵、冷卻水泵和冷卻塔一對(duì)一運(yùn)行。

To avoid the impediment of immunosuppression, clinical trials are directed to the use of autologous hiPSCs-derived RPE.These cells showcased their ability to restore retinal function when grafted into the sub-retinal space, without being mutagenic. In addition, clinical trials provided evidence that the photoreceptor progenitors have the ability to ingrate into the dysfunctional murine and swine retina and partially restore visual function. Nevertheless, the oncogenic likelihood of this procedure remains to be clarified as well as the necessity to be accompanied by supportive strategies of RPE and retinal neurons, to ensure their long-term graft survival.

MSCs primarily originate from the human bone marrow. Clinical studies showcased that bone mesenchymal stem cells (BMSCs) are capable to differentiate into retinal neurons under appropriate induction.Nevertheless, BMSCs’ quantity and ability are susceptible to multiple factors as donor’s gender, age, and radiation therapy history. The application of BMSCs in clinical practice is also hindered by the intrusiveness and inconvenience of bone marrow aspiration.

1.6 統(tǒng)計(jì)方法 統(tǒng)計(jì)不同年齡段女性的高危HPV感染率(各個(gè)年齡組中高危HPV陽(yáng)性病例數(shù)／這一年齡組總?cè)藬?shù)×100%)。高危HPV各亞型感染率指該亞型單獨(dú)或和其他亞型共同出現(xiàn)的感染率，多重感染重復(fù)計(jì)算。例如某患者檢測(cè)結(jié)果為HPVl6、

Corneal epithelium is replenished by limbal epithelial stem cells (LESCs)located in the Vogt palisades. Any disorganization of this microenvironment can cause limbal stem cell deficiency(LSCD). This devastating condition is characterized by unstable corneal epithelium and persistent epithelial defects,leading to chronic pain and progressive visual loss. Very often this condition does not respond to penetrating keratoplasty,and in these cases, limbal stem cell transplantation is used to achieve corneal epithelial regeneration.

公司預(yù)計(jì)下年銷售量為8 500件，兩種經(jīng)營(yíng)模式的收入、成本結(jié)構(gòu)如表11所示。公司計(jì)劃將總資產(chǎn)擴(kuò)大為500萬(wàn)元，假設(shè)公司資產(chǎn)負(fù)債率為50%，負(fù)債利率為8%，固定成本中不包括負(fù)債利息。兩種方案的收益和經(jīng)營(yíng)風(fēng)險(xiǎn)分析，見(jiàn)表11[1]。

Lushowed that by using epidermal growth factor(EGF), taurine, and retinoic acid as inducing reagents, OMMSCs can be induced to differentiate into photoreceptor cellsas well as express a photoreceptor-specific cell marker.The application of OM-MSCs in developing neuron progenitor cells could be a new approach for novel therapies of retinal degenerative diseases.

Furthermore, Daistudied the use of olfactory mucosal cells (OMCs) in rats showcasing elevated intraocular pressure(IOP). Their trials demonstrated that transplantation of OMCs into the optic disc region, reduced to 23% the loss of retinal ganglion cells (RGC) axons at the optic nerve head(ONH). In addition, the pattern electroretinograms (PERG)determined that the decrement of the voltage amplitudes in association with the raised IOP was significantly alleviated.Transplantation of OMCs not only is capable to diminish the loss of RGC axons induced by raised IOP in rats, but also to protect the functional activities of the axons. These findings provide a basis for future human clinical trials. Autografting OMCs from autologous nasal epithelial biopsies could treat or delay glaucoma diseases.

CONCLUSION

Stem cell therapy has become a practical reality and holds a great potential in improving the quality of life of visually impaired people. Current and continuing research increases the possibilities of stem cells applications and carries a promising future in producing novel biological therapies to treat vision loss despite the many challenges faced. Nevertheless, ethical dilemmas arise, particularly in the absence of high-quality evidence, increased marketing claims and lack of transparency regarding patient recruitment. The need to do no harm highlights the importance of rigorous clinical trial processes,appropriate regulation and post-market surveillance to avoid ineffective and unsafe products reaching the community.

International societies are committed to ensuring ethical integrity and adequate scientific standards in clinical trials and therapies concerning stem cell procedures. The International Society for Cellular Therapy (ISCT) and the International Society for Stem Cell Research (ISSCR) are global associations linking pre-clinical research to clinical trials. The Association for Research in Vision and Ophthalmology (ARVO) supports the development of policies that promote stem cell research and therapies. The American Academy of Ophthalmology (AAO) studies and reports the dangers of inadequate quality control of unproven stem cell interventions. With the extremely rapid evolution of ocular regenerative medicine, proper regulation is demanded to avoid ineffective and unsafe products reaching the patients.

The use of treatment outcome registries is taken into account,to provide evidence on the safety and effectiveness of stem cell-based therapies. This methodology could act as a link between clinical trials and ongoing treatments already available on the market. Proper surveillance of these postmarket registries by jurisdictional committees and regulators,could safeguard patients from being misguided.

Progress in regenerative medicine would continue to enhance welfare, while stem cell therapies would manage to improve the patient’s life. In addition, patients should be protected from uncontrolled media violations and deceptive marketing assertions. Ocular regenerative medicine is currently a fastdeveloping area that requires independent, unbiased, and objective scientific evidence in order to ensure patient treatment decisions and outcomes are not mediated by commercial or financial interests.

將結(jié)果帶入SPSS22.0版中,以P<0.05表達(dá)組間差別較大。將兩組患者的不良反應(yīng)發(fā)生率作為計(jì)數(shù)資料,采用百分比表示,卡方檢驗(yàn);將兩組患者的SDS、SAS評(píng)分、住院時(shí)間等作為計(jì)量資料,采用(均數(shù)±標(biāo)準(zhǔn)差)表示,t檢驗(yàn)。

Sotiropulos K: Project development,data management and analysis, manuscript writing and editing;Kourkoutas D: Data analysis, manuscript writing and editing;Almaliotis D: Manuscript editing; Ploumidou K: Manuscript editing; Karampatakis V: manuscript editing.

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