Evaluation of the safety and efficacy of using human menstrual blood-derived mesenchymal stromal cells in treating severe and critically ill COVID-19 patients: An exploratory clinical trial

評估使用間充質(zhì)基質(zhì)干細胞治療重癥和危重癥新冠患者的安全性和有效性：一項探索性臨床試驗

Abstract

The coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was identified in December 2019 and has subsequently spread worldwide. Currently, there is no effective method to cure COVID-19.

Mesenchymal stromal cells (MSCs) may be able to effectively treat COVID-19, especially for severe and critical patients. Menstrual blood-derived MSCs have recently received much attention due to their superior proliferation ability and their lack of ethical problems. Forty-four patients were enrolled from January to April 2020 in a multicenter, open/label,nonrandomized,parallelcontrolled exploratory trial. Twenty-six patients received allogeneic, menstrual blood-derivedMSC therapy,and concomitant medications (experimental group),and 18 patients received only concomitant medications (control group). The experimental group was treated with three infusions totaling 9 × 107 MSCs,one infusion every other day. Primary and secondary endpoints related to safety and efficacy were assessed at various time points during the 1-month period following MSC infusion. Safety was measured using the frequency of treatmentrelated adverse events (AEs). Patients in the MSC group showed significantly lower mortality (7.69% died in the experimental group vs 33.33% in the control group; P = .048). There was a significant improvement in dyspnea while undergoing MSC infusion on days 1, 3, and 5. Additionally, SpO2 was significantly improved following MSC infusion, and chest imaging results were improved in the experimental group in the first month after MSC infusion. The incidence of most AEs did not differ between the groups. MSC-based therapy may serve as a promising alternative method for treating severe and critical COVID-19.

由嚴重急性呼吸綜合癥新冠2 (SARS-CoV-2)引起的扮宠，2019的新冠病毒鬼佣，在2019年12月被確診并且在世界各地廣泛傳播。近期全肮，對于治療COVID-19沒有有效的方法。MSCs治療COVID-19可能有效匹表，特別是重癥和危重癥患者。MSCs由于超強繁殖能力和缺乏倫理問題近期受到大量關(guān)注驼抹。在一項多中心桑孩、開放、不隨機框冀、無平行對照的探索性試驗中從2020年1月到4月入組44例患者流椒。26例患者接受異體經(jīng)血MCS治療和伴隨用藥（試驗組），18例患者只接受伴隨用藥（對照組）明也。試驗組接受三次輸液合計9 × 107 MSCs宣虾，每隔一天輸一次惯裕。根據(jù)MSC注射后一個月內(nèi)的重要時間節(jié)點評估與安全性和有效性有關(guān)的主要及次要指標(biāo)。安全性評估使用與藥物相關(guān)的不良事件绣硝。在MSC組患者顯示顯著的低死亡率(試驗組7.69%蜻势。對照組33.33%，P=0.048)鹉胖。在第1握玛、3、5天接受MSC注射呼吸困難有顯著改善甫菠。另外挠铲，SpO2在MSC注射后有顯著改善，并且在試驗組注射MSC后的第一個月胸部成像結(jié)果得到改善寂诱。大部分AEs發(fā)生率在組間沒有差異拂苹，基于MSC療法可能作為一項治療重癥和危重癥新冠患者有前景的替代療法。

Mortality=death rate

Incidence：

Therapy=treat

Serve as =considered

Promising=hopeful

Alternative=alter+native=改變+原來的痰洒，當(dāng)?shù)厝?替代

翻譯難點瓢棒，

may serve as的翻譯，直譯為作為一項方法為患者服務(wù)丘喻，三個信息點脯宿，它是基于MSC的一項療法，它可能作為一項最有前景的替代療法仓犬，它是為重癥和危重癥新冠患者服務(wù)的嗅绰。

（2）every other day，新知識搀继，每隔一天。

2021年6月8日星期二（第1天翻譯250個詞）

1.introduction

The coronavirus disease 2019 (COVID-19), caused by severe~acute respiratory syndrome coronavirus 2 (SARSCoV2), was initially identified in December 2019 as causing a cluster of respiratory infections.1,2 COVID19 quickly attracted global concern and panic since it is highly contagious.3–5 Due to a lack of adequate awareness in the first few weeks of the outbreak, the number of infected patients increased swiftly, rapidly spreading to more and more countries.6,7 As of January 7, 2021, there have been over 85 929 000 confirmed cases of COVID-19 worldwide,leading to 1 876 100 deaths. Currently, the number of infected patients is still increasing worldwide.

這個新冠疾病2019（COVID-19）翠语，由嚴重急性呼吸綜合冠狀病毒2(SARS-CoV-2)引起的叽躯，在2019年12月被明確定義為引起一系列呼吸道感染。COVID-19由于高度傳染性快速引起全球廣泛關(guān)注和恐慌肌括。由于在爆發(fā)開始的幾周缺乏足夠的意識点骑。感染患者人數(shù)極速增加，傳播到越來越多的國家谍夭。在2021年7月黑滴，在全球已經(jīng)超過了85929000 COVID-19確診病例，導(dǎo)致1876100人死亡紧索。進來袁辈，感染患者的人數(shù)仍然在全球持續(xù)增加。

COVID19 has an incubation period which can range from 1 to 14 days but usually ranges from 3 to 7 days.8 The main symptoms are fever, headache, dry cough, and chest tightness.911 Many patients also experience a sore throat, diarrhea, nasal congestion, and rhinorrhea.12,13

Severe patients often develop expiratory hyperextension and dyspnea 1 week after the onset of the disease. In the most severe cases, patients can quickly develop acute respiratory distress syndrome (ARDS), severe acute lung injury, septic shock, metabolic acidosis, and coagulopathy,as reported in a biopsy and autopsy study.14 COVID-19 can easily cause expiratory dyspnea and ARDS. Of the COVID-19 patients, 13.8% of the cases were severe, 6.1% cases were critical, and about 2.3% cases had fatal outcomes.Since no effective or authorized vaccines are available for preventing COVID19 infections, a breakthrough in the therapeutic strategy is vital for the treatment of COVID19 and especially for severe or critically ill patients who may develop ARDS and/or expiratory dyspnea.17–21 Currently,a few drugs (such as remdesivir and dexamethasone)have shown positive preliminary results in randomized,controlled, open.label, clinical trials.22,23 Even more excitingly, COVID19 vaccines with acceptable safety, tolerability,and immunogenicity have been reported by Zhuet al and Folegatti et al as being effective in initial human clinical trials. Apart from these, many other groups are also developing available vaccines such as the BNT162mRNA vaccine26 sponsored by Pfizer Inc. and BioNTech SE; the chimpanzee adenovirus vectored vaccine ChAdOx1 nCoV-19 (AZD1222) sponsored by AstraZeneca;the mRNA-1273 vaccine29 codeveloped by the Cambridge,Massachusetts-based biotechnology company Moderna,

Inc., and the National Institute of Allergy and Infectious Diseases (NIAID); the adenovirus serotype 26 (Ad26) vaccine30 developed by Janssen Vaccines & Prevention B.V.; the BBIBP-CorV vaccine31 developed by the Beijing Institute of Biological Products; the CoronaVac vaccine developed by Sinovac Life Sciences; and the NVX-CoV2373

Vaccine sponsored by Novavax, Inc. However, no specific drugs have been reported to be absolutely effective in treating COVID-19. Moreover, SARS-CoV-2-induced secondary infections have been reported to induce multiple organ dysfunction syndrome in severe or critically ill patients, and this issue remains a serious problem worldwide.

COVID-19的孵化周期是1-14天珠漂，但一般范圍是3-7天晚缩。最重要的癥狀是發(fā)燒尾膊、頭痛、干咳荞彼、胸悶冈敛。一些患者也會經(jīng)歷咽喉疼痛、腹瀉鸣皂、鼻塞抓谴、流鼻涕。重癥患者通常在發(fā)病一周后出現(xiàn)呼吸性伸展和呼吸困難（onset進攻）寞缝。根據(jù)活檢和尸檢研究報告癌压，在最嚴重的情況下，患者很快發(fā)展成急性呼吸窘迫綜合癥（ARDS）第租，急性重癥肺部損傷措拇，感染性休克、代謝酸中毒慎宾、凝血病丐吓。COVID-19很容易引起呼氣型呼吸困難和ARDS。COVID-19患者中趟据，13.8%病例是重癥券犁，6.1%病例是危重癥，并且大約2.3%病例有致命結(jié)果汹碱。由于沒有有效和權(quán)威的疫苗能夠阻止COVID-19感染粘衬，COVID-19治療尤其改善ARDS和呼氣型呼吸困難的重癥和危重癥患者的治療策略的突破十分關(guān)鍵。近期咳促，在一項隨機稚新、對照、開發(fā)標(biāo)簽的臨床試驗一些藥物顯示了初步的陽性結(jié)果跪腹。甚至更加激動人心的是褂删，具有可接受的安全性、耐受性和免疫原性的COVID-19疫苗被zhu等人和Folegatti等人報道了冲茸。除了這些屯阀，其他研究小組也發(fā)展了可接受疫苗，

例如：（1）Pfizer（輝瑞）和BioNTech SE贊助的BNT162 mRNA苗轴术；

（2）AstraZeneca（阿斯利康）贊助的黑猩猩腺病毒載體疫苗ChAdOx1 nCoV-19 (AZD1222)难衰；

（3）由位于馬薩諸塞州劍橋市的生物技術(shù)公司 Moderna, Inc. 和美國國家過敏和傳染病研究所 (NIAID) 共同開發(fā)的 mRNA-1273 疫苗；

（4）由Janssen Vaccines & Prevention B.V.研發(fā)的腺病毒血清型 26 (Ad26)疫苗

（5）北京生物制品研究所研制的BBIBP-CorV疫苗

（6）Sinovac生命科學(xué)研發(fā)的CoronaVac疫苗

（7）Novavax, Inc贊助NVX-CoV2373疫苗

然而沒有特別的藥物被報道對治療COVID-19明顯有效逗栽。而且盖袭， SARS-CoV-2 引起的繼發(fā)感染被報道可促使重癥和危重癥患者多組織功能障礙綜合癥，這個問題仍然是全球范圍內(nèi)的一個嚴重問題（induce重點謂語翻譯錯誤，誘導(dǎo)苍凛、促使趣席；remain翻譯錯誤，本意是仍然）醇蝴。

Mesenchymal stem cells have been used for almost three decades, and have made great progress.34 According to the recommendations of the International Society for Cell & Gene Therapy (ISCT) in 2019, mesenchymal stem cells should be named mesenchymal stromal cells (MSCs).35 MSC-based cellular therapy has been the subject of an increasing number of studies due to the cells’ self-renewing capacity, multipotent potential, low immunogenicity, anti-inflammatory activity, and ability to home to damaged tissue.More importantly, MSCs have unique immunomodulatory functions of both innate and adaptive immune responses, making them an attractive cell therapy tool.38,39 MSCs regulate adaptive immune responsesmainly through targeting T lymphocytes, B lymphocytes,antigen-presenting cells (APCs), dendritic cells (DCs), natural killer (NK) cells, and regulatory T cells (Tregs).38 MSCs also regulate innate immune responses mainly through targeting DCs, NK cells, innate TH17 cells, neutrophils, monocytes, macrophages, and mast cells.39 Further,MSC-based therapies have shown promising

results in several clinical studies across a variety of diseases.40,41 With the development of stem cell research,researchers have found that after injecting MSCs, the human body activates the host’s innate immune cascade system, such as complement and blood coagulation,which is defined as the instant blood-mediated inflammatory esponse (IBMIR).42 IBMIR is of key importance onsidering the highly procoagulant state of many critically nd severely ill patients in need of MSC therapy.43 SCs can be obtained from various sources, including one marrow (BM), adipose (AD), umbilical cord (UC),placenta, menstrual blood, muscle, dental pulp, Wharton’s elly (WJ), fetal liver, amniotic membrane, amniotic luid, urine etc.44–48 Furthermore, MSC-based treatment has demonstrated promising results in studies on inflammatory lung disease, showing an ability to inhibit alveolar collapse, cell apoptosis, and collagen accumulation in lung tissues.49 The angiotensin-converting enzyme 2 (ACE2) is identified as a receptor of SARS-CoV-2 entering into target cells.50,51 In addition, researchers have demonstrated

thatMSCs do not express ACE2, andMSCs are resistant to SARS-CoV-2 infection as well as when exposed to SARSCoV-2-infected cells.52,53 Additionally, following infusion of allogeneic MSCs into nine patients with ARDS, Wilsonet al54 observed no prespecified side effects including cardiac arrhythmia, hypoxemia, and ventricular tachycardia.Further, our team reported that MSC transplantation significantly lowers the mortality of epidemic Influenza A (H7N9)-induced ARDS patients.55 BM-MSCs have been shown to improve repair after ventilator-induced lung injury, to facilitate the resolution of inflammation, and to restore lung function and structure in ARDS patients.56 With regard to the COVID-19 epidemic, MSCs from different sources (especially UC-MSCs) have been used in clinical trials.53,57 A good proliferation rate plays an important role in clinical application, because stem cell-based treatment is dose-dependent, and usually human clinical research requires millions of stem cells. The doubling time for menstrual blood-derived MSCs is about 20 h, and the doubling time for BM-MSCs is about 40-45 h. Thus, MSCs from menstrual blood can obtain a better yield within a shorter time at early passages.58,59 More importantly, menstrual blood-derived MSCs offer an alternative that is both painless and free of the ethical issues that may arise from BM-MSCs donations.60 Thus, menstrual bloodderivedMSC-

based therapy may be a promising treatment for COVID-19, particularly to combat the inflammatory cytokine storms observed in severe and critical patients.

間充質(zhì)干細胞被使用已經(jīng)有30年了宣肚，并且有很大的進步。在2019年根據(jù)科學(xué)家的解釋推薦細胞聯(lián)合基因治療悠栓，間充質(zhì)干細胞被命名為間充質(zhì)基質(zhì)細胞(MSCs)霉涨。基于MSC的細胞療法惭适，由于自我新生能力笙瑟，多種潛在能力，低免疫原性癞志，抗炎活性往枷，修復(fù)受損組織能力等成為了研究數(shù)量逐增的課題。更重要的是凄杯，MSCs在先天性和適應(yīng)性免疫反應(yīng)错洁，有獨特的免疫調(diào)節(jié)，讓他們成為有吸引力的細胞治療工具戒突。MSCs調(diào)節(jié)適應(yīng)性免疫反應(yīng)主要通過靶向T淋巴細胞屯碴、B淋巴細胞、抗原呈遞細胞膊存、樹突狀細胞导而、自然殺傷細胞和調(diào)節(jié)性T細胞。MSCs也可以調(diào)節(jié)先天性免疫反應(yīng)主要通過靶向DCs隔崎、NK細胞今艺、先天TH17細胞、中性粒細胞爵卒、單核細胞洼滚、巨噬細胞、肥大細胞技潘。更進一步來說，基于MSC治療顯示良好的結(jié)果在通過各種疾病的多項臨床研究中千康。隨著干細胞的研究發(fā)展享幽，研究者發(fā)現(xiàn)接種MSCs之后。人體激活宿主拾弃。IBMIR要至關(guān)重要的考慮到需要MSC治療的許多重癥和危重癥患者的高度凝血狀態(tài)值桩。MSCs可以從不同的來源獲取，包括包括骨髓（BM）豪椿、脂肪（AD）奔坟、臍帶（UC）携栋、胎盤、經(jīng)血咳秉、肌肉婉支、牙髓、沃頓氏膠（WJ）澜建、胎肝向挖、羊膜、羊水炕舵、尿液等何之。此外，基于MSC治療在炎癥性肺部疾病研究中表明有非逞式睿可觀的結(jié)果溶推，顯示了抑制肺泡塌陷、細胞凋亡和肺組織中膠原蛋白積累的能力奸攻。ACE2被鑒定為SARS-CoV-2進入靶細胞的受體蒜危。另外，研究者表明MSCs沒有表達ACE2舞箍，MSC 對 SARS-CoV-2 感染以及暴露于 SARS-CoV-2 感染的細胞具有抵抗力舰褪。

另外，同種異體MSCs注射進入9個ARDS患者, Wilson等人沒有觀察到提前制定的副作用疏橄，包括心律失常占拍、低氧血癥和室性心動過速。

另外捎迫，我們的團隊報告MSCs細胞移植顯著降低流行性甲型流感（H7N9）誘發(fā)的ARDS患者的死亡率晃酒。BM-MSCs也顯示可以提高呼吸機所致肺損傷后修復(fù)，促進炎癥的“再解決”消退窄绒，并恢復(fù) ARDS 患者的肺功能和結(jié)構(gòu)贝次。隨著COVID-19的流行，不同來源的MSCs已經(jīng)使用在臨床試驗中彰导。在臨床試驗中一個良好繁殖率起著非常重要的作用蛔翅，因為基于干細胞治療具有劑量依賴性，并且通常人類臨床研究者要求數(shù)百萬的干細胞位谋。經(jīng)血MSCs的倍增時間約是20h山析，BM-MSCs的倍增時間約是40-45h。因此掏父，在早期的短時間內(nèi)來自經(jīng)血的MSCs可以獲得更好的量笋轨。更重要的是，來自經(jīng)血的MSCs提供一種可替代方案，既無痛和也沒有來自BM-MSCs捐獻者產(chǎn)生的倫理問題（alternative名詞爵政，替代方案）仅讽。因此，基于經(jīng)血來源的MSCs可能是最有前景的治療方法钾挟，對于COVID-19洁灵，特別是對抗在重癥和危重癥患者觀察到的炎癥細胞因子風(fēng)暴。

This study is an exploratory trial to assess the ability of menstrual blood-derived MSCs to treat severe and critically ill COVID-19 patients. To this end, we assessed the safety, therapeutic efficacy, and tolerability of transplanted MSCs with a 1 month follow-up after SARS-CoV-2 infection. In particular, we assessed any improvements in pulmonary

function. Our results not only shed light on the ability ofMSCs to treat COVID-19 patients, but also suggest that MSCs are a promising tool to treat acute or chronic pneumonia in future clinical applications.

這個研究是評估經(jīng)血MSCs治療COVID-19重癥和危重癥患者能力的一項探索性是試驗等龙。在最后处渣，我們評估了SARS-CoV-2感染后MSCs移植一個月隨訪的安全性，治療有效性蛛砰，以及耐受性罐栈。特別是我們評估了肺功能的一些改善。我們的結(jié)果不僅揭示患者MSCS治療COVID-19患者的能力泥畅，也表明在未來臨床應(yīng)用中荠诬，MSCs對于治療急性或慢性肺炎是非常有前景的工具。

(2021年7月6日星期二翻譯完)

2 MATERIALS AND METHODS

2.1 Study design and participants

This was a multicenter, open-label, nonrandomized, and parallel controlled phase I clinical trial performed at two major academic centers in China: the Renmin Hospital of Wuhan University, Wuhan and the First Affiliated Hospital,College of Medicine Zhejiang University, Hangzhou.

The Shulan (Hangzhou) Hospital, affiliated to Zhejiang Shuren University, Shulan International Medical College,Hangzhou also participated in related studies. Eligible patients were 18-75 years old and confirmed to be positive for the SARS-CoV-2 RNA virus by polymerase chain reaction (PCR) analysis performed within biological safety protection level 3 laboratories at Wuhan University and Zhejiang University. Before the initiation of this study, the research protocol, case report form (eCRF), and informed consent form were each obtained and approved by the Ethics Committees of Renmin Hospital of Wuhan University (WDRY2020-K011) and the First Affiliated Hospital,College of Medicine, Zhejiang University in accordance with the Declaration of Helsinki and the criteria of Good Clinical Practice.61 This clinical trial was also registered in the Chinese Clinical Trial Registry (ChiCTR2000029606).

The investigators fully educated each patient’s legal representative

regarding the informed consent form, the detailed therapeutic procedure, aswell as the possible risks and benefits. The patients had the right to withdraw from this clinical study at any time during the clinical trial. Considering the urgency of the COVID-19 epidemic and the operability of the enrollment process, a randomized table was not used for randomization in the research process at Renmin Hospital of Wuhan University, and cases (including severe and critically ill patients) were matched based on similar severity levels and similar timing of enrollment/in the study. This clinical trial was an open study and did not involve blinding or emergency unblinding.

2 材料與方法

2.1 研究設(shè)計和參與者

這是一個在中國兩個重要的學(xué)術(shù)中心開展的多中心位仁、開放柑贞、不隨機、平行對照一期臨床試驗聂抢。武漢大學(xué)人民醫(yī)院和浙江大學(xué)醫(yī)學(xué)院附屬第一醫(yī)院也參與了相關(guān)研究钧嘶。符合資格的病人是18-75歲并且在武漢大學(xué)和浙江大學(xué)在3級生物安全性保護中被RCT分析確認SARS-CoV-2 RNA陽性的。研究開始之前琳疏，這研究的安慰劑有决、eCRF和標(biāo)準(zhǔn)文檔格式被（武漢大學(xué)人民醫(yī)院和浙江大學(xué)醫(yī)學(xué)院附屬第一醫(yī)院的）倫理委員會按照赫爾辛基宣言和GCP標(biāo)準(zhǔn)獲取和同意。該臨床試驗也在中國臨床試驗注冊中心進行注冊空盼。研究者要充分通知每一個患者法律代表书幕，包括標(biāo)準(zhǔn)文檔格式、詳細的治療過程和可能的風(fēng)險受益揽趾。在臨床試驗進行的任何時間患者都可以從臨床研究中退出台汇。考慮到COVID-19疫情的緊急性和入組過程的可操作性篱瞎，隨機表不適用在武漢大學(xué)人民醫(yī)院的研究過程苟呐，案例匹配基于研究中相似的嚴重程度和相近的入組時間。這個臨床試驗是開放性研究俐筋，不涉及設(shè)盲或緊急破盲掠抬。

2021年6月16日星期三（第2天翻譯282個詞）

All patients met the diagnostic criteria for COVID-19 according to the National Health Commission of China (Trial Version 5). Following established clinical guidelines for the diagnosis and treatment of COVID-19, patients can be classified as mild, common, severe, or critical.53,62,63 Only severe and critically ill COVID-19 patients were included in the present study. Severe patients were defined as those with respiratory distress, respiratory rate ≥30 breaths/min; resting oxygen saturation ≤93%; or arterial blood partial pressure of oxygen (PaO2)/fraction of inspiration O2 (FiO2) ≤300 mmHg (1 mmHg = 0.133 kPa). Critical COVID-19 patients were defined as those with respiratory failure who required mechanical ventilation, those who had experienced shock, or those for whom a combination of organ failures necessitatedmonitoring and treatment in the intensive care unit (ICU). Exclusion criteria for this trial were as follows:

(1) severe liver disease;

(2)long-term hemodialysis and severe renal impairment or continuous renal replacement therapy;

(3) comorbidities that might affect the ability of researchers to determine

drug efficacy (mainly malignant tumors, active tuberculosis,interstitial pneumonia, and pulmonary heart disease);

(4) treatment with glucocorticoid medications or other immunosuppressive drugs for longer than 2 weeks;

(5) history of major surgery within 30 days of screening or presence of an unhealed surgical wound;

(6) allergy to any active/inactive ingredients in the study drug;

(7) pregnant or breastfeeding;

(8) previous history of prothrombotic events (venous thromboembolism/stroke);

(9) other circumstances judged by an investigator to preclude participation.

These “other circumstances” leading to exclusion from the study included serious AEs for which the investigator judged that the risk of continuing to participate in the trial was too great, as well as the use of other treatments,without authorization and against medical advice, that could have affected the evaluation. These exclusion criteria followed the National Health Commission of China (Trial Version 5). If a patient met all the inclusion or exclusion criteria, he or she was then enrolled in the experimental group (MSC infusion + concomitant medications) or the control group (concomitant medications).

根據(jù)中國國家健康委員會（試用版5）所有患者均要符合新冠診斷標(biāo)準(zhǔn)。根據(jù)新冠診斷和治療的建立的臨床指導(dǎo)校哎，患者被分為輕度、普通、重癥闷哆、危重腰奋。只有嚴重和危重新冠患者被包含在這個研究中。重癥患者被定義根據(jù)這些呼吸困難抱怔，呼吸頻率大于30次/分鐘劣坊，休息時氧氣飽和度小于等于93%，或者什么的氧氣血液部分壓力動脈血氧分壓(PaO2)/吸入氧分數(shù)(FiO2) 小于等于300 mmHg (1 mmHg = 0.133 kPa)屈留。危重新冠患者被定義為需要借助儀器通氣呼吸失敗衰竭者局冰，經(jīng)歷過休克，在ICU中需要監(jiān)測和治療器官衰竭組合患者灌危。

試驗排除標(biāo)準(zhǔn)如下：

嚴重肝臟疾病

長期什么血液透析和嚴重的什么影響腎臟損害或者不斷的什么腎臟替代治療

什么可能影響研究者決定藥效的能力（重要的是智力問題惡性腫瘤康二，活躍的什么活動性肺結(jié)核，孕婦間質(zhì)性肺炎勇蝙，和肺心臟疾材稹）

超過兩周使用糖皮質(zhì)激素藥物或者免疫抑制藥物

篩選30天內(nèi)有大手術(shù)史，或者存在未愈合的手術(shù)傷口

對研究藥物中任何活性/非活性成分過敏

懷孕或母乳喂養(yǎng)

血栓事件史（靜脈血栓栓塞/中風(fēng)）

通過研究者判斷排除參與的其他的情況

這些其他情況導(dǎo)致研究中排除的包括調(diào)查者判斷為繼續(xù)參與試驗風(fēng)險非常大的嚴重不良事件味混，未經(jīng)授權(quán)使用其他治療和違背藥物說明产雹，影響藥物評估。這些排除標(biāo)準(zhǔn)嚴格遵循中國國家健康委員會（試行版5）如果一個患者滿足所有的排除標(biāo)準(zhǔn)翁锡，他或她會被登記進試驗組蔓挖。（MSC注射+伴隨用藥或?qū)φ战M（伴隨用藥））

2021年6月17日星期四（第3天翻譯331個詞）

2.2 Stratification of disease treatment and concomitant medications

Patients were enrolled and admitted between January and April 2020 at either Renmin Hospital of Wuhan University or the First Affiliated Hospital, College of Medicine,Zhejiang University. A clinical plan was designed for each patient based on their clinical needs. Since the patients

in this clinical study were either severe or critical, concomitant medications were allowed, and the details of all additional treatments were recorded. Drugs that were required to treat other diseases were allowed if an investigator judged that the safety and efficacy of the study medications would not be affected. However, the type and dose of the medications were kept as consistent as possible while prioritizing patient safety. For any concomitant medications or other treatments, detailed medication information was recorded on the original medical record and the eCRF.Of the 44 patients enrolled in this study, 36 were from Wuhan University People’s Hospital, including 20 severe patients and 16 critically ill patients. The remaining eight patients were enrolled from the First Affiliated Hospital,College of Medicine, Zhejiang University including six severe patients and two critically ill patients. Due to the high risk of mortality, patients were given the choice as to which group they were placed in, and 26 patients chose to receive the experimental treatment along with the comprehensive treatment, while 18 patients chose to be included in the control group and receive only the comprehensive treatment. The standard of carewas consistent between the two hospitals.

2.2 疾病治療和伴隨用藥的分層

從2020年1月到4月，在武漢大學(xué)人民醫(yī)院和浙江大學(xué)醫(yī)學(xué)院附屬醫(yī)院患者被安排入組和確認馆衔∥僚校基于他們的臨床需求為每一個患者設(shè)計臨床計劃。無論是重癥還是危重癥的臨床研究患者哈踱，合并用藥都被允許荒适，所有的其他治療詳細信息都要被記錄。如果研究者判斷對研究藥物的安全性和有效性沒有影響开镣，允許使用治療其他疾病的藥物刀诬。但是，在優(yōu)先考慮患者安全性下盡可能保持藥物的類型和劑量一致邪财。對于任何聯(lián)合用藥和其他治療陕壹，詳細的藥物信息被記錄在原始的藥物記錄和eCRF中。

研究中入組的44例患者树埠，36例來自武漢大學(xué)人民醫(yī)院糠馆，包括20例重癥患者和16例危重癥患者。其余的8例患者來自浙江大學(xué)醫(yī)學(xué)院附屬醫(yī)院怎憋，包括6例重癥患者和2例危重癥患者又碌。由于高風(fēng)險死亡率九昧，病人被給與機會選擇他們被參與的組，26個病人選擇在綜合治療接受試驗治療毕匀，18個病人選擇在對照組僅僅接受綜合治療铸鹰。在兩家醫(yī)院保持護理標(biāo)準(zhǔn)一致。

2.3 MSC preparation, cell transplantation, and subsequent observation

Allogeneic, menstrual blood-derived MSCs (no.SC0100919001, no. SC0100919004, and no. SC0100919005 provided by Innovative Precision Medicine (IPM) Group,Hangzhou, China) were obtained from three healthy emale donors (age range, 20-45 years), and the volunteers were educated and provided signed, informed consent before donation, as described in previous studies.55,64 The donation protocol was authorized by the Ethics Committee of Zhejiang University. The mononuclear cells within the menstrual blood were collected and purified, and cell viability was measured prior to seeding for cell culture according to the staining with trypan blue solution (Thermo Fisher Scientific, No. 15250061). MSCs were passaged at 70-80% confluence. Surface-labeled molecules (including CD29, CD34, CD45, CD73, CD90,CD105, CD117, and HLA-DR) were measured using flow cytometry (CytoFLEX LX Flow Cytometer, Beckman) and detailed procedures were described in a previous study.65 Supporting information Table S1 includes detailed information on the antibodies used for surface marker analysis. Detection of the differentiation potential ofMSCs into the osteogenic differentiation medium, chondrogenic differentiation medium, and adipogenic differentiation medium A and B (Cyagen Biosciences) and their detailed information has been reported in a previous study.66 PCR analysis was used to check the ACE2 expression level of the MSCs. The detailed procedures for PCR analysis have been described in a previous study.67 Briefly, cell samples were homogenized in 1 mL of RNAiso Plus (9108, Takara,Japan) to isolate total RNA according to the manufacturer’s instructions. RNA was then reverse-transcribed into cDNA using a FastQuant RT Kit with gDNase (KR106,Tiangen Biotech, China). Then a total of 10 μL sample (1μL cDNA, 5 μL PCR Mastermix [KT201, Tiangen Biotech],1 μL forward primer, 1 μL reversed primer, and 2 μL ddH2O) was used for PCR with 30 cycles. Supporting information Table S2 includes the primer sequences for PCR analysis. The resulting cryopreserved MSCs were shipped frozen to the hospitals in a validated liquid nitrogen (≤?135C) dry shipper. Before their use, MSCs were resuspended in Plasma-Lyte 148 at room temperature by a local laboratory with a specialized cellular therapy center, and the control group was also administered the same volume of Plasma-Lyte 148. The viability of MSCs for the three donors should be >90%, which was a criterion for use in the clinical study guided by the Innovative Precision Medicine (IPM) Group. MSCs were used for treatment at the fifth passage, as described in our previous report.66 Twenty-six patients received MSC transplantation, and18 patients received all treatments exceptMSC transplantation.Complete case enrollment details and disease severity frequencies of the 44 COVID-19 patients included in the present study can be viewed in Table 1. Each COVID-19 patient in the experimental group used the MSC sample from one donor for all three treatment injections. Doctors observed hemodynamic and respiratory parameters at the bedside forat least 1 h toensure that eachpatientwas stable before MSC transplantation. Then, the MSC infusion was initiated using a standard blood filter tube. A researcher remained at each patient’s bedside to continuously monitor the patient for any adverse reactions during the 24 h following treatment. Based on data fromstudies on the use of MSCs to treat H7N9-induced ARDS,55 MSCs were administered as three infusions totaling 9 × 107 MSCs every other day (day 1, day 3, and day 5). Each infusion contained 3 ×107 cells resuspended in 500 mL saline solution and was performed at a speed of 30-40 drops/min for about 15 min,followed by a speed of 100-120 drops/min for 2 h to retain

MSC vitality.

2.3 MSC的制備皂岔，細胞移植蹋笼，和連續(xù)觀察

正如研究前的所述，同種異體躁垛、經(jīng)血來源MSCs的獲取來自三名健康女性捐獻者（年齡范圍20-45歲）剖毯，并且這些志愿者被教育，在捐獻前提供簽署知情同意教馆。這些捐贈協(xié)議被浙江大學(xué)倫理委員會批準(zhǔn)逊谋。經(jīng)血中的單個核細胞被收集和凈化，并且根據(jù)保持藍色情況細胞繁殖被測量加速活玲，在接種細胞培養(yǎng)前根據(jù)盼藍溶液染色測量細胞活性涣狗。（to表示方向和目的，細胞培養(yǎng)接種是什么意思舒憾？暫時沒有得到答案）镀钓。表面標(biāo)記分子被測量使用流式細胞術(shù)，在研究前詳細過程被描述镀迂。支持表S1信息包含在使用表面標(biāo)記分析的抗體上詳細信息丁溅。不同的MSCs潛在保護進入不同的藥物，檢測MSCs向成骨分化培養(yǎng)基探遵、成軟骨分化培養(yǎng)基和成脂分化培養(yǎng)基A和B的分化潛能骡苞，并且他們詳細的信息在研究前被報道（differentiation分化趣斤，medium錯誤翻譯medicine，本意是介質(zhì)，傳媒）档悠。PCR分析被用來檢查MSCs的ACE2表達水平贴铜。PCR分析詳細的過程在研究前被描述涯塔。簡單說找爱，根據(jù)制造商說明樣本細胞在1 mL 的RNAiso Plus中勻漿為了分離總RNA（homogenized本意是同質(zhì)化）。使用具有g(shù)DNase 的FastQuant RT Kit將RNA重復(fù)轉(zhuǎn)錄為cDNA掘殴。10 μL樣本總量用于PCR赚瘦，30個循環(huán)。支持信息表S2包含PCR分析引物序列奏寨。MSCs的冷凍保存結(jié)果在醫(yī)院驗證過的液氮干貨機中風(fēng)干了水分起意。MSCs的冷凍保存結(jié)果使用驗證過的液氮干貨運輸工具冷凍運送到醫(yī)院（to hospitals如何翻譯，主要是ship錯位翻譯為風(fēng)干病瞳，本意是船只揽咕，船的用途是運輸貨物悲酷，所以也有運輸?shù)囊馑迹Ｔ谑褂弥靶暮郑琈SCs在室溫下通過特別專門的細胞治療中心當(dāng)?shù)貦z查實驗室被暫停重新懸浮在Plasma-Lyte 148中舔涎，并且對照組也被管理在Plasma-Lyte 148相同溶劑中（resuspended重新懸浮）逗爹。三個捐獻者MSCs的活性存活率應(yīng)該大于90%，在臨床研究指南中通過IPM小組緊急使用這是通過IPM小組指導(dǎo)的臨床研究中使用標(biāo)準(zhǔn)（viability翻譯為存活率嚎于，criterion翻譯錯誤掘而，等于standard，which is如何翻譯）于购。正如我們報告前沿描述袍睡，MSCs在第5環(huán)節(jié)第5代被使用治療（the fifth passage，第五代肋僧，passage要根據(jù)語境翻譯斑胜，頁數(shù)，段嫌吠，章節(jié)）止潘。

26個患者接受MSC移植，18個患者接受除了MSC移植外所有治療辫诅。完整的病例入組信息和包含在目前研究中44例COVID-19反復(fù)嚴重疾病患者被展示在表1中凭戴，法規(guī)目前研究中44例COVID-19患者的完成病例登記詳細信息和疾病嚴重程度頻數(shù)可以在表1中查看（severity和frequencies翻譯錯誤，嚴重程度和頻數(shù)）炕矮。在試驗組的每一個COVID-19患者使用了來自一個捐獻者的MSC樣本進行了三次治療注射么夫。醫(yī)生在旁邊至少1個小時觀察血液動力學(xué)和呼吸參數(shù)以確保每一個患者在MSC移植前穩(wěn)定。然后MSC開始被注射使用標(biāo)準(zhǔn)血液過濾管肤视。一個研究者保持在每一個病人旁邊為了在24小時內(nèi)跟蹤治療監(jiān)視病人任何不良反應(yīng)档痪。基于從研究中使用MSV治療H7N9-induced ARDS的數(shù)據(jù), MSCs注射三次給藥總量9 × 107 MSCs邢滑，間隔一天（第1腐螟，第3，第5天）殊鞭。每次注射包含3 ×107 細胞重新懸浮在500ml鹽水溶液中遭垛，以30-40滴/秒的速度進行大約15分鐘，然后100-120滴/秒速度進行2個小時保持MSC活性操灿。?

（2021年6月24日星期四翻譯575個單詞）

2.4 Biological measurements and clinical evaluation indices

Laboratory measurements of blood test results, liver function

markers, and inflammatory indicators were carried out at the Medical Inspection Center of the First Affiliated Hospital, College of Medicine, Zhejiang University and Renmin Hospital ofWuhan University. Factors which were investigated for having an association with therapeutic

features or outcomes were as following: (1) baseline characteristics including age, underlying conditions, and clinical symptoms; (2) laboratory data and chest computed tomography (CT) data; and (3) concomitant medications for basic therapy, symptomatic treatment, antiviral treatment,antibacterial treatment, hormone therapy, intestinal

microbial state regulators, extracorporeal blood purification technology, traditional Chinese medicine treatment (including Jinhua Qinggan granules, Lianhua Qingwen capsules [granules], and Shufeng Detoxification capsules[granules]), etc.

The objective of the current study was to evaluate the safety and efficacy of MSC transplantation as a treatment for COVID-19. The primary endpoint of the analysis was survival rate from January to April 2020, and this was based both on the survival of a full analysis set (FAS) and a per protocol set (PPS). The FAS, which was also the effectiveness analysis set, consisted of all 44 subjects who were initially enrolled in this study, which included 26 patients in the experimental group and 18 patients in the control group. Since two cases were initially included in the study who violated the exclusion criteria, the remaining 42 subjects were included in the PPS, which included 24 patients in the experimental group and 18 patients in the control group. The secondary endpoints for this study included measures of effectiveness and tolerability,primarily including negative viral test results, time taken to recover from all symptoms, change in chest CT results, change in indicators of inflammation, change in oxygenation index, occurrence of shock, incidence of multiple organ failure, length of hospital stay, number of days in the ICU, and respiratory support status. Before and after the MSC infusion, patients were tested using several laboratory indices, including those related to hematuria routine, liver and kidney function, coagulation function,vital signs, physical examination, oxygenation (FiO2,peripheral oxygen saturation [SpO2], oxygen saturation[SaO2], and PaO2), and inflammatory factors (including interleukin [IL]-6 and C-reactive protein [CRP]). Figure 1 shows the CONSORT diagram (Figure 1A) and detailed infusions within 1 month (Figure 1B) for this clinical study.Safety wasmeasured by the frequency of treatment-related adverse events (AEs) and through careful surveillance of laboratory indices. Clinical datawere obtained on each day of MSC infusion (days 1, 3, and 5) as well as on days 7, 14,and 30 of the posttreatment period.

2.4生物衡量和臨床評估標(biāo)準(zhǔn)

血液測量結(jié)果锯仪、肝功能指標(biāo)、炎癥指標(biāo)的實驗室檢查在浙江大學(xué)醫(yī)學(xué)院第一附屬醫(yī)院和武漢大學(xué)人民醫(yī)院的醫(yī)學(xué)檢查中心進行趾盐。與治療特征和結(jié)局相關(guān)的因素調(diào)查調(diào)查因素如下：

基線特征庶喜，包括年齡小腊，基礎(chǔ)疾病，臨床特征久窟；

實驗室數(shù)據(jù)和胸部CT數(shù)據(jù)

基礎(chǔ)治療秩冈，對癥治療，抗病毒治療斥扛，抗菌治療入问，激素治療、

腸道微生物狀態(tài)調(diào)節(jié)劑稀颁、體外血液凈化技術(shù)芬失、中藥治療等合并用藥。

這個研究的目的是評估MSC移植治療COVID-19的安全性和有效性匾灶。分析的基礎(chǔ)結(jié)局是2020年1月到4月的存活率棱烂，這是基于全分析集存活和安全性分析集的存活率。全分析集也是有效性分析集阶女，與研究中最初入組的44例患者一致颊糜，包含26例試驗組患者和18例對照組患者。由于最初的2例患者被包含在研究的排除標(biāo)準(zhǔn)中（violated什么意思秃踩，two cases were included in the study who violated the exclusion criteria這句話的主句是什么衬鱼，我判斷included是主句，后面有從句引導(dǎo)詞）吞瞪，由于違反了排除標(biāo)準(zhǔn)的兩個病例最初被納入研究中馁启，所以剩下的42例患者被納入PPS中，包括24例試驗組患者和18例對照組患者芍秆。研究的次要目的包含有效性和容忍度耐受性的測量惯疙，基礎(chǔ)主要包含陰性生命病毒檢測結(jié)果（viral錯誤翻譯為生命，生命是vivid）妖啥，所有癥狀恢復(fù)的所需時間霉颠，胸部CT結(jié)果變化，什么的炎癥指標(biāo)變化荆虱，什么的氧合指數(shù)變化蒿偎，震動休克的發(fā)生，多組織失敗多器官衰竭的發(fā)生率怀读，住院時長诉位，ICU天數(shù)，呼吸支持穩(wěn)定狀態(tài)菜枷。MSC注射前后苍糠，使用多項實驗室指標(biāo)對患者進行檢測，包括什么血尿常規(guī)啤誊，肝腎功能岳瞭，什么凝血功能拥娄，生命體征，體格檢查瞳筏，氧合稚瘾，氧氣什么血氧飽和度，和炎癥因子等相關(guān)指標(biāo)姚炕。圖1顯示了臨床研究的CONSORT圖（圖1A）和1個月內(nèi)的詳細注射摊欠。安全性測量使用與治療相關(guān)的不良事件頻率和通過監(jiān)測實驗室指標(biāo)。MSC注射的每一天（第1柱宦、3凄硼、5天）和治療后第7、14和30天獲取臨床數(shù)據(jù)捷沸。

（2021年6月25日星期五翻譯413單詞）

2.5 Statistical analysis

To compare the experimental and control groups, χ? or Fisher’s exact tests were used, as appropriate, for both the FAS analysis and the PPS analysis. The FAS analysis included 44 patients, while the PPS analysis included a subset of 42 patients. The Kaplan–Meier method was utilized

to analyze the survival time of discharged patients,and two-sided 95% exact confidence intervals (CIs) were calculated using the log-rank test. We also calculated a Cox proportional hazards model to assess factors affecting survival, adjusting for gender, and age as covariates.For the length of hospitalization and ICU stay, aWilcoxon rank sum testwas used to compare the differences between groups. For analysis of inflammatory indices, (including CRP and IL-6) and oxygenation indices (FiO2, SpO2, SaO2,and PaO2), Wilcoxon rank sum tests were used comparing the experimental group before and after the MSC infusion.Statistical analysis was carried out using SAS9.4. P value < .05 was considered as statistically significant.

2.5 統(tǒng)計分析

根據(jù)試用性，對全分析集和PPS分析集比較實驗組和對照組狐史，使用卡方或者Fisher精確檢驗痒给；全分析集包括44例患者，而PPS分析集包含42例患者的子集骏全。這個Kaplan–Meier的方法是用來分析未改變的患者的生存時間苍柏，95%雙側(cè)精確置信區(qū)間使用對數(shù)秩檢驗。為了評估影響生存的因素我們也計算一個COX風(fēng)險比例姜贡，調(diào)整性別和年齡作為協(xié)變量试吁。對于住院時間和ICP停留時間，使用Wilcoxon秩和檢驗比較組間不同楼咳。為了分析什么驗證指數(shù)和氧氣吸入氧和指數(shù)Wilcoxon秩和檢驗用來比較試驗組MSC注射前和注射后熄捍。統(tǒng)計分析使用SAS9.4，P值小于0.05被認為有統(tǒng)計學(xué)意義母怜。

2021年6月17日星期四（第3天翻譯155個詞）

3 RESULTS

3.1 Patient characteristics and MSC treatment

The viability of MSCs for the three donors was >90%(91% for SC0100919001 and SC0100919004, and 92% for SC0100919005). MSCs strongly expressed CD29, CD73,CD90, and CD105; and MSCs were negative for CD34,CD45, CD117, and HLA-DR (Supporting information Figure S1). Furthermore, MSCs can be successfully induced into osteogenic, adipogenic, and chondrogenic cells through the specific medium, and the representative picture for each kind of differentiated cells is shown in Supporting information Figure S2. A representative electrophoretogram with two pairs of primers is shown in Supporting

information Figure S3. MSC ACE2 expression was negative according to PCR analysis, with ddH2O as a negative control and 293T cells as a positive control.Twenty-six patients were included in the experimental

group and treated with MSC transplantation and combination therapy. Among these, 16 (61.5%) were classified as severe and 10 (38.5%) were classified as critical.Eighteen patients were included in the control group and received only combination therapy. Among these, 10(55.6%) were classified as severe and 8 (44.4%) were classified as critical (Table 2). The experimental group contained 17 males (65.38%) and nine females (34.62%), while the control group contained 13 males (72.22%) and five females (27.78%). The mean and median age of the experimental group was 58.31 ± 12.49 and 57.50 years, respectively, while the mean and median age of the control group was 61.11 ± 11.03 and 64.00 years, respectively. There were no statistically significant differences with regards to gender or age (P > .05). There were no significant differences

(P > .05) in clinical symptoms (including fever, expiratory dyspnea, sore throat, diarrhea, and chest tightness) between the experimental group and the control group during the baseline period (Table 2). The symptom of cough showed a significant improvement following MSC infusion on day 1 (P = .037) compared to that of the control group, but no differences were found at other times points. During the period of MSC infusion (days 1, 3,and 5) and the post-treatment period (days 7, 14, and 30), the symptoms of fever, cough, sore throat, diarrhea, and chest tightness were not significantly different (Table 2).There was a significant improvement in expiratory dyspneawhile undergoingMSCinfusion on day 1 (P=.016), day 3 (P = .040), and day 5 (P = .031) compared to the control group (Table 2), but there were no significant differences on day 7 (P = .631), day 14 (P = .635), or day 30 (P = 1.000).

3.1患者特征和MSC治療

對于三個捐獻者MSCs的存活率達到90%（SC0100919001 和SC0100919004是91余耽，SC0100919005達到92%）。

MSCs的強烈表達CD29, CD73,CD90, CD105; MSCs對 CD34,CD45, CD117, HLA-DR呈陰性（支持信息表S1）苹熏。

更重要的是此外碟贾，MSCs通過特殊的藥物特定的培養(yǎng)基可以成功的進入這些細胞誘導(dǎo)成成骨細胞、脂肪細胞和軟骨細胞轨域；每一個分化的細胞代表圖片在顯示在支持信息圖2中袱耽。一個帶著兩雙什么的兩對引物的代表性電泳圖顯示在支持信息圖S3中。在ddH2O作為陰性對照和293T 細胞陽性對照下干发，根據(jù)PCR分析MSC ACE2的表達是陰性朱巨。

26例患者包括在試驗組和MSC移植以及合并治療；在這些里面铐然，16例（61.5%）被分為重癥和10例（61.5%）被分為危重癥蔬崩。18例患者在對照組恶座，且只接受合并治療；在這些里面沥阳，10例（55.6%）被分為重癥跨琳，8例（44.4%）被分為危重癥（見表2）。試驗組中包括17例男性（65.38%）和9例女性（34.62%）桐罕，對照組中包括13例男性*（72.22%）和5例女性（27.78%）脉让。試驗組年齡的平均值和中位數(shù)分別是58.31 ± 12.49 和57.50 歲；對照組的年齡的平均值和中位數(shù)分別是61.11 ± 11.03 和 64.00 歲功炮。關(guān)于性別或年齡沒有顯著統(tǒng)計學(xué)差異（P>0.05）溅潜。在研究初期試驗組和對照組在臨床癥狀上（包括發(fā)燒，呼吸性呼吸困難薪伏，咽喉疼痛滚澜，什么腹瀉，胸部胸悶）沒有顯著統(tǒng)計學(xué)差異（表2）嫁怀∩杈瑁咳嗽的癥狀在MSC注射的第一天與對照組相比有顯著提高（P=0.037），但在其他時間點沒有差異發(fā)現(xiàn)塘淑。在MSC注射期間（第1萝招、3、5天）和注射治療期后（第7存捺、14槐沼、30天），發(fā)燒捌治、咳嗽岗钩、咽喉痛、腹瀉具滴、胸悶這些癥狀沒有顯著差異（見表2）凹嘲。當(dāng)經(jīng)過MSC注射第1天（P=0.016）、第3天（p=0.04）构韵、第5天（p=0.031）與對照組相比呼吸性呼吸困難有顯著提高周蹭，但在第7（P=0.631）、第14（p=0.635）疲恢、或第30天（p=1.000）沒有顯著性差異凶朗。

3.2 Analysis of the use of concomitant medications

As per the principles of treatment for severe and critically ill COVID-19 patients, combination drugs were allowed to treat the patients as effectively as possible. Concomitant medications were mainly used for symptomatic treatment,antiviral treatment, antibacterial treatment, hormone therapy, intestinal microbial state regulation, extracorporeal blood purification, traditional Chinese medicinal treatment, and basic disease treatment, among others.As shown in Table 3, there were no statistical differences in the types of combined medications between the two groups (P > .05). However, there were intragroup differences with regard to concomitant treatment using extracorporeal blood purification both in the experimental group (P < .001) and in the control group (P = .015). Specifically,concomitant treatmentwith an extracorporeal blood purification system was employedmore often for critically ill patients than for severe patients in both the experimental and the control groups.

3.2 使用合并用藥分析

對于重癥和危重癥COVID-19患者每一個治療原則，在盡可能有效性的情況下合并要是被允許治療病人的显拳。合并用藥主要用于對癥治療棚愤、抗病毒治療、抗菌治療、激素治療宛畦、腸道微生物狀態(tài)調(diào)節(jié)瘸洛，體外血液凈化，中藥治療次和、基礎(chǔ)疾病治療反肋、以及其他的。如表3所示踏施，在兩組間合并用藥沒有統(tǒng)計學(xué)差異（P>0.05）石蔗。然而，這里關(guān)于使用體外血液凈化合并治療有組間差異畅形，在試驗組（P<0.001）對照組（P=0.015）养距。值得注意的是，不管是試驗組還是對照組體外血液凈化合并治療的應(yīng)用日熬，危重癥患者一般多于重癥患者棍厌。

3.3 Assessment of the efficacy of MSC infusion after 1-month follow-up

The primary endpoint for this studywas the survival rate of severe and criticalCOVID-19 patientswith or without MSC infusion. Treatment efficacy was assessed throughout the study period both in the FAS and the PPS. As shown inTable 4, the survival rate for the experimental group was 92.31% (24/26) for the FAS, while the survival rate for the control group was 66.67% (12/18). This difference in survival rate was statistically significant (P = .048). Similar results were obtained when the PPS was analyzed separately.The survival curves for the FAS and the PPS are presented in Figure 2A, B, respectively. Our results suggest that MSC infusion exerts a positive therapeutic effect on the survival rate of severe and critically ill COVID-19 patients.

For severe patients, a Cox proportional hazards model of the FAS showed that the hazards ratio (HR) for the experimental group relative to the control group was .10 (95% CI, 0.00-1284.41), which was not statistically significant (P = .437; Supporting information Table S3). Using gender and age as covariates, the adjusted HR (95% CI) was .00 (0.00–∞), which was also not statistically significant (P = .970). For critically ill patients, the HR (95% CI) for the experimental group relative to the control group was .34 (.06-1.88; P = .218), and the adjusted HR (95% CI) with gender and age as covariates was .11 (.01-.89), which was statistically significant (P = .039). Similar results were obtained when the PPS was analyzed (Supporting information Table S4): the adjusted HR (95% CI) with gender and age as covariates was .12 (0.00–.95), which was statistically significant (P = .047). Thus, our results suggest that

MSC transplantation increases survival more for critically ill patients than for severe patients when adjusting for gender and age.

The secondary endpoints for this study mainly included negative viral test results, time taken to recover from all symptoms, length of hospital stay, number of days in the ICU, occurrence of shock, incidence of multiple organ failure, change in chest CT results, and respiratory support status. Detailed results are presented in Table 5. The average time taken to recover from viral infection, as indicated by a negative viral test, was 15.75 ± 13.71 and 18.31 ± 9.86 days for the experimental and the control groups, respectively. Statistical analysis showed that these differences were not significant (P = .251). The average time to improvement for the experimental group and the control groupwas 3.00 ± 3.05 and 8.80 ± 10.77 days, respectively,meaning that the average time taken to improve for the experimental group was 5.8 days shorter than that for the control group, and this difference was statistically significant (P = .049), indicating that MSC infusion was able to shorten the time required for treatment. There was no significant difference in either the length of hospital stay or in the number of days in the ICU (both P > .05), and there was also no significant difference in the occurrence of shock or multiple organ failure between the groups (both P > .05). Further, there was no statistical difference in the use of respiratory support between the two groups (Table 6). One month after MSC infusion, 20 patients in the experimental group and 12 patients in the control group underwent chest CT by three respiratory physicians who were blinded to the treatment group; 17 (85.00%) patients in the experimental group had improved,

while 3 (15.00%) patients showed no significant change. In contrast to the experimental group, six (50.00%) patients in the control group had improved, while the other six (50.00%) showed no significant change. Figure 3 shows representative CT scans documenting lung improvement at various time points for both the control and the experimental group. The difference in the improvement of chest imaging results in the first month after MSC infusion was significant. Representative CT images of both groups at post-treatment days 7, 14, and 30 are shown in Figure 3. Together, these results suggest that the relative improvement rate was higher for the experimental group during the 1 month after MSC infusion than it was for the control group.

Additionally, inflammatory indices (including CRP and IL-6) and oxygenation indices (FiO2, SpO2, SaO2, and PaO2) were analyzed before and after MSC infusion (Table 7), and there were no significant differences in CRP (P = .486), IL-6 (P = .375), FiO2 (P = .174), and SaO2 (P = .068). Interestingly, SpO2 was significantly improved following MSC infusion, from 94.72 ± 3.4% before treatment to 96.04 ± 5.93% after treatment (P < .001). Moreover,PaO2 was significantly improved following MSC transplantation,from 78.89 ± 25.86 mmHg before treatment to 95.62 ± 39.49 mmHg after treatment (P = .015).

3.3評估MSC注射一個月后追蹤的有效性

研究主要結(jié)局是重癥和危重癥COVID-19患者注射MSC和沒有注射的存活率。治療有效性基于研究期的FAS和PPS集評估竖席。如表4所示定铜，在FAS中試驗組的存活率是92.31%（24/26），對照組存活率是66.67%（12/18）怕敬。在存活率的差異具有統(tǒng)計學(xué)顯著意義。在PPS分析中獲取的結(jié)果一致帘皿。對于FAS和PPS存活周期分別見數(shù)據(jù)2A,B东跪。我們的結(jié)果表明MSC注射在重癥和危重癥COVID-19患者存活率上有積極的治療影響。

對重癥患者鹰溜，F(xiàn)AS的COX風(fēng)險比例模型顯示試驗組相對于對照組的HR是0.1（95%CI虽填，0.00-1284.41），無統(tǒng)計學(xué)顯著意義（P=0.437.支持信息表3）曹动。使用性別和年齡作為協(xié)變量斋日，調(diào)整后的HR（95%CI）是0.00（0.00–∞），也沒有統(tǒng)計學(xué)顯著意義（p=0.970）墓陈。對于危重癥患者恶守，試驗組相對與對照組的HR是0.34（0.06；P=0.218）贡必，使用性別和年齡作為協(xié)變量調(diào)整后HR（95%CI）是0.11（0.01-0.89）兔港，是有統(tǒng)計學(xué)顯著意義的（P=0.039）。PPS集分析獲取的結(jié)果相同（支持信息表4）仔拟；性別和年齡作為協(xié)變量調(diào)整后HR（95%CI）是0.12（0.00-0.95）衫樊，有統(tǒng)計學(xué)顯著意義（P=0.047）。因此，我們的結(jié)果表明當(dāng)調(diào)整性別和年齡科侈，危重癥患者比重癥患者MSC移植增加存活载佳。

研究的次要結(jié)局包括陽性病毒檢測結(jié)果、所有癥狀恢復(fù)花費時間臀栈，住院時長蔫慧，ICU天數(shù)，休克的發(fā)生挂脑，多組織衰竭發(fā)生率藕漱，胸部CT結(jié)果的改變，呼吸支持狀態(tài)崭闲。詳細的結(jié)果顯示在表5中肋联。從病毒感染到恢復(fù)耗費的平均時間，通過陽性病毒檢測作為發(fā)生刁俭，試驗組和對照組分別是15.75 ± 13.71和18.31 ± 9.86天橄仍。統(tǒng)計分析顯示差異性沒有顯著意義（P=0.251）。試驗組和對照組改善的平均時間分別是3.00 ± 3.05 和 8.80 ± 10.77天牍戚，意味著得到改善平均時間試驗組比對照組短5.8天侮繁，這個差異是有統(tǒng)計學(xué)意義的（P=0.049），表明MSC注射是能夠縮短治療所需時間的如孝。住院時長和ICU天數(shù)（P值都>0.05）都沒有顯著差異宪哩，且休克和多組織衰竭的發(fā)生也是沒有顯著性差異的。還有第晰，兩組間使用呼吸支持也沒有統(tǒng)計學(xué)差異（見表6）锁孟。MSC注射后一個月，試驗組20例患者和對照組12個患者在對照組經(jīng)歷胸部CT通過三種對治療組設(shè)盲的物理呼吸茁瘦。17（85%）患者有改善品抽，然而3（15%）顯示沒有顯著變化。與試驗組相比甜熔，對照組6（50%）患者得到改善圆恤，當(dāng)其他6例顯示沒有顯著改變。表3展示了代表性CT掃描腔稀，記錄在對照組和試驗組的多時間點的肺部改善盆昙。在MSC注射的第1個月胸部成像結(jié)果改善的差異是有顯著意義的。在治療后第7焊虏、14和30天兩組代表性CT成像被顯示在表3中弱左。總之炕淮，這些結(jié)果表明拆火，在MSC注射后1個月期間相關(guān)改善率試驗組比對照組更高.

另外，在MSC注射前后炎癥指數(shù)(including CRP and IL-6)和氧合指數(shù)（FiO2, SpO2, SaO2, and PaO2）被分析（見表7）.在CRP和 SaO2上沒有顯著性差異(P = .486), IL-6 (P = .375), FiO2 (P = .174), (P = .068)。有趣的是们镜，MSC注射后SpO2有顯著提升币叹，從治療前的94.72 ± 3.4%到治療后的96.04 ± 5.93%（P < .001）。甚至模狭，PaO2在MSC移植后有顯著增加颈抚，從治療前78.89 ± 25.86 mmHg增加到治療后95.62 ± 39.49 mmHg（P=0.015）。

（2021年6月29日星期二翻譯784個詞）

3.4 Presence of AEs following treatment

The severity of each AE related to COVID-19 infection was graded from 1 to 5 for both the experimental group and the control group, and a summary of all AEs experienced by the 44 COVID-19 patients included in the current study are presented in Supporting information Table S5. According to the statistical analysis, 76.92% (20/26) of the patients experienced a total of 56 AEs in the experimental group, including 40 AEs of grade 1, 6 AEs of grade 2, 3 AEs of grade 3, 4 AEs of grade 4, and 3 AEs of grade 5. In contrast, 100.00% (18/18) of the patients experienced a total of 59 AEs in the control group, including 39 Aes of grade 1, 5 AEs of grade 2, 3 AEs of grade 3, 7 AEs of grade 4, and 5 AEs of grade 5 (Supporting information Table S5). During the study period, no patients in either the experimental group or the control group showed Aes that necessitated withdrawal from the study. A detailed analysis of the AEs observed in the experimental group and the control group is presented in Table 8. As shown in Table 8, 10 of the 56 AEs observed in the experimental group were grade 3 or higher, while 15 of the 59 AEs observed in the control group were grade 3 or higher. Except for a difference in the incidence of high blood pressure between the two groups, there were no significant differences (P > .05) between the groups for any AEs related to the measured clinical indices including the blood test results, liver function markers, blood lipid levels,renal function,myocardial enzymes, electrolyte disturbances,and clinical symptoms. High blood pressure was observed in two (3.57%) patients in the experimental group and six (10.18%) patients in the control group (P = .048).In summary, the frequency of each AE was statistically similar between the two groups, except for the AE related to high blood pressure, which was more common in the control group. Further, the experimental group showed a lower incidence of AEs (76.92%) than the experimental group (100.00%), but the difference was not statistically significant. Together, these results suggest that the MSC infusion protocol used in this study showed good safety outcomes.

3.4 治療中AEs情況

在試驗組和對照組中嚼鹉，每一個與COVID-19感染相關(guān)AE的嚴重程度被等級劃分從1到5贩汉，通過本研究的44例COVID-19患者經(jīng)歷的所有Aes的總和被顯示在支持信息表S5中。根據(jù)統(tǒng)計分析锚赤，在試驗組中76.92% (20/26)的患者經(jīng)歷了56次AEs匹舞，其中有40個AE是等級1，3個AE是等級2线脚，3個AE是等級3赐稽，4個AE是等級4，3個AE是等級5浑侥。相比之下姊舵，對照組100.00% (18/18)的患者經(jīng)歷了59次AEs，包括39次AE等級1寓落，5次SE等級2（支持信息表5）括丁。在研究期間，無論是試驗組還是對照組沒有患者顯示由于AE需要從研究中退出試驗伶选。在試驗組和對照組中一個觀察AE的詳細分析顯示在表8中躏将。如表8所示，在試驗組中觀察的56次AEs中的10次是等級3或更高考蕾，然而對照組中觀察的59次AEs中的15次是等級三或更高，除了在兩組中高血壓不同会宪，肖卧，這里沒有顯著性差異（P>0.05），在組中任何與AE相關(guān)的臨床測量指標(biāo)掸鹅，包括血液檢測塞帐，肝功能指標(biāo)，血脂水平巍沙，腎功能葵姥，心肌酶，電解質(zhì)紊亂句携，臨床癥狀榔幸。高血壓被觀察到有2例患者（3.57%）在試驗組，有6例患者（10.18%）在對照組（P=0.048）.總的來說，在兩組中每一個AE的頻數(shù)具有統(tǒng)計相似性削咆，除了與高血壓相關(guān)的AE牍疏，在對照組有更多的共同點。進一步來說拨齐，試驗組的AEs（76.92%）比對照組的（100%）顯示更低的發(fā)生率鳞陨；但是這個差異無統(tǒng)計學(xué)意義≌巴铮總之厦滤，這個結(jié)果表明，在研究中使用MSC注射方案顯示有更好的安全性結(jié)局歼狼。

（2021年6月29日星期二翻譯374個詞）

4 DISCUSSION

The initial symptoms of COVID-19 are often fever, cough,sputum, and shortness of breath. These in turn can lead to dyspnea, ARDS, lung injury, shock, and eventual multiple organ failure.68,69 An autopsy study by Xu et al reported that a deceased COVID-19 patient showed a large amount of sputum, presumably causing severe ARDS, as well as a large number of inflammatory factors in the lung tissue.13 This suggests that effective treatment ofARDS and prevention of multiple organ failure is a key strategy in preventing mortality in COVID-19 patients. Severe ARDS causes breathing difficulties, and it has been suggested that resolving

breathing difficulties in COVID-19 patients in a timely manner may make it is possible to inhibit COVID-19 progression. Interestingly, we found that patients treated with MSCs experienced immediate and dramatic relief from breathing difficulties associated with COVID-19 on day 1 (P = .016), day 3 (P = .040), and day 5 (P = .031) compared with the control group. Further, our results show that both SpO2 and PaO2, indices associated with oxygenation levels, were significantly improved after MSC infusion. These results further support the use ofMSC infusion

as amethod to combat ARDS and expiratory dyspnea, particularly for critically ill COVID-19 patients.

Multiple complications of the COVID-19 epidemic make it difficult to fully treat patients with this disease. Many interventions in China and around the world have proven effective to reduce the epidemic and prevent the virus from continuing to spread.5,70–74 Although effective social distancing can mitigate the virus’s persistence, treating COVID-19 is also an important strategy toward ending the current pandemic.

X-ray and chestCT imaging results ofCOVID-19 patients in the ICU have been shown to reveal the presence of pneumonia,known as novel coronavirus pneumonia. These imaging results reveal bilateral, multilobular involvement as well as subsegmental consolidation.75 In the present study, we observed a statistically significant difference in the rate of improvement of chest CT results in the first month after MSC infusion. A total of 85.00% of patients with MSC treatment showed improved chest CT results, compared with only 50.00% of patients in the control group. These results provide further evidence for the efficacy of MSC infusion.

4討論

COVID-19初期的癥狀是發(fā)燒掏导，咳嗽，咳痰蹂匹、呼吸短促碘菜。這些反過來會導(dǎo)致呼吸困難、ARDS限寞，肺損傷忍啸，休克，最終的多組織衰竭履植。XU等人的尸檢報告稱一名死于COVID-19患者顯示大量的什么痰计雌，造成推測為嚴重ARDS，和肺部什么組織中大量的炎癥因子玫霎。這表明ARDS的有效治療和阻止多組織衰竭是阻止預(yù)防COVID-19患者死亡的鑰匙關(guān)鍵策略凿滤。嚴重的ARDS造成呼吸困難，它已經(jīng)表明及時解決COVID-19患者呼吸困難使抑制COVID-19發(fā)展成為可能庶近。有趣的是翁脆，我們發(fā)現(xiàn)接受MSCs患者治療在第1天（P=0.016），第3天（P=0.040）鼻种，第5天（P=0.310）反番，與對照組相比，與COVID-19相關(guān)的呼吸困難經(jīng)歷什么和什么立即和顯著的緩解叉钥。而且罢缸，我們結(jié)果顯示與氧合水平閑逛指標(biāo)SpO2 和 PaO2，在MSC注射后有顯著提高投队。這些結(jié)果更加支持MSC注射使用作為一種對抗ARDS和呼氣型呼吸困難的方法枫疆，尤其是對危重癥COVID-19患者。

COVID-19“的什么多復(fù)雜的”流行的多種并發(fā)癥讓完全治療這種疾病患者更加困難敷鸦∠⑿ǎ”大多數(shù)”中國和世界”研究者”的很多干預(yù)措施已被證明可以有效減少流行和“阻止“防止病毒繼續(xù)傳播”已經(jīng)證明有效”寝贡。雖然有效的社會社交距離可以”減少病毒傳播“減輕病毒的持久性，但是治療COVID-19也是結(jié)束當(dāng)前疾病流行重要的策略钞螟。

COVID-19患者在ICU的X射線和胸部成像結(jié)果顯示“什么“存在肺炎兔甘，即“新冠病毒的什么出名”新型冠狀病毒肺炎。這些成像結(jié)果“什么鳞滨，什么洞焙，和什么”顯示雙側(cè)，多小葉受累以及亞節(jié)段實變拯啦。在本研究中澡匪，我們觀察到在MSC注射后第一個月胸部CT結(jié)果的改善率有統(tǒng)計學(xué)顯著差異。接受MSC治療患者的85%顯示胸部CT有改善褒链，相比之下對照組只有50%的患者表現(xiàn)出改善唁情。這些結(jié)果為MSC注射有效性提供了“更多”進一步證據(jù)。

Currently, an effective vaccine would be the best way to combat SARS-CoV-2 infection, and many groups have presented preliminary basic and clinical data related to vaccine development.24,25,76–78 However, assessing the safety and efficacy of any vaccine will take a relatively long period of time. Apart from targeted vaccine development, other therapeutic strategies are being developed in the race against time to end the global pandemic. Remdesivir, an inhibitor of the viral RNA-dependent, targets nascent viral RNA chains, resulting in premature termination of the viral life cycle.23,79 Beigel et al reported that remdesivir can significantly shorten the recovery time of COVID-19 patients with lower respiratory tract infections in a doubleblind, randomized controlled clinical study.80 Corticosteroids are important immunomodulators for the clinical treatment of SARS.81 Studies have shown that compared with patients with mild to moderate disease, patients with severe SARS-CoV-2 have higher levels of proinflammatory cytokines in serum samples.82 A meta-analysis indicated that the mortality ratewas reduced in severe COVID-19 patients treated with corticosteroids.83 More recently, the recovery collaborative group reported that dexamethasone (a type of corticosteroid) significantly decreased 28-day mortality in patients hospitalized with COVID-19.22 Corticosteroids have therefore become a potential treatment

for COVID-19 patients. Compared with remdesivir

or corticosteroids, MSC infusion has the potential to significantly reduce dyspnea in a relatively short period of time, as MSCs act by targeting secretory factors and MSCreleased extracellular vesicles deliver microRNA, mRNA, or DNA.84,85 The main methods currently employed to treat COVID-19 patients (especially severe and critically ill patients) include the following:

(1) convalescent plasma therapy;

(2) antiviral drug therapy;

(3) traditional Chinese and western medicine;

(4) MSC-based therapy;

(5)immune-mediated therapy.

Studies on these methods have accelerated the screening of effective drugs, explored new treatment methods, and attempted to prevent mild cases from progressing in severity.

目前甫匹，有效的疫苗是對抗SARS-CoV-2感染最好的方式甸鸟，并且很多小組“有什么基礎(chǔ)和與”已經(jīng)提交了與疫苗發(fā)展相關(guān)初步措施和臨床數(shù)據(jù)。然而兵迅，評估疫苗的安全性和有效性會需要相當(dāng)長的時間周期抢韭。除了靶向疫苗的開發(fā)忘苛，為了結(jié)束全球流行這場與時間對抗的長跑中其他的治療策略也在逐步發(fā)展驹尼。Remdesivir 是一種病毒 RNA依賴性抑制劑，靶向新生的病毒 RNA 鏈硼一，導(dǎo)致病毒生命周期提前終止扯夭。Beigel等人報告說鳍贾，在一項雙盲、隨機對照臨床研究中Remdesivir可以顯著縮短COVID-19下呼吸道感染康復(fù)時間交洗。Corticosteroids 是一種很重要的免疫調(diào)節(jié)劑對于SARS的臨床治療骑科。研究顯示與“什么”輕度至中度疾病患者相比，重癥SARS-CoV-2患者在“什么”血清樣本中“有更高的什么”促炎細胞因子水平更高构拳。一項meta分析表明在重癥COVID-19患者“什么”接受皮質(zhì)類固醇治療中死亡率有所減少咆爽。近期，康復(fù)collaborative組報告了dexamethasone（一種皮質(zhì)類固醇）顯著降低COVID-19住院患者28天死亡率隐圾。因此Corticosteroids已成為新冠患者的潛在治療方法。與remdesivir 或 corticosteroids相比掰茶，MSC注射在相對較短的時期內(nèi)有一個顯著減少呼吸困難暇藏，當(dāng)MSC通過靶向“什么因素”分泌因子和MSC釋放的細胞外囊泡傳遞microRNA, mRNA,or DNA的MSC起作用。對治療新冠患者近期應(yīng)用主要方法包括如下：

“什么”恢復(fù)期血漿治療

抗病毒藥物治療

中西藥結(jié)合

基于MSC治療

免疫介導(dǎo)治療

這些研究方法“什么”加速有效藥物篩選濒蒋，探索新的治療方法和嘗試在嚴重過程中阻止“什么的案例”輕癥病情惡化盐碱。

（2021年6月30日星期三翻譯654個詞）

The current clinical study reports that MSC infusion enhanced the survival rate for severe or critically ill COVID-19 patients in both the FAS (92.31% survival in the experimental group vs 66.67% in the control group;P = .048) and the PPS (95.83% survival in the experimental group vs. 66.67% in the control group; P = .031).Our results agree with a previous report detailing the potential for MSC infusion to treat critically ill COVID-19 patients by alleviating acute respiratory dysfunction and pulmonary fibrosis.91 There are some other sources of MSCs used in clinical studies for treating COVID-19, and these are included in a systematic review and metaanalysis. Leng et al53 used UC-MSCs to treat seven COVID-19 patients, and had three control patients who were infected with COVID-19; there was 100% survival in the experimental group versus 66.67% survival in the control group. Shu et al95 investigated 12 severe COVID-19 patients using UC-MSCs as the experimental group, and 29 severe COVID-19 patients using a placebo as the control group. Their results showed a 100% 28-day survival rate in the experimental group versus 89.66% survival in the control group. More recently, Meng et al57 performed a study in 18 hospitalized patients with moderate to severe COVID-19 pulmonary disease, 9 of whom were treated with UCMSCinfusions andwith 9 control patients. All patients survived survived both in the experimental group and in the control group, but the degree of severity of COVID-19 (moderate to severe) might have affected the outcome. Leng et al,53 Shu et al,95 and our study enrolled more severe or critically ill COVID-19 patients. Recently, Sánchez-Guijo et al investigated 13 severe COVID-19 patients using an ADMSCs infusion for a clinical study that was nonrandomized and without a control; the results showed 84.62% survival using an AD-MSCs infusion. Therefore, MSC transplantation from different sources appears to be a candidate method to improve outcomes for critical cases. Coagulopathy and thromboprophylaxis are very common after MSC infusion.15 Current clinical data indicate that MSCs from human menstrual blood does not clot in patients, which is very favorable for intravenous infusions. One possible reason could be that MSCs express a low level of procoagulant tissue factor TF/CD142, which needs to be systematically investigated and verified in future preclinical studies.Although different sources of MSCs have been investigated for effectiveness in treating COVID-19, more optimized treatment strategies are critical to evaluate and control blood compatibility, optimize cell transfusion, and monitor the real-time dynamics of cells in the body to develop safer and more effective MSC treatments.43 Several concomitant treatments have been shown to exert a synergistic rolewith MSC transplantation. Of note, Peng et al reported that intravenous infusion of convalescent plasma as a treatment for severe COVID-19 may have synergistic characteristics with MSC transplantation in inhibiting cytokine storms,promoting lung injury repair, and recovery of pulmonary function. Although further study is required to establish safety and efficacy, the current body of evidence suggests that MSC transplantation might be an effective treatment for severe and critical COVID-19.

最近的臨床研究表明MSC注射對重癥和危重癥COVID-19患者在FAS（試驗組存活率92.31%VS對照組存活率66.67%把兔；P=0.048）和PPS（試驗組存活率95.83%VS對照組存活率66.67%；P=0.031）中都提升了存活率瓮顽。我們的結(jié)果“同意一個什么的報告”和之前的一份報告一致县好，詳細說明了MSC注射通過緩解急性呼吸阻礙和肺部纖維化來治療危重癥COVID-19患者潛力。這兒有一些其他來源的MSC用于治療COVID-19臨床研究暖混，這些都包含在系統(tǒng)綜述和薈萃分析中缕贡。

Leng等人使用UC-MSCs治療了7例COVID-19患者，有3例感染COVID-19對照組患者拣播，在試驗組中有100%的存活率晾咪，相對來說在對照組中有66.67%存活。Shu等人調(diào)查了12例使用UC-MSCs的 COVID-19重癥患者作為試驗組贮配，29例重癥患者使用安慰劑的COVID-19重癥患者作為對照組谍倦。他們的結(jié)果顯示試驗組28天內(nèi)存活率是100%，相對來說對照組存活率是89.66%泪勒。近期昼蛀，Meng等人實行了在18個住院患者緩解重癥新冠肺部疾病，其中9個UC-MSC注射治療圆存，9個對照患者叼旋。在試驗組和對照組中所有患者都存活了，但是COVID-19的嚴重程度可能會影響結(jié)局辽剧。Leng和Shu等人和我們的研究招募了更重癥和危重癥新冠患者送淆。最近，Sánchez-Guijo等人調(diào)查了13例使用ADMSCs新冠患者進行了一場不隨機無對照的臨床試驗怕轿。這個結(jié)果顯示使用AD-MSCs注射的有84.62%存活率偷崩。因此，從不同來源的MSC移植顯示是一種“什么樣的”改善危重癥案例結(jié)果候選方法撞羽。在MSC注射后阐斜， Coagulopathy 和thromboprophylaxis是非常普遍的。近期臨床數(shù)據(jù)顯示來自人類經(jīng)血的MSC在患者中沒有凝塊诀紊，“它對于什么注射是非常好的”它非常有利于靜脈注射谒出。一個可能的原因可能是MSC表達“出什么因素”低水平促凝血因子TF/CD142，在未來臨床研究上是需要更加系統(tǒng)調(diào)查和“多樣化”驗證邻奠。雖然MSCs不同來源治療COVID-19有效性已經(jīng)有研究笤喳，但更加優(yōu)化的策略是非常重要的去評估和控制血液什么相容性，優(yōu)化細胞輸注碌宴，檢測體內(nèi)細胞的實時動態(tài)杀狡，發(fā)展更加安全和有效的MSC治療。

許多合并治療顯示MSC移植發(fā)揮了協(xié)同作用角色贰镣。值得注意的是呜象，Peng等人報道了膳凝，作為重癥新冠治療方法恢復(fù)期血漿靜脈注射，與在抑制細胞因子風(fēng)暴的MSC細胞移植促進肺損傷修復(fù)和肺功能修復(fù)上可能有協(xié)同作用特性恭陡。雖然研究要求進一步的建立安全性和有效性蹬音，目前證據(jù)表明MSC移植可能對重癥和危重癥COVID-19患者治療有效。

In the current study, it was observed that many clinical symptoms were ameliorated in the 1-month period following MSC transplantation with combined therapy. No cases of pulmonary embolism were observed in the patients who underwent MSC infusion, although this side-effect is considered to be the main concern regarding MSC safety. Rather, our results indicate that MSC therapy is a safe and effective therapeutic strategy to rescue severe and critical lung problems induced by SARS-CoV-2. Thus, in the present study, preliminary clinical datawere provided with regard to the short-term safety (1-month follow-up) and therapeutic effect of MSC transplantation to treat severe and critically ill patients.

在本研究中休玩，觀察到在MSC移植和合并治療一個月期間許多臨床癥狀有所改善著淆。“即使是重點關(guān)注MSC安全性作為單個影響哥捕，在經(jīng)歷MSC注射的患者中也沒有觀察到肺栓塞案例”牧抽，在接受MSC注射的患者中沒有觀察到肺栓塞病例，盡管這種副作用被認為是MSC安全性的主要問題遥赚。而且扬舒，我們的結(jié)果表明MSC治療是安全有效的治療策略對于減輕SARS-CoV-2感染的重癥和危重癥肺部問題。因此凫佛，在本研究中“什么”初步臨床數(shù)據(jù)作為治療重癥和危重癥患者短期安全性和MSC移植的治療效果被提供讲坎。

The current study does not provide long-term evidence related to MSC-induced AEs. In a previous report from our group, MSCs were used to treat 17 H7N9 patients and four of those patients were followed up for 5 years without observing any AEs.55 In the current study of severe and critical COVID-19 patients, almost no significant differences in the occurrence of AEs in the short term were observed between the control group and the group receiving MSC transplantation. Hence, this study found that MSC infusion is associated with good safety outcomes. In addition,although the potential application value of MSC treatment in COVID-19 is obvious, the innate and adaptive immune compatibility test of MSC is incorporated into the current cell detection system, and the establishment of strict monitoring standards for biosafety and effectiveness with regard to the coagulopathy and thromboprophylaxis is crucial.15 Researchers should also pay strict attention to obtaining a suitable source, the product quality, monitoring of the various physiological and biochemical indicators of patients throughout the process, and strict prevention of unnecessary safety hazards.

本研究沒有提供與MSC誘導(dǎo)AEs相關(guān)的長期證據(jù)。在我們組的一個前期報告愧薛，使用MSCs治療的17例H7N9患者晨炕，其中4例被追蹤5年沒有被觀察到AEs。在本研究中毫炉，重癥和危重癥新冠患者瓮栗，在對照組和接受MSC移植組中短期觀察的AEs發(fā)生率幾乎沒有顯著性差異。因此瞄勾，MSC注射和良好的安全性結(jié)局有關(guān)费奸。另外，雖然在MSC治療COVID-19的潛在應(yīng)用價值是明顯的进陡，“MSC的什么和什么的免疫能力測試是什么進入細胞方向系統(tǒng)”MSC的先天和適應(yīng)性免疫相容性測試是納入現(xiàn)有細胞檢測系統(tǒng)愿阐，并且在凝血功能障礙和血栓預(yù)防上生物安全性和有效性嚴格的早期標(biāo)準(zhǔn)建立是至關(guān)重要的。研究者應(yīng)該更加嚴格的關(guān)注為了獲取一個合適的來源趾疚，產(chǎn)生質(zhì)量缨历，監(jiān)控患者多種生理和生化指標(biāo)，并且嚴格阻止不需要的安全性隱患糙麦。

Recently, Leng et al53 published a clinical study on using MSCs to treat COVID-19 patients. This study investigated inflammatory and immune functioning as well as adverse effects in seven patients for 14 days post-MSC transplantation. The authors reported that MSCs appeared to significantly improve the functional outcomes of all seven patients without any observed AEs. However, more clinical data are still needed to identify any short-term adverse reactions following MSC administration. Zheng et al recently reported that 12 patients with moderate to severe ARDS did not experience any infusion-related reactions or serious treatment-related AEs following MSC transplantation. Additionally, Meng et al57 performed a study that included 18 hospitalized patients with moderate to severe COVID-19 pulmonary disease, nine of whom were treatedwithUC-MSC infusions. According to their results, intravenous UC-MSC infusion was safe and well-tolerated throughout the 1-month follow-up period. Although longterm follow-up data regarding the tolerability and safety of MSC transplantation are lacking, MSC transplantation may still be an effective method treatment for COVID-19, especially for severe and critical cases.

近期辛孵，leng等人發(fā)表了關(guān)于使用MSCs去治療COVID-19患者臨床研究。這個研究調(diào)查了7個患者MSCs移植14天后的炎癥和免疫功能以及不良事件赡磅。這個作者報道了MSCs顯示出顯著提高功效結(jié)局

7例患者中沒有觀察到任何AE魄缚。然而，更多的臨床數(shù)據(jù)需要去發(fā)現(xiàn)MSC管理中任何短期不良反應(yīng)仆邓。Zheng等人近期報道12例中中度患者在MSC移植中沒有經(jīng)歷任何注射相關(guān)反應(yīng)或者嚴重治療相關(guān)AEs鲜滩。另外，Meng等人開展了一個研究节值，包括18例住院帶有中重度COVID-19肺部感染患者徙硅，其中9例使用UC-MSC注射治療。根據(jù)他們的結(jié)果搞疗，在整個1個月的隨訪期靜脈注射是安全和良好的耐受性嗓蘑。雖然作為MSC移植的耐受和安全性長期隨訪數(shù)據(jù)是缺乏的，MSC移植對COVID-19特別是重癥和危重癥案例仍然是有效治療方法匿乃。

There are several limitations to this exploratory trial. First and foremost, one patient experienced an AE over grade 3 associated with severe abnormal liver function. More data from a larger study are needed to determine whether MSC infusion can lead to an infusion reaction in certain patients, especially as infusion reactions can cause shortness of breath. Since only 26 patients were treated with MSCs in the current study and because we used a multicenter, open, and parallel-controlled study design, our study may not have detected important AEs associated with MSC treatment. Secondly, we should stress that this clinical trial did not use a standard design owing to the unique nature of the COVID-19 outbreak and the ethical limitations associated with treating severe COVID-19 patients. Thirdly, we reported that the experimental group showed greater improvements in CT results than the control group during the 1-month study period. However, we only observed significant improvements in expiratory dyspnea for the experimental group versus the control group on days 1, 3, and 5, and no differences were observed between the groups at other time points with regard to lung function. We speculate that MSCs exert a short-term effect to improve expiratory dyspnea, while the long-term improvements were more due to the actions of concomitant medications. Moreover, we did not observe significant changes in the levels of inflammatory factors following MSC infusion, even though mortality was significantly decreased in the experimental group. Therefore, the mechanism by whichMSCs reduce mortality should be investigated in future more comprehensive clinical trials. Moll et al investigated that the differences between fresh and cryopreserved MSCs were limited but significant.100 FreshMSCs are the best choices, however, considering the COVID-19 outbreak, the lack of sufficient donors for providing menstrual blood in a short time, and the consistent standard of care between the two hospitals situated in different cities, freshly thawed MSCs were used in this study. Finally, the small sample size of the current study limited our ability to obtain enough clinical data to draw strong conclusions.

這是探索性試驗的一些限制桩皿。首先也是最重要的，一個患者經(jīng)歷了一次與嚴重非正常肺功能超過等級3的AE幢炸。需要來自更大研究更多的數(shù)據(jù)去確認MSC注射是否會導(dǎo)致在確定患者上的注射反應(yīng)泄隔，尤其是作為注射反應(yīng)會引起呼吸短促。由于在本研究中僅僅只有26個患者接受MSCs治療宛徊，因為我們采用的是多中心佛嬉、開放、和平行對照研究設(shè)計闸天，我們的研究沒有發(fā)生與MSC治療相關(guān)的嚴重AEs暖呕。第二，我們需要強調(diào)這個臨床研究沒有使用標(biāo)準(zhǔn)設(shè)計苞氮，由于COVID-19爆發(fā)獨特性質(zhì)和與治療重癥COVID-19患者倫理限制（owing有歸因于的意思）湾揽。第三，我們報告了在一個月研究期內(nèi)試驗組比對照組在CT結(jié)果上顯示更大的改善笼吟。然而库物，我們僅僅只觀察到試驗組對于對照組在第1、3赞厕、5天呼氣型呼吸困難有顯著改善艳狐，根據(jù)肺部功能組之間在其他時間點上沒有觀察到差異。我們推測MSCs對于改善呼吸性呼吸苦難發(fā)揮了短期效果皿桑，然而長期效果更加取決于合并用藥的反應(yīng)毫目。甚至，MSC注射后诲侮。在炎癥因子水平上我們并沒有觀察到顯著變化镀虐，即使在試驗組死亡率有顯著減少。因此沟绪，通過MSC減少死亡率這種機制在未來更多綜合臨床試驗中應(yīng)該被研究刮便，新鮮的和冷凍保存的MSCs之間的差異是有限的但很重要。新鮮的MSCs是最好的選擇绽慈，然而恨旱，考慮到COVID-19爆發(fā)辈毯，短期內(nèi)提供經(jīng)血足夠捐獻者缺乏，不同城市兩個醫(yī)院間一致標(biāo)準(zhǔn)的護理標(biāo)準(zhǔn)搜贤，本研究使用了新鮮解凍的MSCs谆沃。最后，本研究的小樣本限制我們獲取足夠臨床數(shù)據(jù)去挖掘更有力結(jié)論的能力仪芒。

5 CONCLUSIONS

This prospective and systematic study assessed the ability of menstrual blood-derived MSCs to treat both severe and critically ill COVID-19 patients. The results of this multicenter,open, and parallel-controlled clinical study suggest that menstrual blood-derived MSC transplantation significantly lowers the mortality of severe and critical SARSCoV-2-induced COVID-19. Menstrual blood-derivedMSCs may act by alleviating the breathing difficulties caused by COVID-19 and reducing the symptoms of ARDS or expiratory dyspnea. Although the body of research on MSCs is still in its infancy and lacks important long-term safety information, MSC-based therapymay serve in future clinical applications as an alternativemethod for the treatment of COVID-19.

5.結(jié)論

這個前言且系統(tǒng)的研究評估了經(jīng)血來源MSCs治療重癥和危重癥COVID-19患者的能力唁影。這個多中心、開放掂名、平行對照臨床研究結(jié)果表明對于經(jīng)血來源MSC移植治療治療重癥和危重癥COVID-19患者顯著降低死亡率（lower是動詞据沈，降低）。經(jīng)血來源MSCs可能起作用是通過緩解由COVID-19引起的呼吸困難和減少減輕ARDS或者呼吸性呼吸困難的癥狀饺蔑。盡管在MSCs研究主題仍然它的初期在起步階段且缺乏重要長期的安全性信息锌介，在未來臨床應(yīng)用中MSC基礎(chǔ)基于MSC治療可能會作為一項治療COVID-19有前景的替代方案（serve as作為什么服務(wù)于什么）。