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梁月英,李舒婷,朱家源,谢肖霞,谢小英(中山大学附属第一医院烧伤科,广东广州,510080)

[基金项目] * 本课题为国家自然科学基金项目,项目编号为 30973128。

[收稿日期] 2014-04-12

[作者简介] 梁月英(1971-),女,广东台山人,主管护师,本科,主要从事烧伤临床护理工作。

[通信作者] 朱家源,教授,Enail:该Email地址已收到反垃圾邮件插件保护。要显示它您需要在浏览器中启用JavaScript。

[摘要] 目的 观察 ReCell 细胞再生技术在糖尿病足治疗中的应用效果,并探讨其主要的护理措施。

                方法 按治疗方法将 40 例糖尿病足患者分为对照组与实验组,每组各 20 例,在创面床准备后,对照组采用传统的游离皮片移植手术方法,实验组采用 ReCell 细胞再生技术 + 游离皮片移植术,观察两组患者皮片成活情况及治疗时间。

                结果 实验组患者皮片存活情况优于对照组;治疗时间较对照组短,两组比较,差异均有统计学意义(P<0.05)。

                结论 在糖尿病足患者治疗中应用 ReCell 细胞再生技术可使皮瓣成活率高,缩短治疗时间。完善相关护理措施是 ReCell 细胞再生技术取得成功的根本保证。

[关键词] ReCell 细胞再生技术;糖尿病足;护理

[中图分类号]R473.6 [文献标识码]A [文章编号]1671-8283(2014)07-0009-03 [DOI]10.3969/j.issn.1671-8283.2014.07.003

Application of ReCell technique in the treatment of diabetic foot Liang Yueying, Li Shuting, Zhu Jiayuan, Xie Xiaoxia, Xie Xiaoying//Modern Clinical Nursing,-2014,13(07):09.

[Abstract] Objective To investigate the effect of ReCell technique on diabetic foot and explore main nursing measures.

Methods Forty patients with diabetic foot were divided into control group(n=20)and experiment group(n=20)according to admissiontime. With wound bed prepared, the control group received only free skin flap grafting and the observation group was managed with ReCell technique plus free skin flap grafting.The skin flap survival and treatment time in two groups were observed and compared.

Result The skin flap survival in the experiment group was better than the control group and the treatment time was shorter.

Conclusion ReCell technique is effective in raising the survival rate of skin flap in diabetic foot and shortening treatment time.The improvement of nursing measures is critical for the success of ReCell technique.

[Key words] ReCell technique; diabetic foot; nursing

黄韶斌 1 胡志成 1 张逸 2 唐冰 1 王鹏 1 徐海琳 1 王志勇 1 董云先 1 程璞 1 荣燕超 1 吴军 3 朱家源 1

1 中山大学附属第一医院烧伤科,广州 510080;

2 南通大学附属医院烧伤整形外科226001;

3 深圳大学第一附属医院烧伤整形科 518037

通信作者:朱家源,Email:该Email地址已收到反垃圾邮件插件保护。要显示它您需要在浏览器中启用JavaScript。

【摘要】

目的 探讨异体表皮干细胞(ESC)对裸鼠全层皮肤缺损创面移植异体全厚皮成活的影响及其机制。方法 采用实验研究方法。采用酶消化法从 1 只 4 周龄雄性 BALB/c-NU 裸鼠(品系、鼠龄与性别下同)中获取培养 7 d 呈铺路石状原代 ESC,其第 3 代细胞经流式细胞仪鉴定阳性表达ESC 标志物 CD44 且阴性表达 CD45,经免疫荧光法鉴定阳性表达 ESC 标志物 p63 与整合素 6α 且阴性表达 CD71。取对数生长期的第 3 代 ESC 进行后续实验。取 26 只裸鼠,采用随机数字表法平均分为磷酸盐缓冲液(PBS)组和 ESC 组,在每只裸鼠背部制备全层皮肤缺损创面后,2 组创面分别喷涂等体积的 PBS、ESC,创面上均移植从另外 4 只裸鼠背部切取制备的全厚皮。分别取 2 组 10 只裸鼠,观察术后0(即刻)、3、7、14、21 d 创面愈合与皮片存活情况并计算术后 3、7、14、21 d 皮片存活比和皮片收缩率(样本数为各时间点存活皮片数),采用激光散斑血流成像仪检测术后 3、7、14 d 皮片的血流灌注情况并计算各时间点 ESC 组与 PBS 组裸鼠皮片血流灌注比(样本数为各时间点 2 组配对均存活皮片对数);分别取 2 组剩余 3 只裸鼠,取术后 7 d 皮片组织,分别采用实时荧光定量反转录 PCR 法和蛋白质印迹法检测组织中肿瘤坏死因子 α(TNF-α)、白细胞介素 8(IL-8)、IL-10、Ⅰ型胶原、Ⅲ型胶原与基质金属蛋白酶 9(MMP-9)的 mRNA 和蛋白表达。对数据行 Log-rank 检验、重复测量方差分析、单因素方差分析、独立样本 t 检验与 Bonferroni 校正。

结果 以术后 0 d 情况为参照,2 组裸鼠术后 3、7、14、21 d 创面逐渐愈合,皮片收缩情况逐渐明显,其中 PBS 组裸鼠创面收缩愈合情况较 ESC 组明显。术后3 d,ESC 组 1 只裸鼠皮片移植失败,PBS 组 3 只裸鼠皮片移植失败;术后 7 d,PBS 组又有 1 只裸鼠皮片移植失败。2 组裸鼠术后 3、7、14、21 d 皮片存活比相近(P>0.05)。术后 3、7、14、21 d,ESC 组裸鼠皮片收缩率分别为(9.2±0.4)%、(19.7±1.2)%、(53.6±3.5)%、(62.2±5.1)%,显著低于 PBS 组的(11.0±0.9)%、(47.8±2.8)%、(86.1±7.1)%、(89.7±9.0)%(t=5.719、26.650、11.940、7.617,P<0.01)。术后 3、7、14 d,2 组裸鼠皮片均有血流灌注信号;ESC 组与 PBS 组裸鼠皮片血流灌注比均值均大于 1,3 个时间点总体比较,差异无统计学意义(P>0.05)。术后 7 d,与 PBS 组比较,ESC 组裸鼠皮片组织中 TNF-α、IL-8、Ⅰ型胶 原 和 Ⅲ 型 胶 原 的 mRNA(t=2.823、2.934、2.845、2.860,P<0.05)和 蛋 白 表 达 均 明 显 降 低 ,IL-10 和MMP-9 的 mRNA(t=3.877、2.916,P<0.05)和蛋白表达均明显升高。

结论 异体 ESC 可减轻裸鼠全层皮肤缺损创面移植异体全厚皮片收缩,促进移植皮片与创面之间新生血管的形成,减轻炎症反应,降低胶原蛋白表达,促进 MMP-9 的表达,从而提高移植皮片的成活质量。

【关键词】 干细胞移植; 皮肤移植; 伤口愈合; 表皮干细胞

基金项目:国家自然科学基金面上项目(81871565);广东省科技计划(2016B090916001);广东省基 础 与 应 用 基 础 研 究 基 金(2019A1515012208);中 山 大 学 临 床 医 学 研 究 5010 计 划(2013001、2018003);中山大学高校基本科研业务费(19ykpy66)

      Effects and mechanisms of allogeneic epidermal stem cells on the survival of allogeneic full-thickness skin grafts in nude mice with full-thickness skin defect wounds Huang Shaobin1, Hu Zhicheng1, Zhang Yi2, Tang Bing1, Wang Peng1, Xu Hailin1, Wang Zhiyong1Dong Yunxian1, Cheng Pu1, Rong Yanchao1, Wu Jun3, Zhu Jiayuan1

1 Department of Burns, the First Affiliated Hospital of Sun Yat-sen University, Guangzhou 510080, China

2 Department of Burns and Plastic Surgery, Affiliated Hospital of Nantong University, Nantong 226001, China ;;

3 Department of Burns and Plastic Surgery, the First Affiliated Hospital of Shenzhen University, Shenzhen 518037, China Corresponding author: Zhu Jiayuan,

Email: 该Email地址已收到反垃圾邮件插件保护。要显示它您需要在浏览器中启用JavaScript。

【Abstract】

      Objective To investigate the effects and mechanisms of allogeneic epidermal stem cells (ESCs) on the survival of allogeneic full-thickness skin grafts in nude mice with full-thickness skin defect wounds. Methods Experimental research methods were applied. Primary ESCs that appeared paving stone-like after being cultured for 7 d were obtained by enzymatic digestion method from one 4-week-old male BALB/c-NU nude mouse (the same strain, age, and sex below). The cells of third passage were identified by flow cytometry to positively express ESC marker CD44 and negatively express CD45, meanwhile, the positive expression of ESC markers of p63 and integrin 6α, and negative expression of CD71 were identified by immunofluorescence method. The ESCs of third passage in the logarithmic growth phase were used for the following experiments. Twenty-six nude mice were equally divided into phosphate buffered saline (PBS) group and ESCs group according to the random number table. A full -thickness skin defect wound was made on the back of each nude mouse, and then the wounds of the two groups were sprayed with equal volumes of PBS and ESCs, respectively. The wounds were transplanted with full -thickness skin grafts cut from the backs of 4 other nude mice. Each ten nude mice from the two groups were selected, the wound healing and skin survival on post surgery day (PSD) 0 (immediately), 3, 7, 14, and 21 were observed, and the survival ratio and shrinkage rate of skin grafts on PSD 3, 7, 14, and 21 were calculated (the number of sample was the number of surviving skin grafts at each time point); the blood perfusion in the skin grafts on PSD 3, 7, and 14 was detected by the laser speckle blood flow imager, and the blood flow ratio of nude mice skin grafts in ESCs group to PBS group at each time point was calculated (the number of sample was the pair number of surviving skin grafts in group pairing at each time point). The skin graft tissue of each 3 nude mice remained in the two groups were collected on PSD 7, and the mRNA expressions and protein expressions of tumor necrosis factor α (TNF-α), interleukin 8 (IL-8), IL-10, type Ⅰ collagen, type Ⅲ collagen, and matrix metalloproteinase 9 (MMP-9) in the tissue were detected by real-time fluorescent quantitative reverse transcription polymerase chain reaction and Western blotting, respectively. Data were statistically analyzed with Log-rank test, analysis of variance for repeated measurement , one-way analysis of variance , independent sample test, and Bonferroni correction. Results Taking the condition on PSD 0 as a reference, the wounds of nude mice in the two groups healed gradually on PSD 3, 7, 14, and 21, and the shrinkage of skin grafts was gradually obvious. Among them, the shrinkage healing of wound of nude mice in PBS group was more significant than that in ESCs group. On PSD 3, the skin graft of 1 nude mouse failed in ESCs group, while the skin graft of 3 nude mice failed in PBS group. On PSD 7, the skin graft of another nude mouse failed in PBS group. The survival ratio of skin grafts of nude mice in the two groups was similar on PSD 3, 7, 14, and 21 (P>0.05).On PSD 3, 7, 14, and 21, the shrinkage rates of skin grafts of nude mice in ESCs group were (9.2±0.4)% , (19.7±1.2)% , (53.6±3.5)% , and (62.2±5.1)% , respectively, which was significantly lower than (11.0±0.9)% , (47.8±2.8)% , (86.1±7.1)% , and (89.7±9.0)% in PBS group (t=5.719, 26.650, 11.940, 7.617, P<0.01). On PSD 3, 7, and 14, blood perfusion signals were observed in the skin grafts of nude mice in the two groups. The average blood perfusion ratios of the skin grafts of nude mice in ESCs group to PBS group were greater than 1, and there was no statistically significant difference in the overall comparison of 3 time points (P>0.05). On PSD 7, compared with those of PBS group, the mRNA and protein expressions of TNF-α, IL-8, type Ⅰ collagen, and type Ⅲ collagen in the skin graft tissue of nude mice in ESCs group were significantly reduced, while the mRNA and protein expressions of IL-10 and MMP-9 in the skin graft tissue of nude mice in ESCs group were significantly increased (in mRNA comparison, t=2.823, 2.934, 2.845, 2.860, 3.877, 2.916, P<0.05). Conclusions Allogeneic ESCs can reduce the shrinkage of allogeneic full-thickness skin grafts transplanted on full-thickness skin defect wounds in nude mice, promote the formation of new blood vessels between the skin graft and the wound , reduce inflammation and collagen protein expression, and promote the expression of MMP -9, thus improving the survival quality of skin grafts.

【Key words】 Stem cell transplantation; Skin transplantation; Wound healing; Epidermal stem cells

Fund program : General Program of National Natural Science Foundation of China (81871565); Science and Technology Planning Project of Guangdong Province of China (2016B090916001); Foundation for Basic and Applied Basic Research of Guangdong Province of China (2019A1515012208); Sun Yat-sen University Clinical Research 5010 Program (2013001, 2018003); Fundamental Scientific Research of Sun Yat-sen University (19ykpy66)

Shaobin Huang1,2† , Zhicheng Hu1† , Peng Wang1 , Yi Zhang3 , Xiaoling Cao1 , Yunxian Dong1 , Pu Cheng1 , Hailin Xu1, Wenkai Zhu4 , Bing Tang1* and Jiayuan Zhu1*

* Correspondence: 该Email地址已收到反垃圾邮件插件保护。要显示它您需要在浏览器中启用JavaScript。; 该Email地址已收到反垃圾邮件插件保护。要显示它您需要在浏览器中启用JavaScript。 † Shaobin Huang and Zhicheng Hu contributed equally to this work.

1 Department of Burn, First Affiliated Hospital of Sun Yat-sen University, Guangzhou 510080, Guangdong Province, People’s Republic of China Full list of author information is available at the end of the article

Abstract

Background: Full-thickness wounds severely affect patients’ life quality and become challenging problems for clinicians. Stem cells have great prospects in the treatment of wounds. Our previous study confirmed that autologous basal cell suspension could promote wound healing, and epidermal stem cells (ESCs) were detected in the basal cell suspension. Herein, this study aimed to explore the effect of ESCs on full-thickness wounds.

Methods: Rat ESCs were isolated and expanded and then were transfected with lentivirus to stably express enhanced green fluorescent protein. The experimental rats were randomly divided into 2 groups: in the ESC group, the rat ESCs were sprayed on the full-thickness wounds of rats; in the control group, phosphate-buffered saline was sprayed the on the wounds. Next, wound healing and neovascularization were evaluated. Colonization, division, and differentiation of ESCs on the wound were analyzed by immunofluorescence.

Results: The rat ESCs colonized, divided, and proliferated in the wound. Additionally, rat ESCs around blood vessels differentiated into vascular endothelial cells and formed a lumen-like structure. Compared with the control group, the ESC group showed enhanced angiogenesis and accelerated wound healing.

Conclusions: Our study confirmed that rat ESCs are safe and effective for treating full-thickness wounds. Additionally, under certain conditions, ESCs can differentiate into vascular endothelial cells to promote angiogenesis and wound healing.

Keywords: ESCs, Cell differentiation, Angiogenesis, Full-thickness wounds

Z .-C. Hu1, D. Chen2, D. Guo3, Y.-Y. Liang1, J. Zhang1, J.-Y. Zhu1 and B. Tang1

Departments of 1Burn Surgery and 2Hepatobiliary Surgery, The First Affiliated Hospital of Sun Yat-sen University, and 3Department of Plastic Surgery, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, China

Correspondence to: Dr B. Tang, Department of Burn Surgery, The First Affiliated Hospital of Sun Yat-sen University, 58 Zhongshan Road II, Guangzhou 510080, China (e-mail: 该Email地址已收到反垃圾邮件插件保护。要显示它您需要在浏览器中启用JavaScript。)

Background: Treatment of chronic wounds using traditional surgical procedures is challenging because of the low graft take rates. This study investigated the combination approach of split-thickness autografts with harvested skin cell suspension for chronic wound treatment.

Methods: This randomized clinical trial enrolled patients with chronic wounds between March 2012 and December 2013. Patients who were assigned randomly to the active treatment received a split-thickness autograft combined with harvested skin cell suspension. Control patients received the split-thickness autograft alone. The primary outcome was the rate of complete wound closure by postoperative day 28. Analysis was by intention to treat. Patients who achieved wound closure were followed up for a minimum of 6 months to evaluate the quality of healing.

Results: A total of 88 patients were included, 44 in each group. More patients achieved complete wound closure in the skin cell group than in the control group (41 versus 34 patients; P = 0035). Complete wound closure was observed at a median of 14 (95 per cent c.i. 120 to 160) days in the skin cell group and 20 (157 to 243) days in the control group (P = 0001). The skin cell group had significantly fewer complications (4 versus 11 patients; P = 0047). The autografted sites displayed better physical attributes and a reduced tendency for wound recurrence in the skin cell group.

Conclusion: Complementary split-thickness autologous skin grafting with autologous skin cells harvested using ReCell® (Avita Medical, Cambridge, UK) technology improved the healing rate of chronic wounds. Registration number: UMIN000011966 (http://www.umin.ac.jp/ctr).