Department of Urology, School of Medicine, Kyungpook National University, Daegu, Korea.
BMC Med. 2012 Aug 21;10:94. doi: 10.1186/1741-7015-10-94.
Stem cell injection therapies have been proposed to overcome the limited efficacy and adverse reactions of bulking agents. However, most have significant limitations, including painful procurement, requirement for anesthesia, donor site infection and a frequently low cell yield. Recently, human amniotic fluid stem cells (hAFSCs) have been proposed as an ideal cell therapy source. In this study, we investigated whether periurethral injection of hAFSCs can restore urethral sphincter competency in a mouse model.
Amniotic fluids were collected and harvested cells were analyzed for stem cell characteristics and in vitro myogenic differentiation potency. Mice underwent bilateral pudendal nerve transection to generate a stress urinary incontinence (SUI) model and received either periurethral injection of hAFSCs, periurethral injection of Plasma-Lyte (control group), or underwent a sham (normal control group).For in vivo cell tracking, cells were labeled with silica-coated magnetic nanoparticles containing rhodamine B isothiocyanate (MNPs@SiO2 (RITC)) and were injected into the urethral sphincter region (n = 9). Signals were detected by optical imaging. Leak point pressure and closing pressure were recorded serially after injection.Tumorigenicity of hAFSCs was evaluated by implanting hAFSCs into the subcapsular space of the kidney, followed two weeks later by retrieval and histologic analysis.
Flow activated cell sorting showed that hAFSCs expressed mesenchymal stem cell (MSC) markers, but no hematopoietic stem cell markers. Induction of myogenic differentiation in the hAFSCs resulted in expression of PAX7 and MYOD at Day 3, and DYSTROPHIN at Day 7. The nanoparticle-labeled hAFSCs could be tracked in vivo with optical imaging for up to 10 days after injection. Four weeks after injection, the mean LPP and CP were significantly increased in the hAFSC-injected group compared with the control group. Nerve regeneration and neuromuscular junction formation of injected hAFSCs in vivo was confirmed with expression of neuronal markers and acetylcholine receptor. Injection of hAFSCs caused no in vivo host CD8 lymphocyte aggregation or tumor formation.
hAFSCs displayed MSC characteristics and could differentiate into cells of myogenic lineage. Periurethral injection of hAFSCs into an SUI animal model restored the urethral sphincter to apparently normal histology and function, in absence of immunogenicity and tumorigenicity.
人们提出干细胞注射疗法来克服填充剂疗效有限和不良反应的问题。然而，大多数方法都存在显著的局限性，包括采集过程痛苦、需要麻醉、供体部位感染以及细胞产量通常较低。最近，人羊膜间充质干细胞（hAFSCs）被提议作为一种理想的细胞治疗来源。在这项研究中，我们研究了经尿道周围注射 hAFSCs 是否可以恢复小鼠模型中的尿道括约肌功能。
收集羊水并收获细胞，分析其干细胞特征和体外成肌分化潜能。小鼠行双侧阴部神经切断术以建立压力性尿失禁（SUI）模型，并接受经尿道周围注射 hAFSCs、经尿道周围注射 Plasma-Lyte（对照组）或假手术（正常对照组）。为了进行体内细胞追踪，将细胞用含有罗丹明 B 异硫氰酸酯（MNPs@SiO2（RITC））的硅涂层磁性纳米颗粒标记，并注入尿道括约肌区域（n = 9）。通过光学成像检测信号。注射后连续记录漏点压和闭合压。通过将 hAFSCs 植入肾脏被膜下腔，两周后取出并进行组织学分析，评估 hAFSCs 的致瘤性。
流式细胞分选显示，hAFSCs 表达间充质干细胞（MSC）标志物，但不表达造血干细胞标志物。hAFSCs 诱导成肌分化，第 3 天表达 PAX7 和 MYOD，第 7 天表达 DYSTROPHIN。用光学成像可在注射后长达 10 天内追踪到纳米颗粒标记的 hAFSCs。注射后 4 周，hAFSC 注射组的平均 LPP 和 CP 明显高于对照组。体内注射 hAFSCs 后，神经元标志物和乙酰胆碱受体的表达证实了神经再生和神经肌肉接头的形成。hAFSCs 的注射未引起体内宿主 CD8 淋巴细胞聚集或肿瘤形成。
hAFSCs 具有 MSC 特征，并可分化为肌源性谱系的细胞。在 SUI 动物模型中，经尿道周围注射 hAFSCs 可使尿道括约肌恢复正常的组织学和功能，且无免疫原性和致瘤性。
