雌激素缺乏对大鼠根尖牙乳头干细胞增殖及牙向/骨向分化能力的影响

口腔生物医学 ›› 2023, Vol. ›› Issue (3): 156-161.

雌激素缺乏对大鼠根尖牙乳头干细胞增殖及牙向/骨向分化能力的影响

李瑶¹,曹灵²,³,夏涵⁴,周俊波⁴,⁵

1. 1. 南京市中西医结合医院口腔科；2. 南京医科大学江苏省口腔疾病研究重点实验室
2. 南京医科大学附属口腔医院儿童口腔预防科
3. 南京医科大学附属口腔医院儿童牙病科
4. 南京市中西医结合医院口腔科
5. 南京医科大学口腔医学院

收稿日期:2023-01-17 修回日期:2023-04-17 出版日期:2023-09-25 发布日期:2023-09-21
通讯作者: 周俊波 E-mail:syusukeyao@163.com
基金资助:
国家自然科学基金资助项目;国家自然科学基金资助项目

Effects of estrogen deficiency on the proliferation and odonto/osteogenic differentiation of rat stem cells from the apical papilla

1. 1. Department of Stomatology, Nanjing Integrated Traditional Chinese and Western Medicine Hospital, Nanjing 210014, China; 2. Jiangsu Province Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing 210029, China
2. Department of Pediatric and Preventive Dentistry, The Affiliated Stomatological Hospital of Nanjing Medical University
3.
4. Department of Stomatology, Nanjing Integrated Traditional Chinese and Western Medicine Hospital

Received:2023-01-17 Revised:2023-04-17 Online:2023-09-25 Published:2023-09-21

摘要/Abstract

摘要： 目的：探讨雌激素缺乏对大鼠根尖牙乳头干细胞（stem cells from the apical papilla，SCAPs），增殖及牙向/骨向分化能力的影响。方法：将10只8周龄雌性SD大鼠随机分为假手术组（Sham）和卵巢去势组（OVX），每组各5只。全麻下OVX组摘除双侧卵巢，Sham组行相同切口，保留双侧卵巢。1个月后处死大鼠，分离切牙根尖牙乳头组织，酶消化法分离培养两组SCAPs，采用甲基噻唑基四唑（methyl thiazolyl tetrazolium，MTT）法、碱性磷酸酶（alkaline phosphotase，ALP）活性检测、蛋白质印迹法（Western blot）、实时定量反转录聚合酶链反应(reverse transcription PCR，RT-PCR)检测雌激素缺乏对大鼠SCAPs增殖及成牙/成骨向分化能力的影响。将两组细胞分别以明胶海绵为载体植入雌性SD大鼠肾被膜下，8周后取出植入组织，HE(hematoxylin and eosin)染色观察其矿化能力。结果；MTT结果显示OVX组SCAPs增殖能力显著下降，差异具有统计学意义（P<0.01）。ALP活性检测结果显示OVX组SCAPs的ALP活性低于Sham组，差异具有统计学意义（P<0.01）。RT-PCR和Western blot结果均表明OVX组SCAPs的成牙/成骨相关指标表达低于Sham组，差异具有统计学意义（P<0.01）。体内移植结果显示Sham组形成较规则的牙髓-牙本质样复合体，而OVX组形成不规则结构。结论：雌激素缺乏降低了大鼠SCAPs的增殖及牙向/骨向分化能力。

关键词: 雌激素, 根尖牙乳头干细胞, 增殖, 牙向/骨向分化

Abstract: Objective：To evaluate the effects of estrogen deficiency on the proliferation and odonto/osteogenic differentiation of rat stem cells from the apical papilla(SCAPs). Methods: Ten 8-week-old female SD rats were randomly divided into two groups, sham-operation group(Sham) and ovariectomized group(OVX), 5 rats in each group. All rats were anesthetized and both ovaries of OVX-rats were gently removed. Sham-operation rats were treated with the same incisions to expose the ovaries but without any hurt to them. One month after the operation, the apical papillae of rat incisors were gently separated and SCAPs were isolated and incubated. Methyl thiazolyl tetrazolium(MTT) assay, alkaline phosphotase(ALP) activity, real-time reverse transcription（RT-PCR）and western blot assay were respectively used to determine the proliferation and odonto/osteogenic differentiation potential of rat SCAPs. Cell pellets of OVX-SCAPs and Sham-SCAPs were respectively transplanted into the renal capsules of immunodeficient female rats. After in vivo culture for 8 weeks, the retrieved implants were processed for hematoxylin and eosin (H & E) staining. Results: MTT results showed that OVX-SCAPs exhibited the decreased proliferative activity as compared with Sham- SCAPs. ALP activity of OVX- SCAPs was lower than that of Sham- SCAPs (P<0.01). Real-time RT-PCR and Western blot results showed that the expression of odonto/osteogenic markers (Dspp/DSP, Osx/OSX, Ocn/OCN) in OVX- SCAPs was significantly inhibited as compared with Sham- SCAPs (P<0.01). Sham-SCAPs implants showed the typical dentin-pulp like complex, while the retrieved implants of OVX-SCAPs exhibited unregular tissue. Conclusions: Estrogen deficiency significantly inhibits the proliferation and odonto/osteogenetic potential of rat SCAPs.

Key words: Estrogen, Stem cells from the apical papilla, proliferation, odonto/osteogenetic differentiation

李瑶曹灵夏涵周俊波. 雌激素缺乏对大鼠根尖牙乳头干细胞增殖及牙向/骨向分化能力的影响[J]. 口腔生物医学, 2023, (3): 156-161.

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