Intermittent PTHrP(1-34) Exposure Augments Chondrogenesis and Reduces Hypertrophy of Mesenchymal Stromal Cells
Intermittent PTHrP(1-34) Exposure Augments Chondrogenesis and Reduces Hypertrophy of Mesenchymal Stromal Cells
复制标题
DOI：
10.1089/scd.2014.0101
复制
发表时间：
2014-10-15
影响因子：
4
通讯作者：
Richter, Wiltrud
中科院分区：
文献类型：
--
作者：
Fischer, Jennifer;Aulmann, Antje;Richter, Wiltrud
表型不稳定和过早肥大阻碍了人间充质间质细胞（MSCs）用于软骨再生。本研究的目的是研究间歇性补充甲状旁腺激素相关蛋白（PTHrP），而不是持续治疗，是否能有益地影响MSC软骨形成，并探索分解代谢和合成代谢反应的分子机制。在高密度培养中进行软骨诱导的人MSCs在蛋白多糖、II型胶原和X沉积之前，持续或通过6小时脉冲（每周3次）接受PTHrP（1-34）、forskolin、dbcAMP或PTHrP(7-34)；基因表达;测定碱性磷酸酶（ALP）活性。虽然持续应用PTHrP（1-34）抑制MSCs的软骨形成，但脉冲应用在6周后显著增加了2型胶原（COL2A1）基因表达以及II型胶原、蛋白多糖和DNA含量。脉冲PTHrP对10型胶原蛋白（COL10A1）基因表达影响不大，但对印度刺猬蛋白（IHH）表达和ALP活性有显著下调。PTHrP暴露对ALP活性的反应比COL2A1调节更快，这表明信号持续时间对分解代谢反应和合成代谢反应至关重要。通过forskolin刺激环腺苷单磷酸(cAMP)/蛋白激酶A （PKA）信号通路可以复制两种治疗模式的主要效果，而应用能够蛋白激酶C （PKC）信号通路的PTHrP（7-34）无效。脉冲PTHrP暴露于MSCs刺激软骨形成并减少软骨内分化，明显地将软骨基质沉积与肥厚标记物表达脱钩。cAMP/PKA是触发两种治疗方式相反作用的主要信号通路。间歇应用PTHrP是改善MSCs软骨形成的重要新手段，可视为MSCs软骨缺损再生的一种支持性临床治疗模式。
Phenotype instability and premature hypertrophy prevent the use of human mesenchymal stromal cells (MSCs) for cartilage regeneration. Aim of this study was to investigate whether intermittent supplementation of parathyroid hormone-related protein (PTHrP), as opposed to constant treatment, can beneficially influence MSC chondrogenesis and to explore molecular mechanisms below catabolic and anabolic responses. Human MSCs subjected to chondrogenic induction in high-density culture received PTHrP(1-34), forskolin, dbcAMP, or PTHrP(7-34) either constantly or via 6-h pulses (three times weekly), before proteoglycan, collagen type II, and X deposition; gene expression; and alkaline phosphatase (ALP) activity were assessed. While constant application of PTHrP(1-34) suppressed chondrogenesis of MSCs, pulsed application significantly increased collagen type 2 (COL2A1) gene expression and the collagen type II, proteoglycan, and DNA content of pellets after 6 weeks. Collagen type 10 (COL10A1) gene expression was little affected but Indian hedgehog (IHH) expression and ALP activity were significantly downregulated by pulsed PTHrP. A faster response to PTHrP exposure was recorded for ALP activity over COL2A1 regulation, suggesting that signal duration is critical for catabolic versus anabolic reactions. Stimulation of cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) signaling by forskolin reproduced major effects of both treatment modes, whereas application of PTHrP(7-34) capable of protein kinase C (PKC) signaling was ineffective. Pulsed PTHrP exposure of MSCs stimulated chondrogenesis and reduced endochondral differentiation apparently uncoupling chondrogenic matrix deposition from hypertrophic marker expression. cAMP/PKA was the major signaling pathway triggering the opposing effects of both treatment modes. Intermittent application of PTHrP represents an important novel means to improve chondrogenesis of MSCs and may be considered as a supporting clinical-treatment mode for MSC-based cartilage defect regeneration.