神経堤細胞を介して誘導したヒトiPS細胞由来間葉系幹細胞を用いたラット喉頭軟骨再生

京都大学新制・課程博士博士(医学)甲第23417号医博第4762号新制||医||1052(附属図書館)京都大学大学院医学研究科医学専攻(主査)教授 松田 秀一特定拠点, 教授 妻木 範行, 教授 安達 泰治学位規則第4条第1項該当Doctor of Medical ScienceKyoto UniversityDFA

In vivo regeneration of rat laryngeal cartilage with mesenchymal stem cells derived from human induced pluripotent stem cells via neural crest cells

The laryngotracheal cartilage is a cardinal framework for the maintenance of the airway for breathing, which occasionally requires reconstruction. Because hyaline cartilage has a poor intrinsic regenerative ability, various regenerative approaches have been attempted to regenerate laryngotracheal cartilage. The use of autologous mesenchymal stem cells (MSCs) for cartilage regeneration has been widely investigated. However, long-term culture may limit proliferative capacity. Human-induced pluripotent stem cell-derived MSCs (iMSCs) can circumvent this problem due to their unlimited proliferative capacity. This study aimed to investigate the efficacy of iMSCs in the regeneration of thyroid cartilage in immunodeficient rats. Herein, we induced iMSCs through neural crest cell intermediates. For the relevance to prospective future clinical application, induction was conducted under xeno-free/serum-free conditions. Then, clumps fabricated from an iMSC/extracellular matrix complex (C-iMSC) were transplanted into thyroid cartilage defects in immunodeficient rats. Histological examinations revealed cartilage-like regenerated tissue and human nuclear antigen (HNA)-positive surviving transplanted cells in the regenerated lesion. HNA-positive cells co-expressed SOX9, and type II collagen was identified around HNA-positive cells. These results indicated that the transplanted C-iMSCs promoted thyroid cartilage regeneration and some of the iMSCs differentiated into chondrogenic lineage cells. Induced MSCs may be a promising candidate cell therapy for human laryngotracheal reconstruction

Laryngeal Cartilage Regeneration of Nude Rats by Transplantation of Mesenchymal Stem Cells Derived from Human-Induced Pluripotent Stem Cells

Previous studies transplanted human-induced pluripotent stem cells (hiPSCs)-derived mesenchymal stem cells (iMSCs) into thyroid cartilage defect of X-liked severe combined immunodeficiency (X-SCID) rats and confirmed transplanted cell survival and cartilage regeneration. Thus, this study aimed to investigate the contribution of iMSC transplantation to thyroid cartilage regeneration of nude rats. iMSCs were induced from hiPSCs via a neural crest cell lineage. Then, clumps formed from an iMSC/extracellular matrix complex were transplanted into thyroid cartilage defects in nude rats. The larynx was removed and histological and immunohistochemical analyses were performed 4 or 8 weeks after the transplantation. Human nuclear antigen (HNA)-positive cells were observed in 11 of 12 (91.7%) rats, which indicated that transplanted iMSCs survived in thyroid cartilage defects in nude rats. HNA-positive cells co-expressed SOX9, and type II collagen was identified around HNA-positive cells in 8 of 12 rats (66.7%), which indicated cartilage-like regeneration. Cartilage-like regeneration in nude rats in this study was comparable to the previous report on X-SCID rats (HNA-positive cells were observed in all 14 rats and cartilage-like regeneration was observed in 10 of 14 rats). This result suggests that nude rats could be an alternative to X-SCID rats in thyroid cartilage regeneration experiments using iMSCs, and this nude rat cartilage transplantation model may develop cartilage regeneration research concerning fewer problems such as infection due to immunosuppression

Peroxisome Proliferator-Activated Receptor-γ Agonist Attenuates Vocal Fold Fibrosis in Rats via Regulation of Macrophage Activation

Macrophages aid in wound healing by changing their phenotype and can be a key driver of fibrosis. However, the contribution of macrophage phenotype to fibrosis following vocal fold injury remains unclear. Peroxisome proliferator-activated receptor-γ (PPARγ) is expressed mainly by macrophages during early wound healing and regulates the macrophage phenotype. This study aimed to evaluate the effects of pioglitazone, a PPARγ agonist, on the macrophage phenotype and fibrosis following vocal fold injury in rats. Pioglitazone was injected into the rats' vocal folds on days 1, 3, 5, and 7 after injury, and the vocal fold lamina propria was evaluated on days 4 and 56 after injury. Moreover, THP-1-derived macrophages were treated with pioglitazone, and the expression of pro-inflammatory cytokines under lipopolysaccharide/interferon-γ stimulation was analyzed. The results revealed that pioglitazone reduced the expression of Ccl2 both in vivo and in vitro. Furthermore, pioglitazone decreased the density of inducible nitric oxide synthase+ CD68+ macrophages and inhibited the expression of fibrosis-related factors on day 4 after injury. On day 56 after injury, pioglitazone inhibited fibrosis, tissue contracture, and hyaluronic acid loss in a PPARγ-dependent manner. These results indicate that PPARγ activation could inhibit accumulation of inflammatory macrophages and improve tissue repair. Considered together, these findings imply that inflammatory macrophages play a key role in vocal fold fibrosis

Expression of FOXI1 and POU2F3 varies among different salivary gland neoplasms and is higher in Warthin tumor

Abstract Purpose Salivary gland tumors are histologically diverse. Ionocytes and tuft cells, rare epithelial cells found in normal salivary glands, might be associated with salivary tumors. Here, we explored the expression of FOXI1 and POU2F3, master regulators of ionocytes and tuft cells, respectively, for common salivary neoplasms using immunohistochemistry. Methods We analyzed normal salivary tissues and nine salivary gland tumors; Warthin tumors (WT), pleomorphic adenomas (PA), basal cell adenomas, and oncocytomas were benign, whereas mucoepidermoid, adenoid cystic, acinic cell, salivary duct carcinomas, and polymorphous adenocarcinomas were malignant. Results Normal salivary glands contained a few FOXI1- and POU2F3-positive cells in the ducts instead of the acini, consistent with ionocytes and tuft cells, respectively. Among the benign tumors, only WTs and PAs consistently expressed FOXI1 (10/10 and 9/10, respectively). The median H-score of WTs was significantly higher than that of PAs (17.5 vs. 4, P = 0.01). While WTs and PAs harbored POU2F3-positive cells (10/10 and 9/10, respectively), the median H-score was higher in WTs than in PAs (10.5 vs 4, respectively). Furthermore, WTs exhibited a unique staining pattern of FOXI1- and POU2F3-positive cells, which were present in luminal and abluminal locations, respectively. Whereas none of the malignant tumors expressed FOXI1, only adenoid cystic carcinoma consistently expressed POU2F3 (5/5), with a median H-score of 4. Conclusion The expression patterns of the characteristic transcription factors found in ionocytes and tuft cells vary among salivary gland tumor types and are higher in WT, which might be relevant for understanding and diagnosing salivary gland neoplasms

sj-tiff-3-cll-10.1177_09636897241228026 – Supplemental material for Changes in the Proportion of Each Cell Type After hiPSC-Derived Airway Epithelia Transplantation

Supplemental material, sj-tiff-3-cll-10.1177_09636897241228026 for Changes in the Proportion of Each Cell Type After hiPSC-Derived Airway Epithelia Transplantation by Masayuki Kitano, Yasuyuki Hayashi, Hiroe Ohnishi, Hideaki Okuyama, Masayoshi Yoshimatsu, Keisuke Mizuno, Fumihiko Kuwata, Takeshi Tada, Yo Kishimoto, Satoshi Morita and Koichi Omori in Cell Transplantation</p