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Tian Yanpeng,Liu Yibin,Xiao Yanlai,Li Zhongkang,Zhang Mingle,Chen Liang,Li Zhen,Zhang Wangchao,Zhang Zhiqiang,Kong Desheng,Meng Li,Du Yanfang,Zhang Jingkun,Gao Jingui,Huang Xianghua 한국조직공학과 재생의학회 2024 조직공학과 재생의학 Vol.21 No.2
Background: Mayer-Rokitansky-Küster-Hauser (MRKH) syndrome is a severe congenital disorder characterized by vaginal hypoplasia caused by dysplasia of the Müllerian duct. Patients with MRKH syndrome often require nonsurgical or surgical treatment to achieve satisfactory vaginal length and sexual outcomes. The extracellular matrix has been successfully used for vaginal reconstruction. Methods: In this study, we developed a new biological material derived from porcine vagina (acellular vaginal matrix, AVM) to reconstruct the vagina in Bama miniature pigs. The histological characteristics and efficacy of acellularization of AVM were evaluated, and AVM was subsequently transplanted into Bama miniature pigs to reconstruct the vaginas. Results: Macroscopic analysis showed that the neovaginas functioned well in all Bama miniature pigs with AVM implants. Histological analysis and electrophysiological evidence indicated that morphological and functional recovery was restored in normal vaginal tissues. Scanning electron microscopy showed that the neovaginas had mucosal folds characteristics of normal vagina. No significant differences were observed in the expression of CK14, HSP47, and α-actin between the neovaginas and normal vaginal tissues. However, the expression of estrogen receptor (ER) was significantly lower in the neovaginas than in normal vaginal tissues. In addition, AVM promoted the expression of β-catenin, c-Myc, and cyclin D1. These results suggest that AVM might promotes vaginal regeneration by activating the β-catenin/c-Myc/cyclin D1 pathway. Conclusion: This study reveals that porcine-derived AVM has potential application for vaginal regeneration. Background: Mayer-Rokitansky-Küster-Hauser (MRKH) syndrome is a severe congenital disorder characterized by vaginal hypoplasia caused by dysplasia of the Müllerian duct. Patients with MRKH syndrome often require nonsurgical or surgical treatment to achieve satisfactory vaginal length and sexual outcomes. The extracellular matrix has been successfully used for vaginal reconstruction. Methods: In this study, we developed a new biological material derived from porcine vagina (acellular vaginal matrix, AVM) to reconstruct the vagina in Bama miniature pigs. The histological characteristics and efficacy of acellularization of AVM were evaluated, and AVM was subsequently transplanted into Bama miniature pigs to reconstruct the vaginas. Results: Macroscopic analysis showed that the neovaginas functioned well in all Bama miniature pigs with AVM implants. Histological analysis and electrophysiological evidence indicated that morphological and functional recovery was restored in normal vaginal tissues. Scanning electron microscopy showed that the neovaginas had mucosal folds characteristics of normal vagina. No significant differences were observed in the expression of CK14, HSP47, and α-actin between the neovaginas and normal vaginal tissues. However, the expression of estrogen receptor (ER) was significantly lower in the neovaginas than in normal vaginal tissues. In addition, AVM promoted the expression of β-catenin, c-Myc, and cyclin D1. These results suggest that AVM might promotes vaginal regeneration by activating the β-catenin/c-Myc/cyclin D1 pathway. Conclusion: This study reveals that porcine-derived AVM has potential application for vaginal regeneration.
( Ning Yang ),( Zhenmin Ling ),( Liang Peng ),( Yanlai Liu ),( Pu Liu ),( Kai Zhang ),( Aman ),( Juanjuan Shi ),( Xiangkai Li ) 한국미생물생명공학회(구 한국산업미생물학회) 2018 Journal of microbiology and biotechnology Vol.28 No.9
Fusarium oxysporum trypsin (FOT) is a fungal serine protease similar to mammal trypsin. The FOT could be successfully expressed in Pichiapastoris by engineering the natural propeptide APQEIPN. In this study, we constructed two recombinant enzymes with engineered amino acid sequences added to the N-terminus of FOT and expressed in P. pastoris. The N-terminal residues had various effects on the structural and functional properties of trypsin. The FOT, and the recombinants TE (with peptide YVEF) and TS (with peptide YV) displayed the same optimum temperature (40°C) and pH (8.0). However, the combinants TE and TS showed significantly increased thermal stability at 40°C and 50°C. Moreover, the combinants TE and TS also showed enhanced tolerance of alkaline pH conditions. Compared with those of wildtype FOT, the intramolecular hydrogen bonds and the cation π-interactions of the recombinants TE and TS were significantly increased. The recombinants TE and TS also had significantly increased catalytic efficiencies (referring to the specificity constant, k<sub>cat</sub>/K<sub>m</sub>), 1.75- fold and 1.23-fold than wild-type FOT. In silico modeling analysis uncovered that the introduction of the peptides YVEF and YV resulted in shorter distances between the substrate binding pocket (D174, G198, and G208) and catalytic triad (His42, Asp102, and Ser180), which would improve the electron transfer rate and catalytic efficiency. In addition, N-terminal residues modification described here may be a useful approach for improving the catalytic efficiencies and characteristics of other target enzymes.