The organotypic retinal explant culture has been established for more than a decade and offers a range of unique advantages compared with in vivo experiments and cell cultures.However,the lack of systematic and contin...The organotypic retinal explant culture has been established for more than a decade and offers a range of unique advantages compared with in vivo experiments and cell cultures.However,the lack of systematic and continuous comparison between in vivo retinal development and the organotypic retinal explant culture makes this model controversial in postnatal retinal development studies.Thus,we aimed to verify the feasibility of using this model for postnatal retinal development studies by comparing it with the in vivo retina.In this study,we showed that postnatal retinal explants undergo normal development,and exhibit a consistent structure and timeline with retinas in vivo.Initially,we used SOX2 and PAX6 immunostaining to identify retinal progenitor cells.We then examined cell proliferation and migration by immunostaining with Ki-67 and doublecortin,respectively.Ki-67-and doublecortin-positive cells decreased in both in vivo and explants during postnatal retinogenesis,and exhibited a high degree of similarity in abundance and distribution between groups.Additionally,we used Ceh-10 homeodomain-containing homolog,glutamate-ammonia ligase(glutamine synthetase),neuronal nuclei,and ionized calcium-binding adapter molecule 1 immunostaining to examine the emergence of bipolar cells,Müller glia,mature neurons,and microglia,respectively.The timing and spatial patterns of the emergence of these cell types were remarkably consistent between in vivo and explant retinas.Our study showed that the organotypic retinal explant culture model had a high degree of consistency with the progression of in vivo early postnatal retina development.The findings confirm the accuracy and credibility of this model and support its use for long-term,systematic,and continuous observation.展开更多
We investigated the androgen receptor(AR) gene mutation profiles of Chinese patients exhibiting severe androgen insensitivity syndrome(AIS) phenotypes. The present study enrolled 28 patients with genetically diagnosed...We investigated the androgen receptor(AR) gene mutation profiles of Chinese patients exhibiting severe androgen insensitivity syndrome(AIS) phenotypes. The present study enrolled 28 patients with genetically diagnosed AIS, who presented with severe phenotypes(Prader grade 0–3). Patients and some family members were screened via amplification and sequencing of their AR exons 1–8, including the corresponding intronic flanking regions. Luteinizing(LH), follicle-stimulating(FSH), and testosterone(T) hormone levels were found to be slightly, but not significantly, higher in patients with complete androgen insensitivity syndrome(CAIS) than in patients with partial androgen insensitivity syndrome(PAIS)(P>0.05). We identified 24 different AR mutations, including 12 that were novel. Ten patients(cases 2, 3, 10, 28, 11, 12, 19, 20, 24, and 25) were found to carry five recurrent mutations(p.Y572 S, p.P914 S, p.S176 R, p.Y782 N, and p.R841H); of these, p.Y572 S, p.S176 R, and p.Y782 N were novel. Among the mutations identified in patients with CAIS, six(66.7%) were characterized as single-nucleotide missense mutations, and six(66.7%) were found to be located in the AR ligand-binding domain(LBD). Among the mutations identified in patients with PAIS, 15(93.8%) were found to be missense, and 11(68.8%) were found to be located in the LBD. Patients 10 and 28 were determined to harbor the same missense mutation(p.P914S), but were diagnosed with CAIS and PAIS, respectively.Sex hormone levels were slightly, but not significantly, elevated in patients with CAIS compared to those with PAIS. Missense mutations spanning AR exons 1–8 were the predominant form of identified mutations, and these were mostly located in the AR LBD. Approximately 50% of the identified mutations were novel, and have enriched the AR gene-mutation database. Patients harboring identical mutations were in some instances found to exhibit divergent phenotypes.展开更多
基金supported by the National Natural Science Foundation of China,Nos.81901156(to ZZ),82271200(to ZZ),82171308(to XC)the Fundamental Research Funds for the Central Universities,No.xzy012022035(to ZZ)+1 种基金the Natural Science Foundation of Shaanxi Province,Nos.2021JM-261(to QK),2023-YBSF-303(to ZZ)Traditional Chinese Medicine Project of Shaanxi Province,No.2019-ZZ-JC047(to QK)。
文摘The organotypic retinal explant culture has been established for more than a decade and offers a range of unique advantages compared with in vivo experiments and cell cultures.However,the lack of systematic and continuous comparison between in vivo retinal development and the organotypic retinal explant culture makes this model controversial in postnatal retinal development studies.Thus,we aimed to verify the feasibility of using this model for postnatal retinal development studies by comparing it with the in vivo retina.In this study,we showed that postnatal retinal explants undergo normal development,and exhibit a consistent structure and timeline with retinas in vivo.Initially,we used SOX2 and PAX6 immunostaining to identify retinal progenitor cells.We then examined cell proliferation and migration by immunostaining with Ki-67 and doublecortin,respectively.Ki-67-and doublecortin-positive cells decreased in both in vivo and explants during postnatal retinogenesis,and exhibited a high degree of similarity in abundance and distribution between groups.Additionally,we used Ceh-10 homeodomain-containing homolog,glutamate-ammonia ligase(glutamine synthetase),neuronal nuclei,and ionized calcium-binding adapter molecule 1 immunostaining to examine the emergence of bipolar cells,Müller glia,mature neurons,and microglia,respectively.The timing and spatial patterns of the emergence of these cell types were remarkably consistent between in vivo and explant retinas.Our study showed that the organotypic retinal explant culture model had a high degree of consistency with the progression of in vivo early postnatal retina development.The findings confirm the accuracy and credibility of this model and support its use for long-term,systematic,and continuous observation.
基金supported by the Public Health Project for Residents in Beijing (Z151100003915103)the National Key Research and Development Program of China (2016YFC0901505)
文摘We investigated the androgen receptor(AR) gene mutation profiles of Chinese patients exhibiting severe androgen insensitivity syndrome(AIS) phenotypes. The present study enrolled 28 patients with genetically diagnosed AIS, who presented with severe phenotypes(Prader grade 0–3). Patients and some family members were screened via amplification and sequencing of their AR exons 1–8, including the corresponding intronic flanking regions. Luteinizing(LH), follicle-stimulating(FSH), and testosterone(T) hormone levels were found to be slightly, but not significantly, higher in patients with complete androgen insensitivity syndrome(CAIS) than in patients with partial androgen insensitivity syndrome(PAIS)(P>0.05). We identified 24 different AR mutations, including 12 that were novel. Ten patients(cases 2, 3, 10, 28, 11, 12, 19, 20, 24, and 25) were found to carry five recurrent mutations(p.Y572 S, p.P914 S, p.S176 R, p.Y782 N, and p.R841H); of these, p.Y572 S, p.S176 R, and p.Y782 N were novel. Among the mutations identified in patients with CAIS, six(66.7%) were characterized as single-nucleotide missense mutations, and six(66.7%) were found to be located in the AR ligand-binding domain(LBD). Among the mutations identified in patients with PAIS, 15(93.8%) were found to be missense, and 11(68.8%) were found to be located in the LBD. Patients 10 and 28 were determined to harbor the same missense mutation(p.P914S), but were diagnosed with CAIS and PAIS, respectively.Sex hormone levels were slightly, but not significantly, elevated in patients with CAIS compared to those with PAIS. Missense mutations spanning AR exons 1–8 were the predominant form of identified mutations, and these were mostly located in the AR LBD. Approximately 50% of the identified mutations were novel, and have enriched the AR gene-mutation database. Patients harboring identical mutations were in some instances found to exhibit divergent phenotypes.