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1.
J Chin Med Assoc ; 87(3): 261-266, 2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38305450

RESUMO

BACKGROUND: Leber hereditary optic neuropathy (LHON) is mainly the degeneration of retinal ganglion cells (RGCs) associated with high apoptosis and reactive oxygen species (ROS) levels, which is accepted to be caused by the mutations in the subunits of complex I of the mitochondrial electron transport chain. The treatment is still infant while efforts of correcting genes or using antioxidants do not bring good and consistent results. Unaffected carrier carries LHON mutation but shows normal phenotype, suggesting that the disease's pathogenesis is complex, in which secondary factors exist and cooperate with the primary complex I dysfunction. METHODS: Using LHON patient-specific induced pluripotent stem cells (iPSCs) as the in vitro disease model, we previously demonstrated that circRNA_0087207 had the most significantly higher expression level in the LHON patient-iPSC-derived RGCs compared with the unaffected carrier-iPSC-derived RGCs. To elaborate the underlying pathologies regulated by circRNA_008720 mechanistically, bioinformatics analysis was conducted and elucidated that circRNA_0087207 could act as a sponge of miR-548c-3p and modulate PLSCR1/TGFB2 levels in ND4 mutation-carrying LHON patient-iPSC-derived RGCs. RESULTS: Using LHON iPSC-derived RGCs as the disease-based platform, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis on targeted mRNA of miR-548c-3p showed the connection with apoptosis, suggesting downregulation of miR548c-3p contributes to the apoptosis of LHON patient RGCs. CONCLUSION: We showed that the downregulation of miR548c-3p plays a critical role in modulating cellular dysfunction and the apoptotic program of RGCs in LHON.


Assuntos
MicroRNAs , Atrofia Óptica Hereditária de Leber , Humanos , Atrofia Óptica Hereditária de Leber/genética , Atrofia Óptica Hereditária de Leber/patologia , RNA Circular/genética , Mitocôndrias , Apoptose , Mutação , MicroRNAs/genética , MicroRNAs/metabolismo , Fator de Crescimento Transformador beta2/genética , Fator de Crescimento Transformador beta2/metabolismo
2.
J Chin Med Assoc ; 87(2): 163-170, 2024 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-38132887

RESUMO

BACKGROUND: The potential of induced pluripotent stem cells (iPSCs) in revolutionizing regenerative medicine cannot be overstated. iPSCs offer a profound opportunity for therapies involving cell replacement, disease modeling, and cell transplantation. However, the widespread application of iPSC cellular therapy faces hurdles, including the imperative to regulate iPSC differentiation rigorously and the inherent genetic disparities among individuals. To address these challenges, the concept of iPSC super donors emerges, holding exceptional genetic attributes and advantageous traits. These super donors serve as a wellspring of standardized, high-quality cell sources, mitigating inter-individual variations and augmenting the efficacy of therapy. METHODS: In pursuit of this goal, our study embarked on the establishment of iPSC cell lines specifically sourced from donors possessing the HLA type (A33:03-B58:01-DRB1*03:01). The reprogramming process was meticulously executed, resulting in the successful generation of iPSC lines from these carefully selected donors. Subsequently, an extensive characterization was conducted to comprehensively understand the features and attributes of these iPSC lines. RESULTS: The outcomes of our research were highly promising. The reprogramming efforts culminated in the generation of iPSC lines from donors with the specified HLA type. These iPSC lines displayed a range of distinctive characteristics that were thoroughly examined and documented. This successful generation of iPSC lines from super donors possessing advantageous genetic traits represents a significant stride towards the realization of their potential in therapeutic applications. CONCLUSION: In summary, our study marks a crucial milestone in the realm of regenerative medicine. The establishment of iPSC lines from super donors with specific HLA types signifies a paradigm shift in addressing challenges related to iPSC cellular therapy. The standardized and high-quality cell sources derived from these super donors hold immense potential for various therapeutic applications. As we move forward, these findings provide a solid foundation for further research and development, ultimately propelling the field of regenerative medicine toward new horizons of efficacy and accessibility.


Assuntos
Células-Tronco Pluripotentes Induzidas , Humanos , Reprogramação Celular , Diferenciação Celular , Terapia Baseada em Transplante de Células e Tecidos
3.
J Adv Res ; 2023 Aug 07.
Artigo em Inglês | MEDLINE | ID: mdl-37557954

RESUMO

BACKGROUND: Modifications of lipid metabolism were closely associated with the manifestations and prognosis of coronavirus disease of 2019 (COVID-19). Pre-existing metabolic conditions exacerbated the severity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection while modulations of aberrant lipid metabolisms alleviated the manifestations. To elucidate the underlying mechanisms, an experimental platform that reproduces human respiratory physiology is required. METHODS: Here we generated induced pluripotent stem cell-derived airway organoids (iPSC-AOs) that resemble the human native airway. Single-cell sequencing (ScRNAseq) and microscopic examination verified the cellular heterogeneity and microstructures of iPSC-AOs, respectively. We subjected iPSC-AOs to SARS-CoV-2 infection and investigated the treatment effect of lipid modifiers statin drugs on viral pathogenesis, gene expression, and the intracellular trafficking of the SARS-CoV-2 entry receptor angiotensin-converting enzyme-2 (ACE-2). RESULTS: In SARS-CoV-2-infected iPSC-AOs, immunofluorescence staining detected the SARS-CoV-2 spike (S) and nucleocapsid (N) proteins and bioinformatics analysis further showed the aberrant enrichment of lipid-associated pathways. In addition, SARS-CoV-2 hijacked the host RNA replication machinery and generated the new isoforms of a high-density lipoprotein constituent apolipoprotein A1 (APOA1) and the virus-scavenging protein deleted in malignant brain tumors 1 (DMBT1). Manipulating lipid homeostasis using cholesterol-lowering drugs (e.g. Statins) relocated the viral entry receptor angiotensin-converting enzyme-2 (ACE-2) and decreased N protein expression, leading to the reduction of SARS-CoV-2 entry and replication. The same lipid modifications suppressed the entry of luciferase-expressing SARS-CoV-2 pseudoviruses containing the S proteins derived from different SARS-CoV-2 variants, i.e. wild-type, alpha, delta, and omicron. CONCLUSIONS: Together, our data demonstrated that modifications of lipid pathways restrict SARS-CoV-2 propagation in the iPSC-AOs, which the inhibition is speculated through the translocation of ACE2 from the cell membrane to the cytosol. Considering the highly frequent mutation and generation of SARS-CoV-2 variants, targeting host metabolisms of cholesterol or other lipids may represent an alternative approach against SARS-CoV-2 infection.

4.
Oxid Med Cell Longev ; 2023: 8753309, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36644580

RESUMO

Radiotherapy (RT) is currently only used in children with high-risk neuroblastoma (NB) due to concerns of long-term side effects as well as lack of effective adjuvant. Calreticulin (CALR) has served distinct physiological roles in cancer malignancies; nonetheless, impact of radiation on chaperones and molecular roles they play remains largely unknown. In present study, we systemically analyzed correlation between CALR and NB cells of different malignancies to investigate potential role of CALR in mediating radioresistance of NB. Our data revealed that more malignant NB cells are correlated to lower CALR expression, greater radioresistance, and elevated stemness as indicated by colony- and neurospheroid-forming abilities and vice versa. Of note, manipulating CALR expression in NB cells of varying endogenous CALR expression manifested changes in not only stemness but also radioresistant properties of those NB cells. Further, CALR overexpression resulted in greatly enhanced ROS and led to increased secretion of proinflammatory cytokines. Importantly, growth of NB tumors was significantly hampered by CALR overexpression and was synergistically ablated when RT was also administered. Collectively, our current study unraveled a new notion of utilizing CALR expression in malignant NB to diminish cancer stemness and mitigate radioresistance to achieve favorable therapeutic outcome for NB.


Assuntos
Calreticulina , Neuroblastoma , Criança , Humanos , Adjuvantes Imunológicos , Calreticulina/genética , Calreticulina/metabolismo , Linhagem Celular Tumoral , Neuroblastoma/patologia , Neuroblastoma/radioterapia , Tolerância a Radiação
5.
J Pharm Biomed Anal ; 219: 114877, 2022 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-35717702

RESUMO

With the limited sample volume, the droplet-based microfluidic becomes attractive in biomedical diagnosis, especially for measuring multiple analytes. Usually, for multiplexing by parallel processing, a larger sample volume is required. In our previous study, simultaneously detecting two analytes from a single droplet was first achieved by measuring different fluorescence wavelengths for different analytes. However, the number of detectable analytes could be limited by the spectral resolution of fluorescence. Here a different approach is proposed for multiplexing by sharing a single droplet in multiple sub-assays. Therefore, only a single-type reporter, i.e., the fluorescence with the same wavelength, is needed for detection of different analytes from a single sample droplet, called single-type reporter multiplexing (STRM). The standard curves of two analytes, human IL-1ß and human TNF-α, are demonstrated. The required sample volume for one measurement is only 520 nL; the total duration of the on-chip process is less than 50 min. The limits of detection (LOD) of human IL-1ß and human TNF-α are about 1.14 and 0.97 pg/mL, respectively. It is shown that the proposed bead-based digital microfluidic immunoassay can achieve multiple analytes detection with low LOD from a single sample droplet using the single-type reporter, which has never been achieved before.


Assuntos
Microfluídica , Fator de Necrose Tumoral alfa , Humanos , Imunoensaio , Limite de Detecção
6.
Biosens Bioelectron ; 150: 111851, 2020 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-31740257

RESUMO

The measurement of growth factors released in a culture medium is considered to be an attractive non-invasive approach, apart from the embryo morphology, to identify the condition of an embryo development after fertilization in vitro (IVF), but the available embryo culture medium in the current method is only a few microlitres. This small sample volume, also of small concentration, makes difficult the application of a conventional detection method, such as an enzyme-linked immunosorbent assay (ELISA). A reliable detection of the growth factor from each embryo culture medium of such a small concentration hence remains a challenge. Here for the first time we report the results of measurement of not just one, but two, growth factors, human IL-1ß and human TNF-α, from an individual droplet of embryo culture medium with a bead-based digital microfluidic chip. The required sample volume for a single measurement is only 520 nL; the total duration of the on-chip process is less than 40 min. Using the culture media of human embryos with normal morphologic features, we found that the concentrations of TNF-α change little from day 3 to day 5-6, but the concentrations of IL-1ß for some embryos might double from day 3 to day 5-6. For other embryos even with similar normal morphologic features, some growth factors, such as IL-1ß, might exhibit different expressions during the culture period. Those growth factors could serve to distinguish the development conditions of each embryo, not merely from an observation of embryo morphology.


Assuntos
Técnicas Biossensoriais , Interleucina-1beta/isolamento & purificação , Microfluídica , Fator de Necrose Tumoral alfa/isolamento & purificação , Meios de Cultura/química , Feminino , Humanos , Interleucina-1beta/genética , Análise de Sequência com Séries de Oligonucleotídeos , Fator de Necrose Tumoral alfa/genética
7.
J Healthc Eng ; 2017: 3619403, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29065588

RESUMO

Interlocking nailing is a common surgical operation to stabilize fractures in long bones. One of the difficult parts of the surgery is how to locate the position and direction of a screw hole on the interlocking nail, which is invisible to the naked eye after insertion of the nail into the medullary canal. Here, we propose a novel two-stage targeting process using two passive magnetic devices to locate the position and direction of the screw hole without radiation for the locking screw procedure. This involves a ring-shape positioning magnet inside the nail to generate a magnetic field for targeting. From the accuracy test results of these two-stage targeting devices, the search region can be identified in less than 20 seconds by the 1st-stage targeting device, while the total targeting time to locate the drilling position and direction takes less than 4 minutes, with 100% successful rate in 50 attempts. The drilling test further combines the two-stage targeting process and drilling process on the swine tibia, and it is shown that a 100% successful rate is achieved in all 10 attempts, where the total time needed is less than 5 minutes.


Assuntos
Pinos Ortopédicos , Fixação Intramedular de Fraturas/instrumentação , Magnetismo , Fraturas da Tíbia/cirurgia , Animais , Humanos , Suínos
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