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Aging and age-related diseases are intricately associated with oxidative stress and inflammation. Nonsteroidal anti-inflammatory drugs (NSAIDs) have shown their promise in mitigating age-related conditions and potentially extending lifespan in various model organisms. However, the efficacy of NSAIDs in older individuals may be influenced by age-related changes in drug metabolism and tolerance, which could result in age-dependent toxicities. This study aimed to evaluate the potential risks of toxicities associated with commonly used NSAIDs (aspirin, ibuprofen, acetaminophen, and indomethacin) on lifespan, healthspan, and oxidative stress levels in both young and old Caenorhabditis elegans. The results revealed that aspirin and ibuprofen were able to extend lifespan in both young and old worms by suppressing ROS generation and enhancing the expression of antioxidant SOD genes. In contrast, acetaminophen and indomeacin accelerated aging process in old worms, leading to oxidative stress damage and reduced resistance to heat stress through the pmk-1/skn-1 pathway. Notably, the harmful effects of acetaminophen and indomeacin were mitigated when pmk-1 was knocked out in the pmk-1(km25) strain. These results underscore the potential lack of benefit from acetaminophen and indomeacin in elderly individuals due to their increased susceptibility to toxicity. Further research is essential to elucidate the underlying mechanisms driving these age-dependent responses and to evaluate the potential risks associated with NSAID use in the elderly population.
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Hepatocyte transplantation has the potential to treat acute liver failure and correct liver-based metabolic disorders. Proliferating human hepatocytes (ProliHHs) provide a large-scale source as an alternative to primary human hepatocytes. However, host rejection led to inefficient graft survival and function, which hindered the clinical application of cell therapy. Herein, we employed the lentiviral system to overexpress immunomodulatory factors programmed death-ligand 1 (cluster of differentiation 274) (CD274) and cluster of differentiation 47 (CD47) in ProliHHs. CD47+274 overexpression inhibited macrophage and T cell responses in vitro. After transplantation into mice via the spleen without immunosuppression, CD47+274 ProliHHs accumulation in the liver significantly increased for 48 hours compared with ProliHHs. Consistent with the in vitro results, CD47+274 ProliHHs were less aggregated and infiltrated by macrophages and also recruited fewer T cells in the liver. Seven days after transplantation, the human albumin level of engineered ProliHHs doubled compared with control group. CD47+274 ProliHHs further ameliorated the liver injury induced using concanavalin A. Overall, our results suggested CD47+274 overexpression reduced innate and adaptive immune responses during hepatocyte transplantation, and the survival rate and graft function of transplanted hepatocyte-like cells were all significantly improved.
Assuntos
Antígeno CD47 , Hepatopatias , Animais , Humanos , Camundongos , Antígeno B7-H1/metabolismo , Hepatócitos , Imunidade , Hepatopatias/metabolismoRESUMO
BACKGROUND AND AIMS: Hepatocyte transplantation has been demonstrated to be effective to treat liver metabolic disease and acute liver failure. Nevertheless, the shortage of donor hepatocytes restrained its application in clinics. To expand human hepatocytes at a large scale, several dedifferentiation-based protocols have been established, including proliferating human hepatocytes (ProliHH). However, the decreased transplantation efficiency of these cells after long-term expansion largely impedes their application. APPROACH AND RESULTS: We found that accompanied with dedifferentiation, long-term cultured ProliHH (lc-ProliHH) up-regulated a panel of chemokines and cytokines related to innate immunity, which were referred to as dedifferentiation-associated inflammatory factors (DAIF). DAIF elicited excessive macrophage responses, accounting for the elimination of lc-ProliHH specifically during engraftment. Two possible strategies to increase ProliHH transplantation were then characterized. Blockage of innate immune response by dexamethasone reverted the engraftment and repopulation of lc-ProliHH to a level comparable to primary hepatocytes, resulting in improved liver function and a better survival of fumarylacetoacetate hydrolase-deficient mice. Alternatively, rematuration of lc-ProliHH as organoids reduced the expression of DAIF and led to markedly improved engraftment. CONCLUSIONS: These results revealed that lc-ProliHH triggers exacerbated macrophage activation by DAIF and provided potential solutions for clinical transplantation of lc-ProliHH.
Assuntos
Hepatócitos , Fígado , Humanos , Camundongos , Animais , Hepatócitos/metabolismo , Fígado/metabolismo , Citocinas/metabolismo , Quimiocinas/metabolismo , Macrófagos/metabolismoRESUMO
Sulfur mustard (SM) is a highly toxic chemical warfare agent that causes acute lung injury (ALI) and/or acute respiratory distress syndrome (ARDS). There are no effective therapeutic treatments or antidotes available currently to counteract its toxic effects. Our previous study shows that bone marrow-derived mesenchymal stromal cells (BMSCs) could exert therapeutic effects against SM-induced lung injury. In this study, we explored the therapeutic potential of BMSC-derived exosomes (BMSC-Exs) against ALI and the underlying mechanisms. ALI was induced in mice by injection of SM (30 mg/kg, sc) at their medial and dorsal surfaces. BMSC-Exs (20 µg/kg in 200 µL PBS, iv) were injected for a 5-day period after SM exposure. We showed that BMSC-Exs administration caused a protective effect against pulmonary edema. Using a lung epithelial cell barrier model, BMSC-Exs (10, 20, 40 µg) dose-dependently inhibited SM-induced cell apoptosis and promoted the recovery of epithelial barrier function by facilitating the expression and relocalization of junction proteins (E-cadherin, claudin-1, occludin, and ZO-1). We further demonstrated that BMSC-Exs protected against apoptosis and promoted the restoration of barrier function against SM through upregulating G protein-coupled receptor family C group 5 type A (GPRC5A), a retinoic acid target gene predominately expressed in the epithelial cells of the lung. Knockdown of GPRC5A reduced the antiapoptotic and barrier regeneration abilities of BMSC-Exs and diminished their therapeutic effects in vitro and in vivo. BMSC-Exs-caused upregulation of GPRC5A promoted the expression of Bcl-2 and junction proteins via regulating the YAP pathway. In summary, BMSC-Exs treatment exerts protective effects against SM-induced ALI by promoting alveolar epithelial barrier repair and may be an alternative approach to stem cell-based therapy.
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Lesão Pulmonar Aguda/terapia , Exossomos/transplante , Células-Tronco Mesenquimais/citologia , Transdução de Sinais/efeitos dos fármacos , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/metabolismo , Animais , Apoptose/fisiologia , Linhagem Celular , Células Epiteliais/metabolismo , Técnicas de Inativação de Genes , Pulmão/metabolismo , Pulmão/patologia , Masculino , Camundongos Endogâmicos ICR , Camundongos Knockout , Gás de Mostarda , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Proteínas de Sinalização YAP/metabolismoRESUMO
BACKGROUND & AIMS: AT-rich interaction domain 1a (Arid1a), a component of the chromatin remodeling complex, has emerged as a tumor suppressor gene. It is frequently mutated in hepatocellular carcinoma (HCC). However, it remains unknown how Arid1a suppresses HCC development and whether Arid1a deficiency could be exploited for therapy, we aimed to explore these questions. METHODS: The expression of Arid1a in human and mouse HCCs was determined by immunohistochemical (IHC) staining. Gene expression was determined by quantitative PCR, ELISA or western blotting. Arid1a knockdown HCC cell lines were established by lentiviral-based shRNA. Tumor angiogenesis was quantified based on vessel density. The regulation of angiopoietin (Ang2) expression by Arid1a was identified by chromatin immunoprecipitation (ChIP) assay. The tumor promoting function of Arid1a loss was studied with a xenograft model in nude mice and diethylnitrosamine (DEN)-induced HCC in Arid1a conditional knockout mice. The therapeutic values of Ang2 antibody and sorafenib treatment were evaluated both in vitro and in vivo. RESULTS: We demonstrate that Arid1a deficiency, occurring in advanced human HCCs, is associated with increased vessel density. Mechanistically, loss of Arid1a causes aberrant histone H3K27ac deposition at the angiopoietin-2 (Ang2) enhancer and promoter, which eventually leads to ectopic expression of Ang2 and promotes HCC development. Ang2 blockade in Arid1a-deficient HCCs significantly reduces vessel density and tumor progression. Importantly, sorafenib treatment, which suppresses H3K27 acetylation and Ang2 expression, profoundly halts the progression of Arid1a-deficient HCCs. CONCLUSIONS: Arid1a-deficiency activates Ang2-dependent angiogenesis and promotes HCC progression. Loss of Arid1a in HCCs confers sensitivity to Ang2 blockade and sorafenib treatment. LAY SUMMARY: AT-rich interaction domain 1a (Arid1a), is a tumor suppressor gene. Arid1a-deficiency promotes Ang2-dependent angiogenesis leading to hepatocellular carcinoma progression. Arid1a-deficiency also sensitizes tumors to Ang2 blockade by sorafenib treatment.
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Inibidores da Angiogênese/farmacologia , Angiopoietina-2/metabolismo , Carcinoma Hepatocelular , Neoplasias Hepáticas , Neovascularização Patológica/tratamento farmacológico , Proteínas Nucleares , Sorafenibe/farmacologia , Fatores de Transcrição , Animais , Antineoplásicos/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/metabolismo , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Genes Supressores de Tumor , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Camundongos , Estadiamento de Neoplasias , Proteínas Nucleares/deficiência , Proteínas Nucleares/metabolismo , Fatores de Transcrição/deficiência , Fatores de Transcrição/metabolismoRESUMO
Borealin, a member of the chromosomal passenger complex, plays a key regulatory role at centromeres and the central spindle during mitosis. Loss of Borealin leads to defective cell proliferation and early embryonic lethality. The in vivo functions of Borealin in mammalian postnatal development, tissue homeostasis, and tumorigenesis remain elusive. We specifically analyzed the role of Borealin in regulating postnatal liver development, damage-induced liver regeneration, and liver carcinogenesis using mice carrying conditional Borealin alleles. Perinatal loss of Borealin caused increased genome ploidy and enlarged cell size in hepatocytes, likely due to the impaired function of the chromosomal passenger complex in mitosis. Borealin deletion also showed attenuated expansion of Sox9+HNF4α+ progenitor-like cells in liver regeneration during 3,5-diethoxycarbonyl-1,4-dihydrocollidine diet-induced liver injury. Moreover, ΔN90-ß-Catenin and c-Met-induced hepatocarcinogenesis development was largely impeded by Borealin deletion. These findings indicate that Borealin plays a key role in liver development, regeneration, and tumorigenesis and suggests that Borealin could be a potential target for related liver diseases.
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Proteínas de Ciclo Celular/deficiência , Proteínas Cromossômicas não Histona/deficiência , Hepatócitos/metabolismo , Neoplasias Hepáticas/metabolismo , Regeneração Hepática , Fígado/metabolismo , Células-Tronco/metabolismo , Animais , Proteínas de Ciclo Celular/metabolismo , Tamanho Celular , Proteínas Cromossômicas não Histona/metabolismo , Fator 4 Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/metabolismo , Hepatócitos/patologia , Fígado/patologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Mutantes , Ploidias , Proteínas Proto-Oncogênicas c-met/genética , Proteínas Proto-Oncogênicas c-met/metabolismo , Fatores de Transcrição SOX9/genética , Fatores de Transcrição SOX9/metabolismo , Células-Tronco/patologiaRESUMO
UNLABELLED: Hepatocellular carcinoma (HCC) is a cancer lacking effective therapies. Several measures have been proposed to treat HCCs, such as senescence induction, mitotic inhibition, and cell death promotion. However, data from other cancers suggest that single use of these approaches may not be effective. Here, by genetic targeting of Survivin, an inhibitor of apoptosis protein (IAP) that plays dual roles in mitosis and cell survival, we identified a tumor necrosis factor alpha (TNFα)-mediated synergistic lethal effect between senescence and apoptosis sensitization in malignant HCCs. Survivin deficiency results in mitosis defect-associated senescence in HCC cells, which triggers local inflammation and increased TNFα. Survivin inactivation also sensitizes HCC cells to TNFα-triggered cell death, which leads to marked HCC regression. Based on these findings, we designed a combination treatment using mitosis inhibitor and proapoptosis compounds. This treatment recapitulates the therapeutic effect of Survivin deletion and effectively eliminates HCCs, thus representing a potential strategy for HCC therapy. CONCLUSION: Survivin ablation dramatically suppresses human and mouse HCCs by triggering senescence-associated TNFα and sensitizing HCC cells to TNFα-induced cell death. Combined use of mitotic inhibitor and second mitochondrial-derived activator of caspases mimetic can induce senescence-associated TNFα and enhance TNFα-induced cell death and synergistically eliminate HCC. (Hepatology 2016;64:1105-1120).
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Carcinoma Hepatocelular/etiologia , Morte Celular , Senescência Celular , Neoplasias Hepáticas/etiologia , Mitose , Fator de Necrose Tumoral alfa/fisiologia , Animais , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/genética , Humanos , Proteínas Inibidoras de Apoptose/genética , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Masculino , Camundongos , SurvivinaRESUMO
BACKGROUND: Hypercholesterolemia arising from abnormal lipid metabolism is one of the critical risk factors for coronary artery disease (CAD), however the roles of genetic variants in lipid metabolism-related genes on premature CAD (≤ 60 years old) development still require further investigation. We herein genotyped four single nucleotide polymorphisms (SNPs) in lipid metabolism-related genes (rs1132899 and rs5167 in APOC4, rs1801693 and rs7765781 in LPA), aimed to shed light on the influence of these SNPs on individual susceptibility to early-onset CAD. METHODS: Genotyping of the four SNPs (rs1132899, rs5167, rs1801693 and rs7765781) was performed in 224 premature CAD cases and 297 control subjects (≤ 60 years old) using polymerase chain reaction-ligation detection reaction (PCR-LDR) method. The association of these SNPs with premature CAD was performed with SPSS software. RESULTS: Multivariate logistic regression analysis showed that C allele (OR = 1.50, P = 0.027) and CC genotype (OR = 2.84, P = 0.022) of APOC4 rs1132899 were associated with increased premature CAD risk, while the other three SNPs had no significant effect. Further stratified analysis uncovered a more evident association with the risk of premature CAD among male subjects (C allele, OR = 1.65, and CC genotype, OR = 3.33). CONCLUSIONS: Our data provides the first evidence that APOC4 rs1132899 polymorphism was associated with an increased risk of premature CAD in Chinese subjects, and the association was more significant among male subjects.
Assuntos
Apolipoproteínas C/genética , Povo Asiático/genética , Doença da Artéria Coronariana/genética , Etnicidade/genética , Estudos de Associação Genética , Predisposição Genética para Doença , Polimorfismo de Nucleotídeo Único/genética , Análise de Variância , Estudos de Casos e Controles , Doença da Artéria Coronariana/sangue , Feminino , Humanos , Lipídeos/sangue , Lipoproteína(a)/genética , Masculino , Pessoa de Meia-Idade , Análise Multivariada , Fatores de Risco , Fumar/efeitos adversosRESUMO
UNLABELLED: Hepatocytes possess a remarkable capacity to regenerate and reconstitute the parenchyma after liver damage. However, in the case of chronic injury, their proliferative potential is impaired and hepatic progenitor cells (HPCs) are activated, resulting in a ductular reaction known as oval cell response. Proapoptotic and survival signals maintain a precise balance to spare hepatocytes and progenitors from hyperplasia and cell death during regeneration. Survivin, a member of the family of inhibitor of apoptosis proteins (IAPs), plays key roles in the proliferation and apoptosis of various cell types. Here, we characterized the in vivo function of Survivin in regulating postnatal liver development and homeostasis using mice carrying conditional Survivin alleles. Hepatic perinatal loss of Survivin causes impaired mitosis, increased genome ploidy, and enlarged cell size in postnatal livers, which eventually leads to hepatocyte apoptosis and triggers tissue damage and inflammation. Subsequently, HPCs that retain genomic Survivin alleles are activated, which finally differentiate into hepatocytes and reconstitute the whole liver. By contrast, inducible ablation of Survivin in adult hepatocytes does not affect HPC activation and liver homeostasis during a long-life period. CONCLUSION: Perinatal Survivin deletion impairs hepatic mitosis in postnatal liver development, which induces HPC activation and reconstitution in the liver, therefore providing a novel HPC induction model.
Assuntos
Hepatócitos/citologia , Proteínas Inibidoras de Apoptose/genética , Fígado/crescimento & desenvolvimento , Proteínas Repressoras/genética , Células-Tronco/citologia , Animais , Apoptose/fisiologia , Diferenciação Celular , Fígado/patologia , Regeneração Hepática/fisiologia , Camundongos , SurvivinaRESUMO
miRNAs are small non-coding RNAs that play an important role in numerous physiological processes. Common single nucleotide polymorphisms (SNPs) in pre-miRNAs may change their property through altering miRNAs expression and/or maturation, resulting in diverse functional consequences. To date, the role of genetic variants in pre-miRNAs on coronary artery disease (CAD) risk remains poorly understood. Here we aimed to evaluate the influence of three common SNPs in pre-miRNAs (miR-146a rs2910164 G>C, miR-196a2 rs11614913 C>T, miR-499 rs3746444 T>C) on individual susceptibility to CAD in a Chinese population of 295 CAD patients and 283 controls. Genotyping was performed using polymerase chain reaction-based restriction fragment length polymorphism (PCR-RFLP) method. In a logistic regression analysis, we detected an association of rs2910164 in pre-miR-146a with the CAD risk; compared with the GG homozygotes, the GC heterozygotes [odds ratio (OR)=1.89, 95% confidence interval (CI)=1.06-3.36, P=0.029] and the CC homozygotes (OR=1.83, 95% CI=1.01-3.32, P=0.046) genotype were statistically significantly associated with the increased risk for CADs. As we used further genotype association models, we found a similar trend of the association in recessive model (OR=1.86, 95% CI=1.09-3.19, P=0.023). We also found that the genotypes of miR-146a rs2910164 were associated with its mature miRNA expression by analyzing 23 PBMC samples from CAD patients. Individuals carrying rs11614913 GC or CC genotypes showed 3.2-fold higher expression compared to GG genotype carriers (P<0.05). We observed no association of the other two SNPs in miR-196a2 (rs11614913) and miR-499 (rs3746444) with the CAD incidence. Our data provide the first evidence that the miR-146a rs2910164 polymorphism is associated with increased risk of CAD in Chinese Han population, which may be through influencing the expression levels of the miRNA.
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Doença da Artéria Coronariana/genética , MicroRNAs/genética , Idoso , Povo Asiático/genética , Estudos de Casos e Controles , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , RiscoRESUMO
BACKGROUND: Abnormal lipids is one of the critical risk factors for myocardial infarction (MI), however the role of genetic variants in lipid metabolism-related genes on MI pathogenesis still requires further investigation. We herein genotyped three SNPs (LRP6 rs2302685, LDLRAP1 rs6687605, SOAT1 rs13306731) in lipid metabolism-related genes, aimed to shed light on the influence of these SNPs on individual susceptibility to MI. METHODS: Genotyping of the three SNPs (rs2302685, rs6687605 and rs13306731) was performed in 285 MI cases and 650 control subjects using polymerase chain reaction-ligation detection reaction (PCR-LDR) method. The association of these SNPs with MI and lipid profiles was performed with SPSS software. RESULTS: Multivariate logistic regression analysis showed that C allele (OR = 1.62, P = 0.039) and the combined CT/CC genotype (OR = 1.67, P = 0.035) of LRP6 rs2302685 were associated with increased MI risk, while the other two SNPs had no significant effect. Further stratified analysis uncovered a more evident association with MI risk among younger subjects (≤60 years old). Fascinatingly, CT/CC genotype of rs2302685 conferred increased LDL-C levels compared to TT genotype (3.0 mmol/L vs 2.72 mmol/L) in younger subjects. CONCLUSIONS: Our data provides the first evidence that LRP6 rs2302685 polymorphism is associated with an increased risk of MI in Chinese subjects, and the association is more evident among younger individuals, which probably due to the elevated LDL-C levels.
Assuntos
Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Infarto do Miocárdio/genética , Idoso , Feminino , Predisposição Genética para Doença/genética , Genótipo , Humanos , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único/genéticaRESUMO
Aging, a complex biological process influenced by genetic, environmental, and pharmacological factors, presents a significant challenge in understanding its underlying mechanisms. In this study, we explored the divergent impacts of metformin treatment on the lifespan and healthspan of young and old C. elegans, demonstrating a intriguing "elixir in youth, poison in elder" phenomenon. By scrutinizing the gene expression changes in response to metformin in young (day 1 of adulthood) and old (days 8) groups, we identified nhr-57 and C46G7.1 as potential modulators of age-specific responses. Notably, nhr-57 and C46G7.1 exhibit contrasting regulation patterns, being up-regulated in young worms but down-regulated in old counterparts following metformin treatment. Functional studies employing knockdown approaches targeting nhr-57, a gene under the control of hif-1 with a documented protective function against pore-forming toxins in C. elegans, and C46G7.1, unveiled their critical roles in modulating lifespan and healthspan, as well as in mediating the biphasic effects of metformin. Furthermore, deletion of hif-1 retarded the influence of metformin, implicating the involvement of hif-1/nhr-57 in age-specific drug responses. These findings underscored the necessity of deciphering the mechanisms governing age-related susceptibility to pharmacological agents to tailor interventions for promoting successful aging.
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Alginate-encapsulated hepatocyte transplantation is a promising strategy to treat liver failure. However, its clinical application was impeded by the lack of primary human hepatocytes and difficulty in controlling their quality. We previously reported proliferating human hepatocytes (ProliHHs). Here, quality-controlled ProliHHs were produced in mass and engineered as liver organoids to improve their maturity. Encapsulated ProliHHs liver organoids (eLO) were intraperitoneally transplanted to treat liver failure animals. Notably, eLO treatment increased the survival of mice with post-hepatectomy liver failure (PHLF) and ameliorated hyperammonemia and hypoglycemia by providing liver functions. Additionally, eLO treatment protected the gut from PHLF-augmented permeability and normalized the increased serum endotoxin and inflammatory response, which facilitated liver regeneration. The therapeutic effect of eLO was additionally proved in acetaminophen-induced liver failure. Furthermore, we performed assessments of toxicity and biodistribution, demonstrating that eLO had no adverse effects on animals and remained non-tumorigenic.
Assuntos
Falência Hepática Aguda , Falência Hepática , Humanos , Camundongos , Animais , Falência Hepática Aguda/terapia , Falência Hepática Aguda/induzido quimicamente , Distribuição Tecidual , Células Cultivadas , Hepatócitos , Fígado , Falência Hepática/terapia , Falência Hepática/metabolismo , Organoides/metabolismoRESUMO
Cell-based ex vivo gene therapy in solid organs, especially the liver, has proven technically challenging. Here, we report a feasible strategy for the clinical application of hepatocyte therapy. We first generated high-quality autologous hepatocytes through the large-scale expansion of patient-derived hepatocytes. Moreover, the proliferating patient-derived hepatocytes, together with the AAV2.7m8 variant identified through screening, enabled CRISPR-Cas9-mediated targeted integration efficiently, achieving functional correction of pathogenic mutations in FAH or OTC. Importantly, these edited hepatocytes repopulated the injured mouse liver at high repopulation levels and underwent maturation, successfully treating mice with tyrosinemia following transplantation. Our study combines ex vivo large-scale cell expansion and gene editing in patient-derived transplantable hepatocytes, which holds potential for treating human liver diseases.
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Sistemas CRISPR-Cas , Edição de Genes , Terapia Genética , Hepatócitos , Hepatopatias , Hepatócitos/metabolismo , Hepatócitos/transplante , Sistemas CRISPR-Cas/genética , Humanos , Animais , Hepatopatias/terapia , Hepatopatias/genética , Hepatopatias/patologia , Camundongos , Terapia Genética/métodos , Tirosinemias/terapia , Tirosinemias/genética , Proliferação de Células , HidrolasesRESUMO
Cholestatic liver injuries, characterized by regional damage around the bile ductular region, lack curative therapies and cause considerable mortality. Here we generated a high-definition spatiotemporal atlas of gene expression during cholestatic injury and repair in mice by integrating spatial enhanced resolution omics sequencing and single-cell transcriptomics. Spatiotemporal analyses revealed a key role of cholangiocyte-driven signaling correlating with the periportal damage-repair response. Cholangiocytes express genes related to recruitment and differentiation of lipid-associated macrophages, which generate feedback signals enhancing ductular reaction. Moreover, cholangiocytes express high TGFß in association with the conversion of liver progenitor-like cells into cholangiocytes during injury and the dampened proliferation of periportal hepatocytes during recovery. Notably, Atoh8 restricts hepatocyte proliferation during 3,5-diethoxycarbonyl-1,4-dihydro-collidin damage and is quickly downregulated after injury withdrawal, allowing hepatocytes to respond to growth signals. Our findings lay a keystone for in-depth studies of cellular dynamics and molecular mechanisms of cholestatic injuries, which may further develop into therapies for cholangiopathies.
Assuntos
Colestase , Hepatócitos , Animais , Camundongos , Colestase/genética , Colestase/patologia , Colestase/metabolismo , Hepatócitos/metabolismo , Fígado/metabolismo , Fígado/lesões , Fígado/patologia , Proliferação de Células/genética , Ductos Biliares/metabolismo , Regeneração Hepática/genética , Camundongos Endogâmicos C57BL , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Transdução de Sinais , Masculino , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta/genética , Transcriptoma , Modelos Animais de Doenças , Análise Espaço-TemporalRESUMO
BACKGROUND: STIM1 (stromal interaction molecule 1) regulates store-operated calcium entry and is involved in pulmonary artery vasoconstriction and pulmonary artery smooth muscle cell proliferation, leading to pulmonary arterial hypertension (PAH). METHODS: Bioinformatics analysis and a 2-stage matched case-control study were conducted to screen for noncoding variants that may potentially affect STIM1 transcriptional regulation in 242 patients with idiopathic PAH and 414 healthy controls. Luciferase reporter assay, real-time quantitative polymerase chain reaction, western blot, 5-ethynyl-2'-deoxyuridine (EdU) assay, and intracellular Ca2+ measurement were performed to study the mechanistic roles of those STIM1 noncoding variants in PAH. RESULTS: Five noncoding variants (rs3794050, rs7934581, rs3750996, rs1561876, and rs3750994) were identified and genotyped using Sanger sequencing. Rs3794050, rs7934581, and rs1561876 were associated with idiopathic PAH (recessive model, all P<0.05). Bioinformatics analysis showed that these 3 noncoding variants possibly affect the enhancer function of STIM1 or the microRNA (miRNA) binding to STIM1. Functional validation performed in HEK293 and pulmonary artery smooth muscle cells demonstrated that the noncoding variant rs1561876-G (STIM1 mutant) had significantly stronger transcriptional activity than the wild-type counterpart, rs1561876-A, by affecting the transcriptional regulatory function of both hsa-miRNA-3140-5p and hsa-miRNA-4766-5p. rs1561876-G enhanced intracellular Ca2+ signaling in human pulmonary artery smooth muscle cells secondary to calcium-sensing receptor activation and promoted proliferation of pulmonary artery smooth muscle cells under both normoxia and hypoxia conditions, suggesting a possible contribution to PAH development. CONCLUSIONS: The potential clinical implications of the 3 noncoding variants of STIM1, rs3794050, rs7934581, and rs1561876, are 2-fold, as they may help predict the risk and prognosis of idiopathic PAH and guide investigations on novel therapeutic pathway(s).
Assuntos
Artéria Pulmonar , Molécula 1 de Interação Estromal , Humanos , Molécula 1 de Interação Estromal/genética , Molécula 1 de Interação Estromal/metabolismo , Masculino , Feminino , Estudos de Casos e Controles , Pessoa de Meia-Idade , Adulto , Proteínas de Neoplasias/genética , Miócitos de Músculo Liso/metabolismo , Hipertensão Pulmonar Primária Familiar/genética , Hipertensão Pulmonar Primária Familiar/fisiopatologia , Predisposição Genética para Doença , Músculo Liso Vascular/metabolismo , Regulação da Expressão Gênica , Proliferação de Células/genética , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Stem cell-independent reprogramming of differentiated cells has recently been identified as an important paradigm for repairing injured tissues. Following periportal injury, mature hepatocytes re-activate reprogramming/progenitor-related genes (RRGs) and dedifferentiate into liver progenitor-like cells (LPLCs) in both mice and humans, which contribute remarkably to regeneration. However, it remains unknown which and how external factors trigger hepatocyte reprogramming. Here, by employing single-cell transcriptional profiling and lineage-specific deletion tools, we uncovered that periportal-specific LPLC formation was initiated by regionally activated Kupffer cells but not peripheral monocyte-derived macrophages. Unexpectedly, using in vivo screening, the proinflammatory factor IL-6 was identified as the niche signal repurposed for RRG induction via STAT3 activation, which drove RRG expression through binding to their pre-accessible enhancers. Notably, RRGs were activated through injury-specific rather than liver embryogenesis-related enhancers. Collectively, these findings depict an injury-specific niche signal and the inflammation-mediated transcription in driving the conversion of hepatocytes into a progenitor phenotype.
Assuntos
Interleucina-6 , Células de Kupffer , Animais , Humanos , Camundongos , Diferenciação Celular , Hepatócitos/metabolismo , Interleucina-6/metabolismo , Células de Kupffer/fisiologia , Fígado , Regeneração Hepática/fisiologiaRESUMO
Liver resection is the first-line treatment for primary liver cancers, providing the potential for a cure. However, concerns about post-hepatectomy liver failure (PHLF), a leading cause of death following extended liver resection, have restricted the population of eligible patients. Here, we engineered a clinical-grade bioartificial liver (BAL) device employing human-induced hepatocytes (hiHeps) manufactured under GMP conditions. In a porcine PHLF model, the hiHep-BAL treatment showed a remarkable survival benefit. On top of the supportive function, hiHep-BAL treatment restored functions, specifically ammonia detoxification, of the remnant liver and facilitated liver regeneration. Notably, an investigator-initiated study in seven patients with extended liver resection demonstrated that hiHep-BAL treatment was well tolerated and associated with improved liver function and liver regeneration, meeting the primary outcome of safety and feasibility. These encouraging results warrant further testing of hiHep-BAL for PHLF, the success of which would broaden the population of patients eligible for liver resection.
Assuntos
Falência Hepática , Fígado Artificial , Humanos , Animais , Suínos , Hepatócitos , Falência Hepática/cirurgia , Regeneração HepáticaRESUMO
BACKGROUND: Intrauterine adhesions (IUA) develop in up to 20% of women with a history of abortion. After hysteroscopic adhesiolysis, balloon stents are usually placed for seven days to prevent recurrence. The efficacy of prolonged use (30 days) of balloon stents has not been determined. METHODS: The trial was conducted from June 2019 to March 2021. Ninety-one patients who underwent hysteroscopic adhesiolysis for moderate or severe IUA were randomized to receive a 30-day placement of a balloon stent (n = 44) or an intrauterine device (IUD) (n = 47). The primary outcomes were the ongoing pregnancy and miscarriage rates assessed at 15-19 months. The secondary outcomes were the recurrence of IUA and the American Fertility Society (AFS) intrauterine adhesion scores at the first and second hysteroscopies, the diagnosis of new chronic endometritis at the second-look hysteroscopy, and the vaginal/uterine microbiome assessed using 16S rRNA sequencing. The trial was registered at Chinese Clinical Trial Registry (ChiCTR1900023306). FINDINGS: The ongoing pregnancy rates (balloon 56·4% versus IUD 57·1%) and miscarriage rates (balloon 10·3% versus IUD 22·9%) were not significantly different between the groups. No differences in the recurrence of IUA, reduction of AFS scores, or new endometritis rates were detected. The bacterial load in the uterus and vagina increased in the IUD group but not in the balloon group. INTERPRETATION: Balloon placement has a similar effect on ongoing pregnancy rates as intrauterine device (IUD) placement. Patients who underwent balloon placement had a lower miscarriage rate, although the difference was not significant. There were no differences in the recurrence rate of IUA, reduction of American Fertility Society scores, or rate of new chronic endometritis. The balloon stent has less effects on the uterine microbiota. FUNDING: National Natural Science Foundation of China (81802593).
RESUMO
OBJECTIVE: To explore the clinical characteristics and possible pathological mechanisms of vertigo due to enlarged vestibule with lateral semicircular canal dysplasia. METHODS: A retrospective review was conducted for 5 cases of peripheral vertigo due to enlarged vestibule with lateral semicircular canal dysplasia. Their characteristics of medical history, precipitating factors, course of vertigo, auditory tests, vestibular tests and imaging examine results were analyzed. RESULTS: The clinical characteristics were as follows. (1) Specifics of medical history: 4 cases suffered delays in gross motor development and potential equilibrium dysfunctions. One case failed to recount an earlier medical history, but could maintain normal hearing and vestibular functions for a long time in adulthood. (2) Most cases could identify the precipitating factors of initial attacks, such as head-bumping, nose-blowing and constipation, etc. resulting in sudden rises of intracranial or abdominal pressures. (3) Paroxysmal vertigo and progressive hearing loss were mimicking Meniere disease or large vestibular aqueduct syndrome. But its course of vertigo was different from those of Meniere disease and large vestibular aqueduct syndrome with regards to hearing levels and audiograms. (4) Some cases had positional vertigo. But the results of Dix-Hallpike and Roll tests were different from benign paroxysmal positioning vertigo (BPPV). (5) The inner ear imaging showed enlarged vestibule with lateral semicircular canal dysplasia. CONCLUSION: The enlarged vestibule with lateral semicircular canal dysplasia is a rare etiology of peripheral vertigo. The history of delays in gross motor development and potential equilibrium dysfunctions in childhood may offer important diagnostic clues. And audiological and vestibular tests, high-resolution computed tomography and magnetic resonance may help to ascertain the diagnosis.